Pediatric Retina 3rd Ed.

PEDIATRIC RETINA
THIRD EDITION
PEDIATRIC RETINA
THIRD EDITION
M. Elizabeth Hartnett, MD, FACS, FARVO
Distinguished Professor in Ophthalmology and Visual Sciences
University of Utah
Calvin S. and JeNeal N. Hatch Presidential Endowed Chair in
Ophthalmology and Visual Sciences
Director of Pediatric Retina
Vitreoretinal Medical and Surgical Service
Adjunct Professor of Pediatrics Adjunct Professor of Neurobiology and
Anatomy Principal Investigator, Retinal Angiogenesis Laboratory
John A. Moran Eye CenterSalt Lake City, Utah
Section Editors
Arlene V. Drack, MD
Ronald Keech Professor of Pediatric Genetic Eye Disease
Research
Institute for Vision Research
Department of Ophthalmology and Visual Sciences
University of Iowa
Iowa City, Iowa
Antonio Capone Jr, MD, FACS

Professor of Ophthalmology
Oakland University William Beaumont Hospital School of Medicine
Co-President/Partner
Associated Retinal Consultants (ARC), PCAuburn Hills, Michigan
Michael T. Trese, MD
Clinical Professor
Department of Ophthalmology
Oakland University William Beaumont School of Medicine
Rochester, Michigan
Cynthia A. Toth, MD
Joseph AC Wadsworth Professor of Ophthalmology
Duke University
Durham, North Carolina
George Caputo, MD
Chairman
Rothschild Hospital Foundation
Paris, France
Acquisition Editor: Chris Teja
Development Editor: Eric McDermott
Editorial Coordinator: Cody Adams
Marketing Manager: Phyllis Hitner
Project Production Manager: Kirstin Johnson
Manufacturing Coordinator: Beth Welsh
Design Coordinator: Steve Druding
Production Service: SPi Global
Third Edition
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From MEH: To Bill, my love and source of creativity and fun, and
to my parents and family who always have supported and
encouraged me.
From Michael Trese: To Caron who keeps our whole family
together and inspires my work.
From Antonio Capone Jr: In honor of my parents, and with love to
my family.
From Arlene Drack: To Bill, Anya, and Elise for love and support,
and in honor of my parents, Earle and Mary Pagotto Drack.
From Cynthia Toth: To David for your love and support.
From George Caputo: To my family that I thank for their love and
support.
Generally from all: To our patients, mentors, and students who
inspire us to continue in our pursuits.
In the memory of Yomtov Robert Barishak (deceased 2018) and
Mina Millicent Chung (deceased 2020), whose invaluable
contributions enrich pediatric retinal clinicians and scientists
worldwide.
CONTRIBUTORS
Thomas M. Aaberg Sr, MD, MSPH

Emeritus Chairman and Professor
Emory University School of Medicine
Atlanta, Georgia
James D. Akula, PhD

Assistant Professor
Boston Children’s Hospital and Harvard Medical School
Boston, Massachusetts
Tomas S. Aleman, MD
Associate Professor
University of Pennsylvania Perelman School of Medicine
Philadelphia, Pennsylvania
Tala Al-Khaled, BA
Medical Student Researcher
Illinois Eye and Ear Infirmary
University of Illinois at Chicago
Chicago, Illinois
David Ancona-Lezama, MD
Tecnologico de Monterrey, Escuela de Medicina y Ciencias de la Salud
Institute of Ophthalmology and Visual Sciences
Hospital Zambrano Hellion
Monterrey, Nuevo Leon, Mexico
Stephen D. Anesi, MD
Physician
Massachusetts Eye Research and Surgery Institution
Waltham, Massachusetts
Samuel Asanad, BS
Medical Student
David Geffen School of Medicine
University of California Los Angeles
Los Angeles, California
Isabelle Audo, PU, PH

Professor
Department of Genetics
Sorbonne University
Paris, France
Laura Bagdonaite-Bejarano, MD
Clinical Fellow
Yomtov Robert Barishak, MD

Professor
Faculty of Medicine
University of Tel Aviv
Tel Aviv, Israel
Chrysanthi Basdekidou, MD
Pediatric OphthalmologistHead of the Pediatric Ophthalmology Department
Ophthalmica Institute
Thessaloniki, Greece
Jean Bennett, MD, PhD

Professor
Audina M. Berrocal, MD
Professor of Clinical Ophthalmology
Medical Director of Pediatric Retina and ROP
Vitreoretinal Fellowship Director
Bascom Palmer Eye Institute
Miller School of Medicine
University of Miami
Jackson Memorial Health System
Miami, Florida
Jesse L. Berry, MD
Associate Professor of Ophthalmology
Clinical Scholar
Keck School of Medicine at University of Southern California
Associate Director, Ocular Oncology
The Vision Center at Children’s Hospital Los Angeles
USC Roski Eye Institute
Saptarshi Biswas, PhD

Research Assistant
Department of Ophthalmology and Visual Science
State University of New York Upstate Medical University
Syracuse, New York
Andrew Blaikie, MBChB, BSc, Med Sci(path), FRCOphth
Consultant Ophthalmic Surgeon
Queen Margaret Hospital
Fife, United Kingdom
Colin A. Bretz, PhD

Postdoctoral Fellow
John A. Moran Eye Center
The University of Utah
Salt Lake City, Utah
Diana Schorry Brightman, PhD, MS, CGC

Licensed, Certified Genetic Counselor
Human Genetics
Cincinnati Children’s Hospital Medical Center
Cincinnati, Ohio
Meghan M. Brown, BS
MD Candidate
Rochester, Michigan
Beaumont Hospital
Royal Oak, Michigan
William J. Brunken, PhD, FARVO

Director of the Center for Vision Research
Professor and Vice Chair for Research, Ophthalmology and Visual Sciences
Professor of Neuroscience and Physiology
Syracuse, New York
Charles M. Calvo, MD
Vitreoretinal Specialist
Retina Consultants of Nevada
Las Vegas, Nevada
John Peter Campbell, MD, MPH

Assistant Professor of Ophthalmology
Oregon Health and Science University
Portland, Oregon
Antonio Capone Jr, MD, FACS

Oakland University William Beaumont Hospital School of Medicine
Co-President/Partner
Associated Retinal Consultants (ARC), PC
Auburn Hills, Michigan
Megan E. Capozzi, PhD

ScientistDuke Molecular Physiology Institute
Duke University School of Medicine
George Caputo, MD
Chairman
Rothschild Hospital Foundation
Paris, France
Joseph Carroll, PhD

Professor
Medical College of Wisconsin
Milwaukee, Wisconsin
R. V. Paul Chan, MD, MSc

The John H. Panton, MD Professor of Ophthalmology
Illinois Eye and Ear Infirmary
University of Illinois at Chicago
Chicago, Illinois
Emmanuel Y. Chang, MD, PhD

Pediatric Retina SpecialistRetina and Vitreous of Texas
Houston, Texas
Michael F. Chiang, MD
Knowles Professor of Ophthalmology & Medical Informatics and Clinical
Epidemiology
Associate Director, Casey Eye Institute
Oregon Health & Science University
Portland, Oregon
Itay Chowers, MD
Professor and Chairman
Hadassah–Hebrew University Medical Center
Jerusalem, Israel
Mina Chung, MD
Flaum Eye Institute
University of Rochester
Rochester, New York
Christopher D. Conrady, MD, PhD

Clinical Lecturer—Retina and Uveitis
University of Michigan
Ann Arbor, Michigan
Razek Georges Coussa, MD
Physician
Cleveland Clinic
Cleveland, Ohio
Donnell J. Creel, PhD

Research Professor in Ophthalmology and Visual Sciences
Neuroscience Program
University of Utah
Director, Division of Clinical Electrophysiology
University of Utah School of Medicine
Catherine A. Cukras, MD, PhD

Head, Unit on Clinical Investigation of Retinal Disease
Director, Medical Retina Fellowship Program
National Eye Institute
National Institutes of Health
Bethesda, Maryland
Patricia A. D’Amore, PhD, MBA, FARVO

Charles L. Schepens Professor of Ophthalmology, Professor of Pathology
Vice Chair of Basic and Translational Research, Department of
Ophthalmology
Harvard Medical School
Director of Howe Laboratory
Associate Chief of Basic and Translational Research
Mass. Eye and Ear
Senior Scientist
Director of Research
Schepens Eye Research Institute
Lauren A. Dalvin, MD
Assistant Professor
Mayo Clinic Minnesota
Rochester, Minnesota
Hanna M. De Bruyn, MS
Researcher
Department Ophthalmology
Boston Children’s Hospital
Paola Dorta Salfate, MD, MHA, MBA

Medical Director
KYDOFT Foundation
Santiago, Chile
Arlene V. Drack, MD
Ronald Keech Professor of Pediatric Genetic Eye Disease Research
University of Iowa
Iowa City, Iowa
Kimberly A. Drenser, MD, PhD

Vitreoretinal Surgeon
University of Illinois at Chicago College of Applied Health Sciences
Novi, Michigan
Alina V. Dumitrescu, MD
Assistant Professor
Department Ophthalmology
University of Iowa
Iowa City, Iowa
Anna L. Ells, MD, FRCS(C)
Clinical Professor
Department of Surgery
Faculty of Medicine
University of Calgary
Adjunct Professor
Department of Electrical and Computer Engineering
Schulich School of Engineering
University of Calgary
Pediatric Retina Specialist
Alberta Children’s Hospital
Retina Specialist
Calgary Retina Consultants
Calgary, Alberta, Canada
Lisa J. Faia, MD
Associate Professor in Ophthalmology
Rochester, Michigan
Director of Ocular Immunology and Uveitis Service
Associated Retinal Consultants, P.C.
Co-Director of Ocular Immunology and Uveitis Service
Beaumont Eye Institute
William Beaumont Hospital
Vitreoretinal Surgeon
Royal Oak, Michigan
Estephania Feria Anzaldo, MD

PhysicianVitreous Retina DepartmentAsociación Para Evitar La Ceguera en
MéxicoMexico City, Mexico
Philip J. Ferrone, MD
Staff Physician
Long Island Vitreoretinal Consultants
Clinical Associate Professor
Donald and Barbara Zucker School of Medicine of Northwell Hospital
Great Neck, New York
Clarisse M. Fligor, BS
Graduate Student
Department of Biology
Indiana University Purdue University at Indianapolis
Indianapolis, Indiana
C. Stephen Foster, MD, FACS, FACR, FARVO

Professor
Anne B. Fulton, MD
Professor
Clare E. Gilbert, FRCOpth, MD, MSc

Professor of International Eye Health
Clinical Research
London School of Hygiene & Tropical Medicine
London, United Kingdom
Morton F. Goldberg, MD, FACS, FARVO, FAOS

Green Professor of Ophthalmology
Director Emeritus
Wilmer Eye Institute
Johns Hopkins Medicine
Baltimore, Maryland
Mrinali P. Gupta, MD
Director, Medical Retina Fellowship Program
Weill Cornell Medical College
New York, New York
Ronald M. Hansen, PhD

Assistant Professor
Anna-Lena Hård, MD, PhD

Professor
Goteborgs universitet Institutionen for neurovetenskap och fysiologi
Gothenburg, Hovås, Sweden
Roger P. Harrie, MD
Adjunct Professor of Ophthalmology
Director of Ophthalmic Echography
John A Moran Eye Center
University of Utah
M. Elizabeth Hartnett, MD, FACS, FARVO

Distinguished Professor in Ophthalmology and Visual Sciences
University of Utah
Calvin S. and JeNeal N. Hatch Presidential Endowed Chair in
Ophthalmology and Visual Sciences
Director of Pediatric Retina
Adjunct Professor of Pediatrics
Adjunct Professor of Neurobiology and Anatomy
Principal Investigator, Retinal Angiogenesis Laboratory
Ann Hellström, MD, PhD

Professor
Goteborgs universitet Institutionen for neurovetenskap och fysiologi
Gothenburg, Hovås, Sweden
Ronald P. Hobbs, MD
Retinal SpecialistArizona Retina Associates
Mesa, Arizona
George Hoppe, PhD

Research Scientist
Cleveland Clinic Cole Eye Institute
Cleveland, Ohio
Kang-Chieh Huang, MS
Graduate Student
G. Baker Hubbard III, MD

Thomas M. Aaberg, Sr. Professor of Ophthalmology
Vice Chair of Clinical Operations
Director, Retina Service
The Emory Eye Center
Atlanta, Georgia
Dale D. Hunter, MD, JD

Senior Staff Scientist
Syracuse, New York
Laryssa A. Huryn, MD
Director, Ophthalmic Genetics Fellowship Program
Ophthalmic Genetics and Visual Function Branch
National Eye Institute
Bethesda, Maryland
Lea V. M. Hyvärinen, MD, PhD

Professor h.c., Rehabilitation Sciences
TU Dortmund
Dortmund, Germany
Senior Lecturer
Developmental Neuropsychology
University of Helsinki
Helsinki, Finland
Sana Idrees, MD
Vitreoretinal Fellow
University of Rochester
Rochester, New York
Raymond Iezzi Jr, MD

Associate Professor
Mayo Clinic
Marco H. Ji, MD
Research Fellow
Byers Eye Institute
Stanford University School of Medicine
Palo Alto, California
J. Michael Jumper, MD
Partner, West Coast Retina Medical Group
Clinical Professor and Vice Chairman
California Pacific Medical Center
San Francisco, California
Pavlina S. Kemp, MD
Pediatric Ophthalmology and Strabismus
Clinical Assistant Professor of Ophthalmology and Visual Sciences
Clinical Assistant Professor of Pediatrics
University of Iowa Hospitals and Clinics
Director of Medical Student Education
Iowa City, Iowa
John B. Kerrison, MD
Researcher
Department of Genetic Engineering and Molecular Ophthalmology
Baltimore, Maryland
Arif O. Khan, MD
Consultant, Pediatric Ophthalmology & Ocular Genetics
Eye Institute
Cleveland Clinic Abu Dhabi
Abu Dhabi, United Arab Emirates
Professor
Cleveland Clinic Lerner College of Medicine of Case Western University
Cleveland, Ohio
Samer Khateb, MD, PhD

Lecturer in Ophthalmology and Vision Sciences
Hebrew University
Vitreoretinal Diseases and Surgery Specialist
Retina Service
Hadassah Medical Center
Investigator at the Laboratory of Molecular Ophthalmology
Jerusalem, Israel
Matin Khoshnevis, MD
PGY3
Temple University
Sang Jin Kim, MD, PhD

Sungkyunkwan University School of Medicine
Seoul, Jongno-gu, Republic of Korea
Sylvia R. Kodsi, MD
Professor, Chief of Pediatric Ophthalmology and Strabismus
Donald and Barbara Zucker School of Medicine at Hofstra/Northwell
Robert K. Koenekoop, MD, PhD, MSc, FRCS(C), FARVO
Professor of Paediatric Surgery
Human Genetics and Adult Ophthalmology
McGill University Health Centre
Director, McGill Ocular Genetics Laboratory
Chief Paediatric Ophthalmology
Fellowship Director, AAPOS
Principle Investigator for Retinal Therapeutics at Center for Innovative
Medicine
RI-MUHC
Adjunct Professor of Experimental Medicine
McGill University and Optometry Université de Montréal
Montréal, Quebec, Canada
Alaa Koleilat, MS
PhD Candidate
Department of Clinical and Translational Sciences
Mayo Clinic Graduate School for Biomedical Sciences
Nikisha Kothari, MD
Fellow
University of California, Los Angeles
Eric Kunz, BS
Lab Specialist
Baruch D. Kuppermann, MD, PhD

Roger F. Steinert Professor and Chair
School of Medicine
University of California, Irvine
Director, Gavin Herbert Eye Institute
School of Medicine
University of California, Irvine
Joint Appointment, Professor
Department of Biomedical Engineering
The Henry Samueli School of Engineering
Irvine, California
Andrés Kychenthal Bab, MD

Director Vitreoretinal ServiceKYDOFT FoundationSantiago, Chile
Anna La Torre, PhD

Assistant Professor
Department of Cell Biology and Human Anatomy
University of California, Davis
Davis, California
Sailee S. Lavekar, MS
Graduate Student
Michelle E. LeBlanc, PhD

Postdoctoral Research Fellow
Jessica G. Lee, MD
Staff Physician
Long Island Vitreoretinal Consultants
Clinical Assistant Professor
Donald and Barbara Zucker School of Medicine of Northwell Hospital
Associate Adjunct Surgeon
New York Eye and Ear Infirmary of Mount Sinai
New York, New York
Katherine Ann Lee, MD, PhD

Pediatric Ophthalmologist
St. Luke’s Health System
Boise, Idaho
Thomas C. Lee, MD
Retinal Surgeon
Children’s Hospital of Los Angeles
Ana González H. León, MDPhysician

Retina and Vitreous Department
Asociación para Evitar la Ceguera
Mexico City, Mexico
Nathalie Lepvrier-Chomette, MD
Pediatric Ophthalmologist
La Maison Medicale
T. Y. Alvin Liu, MD
Assistant Professor
Johns Hopkins Medicine Wilmer Eye Institute
Baltimore, Maryland
Gerard A. Lutty, PhD
G. Edward and G. Britton Durell Professor of Ophthalmology
Director of the Ocular Angiogenesisi and Vasculogenesis Lab
Wilmer Ophthalmological Institute
Johns Hopkins Hospital
Baltimore, Maryland
Michele C. Madigan, PhD

Associate Professor
Department of Optometry and Vision Science
UNSW, Sydney
Kensington, New South Wales, Australia
Albert M. Maguire, MD
Professor
Niranjan Manoharan, MD
Fellow
Maria Ana Martinez-Castellanos, MDAssistant Professor

Oftalmología. Cirugía de Retina y Vitreo
Cirugía de Retina Pediátrica
Clínica de Enfermedades de los Ojos
Asociación Para Evitar la Ceguera
Mexico City, Mexico
D. Scott McLeod
Technician
Baltimore, MarylandAlice R. McPherson, MD, DSc, FACS, FICSProfessor
of OphthalmologyDepartment of OphthalmologyBaylor College of
MedicineHouston, Texas
Jason S. Meyer, PhD

Associate Professor
Department of Medical and Molecular Genetics
Department of Pharmacology and Toxicology
Stark Neurosciences Research Institute
Indiana University School of Medicine
P. Anthony Meza, MD
Attending Anesthesiologist
Department of Anesthesiology
Beaumont Health System
Bloomfield Hills, Michigan
Mikel Mikhail, MD
Retina Fellow
Associated Retina Consultants
Royal Oak, Michigan
Robert S. Molday, PhD, FRSC, FARVO

Professor of Biochemistry & Molecular Biology and Ophthalmology and
Visual Sciences
University of British Columbia
Canada Research Chair in Vision and Macular Degeneration
Director of the Macular Research Centre
Department of Biochemistry & Molecular Biology
University of British Columbia
Vancouver, British Columbia, Canada
Darius M. Moshfeghi, MD
Professor, Chief of Retina
Byers Eye Institute
Horngren Family Vitreoretinal Center
Stanford University School of Medicine
Palo Alto, California
Stavros N. Moysidis, MD
Retina Fellow
Royal Oak, Michigan
Timothy G. Murray, MD, MBA

Director
Murray Ocular Oncology and Retina
Miami, Florida
Eric Nudleman, MD, PhD

Assistant Professor
Shiley Eye Institute
University of California, San Diego
La Jolla, California
Joan Marie O’Brien, MD

Professor and Chair
University of Pennsylvania
Kean T. Oh, MD
Department of Surgery
Michigan State University College of Human Medicine
Partner
Traverse City, Michigan
Dolly A. Padovani-Claudio, MD, PhDPediatric Ophthalmologist

Vanderbilt University School of Medicine
Nashville, Tennessee
C. K. Patel, MRes(Oxon), FRCOphth

Consultant Paediatric Vitreoretinal Surgeon
Oxford University Hospitals NHS Foundation Trust
Honorary Senior Clinical Lecturer, University of Oxford
Honorary Consultant, Great Ormond Street Hospital for Sick Children,
London
Clinical Director
Oxford Eye Hospital
John Radcliffe Hospitals
Oxford, United Kingdom
Samir N. Patel, MD
Resident Physician
Wills Eye Hospital
Thomas Jefferson University
John S. Penn, PhD

Phyllis G. and William B. Snyder Endowed Professor and Vice Chairman
Professor of Cell & Developmental Biology
Professor or Molecular Physiology & Biophysics
Associate Dean for Faculty Affairs
Professor of Medical Education & Administration
Office of Faculty Affairs
Vanderbilt University School of Medicine
Nashville, Tennessee
Bridget J. Peterson, BA
Researcher
Wanda Pfeifer, OC(C), COMT, CO

Electrophysiology Service Coordinator
Iowa City, Iowa
Supalert Prakhunhungsit, MD
Vitreoretinal Unit
Ophthalmology Department
Faculty of Medicine
Siriraj Hospital
Mahidol University
Bangkok, Thailand
Jan M. Provis, PhD

Emeritus
Department of Neuroscience
ANU
Acton, Canberra, Australia
Aniket Ramshekar, MS
Graduate Student
Thomas A. Reh, PhD

Professor
Biological Structure
School of Medicine
University of Washington
Seattle, Washington
Sasha Rosen, BS
MS IV
Loyola University Health System
Medical School
Maywood, Illinois
Jonathan F. Russell, MD, PhD

Ophthalmology Resident
University of Miami Bascom Palmer Eye Institute
Miami, Florida
Stephen R. Russell, MD
Dana J. Schrage Professor of Macular Degeneration Research
Clinical Director, Carver Family Center for Macular Degeneration
Director, Vitreoretinal Service
Iowa City, Iowa
Ann Saada, PhD

Researcher
Department of Genetic and Metabolic Diseases
Jerusalem, Israel
Sherveen S. Salek, MD
Fellow, Vitreoretinal Surgery
Atlanta, Georgia
Briana L. Sawyer, MS
Cardiovascular Genetic Counselor
Division of Pediatric Cardiology
Department of Pediatrics
University of Utah
Lisa A. Schimmenti, MD, FACMG

Professor of Pediatrics
Mayo Clinic
Chair
Department of Clinical Genomics
Mayo Clinic
Brittni A. Scruggs, MD, PhD

Resident Physician
University of Iowa
Iowa City, Iowa
Jonathan E. Sears, MD
Staff
Cole Eye Institute and Cardiovascular and Metabolic Sciences
Cleveland Clinic
Cleveland, Ohio
J. Sebag, MD, FACS, FRCOphth, FARVO
Senior Research Scientist
Doheny Eye Institute
Professor of Clinical Ophthalmology
Geffen School of Medicine
Founding Director
VMR Institute for Vitreous Macula Retina
Huntington Beach, California
Seongjin Seo, PhD

Associate Professor
University of Iowa
Iowa City, Iowa
Khadija S. Shahid, OD, MPH

Clinical Assistant Professor of Ophthalmology
University of Iowa Carver College of Medicine
Iowa City, Iowa
Carol L. Shields, MD
Director of Ocular Oncology Service
Wills Eye Hospital
Consultant at Children’s Hospital of Philadelphia
Julia P. Shulman, MD
New York Medical College—TouroValhalla, New York
Chair, Department of Ophthalmology
Jamaica Hospital Medical Center
Richmond Hill, New York
Paul A. Sieving, MD, PhD

Director
National Eye institute
Bethesda, Maryland
Aaron B. Simmons, PhD

Postdoctoral Fellow
Lois E. H. Smith, MD, PhD

Department Of Ophthalmology
Senior Associate in Ophthalmology
Boston Children’s Hospital
Abraham Spierer, MD
Clinical Professor
Sheba Medical Center at Tel Hashomer
Ramat Gan, Israel
C. Gail Summers, MD
Professor
Department of Ophthalmology and Visual Neurosciences
Pediatric Ophthalmology and Strabismus
University of Minnesota
Minnesota Lions Children’s Eye Clinic
Minneapolis, Minnesota
Cynthia A. Toth, MD
Joseph AC Wadsworth Professor of Ophthalmology
Duke University
Elias I. Traboulsi, MD, MEd

Stanley Stone Endowed Chair of Pediatric Ophthalmology
Head, Department of Pediatric Ophthalmology & Strabismus
Director, The Center for Genetic Eye Diseases
Cole Eye Institute
Cleveland Clinic
Cleveland, Ohio
Michael T. Trese, MD
Clinical Professor
Rochester, Michigan
Karmen M. Trzupek, MS, CGC

Director, Clinical Trial Services
Director, Ocular & Rare Disease Genetics Services
InformedDNA
St Petersburg, Florida
Irena Tsui, MDPhysician

Stein Eye Institute
Doheny Eye Institute
David Geffen School of Medicine
Ronald Reagan Medical Center
Virginia Miraldi Utz, MD

Associate Professor
Cincinnati Children’s Hospital Medical Center
Abrahamson Pediatric Eye Institute
Cincinnati, Ohio
Lejla Vajzovic, MD
Associate Professor
Duke University
Kirstin B. VanderWall, BA
Graduate Student
Juliette Varin, MS
PhD Student
Sorbonne University
Paris, France
Victor M. Villegas, MD
Associate Professor
Universidad de Puerto Rico
San Juan, Puerto Rico
Anand Vinekar, MD, FRCS
Associate Professor and Head of Pediatric Retina
Postgraduate Institute of Ophthalmology
Narayana Nethralaya
Bangalore, India
Albert T. Vitale, MD
Professor
University of Utah Hospital
Mark K. Walsh, MD, PhD

Partner
Retina Associates Southwest
Department of Ophthalmology and Vision Science
University of Arizona College of Medicine
Tucson, Arizona
Haibo Wang, MD, PhD

Research Associate Professor in Ophthalmology and Visual Sciences
University of Utah
Co-investigator Retinal Angiogenesis Laboratory
Geoffrey Weiner, MD, PhD

Student
Shiley Eye Institute
University of California, San Diego
La Jolla, California
Mark E. Wilkinson, OD
Clinical Professor of Ophthalmology
Director, Vision Rehabilitation Service
Iowa Institute for Vision Research
Carver College of Medicine
University of Iowa
Iowa City, Iowa
Sui Chien Wong, MBBS, FRCSEd(Ophth)

Retinal Surgeon
Great Ormond Street Hospital for Children NHS Foundation Trust
Edward H. Wood, MD
Retina Fellow
Royal Oak, Michigan
Lauren M. Wright, MD
Vitreoretinal Surgery Fellow
Boston Medical Center
Wei-Chi Wu, MD, PhD

Retina Specialist
Chang Gung Memorial Hospital
Guishan, Taoyuan, China
Jinling Yang, PhD

Postdoctoral Research Fellow
Yoshihiro Yonekawa, MD
Wills Eye Hospital/Mid Atlantic Retina
Patrick Yu-Wai-Man, PhD

Researcher
Department of Clinical Neurosciences
University of Cambridge
Christina Zeitz, PhD, HDR

Research Director INSERM
Correspondent
Institut de la Vision
Group Leader of Team S6 A-Z
Institut de la Vision
Team S6 A-Z
INSERM, UMR_S968
CNRS, UMR_7210
Sorbonne University
Paris, France
FOREWORD
The textbook, Pediatric Retina, by M. Elizabeth Hartnett and authors

provides a comprehensive resource for pediatric retina specialists, retina
specialists, pediatric ophthalmologists, and trainees and clinicians in retina
who care for infants and children. The book is divided into sections covering
development, general assessment and visual rehabilitation of the infant and
child, imaging, disorders and genetic understanding of diseases,
pathophysiology and pharmacotherapies, tumors, uveitis, retinopathy of
prematurity, and medical and surgical management of diseases. This
organization allows one to focus on a condition by its genetics,
pathophysiology, or current strategies to manage and treat the condition. The
table of contents and text refer the reader to other chapters about the
condition, while the e-book provides links that bring the reader to related
chapters or images.
Pediatric retinal diseases comprise a number of conditions associated
with one or more causes: developmental abnormalities, infections, premature
birth, metabolic abnormalities, trauma, all of which can be influenced by
genetic variation. Most pediatric retinal conditions are rare, so the appearance
on clinical evaluation may not immediately trigger a diagnosis to the
examining clinician. The Atlas provides retinal appearances grouped by
descriptors that refer the reader to chapters in the text. New in this edition is a
bundled e-book that provides straight-forward ability to click reference or
figure links within the text to allow easy reading and comprehension, as well
as links to videos of surgery.
Pediatric Retina is an invaluable compendium with a bundled handy and
practical electronic version for any specialist caring for infants and children
with retinal diseases.
Alice R. McPherson, MD
PREFACE
Welcome to the third edition of Pediatric Retina. This expanded edition

includes 82 chapters, 8 atlases and an appendix of procedural videos by
internationally renowned experts that are organized into a table of contents
comprising 10 sections on development, visual assessment and rehabilitation,
imaging, genetics, pathophysiology, tumors, uveitis, retinopathy of
prematurity, and principles of and management of surgery for pediatric
retinal conditions. In this edition are new chapters on telemedicine,
addressing the needs of pediatric retina internationally, gene therapy, basic
mechanisms of disease, and methods to regenerate the retina. The atlases
provide retinal images of various pediatric retinal conditions grouped by
descriptive headings that allow the reader to navigate to structured chapters
and identify the condition or learn about others from a comprehensive
differential diagnosis in each chapter. These features guide the reader to a
diagnosis and recommended management of a particular condition.
This third edition includes access to a bundled e-book with numerous
cutting-edge chapters on stem cells and organoids in studying pediatric
retinal conditions, resources to diagnose and understand the effects of genetic
mutations on protein function, and biologic events that lead to pathology in
pediatric retinal conditions, as well as strategies to optimize screening and
treatment of ROP in several emerging countries. Within the e-book, is a
special section of surgical videos that provides visual guidance on surgical
techniques or strategies to address pediatric retinal conditions. The e-book
includes all chapters from the print version with links to figures, videos, and
other chapters for optimal functionality.
We trust this edition will provide a comprehensive resource for pediatric
and general retina specialists, pediatric ophthalmologists, clinician–scientists,
geneticists, trainees, and clinicians who care for infants and children and,
within the same text, provide new information to the experienced pediatric
retinal practitioner in an accessible way.
ACKNOWLEDGMENTS
In honor of the memory of Jonathan Pine, who was open to the idea of the
first edition of Pediatric Retina and encouraging as we developed the table of
contents. He understood the need for education in pediatric retinal conditions
that required bringing together experts in genetics (knowledge that was
emerging then), imaging, science, visual rehabilitation, surgery, and other
fields as well as the importance of an atlas of images that would alert the
reader to chapters describing conditions with similar fundus appearances. He
strongly supported the second edition. The third edition has expanded greatly
as our knowledge of pediatric retina conditions, their pathophysiology,
genetics, and science surrounding them has increased and technology in
imaging has allowed us to visualize retinal changes over time and test
management strategies. Furthermore, advances in publishing now allow us to
provide the e-book along with the print version.
There are many individuals to thank—
To all our section editors: Antonio Capone Jr, George Caputo, Arlene
Drack, Cynthia Toth, and Michael Trese for their exhaustive work to enhance
their sections, assuring current content. To all the authors of the third edition
and previous editions, who are experts providing knowledge, careful review
of the literature, their own research data, and input from their patients.
Thanks for careful proofing of each chapter’s material to meet the book’s
organization and need for accurate and comprehensive material. The
collective knowledge will allow us to develop better future treatments and
provide outstanding care to pediatric retina patients worldwide.
To my administrative coordinator, Maria Isabel Gomez, who was
essential in the entire process and assured on my end that the organization of
chapters and images were in order.
To the many photographers who provided outstanding images of retinal
conditions, angiographic studies, ultrasonography, and optical coherence
tomography. I especially thank Melissa Chandler CRA, OCT-C, James
Gilman, CRA, ROPS, and Glen Jenkins, CRA, OCT-C, COA for cover
design images. The work of the photographer is technical as well as an art.
We recognize the importance of engaging the small child to catch the most
instructive snapshot and that it is a skill. We have acknowledged their images
throughout the book and thank them here collectively.
To the staff at Wolters Kluwer Health who have been dedicated to create
this multi-dimensional edition. They have coordinated the drafts and videos
into a seamless end product and have been organized in their execution. I
thank specifically Chris Teja, the acquisition editor, who supported the third
edition and continued to see it through; Cody Adams, who has been
organized in assuring the drafts and images of the chapters are in order; and
Eric McDermott, who has organized the final aspects of the print edition and
the e-book edition; and thanks to Kirstin Johnson, Arunmozhivarman
Shenbagakutti and SPi Global for guiding the book through production. It has
been a pleasure to work with them all, and without their hard work this book
could not have happened.
We all, as pediatric retina specialists, recognize the importance of our
patients who not only encourage us to do our best and to find better
treatments for their rare conditions but also show us how valuable every
aspect of vision is in life. They keep us moving forward.
CONTENTS
Contributors
Foreword
Preface
Acknowledgments
SECTION I
DEVELOPMENT OF THE EYE AND RETINA
1 Embryology of the Retina and Developmental Disorders
Yomtov Robert Barishak and Abraham Spierer
2 The Hyaloidal Vasculature and Its Role in Development
Charles M. Calvo, Ronald P. Hobbs, and M. Elizabeth Hartnett
3 Vitreous and Developmental Vitreoretinopathies
Samuel Asanad and J. Sebag
4 Vitreous Biochemistry and Pharmacologic Vitreolysis
Sasha Rosen, Matin Khoshnevis, and J. Sebag
5 Retinal Vascular Development
Michelle E. LeBlanc, Jinling Yang, and Patricia A. D’Amore
6 Human Choriocapillaris Development
Gerard A. Lutty and D. Scott McLeod
7 Development of Cone Photoreceptors and the Fovea Centralis
and the Impact of Prematurity
Jan M. Provis and Michele C. Madigan
8 Retinal Development
Thomas A. Reh and Anna La Torre
SECTION II
ASSESSMENT AND REHABILITATION OF
VISUAL FUNCTION IN PEDIATRIC RETINAL
CONDITIONS
9 Assessment of Vision and Retinal Function in Infants and

Children
Anne B. Fulton, Ronald M. Hansen, Bridget J. Peterson, Laura
Bagdonaite-Bejarano, and James D. Akula
10 ERG Waveforms in Inherited Pediatric Retinal Diseases
Hanna M. De Bruyn, Wanda Pfeifer, Arlene V. Drack, Ronald M.
Hansen, and Anne B. Fulton
11 Amblyopia in Pediatric Retinal Conditions
Alina V. Dumitrescu and Pavlina S. Kemp
12 Workup of Infantile/Congenital Nystagmus
Brittni A. Scruggs and Arlene V. Drack
13 Early Intervention and Rehabilitation in Infants and Children
With Visual Impairment
Lea V. M. Hyvärinen
14 Low Vision Management of Children and Teens With Retinal
Disorders
Mark E. Wilkinson and Khadija S. Shahid
SECTION III
IMAGING OF THE INFANT AND CHILD EYE
AND RETINA
15 Retinal Photography and Multi-Wavelength Imaging in
Infants and Children
C. K. Patel and Andrew Blaikie
16 Optical Coherence Tomography in Infants and Children
Lejla Vajzovic, Anand Vinekar, and Cynthia A. Toth
17 Fluorescein Angiography in Pediatric Retinal Diseases
Nikisha Kothari, Niranjan Manoharan, and Irena Tsui
18 Ultrasonographic Imaging in Infants and Children
Roger P. Harrie
19 Imaging Analysis in Infants and Children
Sang Jin Kim, John Peter Campbell, and Michael F. Chiang
20 Image Storage and Retrieval and Telemedicine
Marco H. Ji, Darius M. Moshfeghi, and Antonio Capone Jr
SECTION IV
GENETICS AND DEVELOPMENTAL
DISORDERS IN PEDIATRIC RETINA
21 Genetic Counseling for Pediatric Retinal Diseases

Karmen M. Trzupek and Briana L. Sawyer
22 Anomalies of the Optic Nerve
Chrysanthi Basdekidou and George Caputo
23 Albinism
Katherine Ann Lee, Sylvia R. Kodsi, Joseph Carroll, and C. Gail
Summers
24 Neuronal Ceroid Lipofuscinoses and Lysosomal Storage
Diseases
Sana Idrees and Mina Chung
25 Retinal Manifestations of Metabolic Disease in Children
Arif O. Khan
26 Generalized Retinal and Choroidal Diseases
Meghan M. Brown and Kean T. Oh
27 Leber Congenital Amaurosis
Razek Georges Coussa, Robert K. Koenekoop, and Elias I. Traboulsi
28 Gene Therapy
Tomas S. Aleman, Albert M. Maguire, and Jean Bennett
29 Congenital Stationary Night Blindness (CSNB)
Christina Zeitz, Juliette Varin, and Isabelle Audo
30 Stargardt Macular Dystrophy/Fundus Flavimaculatus: From
Disease and Gene Discovery to Identification of Disease
Pathways to the Development of Therapies
Robert K. Koenekoop and Thomas M. Aaberg Sr
31 Mitochondrial, Peroxisomal, Glycosylation Disorders
Samer Khateb, Ann Saada, Patrick Yu-Wai-Man, John B. Kerrison,
and Itay Chowers
32 Bardet-Biedl Syndrome
Seongjin Seo and Arlene V. Drack
33 Usher Syndrome
Alaa Koleilat, Raymond Iezzi Jr, and Lisa A. Schimmenti
34 X-Linked Retinoschisis
Catherine A. Cukras, Laryssa A. Huryn, Michael T. Trese, and Paul A.
Sieving
35 Norrie Disease
Kimberly A. Drenser
36 Incontinentia Pigmenti
T. Y. Alvin Liu and Morton F. Goldberg
37 High Myopia and Vitreoretinopathies
Sherveen S. Salek and G. Baker Hubbard III
SECTION V
PATHOPHYSIOLOGY IN RETINAL DISEASES
38 Mechanisms of VEGF Signaling in Developmental and

Pathologic Angiogenesis Related to Retinopathy of
Prematurity
M. Elizabeth Hartnett, Haibo Wang, Aaron B. Simmons, Colin A.
Bretz, Eric Kunz, and Aniket Ramshekar
39 Weight Gain and Retinopathy of Prematurity
Ann Hellström, Lois E. H. Smith, and Anna-Lena Hård
40 Oxygen as a Pathogenic Factor in Retinopathy of Prematurity
(ROP)
Dolly A. Padovani-Claudio, Megan E. Capozzi, Colin A. Bretz, and
John S. Penn
41 Oxidative Mechanisms and Signaling Pathways Related to
Pathogenesis of Retinopathy of Prematurity
Haibo Wang and M. Elizabeth Hartnett
42 Signaling Pathways in Wnt and Effects on Disease
Eric Nudleman, Geoffrey Weiner, and Kimberly A. Drenser
43 Hypoxia-Inducible Factors and Prevention of ROP
Jonathan E. Sears and George Hoppe
SECTION VI
TUMORS
44 Retinoblastoma
Jesse L. Berry and Joan Marie O'Brien
45 Treatment of Retinoblastoma
Lauren A. Dalvin, David Ancona-Lezama, and Carol L. Shields
46 Tumors in Infants and Children
Victor M. Villegas and Timothy G. Murray
SECTION VII
UVEITIS IN INFANTS AND CHILDREN
47 Uveitis and Endophthalmitis Affecting Infants and Children

Christopher D. Conrady, Stephen D. Anesi, C. Stephen Foster, and
Albert T. Vitale
48 Surgical Approaches to Uveitis
Stavros N. Moysidis, Edward H. Wood, Baruch D. Kuppermann, and
Lisa J. Faia
49 Uveitis Masquerade Syndromes
Mikel Mikhail and Lisa J. Faia
SECTION VIII
RETINOPATHY OF PREMATURITY
50 Worldwide Causes of Childhood Blindness
Clare E. Gilbert and Nathalie Lepvrier-Chomette
51 Education and Management of Retinopathy of Prematurity
Worldwide
Tala Al-Khaled, Samir N. Patel, and R. V. Paul Chan
52 Clinical Trials and Management of Retinopathy of Prematurity
Julia P. Shulman and M. Elizabeth Hartnett
53 Anti-VEGF Treatment in Retinopathy of Prematurity
Anna L. Ells, Wei-Chi Wu, and Darius M. Moshfeghi
54 Evolution of Stage 4 Retinopathy of Prematurity
Antonio Capone Jr and Michael T. Trese
55 Treatment of Stages 4 and 5 Retinopathy of Prematurity
Antonio Capone Jr, Michael T. Trese, and M. Elizabeth Hartnett
SECTION IX
PRINCIPLES OF SURGERY FOR PEDIATRIC
RETINAL CONDITIONS
56 General Surgical Considerations and Preoperative

Management in Infants and Children
Michael T. Trese and Antonio Capone Jr
57 Anesthesia for Infants and Children
P. Anthony Meza
58 External and Minor Surgical Procedures
Emmanuel Y. Chang, Antonio Capone Jr, and Michael T. Trese
59 Anterior Segment Surgery Combinations With Posterior
Segment Diseases
Mark K. Walsh
60 Surgical Approaches to Infant and Childhood Retinal
Diseases: Scleral Buckles
Jessica G. Lee and Philip J. Ferrone
61 Surgical Approaches to Infant and Childhood Retinal
Diseases: Intravitreal Surgery
Jonathan E. Sears
62 Endoscopic Vitreous Surgery in Infants and Children
Sui Chien Wong and Thomas C. Lee
SECTION X
MANAGEMENT OF VITREORETINAL
CONDITIONS OF INFANTS AND CHILDREN
63 Pediatric Rhegmatogenous Retinal Detachment
Lejla Vajzovic
64 Coats Disease
J. Michael Jumper
65 Persistent Fetal Vasculature Syndrome
Supalert Prakhunhungsit and Audina M. Berrocal
66 Pediatric Vitreoretinopathies: Familial Exudative
Vitreoretinopathy, Norrie Disease, and Incontinentia Pigmenti
Lauren M. Wright and Yoshihiro Yonekawa
67 Epiretinal Membrane and Combined Hamartoma of the Retina
and Retinal Pigment Epithelium
Cynthia A. Toth
68 Surgery for Gene Therapy
Jonathan F. Russell, Albert M. Maguire, and Stephen R. Russell
69 Childhood Ocular Trauma
Mrinali P. Gupta and Philip J. Ferrone
70 Nonaccidental Head Trauma
George Caputo and Wei-Chi Wu
ATLAS
ATLAS A Hemorrhages
George Caputo
ATLAS B Macula
George Caputo
ATLAS C Pigmentary Changes
George Caputo
ATLAS D Posterior Segment Masses
George Caputo
ATLAS E Flecks and Spots
Michael T. Trese, Antonio Capone Jr, and M. Elizabeth Hartnett
ATLAS F Abnormal Retinal Vasculature
George Caputo
ATLAS G Retinal Detachment and Schisis
George Caputo
ATLAS H Optic Nerve
George Caputo
Index
APPENDIX OF PROCEDURAL VIDEOS

George Caputo
EBOOK CHAPTERS
e1 Basics of Genetic Testing and Methodology

Virginia Miraldi Utz and Diana Schorry Brightman
e2 Selected Pediatric Retinal Differential Diagnosis Based on
Clinical Symptoms and Signs Virginia
Virginia Miraldi Utz and Elias I. Traboulsi
e3 Online Resources for Pediatric Retinal Disorders
Arlene V. Drack
e4 Genetic Mutations and Related Proteins Associated With
Inherited Retinal Diseases
Robert S. Molday
e5 Extracellular Matrix Regulation of Vascular Development in
the Retina
Saptarshi Biswas, William J. Brunken, and Dale D. Hunter
Visual Anomalies Associated With Reduced Retinal
e6
Pigmentation
Donnell J. Creel
e7 Applications for Human Pluripotent Stem Cell-Derived
Retinal Cells in Development, Disease and Cellular
Replacement
Kirstin B. VanderWall, Clarisse M. Fligor, Sailee S. Lavekar, Kang-
Chieh Huang, and Jason S. Meyer
e8 Human pluripotent stem cell-derived retinal organoids as in
vitro models of retinogenesis and therapeutic tools for retinal
degeneration
Clarisse M. Fligor, Kang-Chieh Huang, Kirstin B. VanderWall, Sailee
S. Lavekar, and Jason S. Meyer
e9 Regenerative Medicine
Michael T. Trese, Edward H. Wood, Antonio Capone Jr, and Kimberly
A. Drenser
e10 International ROP: Retinopathy of Prematurity in Chile
Andrés Kychenthal Bab and Paola Dorta Salfate
e11 International ROP: Retinopathy of Prematurity in Mexico
María Ana Martínez-Castellanos, Ana González H. León, and
Estephania Feria Anzaldo
e12 International ROP: Retinopathy of Prematurity in India
Anand Vinekar
SECTION 1
Development of the Eye and Retina
1
Embryology of the Retina and
Developmental Disorders
Yomtov Robert Barishak and Abraham Spierer
The development of the retina (Tables 1-1 to 1-3) starts at the 4th week of
gestation with the invagination of the optic vesicle and formation of the optic
cup (Fig. 1-1) with the two layers of the optic cup that constitute the
rudimentary basis or anlage of the retina: the external layer of the retinal
pigment epithelium (RPE) and the internal layer of the sensory retina. The
invagination involves the ventrocaudal wall of the optic vesicle and causes
the formation of the embryonic fissure, which permits the hyaloid artery to
enter the developing optic cup cavity. Invagination also involves the optic
stalk and progressively occludes the optic vesicle cavity. The apposition of
the inner layer of the optic cup (the anlage of the sensory retina) to the
external layer of the optic cup (the anlage of the RPE) allows the axons of the
first ganglion cells to penetrate into the optic stalk. The narrowing optic
vesicle cavity becomes the potential subretinal space. At the end of the 4th
week, the vessels surrounding the neural tube spread over the optic cup and
the cells of the external layer of the cup, the prospective RPE cells. The RPE
cells acquire pigmentation. At this stage, the optic cup is surrounded by the
secondary mesenchyme as migration of neural crest cells into the primary
mesenchyme has already taken place.
TABLE 1-1 Stages of retinal development

RPE, retinal pigment epithelium.
TABLE 1-2 Time line of retinal development:

organogenesis
RPE, retinal pigment epithelium.
TABLE 1-3 Time line of retinal development:

differentiation
IRBP, interstitial retinol-binding protein.

FIGURE 1-1 A 4-week-old embryo. Invagination of the
optic vesicle causes the formation of the optic cup and the
embryonic fissure. (From Embryology of the eye [film].
San Francisco: American Academy of Ophthalmology,
1950. Copyright © 2020 American Academy of
Ophthalmology. Courtesy of Dr. Michael Hogan.)
At the 5th week, the RPE cell is made up of two to three layers of columnar,
pseudostratified pigmented cells attached one to another by junctional
complexes, the anlage of the membrane of Verhoeff. The inner layer of the
optic cup is the prospective sensory retina and is made up of an external layer
of nuclei, called the proliferative or germinative zone, and an anuclear,
marginal zone. The outermost cells of the germinative zone have cilia that
project toward the potential subretinal space and are joined to one another by
zonula adherens, the anlage of the external limiting membrane of the retina.
The anuclear, marginal zone is covered internally by a basal lamina, the
anlage of the inner limiting membrane of the retina. Four zones can be
distinguished at the distal portion of the optic cup. The first zone is the one
that constitutes the contact area between the outer retinal pigment epithelial
layer and the inner retinal neuroblastic layer. The second zone constitutes the
anlage of the marginal sinus of the iris. The third is a zone of progenitor cells,
and the fourth will be the anlage of the RPE and the neural retina (1).
The 6th week is characterized by the closure of the embryonic fissure.
The closure starts at the center of the fissure and proceeds anteriorly toward
the anterior rim of the optic cup and posteriorly into the optic stalk.
During the 7th week, the embryonic fissure closes completely and the
anterior notch at the anterior rim of the optic cup and the posterior notch at
the site of the prospective optic disc around the hyaloid artery disappear. The
RPE cells extend posteriorly as the layer of the outer cells of the optic stalk,
the precursor of the peripheral glial mantle of the optic nerve, which later
develops into the barrier between the axons of the optic nerve and the
surrounding mesenchyme (2). The RPE cells also differentiate by developing
apical villi, basal infoldings, smooth and rough endoplasmic reticulum,
ribosomes, premelanosomes, and melanosomes (3). In the sensory retina,
germinative cells proliferate and migrate inward giving rise to the outer
neuroblastic layer, the inner neuroblastic layer, and the layer of Chievitz in
between (Fig. 1-2). The outermost cells of the outer neuroblastic layer are the
anlage of the photoreceptors. At the posterior pole, around the future optic
disc, the innermost cells of the inner neuroblastic layer migrate inward. The
first cells to migrate inward are the ganglion cells.
FIGURE 1-2 A 7-week-old embryo. Cells from the
common neuroblastic layer of the retina arrange
themselves into an outer and an inner neuroblastic layer
causing the appearance of the layer of Chievitz in between.
(From Embryology of the eye [film]. San Francisco:
American Academy of Ophthalmology, 1950. Copyright
© 2020 American Academy of Ophthalmology. Courtesy
of Dr. Michael Hogan.)
At the 8th week, the first ganglion cells extend axons toward the optic stalk
(Fig. 1-3). These axons will develop into the future nerve fiber layer. The
axons extend to and penetrate the optic stalk requiring first the apoptosis
(programmed cell death) of the primitive neuroectodermal cells that fill the
optic stalk (4). As more ganglion cells differentiate and axons penetrate into
the optic stalk, a Bergmeister papilla forms as a conic mass of glial cells
covering the future optic disc area (5). Migrating retinal glioblasts, the future
retinal astroglia, arrange themselves along the axons and on the internal
limiting membrane (6).
FIGURE 1-3 A 7-week-old embryo. Migration inward of
the ganglion cells and appearance of their axons. (From
Embryology of the eye [film]. San Francisco: American
Academy of Ophthalmology, 1950. Copyright © 2020
American Academy of Ophthalmology. Courtesy of Dr.
Michael Hogan.)
At the 3rd month, the differentiation of the retina starts at the posterior pole
and progresses gradually toward the periphery; the peripheral retina
differentiates a short time after birth. Proliferation starts in the outer cells of
the neuroblastic layer and progresses inward, while differentiation starts at
the inner cells of the neuroblastic layer and extends outward. That is why the
ganglion cells are the first to differentiate and the outermost cells, the anlage
of the photoreceptors, are the last to differentiate (7). The differentiation in
the inner neuroblastic layer manifests as the formation of a separate layer of
ganglion cells, the formation of dendrites by the ganglion cells, the migration
inward of the amacrine cells, and the appearance of glycogen granules in the
cytoplasm of primitive Müller cells (8). The outer neuroblastic layer
differentiates by an inward migration of its most internal cells, the future
bipolar cells, and of those external to the bipolar cells, the future horizontal
cells. The migration of the future bipolar and horizontal cells causes the
occlusion of the layer of Chievitz and the formation of the external plexiform
layer (the internal plexiform layer was formed as a result of the migration of
the ganglion cells). The outermost cells of the outer neuroblastic layer are
connected one to the other with adherent junctions, which make up the future
external limiting membrane.
At the 4th month, all the major constituents of the retina are present (Fig.
1-4). The ora serrata appears as a line of demarcation at the peripheral retina.
At the posterior pole, the outer nuclear layer becomes the precursor of the
rod-free zone, the future fovea. The inner plexiform layer acquires ribbon and
conventional synapses. As neurons mature, so do primitive Müller cells,
which acquire more glycogen, intermediate filaments, myelin-associated
protein, and hyaluronic acid. The differentiation of the photoreceptors
manifests by the production of F-actin and alpha-tubulin, components of the
microtubules. At this point, there is not yet a subretinal space present. The
most important event occurring at this stage is the appearance of retinal
vessels. Cells originating inside the optic disc from the walls of the two
venous channels located on either side of the hyaloid artery and from the
adventitia of the hyaloid artery proliferate and migrate into the inner retina.
They differentiate into endothelial cells, which first make cords and then
canalize to form capillaries. These cells possess a vascular precursor marker,
such as CD39, and have been named angioblasts. The initial process of
retinal vascularization is attributed to the formation of de novo vessels from
precursor vascular cells and is thus believed due to vasculogenesis (9).
FIGURE 1-4 A 4-month-old human fetus. All the major
constituents of the retina are present. Cones (C), rods (R),
external plexiform layer, internal nuclear layer (INL),
internal plexiform layer (IPL), ganglion cell layer (GCL),
nerve fiber layer, and internal limiting membrane can be
clearly distinguished (×540). (Reprinted from Hollenberg
MJ, Spira AW. Early development of the human retina.
Can J Ophthalmol 1972;7(4):472–491. Copyright © 1972
Canadian Ophthalmological Society. With permission.)
The 5th month is characterized by the conspicuous differentiation of the

photoreceptors: Membrane infoldings become the tubular structures of the
outer segments and rhodopsin and S antigen appear (10). The differentiation
of the apical surfaces of the photoreceptors and of the apical villi of the RPE
cells leads to breakdown of junctions between these cells and the formation
of the subretinal space (Fig. 1-5). Photoreceptors secrete interstitial retinol-
binding protein (IRBP) into the subretinal space. After the photoreceptors
develop, horizontal cells become conspicuous in a row. At the internal layers
of the retina, amacrine and ganglion cells are apparent in their definitive
locations. Apoptosis of cells, including ganglion cells, causes cell debris that
is phagocytosed by surrounding cells that are of unconfirmed origin. The cell
loss is not uniform, being more pronounced at the periphery and causing a
centrifugal gradient in the distribution of ganglion cells in the fetal retina. At
this stage, the fovea could be identified by the presence of a single layer of
cones in the outer nuclear layer. Immunolabeling showed the presence of
synaptic proteins, cone and rod opsins, and Müller cells (11).
FIGURE 1-5 A 5-month-old human fetus. The inset
shows all the layers of the retina. Amacrine cells with
large, light-colored nuclei; Müller cells with dark, angular
nuclei; bipolar cells with small, oval nuclei of medium
density; horizontal cells with large nuclei. ON, outer
nuclear layer; IN, inner nuclear layer; asterisk, outer
plexiform layer. The main figure shows the pigment
epithelium–photoreceptor contact area. The blank arrow
points to the beginning of an inner segment. The thin solid
arrow indicates villi of a retinal pigment epithelial (PE)
cell. There are junctions between the RPE cells and the
photoreceptors. The subretinal space is not yet present. cil,
cilium; ELm, external limiting membrane; m,
mitochondria; N, nucleus of pigment epithelial cell; pi,
pigment granule; arrowheads, microvilli of pigment
epithelial cell (×24,000). (Reproduced with permission
from Ozanics J. Prenatal development of the eye and its
adnexa. In: Jakobiec FA, ed. Ocular anatomy, embryology,
and teratology. Philadelphia: Harper & Row,
1982:11–96.)
The pars plana begins to develop and renders the ora serrata more clearly
distinguishable. Retinal vascularization progresses rapidly. Newly formed
capillaries extend peripherally and form arteries and veins; the arteries and
veins present at the optic disc become the central retinal artery and veins. In
capillaries, cells in contact with the blood flow become endothelial cells, and
those surrounding endothelial cells become pericytes. Processes of astrocytes
attach to the collagenous matrix surrounding the capillaries (see also Chapter
5). A study was performed on the proteome profiles of the vitreous in human
embryos aged 14 to 29 weeks of gestation compared to those in young adults.
During the second trimester, the proteome started to undergo substantial
changes, and that was manifested by a marked decrease in most of the
proteins (12).
At the 6th month, cone photoreceptor differentiation occurs. Cone nuclei
are arranged in a row adjacent to the external limiting membrane, whereas
rod nuclei are located more internally. Tubular structures increase in number,
and mitochondria, ribosomes, and endoplasmic reticulum appear in the
prospective inner segment and primitive cone pedicles. Contact synapses are
apparent, but ribbon synapses have not formed (13). For the first time, the
macula appears as a bulging area with a thickened ganglion cell layer (Fig. 1-
6). Most of the photoreceptors are cones. There is a remnant of the layer of
Chievitz. Ganglion cells mature and accumulate cytoplasm: This maturation
starts at the posterior pole and progresses toward the periphery along with
developing retinal vascularization. Ganglion cells posterior to the edge of
advancing vessels are more mature than those anterior to them (14). Müller
cells are developing and are strongly attached to the internal limiting
membrane. Myelin-associated protein is present throughout all the layers of
the retina. Retinal vascularization continues its rapid progression by a process
of angiogenesis. More capillaries appear. Arterial and venous channels
develop while some of their side branches retract and atrophy, giving rise to
the formation of capillary-free perivascular zones. In all mammals with an
intraretinal capillary system, one can see that the capillary-free zone around
arteries is wider than that around veins. The width of the capillary-free zone
depends on the oxygen concentration in the blood flow. Raising the oxygen
concentration in the blood widens the periarterial capillary-free zone, while
reducing it narrows the capillary-free zone (15). The fetal retina is avascular
until the 4th month (16). With the development of the photoreceptors,
increased oxygen consumption is believed to cause hypoxia, which induces
the process of retinal vascularization. Retinal maturation precedes vascular
outgrowth. During the second trimester, the presence of lutein and its
oxidized forms has been detected in the RPE, retina, and vitreous body. The
antioxidant role of the carotenoids has been suggested as a factor in the
normal development of the retina (17).
FIGURE 1-6 A 6-month-old human fetus. The future

fovea at the central bulging area of the retina. Ganglion
cells present a multilayered arrangement. (From
Embryology of the eye [film]. San Francisco: American
Academy of Ophthalmology, 1950. Copyright © 2020
American Academy of Ophthalmology. Courtesy of Dr.
Michael Hogan.)
At the 7th month, the rods differentiate in the same way cones have
previously (Fig. 1-7). In the cones, tubular structures arrange as lamellar sacs,
and mitochondria aggregate at the ellipsoid. Cone and rod terminals develop
synaptic vesicles and ribbons. As a greater number of photoreceptors
develop, the subretinal space enlarges. The ora serrata appears as a circular
line covered by a fold of peripheral retina, the Lange fold (18). The macula
still lacks a foveal depression, and no new cones are generated in this area.
The fovea forms as a result of two kinds of migration. One wave of migration
takes place in the inner retina and causes the formation of the foveal pit and
foveal slope. It consists of the centrifugal displacement of ganglion cells,
their dendrites and synapses to bipolar cells, and the axons of the
photoreceptors. The axons of the outer plexiform layer elongate and form the
fibers of Henle layer. The second wave takes place in the external
photoreceptor layer and consists of a centripetal migration of photoreceptors,
cones, rods, and RPE cells (19). There is no vascularization within the fovea.
The macular area is encircled by capillaries, which do not proliferate over the
center. An interesting theory has been advanced by Springer (20) regarding
the formation of the foveal pit. He claims that two mechanical parameters are
needed to explain the passive movement of the inner retinal cells and that is
deformability and a force that deforms. Because vasculature is mechanically
stiff in relation to the retina at the foveal avascular zone, the inner retina at
the avascular fovea can be considered deformable. The second parameter is
the onset of the intraocular pressure, which acts on the deformable foveal
avascular zone. As a supporting finding to his theory, Springer argues that the
ciliary body and Schlemm’s canal develop before the foveal pit. So the
internal retinal cells do not have to migrate outward; they are instead pushed
outward. The author cannot offer an explanation for the inward migration of
the cones into the foveal pit.
FIGURE 1-7 A 7-month-old human fetus. Development
of the photoreceptor. A:Two outer segments (OS) of
which one is seen attached to an inner segment (IS) by a
connecting cilium (CC) and the other not; they contain
tubular structures. N, nucleus of pigment epithelial cell.
Inset: A primitive outer segment shows an invaginated
plasma membrane (arrows) at the lateral and apical
surfaces. Some tubular structures show a swelling at their
end. B:The OS contain numerous tubular structures
intermingled with each other; they are surrounded by the
apical surface and processes of RPE cells. T, terminal bar
between two pigment epithelial cells. Inset: Tubular
structures at higher magnification. (Reproduced from
Yamada I. Submicroscopic morphogenesis of the human
retina. In Rohen, ed., The Structure of the Eye, symposium
ii. Stuttgart: Schattauer; 1965, with permission. Copyright
© 1965 Schattauer / Thieme Group.
https://trove.nla.gov.au/work/209461253?
q&versionId=229835068.)
Retinal vascularization advances toward the periphery, but an avascular

retinal zone persists at the periphery. Endothelial cells of the retinal vessels
proliferate, form loops, and do not abut the ora serrata (21).
At the 8th month, photoreceptor differentiation extends to the ora serrata.
The photoreceptors elongate, and lamellar structures in their outer segments
acquire an adult arrangement as their inner segments mature. The subretinal
space extends to the ora serrata. The ganglion cell density remains highest in
the perifoveal region, and ganglion cells are rare in number at the periphery.
The macula continues to thin. Hoshino et al. (22) emphasize the dramatic
acceleration of development in the fovea compared to the development of the
peripheral retina and claim that this change is due to the transcriptional
networks controlling photoreceptor differentiation.
At the 9th month, the retina is well differentiated: RPE cells and
photoreceptors are mature although electrophysiologic and visual functions
continue to develop. The retinal surface increases although mitotic activity
has ceased, and this is due to a transverse shifting of retinal cells. As this
occurs, the RPE cells accommodate the change in retinal surface area by
changing their density and becoming wider. A perimacular reflex soon
becomes annular, followed by the appearance of a concavity and a foveolar
light reflex. Macular pigmentation appears at the 34th gestational week, and
the perimacular annular reflex at the 36th week (23). The foveolar reflex is
due to the deepening of the foveal depression (24). During the formation of
the foveolar depression, not only the ganglion cells but also amacrine,
bipolar, Müller, and horizontal cells move away from the fovea (25).
Functional development accompanies differentiation of the retina and the
appearance of the visual evoked response (26). Hendrickson (27) noted that
the outer plexiform layer of the retina has reached the peripheral edge of the
retina by late gestation whereas the inner plexiform layer has already reached
it by mid gestation. Hendrickson claims that only early intrauterine events
could affect the internal plexiform layer lamination in the central retina.
The inner retinal capillary plexus has extended to the temporal periphery,
and the deeper plexus has arisen from the inner plexus with vessels diving
toward the inner nuclear layer and peripherally extending to the ora serrata.
At birth, the development of the eye is complete except for the macula.
The size of the immature fovea is 5 degrees (28). In postnatal development of
the fovea, cones migrate inwardly and elongate. This elongation involves the
outer segment and inner axon and causes narrowing of the foveal cones. The
cone diameter decreases from 7.5 μm at birth to 2 μm at 45 months of age. As
a result, the foveal cone density increases from 18/100 at birth to 42/100 in
the adult eye. The foveola matures to adult configuration at the age of 45
months (29). The development of the fovea coincides with cortical neuronal
dendritic growth and synapse formation. Continued development of the
central nervous system ensues through 2 years of age but at a slower rate
(26,30).
PATHOLOGIC CONDITIONS
ASSOCIATED WITH ABERRANT
EMBRYOLOGIC DEVELOPMENT
During the 7th week, the embryonic fissure closes completely. Failure of
closure of the anterior (proximal) end of the embryonic fissure can result in
coloboma of the iris, ciliary body, and choroid at the inferonasal quadrant of
the globe (Fig. 1-8A and B). Failure of closure of the posterior end causes
coloboma of the optic disc (31). These colobomas are defined as typical
colobomas; atypical colobomas can be located anywhere in the globe and,
therefore, probably do not result from an embryonic fissure defect.
Abnormally persistent vascularized strands of the tunica vasculosa lentis
prevent normal iris growth and result in atypical coloboma of the iris in the
location where regression of the tunica vasculosa lentis failed (32). Coloboma
of the iris varies from a small notch at the pupillary margin to a large sector
defect. Cases with isolated iris coloboma usually have normal visual acuity.
FIGURE 1-8 Coloboma of the iris (A) and the retina and
choroid (B) in the same eye.
In optical nerve colobomas, there is a white bowl-shaped excavation of the

optic disc, usually located inferonasally (see also Chapter 22). The white-
appearing defect may involve the retina and choroid adjacent to the optic disc
and extend anteriorly to include the ciliary body and iris (33).
Microphthalmos is frequently present in cases of extensive coloboma and is
called colobomatous microphthalmos. Optic nerve coloboma may be
associated with systemic abnormalities, including cutaneous abnormalities
(focal dermal hypoplasia) and CHARGE (Coloboma, Heart anomaly, choanal
Atresia, mental Retardation, Genital and Ear anomalies) syndrome. Midline
facial and midbrain defects may be detected by a magnetic resonance
imaging (MRI) scan. Consultation with a nephrologist may be helpful to
detect life-threatening renal hypoplasia (34). Ocular anomalies found in
patients with chorioretinal colobomas include microcornea, choroidal
detachment, and retinal detachment (34). Retinal detachment may be from
defects in the abnormally developed tissue over the colobomatous defects,
from retinal tears in normal retina, or from a mechanism similar to that with
optic nerve pit and serous detachment of the macula (31). Visual acuity may
be mildly to severely affected depending on the extent of damage to the optic
disc and macula. Coloboma may present as sporadically as unilateral or
bilateral cases, or it may have an autosomal dominant pattern (33).
Coloboma of the lens is characterized by a notching in the equator of the
lens. As discussed previously, lack of absorption of part of the tunica
vasculosa lentis lateralis causes a localized inhibition of growth of the
zonular fibers and contiguous lens while the rest of the lens continues its
normal development. Lack of zonular traction in the defective region causes a
thicker and more spherical region of the lens. A coloboma may occur
separately or in conjunction with colobomas of the ciliary body.
Morning glory anomaly is an enlarged excavated disc, orange in color
with fibroglial tissue at its center. The disc is located at the center of a funnel-
shaped excavation of the posterior fundus. The blood vessels arise from the
periphery of the disc. Peripapillary retinal pigmentation surrounds the disc.
This is usually a unilateral defect, which presents a sporadic occurrence. The
embryonic defect of the morning glory anomaly is unknown although an
abnormal closure of the embryonic fissure was suspected (35). Kindler who
first described this anomaly related it to an abnormal development of
remnants of Bergmeister papilla (36). Visual acuity is usually decreased,
ranging from 20/100 to hand motion. Amblyopia may develop and further
reduce visual acuity (37). Retinal detachment developed in 26% to 38% of
patients (38). The pathogenesis of the retinal detachment has been attributed
to subretinal exudation (36), the source of which has been speculated to be
from the cerebral spinal fluid or vitreous, vitreoretinal traction (39), or
rhegmatogenous retinal detachment (40).
Peripapillary staphyloma is a posterior retinal excavation, at the bottom
of which the optic disc is located. Atrophic pigmentary changes can be seen
at the margin of the defect. Cases of contractile peripapillary staphyloma
have been described; retinal pulsation may follow the respiratory rhythm (41)
or occur at irregular intervals (42). This anomaly usually causes decreased
visual acuity although some cases with normal vision have been also reported
(43).
The PAX gene family plays an important role in mammalian
development. PAX2 gene is expressed during development in the optic and
otic vesicles and in the kidney. The Krd (kidney and retinal defects) mouse
carries a deletion on chromosome 19, which includes the PAX2 locus. The
heterozygous deletion of PAX2 is viable causing a variable, semidominant
phenotype, characterized by structural anomalies in the kidney, and an
appearance of the retina and the optic disc similar to the renal coloboma
(papillorenal) syndrome. Homozygosity causes embryonal death. In the
Krd/+ mouse embryo, these PAX2+ cells undergo abnormal morphogenetic
movements, and the embryonic fissure does not close normally, giving rise to
the retinal and optic disc defects that characterize the renal coloboma
syndrome (44).
Ocular manifestations described in this syndrome include, in addition to
the optic disc coloboma, abnormal vascular pattern of the optic disc (45) and
optic disc dysplasia (46). Extraocular anomalies include renal hypoplasia,
vesicoureteral reflux, and sensorineural hearing loss (47). Mutations in the
PAX2 gene were determined in some patients (48).
Parents and medical staff are often confronted with difficult ethical
decisions involving the birth of a child whose vision might be severely
afflicted.
Prenatal medicine and the understanding of the genetics and pathogenesis
of diseases have applied diagnostic methods as well as fetal therapy. Genetic
screening, gene therapy, and other applications of genetic engineering may be
used for the treatment, cure, or prevention of congenital diseases. It is based
on the concept that applying gene therapy vectors to the fetus in utero may
prevent the development of a congenital disease.
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2
The Hyaloidal Vasculature and Its
Role in Development
Charles M. Calvo, Ronald P. Hobbs, and M. Elizabeth Hartnett
The hyaloidal vasculature has an important role in many aspects of the

development of the eye. The normal growth and function of the eye require
the regression of the hyaloid vasculature. Failure in this process can lead to
blinding pathologies described as persistent fetal vasculature, discussed in
Chapters 3 and 65. The mechanisms involved are complex and only recently
are becoming better understood at a molecular level.
The hyaloidal vasculature is composed of the vasa hyaloidea propria,
tunica vasculosa lentis, and pupillary membrane. The vasa hyaloidea propria
consists of the hyaloid artery entering the embryonic fissure and branching
anteriorly through the vitreous to the lens. The tunica vasculosa lentis is a
capillary network that cups the posterior region of the developing lens, and
the pupillary membrane is an extension of the tunica vasculosa lentis
covering the anterior lens (Figure 2-1). The hyaloidal vasculature is believed
to be involved in the normal growth and maturation of the crystalline lens and
eye and eventually makes up the primary vitreous (1). It is first seen in
humans at approximately the 4th week of gestation and attains its maximum
prominence during the 9th week of gestation. Thereafter it normally
regresses. Not uncommonly, the remnants of the regressed hyaloidal
vasculature can be seen as a Mittendorf dot or a persistent pupillary
membrane in an otherwise normal eye (2).
FIGURE 2-1 Artist diagram of hyaloid artery at its
pinnacle in a 40-mm human embryo. a, pupillary
membrane; b, anterior tunica vasculosa lentis; c, outer set
of vasa hyaloidea propria; d, inner set of vasa hyaloidea
propria; e, main trunk of hyaloid artery emerging from
optic cup. (Reprinted from Mann I. Development of the
human eye, 3rd ed. Australia: Grune & Stratton, 1964.
Copyright © 1964 Elsevier. With permission.)
The structure of the hyaloid artery consists of three layers, the intima, the
media, and the adventitia (3). The intima consists of flattened nonfenestrated
endothelial cells connected by tight junctions that have a basement membrane
and an incomplete layer of pericytes. The media has concentric layers of
smooth muscle with basement membranes around each fiber. The adventitia
contains scattered fibroblasts and collagen. The walls of the vasa hyaloidea
propria and the tunica vasculosa lentis are small capillaries that consist of a
complete layer of nonfenestrated endothelium with intervening tight
junctions between adjacent endothelial cells encircled by a continuous
basement membrane and incomplete layer of pericytes in primates (4). The
formation of the hyaloid vasculature has been proposed to occur by
hemovasculogenesis (5) and becomes most prominent between 8 and 12
weeks. Regression of the hyaloidal circulation begins at 12 weeks (6) with
the vasa hyaloidea propria, followed by the tunica vasculosa lentis, and then
the pupillary membrane. The process of regression finishes at about 35 to 36
weeks of gestation with the complete loss of blood flow in the hyaloidal
artery (2). As the hyaloidal vasculature regresses, there is contemporaneous
development of the retinal vasculature. The primary vitreous retracts and
collagen fibers and hyaluronic acid are produced, making up the secondary
vitreous. The posterior segment is composed largely of the secondary
vitreous by the 6th month of gestation, and the primary vitreous at this point
is reduced to a central extension from the optic disc to the posterior lens
surface known as Cloquet canal (Figure 2-2).
FIGURE 2-2 A:Artist diagram of a 48-mm human
embryo with hyaloidal artery regression and early
formation of primary and secondary vitreous. A, primary
vitreous; B, secondary vitreous being formed by
disappearance of vasa hyaloidea propria. B:Artist diagram
of a 65-mm human embryo demonstrating Cloquet canal
and the contained hyaloid artery in central primary
vitreous. The condensation between primary and
secondary vitreous is seen forming the wall of Cloquet
canal. a, primary; b, secondary vitreous. (Reprinted from
Mann I. Development of the human eye, 3rd ed. Australia:
Grune & Stratton, 1964. Copyright © 1964 Elsevier. With
permission.)
Ida Mann described the regression of the human hyaloid through detailed
observations (2). She noted that as the lens increased in size, the mesh of
intertwined vessels that made up the tunica vasculosa lentis along the
posterior capsule became stretched and developed decreased vessel caliber.
At the same time, the vasa hyaloidea propria branches had reduced caliber at
their proximal ends near the origin at the main hyaloid artery and eventually
lost connection with it, although the distal ends of the vasa hyaloidea propria
remained continuous with vessels on the posterior surface of the lens (Figure
2-3). The atrophy of the vasa hyaloidea propria and of the more central set of
vitreous branches is normally complete by 8½ months. During the 7th month,
atrophy of the anterior tunica vasculosa lentis or pupillary membrane begins.
The atrophy of the pupillary membrane begins centrally over the lens where
the vessels undergo a gradual shrinkage of the vessel walls with decreasing
lumen size and ultimately cessation of blood flow. Once the cessation of
blood flow occurs, the central vessels shrink away (Figure 2-4). By 8½
months, most of the central loops of the pupillary membrane have atrophied.
Endothelial cell processes then fill the lumen, and macrophages form a plug
that occludes the vessel. The cells in the vessel wall then undergo necrosis
and are phagocytized by mononuclear phagocytes (7). Frequently, remains of
the pupillary membrane can be seen as fine strands that extend along the
pupillary margin.
FIGURE 2-3 Artist diagram showing the relation of the
hyaloid vessels on the lens during atrophy of the vasa
hyaloidea propria. 1, remains of the vasa hyaloidea
propria; 2, vessels of tunica vasculosa lentis; 3, pupillary
membrane vessels. Note main trunk of hyaloid posterior to
lens. (Reprinted from Mann I. Development of the human
eye, 3rd ed. Australia: Grune & Stratton, 1964. Copyright
© 1964 Elsevier. With permission.)
FIGURE 2-4 Artist representation of the appearance of

the iris and the pupillary membrane of a 7-month human
fetus. Note the thinner caliber of the central loops of the
pupillary membrane. These eventually shrink away. Not
infrequently, remnants of the pupillary membrane can be
seen at the iris edge. (Reprinted from Mann I.
Development of the human eye, 3rd ed. Australia: Grune &
Stratton, 1964. Copyright © 1964 Elsevier. With
permission.)
Regression of the hyaloid artery and its associated branches usually occurs
completely and without complications. Persistence of the hyaloid vascular
system occurs in 3% of full-term infants and in 95% of premature infants. A
persistent hyaloid artery can be associated with prepapillary or vitreous
hemorrhage (8). Anomalies involving incomplete regression of the
embryonic hyaloid vascular system occur in more than 90% of infants born
younger than 36 weeks of gestation and in over 95% of infants weighing <5
pounds at birth (9).
MOLECULAR EVENTS INVOLVED IN

THE HYALOID VASCULATURE
The events defining the formation of the hyaloidal vasculature remain
incompletely understood. There is evidence that vascular endothelial growth
factor (VEGF) is present within the lens while the hyaloidal system is present
(10). Additionally, there is evidence that many of the growth factors and
cytokines present during retinal vascular development are also seen during
the formation of the hyaloidal vasculature (11), but mechanisms are still
incompletely understood.
A proteomic analysis of the vitreous was conducted in the developing
mouse eye. Unlike in the human, the mouse hyaloidal circulation develops
between embryonic day 10.5 and is completed by embryonic day 13.
Regression begins at postnatal day 1 and is mostly complete between
postnatal day 16 to 30. The investigators found that groups of proteins tended
to shift from those known important in energy metabolism, cell proliferation,
and development at postnatal day 1 to those involved in signal transduction at
postnatal day 16. Throughout the time course, apoptotic and antiangiogenic
proteins were also present (12,13).
Other major pathways are believed to contribute to hyaloidal vascular
regression. It has been observed that macrophages play roles in inducing
programmed cell death and driving the regression of the hyaloid system
(14–16). Mice deficient in lymphomyeloid transcription factor, PU.1, lack
macrophages and have persistence of the hyaloidal vascular system
postnatally (17). Additional observations that deficiency in Wnt7b or in
angiopoietin 2 (Ang2) (18) led to persistence of the hyaloid support the role
of these factors in the regression of the hyaloidal vasculature. Wnt7b is
expressed by perivascular macrophages and activates the canonical Wnt
pathway in adjacent vascular endothelial cells inducing their cell cycle and
leading to increased sensitivity of the endothelial cells to cell death. The
angiopoietin receptor is a tyrosine kinase receptor, Tie2, and its best-
characterized ligands are angiopoietin 1 (Ang1) and angiopoietin 2 (Ang2).
Ang1 is associated with cell survival, in part through the phosphoinositol-3-
kinase–Akt signaling pathway. In contrast to Ang1, Ang2 is an antagonist
and interferes with the effect of Ang1, leading to suppression of survival
signals and thus destabilizing vessels. In the regressing hyaloid, Ang2 plays a
dual role in suppressing endothelial survival factor, Akt, and in stimulating
Wnt7b expression in macrophages (19). Ang2 appears to be expressed by
pericytes. Thus, all three cell types are important in murine hyaloidal
regression: pericytes produce Ang2, which inhibits the PI3-kinase–Akt
(survival signal) in vascular endothelial cells and also promotes Wnt7b
expression in macrophages; this in turn leads to apoptosis of vascular
endothelial cells (17,19). Norrin, a protein expressed by Müller glial cells,
and its receptor, Frizzled4 (Fz4), are also involved in retinal vascular
development through initiation of the Wnt cascade (20). Mutations in genes
coding for Norrin and Fz4 have been found in the conditions familial
exudative vitreoretinopathy and Norrie disease (21).
Another hypothesis for hyaloid vascular system regression is that it is
induced by a loss of one of its survival factors, VEGF. Absence or decrease
of VEGF expression may induce apoptosis of the vascular endothelial cells
(12), whereas an overexpression of VEGF from the lens early in development
leads to increased accumulation of angioblasts and vascular endothelial cells,
indicating that VEGF is important to the stability and maturation of the
hyaloid vascular system (22). However, there are reports that VEGF
expression is enhanced rather than decreased in the lens during hyaloid
vascular system regression and remains so in the adult (23,24). Recently,
investigators have demonstrated that VEGF concentrations can be attenuated
by the expression of VEGF receptors. A soluble form of vascular endothelial
growth factor receptor-1 (VEGFR1), expressed by the cornea, may have an
important role in the sequestration of VEGF leading to regression of the
pupillary membrane (25). Yoshikawa et al. showed that the timed regression
of the hyaloid vasculature in murine eyes did not result from down-regulation
of VEGF expression but that vascular endothelial growth factor receptor-2
(VEGFR2) was markedly up-regulated by retinal neurons prior to the
regression (26). This suggests that VEGF is bound to the neuronal VEGFR2,
thereby reducing the VEGF concentration needed to maintain the hyaloid
vasculature. Further evidence has supported this role of VEGFR2 expression
by retinal neurons in sequestering VEGF (27).
Murine studies show that light is involved in hyaloid regression and
retinal vascular development and that these events require controlled timing.
Mice born and reared in dark conditions have persistent hyaloidal vasculature
and retinal vascular abnormalities (28). Light triggers hyaloid regression, and
the precise timing appears to be immediately following delivery, supporting
the idea that light exposure is involved. Light-responsive melanopsin-
expressing ganglion cells increase retinal hypoxia through their metabolic
activity, which then induces VEGF. These events help to pattern retinal
vascular development. In addition, neuropsin-dependent retinal light
responses, particularly to violet light, suppress vitreous dopamine, whereas
dopamine suppresses the activity of VEGFR2 in hyaloidal endothelial cells
and promotes hyaloid regression (29). These studies show the role of light in
patterning of vitreous and retinal vasculature.
The cell–cell interactions between macrophages, endothelial cells, and
pericytes are crucial for hyaloid vascular system regression. However, it
remains largely unknown how macrophages are activated and interact closely
with vascular endothelial cells, how the Wnt–Ang pathway is activated
during hyaloid regression, and the relevance of studies in mice to humans. In
addition to the aforementioned pathways, other mechanisms have been
proposed in hyaloid vascular system regression and include apoptosis factors
(bcl-2, bax, and bak) (30,31), platelet–endothelial cell adhesion molecules
(32), activin receptor-like kinase-1 (33), and the production of transforming
growth factor beta, a potent inhibitor of vascular endothelial cell proliferation
by the hyalocytes (34). A reduction of endogenous “survival factors” below a
critical threshold may induce apoptosis; these include fibroblast growth factor
and platelet-derived growth factor along with the already mentioned VEGF
(35). Additionally, autophagy has also been shown to have a possible role in
the process of hyaloid regression (36). Difficulties in understanding human
pathophysiology exist because it is not possible to perform mechanistic
studies in human infants and there are no representative animal models.
In conclusion, regression of the hyaloid vasculature is a complex and
synchronized process which has much to be understood. Further work in
understanding these mechanisms may elucidate the vision-threatening
pathologies that occur from any number of errors in this process.
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3
Vitreous and Developmental
Vitreoretinopathies
Samuel Asanad and J. Sebag
Invisible (Figure 3-1) “by design,” vitreous was long unseen as important in
the physiology and pathology of the eye. Recent studies have determined that
vitreous plays a significant role in ocular health (1) and disease (1,2),
including a number of important vitreoretinal disorders that arise from
abnormal embryogenesis and development. Vitreous embryology is presented
in detail in Chapter 1. Notable is that primary vitreous is filled with blood
vessels during the first trimester (Figure 3-2). During the second trimester,
these vessels begin to disappear as the secondary vitreous is formed,
ultimately resulting in an exquisitely clear gel (Figure 3-1). The following
will review vitreous development and the congenital disorders that arise from
abnormalities in hyaloid vessel formation and regression during the primary
vitreous stage and biochemical abnormalities related to secondary vitreous
dysgenesis.
FIGURE 3-1 Vitreous from a 9-month-old child was
dissected of the sclera, choroid, and retina. In spite of the
specimen being on a surgical towel exposed to room air,
the gel state is maintained. (Specimen is courtesy of the
New England Eye Bank.)
FIGURE 3-2 Human primary vitreous featuring the
hyaloid artery (3) arising from the optic disc and
branching to form the VHP (2), which anastomoses with
the TVL and PM (1).
BIOCHEMISTRY OF THE VITREOUS
BODY (SEE ALSO CHAPTER 4)
At birth, vitreous is composed primarily of collagen, while hyaluronan (HA)
synthesis begins after birth. This results in a dense appearance on dark-field
slit microscopy (Figure 3-3), because collagen scatters light intensely. Due to
considerable hydrophilicity, HA generates a “swelling” pressure within the
burgeoning vitreous that contributes to growth of the eye and also spreads
apart the collagen fibrils to minimize light scattering, inducing transparency
(Figures. 3-1 and 3-4). This is evident when comparing dark-field
microscopy of a human embryo (Figure 3-3) with that in a 4-year-old child
(Figure 3-4).
FIGURE 3-3 Dark-field slit microscopy of vitreous from

a 33-week-gestational-age human. The anterior segment is
below where the posterior aspect of the lens can be seen.
Coursing through the central vitreous is the remnant of the
hyaloid artery, destined to become the Cloquet canal.
There is considerable light scattering by the gel vitreous.
FIGURE 3-4 Dark-field slit microscopy of vitreous from

a 4-year-old child. The posterior aspect of the lens is seen
below. Other than the peripheral vitreous cortex that
contains densely packed collagen fibrils, there is little light
scattering within the vitreous body.
There is heterogeneous distribution of collagen throughout the vitreous body.

Chemical (3,4) and light-scattering studies (5) have shown that the highest
density of collagen fibrils is present in the vitreous base, followed by the
posterior vitreous cortex anterior to the retina, and then the anterior vitreous
cortex behind the posterior chamber and lens. The lowest density is found in
the central vitreous and adjacent to the anterior cortical gel. HA molecules
have a different distribution from collagen, being most abundant in the
posterior cortical gel with a gradient of decreasing concentration centrally
and anteriorly (6,7).
*The term "hyaloid" is often employed to reference the anterior and posterior vitreous cortex, but
should be reserved for referring to the embryonic vasculature (see below).
Both collagen and HA are synthesized during childhood. Total collagen
content in the vitreous gel remains at about 0.05 mg until the third decade (8).
As collagen does not appreciably increase during this time but the size of the
vitreous does increase with growth, the density of collagen fibrils effectively
decreases. This could potentially weaken the collagen network and
destabilize the gel. However, since there is active synthesis of HA during this
time, the dramatic increase in HA concentration may stabilize the thinning
collagen network (9).
Bishop (10) proposed that the leucine-rich repeat protein, opticin, is the
predominant structural protein in short-range spacing of collagen fibrils.
Scott (11) and Mayne et al. (12) claimed that HA plays a pivotal role in
stabilizing the vitreous gel via long-range spacing. However, studies (13)
using HA lyase to digest vitreous HA found that the gel structure was not
destroyed, suggesting that HA is not essential for the maintenance of vitreous
gel stability, leading to the proposal that collagen alone is responsible for the
gel state of vitreous (10).
Primary Vitreous
Vitreous vascularization begins with the hyaloid artery entering the “eye”
through the optic cup. By 10 weeks of gestation (WG), the hyaloid system is
well established, branching to form the vasa hyaloidea propria (VHP) with
anastomoses to a dense capillary network, the tunica vasculosa lentis (TVL),
which is posterior to the lens, and the pupillary membrane (PM) adherent to
the anterior surface of the lens and iris diaphragm (14). Maximal
development of the hyaloid vasculature is reached by 12 to 13 WG (15)
although evidence of regression is apparent as early as 11 WG (16). The
hyaloid system shows clear signs of regression by 13 to 15 WG beginning in
the VHP followed by the TVL and then the PM (16–18).
The precise biochemical process involved in formation of the hyaloid
vasculature and its regression is poorly defined. However, recent studies have
revealed critical roles for various growth factors in vascular development.
Notably, hypoxia and vascular endothelial growth factor (VEGF) may trigger
the growth of the TVL and PM (19–22). Thus, decreased VEGF levels may
induce vessel regression. In addition, recent studies using whole-mount
immunostaining in mice have demonstrated the role of neurons in regulating
vessel regression. In particular, retinal neurons were shown to control the
precise switch from the fetal to the postnatal circulatory systems by timely
sequestering VEGF (23). In addition, immunohistochemical studies of human
umbilical vein endothelial cells (EC) and mouse eyes have demonstrated that
autophagy, in addition to apoptosis, may play a role in hyaloid vessel
regression during development, especially under hypoxic conditions. This
suggests that while hypoxia is a critical trigger for ocular development, it can
similarly induce autophagy and apoptosis in vascular EC for regression (24).
Furthermore, transforming growth factor (TGF) has been implicated in ocular
vessel remodeling since all four forms of TGF-β are found in vitreous
hyalocytes (25). There is also evidence that members of the Wnt signaling
pathway, particularly Wnt7b expressed in macrophages, are involved in
normal hyaloid regression (26) in the mouse, although this mechanism has
not been confirmed in humans.
Along with growth factors, a number of transcription factors have been
shown to regulate hyaloid vessel formation and regression. Using a mouse
model, Chen et al. illustrated the mechanistic role of the transcription factor,
Cited2 (Cbp/p300-interacting transactivator, with Glu/Asp-rich carboxy-
terminal domain, 2). They demonstrated the role(s) of Cited2 together with
paired box 6 (Pax6) in the normal formation/regression of the hyaloid
vasculature through negative modulation of hypoxia-inducible factor-1 (HIF-
1) signaling (27). Moreover, proteomic investigations of primary vitreous
composition have shown dynamic changes in protein composition as a
function of vessel formation and regression. In particular, studies of vitreous
from 14 to 20 WG human embryos and young adult human eyes have
identified 1,217 proteins, 768 of which were not previously identified. It was
learned that the protein profile of human embryonic vitreous differs from
young adults and that in the embryo, this changes significantly during the
second trimester (14 to 20 WG). Alteration in protein composition was
shown to happen concurrently with regression of embryonic vitreous vessels,
as well as other developmental changes in the eye. These findings suggest
that the associated changes in proteomic profile of human fetal vitreous
during the second trimester may be related to the regression of the embryonic
vitreous vasculature (28).
Multidimensional liquid chromatography and tandem mass spectrometry
studies examined the proteomic profiles of vitreous substructures (the
anterior vitreous cortex, posterior vitreous cortex, central vitreous body, and
vitreous base) finding distinct molecular profiles associated with each
substructure, perhaps related to their unique functions (29). These recent
findings support previous hypotheses that vitreous may function to prevent
infection and regulate oxidative stress. Concerning the latter, there is
increasing evidence that vitreous regulates lens exposure to oxygen and
oxidative stress (30). Deficiencies in these properties due to inherited
vitreoretinal dystrophies, perinatal vitreous malformation, and vitreous
degeneration in aging may contribute to cataracts via oxidative stress.
Protease signaling, including the blood coagulation cascade and matrix
metalloproteases, may also be a major feature of the vitreous physiology in
health. In understanding disease, this knowledge can help link specific
proteins and their respective protein pathways to pathophysiologic processes
including age-related vitreous degeneration, cataract formation with aging
and after vitrectomy, proliferative vitreoretinopathy, diabetic
vitreoretinopathy, and various inflammatory disorders, such as
endophthalmitis and posterior uveitis (29).
In addition to regulatory factors, the developmental modality of the
human hyaloid vascular system remains controversial. In contrast to previous
studies, which suggested embryonic vitreous vessel formation via
vasculogenesis, current concepts are more inclined toward angiogenesis as
the mode of development. More recent studies examining embryonic and
vascular development support the view that similar to the choriocapillaris, the
hyaloid vasculature develops by hemo-vasculogenesis, which is the process
by which vasculogenesis, erythropoiesis, and hematopoiesis occur
simultaneously from common hemangioblast precursors (31).
Atrophy of the vessels begins posteriorly with dropout of the VHP,
followed by the TVL. Recent studies have detected the onset of apoptosis in
the EC of the TVL as early as day 17.5 in the mouse embryo (19). At the
240-mm stage (7th month) in the human, blood flow in the hyaloid artery
ceases. Regression of the vessel itself begins with glycogen and lipid
deposition in the EC and pericytes (PC) of the hyaloid vessels (32). EC
processes then fill the lumen, and macrophages form a plug that occludes the
vessel. The cells in the vessel wall then undergo necrosis and are
phagocytized by mononuclear phagocytes (33) identified as hyalocytes (34).
Recent studies suggest that the VHP and the TVL regress via apoptosis (35).
Mitchell et al. (19) point out that the first event in hyaloid vessel regression is
EC apoptosis and propose that lens development separates the fetal
vasculature from VEGF-producing cells, decreasing the levels of this survival
factor for vascular endothelium, inducing apoptosis. Following EC apoptosis,
there is loss of capillary integrity, leakage of erythrocytes into the vitreous,
and phagocytosis of apoptotic endothelium by hyalocytes. Meeson et al. (36)
proposed that there are actually two forms of apoptosis that are important in
regression of the fetal vitreous vasculature. The first (“initiating apoptosis”)
results from macrophage induction of apoptosis in a single EC of an
otherwise healthy capillary segment with normal blood flow. The isolated
dying EC project into the capillary lumen and interfere with blood flow. This
stimulates synchronous apoptosis of downstream EC (“secondary apoptosis”)
and ultimately obliteration of the vasculature. Removal of the apoptotic
vessels is achieved by hyalocytes.
Proteomic analysis of embryonic (aged 14 to 20 WG) human vitreous
(vitreomics) revealed that during hyaloid vessel regression, there is a
significant decrease in profilin-1 actin-binding protein and significant
increases in cadherin-2 cell adhesion protein, cystatin-C protease inhibitor,
dystroglycan cell adhesion molecule, clusterin, as well as pigment
epithelium–derived factor (PEDF), both known to have antiangiogenic
influences (37). The presence of dystroglycan and profilin-1 was confirmed
in the hyaloid vessels of 10, 14, and 18 WG embryonic human eyes by
immunolabeling (38). Clusterin was not present in the HA or TVL of 10 and
14 WG eyes, but was found in the HA of 18 WG eyes. Cadherin was not
found in the hyaloid vessels of the 10 WG eyes but was observed in the
hyaloid vessels at 14 and 18 WG (38).
Comparison of the protein composition of the adult human and
developing human vitreous from the second trimester (14 to 20 WG) revealed
314 and 1,213 proteins, respectively, including 78 proteins that were unique
to the adult and 1,002 unique to the fetal vitreous (39). A large fraction of the
fetal vitreous proteins are intracellular and may be involved in the remodeling
of the hyaloid vasculature and retina during the second trimester. Proteins
that were found in greater abundance in fetal vitreous compared to adult
vitreous included low molecular weight (MW) kininogen-1, cystatin-B,
insulin-like growth factor–binding protein (IGFBP)-2 and IGFBP-4,
thrombospondin-1 and thrombospondin-4, thioredoxin, mu-crystallin, alpha
2-antiplasmin, nidogen-2 and growth differentiation factor 8, as well as
secreted proteins PEDF, AMBP (α-1-microglobulin), hemoglobin, and Ig
chains. Low MW extracellular matrix proteins detected in fetal vitreous
included COL5A1, 6A3, 11A1, 15A1, 1A1, 2A1, 4A2, and 18A1 (39).
Bioinformatic analysis of the protein profile of fetal (14 to 20 WG)
human vitreous revealed that most proteins were members of either the free
radical scavenging, molecular transport, or small molecule biochemistry or
connective tissue disorder networks. Those proteins involved in free radical
scavenging, molecular transport, and small molecule biochemistry networks
were all intracellular and decreased during the second trimester. In contrast,
the proteins in the connective tissue disorder networks were all extracellular
and increased during the second trimester. This pattern is consistent with
replacement of the cellular primary vitreous with the acellular collagenous
secondary vitreous. This analysis further revealed that EIF2 signaling and
protein ubiquitination were the top canonical pathways (40). While intriguing
and essential (because they are in humans), these studies lack controls, so
mechanisms remain unclear and more research in this area is needed.
Studies of abnormal hyaloid vessel regression found that germ-line
deletion of Bim (proapoptotic factor) results in persistent hyaloid vasculature,
increased retinal vascular density, and prevents retinal vessel regression in
response to hyperoxia (41). Recently, researchers have sought to determine
whether retinal vascular regression is attributable to Bim expression in EC or
PC using transgenic mice. Interestingly, these studies observed attenuation of
hyaloid vessel regression and postnatal vascular pruning specifically in mice
lacking Bim in EC or PC. In addition, apoptosis and proliferation were also
decreased in the retinal vasculature of BimEC and BimPC mice (42).
Secondary Vitreous
Secondary (avascular) vitreous formation arises from a remodeling of the
earlier primary (vascular) vitreous (43). By the 40- to 60-mm stage in
humans, the VHP has reached maximum maturity and regression begins (22).
The posterior VHP is the first component to degenerate. Regression continues
to move centrally affecting the TVL and finally the hyaloid artery itself.
Although flow through the hyaloid artery decreases during this time ceasing
entirely around the 240-mm stage, the artery may continue to elongate as the
eye lengthens (44). As the primary vitreous regresses, its “remnants” provide
a framework for collagen fibrils of the secondary vitreous to form via
interactive remodeling. As this occurs, the primary and secondary vitreous
coexist in the same space during this transitional developmental period
(43,44).
VITREOUS STRUCTURE
In the human embryo, vitreous structure has a dense homogenous appearance
(Figure. 3-3) primarily due to the aforementioned collagen synthesis during
secondary vitreous formation (45–51). HA synthesis after birth separates
collagen fibrils, inducing transparency (Figure 3-4). In the absence of
diabetes and myopia, vitreous remains largely transparent until around the
fourth or fifth decade when fine, parallel fibers appear coursing in an
anteroposterior direction (Figure 3-5B and C) (2,48–50). The fibers arise
from the vitreous base (Figure 3-5H) where they insert anterior and posterior
to the ora serrata. As the peripheral fibers course posteriorly, they are
circumferential with the vitreous cortex, while central fibers “undulate” in a
configuration parallel with Cloquet canal (51). The fibers are continuous and
do not branch. Posteriorly, these fibers insert into the vitreous cortex (Figure
3-5E and F).
FIGURE 3-5 A–H:Dark-field slit microscopy of human
vitreous in eyes dissected of the sclera, choroid, and retina.
The anterior segment is below and the posterior pole is
above in all images. (Specimens courtesy of the New York
Eye Bank for Sight Restoration. Images used with
permission from Sebag J. The Vitreous – Structure,
Function, and Pathobiology. New York, NY: Springer-
Verlag; 1989:41.)
Ultrastructural studies (52) demonstrated that collagen fibrils are the only
microscopic structures that could correspond to these fibers. These studies
also detected the presence of bundles of packed, parallel collagen fibrils.
Eventually, the aggregates of collagen fibrils attain sufficiently large
proportions so as to be visualized in vitro (Figure 3-5) and clinically. The
areas adjacent to these large fibers have a low density of collagen fibrils
separated by HA molecules and therefore do not scatter light as intensely as
the bundles of collagen fibrils.
Vitreous Base
The vitreous base is a three-dimensional zone extending 1.5 to 2 mm anterior
to the ora serrata, 1 to 3 mm posterior to the ora serrata (53), and several
millimeters into the vitreous body itself (54). The posterior extent of the
posterior border of the vitreous base varies with age (55,56). In a variety of
developmental vitreoretinal disorders, the secondary vitreous never forms
temporally, likely due to the lack of proper formation of the peripheral retina
in this location. Thus, there is a different peripheral terminus bounded by
vitreous gel posteriorly and liquid vitreous anteriorly (see below).
Proteomic studies of different parts of the vitreous body found low
complexity protein networks in the vitreous base relative to the highly
complex networks in the anterior and posterior vitreous cortices. In particular,
the largest network unique to the vitreous base was the glycogen synthase
kinase 3 beta subunit (GSK3 beta) protein complex. Other pathways in the
vitreous base include protein kinase R (PKR) in apoptosis, DNA replication,
methionine–cysteine–glutamate metabolism, high-temperature requirement
A1 (HTRA1) signaling, protein kinase A (PKA) signaling, sulfur
metabolism, and interferon immune response (29). The relationship between
these findings and the different functions of different substructures of the
vitreous body remain to be determined.
Vitreous fibers enter the vitreous base by splaying out to insert anterior
and posterior to the ora serrata (Figure 3-5H). The anterior-most fibers form
the “anterior loop” of the vitreous base, a structure that is important in the
pathophysiology of anterior proliferative vitreoretinopathy (2,57–59). Studies
by Gloor and Daicker (60) showed that cords of vitreous collagen insert into
gaps between the neuroglia of the peripheral retina. They likened this
structure to Velcro (a self-adhesive nylon material) and proposed that this
would explain the strong vitreoretinal adhesion at this site. In the anterior
vitreous base, fibrils interdigitate with a reticular complex of fibrillar
basement membrane material between the crevices of the nonpigmented
ciliary epithelium (61). The vitreous base also contains intact cells that are
fibroblast-like anterior to the ora serrata and macrophage-like posteriorly.
Damaged cells in different stages of involution and fragments of basal
laminae, presumed to be remnants of the embryonic hyaloid vascular system,
are also present in the vitreous base (61). Exposure of the adult immune
system to these previously sequestered fetal remnants (antigens) later in life
may play a role in peripheral uveitis, perhaps better termed “peripheral
anterior vitritis.”
Vitreous Cortex
The vitreous cortex is the peripheral shell of the vitreous body that courses
forward and inward from the anterior vitreous base to form the anterior
vitreous cortex and posteriorly from the posterior border of the vitreous base
to form the posterior vitreous cortex. The anterior vitreous cortex, often
referred to as the “anterior hyaloid face,” begins about 1.5 mm anterior to the
ora serrata. The posterior vitreous cortex is 100 to 110 μm thick (62) and
consists of densely packed collagen fibrils (Figure 3-6, bottom). There is no
vitreous cortex over the optic disc (Figure 3-4A), and the cortex is thin over
the macula due to rarefaction of the collagen fibrils (62). The prepapillary
hole in the posterior vitreous cortex can sometimes be visualized clinically
when the posterior vitreous is detached from the retina. If peripapillary glial
tissue is torn away during posterior vitreous detachment (PVD) and remains
attached to the vitreous cortex around the prepapillary hole, it is referred to as
Vogt or Weiss ring. Vitreous can extrude through the prepapillary hole in the
vitreous cortex (Figure 3-4A) but does so to a much lesser extent than
through the premacular vitreous cortex where it can produce traction and
certain forms of maculopathy (63–65).
FIGURE 3-6 Scanning electron microscopy of the
anterior surface of the ILM of the retina (A) and the
posterior aspect of the posterior vitreous cortex in a human
(B).
According to recent proteomic studies, the protein networks constituting the

vitreous cortex are highly complex relative to adjacent vitreous substructures.
The most represented pathways in the cortex involve macrophage migration
inhibitory factor (MIF)-mediated immune response, regulation of cystic
fibrosis transmembrane conductance regular (CFTR) activity, TGF-β1–
dependent inhibition of CFTR expression, PKA signaling, and activin A in
cell differentiation/proliferation (29). Teleologically, it would seem important
that migration and inhibition occur at the vitreoretinal interface so as to
mitigate against unwanted cell migration into vitreous, thus promoting clarity
within the vitreous body. The other findings may relate to the presence of
hyalocytes.
Hyalocytes
Hyalocytes are mononuclear cells (Figure 3-7) that are embedded in the
vitreous cortex and widely spread apart in a monolayer 20 to 50 μm from the
retina. Quantitative studies of cell density in the bovine (66) and rabbit (67)
vitreous found the highest density of hyalocytes in the vitreous base,
followed by the posterior pole, and the lowest density at the equator.
Hyalocytes are oval or spindle-shaped, are 10 to 15 μm in diameter, and
contain a lobulated nucleus, a well-developed Golgi complex, smooth and
rough endoplasmic reticula, and many large periodic acid–Schiff-positive
lysosomal granules and phagosomes (62,68). Balazs (69) pointed out that
hyalocytes are located in the region of highest HA concentration and
suggested that these cells are responsible for vitreous HA synthesis. There is
evidence to suggest that hyalocytes maintain ongoing synthesis and
metabolism of glycoproteins within the vitreous (70,71). Hyalocytes have
also been shown to synthesize vitreous collagen (72) and enzymes (73).
FIGURE 3-7 Transmission electron micrograph of human
hyalocyte embedded in the posterior vitreous cortex with
its dense matrix of collagen fibrils. (Original magnification
×11,670.)
The phagocytic capacity of hyalocytes has been described in vivo (74) and
demonstrated in vitro (75,76), consistent with the presence of pinocytic
vesicles and phagosomes (68) and surface receptors that bind
immunoglobulin G and complement (76). Hyalocytes become phagocytic
cells in response to inducting stimuli and inflammation. HA may have a
regulatory effect on hyalocyte phagocytic activity (77,78). As an extension of
this phagocytic activity, hyalocytes can be antigen-presenting cells that
modulate intraocular inflammation while also keeping the vitreous clear via
phagocytic activity. Finally, collagen membranes within hyalocytes are
known to contract in the presence of certain stimulants, such as TGF-β2. This
phenomenon may explain the contraction of preretinal cortical vitreous
responsible for a myriad of pathologies with preretinal membranes, especially
macular pucker (79).
With respect to adult macular pathology, recent studies have illustrated
the role of vitreoschisis and anomalous PVD in macular holes and macular
pucker pathogenesis. In these vitreomaculopathies, hyalocytes likely play an
early, if not primary, role in the contractile premacular membrane resulting
from vitreoschisis, since these cells undergo myofibroblastic
transdifferentiation in the posterior vitreous cortex and have contractile
properties in the presence of TGF-β (80). Indeed, histopathologic evaluation
of the vitreoretinal interface in 14 monkey eyes and in vivo optical imaging in
humans suggest that the premacular vitreous cortex containing hyalocytes
may be important in pathologic premacular membrane formation (81). Using
a murine model, Vagaja demonstrated that under varying physiologic and
pathologic conditions, hyalocytes were responsive to aging, hyperglycemia,
locally produced VEGF, and both systemic and ocular-derived toll-like
receptor (TLR) ligands (82). Similar pathophysiologic mechanisms may be at
play in pediatric vitreoretinopathies.
Vitreoretinal Interface
In the child, it is virtually impossible to mechanically detach the posterior
cortical vitreous from the retina, resulting in surgical complications such as
retinal breaks. The interface between the vitreous and retina consists of a
complex formed by the posterior vitreous cortex and the inner limiting
membrane (ILM), which includes the basal lamina of Müller cells (83,84)
(Figure 3-6) (see also Chapter 4). The ILM is composed of type IV collagen
closely associated with glycoproteins (85–87). Immediately adjacent to the
Müller cells is the lamina rara, which is 0.03 to 0.06 μm thick and
demonstrates no species variations nor changes with topography or age. The
lamina densa is thinnest at the fovea (0.01 to 0.02 μm). It is thicker elsewhere
in the posterior pole (0.5 to 3.2 μm) than at the equator or vitreous base. At
the rim of the optic nerve head, the retinal ILM ceases, although the basement
membrane continues as the ILM of Elschnig (88). This membrane is 50 nm
thick and is believed to be the basal lamina of the astroglia in the optic nerve
head. At the central-most portion of the optic disc, the membrane thins to 20
nm, follows the irregularities of the underlying cells of the optic nerve head,
and is composed only of glycosaminoglycans and no collagen (89). This
structure is known as the central meniscus of Kuhnt. Balazs (7) has stated
that the Müller cell’s basal lamina prevents the passage of cells as well as
molecules larger than 15 to 20 nm. Consequently, the thinness and chemical
composition of the central meniscus of Kuhnt and the membrane of Elschnig
may account for, among other effects, the frequency with which abnormal
cell proliferation arises from or near the optic nerve head (57,59).
Zimmerman and Straatsma (90) claimed that there are fine, fibrillar
attachments between the posterior vitreous cortex and the ILM and proposed
that this was the source for the strong adhesion between the vitreous and
retina. The composition of these fibrillar structures is not known, and their
presence has never been confirmed. It is more likely that an extracellular
matrix “glue” exists between the vitreous and retina in a fascial as opposed to
focal apposition (48,85,86), composed of fibronectin, laminin, and other
extracellular matrix components (91). Western blots of separated proteins
derived from dissected vitreoretinal tissues identified two chondroitin
sulfate–containing proteoglycans of approximately 240-kDa MW that are
believed to function as adhesive molecules at the vitreoretinal interface.
These findings formed the rationale for experimental and clinical studies on
pharmacologic vitreolysis (92) using ABC chondroitinase (see Chapter 4).
Studies of human donor eyes and human retinectomy samples have
identified the distribution of the specific collagen subtypes constituting the
vitreoretinal interface. In particular, immunostaining studies have revealed
collagen type VI in the ILM and type VII in several layers of the retina. Both
collagens can anchor matrix components, and type VI may likely be involved
in vitreoretinal attachment. Furthermore, the presence of collagen mRNA in
human retinectomy samples may be an indication of postnatal collagen
production by retinal cells (93). Such knowledge regarding collagen
distribution may serve useful in understanding the pathophysiology of a
spontaneous, mechanical, or pharmacologically induced PVD (see below).
DEVELOPMENTAL ABNORMALITIES
OF VITREOUS
Many developmental vitreoretinal disorders result from abnormalities in the
embryonic vascular system of the vitreous (VHP) and lens (TVL). These
attain maximum prominence during the 9th week of gestation or 40-mm stage
(94). Hyaloid vessel development around the seventh WG appears to be
associated with high expression of VEGF165 (an isoform of VEGF-A).
However, once the hyaloid vascular system reaches completion, the
alternatively spliced antiangiogenic VEGF165b isoform begins to dominate
(95).
Pathologies of the Primary Vitreous
Persistent Fetal Vasculature

Previously known as persistent hyperplastic primary vitreous (PHPV),
persistent fetal vasculature (PFV) refers to the persistence of the hyaloid
vascular system and subsequent development of primary vitreous pathologies
(96). PFV occurs in 3% of full-term infants but in 95% of premature infants
(97), 90% of infants born earlier than 36 WG, and in over 95% of infants
weighing <5 pounds at birth (98). PFV is believed to arise from abnormal
regression and hyperplasia of the primary vitreous (96) and was initially
described as a plaque-like membrane of retrolental fibrovascular connective
tissue adherent and posterior to the posterior lens capsule. The membrane
extends laterally to attach to the ciliary processes, which are elongated and
displaced centrally. Approximately 90% of cases are unilateral; many of the
fellow eyes may have concurrent anomalous development of the anterior
vitreous (99). Experimental data suggest that the abnormality begins at the
17-mm stage of embryonic development (100). The hyperplastic features
result from proliferation of retinal astrocytes and a separate component of
glial hyperplasia arising from the optic nerve head (101). The fibrous
component of the PFV membrane is presumably synthesized by these cells
(102). A recent case report found that collagen fibrils in this fibrous tissue
had diameters of 40 to 50 nm with a cross-striation periodicity of 65 nm. The
investigators concluded that the collagen fibrils differed from those of the
primary vitreous and suggested that they arose either from a different
population of cells or were the result of abnormal metabolism by the same
cells that synthesize vitreous collagen (103). The retina is usually not
involved in anterior PFV, suggesting that the anterior form is due to a
primary defect in lens development and that vitreous changes are all
secondary (104). Font and investigators (105) demonstrated the presence of
adipose tissue, smooth muscle, and cartilage within the retrolental plaque and
suggested that PFV arises from metaplasia of mesenchymal elements in the
primary vitreous.
PFV is predominantly sporadic and nonheritable. However, various
animal and human studies have suggested the possibility of rare autosomal
recessive, X-linked, and dominant variants (106–108). In cases of autosomal
recessive PFV, linkage analysis revealed a possible candidate gene located on
chromosome 10q11-q21 (106,107). As a result of atypical symptoms, the
diagnosis of PHPV is often delayed. Recently, ultrasonography has been
proposed as a valuable adjunctive diagnostic tool. An ultrasound study in 32
PFV subjects revealed three characteristic sonographic patterns: band, regular
triangle, or inverted triangle shapes. Band-shaped linear echoes extended
from the optic disc to the posterior lens capsule. The inverted triangle echo
was seen as a membranous septum with a narrow base extending from the
optic disc to the posterior lens capsule with a wider anterior portion. A
triangle-shaped echo appeared as a membranous septum with a wide base
extended from the optic disc to the posterior lens capsule with a narrower
anterior portion (109).
A broad spectrum of disorders resulting from PFV includes the
following:
a. Persistent hyaloid artery arises from the posterior aspect of the
retrolental plaque in the affected eye and is often still perfused
with blood. The artery presents as a single vessel that may
(fully or in parts) extend from the optic nerve head to the
anterior point of attachment (Mittendorf dot) at the posterior
lens capsule. In severe forms, there can be microphthalmos with
anterior displacement of the lens–iris diaphragm, shallowing of
the anterior chamber, and secondary glaucoma (110,111).
b. Mittendorf dot is a remnant of the anterior fetal vascular system
located at the former site of anastomosis of the hyaloid artery
and TVL. It typically occurs inferonasal to the posterior pole of
the lens and is not associated with any known dysfunction
(111).
c. Brittle star sign is a multipronged opacity seen when an
enlarging Mittendorf dot anastomoses with residual vessels that
connect the hyaloid remnant to the posterior TVL (111).
d. Bergmeister papilla is the occluded remnant of the posterior
portion of the hyaloid artery with associated glial tissue. It
appears as a gray, linear structure anterior to the optic disc and
adjacent retina. This can cause congenital falciform folds of the
retina and, if severe, can cause tent-like retinal folds, leading on
rare occasions to retinal detachment (102,112). Exaggerated
forms can present as prepapillary veils.
e. Persistent pupillary membranes result from failure of PM
regression and vessel involution during the third trimester (7 to
9 months of gestation). These membranes are attached at the
iris collarette on one side and may extend across the anterior
lens surface as free-floating pigmented strands or with focal
attachments to the anterior capsule of the lens or to the iris on
the opposite side. Thickened fibrocellular iris stroma and
pigmented cells are seen on histopathology (111).
f. Vitreous cysts are generally benign lesions that are found in eyes
with abnormal regression of the anterior (113) or posterior
(114) hyaloid vascular system, otherwise normal eyes
(115,116), and eyes with coexisting ocular disease, such as
retinitis pigmentosa (117) and uveitis (118). Some vitreous
cysts contain remnants of the hyaloid vascular system (119),
supporting the concept that the cysts result from abnormal
regression of these embryonic vessels (120,121). Other studies
have shown pigment epithelial origins of vitreous cysts,
although the specific type of pigment epithelium remains
unclear. Recently, the features of a pigmented vitreous cyst
have been revealed by autofluorescence and anterior segment
optical coherence tomography. In that study, the intact retina,
the absence of lipofuscin in the cyst and its location in the
anterior vitreous led to the proposal that the cyst may originate
from the ciliary pigment epithelium rather than the retinal
pigment epithelium (122). Vitreous cysts are usually not
symptomatic and thus do not require surgical intervention.
However, there is a report describing the use of Nd:YAG laser
to rupture a free-floating posterior vitreous cyst (123). Thus, in
symptomatic patients, treatment options include laser
photocystotomy or pars plana vitrectomy with cyst excision
(124).
Differential Diagnoses
The following conditions can closely resemble vitreous pathologies arising
from persistence of the fetal vasculature leading to a presumed diagnosis of
PFV.
a. Familial (Dominant) Exudative Vitreoretinopathy (FEVR):
FEVR was first described in 1969 by Criswick and Schepens
(125) as a bilateral, slowly progressive abnormality that
resembles retinopathy of prematurity (ROP), but without a
history of prematurity or postnatal oxygen administration (see
also Chapter 66). Gow and Oliver (126) identified this disorder
as an autosomal dominant condition with complete penetrance
caused by failure of peripheral retinal vascularization
(111,126). They characterized the course of this disease in
stages ranging from PVD with snowflake opacities (stage I) to
thickened vitreous membranes and elevated fibrovascular scars
(stage II) and vitreous fibrosis with subretinal and intraretinal
exudates, ultimately developing retinal detachment due to
fibrovascular proliferation arising from neovascularization in
the temporal periphery (stage III). Plager et al. (127) reported
the same findings in four generations of three families but
found X-linked inheritance. van Nouhuys (128–130) studied
101 affected members in 16 Dutch pedigrees and 5 patients
with sporadic manifestations, finding that the incidence of
retinal detachment was 21%, with all but 1 case occurring prior
to the age of 30. These were all tractional or combined traction–
rhegmatogenous detachments, and there were no cases of
exudative retinal detachment. van Nouhuys (128) concluded
that the etiology of FEVR lies in premature arrest of
development in the retinal vasculature, since the earliest
findings in these patients were nonperfusion of the peripheral
temporal retina with stretched retinal blood vessels and
shunting with vascular leakage. Thus, van Nouhuys considers
FEVR as a retinopathy with secondary vitreous involvement.
However, Brockhurst et al. (131) reported that vitreous
membrane formation began just posterior to the ora serrata and
that it preceded retinal vessel abnormalities, suggesting a
vitreous origin to this disorder. Others suggested that there may
be a combined etiology involving anomalies of the hyaloid
vascular system, primary vitreous, and retinovascular
dysgenesis (132). Factors distinguishing FEVR from PFV
include evidence of family history and bilateral, though
sometimes markedly asymmetric, disease (111).
b. Retinopathy of Prematurity:
First described in 1944 (133), ROP remains a leading cause of
blindness in children in the United States (134). Vitreous
liquefaction, often unidentified (135) in stages 1 and 2 ROP,
likely occurs as a result of reactive oxygen species (136) but
also because of underlying undeveloped peripheral retina where
immature Müller cells may not support typical vitreous gel
synthesis and may account for the vitreous trough apparent
during surgery for stage 4 ROP. Further, the abnormal
composition of the malformed peripheral vitreous may limit the
inherent ability to inhibit cell invasion (137–139), thereby
permitting neovascularization in stage 3 ROP to grow (140,141)
anteriorly between posterior gel and peripheral liquid vitreous
(see Chapter 4, Figure 4-4) (141). Instability at the interface
between gel and liquid vitreous exerts traction on the
underlying ridge. As ROP advances from stage 3 to 4, the
neovascular tissue grows through the vitreous body along the
walls of the future Cloquet canal or the tractus hyaloideus of
Eisner toward Wiegert ligament on the posterior lens capsule
(142). Primary vitreous cells responding to intraocular
angiogenic stimuli likely proliferate and migrate to help create
the dense central vitreous stalk and retrolental membrane seen
in the cicatricial stages. ROP can be distinguished from PFV by
its bilaterality and occurrence in premature, low–birth weight
infants exposed to high levels of oxygen supplementation (111).
Pathologies of the Secondary Vitreous
Vitreous Collagen Disorders

As vitreous is one of many connective tissues in the body, it is of interest to
consider parallel phenomena occurring in vitreous and connective tissues
elsewhere, especially as related to collagen. Long ago, Gärtner (143) pointed
out the similarities between the intervertebral disc and the vitreous, in which
age-related changes with herniation of the nucleus pulposus were associated
with presenile vitreous degeneration in 40% of cases. He proposed that a
generalized connective tissue disorder resulted in disc herniation and
presenile vitreous degeneration in these cases. Based on these findings,
Gärtner likened herniation of the nucleus pulposus in the disc to prolapse of
vitreous into the retrocortical (preretinal) space by way of the posterior
vitreous cortex following PVD (Figure 3-4D).
Irene Maumenee (144) identified several different disorders with single-
gene autosomal dominant inheritance in which dysplastic connective tissue
primarily involves joint cartilage. In these conditions, there is associated
vitreous liquefaction, collagen condensation, and vitreous syneresis
(collapse). Since type II collagen is common to cartilage and vitreous,
Maumenee suggested that various arthro-ophthalmopathies might result from
different mutations, perhaps of the same or neighboring genes, on the
chromosome involved with type II collagen metabolism. In these disorders,
probably including such conditions as Wagner disease (145), the fundamental
problem in the posterior segment of the eye is that vitreous is liquefied and
unstable, becoming syneretic at an early age, thus promoting anomalous PVD
(146,147), because there is no dehiscence at the vitreoretinal interface in
concert with liquefaction inside the vitreous body. Thus, in these cases,
abnormal type II collagen metabolism causes destabilization of the vitreous
body and results in traction on the retina that can lead to large posterior tears
and difficult retinal detachments.
Marfan Syndrome
In Marfan syndrome, an autosomal dominant disorder featuring poor
musculature, lax joints, aortic aneurysms, and arachnodactyly, there is lens
subluxation, thin sclera, peripheral fundus pigmentary changes, and vitreous
liquefaction at an early age. Myopia, vitreous syneresis, and abnormal
vitreoretinal adhesions at the equator likely account for the frequency of
rhegmatogenous retinal detachment caused by equatorial or posterior
horseshoe tears (148). Marfan syndrome is associated with mutations in the
FBN1 gene that encodes the connective tissue protein fibrillin 1.
Ehlers-Danlos Syndrome
Systemic manifestations: Ehlers-Danlos syndrome has some similarities to
Marfan syndrome, most notably joint laxity, aortic aneurysms, and an
autosomal dominant pattern of inheritance. However, there are as many as six
types of Ehlers-Danlos patients, and both autosomal dominant and autosomal
recessive forms have been described. A further distinction from Marfan
patients is hyperelastic skin and poor wound healing of all connective tissues,
including the cornea and sclera. Ehlers-Danlos syndrome has been associated
with mutations in numerous collagen genes, including COL1A1, COL1A2,
COL3A1, COL5A1, and COL5A2.
Ocular manifestations: Ocular manifestations include lens subluxation,
angioid streaks, thin sclera, and high myopia due to posterior staphyloma.
Vitreous liquefaction and syneresis occur at a young age. Vitreous traction
causes vitreous hemorrhage, perhaps also due to blood vessel wall fragility,
and retinal tears with rolled edges, often causing bilateral retinal detachments
(148).
Stickler Syndrome
Genetics: In 1965, Stickler et al. described a condition in five generations of a
family that was found to be autosomal dominant with complete penetrance
and variable expressivity (149). Stickler syndrome is the most common type
II/XI collagenopathy, arising from mutations in at least three collagen genes.
Although typically inherited in an autosomal dominant fashion, families with
an autosomal recessive pattern of inheritance have been described (150).
Stickler syndrome is most commonly associated with mutation in COL2A1, a
54-exon–containing gene coding for type II collagen. In one instance
(151,152) of a family with typical Stickler syndrome, posterior chorioretinal
atrophy and vitreoretinal degeneration were found, even though they have
been classically associated with Wagner disease. Vitreous findings from that
study validated reports that mutations in the COL2A1 gene result in an
optically empty vitreous with retrolenticular membrane phenotype.
Systemic manifestations: General features of Stickler syndrome are a
marfanoid skeletal habitus and orofacial and ocular abnormalities.
Subsequent studies identified subgroups with short stature and a Weill-
Marchesani habitus. The skeletal abnormalities now accepted as
characteristic of Stickler syndrome are radiographic evidence of flat
epiphyses, broad metaphyses, and especially spondyloepiphyseal dysplasia
(153). In general, the systemic manifestations can vary greatly, even within a
family all possessing the same genotype. Distinguishing features can include
deafness, cleft palate, joint hypermobility, and premature arthritis (136).
Recently, however, an ocular-only subgroup has been identified, which has
no systemic features but a high risk of rhegmatogenous retinal detachments
(154,155).
Ocular manifestations: Ocular abnormalities are high myopia, >−10
diopters in 72% of cases (156), and vitreoretinal changes characterized by
vitreous liquefaction, fibrillar collagen condensation, quadrantic lamellar
cortical lens opacities (157), and a perivascular lattice-like degeneration in
the peripheral retina believed to be the cause of a high incidence (>50%) of
retinal detachment (153). There is some evidence of phenotype/genotype
correlation, which has led to the classification of Stickler syndrome patients
into five subgroups (157). Additionally, the vitreous can exhibit three distinct
phenotypes, membranous (type 1), beaded (type 2), or normal (158). Patients
with abnormalities in the genes coding for type II procollagen and type XI α1
procollagen are the ones who have severe vitreous abnormalities. Patients
with type XI α2 procollagen defects typically present without ocular
manifestations (159). Another study (160) analyzed the ultrastructural
features of a vitreous membrane with multiple fenestrations in a patient with
a Stickler syndrome. A type 2 vitreous phenotype was found in the left eye,
whereas the fellow eye’s vitreous abnormalities appeared to result from
conversion to a type 1 phenotype. In such a conversion, a fenestrated
membrane may represent the posterior vitreous cortex in a complete PVD.
The fenestrated membrane is made of avascular fibrocellular tissue with cells
arranged cohesively around the fenestration. Proliferating Müller cells and
collagen fibrils were shown to be similar to normal vitreous by ultrastructural
examination. The authors concluded that collagen molecules are not
functionally modified, but they are probably quantitatively insufficient during
vitreous development. Aside from high myopia in childhood, the majority of
patients with Stickler syndrome have no vision loss unless they have a retinal
detachment. In addition to premacular vitreous changes, recent studies
evaluated foveal structure in 25 patients with Stickler syndrome using swept-
source optical coherence tomography, finding mild foveal hypoplasia with
persistence of the inner retinal layer (161).
Stickler syndromes are the most common causes of inherited and
childhood retinal detachment; however, no consensus exists regarding the
effectiveness of prophylactic intervention. Recent studies evaluated the long-
term safety and efficacy of the Cambridge prophylactic cryotherapy protocol,
a standardized retinal prophylactic treatment developed to prevent retinal
detachment arising from giant retinal tears in type 1 Stickler syndrome. In
487 patients with type I Stickler syndrome, the Cambridge prophylactic
cryotherapy protocol was not only safe but also markedly reduced the risk of
retinal detachment (162).
Knobloch Syndrome
Knobloch (163) described an autosomal recessive syndrome similar to
Stickler syndrome with hypotonia, relative muscular hypoplasia, and mild to
moderate spondyloepiphyseal dysplasia causing hyperextensible joints. The
vitreoretinopathy is characterized by vitreous liquefaction, veils of vitreous
collagen condensation, and perivascular lattice-like changes in the peripheral
retina. Retinal detachment in patients with Knobloch syndrome has been
explained by loss-of-function mutations in COL18A1, the gene encoding the
α1-chain of collagen XVIII based on findings from one investigation (164)
demonstrating that collagen XVIII is crucial for anchoring vitreous collagen
fibrils to the ILM. New electrophysiologic studies described previously
unreported clinical features. In particular, a recent study of the detailed
phenotypes and molecular genetic findings in seven affected families
revealed pigment dispersion syndrome and glaucoma in addition to cone–rod
dysfunction on electroretinography. Two patients had normal neuroradiologic
findings, emphasizing that some affected individuals have isolated ocular
disease (165).
Wagner Syndrome
First described in 1938 by Wagner in a Swiss family (166), this syndrome is a
rare, dominantly inherited vitreoretinopathy with near-complete penetrance
(167). Caused by mutation of the versican (VCAN) gene in chromosome 5q
(previously known as CSPG2), the hallmark feature of this syndrome is an
optically empty vitreous body composed of strands, membranes, or veils. It
can be particularly difficult to distinguish Wagner syndrome from Stickler
syndrome, especially the ocular subtype, based solely on ocular phenotype.
However, consideration of the vitreous phenotype can assist with
distinguishing these two vitreoretinopathies (168).
Myopia
It has been proposed (169) that myopia unrelated to the aforementioned
arthro-ophthalmopathies should also be considered a disorder of vitreous
collagen. The anomalous PVD (146,147) that results from extensive
liquefaction of vitreous (myopic vitreopathy) and propensity for retinal
detachment due to peripheral retinal traction and myopic peripheral retinal
degeneration suggest that this postulate deserves closer scrutiny. Indeed,
vitreomacular traction has already been identified as an important component
in the pathogenesis of myopia-related pathologies, including myopic foveal
retinoschisis (170). Recent swept-source OCT studies of PVD in 151 highly
myopic eyes found larger posterior precortical vitreous pockets (actually the
bursa premacularis of Jan Worst) compared with normal eyes. In addition,
highly myopic eyes are likely to develop PVD at younger ages, often
anomalous with the posterior vitreous cortex more frequently remaining
attached to the macula inducing traction (171). Although the exact
mechanism for increased vitreous liquefaction in high myopia is unclear,
several theories have been proposed. Dysfunctional Müller cell activity is an
older hypothesis that derived from studies showing abnormal B wave results
on electroretinograms of highly myopic patients (current thinking is that the
B wave does not arise from Müller cells but perhaps from ON-center bipolar
cells) (172). However, the absence of abnormally thick inner limiting laminae
in myopic patients (173) suggests that Müller cells may not be the culprit.
Another study showed that highly myopic patients with ocular pathology
(macular detachments or macular hole) had increased vitreous and serum
levels of transthyretin (TTR) compared to controls (174). Additionally, the
TTR in the macular detachment cases was abnormally stable suggesting a
misfolded protein. TTR is a homotetrameric protein that functions as a carrier
for both thyroxin and retinol-binding protein and has previously been
implicated in several amyloid-related diseases such as vitreous amyloidosis,
Alzheimer disease, and familial amyloidotic polyneuropathy (174,175).
These results suggest that TTR may be a biomarker of myopic vitreopathy
while also playing a role in the pathophysiology of myopic ocular conditions.
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4
Vitreous Biochemistry and
Pharmacologic Vitreolysis
Sasha Rosen, Matin Khoshnevis and J. Sebag
Throughout history, medical therapeutics have advanced as a result of

increased understanding of disease pathogenesis. With limited knowledge,
little could be done. As knowledge increased, surgical procedures were
historically early therapeutic approaches. Further increases in knowledge saw
surgical treatments replaced by medical (usually pharmacologic) therapies.
Advanced understanding of disease pathogenesis leads to prevention, the
ultimate goal of medicine. Modern vitreoretinal surgery is a paradigm of this
evolution. The end of the last millennium witnessed the development of
revolutionary surgical approaches to treat vitreoretinal diseases, that is,
scleral buckle and vitrectomy. At present, expanding knowledge of the
biochemistry, structure, and pathophysiology of vitreous and the vitreoretinal
interface is enabling drug therapies such as pharmacologic vitreolysis (1,2).
Initially, this approach will be used as an adjunct to facilitate and enhance
surgery. The next implementation phase, which has already begun (3), will
replace surgery with drugs to treat vitreomacular diseases. Ultimately,
pharmacologic vitreolysis will be used to prevent disease in high-risk
individuals.
Vitreoretinal surgery in pediatric patients is among the most technically
challenging of all eye surgeries. This is largely due to the solid gel vitreous
structure in youth and firm vitreoretinal adhesion. It is thus no surprise that
the first attempts to develop pharmacologic vitreolysis were in a pediatric
setting where the objective was to facilitate vitreous separation from the
retina during surgery. It is important, however, to consider that the
biochemistry, molecular organization, and structure of vitreous and the
vitreoretinal interface in youth are not the same as in adults and the elderly.
Thus, the experience and agents employed in adult pharmacologic vitreolysis
may not directly translate to pediatric patients. Furthermore, the specific
abnormalities in pediatric vitreoretinal diseases may require tailoring of
pharmacologic approaches to the idiosyncrasies of each condition (2). Thus,
in vitreous as elsewhere, great care must be taken when extrapolating from
the adult to the diseased child.
This chapter reviews the biochemical composition and organization of
the human vitreous (4–6) and, where information is available, describes
differences between adults and youth. A review of the various agents being
developed for pharmacologic vitreolysis will also be presented. The
significant biochemical changes that occur during regression of the hyaloid
vasculature (second trimester) are presented in Chapter 2.
VITREOUS BIOCHEMISTRY
Vitreous is an extended extracellular matrix composed of 98% water and 2%
structural components, primarily collagens and glycosaminoglycans (GAGs).
Biochemical analyses of 27 patients undergoing vitrectomy found significant
amounts of electrolytes and metals including copper, zinc, selenium, and iron
(7). Although this study offered the distinct advantage of analyzing undiluted
human vitreous samples (as compared to previous studies that utilized animal
models or postmortem human samples), the fact that all subjects were
undergoing vitrectomy for specific vitreoretinal diseases suggests the results
may not be applicable to healthy eyes or other diseases (8). Of note, the study
featured a mix of diabetic and nondiabetic patients. The authors noted that in
the case of the diabetic patients, vitreous glucose more closely correlated with
serum HbA1C than did serum glucose, suggesting that subclinical alterations
in vitreous biochemistry may forewarn a risk for diabetic retinopathy and
cardiovascular events (7). These findings confirm previous studies that found
elevated levels of glucose (9) and advanced glycation end products (10,11) in
human diabetic vitreous, underscoring the importance of considering diabetes
effects upon vitreous independent from effects on the retina (12). Indeed,
studies in diabetic children have identified structural differences from
nondiabetics that likely result from glucose-induced cross-linking of vitreous
collagen (13). Oxidative stress is probably also involved in these structural
changes within the vitreous body as well as changes at the vitreoretinal
interface, where extracellular superoxide dismutase has been shown to be
important (14).
Collagens
Collagen content is highest where vitreous is a gel (15). As shown in Figure
4-1, individual vitreous collagen fibrils are organized as a triple helix of three
alpha chains. The major collagen fibrils are thin, uniform, and heterotypic,
consisting of more than one collagen type. Recent studies of pepsinized
forms of collagen confirm that vitreous contains collagen type II (60% to
75%), type IX (15% to 25%), and a hybrid of types V/XI (10% to 25%)
(4,6,16).
FIGURE 4-1 Schematic diagram of collagen fibril

structure in the human vitreous.
(Reprinted from Bishop PN. Structural macromolecules and supramolecular organization of the
vitreous gel. Prog Retin Eye Res 2000;19:323–344. Copyright © 2000 Elsevier Science Ltd. With
permission.)
Type II Collagen
Type II collagen is a homotrimer composed of three identical alpha chains
designated as (α1 [II])3 that constitutes 75% of the total collagen content in
vitreous. When first synthesized as a procollagen and secreted into the
extracellular space, type II collagen is highly soluble. The activity of N-
proteinase and C-proteinase enzymes reduces solubility and enables type II
collagen molecules to cross-link covalently in a quarter-staggered array.
Within this array are likely to be N-propeptides, which probably extend
outward from the surface of the forming fibril (5). This may influence the
interaction of the collagen fibril with other components of the extracellular
matrix. Studies combining immunolocalization with Western blot analysis of
macromolecules extracted from bovine vitreous collagen fibrils found that the
pN-type IIA procollagen is located on the surface of the vitreous collagen
fibrils (17). The findings that type IIA procollagen propeptides bind growth
factors such as transforming growth factor-β1 and bone morphogenic protein-
2 support the concept that growth factors interact with vitreous fibrils to
promote enough cell migration and proliferation to result in proliferative
vitreoretinal disorders, such as proliferative vitreoretinopathy in adults and
retinopathy of prematurity in infants (18).
Type IX Collagen
Type IX collagen is a heterotrimer that is disulfide bonded with an (α1 [IX]
α2 [IX] α3 [IX]) configuration. It is oriented regularly along the surfaces of
the major collagen fibrils in a “D periodic” distribution, where it is cross-
linked onto the fibril surface. Type IX is a member of the fibrillar-associated
collagens with interrupted triple helixes group of collagens. It contains
collagenous regions described as COL1, COL2, and COL3 interspersed
between noncollagenous regions called NC1, NC2, NC3, and NC4 (19,20). In
vitreous, as opposed to cartilage, the NC4 domain is small and not highly
charged, thus not likely to exhibit extensive interaction with other
extracellular matrix components (21). In vitreous, type IX collagen always
contains a chondroitin sulfate GAG chain (19,20), which is linked covalently
to the α2 (IX) chain at the NC3 domain, enabling the molecule to assume a
proteoglycan form. Electron microscopy of vitreous stained with cationic
dyes visualizes the chondroitin sulfate chains of type IX collagen,
occasionally found distributed along the surface of vitreous collagen fibrils
(22) and often bridged between neighboring collagen fibrils. Duplexing of
GAG chains from adjacent collagen fibrils may result in a “ladder-like”
configuration (23). It has been recently suggested that type IX collagen
modulates the spatial arrangement of collagen fibrils by both bridging
together and spacing apart individual fibrils (16,24).
Type V/XI Collagen

Ten percent of vitreous collagen is a hybrid of types V and XI collagens that
is believed to constitute the central core of the major collagen fibrils of
vitreous (25). Type V/XI is a heterotrimer that contains α1 (XI) and α2 (V) in
two chains, while the nature of the third chain is presently not known (26).
Along with type II collagen, type V/XI is a fibril-forming collagen. While the
interaction of the fibril with other extracellular matrix components is
probably influenced by a retained N-propeptide that protrudes from the
surface of the fibril in cartilage (25), it is not known whether this is the case
in vitreous (6). Wenstrup et al. have demonstrated that type V collagen plays
an essential role in the initiation of collagen fibril formation (27), which
suggests that the hybrid V/IX collagen could play a similar role in vitreous
fibril assembly (16).
Type VI Collagen
Although there are only small amounts of type VI collagen in vitreous, the
ability of this molecule to bind both type II collagen and hyaluronan (HA)
suggests that it could be important in organizing and maintaining the
supramolecular structure of vitreous gel.
Glycosaminoglycans
GAGs do not normally occur as free polymers in vivo but are covalently
linked to a protein core, the ensemble called a proteoglycan. A sulfated group
is attached to oxygen or nitrogen in all GAGs except HA. Studies in the
rabbit (28) found a total vitreous GAG content of 58 ng with 13% chondroitin
sulfate and 0.5% heparan sulfate.
Hyaluronan
Although HA is present throughout the body, it was first isolated from bovine
vitreous (29). HA appears in human vitreous after birth possibly synthesized
by hyalocytes (30), although other plausible candidates are the ciliary body
and retinal Müller cells. HA is synthesized at a constant rate in the adult.
Although there is no extracellular degradation, HA levels are in a steady state
because the molecule escapes via the anterior segment of the eye (31).
HA is a long, unbranched polymer of repeating glucuronic acid β-1,3-
N,N-acetylglucosamine disaccharide moieties linked by β 1 to 4 bonds (32),
with a molecular weight of 3 to 4.5 × 106 in adult human vitreous (31). HA is
a linear, left-handed, threefold helix with a rise per disaccharide on the helix
axis of 0.98 nm (33). This periodicity, however, can vary depending on
whether the helix is in a “compressed” or “extended” configuration (34).
Changes in the degree of “extension” of HA could be important in retinal
disease, since the volume of the unhydrated HA molecule is about 0.66
cm3/g, whereas the hydrated specific volume is 2,000 to 3,000 cm3/g (31).
Thus, the degree of hydration has a significant influence on the size and
configuration of the HA molecular network. HA also interacts with the
surrounding mobile ions and can undergo changes in its conformation that are
induced by changes in the surrounding ionic milieu (35). A decrease in
surrounding ionic strength can cause the anionic charges on the
polysaccharide backbone to repel one another, resulting in an extended
configuration of the macromolecule. An increase in surrounding ionic
strength can cause contraction of the molecule and, in turn, the entire vitreous
body. As a result of HA’s entanglement and immobilization within the
vitreous collagen fibril matrix, this mechanical force can be transmitted to the
retina, optic disc, and other structures, such as neovascular complexes. This
can be important in certain pathologic conditions that feature fluctuations in
ionic balance and hydration, such as diabetes (12), especially type I diabetes
in children who have a solid gel vitreous firmly adherent to the retina.
Recently, covalent HA coating of lipoplexes has been proposed for delivery
of retinal gene nanomedicines via intravitreal injection (36).
Chondroitin Sulfate
Vitreous contains two chondroitin sulfate (CS) proteoglycans. The minor
type is actually type IX collagen, which was described earlier. The majority
of vitreous CS is in the form of versican; concentration = 0.06 mg
protein/mL, about 5% of the total protein content (37). This large
proteoglycan has a globular N-terminus that binds HA via a 45-kDa link
protein (38). In human (but not bovine) vitreous, versican is believed to form
complexes with HA as well as microfibrillar proteins, such as fibulin-1 and
fibulin-2 (6).
Heparan Sulfate
This sulfated proteoglycan is normally found in basement membranes and on
cell surfaces throughout the body. It was first detected in bovine vitreous in
1977 (39) and in chick vitreous (as “agrin”) in 1995 (40). However, it is not
clear whether heparan sulfate is a true component of vitreous or a
contaminant from adjacent basement membranes, such as the inner limiting
membrane of the retina (41). As pointed out by Bishop, this may also be the
case for nodogen-1, the aforementioned fibulins, and fibronectin (6).
Noncollagenous Structural Proteins
Fibrillins
Fibrillin-containing microfibrils are more abundant in vitreous than type VI
collagen microfibrils. They are found in vitreous gel as well as in the zonules
of the lens, explaining why in Marfan syndrome, defects in the gene encoding
fibrillin-1 (FBN1 on chromosome 15q21) result in both ectopia lentis and
vitreous liquefaction (6). The latter probably plays a role in the high
incidence of rhegmatogenous retinal detachment in these patients.
Opticin
The major noncollagenous protein of vitreous is opticin, a leucine-rich repeat
(LRR) protein, which is bound to the surface of the heterotypic collagen
fibrils (42). Formerly called vitrican, opticin is believed to be important in
collagen fibril assembly and in preventing the aggregation of adjacent
collagen fibrils into bundles. Thus, a breakdown in this property or activity
may play a role in age-related vitreous degeneration (43). A recent study
attempted to determine the structure, location, and expression of the mouse
opticin gene (Optc) (44). The gene was found to be localized to mouse
chromosome 1, consisting of seven exons. Additionally, in situ hybridization
revealed that opticin mRNA is localized exclusively to the ciliary body
during development and to the nonpigmented ciliary epithelium of the adult
mouse eye. The researchers concluded that opticin may represent a marker
for the differentiation of ciliary body. Besides regulating vitreous collagen
fibrillogenesis, it may also have other functions as demonstrated by its
continued expression in the adult mouse eye. Indeed, Bishop and colleagues
recently demonstrated that opticin is capable of modulating
neovascularization in the posterior segment. In their study, an opticin
knockout mouse and a wild-type mouse were compared in an oxygen-
induced retinopathy model. Although the knockout mouse initially had
normal vascular development, following exposure to high oxygen conditions,
the knockout model developed significantly more preretinal
neovascularization. Additionally, intravitreal injections of opticin into the
wild-type mouse significantly reduced preretinal neovascularization when
exposed to the oxygen-induced retinopathy conditions (45).
Supramolecular Organization
Bishop has emphasized the importance of understanding what prevents
collagen fibrils from aggregating and by what means the collagen fibrils are
connected to maintain a stable gel structure (6). CS chains of type IX
collagen bridge between adjacent collagen fibrils in a ladder-like
configuration spacing them apart (46). This arrangement might account for
vitreous transparency, in that keeping vitreous collagen fibrils separated
would minimize light scattering and allow unhindered transmission of
photons to the retinal photoreceptors. However, depolymerizing with
chondroitinase does not destroy the gel, suggesting that CS side chains are
not essential for vitreous collagen spacing. Complexed with HA, however,
the CS side chains might space apart the collagen fibrils (23,46), although
Bishop believes that this form of collagen–HA interaction is “very weak.”
Instead, he proposes that the LRR protein opticin is the predominant
structural protein in short-range spacing of collagen fibrils. Concerning long-
range spacing, Scott (23) and Mayne et al. (47) have claimed that HA plays a
pivotal role in stabilizing the vitreous gel via this mechanism. However,
studies (48) using HA lyase to digest vitreous HA demonstrated that the gel
structure was not destroyed, suggesting that HA is not essential for the
maintenance of vitreous gel stability, leading to the proposal that collagen
alone is responsible for the gel state of vitreous (6).
Total collagen content in the vitreous gel remains at about 0.05 mg until
the third decade (2). As collagen concentration does not appreciably increase
during this time but the size of the vitreous increases, the network density of
collagen fibrils effectively decreases, potentially weakening the collagen
network and destabilizing the gel. However, since there is net synthesis of
HA during this time, it likely stabilizes the thinning collagen network (31).
Although previous studies suggested the lack of postnatal synthesis of
vitreous collagen (16,49), Wang et al. (50) demonstrated some postnatal
collagen synthesis, indicating that the formation of vitreous collagen is a
more dynamic than previously thought (16). The loss of type IX collagen
over time in concert with augmented surface exposure of type II collagen
causes disruption in the parallel organization of collagen fibrils as well as
subsequent aggregation (16,51). Bishop highlights the constitutional role of
the heterotypic collagen fibrils in promoting the stability of the vitreous gel
and proposes that future studies should aim to further describe all of the
macromolecules present on the surface of vitreous collagen fibrils and the
nature of their interactions with other macromolecules as well as collagen
fibrils (6).
VITREOUS DEVELOPMENT AND

AGING
The embryonic vitreous body is filled with blood vessels until the end of the
second trimester. The biochemical events associated with regression of the
fetal hyaloid vasculature and pathologic anomalies in that process are
presented in Chapter 3. Postnatal vitreous structure is characterized by a clear
solid gel, as demonstrated in Figure 3-1.
Vitreous structure changes with age (Figure 4-2), likely related to
changes in collagen/proteoglycan composition and organization. Based upon
vitreous pentosidine levels in nondiabetic patients, the average half-life of
vitreous collagen is estimated to be about 15 years, which is similar to that of
skin collagen (16,52). In diabetic patients, vitreous levels of advanced
glycation end products increase dramatically (10,52). In all patients, HA and
CS probably play important roles in the structural changes that occur with
aging (10,53). Studies have shown CS to primarily interact with type IX
collagen, possibly related to collagen aggregation (5,54), while HS plays a
role at the vitreoretinal interface (5,53,55).
FIGURE 4-2 Aging changes in human vitreous structure.
Dark-field slit microscopy of fresh unfixed whole human
vitreous body with the sclera, choroid, and retina dissected
off the vitreous body, which remains attached to the
anterior segment. A slit lamp beam illuminates from the
side, creating a horizontal optical section with an
illumination–observation angle of 90 degrees, maximizing
the Tyndall effect. The anterior segment is below and the
posterior pole is above in all specimens. Top panel: The
vitreous bodies of an 11-year-old girl (left) and a 14-year-
old boy (right) demonstrate a homogeneous structure with
no significant light scattering within the vitreous body,
only at the periphery where the vitreous cortex is
composed of a dense matrix of collagen fibrils. The
posterior aspect of the lens is visible at the bottom of each
image. Middle panel: Vitreous structure in a 56-year-old
(left) and a 59-year-old (right) subject features
macroscopic fibers in the central vitreous body with an
anteroposterior orientation. These form when hyaluronan
molecules no longer separate collagen fibrils, allowing
cross-linking and aggregation of collagen fibrils into
visible fibers. Bottom panel: In old age the fibers of the
central vitreous become significantly thickened and
tortuous, as demonstrated in the two eyes of an 88-year-
old woman. Adjacent to these large fibers are areas of
liquid vitreous, at times forming pockets, called lacunae.
(Reprinted by permission from Sebag J, Koss MJ.
Anomalous PVD & vitreoschisis. In: Sebag J, ed. Vitreous
—in health & disease. New York: Springer, 2014:241–
265.)
Anomalous Posterior Vitreous Detachment
Concurrent vitreous gel liquefaction (synchisis) and weakening of
vitreoretinal adhesion are necessary for innocuous posterior vitreous
detachment (PVD) (56). Traction at the vitreoretinal interface results from
liquefaction with collapse of the vitreous body (syneresis) but persistent
vitreous adhesion to the retina. A variety of abnormalities can result,
collectively known as anomalous PVD (APVD) (57–59).
Full-thickness APVD occurs when the entire axial thickness of the
posterior vitreous cortex remains attached to part of the retina
(topographically) with vitreoretinal separation elsewhere. The resulting
vitreoretinal traction can manifest posteriorly as vitreomacular traction
syndrome, while peripherally this results in retinal tears/detachments. Partial-
thickness APVD occurs when there is a split in the posterior vitreous cortex,
known as vitreoschisis (Figure 4-3), a process important in the
pathophysiology of macular pucker, and perhaps other vitreo-maculopathies
as well (60).
FIGURE 4-3 Vitreoschisis. In vivo SD-OCT imaging of

the human vitreoretinal interface demonstrates a split in
the posterior vitreous cortex, called vitreoschisis. (Image
courtesy of Jay S. Duker, MD.)
Although APVD is more often relevant in vitreoretinal disorders of adult and

elderly humans, a similar pathophysiology can occur in pediatric patients
whenever vitreoretinal instability arises from vitreous gel liquefaction with
persistently strong adhesion at the vitreoretinal interface. This is important in
retinopathy of prematurity and other disorders where peripheral vitreous gel
does not form normally. The instability this produces at the junction of the
peripheral liquid vitreous with the gel posterior vitreous causes traction upon
the retina. In retinopathy of prematurity, this occurs along a ridge (Figure 4-
4) where neovascularization and fibrous proliferation can develop. If traction
is severe, there can be consequent tractional retinal detachment. A major
objective of pediatric vitreoretinal surgery is the release of vitreoretinal
adhesion and relief of traction.
FIGURE 4-4 Photomicrograph of peripheral fundus in

retinopathy of prematurity. The lens is in the upper right-
hand corner. Below, a fibrovascular membrane is present
at the interface between the posterior gel vitreous and the
peripheral liquid vitreous. The inset shows the
histopathology that clearly distinguishes between gel (G)
and liquid (L) vitreous. (Courtesy of Maurice Landers,
MD.)
PHARMACOLOGIC VITREOLYSIS
Surgery to mechanically separate the vitreous and retina has historically been
the definitive treatment for conditions affecting the vitreoretinal interface
(61); however, as with all surgical interventions, there exist inherent risks and
limitations, as well as costs. Thus, pharmacologic vitreolysis with an
intravitreal injection that can be performed in an office setting represents a
paradigm shift in the therapeutic approach to diseases of the vitreoretinal
interface (62). This is a new approach, however, and it is important to note
that of the seven agents that have to date undergone testing for use in
pharmacologic vitreolysis, five have either failed or have been discontinued
and one is still being developed (63,64). Only one drug has been approved for
clinical application.
The term pharmacologic vitreolysis (1,2) refers to the use of drugs to
alter molecular structure and induce vitreous gel liquefaction, weaken
vitreoretinal adhesion, and induce innocuous PVD. Further, pharmacologic
vitreolysis may improve intraocular physiology and metabolism (65). To
date, investigators have used pharmacologic vitreolysis in diseases such as
diabetic retinopathy (66), macular holes (67), retinopathy of prematurity (68),
and congenital retinoschisis (69). Since initial attempts all used enzymes as
adjuncts to surgery, the term “enzymatic vitreolysis” was prevalent in the
early literature (70,71). However, in 1998, the term “pharmacologic
vitreolysis” was proposed (1) so that vitreolytic agents could be grouped
according to their mechanisms of action as either “enzymatic” or
“nonenzymatic” (2). It was further proposed that these agents be
subcategorized as either nonspecific agents, such as tissue plasminogen
activator (72), plasmin (73,74), microplasmin (known now as ocriplasmin)
(75,76), and nattokinase (77), or substrate-specific agents, such as
chondroitinase (30,53,78), dispase (79,80), and hyaluronidase (53,81,82).
Since there are only two nonenzymatic agents (urea/Vitreosolve) (83) (no
longer under development) and RGD peptides (those composed of amino
acids, L-arginine, glycine, and L-aspartic acid) (84), it would seem that an
alternative classification might be more useful, especially if based upon
biologic activity. Thus, pharmacologic vitreolysis agents have recently been
reclassified (Table 4-1) based upon the ability to induce liquefaction
(“liquefactants”) or whether they induce dehiscence at the vitreoretinal
interface (“interfactants”) (2). Of note is that several agents have both
liquefactant and interfactant properties.
TABLE 4-1 Pharmacologic vitreolysis

classification based on biologic activity
Note: tPA (tissue plasminogen activator); plasmin, ocriplasmin, nattokinase, vitreosolve are believed to
be both liquefactants and interfactants.
aFormerly known as microplasmin.
bNonenzymatic agents.
Reprinted with permission from Sebag J. Pharmacologic vitreolysis-premise and promise of the first
decade. Retina 2009;29(7):871–874.
Although most of the following information relates to adult vitreoretinal

diseases, there may be valuable insights for the development of pediatric
pharmacologic vitreolysis.
Developing Pediatric Pharmacologic Vitreolysis

An important consideration in developing pediatric pharmacologic vitreolysis
and defining indications for its use is that when implemented clinically, this
treatment paradigm will be performed upon eyes with abnormal vitreous.
This is especially true for myopia and diabetes but also prematurity and
various inherited vitreoretinopathies. Myopic vitreopathy (85) causes
profound alterations in vitreous. It is also known that diabetes induces
significant biochemical (10,11,86,87) and structural (13) effects upon
vitreous that impact pathologic processes in diabetic vitreopathy (12) and
diabetic vitreoretinopathy (88) (see above). Since diabetic vitreous is so
different from normal vitreous, even in young children (13), studies of the
effects of pharmacologic vitreolysis upon normal vitreous in vitro and in
experimental animal models may fail to develop agents that are effective in
human disease. This may partly explain why hyaluronidase (Vitrase) failed in
phase III FDA clinical trials for treating vitreous hemorrhage in diabetic
retinopathy. No preclinical studies were performed using Vitrase on diabetic
vitreous. Another explanation for the failure of Vitrase relates to the fact that
hyaluronidase is not an interfactant, only a liquefactant (Table 4-1). Thus,
while hyaluronidase will liquefy gel vitreous, it will not induce vitreoretinal
dehiscence. This will result in persistent traction upon neovascularization
with subsequent recurrent vitreous hemorrhage and vision loss.
In contrast to the Vitrase experience, a recent study (2,89) investigated
the effects of pharmacologic vitreolysis in diabetic rats. The results showed
that hyaluronidase alone did not induce PVD in any of the eyes tested,
confirming previous studies (81,82). Plasmin alone induced only a partial
PVD in 10/10 (100%) subjects. This is disconcerting since past studies
showed that a partially detached vitreous carries the worst prognosis for
progressive diabetic retinopathy (90–92). Thus, plasmin (and possibly
ocriplasmin that has a similar enzymatic profile) might actually worsen the
prognosis in diabetes by inducing an anomalous PVD (57–59). While
previous investigations using plasmin (66–69,93,94) and ocriplasmin
(74,75,93) claimed to induce total PVD, those experiments were performed in
nondiabetic vitreous. In the case of tractional diabetic macular edema, a small
case series reports more success following plasmin injection (92). Thus,
further work needs to be undertaken in diabetic subjects using these and other
agents to treat diabetic vitreoretinopathy. This may be an even more
important consideration in pediatric vitreoretinal diseases. It could be
limiting, indeed even dangerous, to assume that the experience with
pharmacologic vitreolysis in adults will directly translate to the pediatric
setting.
The safety profile of any agent for pharmacologic vitreolysis is of the
utmost importance. Many of the agents that have been studied have shown
side effects that may preclude them from becoming effective clinical
therapies. For example, early studies on dispase had promising efficacy
results but also showed retinal hemorrhages, ultrastructural cell damage,
cataract formation, and a significant reduction in electroretinography (ERG)
amplitude (80,94,95). More recent studies have shown that the toxic effects
of dispase on the retina are dose-related and that at lower doses, PVD can still
be achieved without significant side effects (96). Similarly, high-dose
intravitreal nattokinase caused preretinal hemorrhages, and ERG changes but
apparently only at doses higher than the necessary therapeutic range (77).
A final important consideration in the development of pediatric
pharmacologic vitreolysis is the issue of combination therapy, since
combinations of two or more pharmacologic vitreolysis drugs may enable
lowering the dose of any single agent, thereby rendering a more favorable
safety profile (97). In the aforementioned studies on diabetic rats, neither
hyaluronidase nor plasmin alone achieved effective pharmacologic
vitreolysis, yet the combination of the two agents induced a total PVD in 8/10
(80%) eyes. A plausible explanation is that the liquefactant hyaluronidase
induced gel liquefaction, while the interfactant properties of plasmin induced
sufficient dehiscence at the vitreoretinal interface to induce total PVD in a
much higher percentage of cases than has been observed to date in any
clinical human trials. Future protocols should consider combination therapy
as well as outcome measures of ocular physiology and metabolism to test the
hypothesis that pharmacologic vitreolysis can induce salubrious physiologic
effects (98,99).
Indications and Contraindications

The first drug approved for clinical pharmacologic vitreolysis is ocriplasmin,
a nonspecific serine protease that is both a liquefactant and an interfactant. As
such, it is active against essential components of the vitreoretinal interface,
including fibronectin and laminin (100). The FDA approved ocriplasmin for
the treatment of symptomatic vitreomacular adhesion in October 2012. The
EU approved ocriplasmin for the treatment of vitreomacular traction and
macular holes in February 2013. The basis for these approvals were clinical
trials that showed ocriplasmin relieving vitreomacular adhesion in 26.5%
versus 10.1% in placebo controls (3,101). Closure of small to medium
macular holes occurred in 40.6% versus 10.6% (101). While BCVA
improvement was noted in patients with successful macular hole closure after
receiving ocriplasmin, BCVA of some patients who did not achieve hole
closure with ocriplasmin still improved. This may be due to resolution of
vitreomacular adhesion, which was noted in half of the subjects.
Analysis of initial postapproval experience using ocriplasmin in clinical
practice has shown success rates of up to 78% (102). The patients best suited
for ocriplasmin pharmacologic vitreolysis are younger than 65 years old, with
no previous eye surgeries (including cataract), no diabetic retinopathy, no
premacular membrane with pucker, vitreomacular adhesion limited to <1,500
μm, and full-thickness macular hole <250 μm in diameter (103). Relative
indications include macular hole diameter 250 to 400 μm, female sex, and
specific OCT findings (i.e., small area of adhesion, “V-shaped” VMT with
wide angles). Relative contraindications include previous vitrectomy, laser or
surgery within prior 3 months, intravitreal injection within prior 6 months,
fibrous proliferation within the ILM, premacular membrane with pucker
(104), history of rhegmatogenous detachment, myopia, proliferative
vitreoretinopathy, macular hole diameter >400 μm, and severe peripheral
retinal degeneration (103).
Jackson et al. (105) also sought to identify factors that predict a favorable
response to ocriplasmin in patients with symptomatic vitreomacular adhesion
and reported best efficacy in younger patients with focal vitreomacular
adhesion and without a premacular membrane with pucker, confirming
previous studies. These findings were again confirmed in studies by Feng et
al. (106). However, in patients with full-thickness macular holes, closure
correlated more closely with hole diameter than the presence or absence of
positive prognostic factors.
Sadda et al. (107) investigated the effect of ocriplasmin on
microperimertry parameters and found that fixation and sensitivity
parameters were better with ocriplasmin compared to sham treatment;
however, their study was limited by a small sample size (N = 27 with 6/27
patients discontinued) and skewed results due to 12/19 patients in the
ocriplasmin group with vitreomacular adhesion resolution following
treatment. The authors argue that larger studies are needed to validate
microperimetry as a marker for vitreomacular adhesion improvement or
resolution (108). Functional improvement beyond visual acuity has also been
demonstrated using 3D Contrast Amsler grid testing (109).
Stalmans et al. note that noninvasive high-resolution OCT may be useful
for studying the vitreomacular interface in patients with vitreomacular
traction or macular hole with focal vitreomacular adhesions, both indications
for pharmacologic vitreolysis, in both diagnosis and management (110).
Whereas Warrow reports similar efficacy of ocriplasmin for symptomatic
vitreomacular traction in the private practice setting as studies conducted in
academic centers (111), he advocates for careful postinjection monitoring via
spectral-domain OCT due to the occasional transient toxicity noted to affect
the outer retina.
Recently, Novack et al. (104) reported on the safety of intravitreal
ocriplasmin for focal vitreomacular adhesions in patients with exudative age-
related macular degeneration (AMD). In a randomized, sham-injection
controlled, double-masked, multicenter, phase II trial (N = 100),
posttreatment safety and tolerability were assessed on posttreatment days 7,
14, and 28 and months 3, 6, and 12. The authors report similar safety for
patients with vitreomacular adhesion and exudative AMD compared to the
established ocriplasmin safety profile, with most adverse effects occurring
within the first week of treatment. Ocriplasmin was not only well-tolerated in
this study population but also reduced the need for intravitreal anti-VEGF
injections, thereby enhancing efficacy and further promoting long-term
safety.
Additionally, ocriplasmin has been shown to be effective in resolving
experimental retinal vein occlusion if injected via a micromanipulator in
close proximity to the site of occlusion (112).
Adverse Events
Ocriplasmin’s enzymatic activity is nonspecific and may dissolve vitreous
proteins and possibly interact with proteins within the retina, choroid, and
lens (100). However, intravitreal injection of ocriplasmin has not been shown
to alter the biomechanical properties of the human inner limiting membrane
by atomic force microscopy, and there is no evidence to suggest any
enzymatic effect on basement membranes (113). Adverse effects of
intravitreal ocriplasmin are almost exclusively transient with recovered visual
acuity. Known complications include subfoveal lucency, ellipsoid zone
disruption, and full-thickness macular hole base enlargement (106). Of these,
only the latter two have been linked to visual decline. Other adverse effects
that are possibly related include acute reduction in visual acuity, ERG
changes, dyschromatopsia, retinal tear or detachment, lens subluxation or
phacodonesis, abnormal pupillary reflex, retinal vascular changes, and OCT
ellipsoid zone alterations (114). In one recently reported case of ocriplasmin-
induced PVD, the patient experienced new clinically significant floaters and
acutely reduced contrast sensitivity function, both of which resolved with
limited vitrectomy (115).
Although most of the reported cases of ocriplasmin-induced acute visual
dysfunction are transient, some patients may not achieve complete resolution
(116–118). According to a report cited in an FDA Advisory Committee
briefing document, nine patients experienced a drastic and abrupt visual
acuity impairment as well as dyschromatopsia and electroretinographic
changes; however, most of those patients had fully recovered vision after a
median time of 2 weeks (118). It has been proposed that the mechanism for
these changes may involve transient traction during vitreomacular separation.
Moreover, a retrospective case series by Singh et al. that analyzed anatomic
visual outcomes of patients treated with ocriplasmin found that the mean time
to ellipsoid loss and the time to return of the ellipsoid zone was 5 days and
29.3 days, respectively, and all of the patients’ inner and outer segment
structures fully recovered (119,120).
In a postapproval retrospective review conducted by the American
Society of Retina Specialists Therapeutic Surveillance Committee, aggregate
data through July 16, 2013, were analyzed to characterize adverse events
associated with ocriplasmin for symptomatic vitreomacular adhesion, and
with each of those adverse events, postapproval safety data were compared to
that of preapproval studies. The study included 999 preapproval injections
and 4,387 postapproval injections. The categories of adverse events included
acute reduction in visual acuity attributable to either worsening of macular
pathology or development of subretinal fluid, electroretinogram changes,
dyschromatopsia, retinal tears and detachments, lens subluxation or
phacodonesis, impaired pupillary reflex, and retinal vessel findings. Ellipsoid
zone findings on OCT were only noted in postapproval data. The study found
that adverse events were typically transient with similar incidences in the pre-
and postapproval groups, though the committee warns of potential
underreporting of postapproval adverse effects (121).
OASIS (122,123) is a 2-year, phase IIIb study of 220 patients that
employed ERG and microperimetry to evaluate untoward effects on visual
function and retinal electrophysiology. All participants underwent spectral-
domain OCT (SD-OCT) imaging, allowing for prospective evaluation of
structural effects. The results showed significant resolution of vitreomacular
adhesion at day 28 in the ocriplasmin group compared to the sham group
(41.7% vs. 6.2%, respectively). In the group treated with ocriplasmin,
presence of focal vitreomacular adhesion or full-thickness macular hole,
absence of premacular membrane with pucker, and pseudophakia were
associated with increased likelihood of successful release of vitreomacular
adhesion. This trial found ocriplasmin to be both safe and effective for long-
term resolution of symptomatic vitreomacular adhesion with improved
outcomes, and no new safety signals were identified when compared to
previous phase 3 trials. The effect of ocriplasmin treatment at 6 months was
sustained at 12 and 24 months.
ORBIT and INJECT are prospective, multicenter studies that will provide
safety and efficacy data for ocriplasmin use in routine clinical settings (124).
They will survey the type, frequency, severity, and treatment relationship of
adverse events during the 12-month study period, mean changes in BCVA,
anatomic changes detected by SD-OCT, and the incidence of vitrectomy or
other ocular procedures. In a preliminary report outlining demographic data
collected over the first 6 months of the ORBIT study, preinjection ocular
symptoms were noted in >3% of study participants, including decreased
visual acuity, metamorphopsia, floaters, central visual field defect/central
black spot, difficulty reading at close distance, photopsia, and eye pain/ocular
discomfort (125).
In a multicenter, retrospective study of persistent macular hole after
ocriplasmin injection that underwent vitrectomy, there were no functional or
anatomical differences when compared to eyes with persistent macular hole
without preoperative pharmacologic vitreolysis (126).
OZONE (127) is a retrospective, postapproval review of 200 patients
treated with ocriplasmin for vitreomacular adhesion imaged with SD-OCT.
This study will collect data on clinical characteristics, the treatment course,
and anatomic changes over the initial 6 months posttreatment with the aim of
characterizing anatomic and symptomatic outcomes after ocriplasmin
injection, including ellipsoid zone changes, subretinal fluid, dyschromatopsia,
electroretinographic changes, and vascular changes.
ACKNOWLEDGMENTS
Sam Asanad of UCLA provided excellent assistance with the biochemistry
section.
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5
Retinal Vascular Development
Michelle E. LeBlanc, Jinling Yang, and Patricia A. D’Amore
PATTERNS OF RETINAL VASCULAR

DEVELOPMENT
Vasculogenesis and Angiogenesis
Vessel formation takes place through the processes of vasculogenesis and
angiogenesis. Vasculogenesis is de novo development of vasculature that
involves the proliferation, differentiation, and organization of blood vessels
from endothelial cell precursors, angioblasts. Angiogenesis involves
migration and proliferation of endothelial cells from an existing blood vessel,
thus sprouting and formation of a new blood vessel. Although the
developmental processes are well documented, the mechanism of the
development of the primary retinal vasculature remains controversial.
The evidence for vasculogenesis is based on nonspecific Nissl staining,
ADPase, and lectin staining that reveal a population of spindle-shaped cells at
the leading edge of developing retinal vessels; these are considered to be
“angioblasts.” These cells are positive for CD39 (ecto-ADPase) and CXCR4
(1,2) but not for other typical markers of endothelial precursors such as
CD31, CD34, NADPH diaphorase, and VEGFR1/2 (3–6). CD39+ angioblasts
co-express CXCR4 in the nerve fiber layer (NFL) near the optic nerve. The
ligand for CXCR4 is stromal-derived factor-1 (SDF-1), which is a hypoxia
inducible factor shown to be expressed in the inner retina at 5 to 7 weeks of
gestation. SDF-1 is believed to act as a chemoattractant to guide vascular
development from the neuroblastic layer to the inner retina (7).
On the other hand, spindle-shaped proliferating astrocytes
(Ki67+/GFAP+) have been identified in advance of developing human retinal
vessels (8). In addition, close association between Pax2+/GFAP− astrocyte
precursors and Pax2+/GFAP+ immature retinal astrocytes with developing
vessels (9) suggests the critical role of astrocytes in vessel growth guidance.
Further support for angiogenesis in the development of the primary
retinal vasculature development comes from mouse models. The expression
of PDGF receptor alpha, a marker of astrocytes, was observed in spindle-
shaped cells. Moreover, transgenic mice expressing GFP under the control of
the glial fibrillary acidic protein (GFAP) promoter reveal extensive astrocyte
invasion into the retina before retinal vascular development. This controversy
is due, in part, to the lack of more specific markers for retinal endothelial
precursors. Furthermore, it is quite likely that the mechanism by which the
superficial retinal vasculature forms varies among species. While a definitive
answer awaits for further studies, it is well accepted that the inner plexuses
are formed via angiogenesis.
Human Retinal Vascular Development

The retina is a highly organized organ that is responsible for receiving and
transporting visual signals to the brain. While the choroidal and vitreous
vasculatures begin development quite early—at 5 to 7 weeks of gestation—
the retinal vasculature develops much later around 16 to 20 weeks of
gestation (see also Chapters 2 and 6). Vascular development in the human
eye is regulated by the oxygen demand of the developing tissues. Coincident
with the regression of hyaloid vessels, the development of the retinal
vasculature provides nutrition and oxygen as well as removes metabolic
wastes from the inner neural retina. It also has a highly ordered architecture
that meets the anatomical needs of neural retina function. When abnormal
retinal development occurs, it leads to visual repair in infants.
Retinal vessels are composed of two laminar but interconnected layers:
the primary superficial layer is located at the interface of the NFL and the
ganglion cell layer (GCL), whereas the deeper layers lie at the border of the
inner nuclear layer (INL); the vessels at the border of INL and inner
plexiform layer (IPL) are the superficial INL plexus (SINL), and those at the
border of INL and outer plexiform layer (OPL) are referred to as the deep
INL plexuses (DINL) (2,5,10,11).
In the human retina, the vitreous and primitive choroidal capillary vessel
develop by hemo-vasculogenesis, which is the development of blood vessels
from hemangioblast progenitors, between 5 and 7 weeks of gestation (WG)
(7). The primary superficial vessels arise from the roots of hyaloid vessels
where optic nerve fibers extend into the retina around 12 to 14 WG.
Rudimentary vascular structures can be observed at 18 WG; however, Ki67+
cells at this time are largely astrocytes and not endothelial cells. At 18 to 20
WG, the vessels form a “butterfly-shaped” plexus that consists of four lobes
representing the territories of the four arteries (2,5,10,11), while the macula
remains undeveloped at this time (7). The primary vessels radially grow
toward the ora serrata shortly before birth. Around 24 WG, the vessels that
are sprouting from the primary layer penetrate the INL and give rise to SINL
and DINL. The branching of these two deep layers also occurs in a “central-
to-peripheral” fashion but extends less peripherally than the primary
plexuses. The outer nuclear layer (ONL) and photoreceptor layer as well as
foveal avascular zone (FAZ) remain avascular and receive nutrition and
oxygen primarily from the choriocapillaris (2,5,10,11).
Whereas human samples are very limited, mouse tissues, on the other
hand, are easily accessible. The mouse retina is vascularized postnatally, and
many aspects of its development recapitulate that of human retinal
vascularization. Moreover, the ability for gene manipulation in the mouse
makes it an extremely useful and informative model to study retinal vascular
development and pathology. Nonetheless, there are key differences between
murine and primate retinal vasculature such as the presence of a fovea and
should be kept in mind when testing angiogenic therapies in murine models
for applications to patients.
The Fovea
One of the unique features of primate retina is the fovea, a structural
depression in the retina temporal to the optic disc (10,11). Although the fovea
begins to form at 22 WG, this structure is not complete until 15 to 45 months
postpartum (12). The fovea is composed of high-density cone photoreceptors
and is specialized for high-resolution vision. In addition, the retinal
vasculature is absent from this specialized area so that there are no vascular
structures present to deflect light and no blood cells moving across the fine
grain foveal cone mosaic. During retinal vascular development, radially
growing vessels appear to avoid this area, also termed as FAZ (10,11). The
avascular properties of the FAZ are attributed to high levels of antiangiogenic
factors including pigment epithelium-derived factor (PEDF) and brain
natriuretic peptide precursor B (NPPB) (13,14). The FAZ forms a complete
circle and measures 500 μm at 35 WG; however, this size is reduced to 170
μm at 37 to 41 WG (15) (see also Chapter 7).
Mouse Retinal Development

At birth, the mouse retina is avascular. The primary plexus grows radially
from the optic nerve head to the periphery right after birth (Figure 5-1). The
retinal vasculature begins to grow along the GCL to form the superficial
vascular plexus but does not grow into the neuronal retina, where neurons
and neuroprogenitor cells are localized (16). At postnatal day 4 (P4), the
deeper layers start to form by sprouting from the superficial layer to form a
polygonal pattern. Around P7, the superficial layer extends close to the
periphery, and by P14, both superficial layer and deeper layers are fully
vascularized. Finally, remodeling of the capillary plexus leads to the
formation of a mature vascular network, which is finalized approximately 3
weeks after birth (16–18).
FIGURE 5-1 Illustration of the relationships among

VEGF expression, astrocyte distribution, and retinal
vasculature in the developing retina. Astrocytes (in green)
migrate from the optic never head to the superficial layer
of the retina. VEGF (in orange) is expressed
correspondingly at the front of astrocyte migration on the
superficial layer and vessel growth (in red) follows. By the
time astrocytes have reached the edge of the superficial
layer, VEGF expression is apparent in the INL close to the
optic nerve head and vessels descended from the
superficial layer form the deep vascular plexus. As VEGF
expression radially migrates, the deeper vasculature layer
develops gradually to the peripheral. Once the vessels are
patent and oxygen is being delivered, levels of VEGF
diminish.
Because this entire process can be visualized using flat mounts, the mouse is
frequently employed for the study of the cellular and molecular regulation of
retinal vascularization. The postnatal vascularization of the mouse retina has
also led to the development of a widely used model of oxygen-induced
neovascularization, with aspects that are reminiscent of the retinopathy of
prematurity as well as proliferative diabetic retinopathy.
Blood–Retinal Barrier Development and Maintenance

In humans, the development of the blood–retinal barrier (BRB) is completed
by midgestation, whereas in mice, BRB formation begins on P7 (19) and is
completed by P10 (20). The BRB is composed of an inner and outer layer.
The inner layer consists of an endothelial layer, which lines the
microvasculature and the neuronal retina consisting of pericytes and glial
cells (21). The inner BRB provides nutrients to the inner two-thirds of the
retina (22) and regulates the transport of molecules across the retinal
capillaries (23). The mature retinal vascular network tightly regulates the
inner BRB to allow for the delivery of nutrition and oxygen for neural cells
and ensures proper visual signal transduction between neural cells (24).
Establishment of a BRB requires is a twofold process, which requires the
formation of tight junction proteins as well as regulation of bulk transcytosis
in order to regulate the influx and efflux of retinal molecules. Prior to BRB
formation, the retinal vasculature is highly permeable and is characterized by
the leakage of proteins and molecules (25). Formation of an intact barrier in
the mouse retina was elegantly examined through transcardiac injection of
dextran tracers of varying molecular weights. An injection of dextran
conjugated dyes at P1 showed diffuse dye leakage from the budding vessel
fronts, confirming that newly formed vessels do not have intrinsic barrier
properties. By P8 and P9, only nascent distal but not mature proximal vessels
exhibited barrier leakage. At P10, all tracers were confirmed within the retina
vasculature, denotating the time of an intact BRB, which took place in a
proximal to distal progression (20). Following similar time points, studies
have shown a correlation between the leakage of dextran conjugated dyes
with an increase in transcytosis, measured by an increase in HRP-filled
vesicles. At P8, when only nascent vessels showed dextran-conjugated dye
leakage, there was also a significant number of HRP-filled vesicles,
suggesting a high rate of endothelial transcytosis. By P10, when the
endothelial cells can regulate vessel permeability, there was little detectable
HRP-filled vesicles, suggesting that a proper BRB is completed only after the
endothelium regulates transcytosis (20).
Blood–Retinal Barrier and the Neurovascular Unit

Disruptions of the neurovascular unit can lead to impaired BRB integrity
(25). The neurovascular unit is essential for proper vascular development,
and it consists of ECs interconnected by pericytes and astrocytes (26). The
importance of the neurovascular unit for the developing BRB is highlighted
by studies demonstrating that ECs lacking pericyte coverage have increased
permeability (27). Although pericytes play an essential role in both the
blood–brain barrier (BBB) as well as the BRB, in the retina, the ratio of
pericytes to ECs is three times that of the BBB, suggesting an important role
of pericytes in regulating the BRB (28). Astrocytes have been shown to guide
the growth of new retinal vasculature during retinal development (29).
Similarly, Van der Wijk et al. (30) has shown that GFAP-positive astrocytes
migrate in front of the growing retinal vasculature during the first postnatal
week, suggesting that a functional neurovascular unit precedes vessel
development and that a functional BRB is complete only after assembly of
the neurovascular unit (25).
Molecular Mediators of the Blood–Retinal Barrier

An intact BRB must prevent leakage while also regulating selective
transcytosis of molecules across the barrier (20). Breakdown of this barrier is
a defining characteristic of many vascular conditions such as diabetic
retinopathy and neurodegenerative diseases of the central nervous system
(CNS) (31). The integrity of the BRB/BBB has presented a challenge for
effective drug delivery highlighted by the fact that systemic drug
administration for ocular disease is not an effective route of administration
due to low drug penetrance. From a therapeutic perspective, understanding
the molecular mechanisms of the BRB may provide insight for novel drug
delivery approaches (32). Accordingly, efforts have been made to identify
molecular regulators, BRB formation, and maintenance during development
and into adulthood.
Mfsd2a
Mfsd2a regulates transcytosis during the BRB formation. Mfsd2a−/− mice
showed an increase in HRP-filled vesicles without a change in vessel
permeability, implicating a role for Mfsd2a in suppressing transcytosis.
Accordingly, Mfsd2a was expressed in proximal vessels with intact tight
junctions but not distal leaky vessels, while at the same time, claudin-5 a tight
junction protein was expressed in the distal vessels. Mfsd2a localization and
phenotype observed in Mfsd2a−/− mice suggest a role for this protein in
regulating the formation of endothelial transcytosis and thus a functional
BRB (20).
Wnt Signaling Pathway

The Wnt/wingless pathway is one of the major regulators of brain and retina
development. In the eye, Wnt interacts with ligands including Norrin/Frizzled
(Fz4), and its activation stabilizes β-catenin, which translocates to the nucleus
and regulates gene expression. Wnt pathways can be regulated through
modulating ligand availability during development. Norrin overexpression or
inhibition has been shown to result in enhanced or inhibited vessel formation,
respectively. Familial exudative vitreoretinopathy, which displays delayed
retinal vascularization, has been shown to be the results of mutations in the
Wnt signaling pathway, specifically low-density lipoprotein receptor-related
protein 5 (LRP5) gene (for review, see (33)). Interestingly, using a mouse
model of LRP5 deletion, the activation of Wnt signaling by lithium has been
shown to rescue the retinal vascular development (34).
Norrin activation of β-catenin requires Tspan12, a member of the
tetraspanin family of surface proteins that include four transmembrane
domains and two extracellular domains. Antibody blockade of Tspan12
interfered with murine retinal vascular development as well as pathologic
vessel growth (35). Fzd7 is also involved in retinal vascularization as
endothelial cell deletion of fzd7 caused delayed formation of the initial
capillary plexus due to dysfunctional tip cell formation and reduced
proliferation of stalk cells (36).
Activation of Wnt pathways contribute to BRB formation by regulating
expression of tight junction genes such as Cladn3 and Glut-1 (37). In the
brain, treatment with Wnt3a or stabilization of β-catenin is shown to increase
the expression of Cladn3, a tight junction protein expressed in ECs, whereas
inhibition of Wnt activity decreased Cldn3 expression (37). Canonical Wnt/β-
catenin activation, specifically Wnt isoforms 7a and 7b (Wnt7a, Wnt7b),
appear to be only required for CNS endothelium angiogenesis. This finding
was based on the observation that Wnt inhibition, during embryonic
development, resulted in vascular malformations exclusively in the CNS
endothelium. Double knockout of Wnt7a:Wnt7b in mice resulted in ventral
vascular malformation with thickening of the vascular plexus during
embryonic development at E10.5, E12.5 (37).
Although the Wnt pathway is essential for proper developmental of the
retinal vasculature, it also has roles in the adult retina. Loss of Norrin/Fz4
activity has been shown to decrease BRB integrity, indicating a requirement
for this pathway in maintaining barrier integrity and vessel plasticity beyond
vascular development (38).
Hedgehog Signaling Pathways

Hedgehog signaling is involved in embryonic morphogenesis, neuronal
guidance, and angiogenesis (39), as well as early vascular development (40).
Hedgehog signaling is activated following the binding of diffusible ligands
such as sonic hedgehog (Shh), desert hedgehog, and Indian hedgehog to co-
receptors smoothened (Smo) and patched. Activation of smoothened triggers
nuclear translocation of transcription factors such as Gli to promote the
expression of target genes (26,41). Shh drives vascular development and
arterial identity in zebrafish (42) and mice (43). Shh increases the expression
of proangiogenic factors such as vascular endothelial growth factor (VEGF)
during coronary (44) and pulmonary vascular development (45), suggesting
that Shh modulates EC barrier properties indirectly by regulating VEGF and
Notch activity (43).
Shh/smoothened activation modulates junction proteins involved in
barrier integrity (26,41). In the presence of VEGF, Shh/Smo activation
increases VEGF-mediated EC permeability, measured by a decrease in the
expression of claudin-5, occludin, and ZO-1 in primary bovine retinal ECs
(26). This work suggests that inhibition of Shh/Smo activity promotes barrier
dysfunction in the absence of pericyte or glial cell contribution, defining a
cell autonomous role for Shh/Smo activity (26).
PDGF-B
It has been shown that pericyte-EC interactions and PDGF-B/PDGFRB
signaling was not essential for BRB integrity in quiescent adult ECs but was
required for the formation of the BRB in retinal development (22).
Retinoic Acid
Retinoic acid (RA) is a metabolite of vitamin A that binds to the CRALBP-2
and signals through 3 nuclear RA receptors and scaffolding retinoid X
receptors. RA is essential for BRB development and maintenance; inhibition
of RA synthesis and RA binding to its receptor resulted in the breakdown of
the BRB in zebrafish larvae and adults (21). It remains unclear if RA acts
directly on ECs to regulate BRB or acts indirectly through neighboring
neuronal cells (21). RA is synthetized through aldehyde dehydrogenase 1
family member A1 (Aldh1a1). Studies have shown that loss of Aldh1a1
results in decreased choroid vasculature due to lower VEGF levels and
decreased Sox9 expression. The effects of Aldh1a1 could be rescued by
overexpression of RA, confirming that the reduction in vasculature and
decreased VEGF levels were due to lower levels of RA. RA synthesized by
Aldh1a1 in the neuronal retina stimulates the RPE to secrete VEGF. This
links RA levels with VEGF expression and proper vessel development (46).
Other Cell Types in Vascular Development

The microvasculature in the retina is composed of the endothelial cell tube,
the abluminal pericyte, and associated astrocyte processes (Figure 5-2).
FIGURE 5-2 Schematic diagram of the cell types

associated with retinal blood vessels. A monolayer of
vascular endothelial cells (in white) forms the innermost
layer of retinal blood vessels and is the primary site of the
blood–retinal barrier. Note the branching of the mature
vessel to form a new vessel, also called angiogenesis.
Pericytes (in light grey) ensheath the endothelial layer of
the newly formed vessel, maintain endothelial cell
quiescence, facilitate blood–retinal barrier formation, and
regulate blood flow. Astrocytes (in dark grey) are star-
shaped cells associated with retinal blood vessels on the
superficial layer and contribute to endothelial tight
junction formation.
Pericytes
Following the formation of the early capillary plexus by the endothelium,
mural cells are recruited and mediate the process of vessel maturation. Mural
cells, the collective term for smooth muscle cells and pericytes, surround and
enclose the newly formed vessels. Mural cells associated with arterioles and
venules have a higher expression of α-smooth muscle actin than pericytes and
enable the contractile feature of those cells. Pericytes, the mural cells that are
associated with capillaries, express lower levels of α-smooth muscle actin,
but higher desmin (an intermediate filament protein) than smooth muscle
cells. However, their abluminal location on the vessel wall is consistent with
a role for the pericyte in regulating blood flow (6,47).
In addition to regulating blood flow, mural cells maintain endothelial cell
quiescence by inhibiting their proliferation and migration and facilitating
formation of tight junctions between endothelial cells (11,48–50). At the
front of newly forming capillary beds, mural cell investment is low, leaving
those endothelial cells vulnerable to regression, growth factor stimulation,
and remodeling (6). Moreover, loss of pericytes leads to instability of mature
vessels (51,52). Retinal capillaries have higher pericyte coverage than brain
capillaries, suggesting stricter regulation of the retinal vasculature compared
to brain capillaries (53).
Astrocytes
In the retina, there are two types of neuroglia cells: astrocytes and Müller
cells. The association of neuroglia cells with retinal vascular development has
been extensively studied. In humans, staining of the developing retina for
GFAP reveals a population of cells extending toward peripheral avascular
areas several hundred microns ahead of the vascular front. Those astrocytes
proliferate and migrate from the optic nerve, lie in front of the developing
primary vascular plexus, and act as a template for retinal vessel growth (8). In
the mouse retina, astrocyte precursors migrate from the optic nerve head and
distribute in the retinal whole mount before retinal vascular development, but
in human, they migrate into the retina during retinal vascular development
(8,18). As the retinal vessels begin to develop, they differentiate radially from
optic nerve to the periphery through retinal ganglion cell–derived PDGFα and
guide vessel growth (54). A scaffold of astrocytes with low GFAP expression
lies under the mature retinal vessels and is thought to facilitate the formation
of endothelial tight junction (55–57).
During mouse vascular development, astrocytes guide the growth of
VEGFR2+ endothelial tip and stalk cells through secretion of a tightly
regulated VEGF gradient (58). Interestingly, pharmacological or genetic
depletion of astrocytes does not cause abnormal vessel growth into the
neuroretina, suggesting an alternative mechanism for directing retinal vessel
growth away from the neuroretina (54,59). Recent work has shown that
neuronal expression of VEGFR2 is higher than receptor expression on ECs.
These high levels of VEGFR2 serve to bind and endocytose VEGF ligand,
thus limiting its availability to the vasculature. In this way, neurons titrate
VEGF expression in the retina during development (16).
Similar to the absence of retinal vasculature in the FAZ, there is also no
astrocyte invasion into this area during development, further supporting the
critical role of astrocytes in retinal vascular development (8). Müller cells are
located in the INL and guide the formation and maturation of the deeper
vessel layers (60).
Microglia
Microglia migrate into the retina along with the developing vessels but are
positioned behind the vascular front in human retina at approximately 14 WG
(56). Distinct from the subpopulation of microglial cells that invade the retina
before vascular development around 10 WG, these cells express the
macrophage marker S22, in addition to microglial markers CD45, MHC I,
and II and remain in close contact with vessels during development as well as
after they have matured. In spite of the documentation of their presence, little
is known about the role that microglia might play in vascular development
(61,62). Disruption of SDF-1/CXCR4 signaling during murine retinal
vascularization resulted in a reduced number of microglia along with
impaired tip cell development, leading the authors to speculate regarding a
role for microglia in tip cell activation (63). Induction of systemic
inflammation in P4 mouse pups by lipopolysaccharide (LPS) led to a
significant increase in retinal vascular density that was preceded by elevated
numbers of microglia in the RLC and OPL layers (64). In adulthood, the
retinas of these exhibited decreased vascular density and impaired retinal
function.
HYPOXIC AND RETINAL VASCULAR

DEVELOPMENT
Physiologic Hypoxia
Regulation of oxygen homeostasis is essential for proper vascular formation
(65), and the role of hypoxia in retinal vascular development is considered a
characteristic of normal vessel development.
1. A retinal vasculature develops in retinas whose thickness is outside the

range of oxygen diffusion. The typical thickness of vascularized
mammalian retinas is 200 to 300 μm. Species, whose retinas are 150 μm
or less, remain avascular and are supported by diffusion from the
choroidal circulation (66). The avascularity of the primate FAZ is
consistent with this, as the foveal depression is about 150 μm (10).
2. Development of the neural retina, which causes a local increase in
metabolic demand, also begins at the optic disc and follows a central-to-
peripheral fashion. Thus, retinal vessel formation follows the pattern of
neural retinal development, thereby relieving the local metabolic
demand. In particular, it has been shown that dysregulated photoreceptor
metabolism due to hyperglycemia leads to alterations in retinal vascular
development due to low adiponectin and that adiponectin administration
could restore the vascular development (67). Similarly, impairment of
retinal endothelial glucose metabolism by the uncoupling protein 2
(UP2) in the murine OIR model led to impaired vascular development,
and restoration of endothelial glucose metabolism by pharmacologic
inhibition of UP2 or overexpression of glut-1 normalized vascular
development (68).
3. Regions of the retina with higher oxygen supplies have a lower density
of retinal vessels. For instance, there is a 100- to 150-μm-wide
periarterial capillary-free zone that gradually diminishes in the transition
to branching arterioles and capillaries.
Taken together, these observations support the concept of “physiologic

hypoxia” as the driving force of retinal vascular development.
VEGF and Hypoxia

The concept of “physiologic hypoxia” also suggests the existence of a retina-
derived “vasoformative factor” (69), since shown to be VEGF, associated
with metabolic changes (oxygen), can mediate cell proliferation and regulate
retinal vessel formation. The fact that oxygen levels could regulate the
expression of VEGF made this protein an attractive candidate for the
vasoformative factor (70).
Hypoxia-Inducible Factor-1 (HIF-1)

Given that loss of oxygen regulation can lead to hypoxic injury, oxygen
sensing mechanisms have evolved to address the oxygen demand of the
growing vasculature. These regulators are the basic helix–loop–helix
transcriptional elements known as the hypoxia response factors (HIFs):
HIF1a, HIF2a, and HIF3a (65). One of the best-characterized transcriptional
regulators of VEGF is HIF-1, a heterodimer of two constitutively expressed
subunits, HIF-1α and HIF-1β (71). In the presence of oxygen, specific proline
residues in HIF-1α are converted to hydroxyproline by prolyl hydroxylase
(PHD), enabling the ubiquitination of HIF-1α followed by its destruction
through proteasome pathway. Under hypoxic conditions, the hydroxylation of
HIF-1α proline residues is limited due to the lack of oxygen molecules and
HIF-1α; therefore, it escapes degradation, is translocated into the nuclei and,
along with HIF-1β and cofactor CBP/P300, regulates VEGF mRNA
transcription (72,73). A role for Sirt1 in the regulation of HIF1α has been
demonstrated. Endothelial-specific deletion of Sirt1 led to delayed and
reduced retinal vascular density in vivo, an effect that was shown to be due to
Sirt1-dependent deacetylation of HIF-1α and resultant up-regulation of
VEGF (74).
In fact, it is the disruption of physiologic hypoxia when premature infants
are administered high oxygen that leads to the pathology referred to as
retinopathy of prematurity. This phenomenon, coupled with the fact that the
mouse retina is avascular at birth (see above), has permitted the development
of a murine model of oxygen-induced proliferative retinopathy (75). In this
model, postnatal mice are placed into a hyperoxic environment, which as for
the premature infants, leads to a disruption of the hypoxia-induced signal and
impaired vessel growth/remodeling. Upon return of the mice to room air, the
tissue senses hypoxia and a compensatory up-regulation of VEGF with
associated vessel growth occurs (Figure 5-3) (76).
FIGURE 5-3 Vaso-obliteration and neovascularization

tufts in the OIR model. Isolectin B4 staining of retinal
vasculature in mouse under normoxia or in different
stages of the OIR model. Left panel: normal vasculature
of a mouse under normoxia at P13, note the normal
arteries, veins, and capillaries; middle panel: the retinal
vasculature of a mouse at P12 that has undergone
hyperoxia during P7 to P12, note the vaso-obliteration in
central region of the retina; right panel: the retinal
vasculature of a mouse at P17 that has undergone
hyperoxia during P7 to P12 and returned to normoxia from
P12 to P17, note the neovascularization tufts in the
periphery of the retina. (Images courtesy of Magali Saint-
Geniez, Schepens Eye Research Institute.)
Hypoxia-Inducible Factor-2
It is well established that HIF-1α modulates hypoxia through maintenance of
VEGF gradients in the developing vasculature (77). However, less is known
regarding the role of hypoxia-mediated HIF-2α in vascular development.
Recent work has shown that deleting HIF-2α in murine neuroprogenitor cells
leads to a decreased number of arteries and veins as well as reduced density
of the deep vascular plexus in the periphery. These findings coincided with a
reduction in proangiogenic VEGF and erythropoietin expression, and an
increase in antiangiogenic endostatin levels. Given that HIF-2α is localized
exclusively to the neuroprogenitor cells, these findings demonstrate cross-talk
between neuroprogenitors and vascular development, through oxygen-
dependent up-regulation of HIF-2α (65).
Vascular Endothelial Growth Factor and Retinal

Vascular Development
Studies have revealed a range of candidate vasoformative factors in the
retina. The most attractive and well studied is VEGF or vascular permeability
factor (VPF). VEGF is a secreted glycoprotein, which has potent angiogenic
activity. In the endothelium, acting through two VEGF receptors (VEGFR1
and VEGFR2), VEGF leads to endothelial cell proliferation and migration as
well as increased endothelial permeability (78–80). In the developing retina,
VEGF is highly expressed at the leading front of forming vasculature, and
levels decline when the retina is fully vascularized. Cells that express VEGF
in the superficial layer are positive for astrocytes markers and for Müller cell
markers in deeper layers (6,81). The expression of VEGFR2, the primary
mediator of VEGF signaling, is spatially correlated with the zone of VEGF
expression, but slightly lags behind temporally and its expression is confined
to endothelial cells in newly formed vessels.
A proper VEGF “gradient” is essential for normal vascular development.
This is highlighted by studies in primates, in which there is no vessel
extension into FAZ despite the high VEGF expression by ganglion cells
during vascular development (82). Meanwhile, retinal capillary density is
greatest just peripheral to the rim of FAZ, where a steep VEGF gradient
appears (83).
VEGF Accessibility
In addition to the regulation of VEGF at the level of its expression, the
distribution of VEGF through its binding to heparan sulfate plays a critical
role in vessel growth and patterning. Alternative splicing of VEGF-A mRNA
gives rise to several variants that have different heparan sulfate–binding
capabilities with the major isoforms being VEGF120, VEGF164, and
VEGF188 in mouse and VEGF121, VEGF165, and VEGF189 in human (79).
VEGF120, lacking two highly charged domains, is the shortest isoform
and most soluble, whereas VEGF188 has the strongest binding ability and
VEGF164 is intermediate. Transgenic mice engineered to express only
VEGF120 isoform exhibit severe defects in vascular outgrowth and
patterning, whereas VEGF188 mice have normal venular outgrowth but
impaired arterial development. However, mice expressing only VEGF164
have normal retinal vascular development (84,85).
VEGF mRNA Stability

Additional to transcriptional regulation mediated by HIF-1, there is clearly, at
least under some conditions, a role for mRNA stabilization in the control of
VEGF mRNA levels. Studies using cultured retinal pigment epithelial cells
revealed that the half-life of VEGF mRNA under normoxia was quite low—
at about 45 minutes (86). However, growth of the cells under hypoxic
conditions led to a dramatic increase in the half-life to nearly 8 hours. The
very rapid turnover of VEGF mRNA under normal circumstances is a
characteristic of cytokines and growth factors and reflects the importance of
maintaining tight control over these potent growth factors. Studies using
tumor models revealed that the inclusion of 3ʹ untranslated sequences from
VEGF could increase the levels of VEGF and revealed that inclusion of
reporter constructs resulted in an increase in a reporter gene in the palisade
cells, which are known to experience hypoxia (87), and was later shown to
require the cooperation of multiple RNA elements (88).
Other Molecular Mediators of Retinal Vascular

Development
Placental Growth Factor

Placental growth factor (PGF) is a member of the VEGF family that binds
VEGFR1, and it has been primarily studies in association with preeclampsia,
but it does play a role in retinal vascularization. Mice with targeted deletion
of Pgf showed disrupted vascular morphology, which in some cases remained
in adulthood (89).
Notch Signaling
Endothelial cells at the front of developing vasculature are divided into tip
and stalk cells. Although these two populations of cells are virtually adjacent
to one another, they are very different morphologically and display distinct
responses to VEGF. Tip cells are, as the name suggests, at the front of the
growing vessel, they rarely divide, and are characterized by their long and
dynamic protrusions, named filopodia, that survey directional cues from
surrounding environment. On the other hand, stalk cells have fewer filopodia,
proliferate, and form lumens. A body of recent work has documented the role
of Notch signaling in coordinating VEGF signaling to specify the cell fate of
tip and stalk cells (59).
Notch is an evolutionary-conserved pathway that is involved in
embryonic development, regulating cell fate determination, tissue patterning,
and morphogenesis by mediating communication between neighboring cells.
Members of the Notch family are cell surface receptors on signal-receiving
cells, heterodimers of extracellular domain, and membrane-bound
intracellular domain. Upon binding with its ligands of the Jagged/Delta
family from the neighboring cell, the Notch intracellular domain (NICD) is
released by ADAM and γ-secretase and translocated into the nucleus where it
regulates gene expression. The extracellular domain of Notch that is bound to
the ligand on the surface of neighboring cells is transendocytosed by the
ligand-expressing cells.
At the growing front of developing vessels, Delta-like 4 (Dll4)/Notch1
pathway is involved in tip and stalk cells specification. Cells with higher
levels of Dll4 act as signal-sending cells. Binding to Notch1 receptor on the
neighboring cells results into Notch-dependent regulation of gene expression,
particularly down-regulation of VEGFR2 and Dll4 and up-regulation of
VEGFR1, the decoy receptor for VEGF. Thus, the neighboring cell, with a
reduced response to VEGF, acquires stalk cell fate, while Dll4-expressing
cells preserve its tip cell fate. During vessel formation, cell fate of tip cells
and stalk cells is dynamic and constantly changing as a function of the
Dll4/Notch ratio between a cell and its neighbor (90,91). Recent studies have
identified the transcription factor Krüppel-like factor 4 (KLF4) is upstream of
Notch; endothelial-specific overexpression of KLF4 resulted in increases in
vessel density, branching, and number of tip cells at P6 that, interestingly,
remodels during the subsequent day to a normal morphology in the adult
(92).
Vascular Remodeling
Remodeling is critical to the establishment of a mature, stable vasculature.
Blood vessels are either formed de novo as a tube, such as the aorta, or as a
primitive plexus, as is the case for the majority of retinal vascularization. In
either circumstance, the vessel is subsequently remodeled to form the final
vascular structure by the addition of wall cells (smooth muscle in larger
vessels and pericytes in the microvasculature—collectively referred to as
mural cells), deposition of a basement membrane, and for larger vessels, the
addition of the adventitia (consisting of fibroblasts and their associated
connective tissues). In addition to the growth factors discussed below,
vitamin D has been shown to contribute to vascular stability. Mice with
global deletion of the vitamin D receptor displayed normal vascular
development but exhibited an increase in the density of both pericytes and
endothelial cells in P42 vitamin D receptor–deficient mice (93).
Platelet-Derived Growth Factor (PDGF B) Signaling
Proliferating endothelial cells secrete PDGF B, which has been shown in
tissue culture studies to act as a chemoattractant and mitogen for mural cells
and their precursors (94,95). Consistent with these findings, mice that are
deficient of PDGF B have reduced numbers of smooth muscle cells in their
arteries and a paucity of pericytes in their microvascular bed (96).
Transforming Growth Factor Beta1 (TGFβ1)

The studies that implicate TGFβ in vessel remodeling are based on tissue
culture models. The analysis of deficiency in TGFβ is complicated by the
pleiotropic nature of the factor and the fact that the mice are either embryonic
lethal or die in the first few postnatal weeks due to uncontrolled inflammation
(97). Once the mural cell precursor has been recruited to the newly formed
capillary, contact between it and the endothelial cells leads to activation of
TGFβ1 (98) via a process that is thought to involve the plasmin cleavage of
the associated latency peptide (99). The activated TGFβ1 has a number of
effects that contribute to the ultimate stabilization of the microvessel
including inhibition of endothelial cell proliferation (98) and migration (99),
as well as inducing the mural cell precursor to differentiate to a smooth
muscle cell/pericyte fate (94). Clear evidence of a role for TGFβ in
maintaining vascular stability in the adult comes from studies in which
conditional and specific disruption of TGFβ signaling in endothelial cells
resulted in the development of choroidal neovascularization (100).
It is important to note that these growth factors are pleiotropic and that
TGFβ has also been shown to play a role in neuronal “pruning” during
mammalian retinal development. Conditional deletion of TGFRII during
development led to markedly increased death of retinal neurons both during
development and postnatally (101). In addition, deletion of just 50% (e.g.,
one allele) of SMAD7, a molecule that is central to downstream signaling of
TGFβ, led to increased proliferation of Müller cell progenitors in mouse
retinas at a time when Müller cells differentiate (P4) (102).
Angiopoitein-1/Tie-2 Signaling
In addition to VEGFR2, a receptor tyrosine kinase, as a master regulator of
angiogenesis, Tie-2 belongs to another class of receptor tyrosine kinases and
is critical for vascular development. Tie-2 was cloned and characterized as a
type I transmembrane protein including intracellular tyrosine kinase domain,
transmembrane domain, three fibronectin type III repeats, and two
immunoglobulin-like loop domains flanking three epidermal growth factor
repeats (103). Angiopoitein-1 and 2 (Ang-1 and Ang-2) were identified as the
endogenous agonist and antagonist, respectively, for Tie-2 (104,105). Tie-2 is
abundantly expressed by endothelial cells in developing vasculature but
lower in the adult vascular system (103,106). Ang1 appears to be expressed
mainly by perivascular and mural cells (107,108). Although the binding of
Ang-1 and Tie-2 leads to tyrosine phosphorylation of Tie-2, it does not
promote the growth of cultured endothelial cells (104). Ang-1 is primarily
expressed following or in immediate vicinity with developing vessel, while
Ang-2 is abundantly expressed at the front of invading vessels, suggesting
Ang-1 is mediating vessel maturation, remodeling, and stabilization, while
Ang-2 is cooperating with VEGF signaling for angiogenesis (105). Gene
deletion studies on Ang-1 and Tie-2 also suggest that they are required for
normal interactions between perivascular cells and endothelial cells
(108–110).
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6
Human Choriocapillaris Development
Gerard A. Lutty and D. Scott McLeod
INTRODUCTION
The human choroid forms the posterior portion of the uveal tract (the iris,
ciliary body, and choroid). The choroid is a thin, pigmented, and highly
vascularized tissue that lies beneath the sensory retina. The outer boundary of
the choroid is the lamina fusca and the inner boundary is Bruch membrane
(BrM), a pentalaminar structure upon which the single layer of retinal
pigment epithelium (RPE) sits. The adult choroidal vasculature has three
layers: the outermost Haller layer with large vessels; Sattler layer of
intermediate vessels in the middle; and the innermost layer being
choriocapillaris (CC), which is composed of broad capillaries with flat
lumens (20 to 50 mm diameter). The CC lies immediately adjacent and
posterior to Bruch membrane. The CC has a lobular pattern and is arranged in
a single layer restricted to the inner portion of the choroid. Feeding arterioles
and draining venules enter at right angles from Sattler layer in the posterior
pole. The choroidal vascular plexus in the periphery and equatorial areas of
choroid have arterioles and veins in the same plane as the CC, and the CC is
in a ladder-like arrangement. The choroidal vasculature is positioned between
two pigmented cell types, outer choroidal melanocytes of neural crest origin
and apical RPE of neuroepithelial origin. The basement membrane of the
adult CC constitutes the most posterior layer of BrM, and the endothelial
cells are fenestrated mostly on the retinal side. Fenestrated endothelium
usually is involved in secretion and/or filtration, for example, kidney
glomerulus. The CC is also sided in terms of receptor expression: VEGF
receptors 1, 2, and 3 (VEGF-R1, R2, R3) are most prominent on the retinal
side of the CC lumens (1). Vascular endothelial growth factor (VEGF)
appears to be secreted from the basal side of the RPE, which probably is
necessary for maintenance of the CC fenestrations (1) and for its survival
(2,3). CC is also one of the few capillary systems in which the endothelial
cells constitutively express ICAM-1 (4).
The adult CC transports nutrients and oxygen to the RPE and
photoreceptors (PRs) and removes waste from the RPE. The CC provides all
of the metabolic requirements for the PRs from serum components to 90% of
the O2 consumed by the PR in darkness (5). The PRs consume more oxygen
per gram of tissue weight than does any cell in the body. The tissue oxygen
level at the inner segments is near zero in the dark (5). Therefore, disruption
in choroidal blood flow would be detrimental to PR function and/or survival.
Abnormalities in the choroidal vasculature result in several congenital and
adult diseases, like choroidal coloboma and age-related macular degeneration
(AMD) (6–8).
This review summarizes our studies on the development of the human
CC from 5.5 until 22 weeks’ gestation (WG). The CC is a primitive vascular
system at 5.5 WG and will have almost an adult morphology by 22 WG.
Development of the CC by Hemo-Vasculogenesis (5.5

to 8 Weeks’ Gestation)
The three mechanisms for vascular development are (a) angiogenesis,
migration and proliferation of endothelial cells (EC) from an existing blood
vessel; (b) vasculogenesis, coalescence, and differentiation of vascular
progenitors or angioblasts; (c) hemo-vasculogenesis, differentiation of
vascular and blood cells, comprising erythropoiesis with expression of the
epsilon chain of hemoglobin or Hb-ε+, and hematopoiesis with CD34+
expression, both from a common progenitor, the hemangioblast. We have
recently found that hemo-vasculogenesis is responsible for the initial human
CC development between 5.5 and 8 WG (9). Erythroblasts, that is nucleated
erythrocytes expressing epsilon hemoglobin (Hbε+), were observed at 5.5 to 7
weeks of gestation (WG) within blood island–like structures in the CC layer
and also within the adjacent choroidal stroma (Figure 6-1). The erythroblasts
were in the walls of primitive lumens, as well as within lumens, and were free
within the choroidal stroma (Figure 6-1). Using the Hbε labeling in
conjunction with endothelial cell markers (CD31, CD34, CD39), we found
that the same cells could coexpress Hbε as well as endothelial cell (CD31,
CD39), hematopoietic (CD34), angioblast (CD39), and hemangioblast
(VEGFR-2 or KDR) markers (Figure 6-2) (9). This suggested that these cells
were hemangioblasts or progenitors derived from hemangioblasts and that
their aggregation and subsequent differentiation were responsible for the
initial CC development by the process of the hemo-vasculogenesis (9). When
CD39, a marker for endothelial cells and angioblasts (10), was used to label
flat mounts of choroid, the blood island–like appearance of the vascular
structures was apparent and erythroblasts were observed both inside and
outside of the islands (Figure 6-3A). We observed only a single layer of
vasculature (CD31+, CD39+, CD34+) at this time, suggesting that the CC
formed independent of a functional blood supply (Figure 6-4A). Vascular
lumens were apparent by 8 WG, and there were very few erythroblasts
associated with the vasculature. The erythroblasts remained only within
formed lumens by 9 WG in flat mounts (Figure 6-3B), when Hbε production
declines (9). The association of erythroblasts with endothelial-like
progenitors is apparent by ultrastructure in the early stages of hemo-
vasculogenesis (Figure 6-5A). The endothelial cell progenitors in some
structures almost enclosed erythroblasts in early lumen formation (Figure 6-
5B), but other structures were only empty slit-like lumens (11–13). CC
development by hemo-vasculogenesis from islands of progenitors seems to
explain how the CC forms without a source of blood (9). Hemo-vasculogenic
development of blood vessels has been observed in embryonic mouse in
several organ systems (14). This process for CC development was recently
observed in Zebra fish as well (15).
FIGURE 6-1 Cross sections of 6.5 WG fetal choroid
demonstrate hemo-vasculogenesis (A–F). Erythroblasts
(bright pink cytoplasm) in the choriocapillaris layer can
form solid cord-like structures (double arrow) without a
lumen (A). Erythroblasts, hematopoetic, and vascular cells
develop in situ with erythroblasts sometimes forming a
lumen (arrow in D). Eventually, the outer cells become
primarily endothelial cells, and the inner cells become
primarily erythroblasts (F). Free erythroblasts
(arrowheads in D–F) are present in stroma of choroid. The
monolayer of RPE cells is visible at the top in (A–C).
(Scale bar = 10 μm; Giemsa stained JB4 sections.) (From
Werner JS, Chalupa LM. The New Visual Neurosciences.
Cambridge, MA: MIT Press, 2013. Copyright © 2013
Massachusetts Institute of Technology, by permission of
The MIT Press.)
FIGURE 6-2 Colocalization of embryonic hemoglobin
(Hb-ε) and endothelial markers (CD31 and VEGFR-2 or
FLK-1) in developing choriocapillaris (CC). A–D:Hb-ε
(green) and CD31 (red) are colocalized in cells of the
developing CC (arrows) and single cells within the
choroidal stroma (arrowhead). E–H:Hb-ε and FLK-1
(red) coexpression in cells lining a developing lumen
(arrows) and in cells located outside of the structure
(arrowhead). (Scale bars = 10 μm; counterstained with
DAPI, blue. (Reprinted by permission from Springer:
Lutty GA, Hasegawa T, Baba T, et al. Development of the
human choriocapillaris. Eye (London) 2010;24:408–415.)
FIGURE 6-3 CD39 immunolabeled flat choroids showing
the pattern of the developing choriocapillaris in embryonic
and fetal human eyes. At 5.5 WG (A), CD39-positive cells
and erythroblasts are organized in blood island–like
structures (circle). At 9 WG (B), highly cellular linear
cord-like structures without apparent lumen have formed.
By 12 WG (double arrows) (C), the capillaries are
narrower, lumens have formed, and cellularity has
decreased markedly.
FIGURE 6-4 CD31 immunolabeling of choroidal sections
from embryonic eyes at 6 WG (A), and fetal eyes at 12
WG (B), and 20 WG (C). Only a highly cellular
rudimentary choriocapillaris (CC) with poorly defined
lumen (arrow) is present at 6 WG (A). At 12 WG (B),
vessels are budding from the CC into the deeper choroid
(arrowhead). At 22 WG (C), the CC (arrow), medium-
size vessels of the Sattler layer (arrowhead) and the larger
outer blood vessels (open arrow) are all present. (Scale bar
= 30 μm.)
FIGURE 6-5 Ultrastructure of the developing embryonic
and fetal human choriocapillaris at 6.5 WG (A,B), 16 WG
(C), and 22 WG (D). In ultrathin sections from 6.5 WG
choroid (A,B), free progenitor cells (M) make contact with
free erythroblasts (EB). In other areas, immature
endothelial cells (ECs) envelop the erythroblasts to form
blood island–like structures (B). The RPE are present in
the upper left hand corner in (A) and (B). By 16 WG (C),
lumens are apparent (asterisk), and the EC nuclei (e) with
more organized chromatin were reduced in volume and
had decreased cytoplasmic projections. By 22 WG (D),
lumens were broad and flat (asterisk), ECs were thin and
fusiform, and definitive pericytes (p) were present on the
outer surface of the capillaries. Fenestrations were present
along the inner aspect of the CC at this stage of
development (inset) on the retinal side of CC lumens
under RPE (top). (Scale bars = 20 μm [A,B]; 2 μm [C]; 4
μm [D].) (From Werner JS, Chalupa LM. The New Visual
Neurosciences. Cambridge, MA: MIT Press, 2013.
Copyright © 2013 Massachusetts Institute of Technology,
by permission of The MIT Press.)
The functional properties of the CC were evaluated by performing

immunohistochemistry of three enzymes known to be important in adult CC
function: carbonic anhydrase IV (CA IV), endothelial nitric oxide synthase
(eNOS), and alkaline phosphatase (APase) (16). CA IV, an ecto-enzyme on
endothelial cells, controls local pH when coupled to the electrogenic sodium
bicarbonate cotransporter (NBC1) (17–19). CA IV and eNOS were expressed
at low levels as early as 8 WG, during hemo-vasculogenesis (5.5 to 9 WG)
(11). eNOS’s function is presumably vasodilation of blood vessels, but
mostly slit-like lumens were present at 8 WG (11). Neuronal NOS (nNOS) on
the other hand was present in the nuclei of vascular progenitors in CC, retina,
and vitreous, as well as in the nucleus of RPE cells and scattered progenitors
throughout choroid (Figure 6-6) (20). APase is a marker for choroidal blood
vessels in adult human (16) and it was present in forming CC already at 7
WG, even though the primitive ECs were just starting to differentiate from
hemangioblasts. However, the observation of high APase activity in the
blood island–like formations is not unexpected, because it is used extensively
as a marker for human embryonic stem cells and other progenitors (21,22).
FIGURE 6-6 Choroid of a 21 WG fetal eye labeled for
vWf (red) and nNOS (green) and DAPI (nuclei) (A), and
shown as single (C,D) and multiple channel (A,B) images
to demonstrate colabeling. Nuclear nNOS expression is
seen in RPE (arrowhead), ECs of CC (arrow), and large
choroidal vessels (paired arrow). Nuclei were
counterstained with DAPI (blue). Scale bar in “A” = 10
μm. (From Werner JS, Chalupa LM. The New Visual
Neurosciences. Cambridge, MA: MIT Press, 2013.
Copyright © 2013 Massachusetts Institute of Technology,
by permission of The MIT Press.)
9 to 12 Weeks’ Gestation
Between 9 and 12 WG, development of intermediate and deeper choroidal
vessels was observed in choroidal stroma. This development was more
advanced in the posterior pole than in the equatorial choroid, suggesting it
started in the posterior pole (11). The forming vessels expressed EC markers
including CD31, CD34, and CD39. Proliferation was observed in some ECs
that were budding from the scleral side of the CC using CD34 and Ki67
double labeling, suggesting that intermediate vessels form by angiogenesis
(9). A rather linear pattern was observed in flat CD39-labeled preparations of
choroid (Figure 6-3B) at 9 WG and then a chicken wire–like pattern of
vessels with a few free CD39-positive cells between the vascular segments at
12 WG (9). This suggested that some angioblasts were still present in the
choroid (10) at 12 WG; however, the pattern of CC was approaching an
adult-like lobular pattern but the density of 12 WG CC was much less than
the adult CC (16).
Adult CC is fenestrated mostly on the retinal side of the CC lumens, so
we investigated the presence and position of fenestrations and their
components at different ages. PV-1, also called PLVAP (plasmalemmal
vesicle–associated protein), is an integral membrane glycoprotein in the
diaphragms of fenestrations (23,24). All vessels were negative for PV-1 at
and before 12 WG. Occasional fenestrations were observed with TEM, but
they were associated with filipodial-like structures both in and around the
lumens (11). Because of their unusual position, these fenestrations were
probably not functional.
We and Sellheyer and Spitznas observed that the CC at 9 WG was
composed of aggregates of progenitor cells with only slit-like lumens (13).
We observed some cells adventitial to cells lining the primitive lumens, but
the two cell types were indistinguishable in ultrastructural appearance in
periphery (11). Some plump progenitor cells that bordered on the lumens had
tight junctions. Some Weibel-Palade bodies, organelles found only in
vascular endothelial cells, were present as well. Central choroid (area from
disc to equator) had more definitive pericyte-like cells on the more developed
vessels (11). Pericytes had a nuclear organization that appeared distinct from
the endothelial cells on the lumens of more mature central blood vessels.
Complex membranous infoldings that resembled filopodial processes
extended into the slit-like lumens from the lumenal cells. At the equator,
some lumens were more open and the filopodia appeared to touch
erythrocytes in the lumen; the plasma membranes of the two cells could not
be discerned. Basal lamina was not observed around these developing vessels
(11).
At 14 WG, pericyte-like cells, progenitors in the ablumenal position, formed
peg-in-socket-like contacts with endothelial cells lining the lumen, a
characteristic of normal adult microvasculature (11). Maturation of these
ablumenal cells was evaluated by localizing two pericyte markers: NG-2, a
glycosaminoglycan present on the surface of pericytes, and alpha smooth
muscle actin (aSMA), present in mature pericytes and smooth muscle cells
(SMC). NG2 immunoreactivity was very prominent at 14 WG, but there was
limited aSMA immunoreactivity.
PV-1 was present at low levels in some areas of CC at 16 WG,
suggesting the presence of some fenestrations. This was confirmed with
TEM, which showed a few fenestrations in the CC, but these were not
continuous in the thin endothelial cell processes on the retinal side of the
lumens (Figure 6-5). The CC in the posterior pole had the most fenestrations
compared to peripheral regions and was most mature morphologically. In the
broader lumens at 14 WG, the number of filopodia in lumens appeared
greatly reduced compared to 11 WG (Figure 6-5C). EC nuclei had less dense
chromatin and were more oval and uniform in shape and BrM organization
was more advanced. The CC and intermediate choroidal vessels, which were
more abundant at 14 WG, had intense APase and prominent CA IV and
eNOS immunoreactivity.
Three layers of blood vessels, as demonstrated with EC markers, were
apparent at 21 WG within the posterior pole (Figure 6-4C). Short
rudimentary inner segments were present at the outer most portion of the
neuroblastic layer, providing the first evidence of PR maturation (11). PV-1
immunoreactivity was present in most of the CC, but it was more intense in
the posterior pole than in the periphery. However, PV-1 was uniformly
intense and more apparent on the retinal side of the CC lumens in the adult
human eye used as a positive control (11). eNOS was prominent in EC
cytoplasm and cell membrane of all choroidal blood vessels. nNOS was
mostly nuclear in both pericytes and SMC labeled with αSMA as well as in
ECs, which were double labeled with von Willebrand factor (vWf) (Figure 6-
6) (20).
CC endothelial cells were thin and fusiform ultrastructurally at 22 WG.
The CC had contiguous areas of fenestrations in the narrow endothelial
processes on the retinal side of the broad lumens (Figure 6-5D). Endothelial
cells had well-formed tight junctions, Weibel-Palade body numbers had
increased, and a continuous basement membrane was present. Collagen and
elastin was clearly deposited in BrM under the RPE basement membrane.
αSMA+ cells were present in the CC, as well as intermediate and large
choroidal blood vessels at 22 WG. Pericytes were apparent by TEM at this
age (Figure 6-5D), NG2 was very prominent, and the pericytes were located
primarily on the scleral side of the CC.
Regulation of Vascular Development

Vascular development in many tissues, including the retina, is known to be
controlled by vascular endothelial growth factor-A (VEGF-A) (25). The most
critical isoform of VEGF-A in pathological angiogenesis is thought to be the
VEGF165 isoform (26). Recently, it has been reported that VEGF-A has two
groups of splice variants from the VEGF-A gene product, VEGFxxx and
VEGFxxxb, in addition to the known isoforms with varying heparin binding
affinities and molecular sizes: VEGF121, 145, 165, 189, 206 (27–29). The splicing
of the VEGFxxxb family members, the antiangiogenic forms, is stimulated
by TGF beta through splicing factor SRp55 (serine/arginine protein 55) (30).
VEGFxxx, on the contrary, is proangiogenic and IGF1 (insulin-like growth
factor 1) and TNF alpha (tumor necrosis factor alpha) up-regulate splicing
through ASF/SF2 (alternative splicing factor/splicing factor 2) (30,31). The
two antagonistic splice variants were recently investigated in developing
choroid. VEGF165 was only prominent in the forming CC during
hemovasculogenesis, while VEGF165b is not present (32). VEGF165
expression increased with time in the forming vasculature and became very
prominent in the basal portion of RPE and in CC at 12 WG. VEGF165 was
very prominent in the basal RPE and choroidal vessels at 17 WG, and
VEGF165b expression was also present (32). By 21 WG, the level of the two
splice variants was comparable, and both appeared to be present in the same
cells (Figure 6-7). VEGF165b localized to the nuclei of RPE and CC
endothelial cells while VEGF165 was present mostly in the cytoplasm of CC
endothelial cells and basal cytoplasm of the RPE (Figure 6-7). Therefore,
VEGF165b is only present as vascular development nears completion while
VEGF165 is prominent during hemo-vasculogenesis and angiogenesis.
FIGURE 6-7 VEGF165 (red) (C) and VEGF165b (green)
(D) coexpression (B) in the choriocapillaris (single arrow),
deeper choroidal vessels (paired arrows) and the RPE
(arrowhead). VEGF165b has an apparent nuclear
localization in the endothelium of blood vessels. In the
RPE, it is associated with the nuclei and in the apical
portion of cells. VEGF165 is diffuse in endothelium and
mostly localized to the basal portion of the RPE. Nuclei
were counterstained with DAPI (blue) (D). Images are
shown in single (C,D) and multiple (A,B) channels to
demonstrate colabeling. Scale bar in “A” = 10 μm.
(From Werner JS, Chalupa LM. The New Visual Neurosciences. Cambridge, MA: MIT Press, 2013.
Copyright © 2013 Massachusetts Institute of Technology, by permission of The MIT Press.)
DISCUSSION AND SUMMARY

The adult human CC is a fenestrated and lobular vasculature that is similar to
kidney glomeruli and unlike the terminal end-arterial vascular bed of the
retina. Therefore, it is reasonable that the mode of CC development would be
unique from the retinal vasculature. Ida Mann provided the first elegant
documentation by paraffin histology and light microscopy of the
development of human CC (33). Sellheyer and Spitznas then demonstrated
with meticulous ultrastructural analysis the primitive appearance of the early
CC (13). Our studies demonstrate that human choroidal vasculature
development progresses from the posterior pole to the periphery and involves
several processes. The initial embryonic human CC forms by hemo-
vasculogenesis: progenitors expressing a hematoendothelial phenoptype (Hbe
as well as CD31, CD34, VEGFR-2) differentiate into endothelial cells
(CD31+, CD39+), pericytes (NG2+), erythroblasts (Hbε+), and hematopoietic
cells (CD34+). This process was first documented in blood islands of the yolk
sac but has recently been described throughout the mouse embryo by
Sequeira Lopez and associates (14). In the fetal period (9 WG and older),
hemo-vasculogenesis was complete and new blood vessels appeared to bud
from the scleral side of the newly formed CC by angiogenesis, demonstrated
by the presence of migrating and proliferating Ki67+ ECs (Table 6-1) (9).
The budding from the CC results in the intermediate blood vessels and
eventually anastomoses with other CC segments and larger vessels, as
observed by Drake and associates in mouse embryo (34). In the ages included
in our studies (5.5 to 22 WG), the CC never reached the lobular pattern in the
posterior pole or ladder pattern in periphery or vascular density of the adult
(16), suggesting that significant expansion and remodeling of the system will
occur after 22 WG. Free CD39+ angioblasts were still present in between
formed segments of CC, so additional vessels may form by vasculogenesis,
with the aggregation and differentiation of angioblasts (CD39+) as occurs in
development of the human retinal vasculature (10,35). Alternatively, the
additional segments may form by angiogenesis, because proliferation is still
occurring in CC at 22 WG.
TABLE 6-1 Time line of choriocapillaris

development
aData from Baba T, Grebe R, Hasegawa T, et al. Maturation of the fetal human choriocapillaris. Invest
bData from Hasegawa T, McLeod DS, Bhutto IA, et al. The embryonic human choriocapillaris
develops by hemo-vasculogenesis. Dev Dyn 2007;236:2089–2100.
cData from McLeod D, Baba T, Bhutto I, et al. Co-expression of endothelial and neuronal nitric oxide
synthases in the developing vasculatures of the human fetal eye. Graefes Arch Clin Exp Ophthalmol
2012;250(6):839–848.
dData from Baba T, McLeod DM, Edwards MM, et al. VEGF165b in the developing vasculatures of
the fetal human eye. Dev Dyn 2012;241:595–607.
WG, weeks of gestation; Hemo-vas, Hemo-vasculogenesis; TEM, Transmission electron microscopy;
SMA, smooth muscle actin; CAIV, carbonic anhydrase IV; APase, alkaline phosphatase; eNOS,
endothelial nitric oxide synthase; nNOS, neuronal NOS; VEGF, vascular endothelial growth factor.
It appears that CC maturation is concomitant with both PR differentiation and

RPE maturation. PR metabolism has increased since newly formed inner
segments are rich in mitochondria at 22 WG. Contiguous fenestrations were
present at this time, and fenestrations will transport solutes and small
molecules to and from the CC. Bruch membrane (BrM) is still very immature
at 22 WG, so additional production of BrM matrix will occur after this.
Investing of adventitial cells is critical for vascular maturation: pericytes
around capillaries and venules, and SMC in the walls of arterioles and
arteries. Little is known regarding the origins, differentiation, and appearance
of contractile, adventitial cells associated with the choroidal vasculature
during embryonic and fetal development. Our observations demonstrate that
lumenal (presumably endothelial cells) and perivascular cells (presumably
pericytes) were indistinguishable ultrastructurally in developing embryonic
human CC, that is, during hemo-vasculogenesis, suggesting that pericytes
and ECs are probably derived from a common progenitor, the hemangioblast.
Although the pericyte marker, NG2, was present at low levels at 7 WG and
more prominently at 12 WG in vascular structures (Table 6-1), αSMA, the
predominant actin isoform found in mature SMC and pericytes (36), was not
present until 22 WG. Our in vitro studies of retinal angioblasts demonstrate
that the same progenitor may differentiate into either an endothelial cell or
pericyte depending on the culture conditions (37). TEM demonstrated that
pericytes were present at 22 WG and were located predominantly on the
scleral side of the CC (Figure 6-5D).
A key event in blood vessel development is lumen formation. The initial
luminal spaces of the early CC were slit-like structures between primitive
endothelial cells, as observed by Sellheyer as early as 6.5 WG (13). Our
studies show that even the erythroblasts can participate in lumen formation
during hemo-vasculogenesis (Figures 6-1 and 6-2) (9). Even at this stage, the
lumenal cells made recognizable junctional complexes (Figure 6-5B) that are
needed for a mature vasculature, suggesting that these lumenal cells were
committed to being endothelial cells. The filopodia-like cytoplasmic
extensions from developing endothelial cells were similar to those observed
by Roy et al. in chick brain and, like in chick, the number of cytoplasmic
extensions decreased as the lumens became broader (38). Angioblasts and
ECs use these processes to touch and interlock with each other (39) and
erythroblasts. ECs finally were fusiform in shape at 22 WG, when the wall of
the vessels became thin and lumens became broad and flat. This probably
reflects changes associated with blood flow. Finally at 22 WG, pericytes have
assumed a flatter morphology and their processes ensheathe the vessel wall
(Figure 6-5D).
The CC is one of the few fenestrated capillary beds in the body. The
pores of CC fenestrations have diaphragms that include a protein recognized
by the PV-1 antibody. Passive transport by fenestrations of some fluids and
macromolecules of the correct charge is critical in providing PR and RPE
with nutrients, ions, and small molecules as well as transport of the waste
from the RPE. It was not until 22 WG that CC had contiguous fenestrations,
which is the time in development when PV-1 immunoreactivity was greatly
increased in CC.
Formation of capillaries by hemo-vasculogenesis may explain the
development of the lobular pattern of CC in that the first blood vessels are
islands. Eventually, the islands connect to each other without any
contribution from blood flow, because intermediate and large blood vessels
are not yet connected as yet (Figure 6-8), similar to the sequence of events in
kidney development (40). Eventually, the original islands and the
interconnecting segments become the lobular system of flat broad capillaries
whose lumens are separated by the intercapillary septa or pillars of matrix,
seen in adult CC (16). The final mature CC is very similar to the capillaries of
kidney glomeruli: large flat, fenestrated capillaries that are lobular in pattern
(40). Functional maturity may occur in advance of structural maturity (Table
6-1). Fenestrations form late in maturation (21 to 22 WG), which is
coordinated with the differentiation of PRs that Hendrickson and coworkers
have reported begins around 24 to 26 WG when inner segments form (41).
The CC will be fenestrated after this mostly on the RPE side (42), which is
critical for its adult function in supporting the viability and survival of PRs
and RPE cells.
FIGURE 6-8 A schematic representation of embryonic
and fetal human choriocapillaris development by the initial
process of hemo-vasculogenesis. Loose progenitor cells
outside of free erythroblasts aggregate to form blood
island–like structures. Cells in the blood islands
differentiate and organize into primitive blood vessels:
endothelial cell–like cells line a lumen filled with
erythroblasts and other blood cells while pericyte-like cells
occupy an ablumenal position. Other progenitors
eventually bridge the new radially oriented vessels, which
become united to each other yielding a chicken-wire
pattern that will become the lobular adult CC. (From
Werner JS, Chalupa LM. The New Visual Neurosciences.
Cambridge, MA: MIT Press, 2013. Copyright © 2013
Massachusetts Institute of Technology, by permission of
The MIT Press.)
The final product of this unique developmental process forebodes

susceptibility to choroidal vascular diseases. Although the volume of blood in
the choroidal vasculature is large compared to the retinal vasculature, the
velocity of red blood cells (RBCs) in CC is 4 times slower than in retinal
capillaries (43,44). The lobular pattern of the CC means that there are no long
linear segments for RBC transit; each time the RBC moves, it meets a
vascular wall adjacent to an intercapillary septum. Because ICAM-1 is
constitutively expressed in CC (4), activated leukocytes with CD11/CD18
can bind to the endothelial cell’s lumenal surfaces, further slowing blood cell
velocity (45). The choroidal vasculature was once thought to have a vast
capillary system with great redundancy, so loss of some capillary segments
would not be detrimental (46,47). However, the loss of capillaries we have
seen in AMD and diabetes and the knowledge that PRs deplete most of the
oxygen provided in the dark suggests that PRs would be at risk with loss in
CC. A reduction in submacular blood flow in AMD subjects has been
documented by the Grunwald group (48,49). Furthermore, RPE may become
hypoxic with adjacent loss in CC up-regulating hypoxia-inducible growth
factors like VEGF (50), which would stimulate the choroidal
neovascularization formation from CC that occurs in wet AMD and diabetic
choroidopathy (8,51).
ACKNOWLEDGMENTS
Grant support: NIH grants EY016151 (GL), EY01765 (Wilmer); RPB
unrestricted funds (Wilmer), the Altsheler-Durell Foundation; and a gift from
the Himmelfarb Family Foundation in the name of Morton F. Goldberg.
Gerard Lutty is an RPB Senior Investigator. The authors acknowledge the
excellent electron microscopy of Rhonda Grebe, and confocal microscopy of
Takuya Hasegawa and Takayuki Baba. Takuya Hasegawa and Takayuki
Baba were Bausch and Lomb Japan Vitreoretinal Research Fellows, and
Takayuki Baba was also a Uehara Memorial Foundation Research Fellow.
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7
Development of Cone Photoreceptors
and the Fovea Centralis and the
Impact of Prematurity
Jan M. Provis and Michele C. Madigan
The first cells to exit the mitotic cycle in the developing human retina are the
foveal cone photoreceptors. Although the genes responsible for specification
of the location of the fovea in the early eyecup have not been identified,
histologic studies show that cone photoreceptors prevail in this “central”
region of the retina from around 10 to 12 weeks of gestation (WG) (1,2).
Very few rods are ever seen at this highly specialized part of the
photoreceptor mosaic (3–5), which we subsequently refer to as the “foveal
cone mosaic.” While the foveal cone mosaic differentiates early, the foveal
depression is not discernable until many weeks later—at approximately 26 to
28 WG (6–9). We also now know that formation of a fovea is critically
dependent upon development of the retinal capillary network in the future
macular region and definition of a foveal avascular zone (FAZ) (10,11). With
the advent of advanced imaging technologies, particularly optical coherence
tomography (OCT), it is possible to track foveal development longitudinally;
this approach is proving its utility in understanding how the fovea is affected
by prematurity and why individuals born prematurely may experience
uncorrectable vision loss (12,13).
FORMATION OF THE FOVEA—

NORMAL DEVELOPMENT
The stages in differentiation and formation of the fovea in primate retinas
have been reviewed in detail (14), modeled using finite element analysis
(15,16), and the key mechanisms, including some molecular factors and their
roles, have been identified (17). In this chapter, we aim to highlight the
temporal relationship between development of the retinal vasculature and
formation of the fovea and to discuss maturation of the photoreceptor mosaic,
as a basis to understand how the fovea appears at full-term birth and how
foveal development and photoreceptor maturation might be affected by
prematurity.
A major step forward in understanding the mechanisms that generate the
fovea has been the recognition that in primates, the fovea forms only after the
FAZ has been defined. It is now clear from studies of monkey retinas that the
first step in the formation of the fovea is the definition of a central “no-go”
zone at the posterior pole of the eye, into which blood vessels do not grow
(10,15). Using a combination of OCT and fluorescein angiography, many
clinical studies have also reported that the fovea is absent in individuals who
lack an FAZ (18–24).
The FAZ forms in the ganglion cell layer (GCL) between 25 and 27 WG
approximately and overlies the foveal cone mosaic. The retina shown in
Figure 7-1 is from a fetus at 26 WG and has been immunolabeled with
antibody to rhodopsin, labeling rods green, and a cocktail of antibodies to
label endothelial cells of the developing retinal vasculature (red). The figure
shows the FAZ forming in a region where there are very few rods; it is closed
on the nasal side but remains open along the temporal raphe region in this
specimen. The fovea begins to form once the FAZ is fully enclosed by
perifoveal capillaries.
FIGURE 7-1 A montage of a whole mounted human fetal

retina at 26 WG, immunolabeled to show rod
photoreceptors (green) and retinal blood vessels (red) and
imaged by confocal microscopy. Developing retinal
vessels approach the incipient fovea from the nasal side
(left) as well as above and below. The FAZ is defined
when the superior temporal and inferior temporal
quadrantic arteries (above and below, respectively) meet
along the raphe in temporal retina sometime between 25
and 28 WG, approximately. FAZ, foveal avascular zone.
(Figure 1 is from a retina labeled by Anita Hendrickson,
imaged by JP.)
The FAZ appears to be specified by local expression of the antiangiogenic

pigment epithelium-derived factor (PEDF) by ganglion cells at the incipient
fovea and in the emerging fovea during its formation (25,26). Those studies
also find that EphA6 (a receptor for the axon guidance factors ephrin-A1 and
-A4) may play a role in vascular patterning around the FAZ and likely
accounts for the slow growth of vessels into the posterior pole of the retina
(27). EphA6 is also highly expressed by ganglion cells at the incipient and
developing fovea, and in other animals, EphA6 has been found to regulate
mapping of ganglion cell axons into the appropriate areas of the visual target
nuclei in the brain. The EphA6 molecule interacts with its ligands, ephrin-A1
and -A4, in a repellant response—such that cells expressing the ligand are
turned away from areas of high receptor expression. In the retina, astrocytes
lead retinal endothelial cells in their migration across the retina and express
the ephrin-A1 and ephrin-A4 ligands.
This finding indicates that astrocytes, and hence retinal vessels, are
repelled from the incipient foveal region where EphA6 is highly expressed,
resulting in the slow growth of retinal vessels into the foveal region during
development (27). Development of the retinal vessels results in a retina that is
scaffolded by a capillary plexus throughout, except at the FAZ. It is proposed
that as a result, the FAZ is more responsive to mechanical forces acting on
the retina. That is, because the FAZ is avascular, it is more deformable and
more elastic than the rest of the vascularized retina (28). Using this
observation as a premise, Springer (1999, 2004) has used finite element
analysis to model the effects of pressure (resulting from intraocular pressure)
and stretch (resulting from growth of the eye) on formation of a fovea. The
findings of modeling, combined with careful observation of the growth
surges in the posterior eye, suggest that intraocular pressure acts on the retina
within the FAZ to initiate formation of the foveal depression, soon after
definition of the FAZ. By closely matching changes in the fovea with
analysis of retinal growth, the findings also suggest that when the retina
enters a period of very rapid growth after birth, the FAZ and the early fovea
are affected by growth-induced stretch, which modifies the profile of the
fovea making it more shallow, and the sides more gently sloping, than in the
intrauterine period (28). Previously it has been proposed that tension on
ganglion cell axons during development, like that which would result from
retinal stretch, may be the mechanism that mediates centrifugal displacement
of ganglion cells that results in excavation of the fovea, during the late
prenatal and postnatal phase of development (29).
DIFFERENTIATION OF CONES—
NORMAL DEVELOPMENT
The maturation of the cone photoreceptor population during intrauterine and
early postnatal development follows a rather surprising trajectory. That is,
even though foveal cone photoreceptors are the first cells to differentiate in
the retina, they are the last to achieve the adult-like characteristics of foveal
cones. This oddity in maturation of foveal cones has been known of for at
least a century (6) but is now achieving clinical significance, since the
delayed maturation of the fovea is now widely recognized as a feature that
distinguishes OCT images from adult and neonatal maculae (30–33). The
differences between foveal cones and those on the foveal rim are illustrated at
mid-gestation (20 WG) and full term (40 WG) in Figure 7-2. At 20 WG,
cones in the foveal cone mosaic (“fovea”) are similar in size and appearance
to their neighbors on the foveal rim, which are interspersed with rods. At 40
WG, cones in the central fovea are only a little more differentiated and
slightly taller than at 20 WG. In contrast, cones on the rim of the fovea at 40
WG are highly differentiated and significantly narrower and taller than those
on the rim at 20 WG as well as those within the fovea at 40 WG.
FIGURE 7-2 Cones from within the fovea, and from the
foveal rim, drawn to scale at mid-gestation (20 WG) and at
full term (40 WG). At 20 WG cones are cuboidal cells
apposed to the RPE layer. They have no axonal process,
and the inner and outer segments are undifferentiated, as
seen by light microscopy. Cones on the foveal rim at 20
WG are beginning to elaborate axonal processes, and
differentiation of the inner and outer segments is slightly
more advanced. At 40 WG, cones on the foveal rim are
adult-like, having distinct axonal processes that are
sandwiched by rod somas as well as distinct inner and
outer segments. However, in the fovea at 40 WG, cones
are relatively unchanged since mid-gestation; inner and
outer segments are becoming evident, and the axonal
processes are beginning to narrow. However, overall, they
are only slightly taller than cones at 20 WG and
significantly shorter than those on the foveal rim, at birth.
(Drawings in Figure 2 are by Renee Argaet.)
Thus the fovea is immature at full-term birth in that first, the outer nuclear
layer (ONL) of the fovea at full term comprises cone photoreceptors of a very
immature morphology, which are not stacked into tiers of cells (as in the
adult fovea); rather, they remain in a monolayer of cuboidal cells and are
little changed in shape or size since mid-gestation. Second (as we will see
below), at full term, the fovea still includes cells in the ganglion cell and
inner nuclear layers.
The cellular mechanisms that mediate morphologic transformation of
cones are not clearly understood. It is known that aggregation and elongation
of photoreceptors in central retina is common among primates, even in those
species that do not have a fovea (34). Narrowing and elongation of cones is
the key mechanism that mediates the accumulation of cones in the fovea and
adjacent macula (35), a process commonly referred to as “cone packing.” By
becoming narrower and more elongated, cones can be packed tightly together
in a space-efficient hexagonal pattern, allowing more cones to be
accommodated per unit area in the foveal cone mosaic, while the cell–cell
relationships that were established early in development are preserved. Thus
changes in cell shape mediate cone packing. Fibroblast growth factor (FGF)
2 signaling via FGF receptors, which are differentially distributed across the
soma, axon, and inner and outer segments of cones during development has a
role in mediating the changes in cell shape which favor cone packing (36,37).
Establishment of a high density of cones in the foveal cone mosaic is the
anatomical basis of high acuity vision, and visual acuity is directly
proportional to the packing density of cones within the foveal cone mosaic.
Developmentally (and evolutionarily), there is a drive toward increased
cone density retina in the foveal cone mosaic. The spatial density of cones in
the foveal cone mosaic increases from around 10 K/mm2 to approximately 30
K/mm2 during intrauterine life (2,35), without newly generated cells being
added to the mosaic. This increase in cone density would be seen to be even
greater if cone density in the parafovea (rather than fovea) was compared
over time, since it is now understood that maturation (narrowing and
elongation) of cones in the fovea, and hence their packing, is delayed until
the postnatal period, while cones adjacent to the fovea achieve adult-like
features prenatally (7,36,38–40).
The spatiotemporal pattern of cone packing over time can be observed in
flat-mounted retinas, using the population of cones sensitive to blue/short-
wavelength light (S-cones) as a population “marker.” S-cones constitute
approximately 6.8% of the cone population within 4 mm of the foveal center
(41). They can be identified in postmortem retinas using an antibody against
the short wavelength–sensitive opsin contained in the outer segments of
mature photoreceptors and also present in the cell bodies of developing S-
cones (4). In Figure 7-3, the spatial density of S-cones along the horizontal
meridian of the retina between the fovea and optic disc is shown at three
different ages—18 WG, 6 weeks postnatal (P 6 weeks), and in the adult (data
from (42)). The graphs show that S-cone density just outside the central fovea
(at ~15% of the fovea-to-optic disc (OD) distance, or ~500 μm eccentricity)
increases between 18 WG (pale blue line) and adulthood (black line), while
S-cone density near the optic disc is significantly reduced over the same
period. The graphs indicate that high cone density near the fovea, and in the
macula in general, is achieved by mass displacement of cones from more
eccentric locations (centripetal displacement, indicated by the arrow), since
there is no detectable cell death during this period, and no photoreceptors are
added to the mosaic (4). Note that the S-cone population cannot be used to
indicate changes in total photoreceptor density closer than approximately 1
mm from the foveal center, because S-cones are absent from the fovea in
adults and throughout development.
FIGURE 7-3 Graphs illustrating the change in distribution
of cones in the fovea-to-optic disc region, between 18 WG
and adulthood. The population of cones sensitive to blue
light (short-wavelength–sensitive or S-cones) has been
used as a marker population, representing approximately
6.8% of the total cone population. Also note that because
S-cones are absent from the fovea, numbers in the 10%
closest to the fovea (left side of the figure) are not
indicative of the total cone population. No photoreceptors
are added to the mosaic in this region after approximately
18 WG, and there is negligible cell death. The graphs
show that early in development, cones are more numerous
near the disc than near the fovea; but during in utero
development, there is mass displacement of cones toward
the fovea (indicated by the arrow). This displacement
continues for some months, postnatal. Thus the
mechanisms that affect displacement of cones toward the
fovea act over quite long distances—at least up to
approximately 3 mm. (Reprinted from Cornish E,
Hendrickson A, Provis J. Distribution of short wavelength
sensitive cones in human fetal and postnatal retina: early
development of spatial order and density profiles. Vision
Res 2004;44(17):2019–2026. Copyright © 2004 Elsevier.
With permission.)
These data, along with morphologic observations of cone development,

suggest that displacement of photoreceptors is achieved as a result of a wave
of photoreceptor elongation, which passes from the disc toward the fovea,
over a period of several months (36,40). Increased oxygen supply to cells
allows for an increase in metabolic rate. In humans and macaques, the
passage of this wave of cone packing/elongation coincides approximately
with the spread of the outer plexus of the retinal blood supply toward the
fovea, suggesting that the wave of cone maturation may be attributable to
delivery of additional oxygen and metabolites by the newly formed outer
retinal plexuses.
Unfortunately, the literature is somewhat confused on the subject of
formation of the deep retinal plexuses in the perifoveal region (10,43,44).
However, the study of developing macaque retina by Provis and Hendrickson
(10) clearly identifies three steps in the formation of the perifoveal capillary
plexus. First, the innermost layer of vessels defines the FAZ; second, the
deep capillary plexuses first form in the vicinity of the optic disc and grow
toward the fovea; and third, the deep capillary plexuses around the fovea
form just before birth and anastomose with the innermost layer of vessels in
the early postnatal period (for a review, see Provis 2001, Fig. 11). The late
completion of the perifoveal anastomosis is consistent with expression of
antiangiogenic PEDF at the fovea, which acts to retard endothelial cell
proliferation, and hence formation of anastomotic branches around the fovea
(25–27). Furthermore, postnatal development of the deep plexus surrounding
the fovea coincides with the maturation of perifoveal cones, followed by
foveal cones, reinforcing the idea that delivery of additional metabolites is
crucial to the final phase of cone maturation.
The significance of these observations is that where normal development
of the retinal blood supply is interrupted—for instance, by prematurity—
there are clear implications for foveal development.
IMPACTS OF PREMATURITY ON
DEVELOPMENT OF RETINAL
VESSELS
One of the major consequences of premature delivery/low birth weight, at
least initially, is reduced expression of vascular endothelial growth factor
(VEGF) in the retina, resulting from exposure to higher levels of oxygen than
would normally occur in utero.
A degree of hypoxia in the retina is normal and drives development of
retinal vessels (45). Under in utero conditions, the unvascularized regions of
the retina express high levels of VEGF under influence of hypoxia inducible
factor (HIF), and VEGF promotes division and migration of retinal
endothelial cells toward the VEGF gradient extending vasculature toward the
ora serrata. This hypoxia is eliminated when infants are placed in oxygen-rich
environments, resulting in slowing or inhibition of further vascularization of
the retina. Uncontrolled growth of retinal vessels can occur when affected
infants are returned to room air promoting the inherently vascular disorder,
retinopathy of prematurity (ROP).
To understand the impact of prematurity on foveal development, we need
first to consider the developmental processes that are active, particularly in
central retina, at a few relevant stages: 24 WG, 28 WG, and 32 WG.
At 24 WG, approximately 50% of the retina is still engaged in
neurogenesis, although not in the region around the posterior pole, including
the emerging fovea where neurons are generated much earlier (46). The
retinal vasculature covers approximately 75% of the surface area of the
retina. At this age, retinal vessels are just approaching the (incipient) foveal
region, and the FAZ is wide open on the temporal side (see Fig. 7-1) (43,46).
The foveal depression has not begun to form. Some of the most vulnerable
infants are born around 24 WG, with increasing numbers of these infants
surviving in the modern intensive care nursery environment, with poor visual
outcomes. This group includes many with a birth weight <1,000 g, who are at
high risk of ROP and three times more likely than those born at full term to
have acuity <6/60, with a range of visual impairments—including strabismus,
amblyopia, and color vision defects—suggestive of anomalies of both the
macula and central visual pathways (12,47,48).
At 28 WG, the FAZ is defined at the GCL/inner plexiform layer interface
by the innermost vascular plexus, but no vessels are present in the deeper
layers around the fovea (43). This stage of human development is
approximately equivalent to the macaque retina at 130 days postconception
(dPC), a time when a greater number of specimens have been studied. At this
age, the foveal depression is easily distinguished, although all of the retinal
layers are still present; there is no evidence of a deep plexus in the immediate
vicinity of the fovea (10), although vessels are present in inner retina within
about a millimeter of the fovea. These features of 130 dPC macaque retina
are consistent with the drawings of Bach and Seefelder (6) of a human fetus
at 28 WG. In both human and macaque, the foveal and perifoveal cones are
cuboidal cells showing little evidence of inner and outer segment formation
or elaboration of their axonal processes. It is likely that slow development of
foveal cones is related to absence of the outer layer of retinal vessels.
By 32 WG, vessels are present in the deeper plexuses within 200 μm of
the FAZ margin in human specimens, although anastomoses between the
different layers are not detected within about 500 μm of the FAZ margin.
These vessels grow very slowly toward the FAZ margin—most likely under
the control of the antiangiogenic factor PEDF, which is expressed at the
fovea at this time. These vessels do not form the anastomoses that will
complete development of the perifoveal capillary bed until the early postnatal
period.
Consistent with the arrival of the outer layer of vessels, cones of the
perifovea and on the foveal rim are distinctly elongated at 32 WG, although
those in the foveal center remain cuboidal, with rudimentary inner segments
evident in histologic sections. Maturation of foveal cones takes place over a
longer period in the postnatal period.
THE FOVEA AT FULL TERM WITH

INSIGHTS FROM OCT
The typical features of a normal full-term retina are shown in Figure 7-4,
which comprises an OCT image along with a photomicrograph scaled to
mimic the aspect ratio (x:y = 1:4.3) of the OCT image (Fig. 7-4A), and the
same histologic section shown at full resolution (Fig. 7-4B). Typically, OCT
scans 6 to 9 mm of the retina centered on the fovea—a considerably more
extensive region of retina than is usually observed in photomicrographs. The
specimen shown in Figure 7-4B comprises three adjacent fields of view
imaged using a 20× objective lens and montaged to show 2.4 mm of retina
centered on the fovea, compared with 6.3 mm of retina represented by the
OCT image (Fig. 7-4A). Within the fovea of a full-term retina, the GCL,
inner plexiform, and inner nuclear layers are still present. Ganglion cells and
bipolar cells complete their centrifugal migrations out of the fovea during the
first several postnatal weeks so that in the adult retina, an accumulation of
cone photoreceptor cell bodies lines the floor of the fovea—save for a few
ganglion cells that might be present in a normal adult fovea. Another
prominent feature at the fovea is the presence of a single band of
hyperreflectivity at the level of the retinal pigment epithelium (RPE),
compared with multiple hyperreflective bands toward the edge of the image
(2.5 to 3.0 mm eccentricity). This difference reflects the relatively immature
state of foveal cones at birth (see also Fig. 7-2).
FIGURE 7-4 Sections through the fovea at 40 WG using
OCT and by histology. A:The layers of the retina are
labeled using conventional terminology (left), and the
hyperreflective bands 0 to 4 are indicated to the right.
Band 0 corresponds to the layer of cone pedicles (the outer
plexiform layer), which is continuous through the fovea in
the newborn retina but absent in the adult due to the
outward displacement of bipolar cells from the fovea.
Bands 1 and 2 are distinct about 1.5 mm or more from the
central fovea; band 1 represents the external limiting
membrane, and band 2 is the more dense part of the inner
segment, the ellipsoid. Bands 3 and 4 each represent
components of the RPE; they are seen separately in the
adult fovea but not at 40 WG, because the photoreceptors
are not fully elongated. The key shows the foveal region of
a 40 WG, stained with H&E and scaled to the same aspect
ratio used for the OCT (1x:4.3y). B:The same section
enlarged to show cellular details. Note the elongation of
cones and Henle fibers on the foveal rim (arrowheads),
compared to the central fovea where the cones are neither
elongated nor packed. GCL, ganglion cell layer; HFL,
Henle fiber layer; INL, inner nuclear layer; IPL, inner
plexiform layer; NFL, nerve fiber layer; ONL, outer
nuclear layer; OPL, outer plexiform layer. (OCT image
courtesy Adam Dubis and Joe Carroll.)
Five bands of hyperreflectivity have been identified in mature outer retina

(32,49). Band 0 corresponds to the layer of cone pedicles, which constitutes
the OPL in the vicinity of the fovea (30). In adults, band 0 is absent from the
fovea owing to lateral deflection of the cone/bipolar synapses. In the example
shown in Figure 7-4A, band 0 can be seen distinctly in the parafovea and in
the central fovea, consistent with the presence of cone synapses onto bipolar
cells that have not yet migrated out of the fovea. Band 1 is the external
limiting membrane (ELM), formed by the outer processes of Müller cells and
positioned immediately sclerad to the photoreceptor cell bodies. In this
example, band 1 is distinct in the perifovea but becomes less distinct with
proximity to the fovea and, within the fovea, merges into the single band of
intense hyperreflectivity. This is due to the very short inner segments of
foveal cones at 40 WG, resulting in a separation of the ELM from the RPE
elements that is not resolvable in the OCT scan (see Figs. 7-2 and 7-4B).
Band 2 occurs at the level of the inner segment. Like band 1, it is distinct in
the perifovea but merges with bands 1 and 3 to 4 closer to the fovea where
the cones have short inner segments. Bands 3 and 4 represent RPE elements
and are not distinct from each other in neonatal macular OCT images.
ANATOMY OF THE FOVEA

ASSOCIATED WITH PREMATURITY
OCT studies find that the FAZ is 20% to 50% smaller in people born
prematurely (31,50) who have a more shallow foveal pit and thicker retina
(33,51–54). There is no simple explanation for this observation, although
some links to retinal vascular development can be identified.
As outlined in this chapter, the present understanding is that
antiangiogenic as well as guidance factors regulate vascular patterning
around the fovea and define the FAZ. We also know that retinal vascular
growth is initially delayed by exposure to an oxygen-enriched environment.
Thus, in premature infants who spend several days or weeks of the early
postnatal period in an incubator, the formation of the perifoveal plexus is
likely to be delayed by an equivalent number of days or weeks. This may
result in the FAZ forming at a later stage (e.g., 30 WG rather than 28 WG),
when antiangiogenic and guidance factors are in natural decline, thus
permitting the perifoveal capillaries to “overgrow,” resulting in a smaller
FAZ.
Presence of a more shallow foveal depression, and a thicker central
fovea, in premature infants is strongly supported by a number of studies
(23,31,33,52–58). OCT demonstrates that increased retinal thickness at the
fovea in individuals born prematurely is related to persistence of the GCL
and inner nuclear layer in the central fovea—indicating reduced centrifugal
displacement of inner retinal neurons out of the central fovea in these cases.
OCT findings also suggest that individuals born prematurely may have
increased thickness of the layer comprising photoreceptor cell bodies (ONL)
and their axons (Henle fibers) (53), although caution should be exercised
because foveal morphology also varies with gender and race (59,60). Further,
it is reported that retinal photoreceptor development may be compromised in
infants born before 32 WG (61).
A long-term follow-up study of 150 children born prematurely between
2001 and 2007 (52) confirms increased thickness of the retina, which is not
associated with functional loss, per se. However, the study reports anomalies
of macular development in approximately one-third of subjects—including a
thinner ONL—and absence or poor development of the fovea (hypoplasia),
which was associated with functional loss (52).
Further investigations that take morphogenetic variations into account,
and which correlate FAZ diameter with both (a) time spent in oxygen
enrichment, and (b) with gestational age at birth, are needed to explore more
fully the variables that contribute to FAZ size and shape. Broadly, we can
predict that because by 28 WG the FAZ is already defined, and the fovea is in
the early stages of development, differences in FAZ size and foveal
morphology in infants born after this time point would be less pronounced
than in those born earlier. Indeed, two studies have now reported that retinal
thickness at the fovea is correlated with gestational age (57,62), including an
analysis indicating that 28.9 weeks is a “critical gestational age” for
premature birth in terms of impact on fine foveal structure (57).
A further impact of prematurity is on thickness of the choroid, mainly in
temporal retina and particularly at the eccentricity of the fovea (63–65). A
thinner choroid at the fovea is correlated with lower birth weight and highly
correlated with incidence and degree of ROP (64,65). There is no clear link
between visual acuity and a thinner choroid (63,65). However, the critical
importance of choroidal oxygen supply to photoreceptors suggests that
different measures of visual dysfunction are needed to fully explore the
impact of a compromised choroid, which could impact photoreceptor
maturation, and make photoreceptors more vulnerable to oxidative damage,
including the impact of diet, smoking, and aging.
Generally, studies to date indicate that the morphologic changes to the
fovea associated with premature birth are not well correlated with visual
acuity. A poorly developed or absent fovea in an infant born prematurely or
at very low birthweight, but not affected by ROP, does not necessarily
correlate with poor visual acuity (31,50,57,62). Some data suggest that
premature birth is associated with changes in photoreceptor responses and
other subtle neurovascular anomalies (66).
Relatively little attention has been paid to the effects that premature birth
may have on the target visual centers in the brain. Expression of both VEGF
and PEDF are modulated by oxygen (67,68), and up-regulation of Eph
receptor signaling pathways has also been identified in models of oxygen-
induced retinopathy (69). We have previously identified high levels of
expression of EphA6 in ganglion cells at the incipient fovea in both fetal
human and macaque retinas (26) and find that expression of EphA6 at the
fovea intensifies and becomes more localized at the fovea after definition of
the FAZ (27). Eph/ephrin signaling has a key role in the mapping of retinal
projections in small mammals (70–72). In humans EphA signaling also
participates in mapping the projection of the retina onto the dorsal lateral
geniculate nucleus (73), which commences in utero but continues into the
postnatal phase of development (74). In this context, expression of EphA6 by
ganglion cells in the immediate vicinity of the fovea is likely to contribute to
or reinforce retinogeniculate mapping strategies, pre- and postnatal in
humans; changes in Eph/ephrin signaling resulting from prematurity may
affect visual function as a result of changes to the retina and targets in the
brain.
The rapid progress of tensor diffusion imaging (TDI) means that it will
soon be possible to visualize major brain tracts and evaluate the effects of
prematurity in the brain, including visual pathways (68,75–77). A key
element of analyses of visual pathways will be evaluation of central retinal
development by OCT, including FAZ diameter and foveal morphology,
where deviations from the norm may serve as correlates of downstream
modifications of brain structure, identified by TDI.
ACKNOWLEDGMENTS
Figure 7-1 is from a retina labeled by Anita Hendrickson, imaged by JP.
Drawings in Figure 7-2 are by Renee Argaet. The OCT image in Figure 7-4
was kindly provided by Adam Dubis and Joe Carroll. Thanks to all those who
have donated fetal tissue for study.
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8
Retinal Development
Thomas A. Reh and Anna La Torre
During the past few decades, the breadth of knowledge in the field of retinal
development has expanded dramatically. In this chapter, we attempt to
provide an overview of some of the main features of retinal development,
both classic and recent, but a comprehensive review of this rapidly growing
field is not possible in a single chapter. The reader is, therefore, referred to a
number of excellent review articles that cover specific areas in more detail. In
addition, in this volume, there are several other reviews that contain relevant
and complementary information about other aspects of eye development.
EMBRYOLOGY OF THE EYE

The development of the eye is a complicated process that requires the
coordination of many tissues. Some of the mechanisms that pattern the
developing retinal domains involve interactions with the surrounding tissues,
whereas others appear to follow intrinsic programs. Not surprisingly,
mutations in a variety of genes lead to congenital defects in ocular
development. Before describing any of these conditions in detail, a short
description of the embryology of the eye is necessary (see Chapter 1).
In humans, prior to overt differentiation, the retina primordium can be
identified in the anterior neuroectoderm as two small grooves on each side of
the developing telencephalon as soon as gestation day 22 (1) (Figure 8-1).
By 25 days of gestation, as the neural folds fuse to form the forebrain, the
optic grooves evaginate to form two distinct outpocketings that are now
called optic vesicles (Figure 8-1). The embryonic ectoderm lies adjacent to
the optic vesicle at the earliest stages of development, but soon a layer of
neural crest–derived cells migrates in and surrounds the developing optic
vesicles. Together with somitomeric (paraxial) mesodermal cells, these neural
crest–derived cells form the periocular mesenchyme and contribute to many
ocular tissues, including the corneal endothelium, the retinal
vasculature/hyaloid artery, the choroid, and part of the ciliary body (2). As
development proceeds, the optic vesicles fold in on themselves to form cup-
like structures, aptly named the optic cups (Figure 8-1). These
morphogenetic processes result in the two-tissue–layered structure of the
optic cup, with the outer layer developing into the retinal pigmented
epithelium and the inner layer giving rise to the neural retina. At this stage,
the lens has formed from a similar invagination from the epidermal/ectoderm
layer. First, a thickened surface ectoderm, the preplacodal region, forms at
the area where the optic vesicles are closer to the ectoderm. As the lens
placode grows and invaginates below the level of the surrounding ectoderm,
the lens pit develops. Finally, by day 33, the lens placode detaches from the
surface ectoderm becoming the lens vesicle, now sitting within the optic cup.
The ectodermal cells restore the external surface by means of proliferation to
give rise to the corneal epithelium. Perturbations affecting the development
of the surrounding tissues have profound effects on retinal development. For
example, failure in normal development of the lens in the condition
congenital primary aphakia is associated with microphthalmia, the neural
retina becoming highly folded within the vitreal cavity, and absence of the
iris (3). Similarly, environmental factors such as deficiency of vitamin A,
radiation, and exposure to pathogens can also lead to severe abnormalities in
ocular development. These fundamental morphogenetic processes occur
between 22 and 35 days of gestation. As noted above, the optic cups begin as
evaginations from the diencephalon and remain connected to the brain
through the optic stalks, which will later form the substrate for the growing
axons of the retinal ganglion cells (i.e., optic nerves). As the optic cup grows
around the stalk, the edges fuse to form the ventral (or embryonic) fissure. In
humans, the ventral fissure normally “closes” around day 33; however,
failure of this fissure to close properly leads to coloboma. The optic fissure
facilitates the entrance of mesenchymal cells into the developing eye
chamber, which will form the hyaloid artery, the main blood supply to the
developing retina and lens. Although the hyaloid artery regresses prior to
birth, a central retinal artery takes its place. In some conditions, the hyaloid
artery fails to regress completely, and this can lead to vitreous hemorrhage
and persistent fetal vasculature (PFV) formerly known as persistent
hyperplastic primary vitreous (PHPV) (4) (see Chapters 2 and 65).
FIGURE 8-1 The retinas of vertebrates develop from the
anterior neural tube. Initially, small grooves are formed,
and, as the neural folds fuse, the optic vesicles form
(green). The optic vesicles become patterned into domains
of retinal pigment epithelium (RPE, yellow), neural retina
(green), and the optic stalk connecting the developing
retina to the brain. Next, a distinct optic cup forms as the
retina folds into the RPE to become a two-layered
epithelium; the most distal parts, near the rim of the cup,
give rise to the ciliary epithelium (gray). The embryonic
eye continues to grow, and even more distally, the iris
develops at the rim of the cup (dark green).
THE EYE FIELD

By the 3rd week of embryonic development, the three primary germ layers—
ectoderm, mesoderm, and endoderm—have formed. During gastrulation,
neural induction transforms a region of dorsal ectoderm into the neural plate,
a thickened epithelium that contains the precursors of the central nervous
system (CNS). The specification of the different regions of the CNS occurs
around the same time, and the current data support a model in which the same
factors that induce neural tissue specify the cells to anterior identities,
telencephalon, and diencephalon. More posterior regions of the CNS are
induced by a second set of factors, sometimes called “transformers,” which
promote more posterior fates. Indelibly labeling the cells in various regions
of the neural plate in amphibians with vital dyes first allowed embryologists
to map the fates of the different regions of the developing embryos, and along
with transplantation studies, they were able to show that the cells of the
neural plate are specified very early in development to give rise to different
parts of the CNS. The region of the neural plate that is specified to produce
the optic cups and their derivatives is called the eye field. The eye field forms
as a single domain at the anterior end of the neural plate but is soon split into
two domains by the action of sonic hedgehog (SHH), a factor released by
cells of the subjacent prechordal mesoderm. Mice lacking the Shh gene
develop cyclopia since the eye field is not repressed at the midline (5). In
humans, mutations in the SHH gene cause congenital holoprosencephaly and
several ocular defects, including cyclopia and synophthalmia (5,6).
The eye field is specified to produce the optic cup by the expression of a
group of transcription factors, known as the eye field transcription factors
(EFTFs). These proteins bind to DNA and activate genes necessary for eye
development (7). The first of these EFTFs to be identified was PAX6, a
member of the paired class of homeodomain proteins; transcription factors of
this family are involved in specifying the different regions of the embryo,
including the neural tube. The eye field cells express PAX6, as do the cells of
the optic vesicle and the cells in the developing lens. Pax6 deletion in mice
causes a failure of eye development around the optic vesicle stage, and PAX6
mutations in humans cause a range of eye defects, including microphthalmia,
Peters anomaly, foveal hypoplasia, and aniridia (8). A homologous gene
called eyeless was identified in Drosophila and is required for eye
development in flies (9): Misexpression of eyeless in regions of the
developing embryo that normally do not express eyeless induces ectopic eyes
in various regions of the fly, like the leg or antenna (10). The fact that
PAX6/eyeless has a conserved role in eye development across millions of
years of evolution has led to the idea that PAX6/eyeless is a master control
gene for eye development (11). Although PAX6 is clearly an important factor
in eye development, it is only one of many homeodomain transcription
factors necessary for normal eye development. The coordinated actions of
many genes together define the eye field identity. There is evidence for
EFTFs acting as complexes and also as a cross-regulatory network. RAX1, a
paired-type homeodomain transcription factor, is required at a very early
stage in eye development; in mice, deletion of Rax nearly fully prevents eye
development (12), and patients with RAX1 mutations exhibit anophthalmia
and other cerebral malformations (13). Experimental overexpression of Rax
induces Pax6 expression, which subsequently induces expression of many
other EFTFs. In fact, overexpression of each of the EFTFs has been done,
and nearly all can induce ectopic eye–like tissues; interestingly, the
coordinated overexpression of all of them (Pax6, Rax1, Six3, Six6, Lhx2,
Nr2e1, and Tbx3) is sufficient to induce ectopic eyes in Amphibia, even in
nonneural areas. These experiments in frogs, together with similar studies in
mice, support the hypothesis that the EFTFs form a cross-regulatory, feed-
forward loop (7). Since similar genes are needed for eye formation in
Drosophila and vertebrates, investigators have suggested that a common
ancestor used some similar pathway (11). Although there are many
differences between the compound eyes of arthropods and the camera-type
eyes of vertebrates, the conservation of the genes that control the
development of photosensitive organs is striking.
PATTERNING THE DOMAINS OF THE

OPTIC CUP
The next stage in eye formation in vertebrates requires several inductive
interactions. The neural retina and the retinal pigment epithelium (RPE) both
originate from the optic vesicles, but they ultimately become quite distinct;
the neural retina is a multilayered structure with neurons, including the light-
responsive photoreceptors, whereas the RPE is a single epithelial layer of
pigmented, nonneural, cuboidal cells. In addition to their morphologic
differences, these two domains can also be distinguished from a very early
stage of development by differences in the expression of distinct transcription
factors. For example, VSX2/CHX10 is expressed specifically in the neural
retina, whereas a different transcription factor, MITF, defines the domain of
the presumptive RPE. These transcription factors play important roles in the
specification of these two distinct tissues: loss-of-function mutations in Mitf
cause the RPE to be respecified into the neural retina, forming a duplicated
retina; by contrast, mutations in Vsx2/Chx10 cause the neural retina to
express RPE genes (for review, see (14)). Other transcription factors are also
critical for RPE and neural retinal development. For example, OTX2 is
expressed in the RPE, and up-regulation of Otx2 to the neural retinal domain,
along with the signaling molecule Wnt, converts the presumptive neural
retina into a second RPE layer (15). OTX2 mutations in humans can lead to
severe ocular malformations, including clinical anophthalmia,
microphthalmia, and early-onset retinal dystrophy (16,17).
Besides the interactions within the optic vesicles, the surrounding tissues,
as well as domains of the neuroepithelium immediately adjacent to the
vesicles, play important roles in patterning of the retina and RPE, the ciliary
epithelium and iris. If the optic vesicles are experimentally isolated from the
surrounding tissues, they fail to form complex eyes and instead develop
primarily as neural retina. Likewise, transplantation experiments have shown
that if an optic vesicle is placed in a position in an embryo adjacent to the
developing hindbrain, a second neural retina is formed from the presumptive
RPE layer. Together, these data revealed that the development of the RPE is
regulated by signaling molecules secreted by the mesenchymal (neural crest–
derived) tissue that surrounds the optic vesicle. Accordingly, the addition of
the signaling factor Activin (18) or other members of the TGF-beta
superfamily (19) to cultures of avian embryo optic vesicles can substitute for
the extraocular tissue to promote RPE development. SHH is also involved in
the specification of the RPE domain, specifically in the ventral optic vesicle
as inhibition of Shh in frog and mouse embryos causes defects in ventral RPE
formation (20).
The specification and growth of the neural retina domain also require
several signaling molecules. Fibroblast growth factors (FGFs) are important
for the proper development of the neural retina. Several different FGFs are
expressed in the lens ectoderm and the neural retinal domain (21,22).
Overexpression and inhibition experiments in zebrafish, chick, and mouse
embryos support key roles of FGFs in patterning the neural retinal domain.
For example, the addition of exogenous FGF to chick embryo optic vesicle
cultures induces the presumptive RPE layer to become a second neural retina
(22), whereas blocking FGFs with antibodies in similar cultures prevents
neural retina formation. Inhibition of FGF signaling in mice by conditional
deletion of Shp2, which is a necessary component for FGF signal
transduction, causes the affected region of the neural retina to develop as
RPE instead (23). The model of ocular patterning that has emerged from
these, and other studies, is as follows: FGF promotes neural retinal
specification in the optic cup, in part by inducing VSX2. VSX2, in turn,
represses MITF in the retinal domain. The presumptive RPE is then specified
by several signaling molecules, including SHH, WNT, and Activin/BMP,
some of which are derived from the surrounding mesenchyme (Activin,
Indian hedgehog), and some of which (SHH, BMP7) come from within the
neuroepithelium (14).
HISTOGENESIS OF THE RETINAL

CELL TYPES
After the primary domains of the optic vesicle have been specified, their
growth accelerates, particularly in the neural retina. The neurons and Müller
glial cells of the retina are generated by progenitor cells through multiple
mitotic cell divisions. These progenitors divide rapidly and, in humans,
produce hundreds of millions of cells between the 10th and the 24th
gestational week. In the early part of retinal development, there is a nearly
exponential increase in cell numbers, reflecting the symmetric cell divisions
of the progenitors; however, some of the cell divisions also produce
postmitotic neurons, even at very early stages of retinal histogenesis. The
different types of cells in the retina are produced by the progenitor cells in an
unvarying sequence. The regular “birth order” of retinal cells was first
described by Richard Sidman, using the 3H-thymidine “birthdating”
technique (Figure 8-2) (24). The production of the different types of retinal
cells can be divided into two phases. In the early phase, most of the ganglion
cells, cones, and horizontal cells are produced, whereas most of the rod
photoreceptors, the bipolar cells, and the Müller glial cells are produced in a
second phase (25,26). Amacrine cells do not fall as neatly into one or the
other phase. Although there is clear regularity in histogenesis in the retina,
there is also a central-to-peripheral gradient, such that peripheral retina may
still be producing the early cell types (i.e., ganglion cells) while central retina
is producing later cell types.
FIGURE 8-2 Each retinal cell type is generated over a
slightly different time course during retinal histogenesis.
The graph at the top shows the running total of each cell
type as a function of age in the rat. The lower figure shows
schematically the addition and integration of new cells
during the histogenesis of the retina at three different ages,
corresponding to the chart of birthdating above. (A: From
La Vail MM, Rapaport DH, Rakic P. Cytogenesis in the
monkey retina. J Comp Neurol 1991;309(1):86–114.
Copyright © 1991 Wiley‐Liss, Inc. Modified by
permission of John Wiley & Sons, Inc.)
The progenitor cells of the retina are for the most part able to generate all the
different types of cells and, therefore, have been called multipotent
progenitors (27,28). The multipotential nature of these cells has been
demonstrated by tracking the lineages of individual cells, after infection of
progenitors with a retrovirus containing a reporter gene (27,28) or following
injection of the cells with a permanent dye (29,30). More recently, it has been
possible to track the lineages of progenitors that express specific transcription
factors using inducible Cre-recombinase methods (31,32). These studies have
shown that progenitors can produce multiple different types of retinal
neurons and Müller glia up to their last cell division; however, there do
appear to be at least three distinct types of progenitors in the mouse: (a)
progenitors that express Ascl1 can produce moderate-sized clones with all
types of retinal neurons, except ganglion cells (Figure 8-3); (b) Neurog2-
expressing progenitor cells produce clones with no more than two cells. For
that reason, these progenitors are likely in their last cell division and in
contrast with Ascl1-expressing progenitors; Neurog2 cells can make ganglion
cells; and (c) true multipotent progenitor cells can make large clones
containing all cell types (31).
FIGURE 8-3 Tracking the lineages of retinal cells shows
that there are several types of multipotent progenitor cells
that underlie cellular diversity. The schematic diagram
shows examples of lineages of (1) a retinal progenitor that
expresses Ascl1 (green) that can produce a clone of cells
that include all cell types (HORizontal cells, CONes,
AMAcrine cells, BIPolar cells, ROD photoreceptor cells
and Müller glia), except ganglion cells, which are
produced by (2) an earlier progenitor (black) that is
competent to produce ganglion cells, and then produce an
Ascl1+ progenitor. The diagram also shows the results of
tracking the Neurog2-expressing progenitor lineage in red.
Neurog2 is expressed in both the Ascl1+ and Ascl1−
progenitors but only at the last division of the progenitor
cells. The panel on the right shows an example of a clone
containing rods (r), a bipolar cell (b), and an amacrine cell
(a) derived from the Ascl1 lineage.
The fact that the different types of retinal cells are generated in a sequence
has led to several hypotheses about the nature of the mechanisms that
underlie this phenomenon. One proposal is that each cell type induces the
progenitor cells to produce the cell type that follows in the sequence. In this
model, the first cell type generated by the progenitor cells, the retinal
ganglion cells, produces a signal to the progenitor cells to direct them to
begin producing horizontal cells, the next cell type in the sequence; the
horizontal cells in turn signal to the progenitors to instruct them to make cone
photoreceptors, and so on. In other words, this model proposes that
progenitors can be influenced by their surrounding microenvironment to form
the types of cells they produce. One of such factors is the signaling molecule
SHH, mentioned earlier in the chapter in the context of the eye field. SHH is
expressed by the retinal ganglion cells, regulates the rate of proliferation and
timing of cell cycle exit of the progenitor cells, and consequently determines
the number of ganglion cells in the retina (33). An alternative model to
explain the sequential production of the different retinal cell types proposes
that the progenitors undergo a progressive change in their “internal state,”
like a clock ticking through the different cell fates (34,35). In this model, the
progenitor cells progressively change in their competence to produce
different types of cells in a cell-autonomous manner; their initial state has a
bias for producing retinal ganglion cells, but after a day, some fraction of the
progenitor cells shift their competence to become biased to produce later cell
types, for example, horizontal cells, followed by cone photoreceptors, and so
on. Cell culture experiments provide support for this type of mechanism;
progenitor cells isolated from the early stages of retinal development
predominantly differentiate into early cell types (36), whereas culturing the
progenitor cells with retinal cells from later stages of development biases
them to adopt later cell identities (37–39). The molecular mechanism
underlying such a progressive shift in competence could be a cascade of
transcription factors, and Ikaros genes have been shown to be involved in this
process (40). Recent data have also implicated miRNAs in regulating the
developmental timing. Deletion of the DICER1 gene, which is necessary for
miRNA production as well as the inhibition of 3 miRNAs (Let-7, miR-9, and
miR-125b), leads to a failure in the progenitor cells to progress beyond the
early state, and they continue to generate early cell fates, like ganglion cells,
cones, and horizontal cells for the entire period of retinogenesis (Figure 8-4)
(41–43).
FIGURE 8-4 MicroRNAs are required during retinal
development to allow the progenitors to make late retinal
cell types. When the Dicer gene is knocked out in mouse
retinas, the progenitors overproduce the early-generated
ganglion cells (Brn3+) and do not progress to the state
where they can generate late cell fate types like rods,
bipolar cells, and Müller glia.
The mechanisms by which the progenitors progressively change in their

competence are not understood, but quite a bit is known about the
transcription factors that are necessary for the differentiation of different
types of retinal cells. One transcription factor discussed earlier in the chapter
as important in the eye field, PAX6, is also critical for cell fate determination
later in retinal development. Conditional deletion of Pax6 in retinal
progenitors causes a dramatic restriction in their competence such that they
only produce amacrine cells (44). Conversely, deletion of the transcription
factor FoxN4 produces the complementary result; these mutant progenitor
cells produce all cell types except amacrine cells (45). Other transcription
factors expressed by progenitors also play important roles in the production
of cell diversity in the developing retina. For example, deletion of the gene
Atoh7 (Math5) causes a dramatic reduction in ganglion cell production in
mice (46), and individuals with disrupted ATOH7 expression have
nonsyndromic congenital retinal nonattachment (NCRNA (47)), a severe
form of blindness involving the lack of optic nerves and abnormal retinal
vasculature. Similarly, in mice, conditional deletion of Otx2, another EFTF,
leads to a loss of bipolar and photoreceptor cells (48–50); the transcription
factors Ptf1a, NeuroD1, and Math3, expressed in amacrine cells, are
necessary for amacrine cell development (45); the Nrl transcription factor is
required for rod photoreceptor development, and loss of this gene causes the
progenitor cells to produce as short-wavelength–sensitive cones instead (51);
mutations in the Chx10/Vsx2 gene result in an absence of nearly all bipolar
cells (52); conditional deletion of the Prdm1/Blimp transcription factor
switches the photoreceptors to bipolar cells (53); and deletion of Ascl1, a
bHLH transcription factor, results in an increase in the Müller glia and a
concomitant reduction in late-born retinal neurons: bipolar cells and rods
(38,54,55). The way in which these transcription factors interact remains a
significant challenge for the field.
Other cells that are resident in the retina, but not derived from the
multipotent retinal progenitors, include astrocytes, microglia, and vascular
endothelial cells. Microglia in the CNS, including the retina, are derived from
the mesodermal lineages during embryonic development, not from the neural
tube (56,57). Microglia are highly motile, phagocytic cells and are
responsible for digesting those retinal cells that have undergone apoptotic cell
death. Astrocytes are the second type of glial cell present in the retina, but not
derived from the retinal progenitor cells. These cells migrate along the
developing optic nerve from the brain and, once they have entered the retina,
migrate across the retinal surface and, in some species, along with the
vascular endothelial cells (58,59).
INNER RETINAL DEVELOPMENT

Once they have been generated, the retinal cells must migrate to their proper
positions, project axons, and dendrites and make synaptic connections with
the other neurons. These steps are critical for proper organization and
function of the retina, but not as much is known about the molecular
mechanisms that control them. The ganglion cells are the first to be generated
and also the first to differentiate. Following the last mitotic division at the
scleral surface, the ganglion cells migrate toward the vitreal surface to finally
form the ganglion cell layer (GCL). As they migrate, the ganglion cells take
on morphologic features characteristic of their cell type. They elongate axons
from their vitreal processes even while still migrating (60,61). In the
developing cerebral cortex, neuroblasts migrate along radially arranged
progenitor cells, and much is known about the molecules necessary for their
migration (62). Although similar molecules might mediate ganglion cell
migration, there are no radial glial cells in the retina. Müller cells are not
generated before ganglion cell migration. Instead, the newly generated
ganglion cells may be using the surrounding retinal progenitor cells as a
scaffold for migration (60) or migrate using a different mode of migration
called somal translocation, in which the newly developed postmitotic
ganglion cells extend a cytoplasmic process toward the vitreal surface while
maintaining contact with the apical or outer surface (63). The cell body then
translocates as the apical process slowly retracts until the cell reaches its final
position in the GCL.
As the ganglion cells reach the GCL, they begin to extend dendrites and
form the inner plexiform layer (IPL). Their development has been well
characterized in several species, including primates (64,65). The initial
dendrites have a simple morphology, with only a single primary filopodial
process. Soon, these simple processes begin to elaborate, forming a complex
dendritic arbor. The dendritic arbors of ganglion cells can initially span the
width of the IPL; however, in some types of ganglion cells, the dendrites
grow only in specific sublaminae from the outset (66). The dendritic arbors
are remodeled and pruned as the ganglion cells mature and their dendritic
processes typically become restricted to one sublamina, depending on the ON
or OFF subtype of the ganglion cells. As the retina continues to expand,
primarily by stretch, the overall arbor size of the ganglion cells expands to its
mature extent.
The growing ganglion cell dendrites interact with the amacrine cells that
have completed their migration to the INL. Amacrine cell migration is
orchestrated by Fat3, an atypical cadherin, and the cytoskeletal regulators
Ena/VASP (67). The subsequent growth of amacrine and ganglion cell
dendrites is regulated by some adhesion molecules, particularly N-cadherin
and NCAM (68). Retinal neurons grown on specific CAM substrates display
characteristic morphologies depending on the type of neuron and the specific
adhesion molecule used for the substrate (69). Cells grown on N-cadherin
formed highly branched arborizations similar to those observed in vivo. By
contrast, cells grown on L1 substrate extended axon-like processes but few
dendrites. Nevertheless, these more generally expressed generic CAMs
cannot account for the highly specific morphologies and spatial arrangements
of the many different types of retinal neurons. More recently, other adhesion
molecules with more restricted expression have been implicated in the
development of dendrites in the retina. Dscam (Down syndrome cell adhesion
molecule), Dscaml, Sidekick-1, and Sidekick-2 are all required in the
developing chick retina to specify the dendritic position within sublaminae in
the IPL (70). In the mouse retina, the axon guidance molecules Sema6A and
PlexinA4 are necessary for stratifying the dendrites of specific types of
amacrine and ganglion cells to particular sublaminae of the IPL (71).
The dendritic arbors of retinal cells are often organized for maximum
coverage and minimal overlap. This phenomenon is called “tiling.” For
ganglion cells, each subclass “tiles” the retinal surface (72). Experimentally
depleting a region of the retina of ganglion cells causes the neighboring
ganglion cells to sprout dendrites into the depleted areas. The converse
experiment, increasing the density of ganglion cells by preventing their
normal developmental cell death, causes them to have smaller dendritic
arbors than normal (73). Together, these results support the idea that
interactions among each subclass of ganglion cells regulate dendritic growth,
although the molecules that underlie these interactions are for the most part
unknown. An early idea was that the ganglion cells compete for a dendrite-
promoting factor; when their density is reduced, the remaining ganglion cells
have access to more of the factor, and when their density is greater than
normal, the cells receive proportionally less factor. An alternative idea is that
ganglion cell dendrite growth is limited by “contact inhibition,” where direct
contact between two cells of the same subclass causes their growth cones to
collapse and stop extending. Evidence for contact inhibition comes from an
analysis of the phenotype of Dscam and Dscaml mouse mutants. Although
these molecules are not required for the sublaminar specificity of the
dendrites in mice, they are critical for tiling of the dendrites and an orderly
arrangement of the cell bodies of some ganglion cells. The ganglion cells of
Dscam knockout mice form large clumps instead of being distributed across
the retina, and their dendrites are fasciculated together in bundles rather than
as tiles on the retinal surface (74).
The bipolar cells are the last neurons generated by the progenitor cells.
After bipolar cells are born, they migrate toward the vitreal surface to then
backtrack their path toward the apical side using somal translocation and
maintaining neuroepithelial-like processes that extend the whole thickness of
the retina (75). The final position of the bipolar cells is regulated by a
complex signaling pathway involving Reelin/Dab1 and the E3 ubiquitin
ligase CRL5 (76).
The next stage in the development of the plexiform layers reflects the
dendritic growth and synaptogenesis of the bipolar cells, amacrine cells, and
ganglion cells. Conventional synapses between amacrine cells and ganglion
cells tend to develop before the ribbon synapses form between bipolar cell
axons and the ganglion cell dendrites. Moreover, the initial synapses between
amacrine and ganglion cells mediate horizontal interactions among inner
retinal neurons, which are necessary for ganglion cell axons to segregate into
eye-specific domains in the lateral geniculate nucleus (LGN). Shatz, Wong,
and collaborators demonstrated in ferret retina that waves of spontaneous
activity spread between the ganglion cells before most of the bipolar cells
have been born (77). Thus, horizontal spread of activity predominates in the
early retina, whereas only later in development when bipolar and
photoreceptor synapses are formed, does the vertical connectivity of the
mature retina develop. The primate fovea differs from the rest of the retina in
this respect. In this specialized region, where there are no rod photoreceptors,
the ribbon synapses between ganglion cell dendrites and cone bipolar cells
are the first synapses to form in the IPL (78).
As the functional circuits develop between the retinal cells, synaptic
transmitters and receptors begin their expression (for more in-depth review,
see (79,80)). The primary neurotransmitters of the retina, glutamate, gamma-
aminobutyric acid (GABA), and glycine are present in specific types of
retinal neurons prior to the development of morphologic evidence of
synapses. In addition to their production of neurotransmitters and receptors,
many different types of channels are expressed in retinal cells within a short
time after their production by the progenitor cells. Ganglion cells in the
mouse express their characteristic Ca2+, Na+, and K+ channels by E15, within
a few days of their genesis. Expression of ion channels and neurotransmitter
receptors in very immature neurons has led many investigators to suggest that
neural activity may shape the formation of specific connections in the circuit.
Genetic inhibition of bipolar cell transmitter release does not alter the overall
morphology of the dendrites of ganglion cells (81), but neurotransmitters
appear to play a role in regulating dendrite growth in the outer plexiform
layer (OPL) (see below).
GANGLION CELL DEATH

Many of the neurons that are produced in the CNS during development are
later eliminated by a process called programmed cell death or apoptosis. In
the retina, programmed cell death is best characterized in the ganglion cells
(82), although other cell types also undergo this process (83). Approximately
half of the ganglion cells initially produced by the progenitor cells
subsequently die. Ganglion cell death occurs when their axons reach their
targets in the brain. This loss of neurons is thought to provide a mechanism
for numerical matching of pre- and postsynaptic populations. The neurons in
the targets presumably produce sufficient trophic factor to support a limited
number of ganglion cells. It has also been proposed that different targets
produce unique factors to provide a mechanism for maintaining specific
connections (84).
The neurotrophins, including nerve growth factor, brain-derived
neurotrophic factor (BDNF), neurotrophin 3 (NT3), and NT4/5, are thought
to be important in ganglion cell survival. BDNF and NT3 can promote
ganglion cell survival in several in vitro and in vivo assays and are both
expressed in their targets in the brain (85). Both BDNF and NT3 promote
ganglion cell survival in cell cultures of retinal cells or when injected into
ganglion cell targets in the brain. Receptors for BDNF, NT3, TrkB, and TrkC
are expressed by the ganglion cells (86). It is thought that as the ganglion cell
axons reach their targets, the supply of neurotrophins is limited, and many
will insufficiently activate their Trk receptors and undergo apoptosis.
Although this scheme fits with currently available data, other evidence
implicates additional growth factors in regulating ganglion cell number
(87,88). Transcription factors are also important for ganglion cell survival.
The Brn3 genes, members of the POU-domain class of transcription factors,
are expressed by the ganglion cells. Targeted disruption of the Brn genes in
mice causes the loss of nearly all of the retinal ganglion cells (89–91). The
Brn genes may function by regulating the expression of the neurotrophins and
the Trk receptors (89,92).
PHOTORECEPTOR DEVELOPMENT
The photoreceptors develop somewhat later than the ganglion cells in most
species and somewhat ahead of the bipolar cells and Müller glia. A few key
points of photoreceptor development are reviewed here. Our knowledge of
photoreceptor development comes from studies of a wide variety of
vertebrates from fish to humans, but studies of the rhesus monkey have been
particularly informative of events in human photoreceptor development.
Rhesus monkeys have a fovea similar to the human, whereas the mouse,
another model organism for retinal developmental studies, does not have a
fovea and has relatively fewer cones. Thymidine birthdating studies have
shown that foveal cones are born between gestational days E38 and E50 in
the monkey. Although the cone photoreceptors are produced by the
progenitor cells prior to the rod photoreceptors, rods express rhodopsin
before the cones express their characteristic opsins (93,94); the first foveal
cones are generated on fetal day 38 and do not express S-opsin until fetal day
75. The delay between birthdate and opsin expression occurs in other
vertebrates as well. This phenomenon suggests that the regulation of
specification to cone fate and the factors that regulate opsin expression are
distinct; data in mice support this idea.
As noted previously, retinal cells are arranged in mosaics that cover the
retinal surface like tiles. The mosaics of cone photoreceptors can reach a high
degree of order in some species with almost a crystalline array of the
different subtypes. The factors that control the formation of these mosaics are
still unknown, although some of the same types of CAMs important for inner
retinal neuron organization may also function in the formation of the cone
mosaics. In primates and mice, the first cone photoreceptors to express their
opsins are the S-opsin–expressing cones (93). S-opsin expression begins in
the central retina and sweeps to the periphery. After much of the retinal
surface contains S-opsin–expressing cones, the L-/M-opsin cones in the
central retina begin to express opsin (95) and also sweep from central to
peripheral retina following the S-opsin cones (96).
The molecular mechanisms that control the expression of opsins and
other aspects of photoreceptor differentiation are understood to some extent;
several different transcription factors play critical roles at each stage of
development. OTX2, a transcription factor described earlier in the context of
eye field formation, is expressed in photoreceptors very early in their
specification (48,49). Deletion of Otx2 in retinal progenitors in mice prevents
them from adopting the photoreceptor fate, and instead they develop into
amacrine cells. OTX2 then induces the expression of cone–rod homeobox
gene (CRX) in the early developing photoreceptors, which is required for the
expression of photoreceptor-specific genes, like the opsins (97). The
immature proto-photoreceptors are then specified to become either rods or
cones; if the cells express neural retina leucine (NRL) zipper transcription
factor, they differentiate as rods (51). Photoreceptors that do not express NRL
differentiate as cones; the specification of cone subtype is determined by two
additional transcription factors, thyroid hormone receptor-β2 (TRβ2) and
RXR-gamma (98,99). These transcription factors act as heterodimers (or
possibly homodimers) and promote M-opsin expression but inhibit S-opsin
expression. TRβ2 knockout mice do not have M-opsin–expressing cones but
only contain S-opsin cones (99). RXR-gamma knockout mice contain cones
that all express both M-opsin and S-opsin. In vitro studies have identified
several factors that regulate photoreceptor development. Most of the studies
have focused on rod photoreceptors. Rod photoreceptors do not develop well
in low-density cultures, but the addition of specific signaling molecules
promotes rhodopsin expression in vitro (14). These soluble signaling factors
can be used to promote photoreceptor development in embryonic stem cell–
derived retinal cells (100,101).
After the cones and rods have begun their differentiation, they begin to
form synapses with the horizontal cells and bipolar cells in the OPL. In the
monkey, for example, the first cones have begun to make synaptic
connections with bipolar cells in the OPL by E55, more than 2 weeks after
they had been generated. The molecules that regulate synapse formation
between the photoreceptors and their targets are not known, but there is
evidence that the dendrites in the OPL interact to define their receptive field
extent. The dendritic extent of horizontal cells is a function of the ratio of
rods to cones. Experimentally reducing cone number during retinal
development in mice increases the branching of horizontal cell dendrites.
Ribbon synapses, synaptic specializations involving rod and cone pedicles
with horizontal and bipolar cell dendrites, require normal activity in the cells:
If phototransduction is blocked in cones during the period of ribbon synapse
formation, the cone bipolar cells make synapses with rods instead of cones
(102). Conversely, inhibiting transmitter release from rods during this period
causes the rod bipolar cells and the horizontal cells to produce aberrant
processes that extend into the outer nuclear layer (ONL) and form synapses
with cones (103,104).
FOVEAL DEVELOPMENT
The differentiation of the fovea is among the last major events in retinal
development. The fovea, a small, avascular depression, contains only cone
photoreceptors and Müller glia. Foveal development in primates has been
extensively characterized (105–108). Initially, among the thickest regions of
the retina, the fovea becomes transformed into a pit or depression by cell
migration and deformation. Between the 24th and 26th fetal week in humans,
the ganglion cells begin to migrate away from the center of the fovea, and the
amacrine cells and bipolar cells follow. By the time the fovea forms, the
synapses have already been formed in the plexiform layers; therefore, since
the cone photoreceptors remain in the fovea while the inner retinal neurons
migrate away, the processes of the bipolar cells become extremely elongated.
At birth in humans, the GCL and INL constitute only a single layer of cells.
However, after birth, the inner retinal neurons continue to migrate radially
from the fovea and pile up around it, whereas the photoreceptors become
more concentrated and the ONL increases in thickness. By 4 years of age in
humans, there are six to seven layers of cone nuclei in the foveal ONL. The
molecular mechanisms that drive this very dramatic rearrangement of cells
are still unknown (see also Chapter 7).
CONCLUSIONS
The development of the retina is a remarkably complicated process, and
although quite a lot is known about the cellular and molecular events that
occur during histogenesis and early specification, there are many gaps in our
knowledge. In other aspects of development, like the formation of specific
connections among the retinal cells, very little is known. Nevertheless, there
has been amazing progress in the past 20 years, and the findings have had and
will continue to have an impact on our understanding of developmental
diseases of the eye. Moreover, technologies for producing retinal cells and
organized retinal tissue from human embryonic stem cells and induced
pluripotent stem cells combined with experimental animal studies of retinal
development have the potential to inform both normal retinal development
and developmental pathologies.
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SECTION II
Assessment and Rehabilitation of
Visual Function In Pediatric Retinal
Condition
9
Assessment of Vision and Retinal
Function in Infants and Children
Anne B. Fulton, Ronald M. Hansen, Bridget J. Peterson, Laura
Bagdonaite-Bejarano, and James D. Akula
This chapter presents key features of development of normal visual and

retinal function and illustrates their application to characterization of the
pediatric retina with disease. The purpose of the functional measures is to
contribute to the diagnosis, to define the severity of the patient’s deficit, and
to track changes over time and in response to therapeutic interventions. Thus,
assessment of vision and retinal function supports good management of
infants and children with retinal disease, including preparation for definitive
therapy as new gene-based therapies become available. Furthermore, it is the
combination of assessments of function, retinal structure, and clinical features
that has led to new, fundamental knowledge about pediatric retinal disease
and will continue to do so.
The numeric data obtained by measures of acuity and retinal function
provide the basis for comparing a pediatric patient’s visual and retinal
function to normal function for age as well as monitoring an individual
patient for change over time. Significant developmental changes in visual
acuity and retinal function normally occur during infancy. Numeric
information about the parameters of normal function, including the variability
of these measures, is needed for interpretation of a young patient’s data.
Selected for presentation in this chapter are the electroretinographic (full-
field ERG; multifocal ERG) and psychophysical (visual acuity; dark-adapted
visual threshold [DAT]) functions suitable for assessment of infants and
children with retinal disease. The development of normal function is outlined.
Clinical applications are illustrated by data from patients with retinopathy of
prematurity (ROP) and from those with the inherited retinal disorders: Leber
congenital amaurosis (LCA), achromatopsia (ACHM), and congenital
stationary night blindness (CSNB).
The patients’ results are presented against templates for normal. See also
the appended “Atlas of ERG Responses” for sample ERG records.
Infants and children with retinal disease are brought to ophthalmologists
because of subnormal visual behavior. Many present with nystagmus,
particularly in early infancy when the oculomotor system is immature.
Anxious parents and physicians share concern about the most basic dilemma:
Is this child blind? Is this child’s visual impairment due to disease of the eye,
the brain, or both? Early in the evaluation, imaging of the brain may be
justified to identify, or exclude, brain conditions that put the patient at high
risk for morbidity and mortality.
DEVELOPMENT OF NORMAL VISUAL

ACUITY
Born of a quest to understand development of the human brain and vision,
investigators launched studies of infants’ vision, often backed up by detailed
studies of the immature simian visual system. The most common of these
characterized the maturation of human visual acuity using behavioral
procedures. In this chapter, the results of many studies of acuity have been
compiled to provide the normal template against which data from infant and
child patients can be compared. Remarkably, the compiled data (Figure 9-1)
show a continuum of acuity development from infancy to adulthood, through
grating acuity in infancy, obtained using preferential looking procedures and
grating stimuli in infants (1–3), matching tasks and a few simple symbols in
young children (1,4) and naming letters in older children and adults (1,5–9).
Data from each of these studies cited in Figure 9-1 have been used to
calculate the limits of normal from infancy to adulthood. Knowledge of these
limits facilitates comparison to patients’ data, illustrated later in this chapter
by data from children with LCA, achromatopsia, and CSNB.
FIGURE 9-1 Development of normal visual acuity. Mean
monocular acuity obtained at the indicated ages using the
Teller Acuity Cards (triangles) (1–3), HOTV letters
(squares) (1,4), and ETDRS letters (circles) (4–8) are
shown. The smooth red curve is a logistic growth curve fit
to these data.
Visual acuity, the most frequently measured visual threshold, is 20/20 in a

healthy adult. Correct identification of a letter with a gap width of 1 minute
of arc at 20 feet is designated 20/20 acuity; this is equivalent to 1.0 decimal
or 0.0 log minimum angle of resolution (MAR) (Table 9-1). At age 10 weeks,
an average normal acuity is 20/300, well below that of the adult (2). During
infancy and early childhood, acuity matures (Table 9-2). The minimum
physical attribute of the stimulus that is reliably detected (MAR) decreases
during development.
Table 9-1 Acuity conversion metrics
Cpd, cycles per degree; Dec VA, decimal visual acuity; MAR, minimum angle of resolution; min arc,
minutes of arc.
Table 9-2 Visual acuity development: mean acuity

and 95% prediction interval
aThe prediction limit is the range of acuity values expected in 95% of the normal population.
Normal, 20/20 acuity, is presumed to be mediated by the cones and cone-

mediated pathways in the central retina. The anatomy of the infant’s fovea
predicts better acuity than is measured (10). Therefore, neural processes
central to the photoreceptors must limit the measured acuity (10,11). Shortcut
procedures, such as the Teller Acuity Cards, are available for day-to-day
clinical testing of infants and young children and have been used in
multicenter studies (12). Other tests using a few symbols and allowing
match-to-kind procedures for testing young children may be used.
DEVELOPMENT OF NORMAL
RETINAL FUNCTION
Noninvasive studies of normal retinal function have linked developmental
changes in function to molecular events in the retinal cells. In pediatric
patients, delineation of retinal activity, on the one hand, yields insights into
the action of protein products of genes and, on the other hand, may explain
visual behavior. Coupled with clinical examination, including
ophthalmoscopy, fundus photography, and retinal imaging, studies of
function help to secure diagnoses, delineate severity of disease, track the
course of retinal disease, and monitor the response to interventions.
Retinal function depends on the physical properties of the photoreceptors,
the retinal cells proximal to the photoreceptors, and the neural connections
among the cells. The properties of retinal cells change during postnatal
development. Electroretinographic and psychophysical measures of retinal
function that are constrained by physical immaturity provide a body of
information about the development of human retinal processes. Age-specific
values for parameters of normal retinal function against which a patient’s
data can be compared are available. ERG and psychophysical procedures are
used to obtain these parameters.
Electroretinography
Full-Field ERG
ERG responses to full-field stimuli assess activity in the retina as a whole.
Full-field stimuli are designed to stimulate all retinal cells. The ERG is
initiated when the stimulating flash of light falls on the retina and the visual
pigment in the photoreceptor outer segment absorbs quanta, thus activating
the biochemical phototransduction cascade. In the rods, the visual pigment is
rhodopsin. The first few milliseconds of the a-wave depend on this
biochemical activity.
Application of a mathematical model of the activation of
phototransduction (13,14) to the a-wave allows one to draw inferences about
the sensitivity and saturated amplitude of the photoreceptor response; this
model is applicable to infants as well as adults (15–18). Rod photoreceptor
sensitivity and saturated amplitude, derived from the a-wave, are low in
young infants. For example, at age 10 weeks postterm, both rod
photoresponse parameters, sensitivity and saturated amplitude, are
approximately half the adult values (15). In normal development, the rod and
rod-driven a- and b-wave parameters are scaled by the rhodopsin content of
the retina (15).
Sample ERG records obtained in scotopic conditions from a dark-adapted
10-week-old infant (awake) and an adult, recorded using Burian-Allen
bipolar corneal contact electrodes, are shown in Figure 9-2. The adult
responds to dimmer stimuli, and the maximum amplitude of the a-wave and
the b-wave is larger than in the infant (15).
FIGURE 9-2 Scotopic ERG responses from normal
subjects. Dark-adapted ERG responses to a series of short-
wavelength (blue) flashes are shown for a 10-week-old
infant (upper left) and an adult (upper right). Stimulus
strength in log scotopic troland seconds (scot td s), a unit
of retinal illuminance, is indicated to the left of the traces;
responses to dim (bottom) and then to increasingly bright
(top) stimuli are shown. This unit takes the area of the
subject’s pupil into account. The calibration bar pertains to
both infant and adult waveforms. In the lower panels, b-
wave amplitude is plotted as a function of stimulus
strength for the infant (left) and adult (right). The smooth
curves plot the equation V = VMAX [I/(I + σ)] where
VMAX is the saturated b-wave amplitude, the flash that
produces a half VMAX, and I the stimulus. In each panel,
the VMAX and log σ are indicated by arrows.
The rod photoreceptors send signals to the rod-driven ON bipolar cells. In

clinical work, much information is derived from the b-wave that is generated
by bipolar and other postreceptor retinal cells (19–21). The stimulus–
response relationship for the b-wave in both infants and adults (Figure 9-2,
lower panels) is summarized by the equation:
V = Vmax (I/(I + σ))
where V is the amplitude of the b-wave and I is the strength of the stimulus,
specified in scotopic troland seconds (scot td s). VMAX is the saturated b-
wave amplitude; σ is the stimulus evoking a response with half the amplitude
of VMAX. Thus, 1/σ is an index of sensitivity (22). Logistic growth curves
summarize the developmental changes in log σ and VMAX (Figure 9-3).
FIGURE 9-3 Development of normal scotopic ERG b-
wave. Logistic growth curves summarizing the
developmental changes in parameters log σ (upper panel)
and VMAX (lower panel) are shown (18). Each parameter
is plotted as a function of age. The curves were fit to data
from 136 healthy control subjects; the 50th percentile is
shown as a heavy, solid red line. After infancy, there is
little change in these parameters.
Initiation of phototransduction in the cones is similar to that in the rods;

however, a few milliseconds after onset of the cone a-wave, activity in the
postreceptor cells intrudes and contributes to the observed a-wave. Modeling
of the infants’ and adults’ cone-mediated responses to full-field stimuli shows
that cone sensitivity and saturated amplitude are approximately 70% of the
adult values at age 10 weeks (17).
The cone-mediated b-wave results from the interaction of responses from
both ON and OFF bipolar cells (23,24). Activity of the bipolar and other
second- and third-order neural cells in the postreceptor retina contributes to
the cone mediated b-wave (19–21). In photopic conditions (Figure 9-4), the
infant’s cone and cone-mediated responses are detectable throughout the
stimulus range, and maximum amplitudes in infants and adults are similar.
FIGURE 9-4 Photopic ERG responses from normal
subjects. Upper panels: Light-adapted responses to a
series of long-wavelength (red) flashes presented on a
steady, white, rod-saturating background are shown for a
representative 4-week-old (left), a 10-week-old (center),
and an adult (right). Stimulus strength in log troland
seconds is indicated to the left of the traces. Lower panel:
Photopic b-wave amplitude (mean +/− SEM) is plotted as
a function of stimulus strength for 4- and 10-week-old
infant and adult control subjects (17). A photopic hill is
apparent in the adults but not in the infants.
The cone-driven b-wave stimulus–response relationship in adults shows a

photopic hill; specifically, with increasing stimulus strength, amplitude
increases to a peak and then decreases (Figure 9-4, right panel). The
amplitude of the cone-driven b-wave in healthy young infants is nearly that
of adults, but over the stimulus range used, does not show the falloff at high
intensity. This lack of the “photopic hill” in infants is attributed to immature
interaction of ON and OFF bipolar cell activity (17).
For reference, data from normal infants and children obtained using the
International Society for Clinical Electrophysiology of Vision (ISCEV)
standard ERG test conditions are available (25,26). A synopsis of these
developmental changes is provided in the appended “Atlas of ERG
Responses.” The ISCEV conditions do not consider the stimulus response
data presented in Figures 9-2 through 9-4. In both scotopic and photopic
conditions, ISCEV responses in infants are smaller than those in adults. It is
essential that these age-specific normal values be considered in interpretation
of infants’ ERG results.
Procedural details can make the difference between success and failure in
pediatric ERG testing. ERG tests that have a high probability of yielding
records suitable for analysis are done in a quiet, dark room free of
distractions. Satisfactory ERG responses are recordable in awake infants. In
typically developing infants, this is readily accomplished up to approximately
age 100 days. In older infants and preschool children, who are too active and
restless for reliable testing in the awake state, a plan for safe testing under
sedation or with brief, light anesthesia may be considered.
Multifocal ERG
The multifocal ERG (mfERG) is used to evaluate functional topography in
the central approximately 40 degrees of retina, including the macula (Figure
9-5, top panel). Responses from a large number of discrete retinal regions,
centered on the fovea, are evoked by multiple contiguous stimuli, the
luminance of which alternates under control of an m-sequence, a
pseudorandom binary sequence (27,28).
FIGURE 9-5 Multifocal ERG responses to a pattern of
hexagons that subtended 43 degrees around a central
fixation cross (11). Upper panel: The cone drive
functional topography of the central retina is represented
by the response density map for a healthy adult. The peak
response is at the fovea; response density decreases with
eccentricity. Middle panel: Representative waveforms
from an infant (red traces) and adult (black traces). These
responses were constructed by averaging the responses in
concentric rings from the center (Ring 1) to the periphery
(Ring 5). These subjects had response amplitude near the
median for their age groups. Lower panel: P1 amplitude
as a function of eccentricity for 10-week-old infants and
adults. The difference in P1 between infants and adults
was greatest in the center (Ring 1) and decreased with
eccentricity (ring number). Amplitude was measured from
the baseline to the first positive peak of the response.
Error bars represent the standard error of the mean.
Frequently, mfERG responses are analyzed by combining responses evoked

by elements in concentric rings centered on the fovea (28). In healthy 10-
week-old infants, the amplitudes of the mfERG responses (Figure 9-5) are
smaller and the implicit times longer than those in adults (11).
These results indicate that the cone-driven activity in the central retina is
significantly immature, in contrast to the relative maturity of peripheral cone-
driven ERG responses to full-field stimuli (17).
The mfERG response depends mainly on the activity of the cone bipolar
cells (29,30). The distribution of amplitudes of the mfERG in 10-week-old
infants show little variation with eccentricity (11). This is in contrast to the
adult’s large central peak and falloff with eccentricity. The explanation for
the infants’ mfERG result lies in the immature distribution of the cone and
cone bipolar cells in the central retina (10,11).
Psychophysics: Development of Normal Scotopic
Retinal Sensitivity
Psychophysical methods investigate the relationship between the stimulus,
specified in precise physical terms, and the subject’s visual response. The
visual response must be demonstrably under stimulus control. Noninvasive
methods are widely used in the assessment of retinal function in adult
patients and are also applicable to pediatric patients (31–35). Typically, for
scotopic testing, the stimuli are relatively small in diameter (2 to 10 degrees)
and thus can probe retinal activity in selected, small patches of retina, in
contrast to the massed response of all retinal cells to full-field stimuli.
Nonetheless, in some clinical research studies, visual responses to full-field
stimuli have their place (36). Because the stimuli are typically brief, often 50
ms or less in duration, a presentation is faster than a saccade, an important
procedural detail if the investigator is to stay in command of the measurement
and the subject’s response is to remain under stimulus control.
Psychophysical Assessment of Rod-Mediated Function

Psychophysical methods that study rod-mediated visual thresholds can be
used to track the course of disease, including, for example, recovery from
mild ROP (37) and worsening retinal sensitivity in LCA (38). The parameters
of the stimulus (size, duration, wavelength, and retinal eccentricity) are
precisely specified. Ocular factors that determine the effectiveness of the
stimulus at the retina include transmissivity of the ocular media, retinal
rhodopsin content, outer segment length, and retinal cell density (cells/mm2).
Importantly, thresholds and the retinal processes these thresholds represent
can be investigated and analyzed using a psychophysical approach even when
the ERG responses are markedly attenuated. Thus, psychophysical
procedures are of great value for following the course of severe retinal
disease.
We use a two-alternative, spatial, forced-choice preferential-looking
method to measure rod-mediated thresholds in dark-adapted infants and
young children. Following 30 minutes of dark adaptation, the infant’s gaze is
directed to a fixation target at the center of a dark screen. An adult observer
reports when the infant is alert and looking at the fixation target. Then, a
stimulus is presented on either the left or the right side of the screen. On
every trial, the observer reports stimulus location (right or left) and receives
feedback. Older subjects point to the stimulus or report verbally on stimulus
location. Rigorous psychophysical procedures with well-documented
properties are used to specify threshold (18). Examples of such procedures
are the method of constant stimuli and staircase methods.
Dark-Adapted Visual Threshold

The DAT is the dimmest test light reliably detected by the subject under
dark-adapted conditions. DATs in normal infants are elevated relative to
those in adults. Examples of DATs, measurable from early infancy onward,
are shown in Figure 9-9. A full-field threshold stimulus test (FST) has been
used effectively in characterization of retinal degenerations and in clinical
trials of gene replacement therapy (36,39,40).
At age 10 weeks, threshold for detecting a 50 ms duration, 10-degree
diameter spot, 20 degrees eccentric to central fixation is approximately one
log unit higher than in adults (41,42) (Figure 9-6). That is, the stimulus must
be 10 times stronger (brighter) for reliable detection by the infant.
FIGURE 9-6 Development of dark-adjusted threshold

(DAT) in healthy control infants. Threshold in log
scotopic troland seconds (log scot td s) is plotted as a
function of age for infants. For comparison, thresholds
from 10- to 16-year-old adolescents and adults are also
shown. The average adult threshold (−3.9 log scot td s) is
indicated by the dashed line; this corresponds to a
threshold elevation (right axis) of zero. Error bars indicate
1 standard deviation.
Spectral sensitivity confirms that these thresholds are rod mediated (43).
Cross-sectional and longitudinal data show that the DAT matures by
approximately age 6 months (16).
Developmental elongation of the rod outer segments and consequent
increase in quantum catch account for threshold maturation in infancy (16,42).
Longitudinal DAT data from patients with a history of mild ROP show
improving retinal sensitivity albeit with a slower course than normal
development (16,18,37), worsening retinal sensitivity in LCA and stable
sensitivity in CSNB. The LCA and CSNB data are shown later in this
chapter.
Psychophysical tests have been used to delve into retinal processes that
are not accessible by ERG procedures. In a number of retinal diseases,
including ROP, results of studies of spatial and temporal summation and
background adaptation disclose abnormal retinal processing that cannot be
deduced from ERG data (33,43–45).
Spatial Summation
Receptive field size determines the ability to integrate visual information over
area (spatial summation). By measuring peripheral retinal thresholds for
detection of spots of light of varying diameters, it is determined that the
receptive field diameter is approximately 1 to 2 degrees in adults, whereas in
10-week-old infants, it is much larger, approximately 6 degrees (16,46,47), the
larger receptive field is a strategy to increase the probability of catch of
quanta despite short outer segments and low rhodopsin content. Furthermore,
the center–surround organization of the immature retina has a larger
excitatory center than does the mature retina (46,47).
Temporal Summation
In the normal, mature retina, threshold decreases with increasing stimulus
duration for stimuli <100 ms, as temporal summation prevails; for stimuli
longer than 100 ms, there is little decrease in threshold (48). Healthy young
infants do not show temporal summation at any duration, possibly due to
immature photoreceptor processes affecting the visual threshold. Abnormal
temporal summation functions in children with a history of ROP are
associated with photoreceptor dysfunction (44,48).
Dark Adaptation after Exposure to Bright Light

In research studies, recovery of retinal sensitivity in infants after exposure to
light can be measured by monitoring ERG recovery, and by recovery of pupil
diameter following a bleaching exposure (49,50). These results suggest that
rod dark adaptation in young infants has the same time course as in adults.
Because of the time needed to complete each threshold measurement, the
time course of dark adaptation has not been measured in infants using
psychophysical methods, and it is not feasible for clinical assessments of
infants and young children.
Background Adaptation
The effect of steady background light on rod-mediated visual threshold
(DAT) has been studied in healthy infants using psychophysical procedures.
Thresholds are measured in the dark-adapted state and then with stimuli
presented on a series of increasingly intense adapting fields resulting in an
increment threshold function. Very dim backgrounds have little effect on
threshold. For brighter backgrounds, log threshold increases linearly over a
several log-unit range of background intensity. The background intensity at
which threshold is elevated 0.3 log units above the dark-adapted level is
called the eigengrau; at that background, the number of thermal and
photoisomerizations is equal. Thus, the eigengrau is an index of intrinsic
retinal noise (31). In healthy retinas, the eigengrau is greater in infants than
adults, and greater in some diseased than healthy retinas (33,45,51,52).
Models of the resulting increment threshold function allow testing of
hypotheses about the photoreceptor or postreceptor site of immaturity or
disease (33,45). The increment threshold function is summarized by the
equation:
log T = log TD + log [(Ao + I)/Ao]
where T is the threshold measured at background I (scotopic trolands), TD is

the DAT, and Ao is the eigengrau. Immaturity or disease limited to the
photoreceptors elevates both TD and Ao.
Immaturity or disease that affects postreceptor processing elevates TD
only. Background adaptation has been studied in patients with a history of
ROP (45,52) and in patients with inherited retinal disorders (33).
Subnormal vision is quite frequent in patients with a history of preterm birth.
Prematurity puts an infant at risk for disturbance not only of the retina but
also of the brain (53) and central visual pathways; the brain abnormality may
have minor to marked impact on vision (54,55). Given that as much as 50%
or more of the brain is involved in vision, brain injury of prematurity is
associated with vision problems due to abnormal processes in the brain. On
the other hand, in some former preterms, visual impairment is mainly due to
ROP. Among the most numerous of ROP patients—those with a history of
mild ROP that required no treatment—best-corrected acuity may be in the
20/40 to 20/25 range rather than 20/20. Deficits in acuity occur even if active
ROP produced no ophthalmoscopically detectable abnormality of the macula.
Subnormal acuity has been thoroughly described (56) both in mild ROP that
never required treatment and in more severe ROP. Although by
ophthalmoscopy the macula may look normal, ocular coherence tomography
(OCT) studies have shown alterations in foveal architecture in former
preterms both without and with ROP (57–60) that, by clinical criteria, never
affected the macula even if the ROP was so severe that laser ablation of the
peripheral avascular retina was given.
The extramacular retina contains the overwhelming majority of rod
photoreceptors, which mature later than the peripheral cones. In the normal
macula, maturation of photoreceptors, both cones and rods, is delayed
relative to peripheral photoreceptors, with the foveal cones being the last to
reach maturity. The macular rods (37) and cones (60) appear to be vulnerable
to the effects of even mild ROP, whereas the extramacular cones, as assessed
by full-field ERG testing, appear little affected by ROP unless the active
disease was severe (61). The amplitude of the mfERG responses, mediated by
the central cones and thus an index of photopic activity in the macula,
remains attenuated many years after active ROP has resolved completely
(62,63) (Figure 9-7). It is thought that the mfERG result is due to prematurity
and ROP interacting with development of the late maturing cells in the
central retina (60).
FIGURE 9-7 Mean N1 to P1 amplitude (±SEM) for
subjects with a history of preterm birth compared to term-
born subjects is plotted for rings 1 (center) to 5 (most
peripheral). The preterm subjects are divided into those
who never had ROP (No ROP), those who had mild ROP
that resolved without treatment, and those who had severe
ROP that required laser ablation of peripheral retina.
Description of the patients is in Altschwager et al. (63).
The stimulus was an array of 103 hexagonal elements that
subtended approximately 23 degrees around a central
fixation target.
Normal DAT mediated by the rods in the macula is delayed in development
compared with the threshold mediated by more peripheral rods (42). The
state of rod maturity (length of outer segments; number of rods per unit area)
and consequent quantum catch account for the thresholds. Mild ROP causes
further delay in development of the central retina, as demonstrated by our
longitudinal study of rod-mediated thresholds (37); threshold did not reach
maturity until age 12 months or later rather than by 6 months, as is the case in
normal development (42).
Rod-mediated ERG responses to full-field stimuli are altered by ROP
(16) and to some extent by prematurity itself (64). The ERG responses from
dark-adapted former preterms show deficits in rod photoreceptor sensitivity
(derived from the a-wave) that are present in infancy and persist in childhood.
In childhood, postreceptor sensitivity (b-wave) improves significantly in mild
ROP but not in severe ROP. We also find this pattern of rod photoreceptor
and rod-driven postreceptor activity in a large cross-sectional sample of
former preterms (18,65), tested in infancy (median age 10 weeks) and in
childhood (median age 10 years). Postreceptor sensitivity does not improve in
childhood in those who had severe ROP. Significant deficits in rod
photoreceptor sensitivity are present in infancy and childhood in both mild
and severe ROP. Low rod sensitivity is a sign of photoreceptor dysfunction
and injury. This is predicted by the onset of active ROP at the age when
photoreceptor development proceeds rapidly with escalating metabolic
demands (Figure 9-8).
FIGURE 9-8 Human rhodopsin growth curve and
retinopathy of prematurity (ROP) onset. The logistic
growth curve represents the developmental increase in
rhodopsin (66). At the time of preterm birth, rhodopsin
content is very low. As the rod outer segment undergoes
developmental elongation and increase in rhodopsin,
phototransduction activity, the circulating current, and
turnover of outer segment material escalate. These
processes require energy. ROP has its onset at the ages of
rapid developmental increase in rhodopsin content of the
retina. The red arrow marks the onset of prethreshold ROP
(67). The photoreceptors are the most oxygen-greedy cells
in the body (68). The energy demands for the
aforementioned processes increase as the outer segment
matures.
The deficit in postreceptor retinal sensitivity (specifically, log sigma of ERG

b-wave (18)) is biologically significant because it predicts deficit in vision,
that is, deficits in rod-mediated visual thresholds (18,65). The improvement
in postreceptor sensitivity in mild ROP is interpreted as a sign of beneficial
reorganization of the postreceptor, rod-driven activity such as we have also
seen in pharmacologically induced photoreceptor injury (66).
Psychophysical testing (43–45) more completely characterizes the
postreceptor functional organization of the retina than does ERG. In brief,
these elegant psychophysical studies on older children with a history of ROP
have shown that the receptive field in mild ROP is larger than normal and
largest of all in those who had ROP so severe as to have required laser
treatment. The background adaptation study shows that noisy operation of
receptor and postreceptor retina limits ROP visual sensitivity. Results of the
background adaptation and temporal summation studies add evidence beyond
that provided by full ERG studies (16,18,64) of persistent rod dysfunction.
Although some of the above-mentioned characteristics of ROP vision are
reminiscent of normal immaturities, the ROP results indicate distinct
alterations in receptor and postreceptor processes rather than lingering
immaturities.
LEBER CONGENITAL AMAUROSIS

LCA is a group of retinal degenerative conditions, each attributable to
changes in a particular gene (67–70) (see also Chapter 44). LCA is typically
symptomatic from early infancy onward and is the diagnosis that comes to
the clinician’s mind when a healthy infant presents as possibly blind. Visual
inattention and roving eye movements or nystagmus are the classic
presenting symptoms in an infant with LCA. These symptoms are neither
specific to nor constant in LCA. Visual acuity, DAT and spherical equivalent
(SE) in some of our patients with LCA are shown in Figure 9-9.
FIGURE 9-9 Visual acuity (VA), dark-adapted visual
threshold (DAT), and spherical equivalent (SE) in patients
with a secure genetic diagnosis of Leber congenital
amaurosis (LCA), achromatopsia (ACHM), or congenital
stationary night blindness (CSNB) are compared to normal
(shaded areas). The different symbols represent the
causative gene (listed in the keys). In each panel, line
segments connect data from an individual. The shaded
areas indicate the 95% prediction limits for VA and SE,
and the range for normal dark-adapted threshold; the
smooth curve within each shaded area plots the normal
average as a function of age. In LCA, typically low VA
worsens with age; DAT also worsens with age; SE is
commonly, but not exclusively, hyperopic. In ACHM, the
prevailing course of acuity is low, but stable; DAT is
normal despite mild deficits in scotopic ERG activity; SE
is often hyperopic. In CSNB, VA is below average and
stable; DAT results document night blindness that is stable
in most but not all; SE is often myopic. Visual acuity is
plotted as the log of the minimum angle of resolution (log
MAR); the right axis indicates the Snellen fraction. Dark-
adapted threshold is plotted as the logarithm of the
difference in threshold from the normal mean; the right
axis gives threshold in troland values. Refraction is plotted
as spherical equivalent, diopters.
A broad spectrum of signs is found in infants and young children in whom a

genetic diagnosis of LCA is ultimately secured. Responses of the pupils to a
hand light may be conspicuously attenuated or quite brisk. Often infants with
LCA have high hyperopia (Figure 9-9) but myopia is also found; we and
others have encountered patients with a genetic diagnosis of LCA who have
myopia in excess of 10 diopters in early childhood. Fundus appearance
ranges from normal to marked abnormality of pigmentation. Normal vascular
caliber is seen in some, but arteriolar attenuation is found in many. Optic
nerve heads are remarkably pink in some, but in others are pale or have
indistinct margins. Either atrophic maculae or thickened maculae have been
demonstrated by OCT (71). In short, the history and examination in the office
can raise the suspicion of LCA but, given the enormous spectrum of clinical
features, will not secure the diagnosis. Prompt testing of retinal function is
the next step in the clinical assessment.
ERG responses are markedly attenuated in both scotopic and photopic
conditions. Some examples are shown in the appended “Atlas of ERG
Responses.” In our patients with a genetic diagnosis of LCA, the amplitude
of the ERG responses is significantly reduced, often <5% of the normal mean
for age, even when visual acuity is quite well preserved and the DAT is
readily measureable.
Acuity is measurable in the majority of our patients with LCA. Acuity
tests provide assessment of function mediated by the central retina.
Procedures and test materials, such as the Feinbloom chart, the Rudimentary
Vision Test (72), and the Teller Acuity Cards (1), facilitate measurement of
very low vision that is sometimes reported as “hand motions” or “count
fingers” vision (73,74). Use of these tests is strongly encouraged because they
yield numeric results that facilitate monitoring of the patient’s course.
Acuity of 20/400 or better throughout the life span is found in
approximately one-quarter of patients with LCA (72). At young ages, we
have found acuity of 20/200 or better in more than half of patients with a
genetic diagnosis of LCA. Some examples are shown in Figure 9-9. In
agreement with Walia et al. (74), we found that the best acuities in our
patients were in those with the RDH12, RPE65, LRAT, and CRB1 forms of
LCA. In some children for whom a genetic diagnosis of LCA is ultimately
secured, vision is so good that the diagnosis is initially overlooked by
experienced pediatric ophthalmologists. Among these patients, visual
impairment may not be recognized until the child reaches the age of
independent ambulation; parents report that the child becomes lost even in
familiar environments.
Although it is a huge advantage to have relatively good acuity for early
learning tasks, these children have reduced contrast sensitivity. For instance,
with typical high-contrast acuity tests, acuity of 20/50 or 20/60 may be
obtained, but the vision may plummet to 20/200 or worse for low-contrast
stimuli. If low-contrast sensitivity is not recognized and reported to the
child’s educators, there are huge difficulties in implementing a satisfactory
educational plan.
Developmental changes in normal contrast sensitivity occur in early
infancy (75). At young ages, contrast sensitivity is assessed in the clinic using
the Ohio contrast cards (76–78).
DAT is measureable in the majority of our patients with LCA even when
the ERG responses are markedly attenuated (<5% of the normal amplitude
for age). Abnormal DAT from an early age is the rule (Figure 9-9).
Progressive loss of retinal sensitivity is indicated by further elevation of the
threshold with increasing age. A worsening of 1 log unit or more over time is
a significant change in an individual’s DAT (79). In those patients who
appear to have no light perception, performance on this two-alternative
threshold test falls to chance, indicating no reliable detection of the stimulus
light. Regional variations of the degeneration across the retina (80) may
affect DAT thresholds. In some patients with LRAT and RDH12, DATs are
markedly elevated while acuities are quite well preserved.
The diagnosis of a particular form of LCA is secured through genetic
testing. Assessments of functions characterize the severity of the disease and
monitor the course of the disease. The genes for LCA encode proteins
involved in phototransduction, the pigment epithelial–photoreceptor
exchange of retinoids, and the structure of photoreceptors and thus can have a
direct effect on the physical properties of the retina that constrain visual
function. Some uncommon syndromic conditions may present in infants and
young children who have the ocular features of LCA. At times, the
differential diagnosis of LCA includes Senior-Løken, Joubert, Alstrom,
cerebello–oculo–renal syndromes, and peroxisomal and mitochondrial
disorders. Some of these disorders are caused by different mutations in the
same genes that cause LCA, for example, NPHP5 and NPHP6 mutations can
cause either isolated LCA or LCA plus renal failure, which is then termed
Senior-Løken syndrome. These conditions are diagnosed by identifying
additional nonocular features and following up with the appropriate genetic
and systemic evaluations.
Gene supplementation for patients with RPE65 (LCA2) through viral
vector (36,81–85) is now an approved treatment. Even after prolonged
blindness, the visual cortex responds to the gene replacement (86) under OCT
guidance as to location of viable cells. Work continues on gene replacement
and other approaches to treat the many other forms of LCA (87–90).
ACHROMATOPSIA
ACHM is a group of congenital, predominantly autosomal recessive retinal
disorders in which there is an absence or paucity of functioning cones.
Clinical features include reduced visual acuity (Figure 9-9), photophobia,
nystagmus, impaired color discrimination, and paradoxical pupil constriction
to dark. Due to the photophobia, acuity may be better when measured in dim
rather than typical room light. Hyperopia is common (Figure 9-9), although a
broad range of refractive errors is reported.
Blue cone monochromatism (BCM) is an X-linked condition with
features similar to achromatopsia but sometimes less severe and typically
showing myopic refractive error. The Berson plates (91) or the Mollon-Reffin
minimal test (92) may facilitate identification of BCM in boys whose clinical
features are suggestive of achromatopsia. Rather than hyperopia, myopia is
typical in BCM. While these conditions have traditionally been reported to be
stationary and to have normal fundus appearance, there have been recent
reports of progressive loss of function (92,93). Structural changes in the
macula (94–97) are reported but are quite stable in both children and adults
(97).
Full-field ERG is indispensable in securing a clinical diagnosis of
achromatopsia and should be considered for any patients showing the clinical
characteristics described above. Sample records for achromatopsia are
included in the appended “Atlas of ERG Responses.” In achromatopsia and
blue cone monochromatism, cone and cone-driven ERG responses are
nondetectable or markedly attenuated. Rod and rod-driven responses have
been reported to be normal or near normal (98,99), but several recent studies
have found abnormal, but recordable, scotopic ERGs (93,94,100–102). One
study found that the ERG abnormalities were accompanied by reduced
expression of rod-specific proteins, including rhodopsin, and disruption of the
functional and structural integrity of rod synaptic terminals (102). Despite
scotopic ERG deficits (100), the dark-adapted thresholds (DATs) are normal
in achromatopsia (Figure 9-9).
Additional evidence for structural abnormalities in patients with
achromatopsia is provided by high-resolution OCT and adaptive optics
scanning laser ophthalmoscopy (SLO) studies. OCT findings include
disruption or loss of cone inner and outer segments, reduced retinal thickness
in the macular region, and foveal hypoplasia (lack of foveal pit formation).
SLO evaluation showed disruption of the foveal photoreceptor mosaic, but
residual cone inner segment structure was present in most patients (94).
Generally, the structural abnormalities advance with age, providing further
evidence that achromatopsia is not completely stationary.
A functional magnetic resonance imaging study showed reorganization of
cortical maps in rod monochromats (103). Cortical regions in V1 that
responded only to signals from cones in normal controls responded to rod-
initiated signals in these individuals, suggesting that early, anomalous retinal
inputs affect the organization of the visual cortex.
The most common molecular causes of achromatopsia are mutations in
CNGA3 and CNGB3. Mutations in GNAT2 and PDE6C are less frequent
causes of achromatopsia. All four genes are expressed in the cone
photoreceptor and are involved in cone phototransduction. CNGA3 and
CNGB3 are cyclic guanosine monophosphate (cGMP)–gated cation channel
genes, GNAT2 is a transducin protein, and PDE6C is a cone
phosphodiesterase. Genetic testing for all four of these genes is available. The
most common molecular causes of BCM are mutations in the opsin gene’s
array of long and medium wavelength-sensitive cone visual pigments, located
adjacently on the X chromosome.
Restoration of visual function in animal models of achromatopsia (mouse
(104) and dog (105)) has been accomplished using adeno-associated virus
(AAV) gene replacement therapy. In the treated animals, recovery of cone
function was demonstrated by an increase in photopic ERG amplitude to
near-normal levels. These results hold promise for future treatment in humans
with achromatopsia. The report of age-dependent changes suggests that,
ideally, treatment should be applied early. ERG recording and imaging
techniques can be useful in identifying patients with relatively well-preserved
cone function and structure, respectively, and in assessing the response to
treatment in clinical trials.
CONGENITAL STATIONARY NIGHT
BLINDNESS (SEE ALSO CHAPTER 34)
CSNB is a group of heritable retinal conditions characterized by night
blindness that is considered nonprogressive (106,107). There is difficulty
adapting to low light levels (night blindness). An infant with CSNB may have
difficulty finding the bottle in dim light; youngsters with CSNB may exhibit
anxiety in dimly lit environments. As shown in Figure 9-9, the DAT, acuity,
and SE vary among the genetic types of CSNB with considerable overlap
(108,109). The DAT measured using psychophysical procedures is typically
quite stable over time (Figure 9-9), although progressive compromise of
night vision has been documented in some (106,110,111).
Moderate deficits in visual acuity, such as 20/50 or 20/75, are common in
CSNB, although a broad range (108,109) of acuity is found (Figure 9-9).
Myopia at an early age is common, although some patients who are destined
to become highly myopic are not myopic in early infancy (Figure 9-9).
The combination of common, nonspecific features—inability to correct
visual acuity with glasses, myopia, nystagmus, paradoxical pupil constriction
to darkness, and reduced vision in the dark, along with the normal fundus—
leads the clinician to include CSNB in the differential diagnosis. Nonetheless,
these common features are not necessarily all present in individuals with
CSNB; patients with TRPM1-associated CSNB, for example, were recently
reported to rarely complain of night blindness in childhood (112). Therefore,
maintaining some level of suspicion of CSNB and being prepared to move
forward with functional phenotyping (below) by ERG and DAT testing
becomes important.
In most cases of CSNB, the fundus appears normal except for changes
associated with high myopia or in small subgroups of CSNB, such as those
with fundus albipunctatus in which white dots are present (RDH5, RLBP1)
and Oguchi disease with adaptation-dependent change in fundus color (SAG,
GRK1) (see Chapter 34). Approximately two-thirds of our patients with
CSNB have nystagmus and strabismus, and paradoxical pupil constriction to
darkness is present in approximately half of individuals with CSNB (34).
Among the patients with CSNB, there is a wide range of SE (Figure 9-9)
from low hyperopia to high myopia; myopia is quite prevalent (34,108).
Functional phenotyping by ERG testing and psychophysical assessment of
dark adaptation and DAT are used to help secure a diagnosis of CSNB. To
differentiate CSNB from degenerative retinal disorders, testing for visual
field defects and abnormal autofluorescent pattern are important adjuncts to
the ERG and psychophysical tests.
Clinical categorization of CSNB into CSNB1 (complete) and CSNB2
(incomplete) is based on ERG and psychophysical data. In CSNB1, the dark-
adaptation curve shows little evidence of rod branch (or break), and the DAT
is elevated (Figure 9-10). CSNB2 shows some evidence of rod recovery and
more modest elevation of the DAT. CSNB1 and CSNB2 are associated with
particular genetic diagnoses (Figure 9-10) and altered bipolar cell activity
(113–115).
Both CSNB1 and CSNB2 show a negative ERG waveform; that is, a
robust negative-going a-wave and a significantly reduced b-wave (Figure 9-
11). In general, the changes are more marked in CSNB1 than in CSNB2.
(Table 9-3) (see also appended “Atlas of ERG Responses”).
FIGURE 9-10 Dark adaptation functions in CSNB and
control subjects. These theoretical curves plot threshold as
a function of time following exposure to a bright adapting
field. The solid black curve represents typical results from
a control subject. The first part of the function represents
cone system adaptation, and the second rod system
adaptation. In CSNB1, there is little or no evidence of rod
function (red curve). In CSNB2, there is some evidence of
rod adaptation (blue curve). Although there is a range of
thresholds and some overlap, threshold at 30 minutes
remains elevated in both CSNB1 and CSNB2. Genes
associated with each type of CSNB are in the boxes to the
right of the graph (39,40).
FIGURE 9-11 Sample negative ERG waveform. The red
trace is from a patient with CSNB1; the b-wave–a-wave
ratio is <1. The black trace is from a control subject; the b-
wave–a-wave ratio is >1. The ERG waveforms are in
response to the ISCEV 3.0 dark-adapted condition (116).
Table 9-3 Summary of ERG and psychophysical
responses in CSNB
ERG, electroretinographic; CSNB, congenital stationary night blindness; ISCEV, International Society
for Clinical Electrophysiology of Vision; DAT, dark-adapted visual threshold.
Raghuram A, Hansen RM, Moskowitz A, et al. Photoreceptor and postreceptor responses in congenital
stationary night blindness. Invest Ophthalmol Vis Sci 2013;54(7):4648–4658; Komaromy AM,
Alexander JJ, Rowlan JS, et al. Gene therapy rescues cone function in congenital achromatopsia. Hum
Mol Genet 2010;19(13):2581–2593; Miyake Y, Yagasaki K, Horiguchi M, et al. Congenital stationary
night blindness with negative electroretinogram. Arch Ophthalmol 1986;104(7):1013–1020; Stunkel
ML, Brodie SE, Cideciyan AV, et al. Expanded retinal disease spectrum associated with autosomal
recessive mutations in GUCY2D. Am J Ophthalmol 2018;190:58–68.
The Riggs-type CSNB shows attenuation of both a- and b-wave in scotopic

conditions but preserved photopic function. Riggs-type ERGs have been
reported in autosomal dominant and autosomal recessive forms and are
associated with gene defects in the rod phototransduction cascade (GNAT1,
PDE6B, RHO, SLC24A1). Patients may have relatively mild night blindness
and normal visual acuity and lack high myopia and nystagmus. A recently
reported autosomal recessive Riggs-type CSNB is caused by mutations in
GUCY2D, and although night blindness is noticed early in life, acuity and
refraction are normal and nystagmus is not present (116). For these reasons,
this relatively uncommon form of CSNB may be overlooked (107).
SUMMARY
ERG and psychophysical studies of visual and retinal function can contribute
key diagnostic information. Furthermore, even after genetic diagnosis is
secured, measures of retinal and visual function are used to delineate the
severity of the condition and to track the course of disease.
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10
ERG Waveforms in Inherited
Pediatric Retinal Diseases
Hanna M. De Bruyn, Wanda Pfeifer, Arlene V. Drack, Ronald M.
Hansen, and Anne B. Fulton
This chapter depicts normal and abnormal full-field electroretinogram (ERG)

waveforms in healthy retinas and in specific genetic retinal disorders. It may
be used as a reference, either to search ERG waveform patterns for those that
match a given patient or to search by disease to review the expected ERG
waveforms in each condition. Exemplar ERG records from normal children
and adults as well as pediatric patients with retinal disorders are presented to
illustrate a wide spectrum of retinal conditions. For retinal disorders, sample
records can represent waveforms at only one point in the patient's course;
thus the waveforms at earlier or later time points may differ markedly from
those depicted in the sample records.
During infancy, the retina undergoes developmental changes that affect the
amplitude and implicit time of the ERG a- and b-wave responses (1,2; see
also Chapter 9). A graphic summary of developmental changes from early
infancy to adulthood is shown below (Figure 10-1). Thereafter, Exemplars
demonstrate ERG records showing responses to full-field brief (<3 ms)
stimuli. The ERG records in Sample records 10-1 to 10-12 were obtained
using a bipolar Burian-Allen electrode after pupil dilation and dark adaptation
(≥20 minutes). Each ERG test started in the dark-adapted condition and was
followed by the light-adapted condition, ending with the flicker response.
Table 10-1 summarizes the stimulus conditions used for each test for Sample
records 10-1 to 10-12. These stimuli are similar to those in the ISCEV
standard protocol (3–5). The ERG records in Sample records 10-13 to 10-20
were obtained using DTL plus electrodes with silver clip reference electrode
on the ipsilateral earlobe. ERG testing started with the light adaptation
conditions followed by dark adaptation testing after ≥20 minutes of dark
adaptation. Testing was performed with dilated pupils and followed ISCEV
standard protocols (3). Table 10-2 outlines the stimulus conditions used for
each test for Sample records 10-13 to 10-20.
FIGURE 10-1 ERG b-wave development. Rod (upper),
maximal (middle), and cone (lower) b-wave amplitudes
are presented from infancy to adulthood. Ninety percent of
healthy individuals for the indicated age groups would be
expected to fall within the solid red lines. (Reprinted by
permission from Springer: Fulton AB, Hansen RM,
Westall CA. Development of ERG responses: the ISCEV
rod, maximal and cone responses in normal subjects. Doc
Ophthalmol 2003;107(3):235–241.)
TABLE 10-1 Stimuli (<3 ms)
TABLE 10-2 Stimuli (<3 ms)

SAMPLE RECORD 10-1 ERG responses in a healthy
adult (blue traces) and a healthy 10-week-old infant (red
traces). In the dark-adapted conditions, the infant’s
responses are smaller and delayed compared to that of the
adult. In the light-adapted condition, the 2.25 cd s/m2
flashes produce b-wave amplitudes and implicit times that
are similar in the adult and in the infant. The infant’s
response to the 30-Hz flicker is smaller and delayed
relative to that in the adult.
SAMPLE RECORD 10-2 Achromatopsia (CNGB3
mutations). In this 2-month-old (black traces), the light-
adapted responses are markedly reduced. In the dark-
adapted eye, the response to the 0.01 cd s/m2 flash is
attenuated, but the response to the 0.23 cd s/m2 flash is
robust and similar to that in the healthy 10-week-old (gray
trace). (From Fulton AB, Hansen RM, Westall CA.
Development of ERG responses: the ISCEV rod, maximal
and cone responses in normal subjects. Doc Ophthalmol
2003;107(3):235–241.)
SAMPLE RECORD 10-3 Alström syndrome (ALMS1
mutations). In this 21-month-old, amplitudes of the a- and
b-wave responses are smaller than normal, and implicit
times are longer than normal. In the light-adapted
conditions, the responses are nondetectable. This is
evidence of cone–rod dysfunction.
SAMPLE RECORD 10-4 Blue cone monochromatism
(OPN1LW mutation). Amplitudes of the responses to the
dark-adapted stimuli are reduced in this 2-year-old boy.
Amplitudes of the light-adapted responses are markedly
reduced, and the implicit time of the 30-Hz flicker
response is delayed.
SAMPLE RECORD 10-5 Cone–rod dystrophy
(Bornholm eye disease, OPN1LW mutation). This 2-year-
old boy with high myopia has rod and cone responses with
markedly reduced amplitudes compared to the control.
Additionally, the 30-Hz flicker response is slightly
delayed.
SAMPLE RECORD 10-6 X-linked congenital stationary
night blindness, CSNB1 (NYX mutation) and CSNB2
(CACNAF1 mutation). Left:NYX. In this 17-year-old
male, the b-wave amplitudes are reduced, while the a-
wave in the 0.23 cd s/m2 condition is similar to that in the
control. The photopic responses are similar to those in the
healthy control. Right:CACNAF1. Responses from an 8-
year-old male in the dark-adapted condition are similar to
those in the patient with NYX, although the b-wave is
slightly more robust than in NYX. Note that in the photopic
conditions, amplitudes are significantly reduced compared
to the adult. (See Chapter 9.)
SAMPLE RECORD 10-7 Leber congenital amaurosis,

LCA (CEP290, RPE65, CRB1 mutations). Patients were 2
years (left), 9 years (center), and 3 months old (right) at
the time of the ERG tests. Even in infancy, the amplitude
of responses in both dark- and light-adapted conditions are
markedly attenuated (<5% of the normal mean) (see
Chapter 27).
SAMPLE RECORD 10-8 Posterior microphthalmos with

retinal folds (MFRP mutations). In both dark- and light-
adapted conditions, this 3-year-old’s response amplitudes
are nearly normal; implicit times are slightly prolonged.
SAMPLE RECORD 10-9 Stickler syndrome (COL11A1
mutation). In this 2-year-old, the amplitudes are
attenuated, and the implicit times are slightly delayed in
both the dark- and light-adapted conditions. The response
to the 30-Hz flicker is both reduced in amplitude and
slightly delayed.
SAMPLE RECORD 10-10 Usher syndrome (MYO7A,
USH2A mutations). Left:MYO7A. Both the dark- and
light-adapted responses in this 14-month-old are reduced
in amplitude and delayed. Right:USH2A. In this 18-
month-old, dark- and light-adapted responses are slightly
smaller and slightly delayed compared to normal. (See
Chapter 33.)
SAMPLE RECORD 10-11 X-linked juvenile
retinoschisis (RS1 mutation). In this 4-year-old boy, the a-
wave responses in the dark-adapted conditions are nearly
normal, but the b-wave responses are attenuated and
delayed. The responses in the light-adapted conditions are
similar to the control.
SAMPLE RECORD 10-12 X-linked retinitis pigmentosa
(RPGR mutation). For this 6-year-old boy, amplitudes in
both the dark- and light-adapted conditions are attenuated,
and the implicit times are prolonged. The response to the
30-Hz flicker is reduced in amplitude and is delayed.
In each figure, the patient’s responses (black traces) are superimposed on

those of an age-appropriate healthy control (gray traces) to facilitate
comparison of the amplitudes and the implicit times of the b-wave. The
records do not represent the spectrum of severity for each disease or the
course of the disease. ERGs are rarely diagnostic and must be taken into
context with other clinical and genetic features. Some ERG patterns are
especially characteristic of a diagnosis and include achromatopsia (Sample
record 10-2), congenital stationary night blindness (Sample record 10-6), and
Leber congenital amaurosis in infancy (Sample record 10-7). Responses from
patients under age 1 year are compared to those from a healthy 10-week-old
infant. In all figures, the red bars indicate flash onset; the black bar placed at
the peak amplitude of the b-wave indicates implicit time (Chapter 9). A
Directory of ERG sample records is shown below.
DIRECTORY Directory of ERG Sample Records

SAMPLE RECORD 10-13 Stargardt disease (ABCA4
mutations). Left:Mild type. This 17-year-old female
presented at age 12 years with visual acuity 20/20 OD and
20/25 OS. By 17 years of age, the vision had decreased to
20/50 OD and 20/100 OS. However, the ERG remained
close to normal with only the b-wave amplitude in the 3.0
dark-adapted condition being slightly reduced. In the light-
adapted conditions, the amplitudes are slightly attenuated
with mild changes in the implicit time. Right:Severe type.
This 13-year-old girl first presented at 8 years of age with
20/300 OD and 20/200 OS vision. At 13 years of age,
vision was 20/200 OD and 20/150 OS (after learning to
use eccentric fixation) and amplitudes in both the light-
and dark-adapted conditions are severely attenuated.
SAMPLE RECORD 10-14 Batten disease (CLN3

mutations). Left:Initial ERG. For this 8-year-old female,
amplitudes in both the light- and dark-adapted conditions
are attenuated. An electronegative 3.0 dark-adapted
standard combined response can still be seen. Vision at
this time was 20/500 OD and 20/300 OS. Right:16-
Month follow-up: 16 months later, vision decreased to
worse than 20/600 OD and 20/400 OS with a barely
discernible electronegative standard combined response
(dark-adapted bright flash) b-wave.
SAMPLE RECORD 10-15 Joubert (INPP5E mutations).

Left:Initial ERG. In this 10-year-old female, amplitudes
in both the light- and dark-adapted conditions are
attenuated or significantly decreased. Vision at that time
was 20/125 OD and 20/100 OS. Right:4-Year follow-up:
Four years later, both light and dark amplitudes decreased
to nondiscernible in all test conditions with a recorded
vision of 20/125 OD and 20/150 OS. Visual fields were
markedly constricted over time.
SAMPLE RECORD 10-16 Enhanced S-cone syndrome
(NR2E3 mutations). Left:For this 8-year-old female, the
amplitudes in both the dark and the light are severely
attenuated, and latencies are delayed. A small A- and b-
wave can be seen on dark- and light-adapted 3.0 testing
and 30-Hz flicker testing. Vision was 20/30 OD and 20/40
OS. Right:This 29-year-old female with mutations in the
same gene has a preserved a-wave on dark and light 3.0
testing with a smaller b-wave and delayed latencies. Dark-
adapted 3.0 standard combined testing has an
electronegative ERG waveform. Vision was CF OD and
20/30 OS. Vision OD may have been affected by
amblyopia. Note that waveforms for LA 3.0 and DA 3.0
are strikingly similar.
SAMPLE RECORD 10-17 Bardet-Biedl syndrome
(BBS1 mutations). Younger sibling. In this 8-year-old
female, amplitudes for both dark and light are severely
attenuated with only a small a- and b-wave waveform on
the dark-adapted 3.0 bright flash. Vision was 20/40 OD
and 20/40 OS. The older sibling, a 14-year-old female,
has amplitudes that are nondiscernible in all test
conditions. Vision was 20/150 OD and 20/80 OS.
SAMPLE RECORD 10-18 X-linked juvenile
retinoschisis (RS1 mutation) with electronegative DA
bright flash. For this 12-year-old male, the dark-adapted
bright flash waveforms are below normal with preserved
a-wave and slightly smaller b-wave with an
electronegative waveform. Light-adapted ERG amplitudes
are within normal range but slightly delayed in latency.
SAMPLE RECORD 10-19 Night blindness.
Left:Autosomal recessive complete congenital
stationary night blindness (TRPM1 mutations). This 15-
year-old male’s dark-adapted ERG has decreased dim
flash amplitudes and an electronegative 3.0 bright flash.
His light-adapted ERG has decreased amplitudes with a
broadened 3.0 a-wave and broadened/double–peaked
flicker waveform. Vision was 20/25 OD and 20/20 OS.
Right:Autosomal recessive congenital night blindness
(GUCY2D mutations). This 17-year-old male has
severely attenuated dark-adapted ERG waveforms. Light-
adapted ERG waveforms are below normal and delayed,
with 30-Hz flicker response relatively preserved. Vision
was 20/20 OD and 20/20 OS.
SAMPLE RECORD 10-20 Autosomal dominant retinitis
pigmentosa (PRPF8 mutation). This 6-year-old female has
decreased dark and light ERG waveforms. Her dark-
adapted 3.0 bright flash has a small a- and b-wave
waveform with delayed latencies. She has a small
preserved b-wave present on the 3.0 light-adapted ERG
that is also delayed in latency. Her 30-Hz flicker ERG
appears nondiscernible. Her vision was 20/25 OD and
20/25 OS.
REFERENCES
prematurity. Prog Retin Eye Res 2009;28:452–482.
2. Hansen RM, Moskowitz A, Akula JD, et al. The neural retina in retinopathy of prematurity.
Prog Retin Eye Res 2017;56:32–57.
3. McCulloch DL, Marmor MF, Brigell MG, et al. ISCEV standard for full-field clinical
electroretinography (2015 update). Doc Ophthalmol 2015;130(1):1–12.
4. Fulton AB, Hansen RM, Westall CA. Development of ERG responses: the ISCEV rod, maximal
and cone responses in normal subjects. Doc Ophthalmol 2003;107(3): 235–241.
5. Moskowitz A, Hansen RM, Akula JD, et al. Rod and rod-driven function in achromatopsia and
blue cone monochromatism. Invest Ophthalmol Vis Sci 2009;50: 950–958.
11
Amblyopia in Pediatric Retinal
Conditions
Alina V. Dumitrescu and Pavlina S. Kemp
Even the most elegant and anatomically successful retinal surgery in a child
can fail to improve vision—or even worsen it—due to amblyopia. Amblyopia
is a preventable, treatable form of vision loss due to lack of visual input from
an eye to the visual cortex during the critical period of visual development.
From birth to 4 months of age, amblyogenic factors are emergencies, and
they continue to be a serious concern until about 9 to 10 years of age. When
considering retinal surgery for children in the amblyopic age range, timing
and type of procedure, as well as postoperative amblyopia treatment, must be
carefully considered in order to achieve an optimal visual outcome.
PREVALENCE
Amblyopia has an estimated prevalence worldwide between 2% and 5% (1),
similarly distributed among racial and ethnic groups. It is most commonly
unilateral but can be bilateral. Typically, amblyopia can only occur from birth
to age 10 years while the visual system is developing and is most efficiently
treated during this important window of time (2,3). Although some treatment
success has been reported at older ages (4,5), development of the disease has
not.
Pediatric retinal disorders are part of a heterogeneous group of
conditions, including many diseases affecting vision early in life. They can be
acquired or inherited and can threaten vision throughout childhood. Patients
with retinal diseases with onset in the first decade of life carry the additional
risk of developing unilateral or bilateral amblyopia with long-term visual
consequences. Amblyopia and its risk factors may result from the retinal
disorder itself or the treatment thereof.
The diagnosis and treatment of amblyopia must be part of the
management plan for children with retinal disorders, starting at diagnosis.
Amblyopia risk should be considered in treatment plans and addressed in
consent for procedures. Retina specialists treating pediatric patients should be
familiar with the principles described in this chapter and may find it helpful
to collaborate with their pediatric ophthalmologist colleagues when caring for
young children with retinal disorders.
ENVIRONMENTAL FACTORS
Amblyopia is caused by abnormal visual input to the brain during the critical
period of visual development (6,7). This can result from various causes, such
as strabismus, refractive error, or visual deprivation (8), each of which can
occur independently, due to retinal disorders or as a result of their treatment.
Visual deprivation, due to corneal, lens or vitreous opacity, retinal
malformation, or degeneration, may result in amblyopia in addition to the
primary organic cause of the vision loss. As a result, retinal disorders have a
much higher risk of vision loss in childhood than if they are acquired in
adolescence or adulthood. Complications from treating retinal disorders or
the necessary postoperative course following successful treatment can also
introduce additional amblyopia risk factors. Environmental factors such as
ocular trauma or infection in childhood can lead to unilaterally or bilaterally
decreased visual acuity. Poor visual acuity at a very young age often results
in nystagmus.
General lack of resources to treat refractive error with glasses or contact
lenses, as well as shortage of medical and surgical expertise, may lead to
amblyopia in situations where it could have otherwise been avoided.
GENETICS
Although amblyopia itself is not considered an inherited disorder, amblyopia
risk factors, such as strabismus, refractive error, and anisometropia, are often
familial. The inheritance of these risk factors seems to be multifactorial and is
not well established (9). Prematurity, associated developmental delays, and
other environmental factors contribute to the risk of developing amblyopia
(10).
Retinal disorders with onset in childhood are often inherited with a well-
established genetic profile. The association of childhood-onset retinal
disorders with amblyopia is common.
WORLDWIDE IMPACT
Due to the treatable nature of amblyopia, the impact of diagnosis and
treatment is tremendous. Every single case of amblyopia, if diagnosed and
treated promptly, can lead to a lifelong improvement in vision (11).
As will be repeatedly emphasized in this chapter, amblyopia that
develops in association with a retinal disorder has its own particularities in
diagnosis and treatment. On the one hand, childhood-onset retinal disorders
are either traumatic or associated with complex ocular pathology and,
therefore, difficult to treat and often are associated with less-than-perfect
vision even when optimally treated. On the other hand, the final visual
outcome is difficult to predict and often surprising in children. However, if
overlying amblyopia develops and is not recognized or treated, the final
visual acuity can be permanently and severely impacted (12,13).
PATHOPHYSIOLOGY
According to the American Association for Pediatric Ophthalmology and
Strabismus (AAPOS), amblyopia is defined as “decreased vision in one or
both eyes due to abnormal development of vision in infancy or childhood.”
(14) Amblyopia is the leading cause of vision loss among children and is
often called “lazy eye” in lay terminology. Vision loss occurs due to
insufficient stimulation of neural pathways between the brain and the eye
during the critical developmental period. This results in abnormal processing
of visual images and leads to decreased visual acuity (15). The brain often
favors one eye due to poor vision in the other eye. Most commonly, there is
no structural problem of the eye or visual pathway. Amblyopia can, however,
occur in eyes with a structural abnormality and cause further reduction in
visual acuity. This is sometimes termed “organic amblyopia.” This type of
amblyopia is especially common in pediatric retinal disorders and may be
missed or not considered because 100% of vision loss is attributed to the
underlying pathology. Every child under the age of 10 years with a retinal
disorder should be considered to have amblyopia as a cofactor in his or her
vision loss.
Unilateral amblyopia is thought to result from competition and inhibition
between the inputs received by neurons processing the visual information
from the two eyes. If the images are not clear (due to blur) or not fusible (due
to strabismus or retinal noncorrespondence), domination of cortical vision
centers by the fixating or better-seeing eye will develop. This further
chronically reduces the responsiveness to input from the nonfixating eye (6).
The Basic and Clinical Science Course, Section 6: Pediatric
Ophthalmology and Strabismus defines unilateral amblyopia as a “two-line
difference in verbal recognition visual acuity measured with linear Allen,
HOTV, or Snellen (not LogMAR acuity) (16).”
Unilateral amblyopia is defined as mild, moderate, or severe, and the
level of severity has implications for visual prognosis and treatment. A two-
line difference but less than a four-line difference is defined as mild
amblyopia. Moderate amblyopia is defined as a four-line difference to less
than six-line difference. A six-line difference or more is defined as severe
amblyopia (15). In preliterate children, mild amblyopia is central, steady,
unmaintained (CSUM) fixation, as compared with central, steady, maintained
(CSM) fixation in the nonamblyopic eye; uncentral, steady, unmaintained
(UCSUM) fixation is considered moderate, and uncentral, unsteady,
unmaintained (UCUSUM) fixation is considered dense amblyopia (8).
Generally, bilateral amblyopia is defined as best-corrected visual acuity
in each eye worse than 20/50 for 3-year-olds and worse than 20/40 in 4- to 5-
year-old children (6).
Critical Period for Visual Development

In general, amblyopia results in lifelong visual loss if it is untreated or
inadequately treated. The disease develops exclusively during the visual
development period and is much more severe when it develops at a young
age; therefore, age at diagnosis and treatment of amblyopia are critically
important.
From observational studies of patients with congenital cataract and from
animal studies, the critical period for visual development in humans was
established (17). For unilateral deprivation, the first 6 to 10 weeks of life are
critical for visual development (17,18). For bilateral deprivation, the first 17
weeks of life are critical. Treatment within this period is the most effective
(19). Reports of significant visual improvement outside the critical age are
the exception, rather than the rule. Severe or complete visual deprivation in
early infancy (within the first 3 months of life) is associated with the
development of nystagmus. Delays in treatment beyond this age usually
result in poor visual outcomes despite anatomic success (20) and irreversible
dense amblyopia in one or both eyes. Less severe amblyopia can develop in
early infancy even with short, constant periods of visual deprivation or
inequality. The potential for development of amblyopia exists with
deprivation as short as 1 week per year of age (21).
The critical period for visual development extends until 9 to 10 years of
age and sometimes beyond. However, the degree of amblyopia, reversibility,
and timing of development vary. Fortunately, at an older age, amblyopia is
less severe and more reversible with prompt treatment.
CLINICAL SYMPTOMS AND SIGNS

Amblyopia presents as reduced visual acuity that is not improved by
refraction either in an anatomically normal eye or in an eye with pathology
but not of a severity equal to the visual deficit in the presence of an
amblyopia risk factor. Unless suspected, amblyopia might not be diagnosed
especially in diseases that are bilateral and asymmetric. Strabismus and
eccentric fixation may or may not be present. Children generally do not
report decreased vision especially if it is unilateral. Decreased vision may be
discovered by parents or teachers or through vision screening at the
pediatrician or community vision screening efforts.
DIAGNOSTIC STUDIES
Visual acuity measurement is the most important step in diagnosing
amblyopia. In infants and children, this critical step is complex and time
consuming (22,23). The examination includes vision assessment with proper
optical correction and identification of all risk factors for amblyopia.
Glasses prescription must be based on cycloplegic retinoscopy (23). The
lenses present in the glasses must be measured at every visit and confirmed to
be the lenses prescribed, because children frequently break or lose glasses
and have them remade but will start using an old pair or even a pair intended
for siblings. Lenses may become dislodged from glasses during cleaning or
play and are often repositioned incorrectly, thereby resulting in switched
anisometropic correction and displaced optical centers.
The choice of visual acuity testing has to be appropriate for age (24). For
young children, usually at least 2 years and younger, fixation preference and
induced tropia testing are the preferred methods for checking visual acuity
(22). Teller (or grating) visual acuity is also used but overestimates vision in
amblyopic eyes (25). For older children, optotypes with symbols or letters are
used. Amblyopic eyes typically test better with single-letter or isolated-letter
acuity than linear acuity, and the first and last letters of a line are more likely
to be identified correctly (26). Due to these considerations, full lines of
optotypes or crowding bars around single letters are recommended, and
Teller acuity should be interpreted judiciously. Care must be taken to test one
eye at a time (6); this is very different than testing in adults, because children
want to please the doctor and do not want to get answers “wrong”; therefore,
children will guess and peek with the better eye. An adhesive eye patch is
recommended, because using a plastic occluder or holding a hand over one
eye does not yield accurate visual acuities in children (6).
Latent nystagmus increases with complete occlusion of one eye and will
degrade monocular visual acuity. In children with nystagmus, vision should
be tested using a fogging lens (+5 to +10 D for phakic eyes) or a translucent
occluder over one eye first and then the other. If the child has a null point,
acuity should be tested with the head in the preferred position to allow
minimum eye movement.
For pediatric patients with retinal disease, the presence of amblyopia
should be suspected if vision is decreased beyond what is expected for the
retinal disease alone and should be interpreted in the context of any identified
amblyopia risk factors. Risk factors for development of amblyopia are high
refractive error, anisometropia, strabismus, and visual deprivation beginning
at <10 years of age. Additionally, occlusion amblyopia can be caused by
penalization treatment of amblyopia and can lead to reversal of amblyopia
laterality.
Diagnostic optical coherence tomography can be used to assess the
macular structure, but a trial of amblyopia therapy may nonetheless be
successful in attaining some visual improvement even with structural
abnormalities.
Refractive Error and Anisometropia

Bilateral high refractive error is a risk factor for bilateral amblyopia.
Classically, hyperopia more than 4 to 5 D, myopia more than 5 to 6 D, and
astigmatism more than 2 to 3 D (16) are described as risk factors for
isometropic amblyopia. Large population studies have also described children
developing bilateral amblyopia with lower bilateral refractive error (27).
Anisometropia is the difference in refraction between the two eyes and is
a risk factor for unilateral amblyopia. AAPOS criteria for amblyogenic
anisometropia include more than 1.5 D difference between the two eyes for
hyperopia; more than 3.5 D difference for myopia; and more than 1.5 D
difference for astigmatism in any meridian (14). However, anisometropia as
minor as 0.5 D has been reported to cause amblyopia (8). Studies show that
age of onset and the degree of anisometropia are the most important factors in
developing refractive amblyopia (28,29). The prevalence of refractive
amblyopia increases with age, and the severity of amblyopia increases with
the difference in refractive error between the two eyes. Amblyopia develops
in about 20% of children with 1 D spherical equivalent of anisometropia and
tends to be very mild with two lines of acuity difference or less. In contrast,
over 60% of children with 4 or more diopters of anisometropia will develop
moderate or severe amblyopia (29).
The type of refractive error is also important in amblyopia risk
assessment. Compared with anisometric myopia, anisometric hyperopia and
astigmatism have a higher risk of amblyopia, and smaller refractive
differences are pathologic. In high myopic anisometropia of 4 D or more, the
rate of amblyopia development is very similar to hyperopic or astigmatic
amblyopia (28).
Strabismus
Although any ocular misalignment can be associated with the development of
amblyopia, unilateral vision loss is more likely to result from nonalternating
and constant strabismus. Esodeviations more commonly cause amblyopia
than exodeviations. Acquired childhood strabismus is more often associated
with amblyopia than is congenital strabismus (6). Development of the
compensatory mechanisms of cross-fixation and alternating fixation help
young children maintain equal vision in both eyes and avoid diplopia.
Unilateral vision loss, such as that from retinal disease, can lead to loss of
binocularity and fusion and can result in sensory strabismus followed by
further visual compromise from strabismic amblyopia. Young children are
more likely to develop sensory esotropia than older children and adults, who
tend to develop sensory exotropia. A child with a latent or intermittent
exotropia, who suffers a vitreous hemorrhage, may develop a manifest
exotropia within a few weeks due to the decreased ability to fuse. This then
worsens the amblyopia caused by the vitreous hemorrhage itself.
Visual Deprivation
Deprivation amblyopia is a consequence of decreased input from one or both
eyes due to anterior segment media opacities, such as corneal scars or edema,
inflammation, cataract, ptosis obscuring the visual axis, and/or vitreous and
retinal hemorrhages, retinal detachment, or retinal scars. Patching or covering
the eye or blurring the vision with plastic shields, atropine, or ointment can
potentially cause amblyopia if the child is young enough and the distortion
persists long enough. Deprivation amblyopia can begin at birth or develop
later. It is the least common form of amblyopia but is often the most severe
and difficult to treat. The degree of amblyopia is correlated with the degree of
visual obstruction and the age of onset. Visual deprivation amblyopia is the
most severe form of amblyopia especially if the onset of deprivation is in
infancy (30).
Severely reduced visual input to the brain from an eye due to vitreous
hemorrhage, cataract, or patching causes dense amblyopia. Reports suggest
dense amblyopia from deprivation begins at approximately a week of
complete occlusion per year of age of the patient (31). There are no
confirmatory studies, but visual outcome data on natural history of untreated
congenital cataract (17) as well as reports on occlusion amblyopia (32) are
supportive of this timeline. For example, a 9-year-old child with a vitreous
hemorrhage or a dense traumatic cataract has about 9 weeks before
amblyopia will be a concern, whereas a 1-year-old child has only 1 week.
Therefore, surgical removal of vitreous hemorrhage is elective in adults,
fairly urgent in school age children, and an emergency in infants and children
of 1 to 2 years of age.
Occlusion amblyopia is an acquired, iatrogenic form of deprivation
amblyopia that can occur in the previously better-seeing eye after therapeutic
patching or cycloplegic penalization. It can also occur due to prolonged
postoperative patching, especially in very young children. The incidence and
severity depend on the length of occlusion time, but occlusion amblyopia
generally is believed to be mild and reversible. In at least one prospective
randomized trial, reversal of amblyopia developed in 1% of children patching
for 6 or more hours per day and in as many as 9% of children given one drop
of topical atropine daily after 6 months of treatment (29,33). A retrospective
study of full-time patching for treatment of unilateral amblyopia reported that
as many as 25% of children developed occlusion amblyopia (33). In nearly
every case, the visual acuity in the fellow eye returned to baseline with
discontinuation of the current patching therapy. Other studies that evaluated
lower doses of patching, <6 hours, and shorter duration of atropine found
lower rates of occlusion amblyopia (21,34). Younger children and those with
higher patching doses should be followed at closer intervals and monitored
for amblyopia to decrease the risk of occlusion amblyopia. Postoperative
patching of the operated eye should be done for the shortest time possible,
and a shield with perforations should be used, if available.
Amblyopia in Pediatric Retina Practice

Having established a basic understanding of amblyopia, this chapter gives
special consideration to the role of inherited, congenital, and acquired
vitreoretinal pathology occurring in the critical period for amblyopia and
visual development from birth to 10 years of age (Table 11-1).
TABLE 11-1 Common causes of amblyopia in
retina practice
General rule of thumb: Amblyopia starts to develop when the inciting condition has been present for
about 1 week per year of age of the patient. For example, in a 2-year-old child, amblyopia will start to
be a factor after 2 weeks of vitreous hemorrhage. This is also approximately the time it will take to start
reversing amblyopia with treatment. For example, a 4-year-old child with amblyopia from a vitreous
hemorrhage will need patching or other penalization therapy for at least 4 weeks before an
improvement in vision will be seen.
Vitreoretinal disorders can occur at any age, but inherited disease frequently
manifests early in life. Visual acuity and long-term visual potential are often
poor due to the underlying disease, as well as nystagmus. Despite visual
limitation due to retinal pathology itself, the presence of refractive error,
anisometropia, strabismus, asymmetric disease, or visual deprivation should
raise the suspicion of concomitant amblyopia.
Retinopathy of prematurity (ROP), its related complications, trauma with
cataract and/or posterior segment involvement, as well as infections and
inflammatory systemic diseases with ocular involvement are also common
early in life. If a disorder or its treatment introduces amblyogenic risk factors,
these should be added to the consent, prognosis, follow-up, and treatment
plan.
MANAGEMENT OF AMBLYOPIA
FREQUENTLY ENCOUNTERED IN
PEDIATRIC RETINAL DISEASES
Refractive Error
Some diseases are associated with bilateral high refractive error. Stickler
syndrome, congenital stationary night blindness, Ehlers-Danlos syndrome,
and others are associated with bilateral high myopia with onset before school
age. Leber congenital amaurosis (LCA), oculocutaneous albinism, juvenile
X-linked retinoschisis, and others are associated with high hyperopia from a
very young age. Although these children will have anatomical reasons for
limitation of vision, for example, from staphylomas or foveal hypoplasia,
they will benefit from optical correction as early as possible to develop their
best vision and avoid bilateral refractive amblyopia.
Separately from the underlying disease process, any child can have
unrelated refractive error. Because of this, children with vitreoretinal disease,
as with all other children, should have an eye examination that includes
cycloplegic refraction and should be prescribed glasses, when appropriate,
despite their underlying diseases. It is generally accepted that myopia of at
least −6 D, hyperopia of at least +5 D without esotropia, and astigmatism
more than 2 D are amblyogenic and need to be corrected (6).
Anisometropia can occur in any child and at any age and is a risk factor
for amblyopia. Additionally, asymmetric or unilateral disease, unilateral
surgery, including scleral buckle, vitrectomy, silicone oil placement, or other
types of unilateral treatment, including laser or cryotherapy, increase the risk
of developing anisometropia and unilateral amblyopia.
Retinopathy of prematurity of all levels of severity is a significant risk
factor for developing refractive error, anisometropia, and strabismus. Long-
term follow-up studies have shown that 26% of premature babies with ROP
severe enough to require peripheral laser treatment underwent additional
retinal surgery; 18% had cataract surgery; 20% developed amblyopia, and
10% developed strabismus requiring surgery (20). Approximately 66% of
eyes with high-risk prethreshold ROP observed during the neonatal period
were myopic in preschool and early school years (35). Pathologic progressive
myopia that presents at a very early age is also more common in children
who have had ROP and affects up to 5% of patients (36). Many children who
were born prematurely have foveal anomalies and hypoplasia on OCT even if
they did not have severe ROP (37,38). For these reasons, it is recommended
that children with a history of prematurity and/or ROP have periodic
evaluations for vision, ocular alignment, and cycloplegic refraction (39)
especially early in life.
Additionally, peripheral laser treatment for ROP can be associated with
retinal or vitreous hemorrhages that can precipitate the development of
deprivational amblyopia if they are large or long-lasting.
Vitreous and retinal hemorrhages in childhood can occur spontaneously
from anemia, leukemia, and thrombocytopenia or juvenile X-linked
retinoschisis or in the context of trauma related to vaginal delivery,
nonaccidental or accidental causes, or surgery. Vitreous hemorrhage in early
childhood is rare and may cause few or no symptoms. In the absence of a
known causative event, it may not be discovered for a long time. Absence of
a red reflex is often the reason for an evaluation. Vitreous and retinal
hemorrhages can clear spontaneously, but the process usually takes months or
even longer in large vitreous hemorrhages due to the formed vitreous of
children. During this time, severe unilateral or bilateral amblyopia may
develop. As described above, the severity of amblyopia depends on the
patient’s age. In very young infants within the first 3 months of life,
deprivation from dense vitreous hemorrhage can lead to irreversible
amblyopia within weeks (31). In older children, deprivation amblyopia can
also occur but will develop more slowly.
Unless the vitreous hemorrhage clears spontaneously, its treatment is
surgical removal. The risks and benefits of such surgery need to be weighed
in the disease context and the age of the infant or child. Very young infants
need clear media as soon as possible to develop vision; however, one of the
common complications of vitrectomy is cataract formation, which occurs
about 20% of the time (40). Cataracts will need to be addressed surgically
early in life and will lead to aphakia or pseudophakia, anisometropia and
refractive error, and the added risk of secondary glaucoma. The same is true
of older children but to a lesser degree.
Young children with trauma, either accidental or nonaccidental, and
retinal and/or vitreous hemorrhages often have associated traumatic cataract,
corneal lacerations, cerebral injuries, and intracranial hemorrhages leading to
additional surgeries, cortical visual impairment, and other disabilities.
Recovery is sometimes slow, and anesthesia risk is high. Therefore, although
the goal to gain as much visual recovery is important, the risks and benefits
of surgery, patient’s age, severity of the ocular disease, and effects from
associated neurologic abnormalities must be weighed. In cases in which the
ocular versus cortical component is unclear, serial visually evoked potential
(VEP) tests and/or electroretinograms (ERG) may be useful to assess
changes.
Regardless of age, etiology, and associated comorbidities, the treatment
of retinal and vitreous hemorrhages is not complete without amblyopia
management.
Lensectomy in children is recommended for congenital or acquired
cataract from trauma or secondary to surgery. Congenital cataracts, especially
when unilateral, are often associated with persistent fetal vasculature and its
associated findings: retinal folds, funnel- or stalk-shaped retinal detachment,
spontaneous vitreous hemorrhage, and a congenitally small eye (see also
Chapter 6). Unilateral congenital cataract in general is highly amblyogenic.
When associated with posterior pole disease, visual prognosis is limited (41).
The decision between aphakia and intraocular lens implant is based on
age, laterality, and access to optical rehabilitation. The postoperative
refractive target is based on the child’s age. Glasses will be needed even if
contact lenses are used, because the loss of accommodation will require near
correction. Near vision is very important for development and schoolwork
and needs to be part of pediatric visual rehabilitation.
Retinal detachment is uncommon in children compared with adults, and
the causes and challenges are also different (see also Chapters 60–63). Only
3% to 7% of all retinal detachments occur in the pediatric population (42).
Risk factors associated with retinal detachment in children are accidental and
nonaccidental trauma (see also Chapters 69 and 70), high myopia, hereditary
vitreoretinopathies such as Stickler syndrome and Marshall syndrome,
familial exudative vitroretinopathy (see also Chapters 3, 35, 37, and 66),
Marfan syndrome (see also chapter 37), ROP (see also Chapters 38–41, 43,
51–55), sickle cell retinopathy, previous intraocular surgery, inflammation
(see also Chapters 47 and 48) and Coats disease (see also Chapter 64). The
most frequent type of pediatric retinal detachment is trauma-associated, and
the most common cause of inherited retinal detachment is from
vitreoretinopathies, which can be spontaneous or related to minor trauma
(described in detail in Chapters 4, 7, 48, 54, 55, and 64). (43). A significant
challenge of unilateral retinal detachment without known major trauma is the
lack of symptoms or complaints from the child, which lead to delayed
diagnosis and treatment.
The treatment of pediatric retinal detachment is complex due to several
factors: the high rate of vision-threatening pathology in the fellow eye in
cases of systemic disorders and the considerable financial, emotional, and
time-based investment of attempted repair. Amblyopia risk is an important
component of this decision. Increased risk of proliferative vitreoretinopathy
and increased frequency of macular involvement due to late presentation are
other important factors (43).
The anatomic success of surgery for retinal detachment due to any
etiology in children tends to be lower than in adults and may require more
than one surgery (44). Postoperative vision is correlated with visual acuity at
presentation and the reason for the detachment, but visual acuity better then
20/200 has been reported in some studies as occurring in <50% of cases (45).
The most common primary repair used in children is a scleral buckle;
however, vitrectomy, laser retinopexy, and combinations of these procedures
may be necessary and are addressed in other chapters (see also Chapters 60–
63). Use of silicone oil may also be needed, because postoperative
positioning is difficult in the pediatric population.
The unilateral use of a scleral buckle or silicone oil induces
anisometropia. Scleral buckle placement can lead to axial myopia. Silicone
oil placement will induce variable changes in refraction based on the status of
the lens in the eye. Generally, silicone oil induces a hyperopic shift in a
phakic eye and a myopic shift in an aphakic eye (46). These refractive errors
should be corrected early postoperatively because they can lead to refractive
amblyopia. Postsurgical strabismus and cataract formation are relatively
common complications of retinal detachment surgery. Postoperative
strabismus tends to be restrictive and can be variable. These are all
amblyogenic factors, which have to be considered and included in the visual
rehabilitation plan.
As mentioned above, the age of the pediatric patient is very important
with younger children having a higher risk of developing denser amblyopia in
a shorter duration of time.
Posterior uveitis is not common in children and represents only 5% to
10% of cases of all uveitis (47). However, uveitis can be challenging to
diagnose and treat due to its bilateral and asymptomatic nature in children
(see also Chapters 47 and 48). Most cases are idiopathic, infectious, or
secondary to systemic inflammatory conditions and, therefore, bilateral.
Cases associated with infection tend to also present unilaterally (47). The
most common infections that cause posterior uveitis are acquired or
congenital toxoplasmic retinochoroiditis, necrotizing herpetic retinitis,
toxocariasis, Bartonella henselae, and nematode-associated neuroretinitis. In
infants, ToRCH-related syndromes (toxoplasmosis, rubella, cytomegalovirus,
herpes virus) and lymphocytic choriomeningitis virus and West Nile virus
congenital infections are the most common causes of congenital posterior
uveitis (48) with Zika virus being a more recent concern (49).
Posterior uveitis can lead to the development of band keratopathy,
chorioretinal scars and epiretinal membrane formation, and choroidal
neovascular membranes, all of which can be amblyogenic especially when
associated with unilaterally decreased vision in young children. Treatment of
uveitis requires long-term steroid use and, in some cases, intra- and
periocular steroid injections in combination with systemic treatment. These
treatments have potential complications, including cataract formation,
increased intraocular pressure or frank glaucoma, and retinal detachment
(50). Consequent surgical procedures, such as lensectomy, vitrectomy, and
filtering procedures, can lead to the development of amblyopia risk factors of
anisometropia from pseudophakia or aphakia or strabismus from setons.
Amblyopia should always be suspected when amblyogenic factors are
present, and a diagnostic trial of patching may be considered especially if
acuity is worse than the retinal anatomy would suggest.
Organic amblyopia can develop in any case in which organic disease is
unilateral or asymmetric.
Chorioretinal colobomas, foveal hypoplasia, inherited retinal dystrophies,
and congenital infections, when unilateral or bilateral but asymmetric, will
lead to the development of amblyopia in the worse-seeing eye. The challenge
in such situations is to determine how much of the visual asymmetry is due to
amblyopia versus how much is organic or structural due to the disease
process. It is also difficult to determine the visual potential of the presumed
amblyopic eye and to decide how much and for how long to treat. Often, a
trial of amblyopia treatment is needed to assess the potential for
improvement. Collaboration with a pediatric ophthalmologist is helpful to
determine visual potential.
Retinal dystrophies are often associated with the development of cystoid
macular edema (CME), which should be suspected if a sudden vision
decrease is noted. Chronic CME leads to epiretinal membrane formation and
additional vision loss. Topical or systemic treatment with carbonic anhydrase
inhibitors may lead to improvement or resolution of edema, which can be
recurrent and sometimes require long-term treatment. When unilateral or
asymmetric, CME can create the circumstances for development of
amblyopia.
LCA and other retinal degenerative disorders of childhood can be
associated with amblyopia. The degree of visual loss associated with LCA is
variable and largely dependent on the gene and mutations involved. This is
also true for other childhood-onset retinal degenerations. Refractive error,
anisometropia, strabismus, as well as nystagmus and abnormal head position
are often associated with these conditions. Diagnosing amblyopia and
treating it are essential.
Gene replacement therapy for LCA is now available for at least one gene
(biallelic RPE65 mutations). Clinical trials and posttrial analysis show that
patients who had low vision through the entire period of critical vision
development and treated with gene replacement therapy responded to the
treatment (51) and patients treated at <10 years of age had similar visual
improvement as older patients treated after age 10. Additionally, a study
using noninvasive multimodal neuroimaging showed normalization along the
visual fibers in the visual pathway of the treated eye to levels comparable to
normally sighted people, and the degree of plasticity and recovery did not
depend on the patient’s age at the time of the treatment (52). The impact of
amblyopia in future gene therapy or stem cell-derived therapy for unilateral
or bilateral retinal dystrophy is yet unknown. There is no study showing a
direct effect of amblyopia on the success of the therapy, but it is inevitable
that we will learn more about the implications of amblyopia as treatments for
more common inherited retinal diseases become available, and children of
various ages are studied.
Oculocutaneous albinism and foveal hypoplasia are often associated with
high hyperopia, nystagmus, and various degrees of esotropia. Treatment of
hyperopia and strabismus are always recommended regardless of degree of
foveal hypoplasia or visual acuity. Often, vision improves with glasses.
Because some diseases have no current treatment options and can lead to
further vision loss due to disease progression, parents and physicians may
question the need to treat amblyopia in such cases. Further knowledge of
amblyopia treatment will help with such decisions.
TREATMENT AND VISION

REHABILITATION IN AMBLYOPIA
Regardless of the cause of amblyopia and the patient’s age, amblyopia
treatment should at least be considered and discussed with the family as well
as the patient, if possible. All considerations should be reviewed including
timing and time investment, safety and visual potential, as well as long-term
consequences. Treatment success tends to decline with increasing in age;
however, an attempt at treatment should be offered to all children including
older children and teenagers (15).
The chances for obtaining good or normal vision in an amblyopic eye
depend on many factors, including the age of onset as well as the cause,
severity, and duration of amblyopia. Success is also influenced by any
concomitant conditions and highly dependent on adherence to treatment
recommendations (53).
The first steps in amblyopia treatment are obtaining clear media and
proper refraction in both eyes. The next step is to the promote the use of the
amblyopic eye, which can be done using part-time occlusion/patching (PTO),
full-time occlusion/patching (FTO), pharmacologic penalization using
atropine 1%, fogging, or binocular treatment. The choice of treatment
modality is mainly based on the density of amblyopia and child’s age as well
as the ability of the child to cooperate and adhere to the follow-up schedule.
Pharmacologic penalization using atropine 1% is only recommended for
mild to moderate amblyopia in children 3 to 15 years of age and is effective
in cases with visual acuity in the amblyopic eye no worse than 20/80 (21).
The principle of treatment is that cycloplegia induced by atropine optically
defocuses the nonamblyopic eye and changes the preference to the amblyopic
eye.
Pharmacologic penalization works best when the nonamblyopic eye is
hyperopic and not optically corrected. Use of atropine for penalization may
be especially effective in the presence of latent nystagmus or for maintenance
treatment (6,21). The usual dose is one drop daily, but various other regimens
have been described. Atropine may have systemic side effects, including dry
mouth and skin, fever, delirium, and tachycardia, which may be more severe
in younger patients. Parents should also be counseled about the severe
reaction and potential lethality to orally ingesting this medication. Therefore,
it is necessary to carefully store the drug away from children at home.
Patching is well established in the treatment of amblyopia. The
mechanism of improvement in visual acuity with patching is believed to be
related with the decrease in neural signals from the nonamblyopic eye under
the patch and the increased signal from the unpatched eye. The recommended
way of patching is to apply an opaque adhesive patch directly to the skin over
the nonamblyopic eye. Proper glasses are worn over the patch. Alternatives,
including a cloth patch mounted on the eyeglass frame or pirate patch, are
less effective, because children can easily look around these types of patches
(6).
The amount of daily patching is dependent on the child’s age and density
of amblyopia. Mild to moderate amblyopia can benefit from PTO of 2 to 6
hours/day, whereas denser amblyopia requires more aggressive regimens, up
to FTO (33,34). Occlusion amblyopia, worsening of strabismus, and skin
irritation from use of adhesive patching have been reported as side effects
regardless of the intensity of the regimen (34,54). These side effects tend to
be reversible without long-term harm.
Most studies recommend PTO up to 6 hours/day, and some studies found
this regimen as effective as FTO (54,55). There is compelling evidence of the
effectiveness of FTO especially in the treatment of moderate to severe
amblyopia, and it is the only treatment modality with a definitive end point in
cases when the visual acuity in the amblyopic eye does not achieve 20/20 or
equal to that of the nonamblyopic eye (56). Use of FTO has an increased risk
of developing occlusion amblyopia, and the follow-up of these patients must
be more frequent. In PTO, the recommended follow-up is every 2 to 3
months (34); for FTO, the recommended follow-up is 1 week for every year
of age of the child. This interval in FTO is considered a “treatment interval”
and used to determine the end point; three treatment intervals without
improvement indicate that patching can be tapered, because FTO beyond this
time is unlikely to result in an improvement in visual acuity (56). Visual
acuity should be regularly assessed during tapering of patching to monitor for
any recurrence of amblyopia.
The role of surgery in amblyopia treatment is to address causative factors,
such as clearing the visual media, which should be done early on. Surgery
addressing strabismus is usually considered after occlusion treatment is at its
end point and the child’s potential for binocularity and, therefore, stable
ocular alignment is maximal. For children with acquired strabismus and
diplopia, the use of prisms to maintain fusion and binocularity prior to
strabismus surgery is recommended.
Binocular therapy has been tried in children with orthotropia or small-
angle strabismus with some preexisting binocularity. Although early
nonrandomized studies were promising, more recent results from a
randomized trial failed to demonstrate that binocular game play at 1 hour per
day was as good as patching 2 hours per day. Research is ongoing, but at this
point, there is insufficient evidence to recommend binocular therapy for
treatment of amblyopia (57).
Other various types of optical penalization, like removing the optical
correction of the better eye or fogging the lens of the better-seeing eye with
filters or other materials, might have a role for mild amblyopia (58). The
prescriber and parents have to keep in mind that children of all ages like to
have the best vision they can at all given times and will try to use their better-
seeing eye. When they have the opportunity, children will look over or under
their glasses making optical penalization less effective.
After the initial treatment, up to 25% of successfully treated children can
have a recurrence of amblyopia when treatment is discontinued. Restarting
treatment and long-term maintenance therapy may be necessary in some
cases (6).
ROLES OF OTHER PHYSICIAN AND

HEALTH CARE PROVIDERS
The diagnosis and treatment of amblyopia can be difficult for patients and
parents who are already facing an ocular disorder that limits vision in one or
both eyes. Amblyopia treatment occasionally spans over months, and often
long-term maintenance treatment is necessary during the visually vulnerable
first decade of life.
Support from trusted pediatric ophthalmologists and pediatricians who
know the family is often very valuable in improving the treatment
compliance and coping with the disease. Particularly important for children
with developmental delays, but also for all children having a hard time
dealing with decreased baseline vision during amblyopia treatment, the
involvement of behavioral therapists and psychologists can be beneficial.
It is important to ask about socioeconomic barriers to treatment, such as
lack of transportation for follow-up visits, inability to afford glasses, patches
or medicine, as well as personal beliefs that may affect the ability to adhere to
recommended treatment. Social workers may help with overcoming some of
these limitations.
ETHICAL CONSIDERATIONS
Dense amblyopia with significant underlying disease is very difficult to treat,
and visual outcomes are often poor. Large differences in vision between the
two eyes allow only limited treatment options and make it more difficult for
the family and child to comply with treatment. Long-term patching for
treatment of amblyopia is more difficult in children with very poor vision in
the amblyopic eye as well as in children with associated morbidities and
developmental problems. Patching can carry a significant social stigma for
children and impact the quality of life for both children and their parents.
Goals of treatment and prognosis should be clear and reasonable. A clear end
point for reevaluation of patching should be discussed at the outset; if a
plateau is reached with no improvement, patching should be discontinued or
tapered off.
It should be emphasized to patients and their parents that even in children
with underlying organic diseases, visual acuity can improve with amblyopia
therapy. In one retrospective review, 51% of patients with underlying organic
abnormities achieved a visual acuity of at least 20/80 with FTO for up to 7
months with younger patients responding faster and obtaining better final
visual acuity (59).
Every age group and etiology for amblyopia have their own treatment
challenges. Young children with dense amblyopia will find it very difficult to
patch, but a close and sustained collaboration with families can lead to
successful results. Older children may be more resilient, but need to be
involved in the treatment and motivated to participate.
Studies have shown somewhat contradictory information about long-term
implications of amblyopia in people’s lives. In one study of 370 children and
adults with unilateral amblyopia, acquired vision loss in the non-amblyopic
eye resulted in severe visual impairment in 23% of patients (60), indicating
the importance of rehabilitating amblyopic eyes whenever possible. A
different study following 1,037 patients with unilateral vision loss due to
amblyopia found no functional impact on childhood motor development,
teenage self-esteem or adult socioeconomic status during follow-up when
compared to peers with good vision in both eyes (61).
Although sometimes unachievable, the ultimate goal of treatment is
lasting equal visual acuity between the two eyes.
FUTURE TREATMENTS
Amblyopia does not resolve spontaneously and will lead to permanent vision
loss in the affected eye unless treated with patching, pharmacologic
penalization, glasses when needed, or surgery when necessary (14).
Use of refractive surgery to correct for anisometropia and refractive
errors has been proposed and is used in selected cases at this time (62). If
long-term safety and efficacy are proven, refractive surgery might be used
more commonly in the pediatric population.
The addition of new treatment modalities with pharmacologic agents like
levodopa/carbidopa alone or in combination with patching or atropine (63)
has been proven to have some efficacy but is still not widely used. Similarly,
the use of citicoline has been described with some efficacy in improving
vision in amblyopic eyes of older patients when used alone or in association
with patching or atropine (64).
New trends and nonoperative interventions using modern technology for
treatment of amblyopia are still under scrutiny; these include the use of liquid
crystal glasses (65), perceptual learning (66), transcranial magnetic
stimulation (67), dichoptic training (68), or acupuncture (69). Small studies
have shown the value of each of these approaches in isolation or in
combination with traditional treatment at various ages.
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12
Workup of Infantile/Congenital
Nystagmus
Brittni A. Scruggs and Arlene V. Drack
Infantile nystagmus syndrome (INS) is an involuntary, oscillatory eye

movement disorder presenting within the first 6 months of life that is
unrelated to medications or other acquired forms of nystagmus. A sign rather
than a diagnosis, INS warrants careful history taking, examination, and a
thorough workup with possible coordination of care with neurology,
pediatric, or genetic services.
INS can be idiopathic or associated with various disease states, such as
albinism, early-onset retinal dystrophies, neurologic states, or low vision (1).
This chapter reviews the main etiologies of INS and highlights a cost-
effective and accurate diagnostic approach that will help with the
management of these complicated cases. Congenital nystagmus is often
associated with ocular diseases that cause deterioration of visual acuity;
however, accurate diagnosis will identify children with high refractive errors
or other treatable etiologies of nystagmus. INS can also be associated with an
underlying systemic or neurologic disease. An accurate diagnosis in these
cases has profound effects on the child’s psychological, social, and
neurologic development.
PREVALENCE
Approximately 14 children per 10,000 live births have INS worldwide (1). In
the United States, the birth prevalence of INS is estimated to be 12 children
per 10,000 live births with a two- to threefold male predominance (2,3). A
study by Nash et al. revealed that 87% of all pediatric patients with
nystagmus can be classified as having INS (2).
No known external environmental factors cause INS. Premature birth and
intrauterine infections or toxins that result in central nervous system or ocular
damage increase the risk of INS. Young maternal age (<18 years) and
intrauterine exposure to alcohol or diabetes increase the risk of optic nerve
hypoplasia, which is an important cause of INS.
GENETICS
INS is a descriptive name for an eye movement disorder with many different
etiologies. In one study of 202 INS patients, the vast majority had a genetic
cause (4). Genes causing INS can result in progressive or stable conditions,
and inheritance can be autosomal dominant, autosomal recessive, X-linked,
mitochondrial, or sporadic. Some of the more common genetic causes will be
discussed in this chapter.
WORLDWIDE IMPACT
Because of the heterogeneity of etiologies, exact worldwide prevalence data
for INS are not known. An estimated 1.4 million children are blind
worldwide affecting about 1.5 per 1,000 children in low-income countries
with high pediatric mortality rates and around 0.3 per 1,000 children in high-
income countries with low pediatric mortality rates (5). Most children with
early-onset blindness develop INS as a result of vision loss, and many have
persistent nystagmus as adults; other children and adults have INS as a
primary condition.
PATHOPHYSIOLOGY
A retrospective study of 202 patients with INS revealed that there are three
major categories of INS etiologies: (a) eye movement/oculomotor disorders,
(b) vision-related disorders, and (c) neurologic disorders (4). The most
common INS etiologies are shown in Figure 12-1. The patients in the
oculomotor disorder category have idiopathic INS, also called congenital
motor nystagmus (CMN), and represent about 10% of all INS patients (4).
CMN is a diagnosis of exclusion, so this diagnosis should never be made
without a complete workup. The pathogenesis of idiopathic CMN is still not
completely elucidated, and no single causative lesion in the ocular motor
control system has been identified. However, there are many connections
between the eye and various brain locations, and defects in fixation, saccades,
the motor neural integrator, the optokinetic reflex, or the pursuit eye
movement systems may contribute to the development of nystagmus (6).
FIGURE 12-1 The percentage of infantile nystagmus

etiologies based on a retrospective study of 202 INS cases.
(Adapted by permission of Taylor and Francis Group,
LLC, Bertsch M, Floyd M, Kehoe T, et al. The clinical
evaluation of infantile nystagmus: what to do first and
why. Ophthalmic Genetics 2017;38:22–33.)
CMN often presents at a few weeks of age as a bilateral, conjugate horizontal
nystagmus that has an accelerating slow phase. Likely due to the
developmental miswiring of the visual system, this disease process
commonly portends a good prognosis due to the foveation strategy of patients
during vision development. The most common hereditary form of CMN is X-
linked, associated with mutations in the FRMD7 gene. This form may not be
completely idiopathic, however, as some mutations have recently been linked
to subtle foveal hypoplasia and optic disc anomalies (7). FRMD7-associated
CMN shows 100% penetrance in male children and 53% in female carriers
(3). CMN patients have good near visual acuity, usually in the normal range,
often associated with a symmetric jerk nystagmus that is absent during sleep.
In these patients, convergence dampens nystagmus and oscillopsia are absent.
Pediatric patients with INS have a high likelihood of having a genetic
cause for their nystagmus with primary defects in the anterior visual pathway.
Some propose that the oscillations are adaptive responses to poor foveal
fixation by a normal ocular motor system (3,8). In the study by Bertsch et al.,
56% of all INS cases were secondary to retinal disorders, most commonly
albinism and Leber congenital amaurosis (LCA) (4). Both X-linked and
autosomal recessive oculocutaneous albinism (OCA) patients have reduced
visual acuity with the X-linked OA patients generally having more peripheral
pigment (in hair, skin, etc.) than their AR counterparts. Even in OCA
patients, those with only partial loss of enzyme activity in the pigment
cascade (previously called tyrosinase positive) have gradual darkening of
their skin, hair, and sometimes retinal pigment epithelium over time; this may
be associated with mildly improved vision and nystagmus. Albinism-affected
patients tend to have photophobia due to diffuse iris transillumination defects,
a hypopigmented fundus, and foveal hypoplasia. There are albinism
syndromes, such as Chédiak-Higashi syndrome and Hermansky-Pudlak
syndrome, that present with nystagmus but also have serious systemic
manifestations, which are detailed further in this chapter.
In the genetic era, the extreme heterogeneity of expression of albinism
has been appreciated. Previously, only patients with complete lack of
pigment could be confidently diagnosed with albinism. With genetic testing,
we have learned that many albinism patients have moderate peripheral
pigment and some have no iris transillumination defects. There are even
albinism patients without nystagmus. As a common cause of INS, albinism
should always be suspected and not ruled out until macula optical coherence
tomography (OCT) and/or genetic testing can be performed.
Patients with vision-related disorders causing INS have a sensory deficit
associated INS. LCA, achromatopsia, blue cone monochromatism, congenital
stationary night blindness (CSNB), Bardet-Biedl syndrome (BBS), and
Joubert syndrome are inherited retinal dystrophies causing poor vision at
birth and are commonly associated with INS (1,4). Ocular disorders with
afferent defects (e.g., optic nerve hypoplasia, foveal hypoplasia, aniridia,
retinopathy of prematurity, and/or large colobomas) and visual deprivation
(e.g., congenital cataracts) can also induce INS (1,4).
Neurologic conditions are the third major cause of INS. Intracranial
tumors, Chiari malformations, hydrocephalus, structural malformations,
metabolic disorders, and various developmental syndromes are in the
differential diagnosis and generally cause deviations in growth and
development in addition to INS. Fewer than 3% of all INS cases in a pediatric
ophthalmology practice had purely neurologic causes (4). A thorough review
of systems, history, growth, head circumference and weight charts, and eye
and neurologic examination guide the clinician to obtain head imaging or
request neurology consultation when appropriate.
CLINICAL SIGNS AND SYMPTOMS

A classification of eye movement abnormalities and strabismus (CEMAS) is
one nomenclature system for nystagmus classification. In this system, the
nystagmus is characterized by waveform type (e.g., direction, amplitude,
frequency). The CEMAS criterion for diagnosing INS is an accelerating slow
phase in an infant (3). This method of classification has low specificity in
diagnosing underlying INS etiologies, which are numerous and may have
similar waveforms (Figure 12-2) (4).
FIGURE 12-2 Nystagmus waveforms seen in patients
with infantile nystagmus. Nystagmus waveform types are
listed on the Y-axis. The most common etiologies of
infantile nystagmus syndrome are provided on the X-axis.
For any given diagnosis, the percentage of patients with
each waveform is provided. The waveform types per
diagnosis may equate to more than 100% given patients
often have more than one waveform type. (Reproduced by
permission of Taylor and Francis Group, LLC, from
Bertsch M, Floyd M, Kehoe T, et al. The clinical
evaluation of infantile nystagmus: what to do first and
why. Ophthalmic Genet 2017;38:22–33.)
Although formal eye movement recordings are rarely performed clinically,

physicians should note the general appearance of the nystagmus as to identify
red flags that increase suspicion of a neurologic disorder (1,9):
Asymmetric or unilateral nystagmus

Gaze-evoked nystagmus
Saccadic intrusions (fast eye movements away from a target)
Seesaw nystagmus
Spasmus nutans (a shimmering and asymmetric nystagmus)
Vertical nystagmus
In INS, there are varied manifestations, but the eye movements are mostly
conjugate (10). Nystagmus with acceleration during the slow phase with a
typical frequency of 2 to 4 Hz is characteristic of INS (Figure 12-3) (6). Jerk
nystagmus with fast corrective saccades and pendular waveforms with slow
initiating and corrective movements also can occur in infantile cases (3,6).
Any saccadic intrusion, or fast eye movement away from a target, should
raise suspicion that the child does not have INS and, instead, may have
opsoclonus secondary to neuroblastoma (3). CSNB often has associated INS
with unusual waveforms, including vertical and opsoclonus-like (4,11).
FIGURE 12-3 Typical nystagmus waveforms in infantile
nystagmus syndrome. A:Nystagmus with acceleration
during the slow phase and fast corrective saccades is
characteristic of jerk-type infantile nystagmus syndrome.
B:Pendular waveforms with slow initiating and corrective
movements also can occur in infantile cases. Most
infantile nystagmus waveforms have a frequency of 2 to 4
Hz. (Reprinted from Richards M, Wong A. Infantile
nystagmus syndrome: clinical characteristics, current
theories or pathogenesis, diagnosis, and management. Can
J Ophthalmol 2015;50(6):400–408. Copyright © 2015
Canadian Ophthalmological Society. With permission.)
It is common for infants to minimize their nystagmus by keeping their eyes in
a null position or zone; such foveation periods minimize visual symptoms
(e.g., oscillopsia) and improve visual acuity (12). Alexander law states that
nystagmus is more pronounced when the gaze is directed toward the side of
the fast-beating component. Thus, a patient with a left jerk nystagmus would
expect to have a left head turn. These abnormal head positions should be
noted in the initial examination; a small lateral head turn is the most common
position adopted as the null point is usually within 10 degrees of fixation.
Acquired forms of nystagmus can often be localized to a specific
anatomic region based on waveform. For example, downbeat nystagmus
usually localizes to the vestibulocerebellum (e.g., cerebellar degeneration,
Chiari malformation, etc.). The same localization is not possible for early-
onset nystagmus forms like INS. The presence of oscillopsia, which is the
perception of visual motion, further differentiates pathologic acquired
nystagmus from INS as patients with INS rarely complain of this.
INS should also be distinguished from latent nystagmus (fusion
maldevelopment nystagmus), which occurs in approximately 50% of patients
with infantile esotropia and, less frequently, in other types of infantile
strabismus. Latent nystagmus is predominantly a bilateral, horizontal jerk
nystagmus elicited by occluding either eye. The slow phase is toward the side
of the occluded eye with an exponential decrease in the slow-phase velocity.
In INS, there is a reversed optokinetic nystagmus (OKN) response in which
the fast-beating phase of the induced nystagmus is in the same direction as
the OKN stimulus; this is compared to the normal OKN response in which
the fast phase beats in the opposite direction to the stimulus movement (13).
DIAGNOSTIC APPROACH
Clinical History
An initial clinic visit for INS should include detailed family, birth, medical,
and developmental histories of the child. The age of nystagmus onset should
be determined given INS is rarely present at birth and more commonly begins
by 3 months of age (3). Onset after 6 months should raise suspicion of an
acquired nystagmus, which has a different workup and is not covered in this
chapter. Relevant vision impairment, strabismus, abnormal head posture,
kidney diseases, and neurologic disorders in the family or child should be
discussed, and pedigrees should be drawn to determine the pattern of
inheritance when appropriate.
Clinicians should be aware that in certain inherited disease states, there
are wide phenotypic variations within a family. For example,
nephronophthisis (NPHP) gene mutations can lead to progressive cystic
kidney disease with or without profound vision loss due to LCA. The
combination of LCA with NPHP is caused by mutations in NPHP5 or
NPHP6 (also called IQCB1 and CEP290) and is referred to as Senior-Löken
syndrome (14). The first symptom is usually nocturia or bed-wetting due to
an inability to concentrate urine. Cerebellar ataxia, skeletal involvement,
congenital oculomotor apraxia, and hepatic fibrosis can also be present in
these patients (14). Conversely, of all patients with idiopathic INS (i.e., no
evidence of ocular or systemic disease), up to 50% have a family history of
nystagmus without other findings.
Gestational age, developmental milestones (e.g., walking, talking), delays
or regressions, and growth curves (e.g., head circumference, weight, height)
should be reviewed. As children get older, abnormalities that suggest the
presence of an underlying ocular or neurologic disease state should be
identified, including:
Neurologic signs or symptoms

Ataxia
Oculomotor apraxia
Seizures
Vertigo, nausea, frequent vomiting
Changing percentiles on growth curves for head circumference,
height, and/or weight
Ocular signs or symptoms
Night blindness
Photophobia
Poor color discrimination
Clinical Examination
A complete pediatric eye examination commonly narrows down the diagnosis
in INS and is indicated for all infants with nystagmus, ideally prior to
imaging or further testing.
Examination findings that may increase the probability of specific ocular
disorders associated with INS are listed in Table 12-1. The examination
should begin with careful external inspection of the child. For example,
albinism often presents with hypopigmentation of the child’s eyes, skin, and
hair secondary to reduced quantity of melanin in melanosomes. The
phenotypic variations of OCA are provided in Figure 12-4.
Neurofibromatosis type I, which can cause optic nerve gliomas and resultant
INS, has characteristic signs of neurofibromas (i.e., peripheral nerve tumors),
café au lait spots, axillary freckling, and/or osseous lesions or deformities.
Lisch nodules of the iris can be seen on slit lamp examination in about 80%
of children by 8 years of age. Obesity may signal a syndrome, such as
Bardet-Biedl or Alström.
TABLE 12-1 Clinical findings of ocular diseases

commonly associated with infantile nystagmus
All findings may not be present in every patient due to variable expressivity.
FIGURE 12-4 Clinical variation of patients with albinism
and infantile nystagmus. X-linked albinism has been called
“ocular albinism.” However, patients have abnormal
macromelanosomes in skin, and while peripheral pigment
may appear normal, it may not provide full sun protection.
Albinism patients commonly have hypopigmented irides
(A, arrows), hair and skin (B), and fundi (C). Autosomal
recessive oculocutaneous syndrome may be associated
with either scattered or diffuse iris transillumination
defects (D and G, arrows), mild or profound
hypopigmentation of the skin and hair (E and H), and
reduced retinal pigment (F and I). The amount of pigment
produced depends on whether the mutations completely or
incompletely block pigment formation, but the patient’s
gene cannot be predicted based on clinical features (D–F
compared to G–I).
A relative afferent pupillary defect (RAPD) increases the likelihood that there
is an optic nerve pathology as the INS etiology, whereas bilateral sluggish
pupils may be present in certain retinal dystrophies, such as LCA.
Paradoxical pupils can be seen in a number of congenital optic nerve and
retinal disorders. It should be noted if the child has good vision or poor
vision; best-corrected visual acuity helps guide the workup, as described later
in this chapter. The preverbal child should be evaluated using the CSM
method: central fixation (C), steadiness of fixation (S), and maintenance of
fixation (M). To determine if the child has central fixation, each eye should
be covered and then uncovered while the child fixates on a target of interest
to determine if fixation is central or eccentric. Similarly, testing for steadiness
is done monocularly to determine if the patient can fixate without nystagmus.
Maintenance is a binocular test; for orthotropic patients, a prism of 14
diopters is placed in front of one eye base down as the child views a target of
interest at near and then at distance. If vision is equal, the eye behind the
prism will deviate upward to view the displaced image; as the second image
is noted in the fellow eye, the eyes will then deviate downward. If one eye
sees better than the other, the eyes will remain in the position corresponding
to the image for that eye. If a child has strabismus, alternately covering the
eyes and watching for movement of which eye is fixating can be done
without a prism. The eyes should be observed in all cardinal gazes to
characterize the nystagmus waveform, to detect the presence of a null zone,
and to identify red flags. Gaze-evoked nystagmus with changes in the plane
of oscillation may be associated with posterior fossa lesions (3).
Cycloplegic refraction may detect high refractive errors and guide further
testing, such as LCA molecular testing in INS patients with high hyperopia
and poor vision. Strabismus and amblyopia are detected in up to 35% and
14%, respectively, of all pediatric patients with nystagmus (2); however,
compared to INS, latent nystagmus is more commonly related to comorbid
amblyopia, strabismus, or syndromic disorders (e.g., Trisomy 21) that disrupt
binocularity (3). Slit lamp or penlight assessment help rule out lens, cornea,
or iris abnormalities. Iris transillumination defects, deficient iris tissue, ovoid
pupils, or ectopic pupils should be investigated further for disease processes
such as albinism or PAX6-associated ocular diseases (e.g., aniridia or partial
aniridia).
The dilated fundus examination should be carefully performed, and there
should be a low threshold to perform an examination under anesthesia (EUA)
if the fundus view is inadequate or if the child is uncooperative in the clinic
setting. The optic nerve should be evaluated for colobomatous changes and
disc size; optic nerve hypoplasia commonly leads to INS and can be
associated with pituitary and/or septum pellucidum defects (septo-optic
dysplasia) secondary to congenital malformation. Brain MRI to assess
pituitary and midline brain anatomy as well as endocrine evaluation is
indicated in cases of isolated optic nerve hypoplasia. Optic nerve hypoplasia
may also be a feature of albinism or aniridia, and if it can be proven that it is
part of these syndromes, brain MRI is usually not needed as these types of
optic nerve hypoplasia are not reported to be associated with pituitary
dysfunction. The combination of optic nerve atrophy and INS warrants head
imaging to rule out intracranial pathology. Foveal hypoplasia suggests the
presence of albinism or aniridia but has also been reported to occur in
Stickler syndrome, familial exudative vitreoretinopathy (FEVR), premature
birth, and as an isolated anomaly. Retinal pigmentation abnormalities,
arteriolar attenuation, and/or blunted foveal light reflex should raise suspicion
for LCA, retinitis pigmentosa (RP), achromatopsia, or other inherited eye
diseases that should be investigated, as described later in this chapter.
Many children with INS will initially present to pediatricians or
neurologists, and they will undergo an MRI before a complete eye
examination is performed. A complete history and a complete pediatric eye
examination with dilation as well as ancillary tests of ocular function are
necessary for all children with INS, even those with a negative MRI. A
negative brain MRI should never be the only test performed on a child with
INS.
General Considerations for Testing

The choice and order of testing for INS should be based on family history,
symptoms and signs, and findings of the eye and neurologic examinations.
Nonspecific “shotgun” testing tends to yield equivocal or nondiagnostic
results; this increases the financial and time burden for families and can put
undue stress on families, especially if false positive or negative results are
obtained. This chapter highlights a cost-effective diagnostic approach
(Figure 12-5) that minimizes these burdens and improves accuracy in
diagnosing causes of INS.
FIGURE 12-5 Diagnostic algorithm for infantile
nystagmus. Red text indicates the most common diseases
and/or genes that should be further investigated given the
clinical findings and testing results. This algorithm is not
comprehensive for all possible diagnoses but can help
narrow the diagnosis to a specific category.
A cycloplegic refraction should be the first step in the diagnostic approach.

The clinician should determine if the child has poor vision or good vision
with best correction and whether nystagmus resolves with optical correction.
Generally uncorrected high myopia greater than about −15 diopters and high
hyperopia greater than around 10 diopters can cause visual deprivation in
infants; once optical correction is worn for several weeks, the nystagmus
improves or resolves. Such high refractive errors may not be isolated,
however, and further workup should be considered for entities such as
connective tissue disorders and retinal dystrophies and degenerations.
Brain Imaging
INS alone does not necessarily warrant obtaining brain imaging given the low
diagnostic yield of MRI testing in the absence of other findings. However, a
low threshold remains for ordering a brain MRI with and without contrast on
a child with INS who has asymmetric or unilateral nystagmus, neurologic
signs, developmental delays, small or large head circumference for age,
and/or changing growth curve percentiles over time. If nystagmus onset was
after 6 months of age, brain MRI should be considered. It should be noted
that dominant optic atrophy usually presents with temporal optic nerve pallor
and slow vision loss without the development of nystagmus. Thus, evidence
of optic nerve atrophy or hypoplasia in patients with INS should warrant
brain imaging. The shimmering, asymmetric nystagmus seen in patients with
spasmus nutans should always be further evaluated given that certain CNS
tumors, especially diencephalic syndrome, can mimic spasmus nutans, which
presents with the triad of nystagmus, head nodding, and torticollis (9,15).
Children with significant perinatal history (e.g., prematurity, traumatic brain
injury, hypoxia, infection, etc.) should be evaluated with a brain MRI to
identify structural causes of cortical visual impairment (3). If a child has a
known ocular etiology of his/her nystagmus but has additional unexplained
neurologic findings, it is prudent to investigate further. For example,
progressive optic atrophy in a patient with known CSNB would be
uncharacteristic for the ocular disease state and may represent an
undiagnosed intracranial disease process.
A negative brain MRI is reassuring that the cause of INS is likely not
neurologic, but it should never be the final test in the workup. A negative
MRI means ocular causes should be explored. Even in the case of spasmus
nutans, which has been described as a developmental, self-resolving
condition of the first few years of life, a normal MRI should be followed by
an electroretinogram (16). There are many reports of CSNB and
achromatopsia presenting as spasmus nutans-like nystagmus that resolves as
the child ages. Conversely, some LCA patients present with a spasmus
nutans-like nystagmus that usually does not resolve over time.
Electroretinography
Electroretinography (ERG) is a powerful diagnostic tool for INS and should
be obtained in any case with a nondiagnostic MRI. It can also be helpful to
obtain prior to head imaging when evaluating patients with poor vision,
nystagmus, and no other findings; the diagnostic yield for these cases has
been reported to be 56% (17). A normal ERG does not rule out all ocular
causes of nystagmus and does not replace a complete ophthalmologic
examination. In general, a flat ERG suggests severe cone and rod disease,
likely secondary to LCA or RP. Abnormal cone responses with relatively
preserved rods are seen in achromatopsia and cone dystrophies, whereas an
electronegative ERG should increase suspicion of CSNB, X-linked juvenile
retinoschisis (XLRS), or Batten disease. ERG may be successfully performed
in awake children in clinic from birth to about 6 months of age and then after
about 3 to 4 years of age depending on the child. A patient, expert clinician is
necessary to perform atraumatic pediatric ERGs. Dawson-Trick-Litzkow
(DTL) electrodes, which allow normal blinking and are less difficult to apply
than contact lens electrodes, greatly reduce the trauma of pediatric (and adult)
ERGs and yield comparable amplitudes with only slightly more background
noise. Jet electrodes and skin electrodes can also be used. Each electrode has
its own pluses and minuses, which should be thoroughly understood by
whomever is interpreting the test to avoid under- and overdiagnosis of retinal
disorders. It should also be remembered that the ERG evolves over the first
year of life. A very low-amplitude ERG at 6 months of age may evolve into a
higher amplitude or electronegative ERG at 18 months of age. If ERG in the
clinic is not possible but is necessary, ERG can be performed under
anesthesia. Amplitudes may be decreased by up to 50% due to anesthesia,
particularly the rod ERG, so this must be taken into account when
interpreting these ERGs.
There is increased crossing of temporal ganglion cell axons in the optic
chiasms of patients with albinism, and this can be detected using a
multichannel visual-evoked potential (VEP) (18). VEP is easier to perform in
children than ERG because electrodes are placed on the head rather than the
eyes. However, detection of abnormal decussation in albinism is variable and
may become undetectable at older ages. This VEP finding has also been
described in aniridia and, therefore, is not specific enough for clinical
diagnosis.
Optical Coherence Tomography

Detailed assessment of the fovea and retinal structure with OCT helps
identify ocular disorders associated with INS, such as foveal hypoplasia and
retinal dystrophies (19). Handheld OCTs can be used clinically in the
operating room during an EUA if the child is not old enough or cooperative
enough to obtain OCT in clinic. OCT with motion tracking technology can
often capture a useful image in children as young as 3 years old in the clinic,
even when nystagmus is present. In combination with genetic testing, OCT
provides a noninvasive and accurate diagnostic approach to detecting
inherited eye diseases.
Molecular Genetic Testing

Ocular abnormalities, retinal structure changes on OCT, and ERG changes all
increase or decrease the pretest probabilities of specific ocular disorders.
Most vision-threatening causes of INS can be attributed to mutations in a
single gene. With high clinical suspicion of a certain inherited disease state,
gene panel testing is often the most cost-effective and timely method for
arriving at a diagnosis (20). For instance, FRMD7 genetic testing may be
performed to establish a diagnosis of X-linked motor nystagmus in young
patients with good vision, a normal eye examination, and nystagmus in a
pedigree consistent with X-linked inheritance, often with manifesting females
(21–23). The diagnostic yield of FRMD7 genetic testing has been shown to
be 20% to 57% in idiopathic INS cases with an X-linked pedigree (24). Care
must be taken to test with both sequencing and deletion/duplication testing as
the latter may be missed on typical panels yet has been documented in
FRMD7. On the other hand, in a child presenting with infantile nystagmus,
poor vision, and no family history, an LCA panel is often the best first step
(25). If the diagnosis is more equivocal, whole exome sequencing may be
indicated. In whole exome sequencing, the entire exome may be sequenced,
and then the genes of interest, for example, all genes known to cause retinal
disease, can be analyzed.
A recent study describes utilizing a “nystagmus gene panel” in 48
infantile nystagmus patients with a diagnostic yield of 58% (26). In this
cohort, causative mutations were detected in FRMD7, GRP143 (OA1), and
PAX6 as the major genetic causes of inherited INS (26). This gene panel did
not include CSNB and other important genes, and some cases of INS are
environmental rather than genetic, which accounts for the relatively low
diagnostic yield.
Confirmation of certain genetic diseases, such as albinism or aniridia,
allows the patient to forgo other testing as MRIs are not routinely needed in
these cases. An accurate diagnosis improves patient-centered care by
incorporating genetic counseling, family planning, and disease-specific
management. Appropriate referrals can also be made according to the genetic
diagnosis.
Eye Movement Testing

Nystagmography is not routinely available in the clinical setting, yet such
testing could lend itself to meaningful research and may ultimately contribute
to the workup of INS. These recordings may be used to determine change in
amplitude or frequency of nystagmus waveforms after surgical or medical
therapies (6). There are no nystagmus waveforms that are specific to an
ocular diagnosis (Figure 12-2). Roving nystagmus, for example, is
overrepresented in LCA but can also be seen in patients with albinism, optic
nerve hypoplasia, and PAX6-associated diseases. Ocular flutter in pediatric
populations is reportedly highly specific for Joubert syndrome, yet this form
of nystagmus is uncommon in these patients (i.e., low sensitivity).
MANAGEMENT
Optical Treatments
INS can result from uncorrected high hyperopia, high myopia, or high
astigmatism; optimal refractive correction and treatment of amblyopia are
essential for these children. Thus, a cycloplegic refraction must be performed
on every child presenting with nystagmus. For adult patients with lifelong
nystagmus and binocular vision, base-out prisms can be used to induce
convergence during the viewing of far targets; convergence-induced
accommodation must be accounted for in the final prescription with the
addition of minus lens power in young patients (6). A recent randomized
controlled trial of contact lens therapy in INS failed to show reduction in
nystagmus. However, contact lenses, which reduce the prismatic effects of
spectacles and keep the optical center of the lens in line with the center of the
pupil, remain an option for children with nystagmus and high refractive
errors. Some clinicians add prisms to correct a small anomalous head position
where the apex is pointed in the direction of the patient’s gaze (6).
For INS patients with poor vision, vision rehabilitation services should be
provided to help these children perform everyday tasks. Use of electronic
devices, large print, tablet computers, and magnification devices can
significantly improve their quality of life (27).
Extraocular Muscle Surgery

The extraocular muscles in INS patients appear to have increased variation in
their muscle fibers and altered proprioceptors compared to controls (6,28).
The significance of this proprioceptive disturbance as it relates to a primary
ocular motor control system defect has not been fully elucidated. Tenotomy
with reattachment of muscles to their original insertion sites has been
reported to minimize eye movements (and increase foveation periods)
documented by nystagmus recordings, possibly by changing the muscles’
proprioceptor function and decreasing the peripheral oculomotor response
(6,29). This procedure does not completely resolve nystagmus, however, so
patients must be carefully counseled about expectations. Alternatively, the
Anderson-Kestenbaum procedure shifts a patient’s null zone into primary
position and corrects abnormal head positions using recession–resection of all
four horizontal rectus muscles or of vertical or cyclotorsional muscles
depending on the abnormal head position. Performed since the 1950s, this
surgery decreases the amplitude and frequency of nystagmus in primary
position and corrects torticollis during active fixation (30). One caveat is that
the resting position of the eyes is shifted to an eccentric position; thus
patients may develop a new head turn when not actively using detailed vision
and may adopt the straight head posture only while reading, driving, etc.
Select patients undergo artificial divergence procedures (i.e., bilateral
medial rectus recessions) that require prolonged convergence of the eyes that
ultimately dampens the nystagmus and broadens the null zone. The
preoperative measurement of fusional amplitudes is important prior to
creating such a divergent resting position (6). Other surgical interventions for
nystagmus include various combinations of muscle recession and resection.
Several studies have demonstrated that the symmetric recession of all four
horizontal extraocular muscles has a significant impact on nystagmus
amplitude and frequency, visual acuity, and head positioning (31);
complications of this procedure may include anterior segment ischemia.
Botulinum toxin injections have been performed into the extraocular muscles
and into the retrobulbar space (32,33). There are no randomized control trials
comparing these surgical techniques and their effects on nystagmus.
Pharmacologic Therapies
Systemic medications are not commonly used in INS given the side effects
that may occur with long-term use. Furthermore, most medical therapies are
generally not effective for afferent visual system problems (i.e., secondary
nystagmus), which constitute the majority of INS. The available treatment
options aim to increase the duration of foveation periods and minimize visual
symptoms that are more common in adult populations (6). Baclofen, which
activates gamma-aminobutyric acid receptors, has been shown to be effective
in improving visual acuity and correcting abnormal head postures in patients
with infantile periodic alternating nystagmus; however, half of these treated
patients developed side effects requiring medication cessation (34). One
double-blind randomized controlled trial in 2007 showed that both
gabapentin and memantine reduce the nystagmus intensity (i.e., amplitude ×
frequency) and slightly improve visual acuity in idiopathic INS patients
without serious side effects (35); however, the mechanism of these effects
remains unknown. Oral and topical carbonic anhydrase inhibitors (e.g.,
brinzolamide, acetazolamide) reportedly suppress INS, but larger studies
showing efficacy are needed (36).
Subretinal Gene Therapy
The blood–retina barrier limits the risk of immune reactions to subretinal
gene therapy; there are several retinal degenerative diseases that cause INS
that may be amenable to such therapy. Clinical trials for these disorders can
be found at www.clinicaltrials.gov.
The causative gene must be known in order to determine if a patient will
qualify for a trial. Only one gene therapy for a retinal degeneration causing
INS is clinically available: Luxturna for RPE65-associated LCA. The Food
and Drug Administration has approved subretinal administration of an adeno-
associated virus gene vector containing a normal copy of the RPE65 gene in
both eyes of patients. This treatment has led to meaningful improvements in
ambulatory vision and nystagmus for children with this blinding condition
(37,38). Not all adults experienced marked improvement; however, some did,
including a 44-year-old patient. The treatment is approved for children as
young as 1 year old (although the youngest patients in the clinical trial were 4
years old) and adults with RPE65-associated LCA or RP.
VISION REHABILITATION
Some patients with INS have normal vision and do not need vision
rehabilitation; however, the majority of INS patients will benefit from special
accommodations at school, use of technology, and special evaluation for
driving licenses, test taking, using a microscope (a microscope with a video
screen is required), and other highly visual tasks. INS patients with best-
corrected visual acuity of <20/40 or any field loss should be referred to a low
vision clinic. Chapter 14 describes the low vision services available in more
detail.
ROLES OF OTHER PHYSICIANS AND

Understanding that INS is a sign rather than a diagnosis is the first step in
caring for these patients. The importance of a diagnosis includes referring
children to the appropriate specialists for other features of their syndrome, if
one exists (Table 12-2).
TABLE 12-2 Some systemic diseases associated

with infantile nystagmus
Making a clinical diagnosis of LCA as the cause of INS is only the
beginning. A molecular genetic diagnosis is necessary to know which
patients to refer for regular renal ultrasounds and kidney evaluations. LCA,
due to mutations in the NPHP genes (CEP290 and IQCB1) may be associated
with Senior-Löken Syndrome, which can have life-threatening consequences
due to early-onset kidney failure. RPGRIP1-associated LCA patients may
also be at risk as this gene participates in the same pathway. Patients with
these mutations should be referred for renal ultrasound and evaluation. BBS
and Joubert syndrome both are ciliopathies that result in retinal dystrophies
with profound systemic manifestations and developmental delays.
BBS is associated with polydactyly, cardiac abnormalities, obesity, and
renal dysfunction (39), whereas Joubert syndrome results in hypotonia, liver
and renal dysfunction, and respiratory issues, among others (40). Chediak-
Higashi syndrome, a form of OCA, can lead to recurrent infections secondary
to profound neutropenia (41). About 4% of albinism patients have
Hermansky-Pudlak syndrome, which includes a bleeding diathesis due to
abnormal platelets and pulmonary fibrosis (42). The medical literature
suggests that Hermansky-Pudlak syndrome occurs mostly in Puerto Rican
albinism patients and that most patients have significant peripheral pigment;
in the age of genetic testing, we know this is not correct. Genetic testing
should be offered to all albinism patients regardless of ancestry or
pigmentation, so appropriate systemic treatment can be given for those with
this syndrome. Optic nerve hypoplasia, which is commonly associated with
septo-optic dysplasia, warrants referral to a pediatric endocrinologist as these
children may have multiple hormonal deficiencies including a deficiency of
endogenous cortisol secretion with febrile illness, which can be fatal if not
treated. All patients with albinism have a greatly increased risk of skin
cancer. In addition to reminding these families to use sun protective clothing
and lotions, patients should be referred to dermatology for skin surveillance
starting in adolescence, sooner if suspicious skin lesions are present. In
general, once a clinical or molecular genetic diagnosis is made, patients and
families should be referred to a medical geneticist or genetic counselor to
discuss inheritance and recurrence risk. Communication with the child’s
pediatrician should be done to aid in the referral and workup process for
syndromes. An overview of the workup and management of several systemic
diseases associated with INS is provided in Table 12-2.
Many of the disorders that cause INS are genetic. Some carry serious
systemic risks that can be mitigated if detected and treated early. One cause
of INS, RPE65 LCA, has a gene therapy treatment available, which is only
effective in patients with viable photoreceptor cells. This means earlier
diagnosis may have some beneficial effect on treatment outcome. Parents of
children with INS may wish to explore options to decrease their risk of
having other affected children. Genetic counseling can only be performed
accurately when the diagnosis is known. In vitro fertilization with
preimplantation genetic testing can be offered only if a molecular genetic
diagnosis is known early in a subsequent pregnancy. Therefore, it behooves
physicians to offer a complete workup to determine the etiology of INS. This
may involve referral to a genetic eye disease specialist or medical geneticist.
Some families may choose not to pursue definitive diagnosis, but it should be
offered.
FUTURE TREATMENTS
Patients with intact outer and inner retinal layers may be candidates for gene
therapy; in these trials, intraocular transplantation of a viral vector containing
the gene of interest leads to the transduction of affected photoreceptor cells,
which may be beneficial. In more advanced disease states, stem cell therapy
to replace lost photoreceptor cells is needed. Data show that cultured human-
induced pluripotent stem cells have the ability to form all retinal cell types,
integrate into retinal layers, and contribute synaptically (43–45). These cells
have substantially improved vision in animal models; when injected into
mouse RP models, these cells survive and differentiate into functional
photoreceptor cells (46). Research is ongoing into this type of treatment and
may offer hope to patients with nystagmus due to photoreceptor loss in the
future.
CONCLUSIONS
INS is an oscillatory eye movement disorder that presents before the age of 6
months. Diagnosis and management of INS requires a clinician to obtain a
thorough history, perform a detailed clinical examination, and order selective
ancillary testing to determine the cause. In many INS cases, there is an
underlying retinal disease that can be readily diagnosed with cycloplegic
refraction, a dilated fundus examination, heightened clinical suspicion, and
appropriate studies, including ERG, OCT, and genetic studies. Neurologic
causes of INS in otherwise developmentally normal healthy children are rare
but should not be overlooked; thus, a brain MRI should be ordered when
clinically indicated. Other specialists should be involved in cases of
syndromes associated with INS. Pharmacologic, surgical, and optical
management of underlying ocular disorders and systemic diseases is an
essential part of addressing the needs of pediatric patients with infantile
nystagmus. As gene therapy and other regenerative medicine techniques
become available, the importance of an accurate clinical diagnosis in these
patients becomes even more important.
ACKNOWLEDGMENTS
The Ronald Keech Professorship supported this work.
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13
Early Intervention and Rehabilitation
in Infants and Children With Visual
Impairment
Lea V. M. Hyvärinen
INTRODUCTION
Changes in retinal structure and functioning begin in infants and children at
varying ages and affect vision. Diseases of the visual system often present in
this age range, some of which also involve hearing, cognitive, and motor
functions. More than 50 groups of diseases with variation in severity are
covered in this book and present a broad range of phenotypes within and
among diagnoses. Several retinal diseases are parts of syndromes or occur in
children, some following intrauterine infections. Therefore, the effects of
diseases on the visual system and on general functioning vary greatly and
require numerous observations, vision tests, and follow-up to define a child’s
needs in the early intervention and later in special education and
rehabilitation. For each child and family and the rehabilitation service/school,
the findings of the clinical examinations should depict, clarify, and support
the functions and skills observed by parents, therapists, and teachers so that
the child’s visual functioning and needs for further development can be
understood and planned correctly.
The problems in early intervention and rehabilitation are in three areas:
1. Early detection and referrals are often delayed.

2. Clinical examination and assessments do not cover all vision problems
and visual functioning.
3. Special education and rehabilitation services do not receive enough
information on multidisabled children because the eye clinics do not
have special tests for “difficult-to-test” children. Students with
ergonomic challenges are best tested at special schools where therapists
and teachers can arrange the test situations to meet the needs of each
student and the visiting opthalmologic/optometric specialists. Teachers
of the visually impaired (TVI) and other special educators can repeat the
tests in the beginning of each school year, which decreases work at
clinics and students traveling to hospitals. Children’s ergonomic
problems are assessed much better at special schools than hospital
clinics, because the student’s own teachers are instrumental during
testing of individual learning strategies.
EARLY DETECTION
Early detection of diseases and disorders of vision requires systematic
observations of infants’ and children’s visual functioning by the medical and
educational services and referrals to pediatric ophthalmologic services. All
pediatric medical services should have awareness campaigns on sensory
functions, and each newly diagnosed visual disease in ophthalmologists’
offices should lead to consultation at pediatric, neuropediatric, and/or
neuropsychological services so that early intervention, rehabilitation, and
educational services can be wisely chosen for each child and family.
Premature infants with or without retinopathy are a large group with
atypical vision; they may require long treatments in hospital with follow-up
and treatment of retinopathy of prematurity (ROP). They may also have
changes in brain functions leading to visual processing disorders and visual
field losses and changes in other sensory, cognitive, and motor functions.
Contact with parents may be limited due to the infant being in an incubator
connected to monitors, and, therefore, development of bonding and
interaction is in danger in these families.
Early intervention has not developed to cover problems in infants’
development of functioning caused by retinal damage and prematurity. The
medical care has improved with standardized treatment. Similarly, early
intervention for visual dysfunction is likely to improve if assessment of
vision starts early and covers several functional areas. Present problems
include delay in diagnoses other than ROP and issues related to treatment and
variation in the testing and training vision. Therefore, further work is needed
to optimize visual development.
In the follow-up of development of vision, the Teller Preferential
Looking Grating test for research laboratories seems to be used most often,
but the values are reported as visual acuity values, which are not correct.
Preference and response to looking at the grating in preferential looking tests
(Teller and LEA Grating tests [www.lea-test.fi]) do not reveal whether the
infant has good detail discrimination, only whether one grating looks more
interesting than another. Optotype values on the Teller Preferential test were
demanded by the ophthalmologists, although Davida Teller explained that
they were not valid to describe visual acuity. The grating can be seen badly
distorted, and yet it is a strong stimulus, preferred to an evenly gray or white
area. Errors in measurement have been noted from the earliest days of grating
acuity (GrA) testing (Figure 13-1): correct GrA cannot be measured
accurately until the child can understand and respond to questions regarding
the intricacies related to visual function. For example, the question of where
he sees the lines: “at the edge or in the center” and “whether lines are straight
or tangled,” the answer can be “There are no lines in the middle.”
Figure 13-1 When the first, 4-degree grating test was

created and was ready for use, there were no children in
the laboratory, so a young man was asked to be the first
“patient.” After the measurement, he was told that he had
symmetric, normal vision in his eyes. He answered: “That
is not possible; one eye is normal, the other is amblyopic. I
cannot read with it.” He drew pictures of gratings as he
saw them with his amblyopic eye; drawing was guided by
his normally sighted eye. In each grating, the distortion
was in the middle and there was enough information at the
edges to answer in what direction the lines were. Thus, the
wrongly measured GrA values of the two eyes were
symmetric. (Reprinted with permission from Springer:
Hess RF, Campbell FW, Greenhalgh T. On the nature of
the neural abnormality in human amblyopia; neural
aberrations and neural sensitivity loss. Pflugers Arch
1978;377(3):201–207.)
Distortions of fine grating lines occur in many conditions that disturb exact
coding of the line information in the retina or in the primary visual cortex
(V1) in the occipital lobe.
The use of grating tests requires training, so that the tester shows first
broad full contrast lines that are easy to see and the child shows with hand or
on answering cards the direction of the lines. When the direction of lines is
understood, the tests are shown according to the instructions (in www.lea-
test.fi). This may be possible first at the age of 4 to 6 years. We should be
aware of the difference between gratings and optotypes and know that the
preferential looking tests do not result in a real grating acuity value but show
the preferential response. These tests are useful to detect a marked preference
for one eye, possibly signifying amblyopia or organic vision loss, but not the
absolute visual acuity of either eye.
Active use of vision on day 1 or 2 has been registered in thousands of
pictures showing stable eye contact with the mother in some pictures even
copying expressions. In Chitrakoot, India, a pilot study on feasibility of
assessment of stable eye contact as a part of the routine examination of facial
structures, eyes, and red reflex by the pediatrician was found to be easily
performed and took <1 minute during the discussion with the mother. Of the
70 newborn infants, all but 1 had typical stable eye contact when it was
initiated by the mother. This nonresponding infant was premature; her
refractive error was +3.50, so test glasses +6 were placed in front of her eyes.
She immediately had stable eye contact. Accommodation had not yet
developed in this premature baby, but the visual response to eye contact was
present. This test is short and easy but requires either training of pediatricians
or access to a pediatric ophthalmologist or optometrist when the pediatrician
examines infants the day before they leave the hospital. Hyperopia and delay
in development of accommodation are assessed in 2 minutes when
noncycloplegic or “dry retinoscopy,” and a set of test glasses +2, +4, +6, and
+8 are used. If a newborn does not make eye contact with a parent on the day
of discharge, dry retinoscopy is performed, and trial lenses of that correction
plus three diopters for near are placed on the baby. The baby is then observed
with the parent again, and if eye contact is better, glasses with this
prescription are dispensed to be used until accommodation develops. If this
does not improve eye contact, an ophthalmology consult should be obtained.
This technique continues to be studied, but enough is now known to
recommend recording stable eye contact in newborns prior to hospital
discharge and suspecting deficient accommodation if it is not present.
Newborn and young infants use the tectopulvinar pathway that transfers
low-contrast information about motion, that is, the infants can see facial
expressions on adult persons’ faces and can even respond during the first
week when the mirror neuron network moves visual information to motor
activation of the baby’s facial muscles resulting in a smile and feeling of
happiness. The brain functions in the infant are activated, and the parents
support the infant response by increased bonding. At 10 to 12 weeks of age,
infants’ visual behaviors depict development best, although individual
variation is great. Some infants are “talkative” at the age of 8 weeks and use
vision and voice; other infants smile and listen until 12 weeks. The visual
milestones are a good guide during the first months (Table 13-1).
Table 13-1 Visual milestones in the first months

after birth
CLINICAL EXAMINATION AND
ASSESSMENT
Clinical examination for rehabilitation changed greatly, when in 1978, the
ICD-9 (1) defined low vision as three levels of visual impairment, based on
visual acuity (VA) values (2). It was meant only for reporting to the World
Health Organization, but this was not clearly expressed. Because there were
definitions of “impaired vision,” VA values were used to restrict activity in
work requiring good vision at low contrast, for example, truck drivers driving
in fog, rain, or snow fall; the limit, VA 20/40, is based on seeing the numbers
on traffic signs. In the United States, use of low-contrast tests was disproved
by insurance companies in the late 1980s, so contrast sensitivity tests were no
longer used. In 2011, contrast sensitivity was deleted from the International
Classification of Functioning, Disability and Health that received approval
from all 191 World Health Organization member states on May 22, 2001,
during the 54th World Health Assembly (WHA 54.21)
(www.who.int/classifications/icf/en/). Despite the change in rules, contrast
sensitivity should be measured in children in addition to visual acuity in order
to understand their visual functioning.
Visual acuity is not synonymous with visual function. Our vision is most
effective in low-contrast areas at VA values lower than 0.3, 20/60 (Figure
13-2). The area below the curve is largest at the range of low VA values.
Therefore, “low vision” is not necessarily “poor vision.” Originally, “low
vision” (introduced by Dr. Eleanor Fay in the early 1970s) covered people
who were not blind but were not fully sighted either. The WHO registered
only blind and sighted, so patients were often in a gray area between the two
extremes. “Low vision” was appreciated as a temporary definition, when the
exact nature of atypical vision could not yet be diagnosed or when it was
between blind and fully sighted. Visual acuity values alone do not define
impairment or the need for rehabilitation services, and designating an
individual as “impaired” may adversely impact them. Visual acuity can be
very low, even 20/2,000 (Figure 13-2C), but awareness of spaces, distances,
and directions can be good. One patient seen in our center had normal
contrast sensitivity at low VA and GrA and good motion perception. He used
Braille at school age, has a university degree, and has continued to travel
independently and perform research in many countries. Low visual acuity can
now be compensated by using technology and high magnification. If other
important visual functions are more or less normal, low VA may not cause
severe impairment. Children should be categorized according to visual acuity
for the purposes of receiving special assessments in school, but a discussion
with patients, parents, and teachers should include that acuity alone does not
define how an individual will function or what they will accomplish.
Figure 13-2 A:Contrast sensitivity curve depicts

functioning at all contrast levels (y-axis) within visual
acuity range (x-axis) used. B:Vision for communication
distance can be assessed with the Hiding Heidi (www.lea-
test.fi) low-contrast test at the age of 3 months (infant
responds to the picture of smiling face so he must see the
smile in the picture). C:Contrast sensitivity curve in 1977
of a boy with visual acuity 20/2,000 (from atrophy of the
occipital lobe) and GrA 4 cycles per degree (cpd). He had
normal contrast sensitivity curve at low grating acuities
until 1 cpd, where the curve bends down and reaches the x-
axis at 4 cpd. The boy had good motion perception.
20/2,000 does not alone define functional blindness.
D:Slope of the contrast sensitivity curve from VA 20/100
on x-axis to unusually high values at low contrast (0.6%
contrast) of a 45-year-old patient with X-linked
retinoschisis who drove a vehicle without any difficulty in
Finland to the age of 70 years. (Reprinted with permission
from Hyvärinen L, Jacob N. What and how does this child
see? Assessment of visual functioning for development
and learning, 2nd ed. Helsinki: VISTEST Ltd., 2013.)
Contrast sensitivity depicts vision at all contrast levels (y-axis) at the visual
acuity (x-axis) range used as in Figure 13-2A, C, D. The slope of the contrast
sensitivity (CS) curve can be quickly defined, because between full and 2.5%
contrast, the slope is close to a straight line that depicts the curve well
(Figure 13-3).
Figure 13-3 Slopes of contrast sensitivity curves of 50

Finnish students in special schools for children with motor
problems.
Most students in Figure 13-3 had cerebral palsy (CP). The slopes varied in
their inclination; some slopes are steep and have almost the same value at full
contrast VA on the x-axis and at the level of 2.5% contrast. Another large
group has slightly more difference between these two values; the inclination
of the slope is a little flatter than in the first group; in the third group, the
slope is very flat, that is, the difference between VA values at full and 2.5%
contrast can be 20/12 to 20/200. The low-contrast visual acuity values add
important information in the assessment of functioning and takes <2 minutes
to measure with the 2.5% chart after the full contrast value is measured. In
this group of 50 students, VA was <20/60 in 12 students; in 10 students, it
was 20/20 or higher. Note that the majority of students had VA > 20/60 (0.3).
In most of the United States, in order to obtain an unrestricted driving license,
visual acuity must be at least 20/40 in one eye; restricted driving, such as
daytime only or off highway only, may be possible at lower acuities. Chapter
14 discusses the low-vision evaulation for driving in more detail.
In 2008, visual acuity limit 20/60 (0.3) was used for the first time to
divide students with visual processing disorders into those with VA < 20/60
(“low vision”), who had “cortical visual impairment” (CVI) who were
eligible to have TVI services, and those with VA > 20/60 who were not
eligible for services. This diagnosis was an educational diagnosis, but was
soon used as if it were a medical diagnosis.
Within a few years, 20/60 became the VA value that divided all children
with visual disorders, so only those with VA < 20/60 were examined in many
university hospitals worldwide. Visual acuity was recently recommended not
to be a divider of services in the United States, but follow-up information is
not yet available. In India, this VA limitation ended earlier this year. Eye care
providers in the United States often measure only distance VA in children.
This decreases the possibility to notice weak accommodation as a potential
common problem in school. Often near work magnification need is based on
distance vision. Near vision is the most used visual function in children in
demanding tasks, especially in school age and should be tested at each visit.
Children with amblyopia or certain visual processing disorders may have
reduced ability to read due to the crowding phenomenon. In this condition,
letters in close proximity appear to overlap each other and become
unreadable. If a patient with the crowding phenomenon is tested using
individual widely spaced optotypes, acuity will be normal, yet they are
unable to read words of the same size. The LEA near vision test (www.lea-
tes.fi) used at 40 cm distance (the length of the cord on the test) has the
standard line test with typical spacing between letters. On the reverse side,
the tests have tighter spacing between the letters at 50% and 25% to detect
crowding. If crowding is detected, large texts can be used to improve reading
ability in schools, even if line acuity is normal. This near test can be used as a
distance test at 2 or 3 m distance, so there are five tests on this small card that
was created especially for developing countries where rehabilitation teams
must travel long distances and bring test materials with them. The small LEA
low-contrast VA test, the 10M test, and the Hiding Heidi test (www.lea-
test.fi) for assessment of communication distance complete the VA testing
material until the students start to use local letters and numbers. These
abstract forms are processed differently from the LEA symbols that are
pictures of concrete objects and thus can be used earlier than abstract letters
and numbers. Letters are composed of short straight and curved lines, so
grating acuity should be tested as early as possible.
Motor problems, such as poor head control, or neurologic impairment of
the visual system may also affect ability to track and process words (3). If a
child has atypical “dyslexia,” a small dome magnifier with an arrow inside
can function as a “reading stick” and may stabilize fixation.
Visual fields can have atypical structure: a hemianopic field can contain
motion perception (4). The tectopulvinar pathway may transfer visual motion
information normally (4) while damage in the optic tract causes loss of
hemifield measured with automated perimetry. The size and structure of the
visual field in retinal degenerations should be assessed in young children (5)
by observing strategies that the child uses in moving, such as searching for
edges with feet or hands. The peripheral visual field may require large stimuli
for the measurement. The child depicts vision of large surfaces in the
environment, which is important. The “blind” areas in automated perimetry
should not be marked with black before motion vision is measured within
them, because movement may still be perceived in these fields.
All measurable functions should be assessed and recorded in teenagers
and young adults with retinal diseases and atypical vision when training for
driver’s licenses. Diagnoses should be based on careful assessment of visual
functioning in clinics specialized in visual functioning in demanding
conditions, so the limits for driving will be individually tested (Figure 13-
2D). VA is the limiting function, but the test may not be standardized with
the international standard, the Landolt C test; VA values based on
nonstandard tests should not be used. See Chapter 14, which discusses
driving with low vision in more detail.
Visual pathways are not well known, especially the tectopulvinar
pathway. The tectopulvinar pathway functions at birth and transfers low-
contrast information in motion; therefore, newborn infants can perceive facial
expressions on adults’ faces. Their mirror neuron networks change visual
information to motor functions in the baby’s facial muscles allowing infants
to smile on the 2nd day of life and feel happiness, which activates brain
functions. Newborn infants’ vision is not poor, “less than 20/200”; they
see motion well in low contrast so they perceive facial expressions.
Assessment of visual functioning requires that all visual areas are
considered (Figure 13-4): (a) Visual information leaving the eyes via the
retinocalcarine pathway changes its content in the LGN nucleus based on
information arriving from specific cortical areas (marked with the thick arrow
from V1 to LGN in Figure 13-4C) so that less information leaves from the
LGN compared with what arrived, and this information is then carried on to
specific cortical areas. (b) Early processing in the occipital cortex (Figure 13-
4B) is the “feature level” coding for colors, forms, length and direction of
lines, depth, movement, motion, stereovision, textures, surface qualities,
object background, figure ground, visual closure, filling in, and visual
illusions. In V1, tactile and haptic information, and auditory space are
coded, so that they can be recognized and used with visual information in the
higher processing functions in the large parietal and temporal networks at the
“object level.” In education, children’s difficulties at the feature level
dominate in preschool and early grades and should be detected, because if
started early, a well-planned training often normalizes functioning.
Figure 13-4 A:The retinocalcarine pathway (blue) with its
lateral geniculate nucleus (LGN) transfers information to
the first visual area, V1; the tectopulvinar pathway (green)
with its pulvinar nucleus (PULV) transfers information to
V5/MT. B:Decoding and recoding of information, colors,
forms (lines, edges), and movement occur in different
parts of V1. C:Greatly simplified connections of functions
between the thalamic nuclei (LGN to V1), superior
colliculus (SC), and pulvinar to V5/MT and all visual
areas in the occipital lobe. Notice the two-way movement
of information and the thick arrow from V1 to LGN.
D:From the early processing in the occipital lobe, the
“feature level visual information” moves into the large
temporal and parietal networks for the “object level”
information processing. NOTE: More information moves
to the occipital lobe from higher cortical areas than from
the occipital lobe toward temporal and parietal lobes
(arrows are thicker toward occipital lobe). (From
Hyvärinen L, Jacob N. What and how does this child see?
Assessment of visual functioning for development and
learning, 2nd ed. Helsinki: VISTEST Ltd., 2013.)
Ophthalmologists usually have limited time for testing at the feature level of
early processing, so rehabilitation therapists, TVI, and others generally
administer and use these tests. At the age of 4 to 5 years, students with
complex processing problems benefit from testing by pediatric
neuropsychologists to get an overall picture of the functions that should be
trained.
Visual Processing at the “Feature Level”
Processing functions at the feature level in the occipital lobe are
especially important in preschool and the first grade in school, so that the
abstract space for forms and math develops. Many teachers observe preschool
and first grade children’s basic visual concepts and eye–hand coordination to
choose additional training. Therefore, the information that is found in clinical
assessments is especially important for the development of these young
students.
Together with special schools, easy-to-use tests were developed for
feature level processing when the first Vision Rehabilitation Centre in
Helsinki, Finland, started in 1976 (Figure 13-5).
Figure 13-5 A–D:Examples of the play situations to

assess children’s way to experience sizes and directions in
eye–hand functions, coordination, and motion. E:The
original “Johansson’s Walking man”-test and Pepi-figure-
in-motion test at full contrast. (Related videos can be
found at LEA-test.fi under Videos of the book tab.)
Visual Processing at the “Object Level”

Figure 13-6 is based on Riitta Hari’s article (6) on brain activities when
looking at a face: Simultaneous processes of face-related information in
several areas of the brain combine information from these networks into
expressions and features in the image.
Figure 13-6 Cortical functions during looking at a face.
Because information related to an object, for example,
face, constantly moves through several specific cortical
areas, young infants perceive the face first as facial
expressions (early motion perception by the tectopulvinar
pathway combined with mirroring); then, recognition of
different parts of the face can be learned with time.
Children with delay in recognition of faces report that eyes
and mouth disappear and reappear in faces at the time
when the child is learning to see them. In face blindness,
the structure of the faces of well-known persons remains
so weak that recognition is based on voice, hair, jewelry,
and any detail that is always present. These children may
not recognize the mouth in a simple drawing of face, even
if they know that the mouth is in the lower part of the face,
and need training with concrete pictures of the face that
the child makes using pipe cleaner for the mouth and large
dark buttons for eyes to use the concepts and words related
to “face.” Their motion perception may be atypical and
disturbed in lipreading.
EARLY INTERVENTION AND

REHABILITATION
Early intervention is an integral part of the assessment and treatment for
visually impaired infants and children and requires close collaborative work
with therapists, special education teachers, and social services to support
children’s development and the needs of their families. More than half of the
infants and children with impaired vision have one or several other
impairments. Therefore, early intervention and later rehabilitation of vision
during special education are recommended. This needs to be carried out in a
coordinated manner in conjunction with a team of educators, therapists, and
physicians and the family. Our framework in early intervention,
rehabilitation, and education of children with impaired vision is now the
International Classification of Functioning, Disability and Health for
Children and Youth Version (ICF-CY) (7).
Early intervention has three parts: (a) initial information; (b) early
intervention during the investigations and treatments in the hospital, which
may last from a day to months; and (c) early intervention when the
infant/child is at home.
INITIAL INFORMATION
Early intervention traditionally starts as a part of the first assessment of an
infant’s or child’s vision when there is a suspicion of vision loss or disease.
The parents are often worried about vision, especially if it affects early visual
communication. They are anxious to have a thorough description of their
infant’s condition. Most young parents have no experience with visual
impairment and may only relate to the alarming concept of total blindness. If
the infant is their first child, they have little knowledge on how to care for an
infant and virtually no information about the care of a visually impaired
infant. In such a delicate situation, it is important that the discussion of the
findings, prognosis, and planned management is undertaken by an
experienced pediatric ophthalmologist or pediatric retinal specialist who is
trained to discuss disability-related matters, knows the support systems, and
connects the parents with an early intervention professional who has the time
and the expertise to talk with the parents immediately after diagnosis. Ideally,
the early intervention professional should be present during the initial
discussion of the diagnosis and continue to support the parents. Parents may
remember only a fraction of what has been said. Terminology has been
frightening and confusing, and parents may feel overwhelmed and grieve the
loss of the perfect baby of their dreams.
Some doctors may contact the family the next day to make sure that the
family has connected with the local services and is not abandoned. If parents
are simply given a phone number to call an early intervention worker
directly, they may be unprepared to make that call for months since the
parents do not know that an early intervention worker assigned to them
would be able to assist them in developing communication and interactive
skills and to help them to enjoy and love their child without the reward of
normal visual communication. Without this assistance, the family may not
know how to care for their infant at the critical time of the early months,
which can lead to parental depression.
EARLY INTERVENTION DURING

TREATMENT
Some infants need surgical care within the first few weeks or months. The
infant in Figure 13-7 was born with cloudy corneas due to congenital
glaucoma. The operations on the left eye failed, and the corneal transplant
became cloudy. The right cornea was operated at the age of 5 weeks and
remained clear until the 15th week. This window of 10 weeks of clear vision
allowed the baby to see details, the parents, and her home. With the help of
neurodevelopmental therapy, she caught up with the delay in motor
development and later developed normally although she could see only bright
colors, shadows, and movements. She is now an adult with a university
degree and a rewarding job. The window of good vision after an operation
can be short, and therefore, it is wise to start supportive training without
delay.
Figure 13-7 A:Opening the eyes did not change the

image, so the baby kept the eyes shut. B:After the
operation of the right eye, she received near-correction
glasses and learned to use vision. C:She caught up with
the delay in motor development and has developed
normally despite her severe loss of vision. (Reprinted with
permission from Hyvärinen L, Jacob N. What and how
does this child see? Assessment of visual functioning for
development and learning, 2nd ed. Helsinki: VISTEST
Ltd., 2013.)
In general, pediatric surgical services and active (re)habilitation start before

and continue immediately after surgical interventions. Growth and
developmental issues are central in the total care, and immediate early
intervention before and after surgery, including ophthalmic surgery, is
important. Anticipated functional vision should be discussed with the family
before surgery, especially whether surgery is likely to prevent further vision
loss, improve vision, or just save the eyeball. If the parents are given false
hopes of the infant/child becoming sighted after surgery alone and do not
understand the importance of visual (re)habilitation (e.g., contact lenses,
spectacles, low-vision aids), they may neglect to provide for their child’s
functional needs. Education about the special care of the severely visually
impaired infant/child must be provided. Disappointment after surgery or a
visit to the hospital may prevent the parents’ motivation and cooperation for
months.
The number of infants with brain damage and other conditions with
several functional losses is increasing in all Western countries and the
developing world. This makes the diagnostic investigations long and
exhausting for the parents who “learn every day—often from a new person—
that their infant has another abnormal structure or function.” Ideally, all
information should be communicated to the parents by one person who
knows the complexity of the situation and has the authority to arrange an
early intervention professional to have time for the family in crisis. The stress
that the infant undergoes during the sometimes painful examinations and
treatments may be detrimental. To counteract the stress, parents should be
able to hold the infant close in quiet and comfort and offered a protected
place in which breast-feeding is not disrupted. Many hospitals have good
routine care for the delivery of “bad news.” The advice on all the aspects of
care of the infant and the family during the diagnostic period should be
collected into a manual for training of personnel. This would lead to a better
standard of care of patients and parents everywhere.
The phrase developmental emergency was first used by Patricia Sonksen
et al. (8) to describe the need of immediate care of an infant and the family, if
diagnosis of impaired vision is made during the first year of life. The
emotional stress may lead to depression in the parents; therefore, the families
of disabled children should be considered as patients during and after
diagnosis and treatment and be supported in the demanding care of their
infants.
EARLY INTERVENTION AT HOME

AND AT SCHOOL
Early intervention services at the local level vary from adequate to none. The
infant’s pediatrician may have never before created a local team for early
intervention that provides for the needs of the infant/child, parents, and
siblings. Such a team is needed. Often, contact with other families with a
similar experience, peer support, helps to decrease anxiety related to the
child’s future. To promote this, contact with the local or national
organizations of the blind may be suggested so that families can learn about
other families with visually impaired infants and children; organizations are
often available online.
There is such a great variation in the organization of early intervention,
rehabilitation, and special education of children with impaired vision in
different countries and even in the different states in countries like the United
States and Germany, that generalizations are not possible. In countries and
states where awareness of the importance of vision in early development and
the role that impaired vision plays in learning prevails, it has led to added
information on vision in the training of therapists and teachers, early referral
by pediatric neurologists to ophthalmologists, and even ophthalmologists in
the pediatric early intervention teams of hospitals. Increasing teaching about
vision and visual functioning in all pediatric medical training programs and
training of teachers, optometrists, and psychologists have been effective ways
of improving the quality of services.
Passive “vision stimulation” is used in many local services. It should be
used only initially when working with an infant who has so little vision that
he or she is unaware of its presence. Even in that early phase, activities
should be multimodal and include motor functions of the hands so that the
visual experience is more concrete and supports learning (Figure 13-8).
Videos like “Leo Learns by Doing” (9) (Good-Lite), which depicts family
activities that led to normal motor and cognitive development of Leo, an
infant with septo-optic dysplasia and only light perception, can be
instrumental in training early intervention workers and families alike in all
cultures and should be easily available to all young parents.
Figure 13-8 Training the infant’s therapist to include

activation of vision in Bobath therapy. Related videos can
be found at LEA-test.fi under “Videos of the book” tab.
(Reprinted with permission from Hyvärinen L, Jacob N.
What and how does this child see? Assessment of visual
functioning for development and learning, 2nd ed.
Helsinki: VISTEST Ltd., 2013.)
Early intervention should consider all developmental areas in each case and
find an answer to the basic question: “How much and what kind of vision is
present for different functions and for the development of each particular
function?” “For which functions should the infant/child be taught to learn
strategies of exploration typical to blind children?”
Training of vision-related brain functions can start during the first
assessment, which also serves as an introduction for the local therapist into
work with visually impaired infants. The local therapist has usually met the
infant at home before the first assessment of visual functioning (Figure 13-8)
and introduced himself or herself to start the assessment. If an infant has been
in hospital for a long time and experienced that a “new voice” means “a new
type of pain,” using the local therapist in the beginning of the assessment
makes the situation easier for the infant. If the infant is relaxed and expects a
pleasant play situation, several functions may be assessed at their best level.
The test situation is also a smooth introduction of a child’s vision into the
therapist’s usual neurodevelopmental therapy.
Visual field can be measured using an illuminated ball, that is, a penlight
illuminating a small plastic ball with uneven surface (Figure 13-8) (e.g., a toy
for kittens with a bell in it). If the infant responds only in the midline, the use
of the visual field can be activated by helping the infant to see his or her
hands as shadows against the illuminated ball. Information about the form
and surface of the ball combined with the movements of the hand and arm
may help the infant to watch his or her hand for the first time. The better
functioning hand is used first; then, the hemiplegic arm and hand are
supported to a position where the infant can see the fingers against the light.
Vision loss affects nearly all areas of development. The most important
areas of functioning in which vision plays a central role are
Communication and interaction
Motor functions and balance
Body awareness, visual spatial concepts, and orientation in space
Auditory spatial concepts, typically built within the normal visual space
Object permanence
Language
Incidental learning
Social skills
Many of the functional areas mentioned above are included in the usual
therapies for infants. Development of visual functions and their use in all
areas of functioning can and should become an integral part of therapies and
enrich the therapist’s work. Therapists need vision for communication with
the infant and are highly motivated to learn about the infant’s visual
functioning in detail. They also have the opportunity to observe the
development of the infant two to three times a week and can teach the parents
to use the training activities several times each day.
Communication and interaction are the first activities on the list, because
they are the most important functions for the emotional development of the
infant and parents and the bonding between them. Tactile exploration of
parents’ faces is started early, especially if there is suspicion that hearing
might also be affected.
Tadoma, the technique of speech reading by feeling the mouth, facial
muscles, and the vibration of the vocal cords with an adult’s hand, is
modified for the little hands to feel the movements of the mouth and the
vibration of the vocal cords in turn (Figure 13-9).
Figure 13-9 Training of Baby Tadoma. A. Hearing was

impaired in this boy with microphthalmia and colobomas
of both eyes. Baby’s hand is guided to adult’s mouth while
she speaks. B. In Bobath therapy, he was guided to explore
his body, which supported the development of normal
spatial concepts and motor functions. C. The more
microphthalmic and less functioning myopic eye was
trained during therapy, and when parents read books to the
boy, built with blocks, or later explored miniature animals
and houses, his visual acuity in the poorer right eye
remained low but would be of some help if something
happened to the left eye. (Reprinted with permission from
If visual communication skills of visually impaired young children are not

carefully assessed, we may be unaware of a child’s poor motion perception.
Due to poor motion perception, the lips appear blurred. During exploration of
the face in front of a magnifying mirror, a well-functioning 6-year-old child
asked, “Does the voice come through the mouth?” and when answered “Yes,
why do you ask?” the answer was “I thought that it comes through the ears.”
In this case, nobody had been aware of the girl’s poor perception of the fast
lip movements. She experienced lip movements since her hand happened to
be close to the lips when she said something. We use lip reading without
thinking about the complex visual information it requires. Lip reading should
be carefully assessed in all infants and children, especially if hearing loss is
present.
Exploring the parents’, siblings’, and caregivers’ faces makes these
important persons concrete and real. Father’s face is especially interesting if
explored before he shaves in the morning, next when one-half of the face has
been shaved, and a third time when the shaving is completed. This experience
gives the infant much to ponder until the father comes home in the afternoon,
again slightly different. Mother’s face should be explored as well, noticing
she doesn’t change like father.
Motor development and vision are closely related. When a normally
sighted infant sees his or her hand and brings it to the mouth, the early
asymmetrical tonic neck reflex is counteracted. If a visually impaired infant
does not see his or her hands, there is no incentive to explore them with the
mouth, and the early reflex pattern may remain. Therefore, playing with the
infants’ hands, guiding them to meet in the midline, and bringing them into
the mouth are important in play therapy.
Body awareness and the possibility of seeing and feeling one’s own body
parts facilitate development of spatial concepts.
Spatial concepts are difficult to create if the visual image of the
environment is unclear. These concepts can be trained early using playmats
with different structure at the two ends (Figure 13-10A). Small spaces are
important for the development of visually impaired infants and children. A
tiny prematurely born infant starts in a shoe box, moves to a boot box, and
then to larger and larger brown boxes (Figure 13-10B). Later, the boxes need
to have a sturdy frame; otherwise, the infant may push the box to lie on its
side. The size of the “little room” should be chosen so that the infant can
reach the walls without stretching his arms straight out because infants do not
extend like that.
Figure 13-10 Examples of easy modifications of

environment to support early development. A:Playmat
with clearly different ends helps in development of
concepts of directions. B:Brown box as a visual, auditory,
and tactile space. Toys and objects hanging on the broad
rubber band facilitate exploration and the development of
object permanence. C:Training listening skills is important
in infancy: the echoes of the infant’s voice from a metal
washing basin on the right and the red plastic wastebasket
on the left are different. Movements cause sounds that are
amplified by the resonance board. (Reprinted with
Ltd., 2013.)
Large “little rooms” are constructed for blind school-age children with
developmental delay. These “rooms” have a clear ceiling, whereas visually
impaired infants and children with useful vision should have a nontranslucent
ceiling and narrow slits up on the sides so that the space is a visual and
auditory space. The walls should have visuotactile areas as landmarks for
exploration. Toys and objects hanging from a thick rubber band do not
disappear when the infant lets them go, which supports the development of
object permanence.
A resonance board makes the infant aware of the movements of the feet
even if they are difficult to see (Figure 13-10C). In therapy, the infant’s
hands are helped to meet in the midline and, from there, to the mouth: this
supports development of motor functions. Hands are also guided to feel feet
and legs and together with them create a closed space around a big ball on the
stomach, a closed space created with the infant’s own body. Notice that this
infant with aniridia has dark glasses to prevent photophobia.
Carrying the baby in a sling gives experiences of movement patterns,
creates opportunities for incidental learning, and increases communication
between the adult and the infant. If the infant without motor problems is
helped to become an active explorer and learner, early development may not
notably differ from that of a sighted child. Exploration takes more time and
requires thinking when vision does not give its usual firm framework for
multisensory experiences. Numerous repetitions of movements are needed to
learn and remember the tactilely and kinesthetically studied structure of
objects, that is, movements of a door to learn how it sounds and mouthing of
objects to explore details with the tip of the tongue and lips. Mouthing and
repetitive movements can often be mistaken for autistic behavior in a visually
impaired infant/child. Autistic children have similar repetitive movements
and actions as visually impaired children. It is not known how often autistic
children are visually impaired and have skillfully learned to use techniques
that compensate for loss of visual information. We should carefully look for
the difference between exploration and meaningless repetitive movements.
CHALLENGING TEST SITUATIONS IN

KINDERGARTEN AND SCHOOL
Therapists and special education teachers are trained to pay attention to well-
supported postures and facilitation of movements. As a part of their work,
they can help in modifying the test situations for children so that these
children can be assessed more completely.
Figure 13-11A shows a boy without good support and proper near
correction unable to be tested. In Figure 13-11B, he reacts to the frame of the
glasses because of hypersensitivity of the facial skin. This should not be
interpreted as a sign that the child does not want to have glasses. In Figure
13-11C, the boy watches the figure-in-motion Pepi test (www.lea-test.fi) and
apparently perceived the dog because his eyes made a quick saccade to the
corner where the dog appeared and then followed the dog across the screen
several times. This figure-in-motion test thus functions also as an opportunity
to observe ocular motor functions.
Figure 13-11 A:This boy’s head fell to the side when he

tried to bring it to the midline. B:Placing spectacles on a
child’s face disturbs the child as long as the frame moves
on the skin of the hypersensitive face. C:With proper head
support and near-correction, the child could participate in
the tests. In this picture, we see that the eyes have
followed the figure-in-motion to the upper right corner.
Related videos can be found at LEA-test.fi under “Videos
of the book” tab. The Pepi test is available for download at
www.lea-test.fi/GAMES. (Reprinted with permission from
When we have to assess difficult-to-test children, it may be helpful to present

tests as games in the daycare or school. The assessment is carried out by the
person who has the best communication with the child (Figure 13-12A and
B). For example, the Panel 16 Color Vision Test was difficult at a school for
children with severe CP but became successful when the team understood
that two people should support the students with irregular arm and hand
movements. Well supported, the students could concentrate on the test
situation while the tester moved one color cap at a time along the row of caps
(Figure 13-12C). The children who could not speak responded with a nod
when the test cap was close to the cap with the next color shade. After the
third cap, this student became as quick as adult patients in choosing the next
color cap.
Figure 13-12 A:This child and her teacher used sign

language to measure visual acuity with single symbols.
The eye specialist placed the test cards on the test surface.
B:A student with a short concentration span was trained to
be tested for visual acuity by the local vision teacher and a
classroom assistant. C:When a student has poorly
controlled hand movements and no speech, nodding was
used as the response that the test cap corresponded to the
cap of nearly the same color. A lamp with high K-value
was used. Related videos can be found at LEA-test.fi
under “Videos of the book” tab. (Reprinted with
Ltd., 2013.)
If the testing is performed at daycare or at school, teachers, therapists, and

classroom assistants should learn to use the simple test games, repeat them,
and thus participate actively in the assessment and early intervention of the
child. The professional team learns to understand the test results and uses the
new knowledge in their work, makes good observations, and discusses the
observations with the family.
An infant with multiple afflictions or disabilities may have several areas
of development in which his impaired vision may cause challenges. For each
infant and child, it is useful to write a list of functions that need to be
specifically addressed during assessments, therapies, and planning of early
intervention. This list becomes the child’s first Profile of Visual Functioning
for transdisciplinary care and carries information between medical services
and early intervention and later between medical services and schools.
HIGH-RISK GROUPS OF INFANTS AND

CHILDREN WITH VISION LOSS
The great majority of visual impairments are congenital or occur soon after
birth, and more than half of the infants with impaired vision have other more
obvious impairments. The largest groups among visually impaired, multiply
disabled infants who can be diagnosed soon after birth are
Infants with birth trauma–related hypotonia that later may lead to

spasticity
Infants with diagnosis of deafness or hearing problems who may have
retinal degeneration
Infants with syndromes known to cause problems in vision; the most
common is Down syndrome (trisomy 21) because of combined
hypotonia and large refractive errors, cataract, and/or nystagmus
Infants with severe prematurity
These at-risk groups should be seen immediately if they have any symptoms
of communication problems at the usual screening ages of 6 to 8 weeks.
Large refractive errors should be corrected early (with undercorrection of
large myopic corrections to encourage focus at near). If an infant has
difficulty in early communication, accommodation should be measured and
near correction given for therapy and to facilitate communication. Infants
with birth trauma and insufficient convergence, especially if they are
hypotonic, nearly always show loss of accommodation; therefore, their needs
of near correction should be addressed and early communication supported.
Since children with CP often develop deformation of their spine, good
ergonomic postures are important. This is also true for visual tasks.
Spectacles should be specially designed to meet the needs related to poor
head posture, which makes the line of gaze pass unusually high up through
the lens when looking at a computer or a TV screen prior to the reading age.
Retinal degeneration in deaf infants may cause night blindness during the
first year but arises more commonly when the child is a few years old. If the
hearing impairment is moderate, assessment of vision may be forgotten until
school age. In populations with Nordic ancestry or of Acadian descent (see
Chapter 23), Usher syndrome type III with progressive hearing and vision
loss in school age is more common (see also Chapter 33). Early intervention
and contact with other families with hearing-impaired children of the same
age are important so that the children learn to live in the two cultures.
Cochlear implants make them semihearing, but there may be situations in
which they cannot use the implant and need to be able to communicate with
sign language. Some hospitals still claim that a child does not have dual
sensory impairment if visual acuity is better than 20/60. These children’s
visual problems are related to photophobia and slow adaptation into low
luminance levels, not to visual acuity. Since retinal sensory cells of animals
with retinitis pigmentosa–like retinal degeneration have shown damage when
exposed to short wavelength light, good filter lenses may delay the
development of retinal changes. Therefore, education as to use of hats and
spectacles that protect against damaging wavelengths is advised.
There is great variation in the progression of retinal changes in genetic
conditions, even within the same family; painting a bleak picture of future
complete blindness is not productive. The family should have opportunities to
meet with other children with retinal degenerations and dystrophies with or
without hearing problems and learn that most children with dual sensory loss,
even deafblindness, are healthy and active. Young children learn from the
older children with the same disease a realistic picture of their future and may
avoid the typical period of depression around the age of 9 years (10).
To care for those with vision impairment particularly in hearing impaired
or with multiple impairments, such as children with rubella or other
syndromes, requires careful observation and training in all vision-related
functions. In this group of children, even major changes in the midline (cleft
lip and palate, etc.) and microphthalmia with colobomas may not prevent the
child from developing into a student with exceptional abilities. Infants and
children with severe communication and developmental delay should have
appropriate glasses, particularly with large progressive near correction if
accommodation is insufficient. The behavior related to changes in color,
contrasts, and movement in the near environment should be observed
carefully so that a well-structured near space is created. Many children are
hypersensitive to touching (tactile defensiveness), which can be often
decreased using the child’s own hands. Covering the hands first with soft
material and progressively increasing the roughness of the material makes it
easier for the child to accept the desensitization.
In all pediatric rehabilitation, not only the infant or child but also the
whole family, including parents, siblings, grandparents, and even neighbors,
should be considered.
PROBLEMATIC BEHAVIORS AND
SITUATIONS
Visually impaired infants may start pressing on their eyes, which is called the
oculodigital reflex. The causes of the oculodigital reflex are not clear, but
self-stimulation seems to be more common when the infant is not occupied
by play or interaction. Eye poking causes pressure atrophy of fat tissue in the
orbit and of the bony rim of the orbit and can also cause keratoconus or
retinal detachment. Since a child usually cannot report a change in vision,
retinal detachment may be picked up so late that surgical results are poor.
Deep-set eyes are a cosmetic problem that affects the child later in life. Often,
the skin around the eye becomes darker. Gently redirecting the preverbal
child’s attention to other activities, or sometimes providing a barrier such as
well-fitting glasses, can decrease this behavior. The child should not be
spanked or scolded. Once children are old enough to understand that the
behavior is harmful, they will usually stop.
Depression is common among children with disabilities. Nearly every
child with a disability due to chronic illness or an impairment goes through a
subtle or obviously expressed depression around the age of 9 years (10). This
happens when children become aware that parents and medical experts are
unable to make them like their peers, and they must accept their impairment
and disability as a feature of their self-image. Since the depression period is
so common, it is wise to talk more about the good functions in vision than
what is lost. Arrangement for age-appropriate devices and training for new
strategies that benefit the child should be put in place. Contact with other
children with similar problems can help the child to accept his or her situation
and to develop compensatory strategies and become an advocate of his or her
own needs. Anxiety is also common in adolescents.
If an infant does not start to recognize family members by their faces at
the age of 7 to 10 months but does recognize them by their voices, the infant
may obtain such a poor quality of visual image that he or she cannot see the
differences in people’s facial features. In infants with neurologic issues, there
may also be a specific loss of the cortical function responsible for face
recognition, called prosopagnosia (www.faceblind.org). An ophthalmologist
and pediatric neurologist should investigate the child’s functioning without
delay. Some infants also have no recognition and interpretation of facial
expressions and, therefore, cannot understand the emotional content of visual
communication. These two deficits in cognitive visual functions are the
socially most important ones and should be observed during therapy and
discussed during each ophthalmologic and neurologic examination of infants
and toddlers. The degree of loss varies; for example, some children are aware
of the presence of eyes and mouth but do not see changes in facial
expressions due to lack of motion perception, whereas others if asked what
they see in the face of their teacher may answer “skin.” They know the facial
structures as tactile information, but visually the face is a featureless surface.
Faces should be explored using tactile information, magnifying mirrors, and
visuotactile pictures, so that the child has a good foundation for imagination.
COMMUNICATION IN EARLY
INTERVENTION FOR TODDLERS AND
PRESCHOOLERS
Vision plays a central role in the communication of a toddler. If we bring
together five or six toddlers for the first time, it takes them less than half an
hour to decide “who is the boss” and “who is the underdog” and what the
ranking order is between these two. There has been no discussion as there is
no speech possible yet, but body language and expressions make the status of
each child quite clear. When a visually impaired toddler is brought to a
daycare center, limitations in visual communication must be described to the
personnel, for example, poor contrast sensitivity and visual acuity, loss of
central visual field, or cognitive problems in recognition of facial features
and/or expressions. A teacher’s aide with good communication skills should
function as the intervener structuring the play situation of the visually
impaired child and explaining to the other children if the visually impaired
child does not respond as expected. Sometimes dark glasses must be
prescribed as a distinguishing feature for the child to help the other children
understand that the child does not see well.
Personnel in daycare centers should have information about the
disadvantages of extraneous noise when the auditory channel is the dominant
sense in that the child may need play time in a quiet corner. The limited
vision of the child is demonstrated to the teachers who present pictures and
objects to a group of children. A good way to do this is with “demonstration
glasses” made of clear kitchen plastic wrap folded over lensless eyeglass
frames as many times as needed to decrease the teacher’s visual acuity to the
level of that of the child when looking through the folded material.
Well-planned pediatric orientation and mobility (O&M) training is key to
integration into the local school and society. If an instructor is unavailable
locally, the child’s therapist should have support from an O&M instructor so
that the child is taught proper strategies in orientation and moving. In such
cases, training during summer camps or weekend gatherings can only
partially compensate for the lack of formal training in O&M, an important
part of special education.
SCHOOL AGE
School age brings a major change in the life of a visually impaired child and
his or her family. Many children go to a local school and are integrated into
the mainstream education programs.
In 1996, the Expanded Core Curriculum for Students with Visual
Impairments was first introduced in the United States (11). While the
common core includes subjects all students learn in schools, for example,
mathematics and science, the Expanded Core includes nine areas that are
considered essential for students with low vision and blindness to function in
schools, homes, and communities. They are compensatory skills (skills that
enable a student to access the general education core curriculum), sensory
efficiency skills (including, but not limited to, the use of low-vision and
multisensory approaches to learning), orientation and mobility skills, social
interaction skills, independent living skills, technology skills, career
education skills, recreation and leisure skills, and self-determination skills.
Ideally, each visually impaired child has an individual educational plan (IEP)
or an individual learning plan (ILP) that covers all requirements of special
learning techniques and identifies who is going to teach these skills. Each
disabled child has the right to study in the least restrictive environment.
Visually impaired children with severe learning disabilities are in special
schools or special classes in local schools in which knowledge of learning
strategies of visually impaired children often may be limited. Recent pilot
studies show that half of the children in such schools may not have glasses or
only have glasses correct for distance vision without correction for much
more important and relevant near vision tasks. If glasses get broken, new
glasses may not be prescribed in a timely manner. There is a need for an
international, well-planned survey on current provision services for visually
impaired children with intellectual disabilities. This survey should include all
children with brain damage–related vision loss, especially children with CP,
because their special educational and rehabilitation needs are not met in
schools in many countries.
Ophthalmologists’ reports together with other physicians’ reports are
important in informing the schools about common multidisabilities of
children with brain damage (12). When the child’s needs are well stated, it
usually is possible to arrange appropriate special education. Among children
with several disabilities, 40% to 80% have vision problems and need
treatment and rehabilitation. It is, therefore, important that these children
have a targeted clinical assessment using tests specifically developed for
each type of disorder.
CEREBRAL VISUAL IMPAIRMENT

Brain damage–related vision loss is often called cerebral or cortical visual
impairment (CVI) (13), although cerebellar functions may also be affected.
There are so many different combinations of possible losses of cortical and
subcortical functions that an overall description of early intervention and
planning of special education is not possible in this book concerning the
retina.
The low-vision team and the child’s neurologist assess all functions and
decide how to train each of them and which compensatory functions will be
important to develop. After a child’s anterior visual impairment has been
assessed, cortical functions are evaluated. The child’s visual functioning is
described covering (a) normal or low visual acuity with or without increased
crowding; (b) contrast sensitivity problems; (c) color vision defects; (d)
photophobia; (e) visual field loss; (f) motion perception changes; (g) ocular–
motor problems; (h) deviation from normal in perception of length and
orientation of lines; (i) difficulty in recognition of facial features; (j)
difficulty in recognition of facial expressions; (k) problems in depth
perception; (l) problems in form perception, either concrete objects or
pictures of concrete objects, abstract forms like numbers, and letters; (m)
problems in perception of surface qualities; (n) problems in picture
perception and comprehension/understanding; (o) difficulty in composing a
whole picture from its parts; (p) problems in spatial awareness and
orientation in space; (q) problems in eye–hand coordination (vision–hand
coordination); or (r) simultanagnosia. When the different losses of functions
are recorded for the report on impairment and disability, the findings are
assessed for their positive aspect to depict what kind of visual functions there
are for visual activities and learning in the four main functional areas,
communication and interaction, spatial orientation and moving, activities of
daily living, and sustained near vision tasks like reading and writing, and in
each specific functional area to support the IEP and the ILP.
The effect of posture on the use of vision should be carefully assessed by
therapists and teachers in all children who have motor problems. Limited
attentional and planning capacity often inhibits children’s performance in
tasks and activities that they would be otherwise good at if the task had been
planned considering the child’s capacity. In clinical situations, this can be
observed especially clearly during measurement of visual field. If Goldmann
perimetry is done as in the examination of typically developing children
asking the child to look at the central target while being ready to respond to a
stimulus somewhere in the peripheral visual field and immediately pressing
the “clicker,” the resulting visual field is smaller than if measured using
reflex saccades. Visual field measurements can be performed using the voice
of the tester as the fixation target, if the child has simultanagnosia, for
example, and bringing test objects in from the sides. Loss of control of motor
functions can occur if a child has a demanding visual task that prevents the
child from attending to a consciously performed motor function, like head
control. Loss of head control is seen so often that it is taken as a normal part
of functioning in children with CP. However, it is a sign of overload. Visual
and general ergonomics of children with limited motor functions require an
entire section devoted to it in recommendations for schools.
Neurophysiologic studies have clearly shown dramatic changes in the
development of the brain if vision did not have its normal role in functioning
and representation in the cortex (14–16). Training of amblyopic eyes has
been a standard treatment now for 40 years. If both eyes have subnormal
vision, training of visual and all vision-related brain functions in infants and
children is logical. By training therapists to include activation of the use of
vision in their work and in their training of parents, the need to increase the
number of therapists is less. Teachers for all children with special needs will
benefit from further training in brain damage–related vision loss and better
understanding of the many unusual behaviors in visually impaired infants and
children. By including vision care in early intervention, ophthalmology can
greatly facilitate the development of infants and children and support their
families.
I would like to end this chapter with two pictures and videos recorded
several years ago during assessment of vision of a severely hypotonic child
with CP (Related videos can be found at LEA-test.fi under “Videos of the
book” tab. [see also Reference 17]). His visual acuity was 1.25 (20/16) with
the LEA symbols line test at distance. He was not accustomed to using his
hyperopic correction (plus lenses) because, due to his poor head control, he
most often looked through the edges of the lenses or past them. He preferred
a 48-point text for reading and very large letters for communication. He had
great difficulties during the tests but did not give up and answered using a
stick taped onto his right wrist (Figure 13-13).
Figure 13-13 A:A very hypotonic student used a stick to

point to the Panel 16 caps and completed the test correctly.
Skillful facilitation by his father helped the boy in the
demanding task. The father was color deficient and thus
could not be a part of the correct result. Note the soft
material at the end of the stick to prevent scratches on the
color surfaces. B:The boy wrote his answers pointing at
large letters. Related videos can be found at LEA-test.fi
under “Videos of the book” tab. (Reprinted with
Ltd., 2013.)
At the end of the examination, I told the boy that he had performed well;
most of his results (in sensory tests) were good. He answered by pointing at
large letters with his stick that he had used during the examination and wrote
an old Finnish proverb:
“Don’t judge a dog by its fur.”
CONCLUSION
Early intervention and rehabilitation for infants and children with atypical
visual development is based on early detection. High-risk groups of infants
and children, such as with CP and other motor problems or Down syndrome,
often have weak accommodation and also large errors in refraction (as in
Down syndrome), which should be corrected early. Ideally, each infant and
child should be observed whether he or she needs help in early development,
so that they can develop knowing how to manage the weaker areas in their
knowledge and find support for further learning.
All infants with delays in visual milestones during the first year should
have their sensory and motor functions carefully assessed and should have
near correction in comfortable frames when needed. When the infant starts to
sit, bifocals with large flat-top reading adds are useful. Visual information in
the newborn is transferred by tectopulvinar pathway as low-contrast motion
information, so infants see facial expressions as fast-moving low-contrast
shadows and may copy expressions during the first week. There are tests to
assess vision for communication and recognition of movement at the age of 3
months, but observations on communication and interaction reveal the most
important features in the infant’s and child’s early development. Therefore
parents, therapists, and teachers should get detailed information on each area
of development and the visual tasks within it and be made aware of what to
observe during development.
Several inherited and acute diseases and accidents can damage the retina,
visual pathways, and brain, so assessment of visual functions and the
development of visual functioning are an integral part in the care of all
infants and children. Early intervention and supporting special education with
clinical examination and interdisciplinary exchange of information on a
child’s needs are important for rehabilitation and participation in school-age
activities. This helps children and their families learn to live with atypical
vision using many specific skills and devices that improve the child’s
functioning and further learning.
The unfortunate definition of low vision as “impaired vision” in the ICD-
9 misguided people to look at what the children did not have instead of what
they have. Choosing one detail, full contrast visual acuity, as the measuring
stick of quality was inadequate. In audiology, the whole frequency area is
tested; the same should be done also in vision, that is, contrast sensitivity,
motion perception, peripheral vision, and more, because vision is a complex
sense.
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9. “Leo learns by doing.” Video produced originally by Finnish Federation of Visually Impaired.
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17. http://www.lea-test.fi
14
Low Vision Management of Children
and Teens With Retinal Disorders
Mark E. Wilkinson and Khadija S. Shahid
Clinical low vision evaluations provided by optometrists or ophthalmologists

trained and experienced in low vision rehabilitation care should be
recommended for all children with a visual impairment, regardless of the
cause of their vision loss, their age, or the severity of their additional
disabilities. That said, even now, optometrists and ophthalmologists may
overlook clinical low vision care as an integral component in the treatment of
children that are visually impaired, if they do not provide these services
themselves.
An effective model for pediatric vision rehabilitation services for children
and students who are visually impaired requires a collaboration of clinical
information and findings from the eye care provider and the clinical low
vision rehabilitation practitioner, in conjunction with educational and
functional information and findings from the teacher of students who are
visually impaired (TVI), classroom teachers, a Certified Orientation and
Mobility Specialist (COMS), the child’s parents, and the children themselves,
as they mature.
Functional and educational outcomes should focus on the student’s
individual needs for school tasks as well as for community and vocational
tasks. Additionally, focus should address concerns about independent travel
abilities. Also, consideration needs to be given to the student’s personal goals
and desires, for example, reading leisure materials and other avocational
activities.
The American Optometric Association’s Clinical Practice Guidelines (1),
the National Eye Institute’s National Eye Health Education Program (2), the
American Academy of Ophthalmology’s Preferred Practice Pattern for Vision
Rehabilitation (3), and the American Academy of Optometry’s Position
Paper on Clinical Low Vision Evaluation and Treatment of Students with
Visual Impairments for Parents, Educators and Other Professionals (4)
recognize the importance of low vision rehabilitation in the treatment and
management of children with visual impairments. These organizations
acknowledge that low vision rehabilitation/habilitation is the main treatment
modality for permanent visual impairment and is the primary strategy to
prevent a visual impairment from becoming a disability or a handicap.
WHAT IS LOW VISION?

The National Eye Institute (5) defines low vision as a bilateral visual
impairment that is not correctable by standard glasses, contact lenses,
medicine, or surgery and that interferes with the person’s ability to perform
everyday activities. Similarly, under the Individuals with Disability in
Education Act (IDEA) Section 300.8(13), visual impairment means an
impairment in vision (after best correction with glasses or contact lenses) that
adversely affects a child’s educational performance (6).
Statistics
United States data estimates that 0.2% of the school-age population is
composed of children with low vision or blindness (7). Of these, only 10% to
15% are considered to be functionally or totally blind (8,9). Despite the low
prevalence of visual impairment in the school-age population, childhood
vision impairment is a significant public health problem as it affects children
across their lifespans.
Eye conditions that result in childhood visual impairment include
congenitally acquired conditions, such as albinism, achromatopsia, aniridia,
congenital glaucoma, congenital cataracts, Leber congenital amaurosis,
nystagmus, optic nerve hypoplasia, and retinopathy of prematurity, as well as
conditions that develop after birth such as cone–rod dystrophy, dominant
optic atrophy, retinitis pigmentosa, and Stargardt disease. Systemic
conditions that result in childhood visual impairment include genetic and
developmental syndromes, such as Usher syndrome and Bardet-Biedl
syndrome, as well as congenital or acquired neurologic conditions that result
in cortical visual impairment or cerebral visual impairment (CVI). The
additional disabilities associated with systemic conditions can impact how the
clinical low vision evaluation is performed and what treatment strategies are
to be employed by the low vision team.
Studies have found that 40% to 66% of children with visual impairments
have additional disabilities that can include cognitive limitations, speech and
language problems, hearing impairment and deafness, and motor and
orthopedic difficulties, such as cerebral palsy, seizures, and autism, among
many other conditions. Despite these multiple impairment challenges, 75% to
80% of this population have some useful level of vision (10,11).
CLINICAL LOW VISION

REHABILITATION EVALUATIONS
A clinical low vision rehabilitation evaluation should be recommended for all
children with low vision as an adjunct to the care they receive from their eye
care providers, both ophthalmologists and optometrists. Whereas the role of
the eye care provider is to maximize a child’s visual capabilities through all
available medical, surgical, and optical means, the role of the clinical low
vision rehabilitation practitioner is to maximize the child’s functional vision
capabilities. The term “functional vision” is used to describe what the child
with low vision is able to do visually with their remaining vision in the real
world, as opposed to in a clinical setting.
Comprehensive clinical low vision rehabilitation care should begin as
soon as a visual impairment has been identified, regardless of a child’s age or
presence of any additional physical or developmental disabilities. A specific
diagnosis along with prognosis is helpful, but lack of a diagnosis should not
delay the referral of a child to low vision clinic. Although infants and
toddlers who are visually impaired will not have low vision devices
prescribed for them, an assessment of their refractive status and current level
of visual functioning will help their educational team plan for early
intervention activities and future educational needs. Additionally, a clinical
low vision rehabilitation evaluation will provide the infant or toddler’s
parents with information concerning their child’s current visual functioning
and expected visual abilities as they mature.
The incorporation of an individualized rehabilitation/habilitation plan
into an Early Intervention Program can provide children with better access to
their visual environment, minimize the impact of visual impairment on
everyday activities, and smooth the transition to school. Ongoing clinical low
vision rehabilitation care throughout the preschool/K-12 educational program
is imperative, because visual needs change throughout their educational years
and into adulthood. Clinical low vision rehabilitation evaluations are
especially important at transition times, such as when entering primary and
secondary school programs or when students are preparing to enter college or
a vocational training program.
A clinical low vision rehabilitation evaluation is a specialized eye
examination that focuses on visual function as well as the prescription of
adaptive equipment to maximize the use of the child’s limited vision. An
optometrist or ophthalmologist trained and experienced in low vision
rehabilitation uses a variety of standard and modified testing techniques to
obtain information about the student’s visual condition including, but not
limited to, best-corrected distance and near visual acuity, central and
peripheral field of vision, focusing ability, range of eye movements, eye
alignment, depth perception, color vision, ability to see low-contrast objects,
and ability to function under different levels of illumination.
When indicated, the clinical low vision rehabilitation practitioner will
prescribe corrective lenses and adaptive devices that enable the student to use
residual vision more effectively and efficiently for learning, as well as for
everyday activities. A report that includes recommendations for optimum
print size and contrast of educational materials, lighting control and glare
reduction strategies, magnification needs (including adaptive devices and
technology), and viewing and mobility strategies to reduce the effects of
central blind spots or significant visual field loss (if present), as well as
contrast enhancement strategies for both reading and writing activities, is
provided. Recommendations are also made regarding field of vision and
acuity-related classroom seating and placement of educational materials to
enhance the use of residual field and reduce nystagmus (if present), as well as
other classroom environmental modifications to enhance the use of the
student’s available vision for learning. For students who experience
discomfort from indoor or outdoor glare, recommendations are made for
specialty lens tints and glare-reducing coatings for their spectacles to increase
visual comfort under a wide range of lighting conditions.
Clinical low vision rehabilitation evaluations should be ongoing and
integrated into the care provided for all children who are visually impaired.
Having available information about the visual abilities of children with visual
impairments is critically important for successful habilitation and
rehabilitation outcomes.
A clinical low vision rehabilitation evaluation can provide the following
information to the ophthalmic practitioner and educational team:
Baseline acuity measurements and general visual functioning level

Assessment of visual skills, that is, whether the extent of vision loss be a
major factor affecting other developmental areas
Assistance for parents and teachers to better understand the child’s
visual condition and visual functioning, that is, “how” he/she sees
Determination as to whether there is a refractive error and whether the
refractive error is significant enough to need corrective lenses
Information and assistance in determining the most appropriate
learning/literacy media
Assessment of low vision devices, technology equipment, or other
adaptations and accommodations that can enhance the student’s
functioning level in school and in their community
Assistance to the educational team members with trial and acquisition of
recommended devices, equipment, or strategies
Assessment of driving vision for acquiring an instructional permit or
driver’s license if appropriate
Provision of timely re-evaluation to address changes in vision and the
positive or negative effect on visual function status, as well as changes
in visual demand as student progress through their educational program
Low vision rehabilitation care must be comprehensive to ensure that children

who are visually impaired have the devices and techniques they need to assist
them in their educational, vocational, self-help, and recreational activities. It
must also be ongoing, since the child’s visual needs may change over time
because of changes in their visual system, the demands of their work or
school, and their personal goals for vision utilization.
The clinical low vision rehabilitation practitioner initiates and maintains
ongoing communication with other members of the low vision team. At the
conclusion of the clinical low vision rehabilitation evaluation, the team
members review the student’s capabilities to be sure that he or she can easily
and efficiently use the devices that have been recommended to accomplish
specified goals. The clinical low vision rehabilitation evaluation also results
in referrals to other services or resources, when appropriate.
Members of the Low Vision Rehabilitation Team

Providing care for children who are visually impaired often requires a
multidisciplinary team that can include the primary eye care practitioner; a
clinical low vision rehabilitation practitioner, either an optometrist or
ophthalmologist who is specifically trained and has experience in low vision
rehabilitation care; a teacher of students with visual impairments; a certified
orientation and mobility instructor; classroom teacher(s); a technology
consultant; vocational rehabilitation counselor; occupational therapist; as well
as physical and speech therapist, depending on the needs of the child.
The clinical low vision rehabilitation practitioner provides the clinical
low vision evaluation. This clinical low vision rehabilitation practitioner will
have had specialized training in low vision rehabilitation either during his or
her optometric or ophthalmologic training or in a fellowship program and
may have gained additional experience in low vision rehabilitation care in a
mentorship program. It is essential that the specialist has had experience
evaluating and caring for children who are visually impaired from a variety of
eye conditions.
The TVI addresses specific educational needs. The COMS reviews the
child’s functional abilities for independent travel, both indoors and outdoors
in varying lighting conditions. Through a COMS evaluation, common
mobility questions can be addressed including whether or not the child is able
to cross a street, walk to school, ride a bike, ride a moped, and/or drive a car.
A COMS will also assess mobility skills in lower lighting and unfamiliar
conditions, to be sure the student has the tools and skills needed for safe
mobility in all environments.
Both the TVI and COMS work together to perform a functional vision
assessment and provide a report to the clinical low vision rehabilitation
practitioner before the clinical evaluation. A functional vision assessment
reviews the use of vision at near and distance, visual field preference,
tracking and scanning abilities, visual attention, ability to reach or move
toward an object, responses to lighting and color, and perceptual abilities.
This information is helpful for the clinical low vision rehabilitation
practitioner to have prior to their assessment of the child.
Technology
Technologic advances to assist students with low vision include optical
character recognition and text-to-speech synthesizers, computer speech
synthesis, variable size fonts, and contrast enhancement devices. The
technology team and rehabilitation specialist will review with the educational
team those options that are appropriate for a given child. The expertise of a
technology consultant is invaluable in answering questions regarding access
to technology and use of technology to increase functioning.
When appropriate, the technology consultant reviews assistive
technology to improve current functioning. The technology consultant
considers low-tech to high-tech solutions, facilitates access to technology,
makes referrals for local follow-up, or provides follow-up in the student’s
home, school, and community. Review of technology options should be a
part of the ongoing process to determine what additional learning and literacy
media will best meet the needs of students with visual impairments. To
address questions, the technology consultant considers factors as outlined in
the Student, Environment, Tasks, and Tools (SETT) framework (12).
SETT:
Student information
The eye condition(s)
Low vision evaluation findings
Functional vision information
Consideration of other impairments Environments
Home
School
Vocational
Community tasks
The activities the student may require assistive technology for
Tools
What are required by the student to function as independently,
efficiently, and competitively as possible
As technology continues to advance, technology consultants are critically

helpful in getting the students the tools they need to be successful in both the
classroom and the community. These consultants may be teachers of the
visually impaired, members of school technology teams, or educational
specialists.
Technology advancements over the past decade have removed significant
barriers for individuals with vision loss, allowing them to engage in activities
that would have been impossible in the past. An added advantage is the
mainstream use of this technology by both individuals with and without
vision loss so as not to stigmatize users who are visually impaired. For
example, despite their small screens and keypads, several features built into
smartphones and tablets make them easily accessible to users who are blind
or visually impaired. Leading the industry are Apple products that provide
easy accessibility to users with vision loss through their VoiceOver and
Zoom programs.
VoiceOver is an audio screen reader that uses text-to-speech to speak out
loud what is printed on the screen, to confirm selections, respond to typed
letters and commands, and integrate keyboard shortcuts that make application
and web page navigation easier. The Zoom app magnifies everything
onscreen from 1.2 to 15 times its original size while maintaining their
original clarity. Additional accessibility options include the “Large Text”
option which allows the user to select a larger font size (20 to 56 point) for
any text appearing on their device, along with a reverse contrast option.
Many individuals with vision loss respond better to visual text that is
displayed as “White on Black.” This reversal of contrast is often the only
accommodation needed for an individual with a visual impairment to easily
read on their phone or tablet. Additionally, there are free or low-cost apps for
smartphones and tablets that allow the device to function like a handheld
video magnifier. The authors currently recommend the Super Vision+ app to
make a smartphone work like a video magnifier. Finally, there are free or
low-cost apps for smartphones and tablets that allow the device to function as
an optical character reader to read printed and handwritten text, identify
currency, and interpret bar codes. Our favorite app is the Seeing AI app.
These apps can be found in the app store for Apple and Android devices.
Note that these apps are constantly evolving and improving so students with
visual impairment should be encouraged to investigate both Apple and
Android (or their smartphone of choice) website for new accessibility
options.
The multidisciplinary team will work together to assess the strengths and
weakness of the student with respect to their educational, vocational, and
avocational pursuits. They will work with the student to help the student
function at their highest potential during their formal education years and
later as adults.
LITERACY
The development of literacy skills for children with visual impairments is one
of the most common functional concerns reported in a clinical low vision
evaluation (13–15). The term literacy is often used, but rarely defined in an
objective and meaningful way (16). Literacy, as defined by Webster’s New
World Dictionary, Third College Edition, is the “state or quality of being
literate, specifically the ability to read and write.” Koenig (16) stated that
“literacy is demonstrated by successful and meaningful application of reading
and writing skills to accomplish desired and required literacy tasks in all
environments.”
Throughout an individual’s life, literacy is demonstrated at different
levels that go beyond the basic stage. For the individual with a visual
impairment, strategies to increase independent and efficient literacy skills
must be determined through a combination of assessment of the individual’s
level of visual functioning, determination of literacy needs, and the ability to
use adaptations and/or devices for successful access (17–26).
For children with visual impairments, the development of functional
literacy skills will be critical to their success in adult life (13–15). Reading
and writing are essential elements in the majority of vocational and
educational activities, as well as for avocational and recreational activities
(14). Koenig and Holbrook state: “There are perhaps few decisions made on
behalf of students with visual impairments that are more crucial, yet subject
to more confusion and controversy, than the decision regarding an
appropriate reading medium. Determination of the appropriate reading
medium is not a concern for those who have no visual impairment (i.e., they
will learn to read visually, using standard-sized print). Nor is it a concern for
those who are totally blind (i.e., they will learn to read using tactile braille
and/or audio reading). Difficulties arise, however, in making decisions for
those students who are visually impaired, but not totally blind” (27).
Most students with visual impairments can read visually using one or
more adaptations. This can include a closer viewing distance (relative
distance magnification), the use of an optical or electronic magnification
device, higher-contrast materials, large print, or digital materials. These
strategies facilitate the effective use of residual vision in achieving literacy
(22,23). The challenge, and a long-standing dilemma, lies in selecting which
of the above adaptations is most appropriate for the student who is visually
impaired. For example, multiple authors have found that eye care
practitioners and teachers often recommend large print books without
collecting objective data or using any systematic process to support the need
for such materials (17,27–38). The authors have noted, and the literature has
shown, that judgments are made about the reading mode for individual
children without regard to the child’s particular needs. As noted by Koenig,
Layton, and Ross, “although most professionals and parents would confirm
the need to make individual decisions for individual students, this philosophy
does not appear to hold for the provision of large print books” (39).
It has been recognized for years that many children, with significantly
reduced distance acuity (20/200 or worse), read standard or even very small
print faster and more effectively than they do large print (28,37,38,40).
Unfortunately, confusion still exists regarding the most appropriate mode of
reading for these students, especially when distance visual acuity is all that is
known or reported about the visual function. Little relationship has been
found between distance acuity and the most efficient mode of reading
(40–43). Sykes stated, “distance visual acuity is unreliable as a guide to
reading ability, although higher distance acuities undoubtedly facilitate
reading speed” (38).
Historical Perspective
As far back as the 1920s, it was thought that if an individual with a visual
impairment used their remaining sight, they would further impair what little
vision remained (44). This archaic thinking led to the practice of blindfolding
all children who were visually impaired to instruct them in braille. In the
1940s, similar thoughts led to the recommendation of providing all children
who were visually impaired with large print to avoid “unnecessary stress on
the eyes” (28). It wasn’t until 1964 that Barraga demonstrated the importance
of using vision from a young age to develop the highest level of visual
functioning (29). And in the 1970s, Sloan and Habel further demonstrated
that the students’ reading speed and comprehension scores were equal when
comparing performance with standard print versus large print (37). It is now
known that reading speed is faster when students are given regular-size print,
with or without the use of vision enhancing devices, compared to reading
large print (39).
Yet large print had been advocated for decades and continues to be
recommended for students with visual impairments. The disadvantages of
this practice are numerous. For example, “large print” suggests bigger font
size, with no standard size indicated and often without considering the need
to further magnify special characters such as fractions, exponential numbers,
or diagram labels. This “one size fits all” approach is never appropriate for
individual rehabilitation treatment. Additionally, as mentioned above, reading
speeds are slower when using large print compared to regular print even if a
vision enhancing device is needed to read regular-size print. This is because
the total head sweep needed to read large print is time-consuming and tiring.
An additional problem with large print materials is their size and weight,
which makes them more difficult to transport between classrooms and from
classroom to home. Finally, it is important to note that once a student
graduates from high school, the availability of large print reading material is
significantly reduced in higher education settings, employment opportunities,
and leisure reading tasks.
Despite these disadvantages, the practice of large print continues to be
recommended by eye or other doctors, often without reasonable information
to validate this recommendation, and by general education teachers and
parents, simply because this has been standard practice for years prior. For
some schools, there is a perception of advocating for a student with visual
impairment by using these specialized large-print textbooks and materials.
And for some TVIs, the recommendation is made when they don’t have any
additional data (from the primary eye care provider) to support a more
appropriate choice.
Optical and/or electronic magnification devices provide students with
immediate access to printed material at their desks. Some devices also allow
access to educational information at extended distances, such as on traditional
or interactive white boards. These devices should be used in conjunction with
the best corrective lenses provided for distance viewing. Options such as
telescopes will improve detail vision at distances beyond arm’s length, such
as reading the board in school and street signs in the community. Electronic
magnification devices (aka closed-circuit television or CCTV) provide access
for both near and distance tasks in the classroom. These devices enhance the
student’s accessibility in the classroom and beyond, resulting in increased
visual independence.
Devices a Student With a Visual Impairment Might

Use for Literacy
Optical and electronic magnification devices often play an important role in
enabling the successful development and maintenance of literacy for students
with a visual impairment. While many devices are available without
prescription, performance with the device is optimal when the device is
individualized for the unique needs and characteristics of each student. When
prescribing these devices, an optometrist or ophthalmologist trained and
experienced in low vision rehabilitation considers the student’s goals,
characteristics of the student’s underlying disease or disorder, and specific
test results, including the student’s visual acuity (distance and near),
refractive error, accommodative amplitude, contrast sensitivity, binocularity,
visual field, working distance, and lighting needs. The prescribed device
incorporates the necessary corrective lenses and magnification power that
enables the student to read educational text.
In order to read with comfort and efficiency, multiple studies have shown
that individuals with vision impairment require greater magnification than is
required to just discern their print size goal (9–13). The difference between a
student’s smallest identifiable threshold print size and the print size that
provides the student with comfortable and efficient reading is known as the
reading reserve. Sufficient magnification is prescribed during a clinical low
vision evaluation to ensure an adequate reading reserve, resulting in faster
and more efficient reading. The amount of reading reserve required for best
performance varies from student to student, even when they have similar eye
conditions. It is therefore imperative that the device magnification power be
individually optimized for each student. The clinical low vision rehabilitation
practitioner uses objective data to make these recommendations.
Least Restrictive Educational Environment

The IDEA states that all students are entitled to a free, appropriate public
education in the least restrictive environment. Often, the use of low vision
devices, prescribed by an optometrist or ophthalmologist trained and
experienced in low vision rehabilitation, constitutes the least restrictive
approach to allowing access to printed materials (17).
Assessment of literacy needs is an ongoing process. It is important to
determine if the visual condition the student has is a stable condition or
progressive. If there is an expectation of visual deterioration in adolescence
or early adulthood, the low vision team must determine if instruction in a new
primary literacy medium is necessary before visual deterioration makes the
current medium ineffective.
Today, best professional practices and federal legislation (IDEA ‘97’
Title 34 CFR, Sec. 300.346 (a)(2)) specifies that educational decisions must
be made by an educational team according to the individual needs and
abilities of each student. These decisions must be based on information
obtained from systematic assessment procedures. The Principle of Least
Restrictive Materials, initially developed by Stratton and later explained by
Koenig, states that “materials should be adapted only to the extent necessary
for efficient learning. If regular materials can be used in conjunction with
environmental adaptations or low vision devices, such an approach is
preferable to using specialized materials” (45).
In the past, large type materials were offered as a solution to problems
associated with access to selected text. However, this may be considered a
more “restrictive” approach to the consumption of print because of
difficulties in obtaining books in large print and the lack of individualized
accommodations. Students who are served only with the provision of large-
type text are not afforded independent access to libraries, newspapers,
information on consumer products, and other printed materials that are not
available in large type. Additionally, their needs to access distance objects
and information, such as chalkboards or projection screens, are often
neglected.
Emphasis should be placed on an individual appraisal of each student
with a visual impairment’s abilities, to ascertain his or her functional use of
vision. An objective measure of reading skills for each individual is
necessary to prevent reliance on a subjective preference for a certain size of
print and to prevent a recommendation based on a single measure of visual
acuity, particularly if it is a measure of distance acuity only. The selection of
the appropriate print media for students with low vision can be accomplished
with the use of the Print Media Assessment Process (PMAP) developed by
Koenig and Holbrook (46). This allows the educational team to look at both
the level of visual functioning and the level of reading efficiency in regular
print, regular print with an optical device, and large print. The decision
regarding the most appropriate reading medium should be based on the
collective judgment of qualified professionals on the educational team
(47–49), including the parents and, as appropriate, the student (32). As the
educational team assembles objective assessment data, they will be able to
consider the profile of each student to determine the literacy medium or
media that most appropriately match the student’s characteristics and needs.
It is important to consider the benefits of using standard print materials,
when possible, for students who are visually impaired. He or she will be
prepared for any type of future employment where the use of standard print is
required, but the same cannot be said of the student trained only with large
print materials (17,50).
For those students who do not have any functional vision or those who
cannot use visual print efficiently, tactile print through braille will provide
access to literacy. Both braille and visual print should be considered equally
valuable as media for literacy skills, with selection of one as the primary
medium based on individual student needs. Auditory literacy is a third
medium for literacy. Corn and Koenig (45) stated that “aural reading (reading
by listening) may be a beneficial supplement to a braille or print literacy
program. However, because it does not allow a means to both read and write,
aural reading should not be considered as a primary literacy medium.
Auditory presentation of a text allows for factual presentation of concepts,
but omits grammatical structure, spelling, and traditional format. Therefore,
the use of auditory materials as a primary learning medium is recommended
only for students having multiple impairments, who cannot read in print or
braille.”
In some cases, it may be appropriate to mix different types of media for
individual cases. For example, it may be necessary to teach braille to
supplement reading and writing tasks for a print reader, or it may be
appropriate to teach print reading and writing for functional activities to
braille readers. Occasionally, large print will be used as a transitional tool
when a student’s primary learning media is being switched from print to
braille. Additional means of sending and receiving information (e.g.,
recorded materials, computers, telecommunication) should be considered for
all students regardless of their primary reading medium. The emphasis should
be placed on developing options for students for both immediate and future
use (27).
An essential consideration for the student with a visual impairment is the
availability of technology, which will increase the student’s options for
efficiently performing reading and writing tasks. The current and future range
of computer and related technology has the potential for increasing a
student’s level of independence by providing more immediate and efficient
access to information (27). The Individuals with Disabilities Education Act
states that the Individual Educational Plan must include assistive technology
needs.
Braille
Despite the body of work promoting the use of available vision for learning
and the use of assistive technology to enhance access to educational
materials, promotion of a braille-based approach to education for a majority
of students with low vision continues to persist among some groups. Section
614 (d)(3)(B)(iii) of IDEA states that “The Individualized Educational Plan
(IEP) Team shall, in the case of a child who is blind or visually impaired,
provide for instruction in braille and the use of braille unless the IEP Team
determines, after an evaluation of the child’s reading and writing skills,
needs, and appropriate reading and writing media (including an evaluation of
the child’s future needs for instruction in braille or the use of braille), that
instruction in braille or the use of braille is not appropriate for the child.”
In 2009 the National Federation for the Blind (NFB) published a report
on literacy titled “The Braille Literacy Crisis in America.” The report
declared that children who are visually impaired or blind requiring braille
instruction were unable to obtain timely and appropriate training. The report
stated that braille instruction should be incorporated into the educational plan
for the majority of children with vision loss. To support their agenda, the
NFB developed the National Reading Media Assessment (NRMA). The
NRMA does not allow the student to use any vision enhancing tools other
than corrective lenses and visors. The use of optical and/or electronic
magnification devices is prohibited during this assessment. Additionally, the
NRMA states that all assessment materials will remain flat on the desk or
table and not held at the habitual or preferred reading distance of the student.
The approach promoted by the NRMA is in direct contrast to IDEA and
Individuals with Disability Improvement Act (IDEIA) of 2004, which
requires that an assistive technology assessment be provided to determine
what tools are needed by the student to access the general education
curriculum. Additionally, the NRMA recommendation that testing materials
should not be brought closer to the eyes is inconsistent with how students
with visual impairments function. Additionally, IDEA allows for
accommodations on tests, which the NRMA does not allow. There continues
to be no published data on the NRMA that proves this assessment
methodology is a valid and/or reliable assessment tool.
Many braille proponents state that a student should learn braille
immediately, because they may need it in the future. The problem with this
approach is that it assumes the world is static and does not consider
technology advances, including gene or stem cell therapies that are
anticipated to be available in the near future for some patients. Additionally, a
student learning both braille and print reading will often only learn braille as
well as they can see it. Finally, trying to learn to read in both print and braille
can take time away from learning other educational areas. Experience
demonstrates that those that truly need to learn braille will learn it. A recent
study of 29 sighted adults naïve to tactile braille reading given a 9-month
course in braille found that the majority could read whole words at 6 words
per minute by the end of the course, demonstrating that learning braille as a
child is not a necessity (51). Novel technologies that combine electronics
with tactile braille reading are being developed (52).
VISION ENHANCEMENT OPTIONS

Magnification is the main treatment option for enhancing the visual
functioning of individuals with vision loss. There are four types of
magnification that individuals with visual impairments employ to enhance
their visual abilities:
1. Relative distance magnification—by holding the materials closer to the

eye, they appear bigger. Children with visual impairments do this
naturally secondary to their large accommodative amplitudes. As the
child matures and amplitudes decrease, they will require the
appropriately powered reading correction or bifocal for this to work
efficiently.
2. Angular magnification—occurs when using a low vision device, such as
a handheld magnifier or telescope.
3. Electronic magnification—is available in handheld or desk- or arm-
mounted electronic magnification devices, computer software, as well as
built-in accessibility options on smartphones and tablets. Electronic
magnification can make the image both larger and with greater contrast.
4. Relative size magnification—makes the object larger, such as with large
print materials. The problem with large print is that it is not readily
available in the myriad of materials that individuals with a visual
impairment need to read on a regular/daily basis (i.e., bank statements,
bills, most other general mail, work-related materials, etc.).
Optical Devices
Optical devices were first prescribed for school-aged children in the early
1960s, about the time when educators were starting to believe that vision
should be developed for efficiency, rather than conserved and saved (29).
Sykes (38) and Sloan and Habel (37) found that students who were visually
impaired performed as well using 10-point type with optical devices as they
did with 18-point type without optical devices.
The Division for the Visually Impaired of the Council for Exceptional
Children published a position paper that stated “any child who can benefit
from the use of a device should receive a clinical low vision rehabilitation
evaluation by an ophthalmologist or optometrist who is knowledgeable in the
prescription of such devices” (53). The position paper also noted that the
evaluation and the optical device(s) can often be obtained at an overall lower
cost than large type books. Therefore, when funds are redirected to clinical
evaluations and prescribed optical devices, the result can be a decreased
burden of cost to taxpayers and an increased accessibility to regular print
materials to the student with visual impairment (53,54).
Corn and Ryser (17) found that students who use optical devices gain
certain advantages for functioning in the sighted world. The use of optical
devices, for those who can benefit from them, should be viewed as the least
restrictive approach to gain access to regular print materials for near and
distance tasks. The use of prescribed telescopic devices gives the student
access to chalkboards, signs, and events in the distance. Corn and Ryser
stated (17) “optical devices should be considered individualized educational
tools that are just as important to a child with low vision as is a brace to a
child with a physical disability or a hearing aid to a child with a hearing
impairment.”
Evaluations by qualified low vision rehabilitation practitioners are critical
to the educational process because they yield a starting point for the
prescription of a low vision device or devices. After the initial prescription,
the TVI teaches the student to use the device and works alongside others on
the educational team to evaluate its usefulness in real-life settings. Proper
prescription and instruction must be a joint effort by clinical low vision
rehabilitation practitioners and educators. Both contribute unique information
to the decision-making process that should not be accomplished separately
(39).
For the student whose primary literacy medium is print, the use of a
handheld or full-sized video magnifier, or an optical device as a tool to
supplement the use of regular print, will likely allow access to all printed
medium needs in all types of settings.
Koenig and Holbrook (27) proposed “the need to fill a student’s ‘toolbox’
with all the ‘tools’ necessary to accomplish the demands of specific tasks and
thereby demonstrate functional literacy.” The choice of tools will be specific
to the needs of each student and may include digital audio media; the
assistance of a sighted reader; optical and electronic magnification; radio
reading service; print (standard or large type); braille; and accessible
technology such as word processing and/or computer applications, speech
synthesis devices, large print terminal displays, and optical recognition
scanners. An individual at the functional level of literacy will determine the
requirements of a given task and then select the tools to accomplish the task
most appropriately and efficiently.
Because of the unique characteristics of students with visual impairments,
no generalizations about reading media are possible. For this reason, the
educational team must provide an individually administered and interpreted
learning media assessment to identify personal differences for individual
students.
The selection of the most efficient reading medium for a given student
must reflect the input from each member of the educational team. Decisions
should consider the individual sensory ability and capabilities of each
student, as well as their immediate and future needs. The need to provide
additional instruction in other reading media should be reviewed through
continuous evaluation as the student’s needs change or expand. When no
educationally relevant differences exist between print options, selection of the
option that is least restrictive is key. When the most efficient option is not the
least restrictive one, the team must build additional skills in the least
restrictive option. By objectively determining each student’s most efficient
print reading option, overall reading efficiency can be assured.
The importance of low vision rehabilitation practitioners and educators
working collaboratively to meet the individual needs of a child with a visual
impairment cannot be overemphasized. This is especially true when a change
in the primary literacy medium is warranted. For example, changing to large
print should not be provided without objective data to support that decision.
A clinical low vision assessment, provided by a low vision rehabilitation
practitioner, is essential to assist the educational team in making literacy
medium decisions.
Eye care practitioners should avoid unilateral decisions. They are only
one of the many professionals responsible for the child’s educational
development. Eye care practitioners must remember that the literacy needs of
children are different than the literacy needs of adults. Most adults who seek
low vision care (because of acquired vision loss) already know how to read
and write. For this reason, adults simply need assistance to use the same
reading and writing skills they developed in their youth. Children, on the
other hand, are just learning to read and write, which requires ongoing,
multidisciplinary services. When a collaborative relationship is developed
between eye care practitioners and educational specialists, timely and
appropriate educational decisions can be made for children with low vision.
PEDIATRIC VISUAL ACUITY TESTING

What is their acuity? To better understand a child’s visual abilities, it is
helpful to quantify their vision beyond, fix, and follow and/or central, steady,
and maintained.
There are a number of different types of visual acuity testing. The most
rudimentary is localization acuity. Localization acuity is the visual
awareness/perception of the location of an object or light. Examples include
light perception with projection as well as awareness of the location of
people/animals, etc.
Detection acuity, also known as minimum perceptible acuity, is the next
level of visual acuity. Detection acuity measures the simple
detection/awareness of objects, not their identification or naming. Detection
acuity can be quantified by having the child retrieve objects of different sizes.
Detection acuity can be estimated using the common object technique. To
use this technique, we use as reference the following sizes presented at 4-m
distance from the patient:
3 mm object = ~4/2 size equivalent (20/10)

30 mm object = ~4/20 size equivalent (20/100)60 mm object = ~4/40 size
equivalent (20/200)
Example: A child consistently finds a 30-mm ball from a distance of 1 m.

Functional VA = 1/20 = 20/400.
Resolution acuity is also known as minimum separable acuity. Resolution
acuity measures threshold at which an individual can discriminate the
separation between critical elements of a stimulus pattern. Examples include
the Landolt C, Tumbling E, or Broken Wheel test. Grating acuity testing is
another type of resolution test, also known as preferential looking. It can be
done with Teller Acuity Cards or LEA paddles. Optokinetic nystagmus
(OKN) is also an example of resolution acuity. It is important to know that to
achieve 20/20 acuity with a standard optokinetic drum, the test distance needs
to be about 9 m from the individual.
OKN drum stripes are 11 mm wide. In comparison, a 20/200 letter
presented at 4 m is approximately 60 mm in size, and a 20/20 letter presented
at 4 m is approximately 6 mm in size.
Resolution acuity is a more simple visual task than recognition acuity,
and some feel resolution acuity overestimates the individual’s visual abilities.
That said, resolution acuity does require the child to attend to the task, and it
allows for gross quantification of visual acuity. This is important for the
clinician to monitor the individual’s visual acuity over time and goes beyond
what can be assessed by measuring basic fixation and following abilities.
However, there are a number of sources of error with resolution acuity. Those
sources of error include poor fixation, strabismus, poor attention, lack of
interest, a preference for looking only to one side, and improper placement of
the individual during testing.
Recognition acuity is also known as Minimum Legible Acuity.
Recognition acuity is most often used clinically and is the highest level of
acuity measurement. Recognition acuity requires the child to understand the
task and requires a subjective response. Examples of recognition acuity charts
include LEA symbols as well as letter (Snellen) and number charts.
Charts Not to Use

Lined pictures (Allen cards) have been found difficult to standardize for
recognizability and identification (children may vary in their picture-naming
ability) (55–58). The International Council of Ophthalmology stated that
Allen cards should be used only as a last resort (57).
The International Council of Ophthalmology also stated that the tumbling
E charts require a sense of laterality that may not be developed in young
children or children with developmental delays (57).
The current consensus is that LEA symbols give the most reliable results
of all recently developed symbol charts, especially for young children
(55–58) (see also Chapter 13).
Distance Acuity Testing

Reliable testing of distance acuity will be at a closer test distance (1 to 2 m or
5 to 10 feet) until the child is school aged.
Near Acuity Testing

Near vision is functionally more important than distance vision for most
individuals, particularly younger students. The student being tested should
hold the reading card at their normal reading distance (with reading Rx on if
used).
Reports
When reporting a child’s visual status, it is important to include the following
elements in the report:
Diagnosis
Prognosis that realistically states the expected course of the visual
condition over time
Eye medications
Color vision deficiencies
Visual acuity: distance
Visual acuity: near with working distance for reading
Recommended reading print size if known
Visual field limitations
Photophobia that requires intervention when indoors as well as outdoors
Restrictions in activities including the need to restrict certain sports
activities, such as small ball sports and/or having an adapted physical
education program. If a student may participate in contact sports only if
Trivex or polycarbonate sports safety glasses are worn, this should be
stated. These lenses are impact and breakage resistant; Trivex may have
superior optics and impact resistance than the older polycarbonate
lenses.
Special contrast needs and/or lighting requirements
Is a spectacle correction needed for general use, distance only, or
reading only?
Optical and/or electronic magnification devices
recommended/prescribed
Next evaluation date
DRIVING WITH A VISUAL

IMPAIRMENT
By the mid-teen years, many individuals who are visually impaired will want
to know if they can visually qualify to drive. At the time of this writing, there
are no uniform qualification vision standards for driving. If a patient has
visual acuity between 20/50 and 20/200, with normal or near-normal visual
fields, he or she may be able to become licensed to drive at least on a limited
basis. Throughout the United States, there are different standards for
restricted driving privileges in a number of categories, including maximum
speed, time of day, and distance from home. In some states, licensure
requires the use of a spectacle-mounted telescope. In others, it simply
requires passing a behind-the-wheel driving test to demonstrate that the
person with a visual impairment has the ability to safely operate a motor
vehicle. With this in mind, it is important for the provider to know the
regulations for driving with a visual impairment in their state and to
continuously monitor and inform their patient of progressive visual acuity or
visual field loss that may disqualify them from driving.
All drivers, including those with a visual impairment, will benefit from
advances in car technology such as advanced driver assistance systems
(ADAS) that are increasingly available on cars at all price points, often with
no additional costs. These systems are designed to assist drivers by providing
information, alerts, and varying levels of control based on the environment
around the vehicle. Additionally, talking GPS technology readily available
both in cars and smartphones helps address concerns about the ability to read
street signs. Driver distraction principles support these audio options because
they allow the driver to maintain their eyes and attention on the road and
traffic around them.
Even if a student will not be able to visually qualify for driving at this
time, and/or is not interested in driving, they should be encouraged to take the
classroom portion of driver’s education at the appropriate age, so they will
know the rules of the road and understand the issues required for safe driving.
BARRIERS TO CARE
There are barriers to accessing low vision rehabilitation services for many
children and young adults with low vision. Those barriers included the cost of
low vision services, lack of availability of low vision practitioners,
geography, as well as the attitudes of parents and teachers about what can be
done to help students with visual impairments. Also, beliefs about what to
expect from students with visual impairments in addition to a lack of
knowledge concerning visual impairments in general can be factors that
impede widespread availability of low vision clinical information.
SUMMARY
Low vision rehabilitation is the only nonsurgical treatment modality for
vision loss. This is why it is essential that children with visual impairments
have access to ongoing clinical low vision rehabilitation care. Ongoing
clinical low vision rehabilitation evaluations, performed by optometrists or
ophthalmologists trained and experienced in low vision rehabilitation, are an
essential component in the educational planning for every child with a visual
impairment. The clinical low vision rehabilitation evaluation provides all
adults involved in the care, education, and habilitation/rehabilitation of
children with visual impairments with critical information about the nature
and severity of a child’s visual impairment and strategies for enhancing the
child’s use of remaining vision. These strategies include corrective lenses,
magnification, and other low vision adaptive devices, as well as services and
accommodations that would increase the child’s access to visual information
at school and during activities of daily living.
Students with visual impairments should have access to prescribed
optical and/or electronic adaptive devices, instruction in the use of prescribed
devices, and recommended habilitation/rehabilitation services throughout
their educational program. Information provided by ongoing clinical low
rehabilitation vision evaluations ensures that Early Intervention Programs for
young children and Individualized Educational Programs for school-age
children are truly individualized for the visual needs of children with visual
impairments and provides these children with the best opportunity for
successful growth and development.
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SECTION III
Imaging of the Infant and Child Eye
and Retina
15
Retinal Photography and Multi-
Wavelength Imaging in Infants and
Children
C. K. Patel and Andrew Blaikie
INTRODUCTION
The chapter considers the basic requirements of visualizing the retina to
record an image with emphasis on historical milestones in photography that
have led us to contemporary methods of retinal imaging. A classification of
devices used in pediatric ophthalmology and retina is presented based on the
wavelength of light used for image acquisition, image quality, ease of use,
and cost. How some of the devices are used will be described in detail. The
utility of these imaging techniques will be illustrated with case histories.
EVOLUTION OF RETINAL IMAGING

Retinal photography yields an en-face image of the retina. It requires the
capture of light reflected from the retina onto a sensor and has built on
historical achievements. Jackman and Webster were the first to use
achromatic light with a camera to publish a fundus photograph in an adult
although with a central illumination artifact (1). Dimmer’s effort resulted in
less artifact but required a very large device and long exposure time relative
to a patient’s blink reflex (1). Nordenson, working with Zeiss, was able to
commercialize a smaller device that yielded color images with a small field
of view (FOV) but was still limited by the long exposure time (1). The arrival
of flash reduced exposure to 1/25 seconds, which facilitated wider adoption
of fundus photography. Filtering achromatic light made it possible to
dynamically assess fluid flux across the choroidal and retinal circulation in
1961 with the advent of fluorescein angiography (2) and a decade later using
indocyanine green (3). The momentum with which these angiographic
techniques were adopted increased rapidly with the evolution of the charged
couple device in 1969, which resulted in the transition from film to digital
imaging. Digital video recording from binocular indirect ophthalmoscopes
(4) allowed still color images of the fundus to be captured in children (5,6). A
limitation of using achromatic light was scattering from media opacity, which
degraded image quality. The introduction of monochromatic light using
scanning lasers in scanning laser ophthalmoscopy (SLO) (7) reduced this
problem and paved the way for innovations, such as the ellipsoid mirror that
Doug Anderson developed for the first ultra-widefield imaging (U-WFI)
device known as the Optomap® (Optomap), commercially available in 2000
(8). The Optomap was intended for use in children but was commercialized
for adults until its utility in neonates and children was realized (9). Digital
imaging coupled to a handheld platform was a major breakthrough that
allowed the development of a camera specifically developed for pediatric
imaging in the supine infant or child. This technology was used in the
RetCam (Massie Laboratories, Dublin, CA), which was introduced in 1999
(10). Subsequent RetCam systems (RetCam 3 and RetCam Shuttle, Natus
Medical Inc., Pleasanton, CA) were developed and together, these cameras
have become the most popular WFI devices for retinal photography in infants
and young children. Throughout this chapter, these will be referred to as
RetCam, except where a distinction is required. Imaging the retina is possible
in premature and other infants, who may be awake, and in older children
under general anesthesia (11). Increasing computer power has allowed the
development of video recording from which the best still images can be
captured and has facilitated improvements in image quality through
averaging algorithms used by devices, such as Spectralis (Heidelberg
Engineering). Recent advances in the use of LED illumination resulted in
handheld systems such as the Neo (Forus Health, Bangalore, India), which
provides a small, low-cost portable imaging system for infant retinal imaging
(12). Noncontact cameras have also been developed for portable handheld
digital imaging albeit with smaller fields of view (13). A more recent
breakthrough has been the use of the mobile (smart) phone camera and
illumination/flash with external separate or attached lenses to enable low-cost
pediatric retinal photography (14).
TECHNOLOGY, ADVANTAGES, AND
LIMITATIONS OF IMAGING UNITS
The wavelength light used to obtain an image of the retina determines what is
seen by the user. True color images are obtained when achromatic light is
used for illumination and color film or a 3-CCD chip is used without filters as
the sensor. This is used in the majority of fundus cameras, including RetCam,
Zeiss tabletop, Pictor, and mobile phone imaging. A red filter illuminates the
retina with green light, which accentuates contrast and highlights normal and
abnormal retinal vessels, hemorrhages, and the nerve fiber layer.
Illumination with monochromatic laser yields grayscale images that
highlight different parts of the posterior segment depending on the absorption
characteristics of cellular interfaces. Illumination with green laser highlights
the vitreoretinal interface, whereas infrared laser penetrates pigment and
improves choroidal visibility in the presence of a heavily pigmented retinal
pigment epithelium (RPE). Combining these two wavelengths yields a
pseudocolor image, for example, in devices such as EasyScan® (EasyScan)
from Hoya (Tokyo, Japan) and the Optos (Dunfermline, Scotland) (Optos)®
platforms. The differential absorption of these wavelengths by variably
oxygenated hemoglobin can provide retinal oximetry images, which is being
tested as a useful biomarker of advanced retinopathy of prematurity (ROP)
(15). Fluorophores in RPE autofluorescence; capturing this fundus
autofluorescence (FAF) through specific illumination and filters is useful in
the diagnosis of some retinal disorders.
Retinal imaging devices can be classified into six main groups as
described in Table 15-1. These different groups are defined largely by the
nature of the hardware and consequently the optical strategy employed to
image the retina. The different optical approaches lead to a range of intrinsic
device characteristics that fall into three main categories: (a) the quality of the
image acquired; (b) the ease with which the device is used; and (c) the initial
purchase price and likely on-going costs of disposable items and
maintenance. These innate differences between devices make them more or
less suitable for different pathologies and resource settings. Sometimes, the
strength of a device for imaging one particular disease can be a weakness’
when imaging a different pathology or limit its use in a lower resource rural
setting where portability and robustness are key. For example, devices that
optimize FOV for ROP screening are typically hard to transport between sites
and may be limited in resolving power when highlighting fine detail of the
optic nerve or macula. Table 15-1 also attempts to describe the device
characteristics of the major groups and consequently their suitability for use
for different pathologies and in different settings.
TABLE 15-1 Classification of pediatric retinal

imaging devices
I, internal angle; V, video: Cost in U.S. dollars defined as high (>20 K), mid (2.5 to 20 K), low (0.3 to
2.5 K), and ultralow (<0.3 K).
The basic elements of image quality are resolution and FOV. There is no
consistency on how best to describe the resolution of imaging devices.
Manufacturers may fail to state objective measures of resolution and simply
use adjectives such as high. When objective statements are made, reference to
the specification of the sensor within the device is made implying that this
will reflect the real resolution of the final image. The specification of the
sensor, however, can be meaningless if the optical system projects an image
of low quality onto a high-resolution sensor. As with a HI-FI system, the
quality of sound generated by expensive big speakers is irrelevant if they are
being driven by a low-quality turntable and amplifier. Manufacturers could
consider stating resolution as the size of the smallest resolvable retinal detail
that can be reliably imaged, such as “10 microns.” Another measure could be
sensor pixels per micron of retina or per degree of field. These approaches
would offer more clinically relevant information that will relate to the
genuine quality of the image generated. Although instinctively a device that
generates images of high resolution would feel like a positive asset, not all
pathologies demand extreme granularity of detail for reliable interpretation
and consequently may be unnecessarily over specified. For instance, one may
not require the same level of fine lateral resolution to assess plus disease as
would be needed to discern diabetic microaneurysms.
The nomenclature around FOV in pediatric retinal imaging has been
reviewed to ensure that devices are comparable (15). Figure 15-1 shows that
three different angles referenced to the nodal point (E,I) and center of the eye
(C) describe the same area of retina being imaged. The central angle is largest
and internal angle smallest.
FIGURE 15-1 The area of the retina to be imaged is

shaded yellow. The three angles subtending this area are
from largest to smallest the central (C), external (E), and
internal (I) angles, respectively. The nodal point of the eye
is marked by the orange arrow.
For fundus photography, a standard FOV is an internal angle of 30 degrees.

The term widefield imaging was coined to describe images taken by RetCam
and ultra-widefield to describe images taken by the Optos platform. A
phantom model of a neonatal eye enabled Fung to determine that the FOV
horizontally was an internal angle of 80, 110, and 130 degrees for RetCam
Shuttle, Spectralis, and Optos 200Tx, respectively (16). Figure 15-2 shows
images of the phantom eye obtained with these systems. Although the Optos
200Tx has the largest FOV horizontally (130 degrees), the Spectralis has
better clarity of the superior and inferior parts of the retina.
FIGURE 15-2 The Phantom eye has rings separated by

10-degree intervals simulating the retina imaged by
RetCam-Shuttle 130-degree lens, Spectralis, Ultra-
widefield Imaging Module, and O-Tx with examples of an
angiogram in the same patient imaged with the latter two
devices.
Several devices have been developed to maximize the FOV recorded. This
technology was driven by the clinical wish to image a specific disease where
peripheral imaging is important such as in ROP, familial exudative
vitreoretinopathy (FEVR), and peripheral retinal degenerations that may
predispose to rhegmatogenous retinal detachment. Group 1 devices include
those devices, tabletop and handheld, that produce images with a FOV 80
degrees or greater. Traditional desktop retinal imaging cameras (group 2)
employ noncontact conventional optics generating a FOV in the range of 30
to 50 degrees that displays the macula and surrounding vessels in a single
unedited image. This FOV is similar to the noncontact handheld devices
(group 3), binocular indirect ophthalmoscopes with video (group 4), as well
as devices that exploit cameras on mobile phones when using a high powered
condensing lens (group 5). Optically, group 5 is akin to a monocular form of
binocular indirect ophthalmoscopy (BIO) but replacing the viewers’ two eyes
with the single camera lens of the phone. Retinal imaging devices that exploit
direct ophthalmoscopy techniques (group 6) are innately limited to a small
FOV of around 5 to 15 degrees depending on nature of the piggyback device,
the diameter of the pupil, and how close the camera is to the iris plane. To
overcome the limitation of a still image with a narrow FOV, assessing a
video sweep of the area of interest is another strategy. Alternatively, software
can automatically stitch frames together either from a number of single
images of different regions of the fundus or from video sweep, although
artifacts can be introduced when a stitched montage is created. The apparent
FOV of innately narrow field devices can then be artificially expanded to
generate a wider FOV.
There are benefits and drawbacks to ease of use for the different camera
systems. Some of the U-WFI cameras (group 1) require considerable
cooperation to reliably acquire an image in a short period of time without
unduly upsetting the child. All devices that require contact with the cornea
require anesthesia and restriction of movement. A minimum requirement is
topical anesthesia combined with swaddling to restrict movement of the baby.
In older children, deep sedation or general anesthesia is needed. Other
devices are restricted to imaging patients upright, requiring the placing of the
child’s head in a cage-like rest (group 2) or requiring the child to peer into a
black hole. This is generally impractical for many preschool children but can
be worthwhile in selected cases. Light handheld noncontact devices can allow
the user to quickly maneuver into position to accommodate a wiggly child,
but motion artifact has the potential to compromise image quality. In this
situation, acquisition of high-resolution video rather than intermittent single
frames offers advantages. Review of the video after the examination allows
editing of the best segments or selection of the most informative single
frames for a second opinion or analysis. Several new devices offer this useful
function, thereby increasing the opportunity for acquiring clinically useful
images.
The retinal cameras vary in weight, portability, and cost. Devices
designed for U-WFI are innately large and heavy making them impractical to
transport swiftly and reliably between sites, and the cost may limit purchase
of a system for each site. The ideal retinal camera for a rural primary eye care
setting in a low-income country would be robust yet light, taking up little
volume and allowing it to be quickly packed up and safely moved between
clinics. There are now a number of handheld devices that meet these criteria,
although they are still relatively expensive with most devices falling into the
mid cost range. The most compact devices are also those that cost the least by
exploiting the cameras of mobile phones. They are, however, dependent on
the optics, software, and sensors of the camera within the phone.
Consequently, the quality of image is variable depending on the make and
model of phone with little published information on the FOV and resolution.
A recent device on the market that is an outlier is the epiCam made by
Epipole Limited (Rosyth, Scotland). This device exploits the power supply
and computing hardware of a mobile phone but uses its own software and
optics. It acquires high resolution at 15 frames per second video in gray scale
and falls into the low-cost range (https://www.epipole.com/epicam-m/ ).
The additional hardware needed to acquire retinal images in this way
varies in cost from 10 to 1,000 British pounds (>$1,200 American dollars)
with the additional cost of acquiring a suitable mobile phone camera.
Thus, there is no overall ideal retinal imaging device; the reality is that
there are cameras for specific settings and specific pathologies with a
constant tradeoff between image quality, portability, and cost. What works
well for a child with ROP under a general anesthetic in a tertiary referral
center in a high-income country may not be the device of choice for a child in
a primary care clinic in rural sub-Saharan Africa with cerebral malaria.
Moore law suggests that technology improves significantly each year with a
paradoxical reduction in cost. It can be hoped that retinal imaging will also
benefit from this trend with higher quality, easier to use, and lower cost
devices, further democratizing access to relevant retinal imaging.
CONSIDERATION FOR IMAGING IN
INFANTS AND CHILDREN
As a general rule, the advantages of innovation in imaging of the retina in
adults have been useful in imaging the retina of children 5 years of age or
more as they are likely to cooperate with the need to maintain steady fixation.
In less cooperative children including infants, the options include cameras
that require contact with the cornea or are noncontact systems. Contact
cameras are useful both at the bedside and in the operating room, as
exemplified by the RetCam. One of the authors (CKP) has direct experience
using the portable version of RetCam-II known as RetCam shuttle. RetCam-
III is the latest iteration and retains a 3-chip charge-coupled device (CCD)
camera yielding a 24-bit color image with the main modifications being a
lighter handheld “gun,” improved review software and extended duration of
video capture with options of modules that allow for fluorescein angiography.
Figure 15-3 shows a RetCam-III being used in the operating room following
the insertion of a lid speculum. The D1300 focusing lens is the most popular
for ROP and attaches to the handheld “gun,” which is placed on a baby’s eye
through a coupling gel. Achromatic light from a ring-based illumination is
focused through the entrance pupil. A foot-pedal attachment is used to
modify illumination and focus and then a foot switch is actuated to capture an
image. The gun has to be rotated about the visual axis to capture images of
the peripheral retina (Figure 15-3A). For assessment of ROP, the standard
examinations consist of either three images (central [C], nasal [N], temporal
[T]) or five captured images if superior and inferior images are also added.
There is an option to record a video of the real-time examination from which
still images can be extracted. The images are then graded for ROP by medical
(11,17) or nonmedical personnel (18). There are now several alternatives to
RetCam-III including the Icon, 3nethra-neo, PanoCam from Phoenix
(Pleasanton, California), Forus (Bengaluru, India), and Visunex (Freemont,
California), respectively. The PanoCam has wireless capability for color
images and fluorescein angiography that is a potential advantage for
telemedicine paradigms.
FIGURE 15-3 RetCam-III being used in the operating
room under general anesthesia showing how it is held
centrally (C) over the eye (A-C, B-C) to obtain disc and
macular views with tilting required (A-T, A-N) for
temporal and nasal vies of the left eye in this patient. Note
that rotation of the device about the visual axis affects the
orientation of the fundal image with respect to the primary
position of the eye.
There are several advantages to the contact cameras. They are portable and
can, therefore, be used easily in different hospital settings including
ophthalmic outpatients, neonatal intensive care units, and the operating room.
Contact cameras yield true color images, which accurately convey critical
diagnostic features of retinal pathology and are also able to image anterior
segment structures. A narrow depth of field can yield inferences about three-
dimensional aspects of pathology as shown by the diagnosis of stage 4 ROP
(Figure 15-4).
FIGURE 15-4 Stage 4b ROP in which preoperatively it is

implied that the ridge is very high because when macular
vessels are focused in the RetCam image, the ridge is
blurred (A) and vice-e-versa (B). (yellow arrow)
Following lens sparing endoscopic vitrectomy, there is
resolution of exudates (green arrows A, before surgery,
and C, after surgery), and disparity in focus is less because
the traction released has allowed the ridge to adopt a more
posterior position (D). (yellow arrow) (Courtesy of Mr.
Chien Wong.)
The contact cameras also have limitations. Ring illumination yields variable
exposure across an image, such that the periphery is less clear and can impact
adversely on telemedicine grading required for ROP (19). Clarity is
compromised in darkly pigmented fundi and in the presence of small pupils
and media opacity. Ocular blood flow is impaired by excess globe pressure,
potentially compromising the diagnosis of plus disease in ROP (20). Retinal
hemorrhages have been ascribed to globe contact (21), and infection control
is of paramount importance when using these devices between patients.
The Optos device, although a tabletop device, also can be used for infant
and child imaging. Initial reports from Oxford (9) and Scotland (22) have
been followed by wider adoption of these devices for use in neonates from
other countries, including India (23) and the United States (24). Figure 15-5
shows the imaging technique in premature infants in the neonatal unit for
ROP screening. The process involves the use of an eyelid speculum, often
taped to the cheek following instillation of local anesthetic eye drops. The
flying baby position is used to securely hold a baby. The neck is extended
slightly as the baby’s face is brought close to the aperture of the Optos
200Tx. The lumbar area of the large more mobile baby requires support to
minimize body movement. Ocular alignment is optimized on the grayscale
monitor with small movement of the head while the photographer clicks a
handheld switch multiple times to capture a series of pseudocolor images.
The cornea is hydrated intermittently. The vestibulo-ocular reflex is used to
encourage ocular alignment if the eye rotates out of the primary position. The
index and ring finger of the hand supporting the chin can also be used to
support an endotracheal tube allowing ventilated babies to undergo imaging
with full medical supervision in the neonatal intensive care unit (15,25).
There is a virtual reality video demonstrating the technique (26).
FIGURE 15-5 Flying baby positioning for Optos ultra-
widefield imaging. The baby is lifted from the cot, held
horizontally while the assistant inserts taped lieberman
style speculums into anesthetized eyes (A). The cornea is
hydrated and the ventral torso of the baby is supported by
the forearm while the thumb and forefinger support the
chin and face (B). Once the baby is rotated, the
contralateral hand supports the occiput with the neck
slightly extended to allow approximation of the eye to the
camera (C, D). It is helpful if the hand supporting the chin
touches the camera to provide proprioceptive feedback
while the grayscale monitor is observed to facilitate ocular
alignment.
By rotating the infant’s eye from the primary position in the flying baby
technique, it is possible to improve the FOV of the superior and inferior
retina with the Optos 200Tx (27) as shown in Figure 15-6. There are
advantages to imaging infants and children with the noncontact ultra-
widefield Optos system. The confocal laser optics yield a more uniform focus
across the FOV. Image capture occurs rapidly over 0.25 seconds. Adequate
ocular alignment can usually be obtained with examination times of 10 to 30
seconds once the flying baby technique is mastered. The FOV is not limited
by pupil size, and the laser illumination less often compromises clarity when
media opacity, such as a corneal scar, is present (28). The narrow 0.3-mm
scanning beam of the Optos and a virtual focal point that is behind the plane
of the anterior lens capsule allow for visualization of the retina in the
presence of an axial cataract. Such a cataract can prevent experts from seeing
the retina using BIO (29). High quality of images with fundus fluorescein
angiography (FFA) is achievable in awake neonates, potentially reducing the
requirement for general anesthesia and reducing morbidity as well as cost
(30). FAF U-WFI uses green laser for excitation, which is less absorbed by
macular pigment and potentially yields better signal from the macula.
Noncontact systems avoid traumatic sequelae and infection risk associated
with contact-based systems.
FIGURE 15-6 The image on the right (B) was taken with
the baby’s left eye rotated approximately 90 degrees and
shows improved visibility of the superior and inferior
retina above and below the arrows, which shows
corresponding vascular landmarks relative to the figure on
left (A).
There are also limitations to the use of the Optos system for infant and child
imaging. There is an instinctive reluctance to consider the use of the flying
baby technique in medically vulnerable groups, such as premature infants.
Although we have demonstrated that it is feasible to image even the most
vulnerable babies, for example, those who are ventilated, there is the potential
for statistically significant morbidity (31) although it is debatable if this is
clinically relevant (32). The Optos 200Tx is not portable, which limits its
potential use across a hospital site in the way that RetCam shuttle could be
used. The Optos California device has a smaller footprint and is mounted on
a table that can be moved but lacks a mobile power source. The FOV varies
with the distance of the baby from the device at the time of image capture
(27). FAF U-WFI does not use image averaging, which can limit quality.
The artifacts produced during pediatric retinal imaging have been
systematically studied and reported (27). They were classified according to
descriptors illustrated in Figure 15-7.
FIGURE 15-7 Dust on the mirror results in dark spots

(white arrows), that are geometrically static while other
retinal features such as a vortex vein (*) move between
consecutively acquired images (A, B). Dust should
regularly be removed with the manufacturers approved
method to avoid this spot artifact. At the margins of the
images, artifacts from the eyelids, eyelashes, and lid
speculum can be seen. The fine red vertical lines in the left
image are from the lid speculum.
There was a significant association of vertical red/black line artifacts

associated with glint reflexes from metallic eyelid speculums. The detection
of a subtle retinal detachment can be difficult with Optos images, because the
depth of focus is large. The selective use of the red and green laser images
can be helpful in highlighting the presence of subretinal fluid, which reduces
visibility of the choroidal circulation (Figure 15-8). The noncontact U-WF
imaging module for the Spectralis (Heidelberg Engineering, Figure 15-9A)
may also be used in any child who cooperates with fixation. It comprises
dedicated software and lens, which attaches to the camera and is
interchangeable with the standard 30- and 55-degree lenses. Confocal SLO
technology produces a high contrast evenly illuminated grayscale infrared
image of the retina out into the far retinal periphery.
FIGURE 15-8 An autistic 7-year-old boy underwent a
scleral buckle and cryotherapy for retinal detachment with
a single temporal break. Postoperative images 2 weeks
later showed an adequate indent and cryopexy (A).
Inspection of the inferonasal retina showed residual
subretinal fluid (*B, C, D), which was not apparent in
fleeting views of the retina with BIO. The subretinal fluid
is accentuated in the red laser channel (*D) because it
masks the choroidal circulation.
FIGURE 15-9 An U-WF imaging module (A) attaches to
the Spectralis (B). The working distance over a baby’s eye
(C) is very short. Alternately, the Spectralis Flex system
(D) images at a greater distance from the infant eye with a
field of view comparable to the tabletop Spectralis (not U-
WF).
The U-WF imaging module can be used to obtain grayscale infrared, FFA
and ICG-A images, individually or simultaneously. Following proof of
concept that it could also acquire optical coherence tomography (OCT)
images when manually held over an infant’s eye (33), we customized an arm
to support it for use in the operating room to acquire OCT scans and
angiograms (34) (Figure 15-9D and E). Heidelberg engineering
subsequently developed a servo-assisted arm called the Flex to perform a
similar function for supine patients, such as infants in the operating room
(35). The platform is, therefore, able to deliver the full range of retinal
imaging to the pediatric patient, including infrared, multicolor, infrared, and
blue FAF imaging. Optical coherence tomography angiography has also
recently become available.
One clear advantage of the Spectralis platform is the integration of OCT
with retinal imaging that incorporates a tracker. This allows for accurate
localization of pathology and facilitates monitoring of diseases in clinical and
research domains. The images are less distorted compared with images from
Optos devices. Image averaging algorithms facilitate higher quality
autofluorescence imaging. Media opacity and pupil size affect image quality
less than systems that use achromatic light.
Limitations to the system are that the device usually requires general
anesthesia to obtain good quality images in infants. It is not portable. Dark
condensation artifact with FFA has been reported and attributed to the close
working distance of the ultra-widefield lens to the eye during anesthesia (36)
(Figure 15-9C). The artifact can be eliminated by generating air currents near
the eye. The macular autofluorescence with blue light is reduced because of
absorption by luteal pigment.
UTILITY OF PEDIATRIC RETINAL

IMAGING
Diagnosis
The diagnosis of retinal disease in pediatric retina has been revolutionized by
RetCam, the Optos, and Spectralis imaging systems, all of which are capable
of widefield retinal imaging and FFA. There are multiple indications for
using the platforms, and several of these are considered below.
Widefield imaging can be useful when there is a small pupil or media
opacities. The diagnostic capability of Optos devices has been reported for a
wide range of conditions referenced in recent review articles, including
chorioretinitis, inherited retinal degeneration, infiltrative disease, such as
leukemia, nevi, neoplasia (retinoblastoma), and retinal vascular diseases, such
as Coats disease, FEVR, ROP, and incontinentia pigmenti (9,15,37). Retinal
imaging alone may not display the extent of nonperfused retina; therefore,
ultra-widefield FFA is critical in FEVR and subclinical FEVR in
asymptomatic relatives of an index case (38).
The Optos devices have particular value in establishing the correct
diagnosis when there is a small pupil precluding adequate indirect
ophthalmoscopy. This is illustrated by a case in which B-scan
ultrasonography suggested a diagnosis of acute tractional retinal detachment
following laser treatment of type 2 ROP. The Optomap image established a
diagnosis of exudative retinal detachment with an absence of fibrosis, which
allowed the correct decision for medical treatment rather than surgery
(Figure 15-10). Similarly, in the presence of a 2-mm pupil that would not
dilate because of extensive posterior synechiae, imaging with the 55-degree
and U-WFI Spectralis lenses confirmed an absence of cystoid macular edema
but the presence of disc swelling in a child with juvenile chromic arthritis.
This information was useful for planning optimal timing of cataract surgery
(Figure 15-11).
FIGURE 15-10 Both eyes were treated with laser
photocoagulation for ROP. The left eye developed retinal
detachment 2 weeks later with a poorly dilating pupil,
which precluded indirect ophthalmoscopy. An ultrasound
scan (A) showed concave retinal detachment suggestion
traction. U-WF imaging confirmed regressed ROP in the
right eye (B) and absence of preretinal fibrotic tissue
causing traction in the left eye (C) confirming that
exudation and not traction was responsible for the retinal
detachment. (From Patel CK, Buckle M. Ultra-widefield
imaging for pediatric retinal disease. Asia Pac J
Ophthalmol (Phila) 2018;7(3):208–214. doi:
10.22608/APO.2018100.)
FIGURE 15-11 A female child with juvenile chronic

arthritis required cataract surgery. Spectralis imaging
through a miosed pupil showed optic nerve swelling (B,
C) absence of cystoid macular edema (A), which could not
otherwise be discerned.
The Optomap is also valuable for ruling out sight-threatening ROP when
media opacity such as anterior polar cataract precludes adequate binocular
indirect fundoscopy (Figure 15-12).
FIGURE 15-12 Fundus visualization by an expert was not

possible with indirect ophthalmoscopy or RetCam in this
preterm baby with anterior polar cataract as a component
of persistent fetal vasculature (A). Imaging with the Optos
confirmed that plus disease was absent (B) that there was
no ROP at the periphery of retinal vascularization that was
seen with FFA (white arrow—C).
Use of a contact camera focusing at different levels in the vitreous cavity can
be helpful in the diagnosis of acute tractional retinal detachment (Figure 15-
4), which is less well seen by Optos devices. The true color representation of
lesions afforded by contact-based cameras that use achromatic light is
important in the differential diagnosis of retinal neoplasia and forms of
uveitis and chorioretinitis.
Multimodal images that also incorporate OCT are very useful
diagnostically. For example, in the management of a child with vitreous
hemorrhage, infrared and OCT imaging of the contralateral eye can suggest a
diagnosis of congenital X-linked retinoschisis (Figure 15-13). FAF imaging
is key to the diagnosis of a retinal dystrophy, such as Stargardt disease, and
acquired problems, such as laser-induced maculopathy.
FIGURE 15-13 A male child presented with strabismus

and vitreous hemorrhage in the left eye as shown with a
RetCam shuttle image (B). Infrared/OCT visualization of
the contralateral fovea (A) offered congenital x-linked
retinoschisis as the most likely diagnosis.
Screening and TreatmentBasic fundus images, WFI, and U-WFI are useful in
the screening and documentation of many retinal conditions in infants and
older children to provide a useful reference as to the location, severity, and
extent of a vitreous or retinal finding, including vitreous or retinal
hemorrhage, choroidal nevus, coloboma, chorioretinal scar, vitelliform
lesion, or retinitis. The RetCam and allied cameras are well established for
the documentation of retinal hemorrhages in the diagnosis of abusive head
trauma in infants (details in Chapter 70). Optomap U-WF images are an
alternative option in the outpatient setting that can be used in conjunction
with a speculum using the flying baby technique when safe to the infant (39).
In heavier larger infants, when visual function is impaired bilaterally from
premacular hemorrhage, it is worth attempting imaging without a speculum
while supporting an infant held vertically to avoid distress. FFA can be
helpful in establishing that a suspected macular obscuring preretinal
hemorrhage is in fact intraretinal schitic hemorrhage with inner retinal vessels
visible on FFA (39). The Spectralis platform is able to show how FFA can
also be valuable in establishing the later complication of peripheral retinal
ischemia (Figure 15-14).
FIGURE 15-14 A pseudocolor Optomap image (A)

showing hemorrhages consistent with abusive head
trauma. Wide-field fundus fluorescein angiography shows
disruption of the macular circulation (B) and peripheral
non-perfusion that can occur and result in neovascular
glaucoma.
RetCam was developed for monitoring the response of retinoblastoma to

surgical treatment so that tumor recurrence could be identified early (40). The
Optos devices can be very useful when following children who have
undergone surgery for retinal detachment (Figure 15-8) and when nystagmus
prevents adequate conventional slit-lamp fundoscopy or BIO (15). The Optos
devices are better than RetCam in older infants, who have undergone surgical
treatment for ROP and require imaging during postoperative follow-up in the
clinic (Figure 15-15). The device has been incorporated into the pathway for
monitoring the response to endoscopic surgery for retinal detachment in the
United Kingdom. Serial FAF images can facilitate monitoring the progress of
inflammatory retinal disease, such as retinal necrosis, and inherited retinal
degenerations, such as Stargardt and Best disease (41–43).
FIGURE 15-15 A premature baby with a birthweight of
660 g and a gestational age of 24 weeks developed stage
4a ROP following laser ablation of the peripheral retina (A
—white arrow) requiring endoscopic lens sparing
vitrectomy. One week following surgery, a total retinal
detachment without obvious breaks was imaged with
RetCam shuttle (3 montages RetCam images (B)). There
was gradual absorption of a presumed effusive
postoperative retinal detachment, which was monitored
using the Optos 200Tx (C—blue arrows, D: partial and
total retinal reattachment, respectively).
Basic fundus images play a role in the management of retinal vascular disease
through documentation of macular exudation, vascular abnormalities, retinal
and vitreous hemorrhage, and location of laser treatment spots. As in adult
disease, retinal images may be examined to screen for or document status of
diabetic or sickle retinopathy. Screening of children with sickle cell is
suggested to commence at 9 years of age for SC genotype and 13 for SS
genotype. Retinal WFI and U-WFI may be useful in the screening and
documentation; however, the additional use of FFA with U-WFI, SD-OCT,
and OCTA have the potential to detect proliferative disease earlier in the
natural history before sight-threatening neovascular complications ensue
(44). Fluorescein angiography is well established as key for image-guided
treatment paradigms allowing for precise dosing of laser treatment in
ischemic and exudative retinopathies, such as ROP, Coats disease, and FEVR
(15,38). FAF can be helpful in the assessment after laser treatment, because
laser treatment sites appear as hypo-autofluorescent spots. Thus FAF images
can be used to locate sites for augmenting laser treatment such as in the
surgical management of optic disc pit maculopathy (Figure 15-16).
FIGURE 15-16 Disc pit maculopathy recurred following
primary treatment with barrier laser photocoagulation with
the OCT showing submacular and intraretinal fluid (A)
with the corresponding 55-degree Spectralis blue FAF
image (C) showing the old laser burns around the disc,
diffuse (black arrow) and punctate (yellow arrow) hyper-
autofluorescence secondary to bisretinoid accumulation.
Six months following vitrectomy, gas tamponade, and
endolaser, there was subtotal resolution of subretinal fluid
(B) with improved vision. The 30-degree blue FAF image
(D) show a missing area of laser (blue arrow) that was
augmented with outpatient slit lamp delivered laser
photocoagulation as the 15-year-old teenager was
cooperative.
The reference standard for case detection in ROP is BIO performed by a

consultant. RetCam has been used for this purpose and shown to be feasible
in numerous pilot studies, but a recent review has failed to endorse it as a
replacement for BIO by a consultant (45). U-WF imaging with the Optos
200Tx was shown as a feasible option in a pilot study of 48 babies (27). A
fundamental obstacle to adopting a comprehensive image-only–based
paradigm for screening in ROP is the inability to determine vascularization
into zone 3, which is one key endpoint in determining when to terminate
screening. The junction of zone 2 and zone 3 is approximately 2 disc
diameters internal to the edge of the image for RetCam when ROP is imaged
temporally (46). If ROP is absent, this approach only works well if it is
possible to visualize where the retinal vessels terminate. Image quality is
often poorest in the periphery with RetCam-III, which makes this unreliable.
Indentation-assisted imaging of babies using oral FFA is able to visualize the
ora serrata nasally using Optomap U-WFI (15). The reliability of this
approach for screening remains to be determined. The topic of ROP
screening and telemedicine are addressed in greater detail in Chapters 20 and
52. There is questionable merit in using RetCam in universal screening for
retinal disease in newborns (47). A similar approach could be pursued using
U-WFI imaging.
RESEARCH
Morphometry, the quantitative analysis of size and shape, is one of the pillars
of scientific endeavor and essential in research and clinical care of retinal
disease. Retinal imaging involves translating a three-dimensional curved
surface into a two-dimensional representation. This process is constrained by
the optics of devices and results in distortion of images, such as exaggeration
of the size of peripheral versus central features. For quantitative analysis, a
variety of grids have been developed to overlay across images for grading
features, such as nonperfusion in diabetic retinopathy (48). Software
modifications have attempted to reduce distortion to facilitate accurate
morphometry, but distortion especially in the far periphery remains a problem
that requires consideration and potentially adjustment during image analysis
(49). This peripheral distortion is pertinent to grading of ROP with WFI and
U-WFI, since the International Classification of Retinopathy of Prematurity
(ICROP) zones are circles centered around the optic nerve. For example,
because of distortion of retinal images, it is not known if it is accurate to
assume that the distance between disc to fovea is the same as the
corresponding distance nasal to the disc, when overlaying circles on images.
Artificial intelligence is now a well-established modality facilitating
pattern recognition. Large datasets of retinal images can be processed by deep
learning algorithms to assist with diagnosis. In pediatric retina, screening for
ROP is a large public health issue, which is focused on the need to make a
diagnosis of plus disease. The current gold standard of indirect
ophthalmoscopy and image-based screening has not yet delivered a
universally accepted effective system. Deep learning may deliver this in the
near future (50) and is addressed in greater detail in Chapter 19.
Additional research measurements from imaging will play a future role in
pediatric disease. One example is the differential reflectance of
oxyhemoglobin and deoxyhemoglobin with the dual-laser wavelength
imaging platform of the Optomap. This technique allows the measurement of
oxygen tension in the retina and has been demonstrated in healthy neonates
(51). Changes in these measures in the retina can be a proxy for disease in the
central nervous system. Thus there is potential for retinal imaging in children
to identify biomarkers of pediatric degenerative brain disease much in the
same way as is being explored for adult disease (52) or for retinal oximetry to
be applied in ROP (15). With the development of handheld oximeters, retinal
oxygen metabolism in pediatric retinal vascular disease should provide new
insights that could prove useful in better understanding disease
pathophysiology and potentially clinically (53).
SUMMARY
The pediatric retina can be interrogated by conventional white light, filtered
light, or with single wavelengths of light, which yield color, pseudocolor,
grayscale, angiographic, and autofluorescence images. Advances in
information technology and physics have delivered platforms that allow us to
overcome the barrier of ocular fixation that has in the past prevented imaging
the retina of infants and children. These advances facilitate clinical care of
children by assisting with diagnosis, follow-up, screening, treatment, and
research to improve outcomes for a population that is the future of every
society.
ACKNOWLEDGMENTS
1. Photographers at Oxford Eye Hospital: Lewis Smith and Jon Brett
2. Peter Charbel Issa for reviewing manuscript
3. Caroline Justice: ROP nurse specialist
4. All trainees that transit through Pediatric retinal service at Oxford
5. Optos, Heidelberg Engineering, and Epipole for their support in
developing instrumentation for pediatric imaging
6. Our families for their support
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16
Optical Coherence Tomography in
Lejla Vajzovic, Anand Vinekar, and Cynthia A. Toth
High-resolution spectral-domain optical coherence tomography (SDOCT) is a

signal acquisition technique that provides detailed images of light scattering
tissues with reported resolution of <5 μm. It has enabled visualization and
analysis of retinal anatomy and clinical evaluation of retinal pathology in the
adult and pediatric retina (1–3).
RETINAL LAYERS ON OCT

SDOCT shows alternating bands of hyper- and hyporeflectivity (Figure 16-
1) that correlate with histologic bands and are labeled from inner to outer:
nerve fiber layer (NFL); ganglion cell layer (GCL); inner plexiform layer
(IPL); inner nuclear layer (INL); outer plexiform layer/photoreceptor synapse
layer (OPL/PSL) (4); outer nuclear layer, which includes Henle fiber layer in
the macula (ONL+HFL) (4); external limiting membrane (ELM), inner
segment band (IS), which is also commonly called the ellipsoid zone (EZ);
outer segments (OS); interdigitation zone (IZ);and retinal pigment epithelium
(RPE), choriocapillaris (CC), choroid (C), and sclera (S) (Figure 16-1) (1–3).
Unlike adults, in infants and children OCT layers rapidly appear, disappear,
and change in thickness at different retinal locations based on the stages of
the eye development as described below. Thus, age should be considered in
the interpretation of pediatric OCT images (Figure 16-2).
FIGURE 16-1 Healthy infant and older child. SDOCT
image of the immature macula/fovea of a 34-week-old
(corrected age) premature infant (top) and mature
macula/fovea of a 17-year-old child (bottom). SDOCT
shows alternating bands of hyperreflective (nerve fiber and
plexiform) and hyporeflectivity (nuclear) layers labeled
from inner to outer: note the internal limiting membrane
(ILM) is not visible as a distinct layer unless elevated off
of the NFL, retinal nerve fiber layer (RNFL); ganglion cell
layer (GCL); inner plexiform layer (IPL); inner nuclear
layer (INL); outer plexiform layer/photoreceptor synapse
layer (OPL/PSL); outer nuclear layer, which includes
Henle fiber layer (ONL+HFL);external limiting membrane
(ELM); inner segment band/ellipsoid zone (IS/EZ); outer
segments (OS); interdigitation zone (IZ); and retinal
pigment epithelium (RPE); choroid (C); and sclera (S).
Note that multiple photoreceptor layers (PRLs) that are not
present at the fovea in the infant are present in the mature
retina. Central foveal thickness is measured from the ILM
to the RPE, the inner retina includes layers from the ILM
to include the INL, and the outer retina extends from the
OPL to the inner border of the RPE and includes the PRL.
Some groups include the OPL as part of the PRL, while
others measure the PRL from the outer aspect of the OPL
to the inner border of the RPE. When macular volumes are
measured, the mean thickness in the 1-mm circular region
centered on the fovea may be called central subfield
thickness or sometimes foveal thickness.
FIGURE 16-2 Foveal development. SDOCT image of the
foveal center of a 35-week-old (top) and 39-week-old
(bottom) premature infant with ROP. Note that inner
retinal layers have migrated further out of the foveal pit in
the older child and that photoreceptor layers such as the
ellipsoid zone (EZ) and outer segments (OS) are more
developed in the older infant. The EZ is not yet present at
the foveal center in the 39-week-old infant.
Terminology
1. Central Foveal Thickness: The thickness of the entire retina extends
from the inner aspect of the internal limiting membrane (ILM) to the
inner aspect of the RPE at the foveal center.
2. Inner Retina: The inner retinal layers (IRLs) include all the retinal layers
from the inner aspect of the ILM to the outer border of the INLs.
3. Outer Retina: The outer retinal layer extends from the inner aspect of the
OPL to the inner border of the RPE.
4. Photoreceptor Layer: The photoreceptor layer (PRL) extends from the
outer aspect of the OPL to the inner border of the RPE.
OCT SYSTEMS FOR PEDIATRIC

IMAGING
The era of pediatric OCT imaging started with the advent of faster OCT
systems, such as spectral domain (SD) (and more recently with swept source
[SS] devices) that have shorter acquisition times and, therefore, are more
advantageous for imaging in pediatric patients. In the past, tabletop systems
were used for imaging young pediatric patients in “flying baby” position, or
tabletop systems were custom adapted for imaging supine infants (5). With
the development of a commercially available portable handheld SDOCT
system (Leica, Research Triangle Park, NC, USA) (6), imaging infants and
children in the clinic, operating room, or hospital setting has become
mainstream with an expansion of use for accurate diagnosis, assessment of
disease stage, and assessment of response to treatment (7,8). Other handheld
or armature OCT devices (9–12) are currently under investigation and
provide additional advantages in pediatric imaging.
The faster OCT systems have also allowed for novel imaging of vascular
flow without the use of injected dyes, which can be especially valuable in the
pediatric population. Optical coherence tomography angiography (OCT-A) is
a noninvasive, high-resolution, depth-resolved imaging modality that enables
superior visualization of retinal microvasculature, with differentiation of
superficial, penetrating, and deep vascular complexes (9,13,14) and of CC
and choroidal vasculature. OCT-A has advanced the study of diseases
affecting retinal vasculature and allows for quantifiable, high-resolution
visualization of areas of retinal ischemia and neovascularization in vascular
diseases especially in the macula. Features such as the area of the foveal
avascular zone (FAZ) can be clearly and reproducibly measured on OCT-A
(9,15). Furthermore, the noninvasive, dyeless nature of OCT-A is ideal for
imaging infants and children and has been shown to allow for frequent
monitoring and follow-up including treatment response. Application of OCT-
A to infants and children to examine pediatric retinal microvasculature
currently involves research handheld or portable OCT-A systems (9,11,16)
and may provide insights in healthy (Figure 16-3) or diseased (Figure 16-4)
microvascular development. Other high-resolution portable OCT systems
even enable imaging of photoreceptor development and inherited diseases
(12).
FIGURE 16-3 Range of foveal avascular zone

morphology in the eyes of term-born children without eye
disease. Optical coherence tomography angiography
(OCT-A) allows for visualization and fine demarcation of
the superficial and deep venous complex (SVC, DVC) of
the foveal avascular zone (FAZ). Note that there is a vast
range of normal FAZ sizes as illustrated in these three
children.
FIGURE 16-4 Retinopathy of prematurity (ROP). OCT-A
illustrates irregular, angular vascular pattern with several
large vessels diving down into the inner retinal layers.
Aberrant vascularization of FAZ is noted here. SVC,
superficial venous complex; DVC, deep venous complex.
Methods and Limitations of OCT Imaging in Infants

and Young Children
Optical coherence tomography imaging is routinely performed in the care of
adult retina patients and older pediatric patients and not as often in the care of
young pediatric patients. The more limited use of OCT imaging is primarily
due to challenges in obtaining an image in a nonsedated, preverbal child or a
verbal child who may not be able to cooperate for fixation or for the duration
of scanning with a tabletop system.
OCT imaging in young children is feasible across multiple settings
including at the inpatient bedside, in clinic while supine or upright, and in the
surgical suite while under anesthesia (17,18). Imaging of an upright or supine
child may be performed with a portable, noncontact handheld SDOCT system
such as the Envisu 2300 (Leica, Research Triangle Park, NC) (Figure 16-5).
The ability to maneuver the handheld probe is useful for awake infant and
child imaging. Nonsedated infants and children may be imaged at the bedside
in an incubator or bassinette or in clinic on an examination table or the
parent’s lap. The handheld OCT system has been shown to provide reliable
measurements in children with and without nystagmus (19). In neurologically
unstable infants or children, SDOCT imaging can often be performed at the
bedside through the undilated pupil (20). One method to stabilize the
handheld system is to hold this near the distal end of the imaging bore and
rest the tip for stability on the back of the fingers of the other hand; those
fingers hold the eyelids open and/or stabilize against the
eyelids/forehead/cheek to maintain a constant distance from the eye [20,
including the published videos] (Figure 16-5A and B). Verbal children
typically 4 to 5 years and above may be imaged by standard tabletop OCT
device by an experienced imager. A stepstool for positioning can be useful in
toddlers, while fixation toys or smart phone held by a second person can be
useful in infants and young children (6) (Figure 16-5C). Finally, pediatric
patients with retinal disease often need to be evaluated under anesthesia in the
operating suite, to allow for complete retinal examination and fluorescein
angiography (FA) and treatment. Imaging with a portable OCT unit is often
valuable in the examination under anesthesia setting. Here, because of the
immobility of the patient, either handheld OCT systems or armature OCT
systems (such as the investigational Flex module of the Spectralis OCT
system, Heidelberg, FRG) may be useful.
FIGURE 16-5 OCT imaging of a child with a handheld
system in nursery (A), in clinic (B), and with a tabletop
system and stepstool (C). The imager holds the handheld
system by the bore and rests against fingers of the opposite
hand. A second person helps to attract the attention of the
toddler at the tabletop system.
The axial length of the infant eye changes by 9 mm from birth to age 10, and
this affects reference arm length for OCT imaging and the scaling factor for
lateral measurements. While this is also relevant for measurements across
different adult eyes, only in children do we need to consider the change in
axial length in the same patient from month to month (6). Particularly, one
should take into account patient’s age and optics of the eye when setting
imaging parameters for infants and young children and measuring ocular
structures on those images (6).
The portable, noncontact, handheld SDOCT unit (Envisu 2300, Leica,
NC) consists of an imaging hand piece connected via a flexible cable to an
SDOCT engine mounted on a rolling cart. The SDOCT system has a
calibrated knob to adjust the reference arm position settings for the axial
length of the eye based on age (6). The handheld probe has a focus correction
adjustment with a range of +10 to −12 D, which limits imaging in the setting
of aphakia. When imaging infant eyes in the clinic or hospital unit, we have
found that the near infrared light of SDOCT imaging is tolerated better by
infants than the clinical exam with white-light indirect ophthalmoscopy. The
illumination from the Envisu system appears to the patient as a faint red line
against a black background. Swept source OCT, which typically uses light
even further in the infrared, is usually not visible to the patient. We have also
found that the operator could readily hold the eyelids open in the awake or
sleeping infant and that a lid speculum was thus not necessary for this
imaging. Adding artificial tears before imaging, especially when under
anesthesia, provides a more stable tear film and often better signal in the OCT
images.
Infant Foveal Development Considerations for OCT

Interpretation
OCT is a valuable tool in the evaluation of retinal development during critical
time in visual development, and preterm birth may impact this development.
One must recognize the normal morphology for age when assessing for
disease, degeneration injury, or abnormal development. In neonates,
especially those born preterm and examined in the nursery before the time of
expected term birth, the RPE and the choroid are thinner (21,22), and the
outer retina is extremely immature especially in the fovea. OS elongate, and
the EZ matures in the third trimester and after term age, creating a visible EZ,
distinct from the RPE. The ELM is visible before the EZ, and the EZ is
visible in almost 50% of healthy infant foveae at term birth (23). The outer
segment–RPE interface continues to mature in the macula from around 13 to
15 months till approximately 5 years of age (24). Before birth, the IRLs,
especially IPL and INL, are present across the foveal pit; they subsequently
shift out of the foveal center. These in vivo changes of inner and outer retinal
layers at the foveal center are consistent with cellular redistributions reported
in histologic studies (1,2,23–26).
Compared to the full-term infant, an older child or adult, the premature
eye has a shallower foveal depression, presence of distinct IRLs at the foveal
center, thinner retinal layers overall, and attenuation of the PRL with a
delayed appearance of the ELM and EZ relative to the term-born infant (Figs.
16-1 and 16-2) (1,27,28). Visual acuity correlates with events relating to
foveal maturation that include IRL fusion and the completion of the EZ in the
first year of life in premature infants (15). An infrequent SDOCT finding in
healthy infants within days of term birth is a pocket of subretinal fluid
centered on the fovea. This resolves in days to weeks and has not been linked
to vision or health problems (29).
FINDINGS FROM OCT IMAGING IN

PEDIATRIC RETINAL DISEASES
OCT images of the vitreous, retina, RPE, and choroid have revealed relevant
information about structural morphology and vascular flow that has been
shown to be critical in the diagnosis and in guiding the care for a wide range
of pediatric retinal diseases (7,8,26,30). The list below includes several
examples of diagnostic information that is only available from OCT imaging
and that is not available on clinical examination in these pediatric retinal
diseases. This topic is broad, and thus the list is of necessity incomplete.
Retinopathy of Prematurity
The current gold standard for retinopathy of prematurity (ROP) diagnosis is
clinical examination performed by trained specialist using indirect
ophthalmoscopy. Additionally, telemedicine has gained traction, and in
certain centers, screenings are facilitated by capturing and reviewing wide-
field fundus photographs using contact camera systems such as RetCam
(Clarity MSI, CA, USA). These bedside examination tools allow for
ophthalmoscopic en face viewing of the premature retina. The cross-sectional
visualization of the retina and its vasculature from OCT imaging in preterm
infants and young children has added relevant information about the effects
of ROP across the developing retina. The previously unrecognized retinal
findings in ROP will be useful in the assessment of clinical outcomes (31).
As noted above, with SDOCT imaging, the premature eye has a
shallower foveal pit, presence of IRLs at the foveal center, thinner outer
retinal layers including attenuated photoreceptors, and absence of inner and
OS (Figure 16-2) (1,25). Notably, 20% to 50% of premature infants have
intraretinal cystoid changes described as macular edema of prematurity
(Figure 16-6) (32–37). Intraretinal cystoid spaces are only rarely seen in term
infants, and if these do occur, they are most common with a systemic disease
such as liver failure (38). Macular cystoid spaces are a subclinical feature
only identified by OCT, although in severe cases of macular edema, the
blunted fovea may be discerned on the clinical examination; in some cases, it
has been shown to leak on FA (39). Furthermore, macular edema of
prematurity has, in some cases, been associated with poorer visual acuity and
neurodevelopmental outcomes (40).
FIGURE 16-6 Macular edema of prematurity in 36-week-
old premature infants. Note the hyporeflective intraretinal
cystoid spaces of so-called macular edema (ME) of
prematurity. They resemble macular edema seen in adult
diseases. Varying degrees of macular edema are illustrated
starting with mild ME (foveal depression persists with
small cystoid spaces), moderate ME (loss of foveal
depression and retinal thickening with columnar-like
cystoid spaces), and severe ME (marked retinal thickening
with columnar-like cystoid spaces).
Outside of the fovea, SDOCT can reveal the three-dimensional aspects of

plus disease, causing retinal deformation (41), and of the development of
extraretinal neovascular buds, bridging networks and plaques (42) (Figure
16-7). With OCT, we can monitor the regression of neovascular complexes
and whether this occurs with or without tractional retinal schisis (43). In stage
4 ROP, SDOCT allows us to localize subretinal fluid and determine whether
there is foveal detachment (stage 4B vs. 4A), which impacts visual prognosis
(31). With OCT imaging, we can readily differentiate retinal detachment
from schisis, and we have learned that ROP-induced tractional retinal schisis
may appear clinically to be a retinal detachment (42,43) (Figure 16-8).
FIGURE 16-7 Three-dimensional OCT images of ROP
with tiny neovascular buds posterior to the vascular–
avascular junction (A) and bridging networks of
neovascularization at the vascular–avascular junction (B)
in stage 3 ROP.
FIGURE 16-8 OCT is useful to distinguish retinoschisis
(arrow, A and B) with persisting outer retinal attachment
from retinal detachment (asterisk, A and B) with no retinal
structures remaining attached to the retinal pigment
epithelial surface.
Early OCT-A allows capture of aberrant retinal perifoveal microvascular

development (44) and also the remodeling process during the critical period
of ROP pathogenesis, particularly in development of the superficial vascular
complex (Figure 16-4) (13,14). OCT-A has been used to image the
neovascular pathology in aggressive posterior ROP (APROP). Conventional
OCT as well as dye-based angiography preclude the detection of deeper
extensions of neovascularization. OCT-A on the other hand detected the
involvement of a “complex” in the deep capillary plexus and outer retinal
layers with corresponding flow outlines of the superficial neovascularization
suggesting a change in nomenclature from “flat” neovascularization to flat
neovascular “complex” in APROP (45). In school-aged children, OCT-A has
been used to visualize the small FAZ associated with the persistence of IRLs
and related to treated ROP (46) and to preterm birth (47).
Familial Exudative Vitreoretinopathy

Familial exudative vitreoretinopathy (FEVR) is a rare, inherited disorder of
retinal vascular development leading to incomplete and anomalous
vascularization of the peripheral retina (further information in Chapters 35
and 66). The anomalous peripheral vascularization is primarily seen on FA.
SDOCT allows for visualization of retinal changes such as foveal
displacement as seen in mild FEVR and then diminished foveal contour,
cystoid macular edema, retinal thickening, ERM formation, and EZ
disruption in severe FEVR (48). SDOCT can be used to diagnose and
monitor subclinical progressive optic nerve head elevation from FEVR and
after vitrectomy, the recovery from anteroposterior traction-induced elevation
(49).
Ultra–wide-field imaging and FA have been used to describe a range of
associated retinal features, such as aberrant peripheral vessels, arterial
tortuosity, telangiectasias, and capillary agenesis in FEVR. While these gross
vascular abnormalities are observed on FA, the OCT-A in FEVR allows us to
visualize abnormalities in the vertical penetration of the deeper retinal
vascular layers due to incomplete angiogenesis. Notable abnormalities
involve both the superficial vascular complex (increased vessel dilation,
presence of vascular curls and loops, and straightening of the macular
vasculature) and the deep vascular complex (disorganized vascular pattern
with stub-like vessel terminations or “end-bulbs”) (50).
Incontinentia Pigmenti
Incontinentia pigmenti (IP) is a rare, inherited retinal vascular disordered
with X-linked dominance (discussed in Chapters 36 and 66). It presents with
ocular findings that include retinal vascular occlusion, neovascularization,
hemorrhages, foveal abnormalities, and retinal detachments, some of which
are readily recognized on FA. However abnormalities such as inner and outer
retinal thinning are identified on SDOCT that are likely related to ischemia
and are not readily picked up on clinical exam or funduscopic imaging and
may explain changes in visual acuity (51,52) (Figure 16-9).
FIGURE 16-9 In incontinentia pigmenti, the OCT retinal

view (left) shows the straightened retinal vessels. The B-
scan (right) shows the focal patch of thinning of the inner
retinal layers superiorly (left side of scan) with severe loss
of nerve fiber layer.
Coats Disease
Coats disease, a retinal vascular disease, is typically unilateral and affects
young boys (more details Chapter 64). Although clinical examination and
fluorescein-guided treatment remains gold standard, SDOCT can be used to
identify and monitor distinct retinal and subretinal features such as
intraretinal or subretinal exudation, intra- or subretinal fluid, outer retinal
layer loss, subretinal fibrosis, and retinal thickening (53,54) (Figure 16-10).
The SDOCT features of subretinal fibrosis and outer retinal atrophy are
associated with poorer visual acuity. Thus, OCT contributes to the prognostic
factors and response to treatment.
FIGURE 16-10 In Coats disease, the OCT image reveals

the location of exudates (hyperreflective spots) within the
inner and deep retinal layers, the absence of subretinal
fluid, and a hyperreflective subretinal nodule within the
fovea.
Pediatric Choroidal Neovascular Membranes

Similar to adult choroidal neovascular membranes (CNVMs), pediatric
CNVMs are readily identified by SDOCT. Findings include subretinal
vascular complex/plaque, subretinal fluid or hemorrhage, and intraretinal
fluid or hemorrhage. Unlike adults, children often do not report visual
disturbances, and thus SDOCT aids in proper diagnosis and allows for
treatment response follow-up (55).
Pediatric Epiretinal Membrane

Pediatric epiretinal membranes, such as secondary to combined hamartomas
of the retina and RPE, have been very well described and are addressed in
detail in Chapter 67. They have characteristic features on OCT that include
hyaloidal thickening and traction, retinal thickening with deep retinal folds,
and disorganization of retinal layers (Figure 16-11). Similar to adults, intact
outer retinal layers reflect visual potential after surgical removal (56–58).
FIGURE 16-11 Pediatric epiretinal membrane with striae
(left) in which the OCT B-scan (right) reveals the
membrane and condensed hyaloid remain attached across
the inner retinal surface and create a deep fold of
thickened retina with small intraretinal cystoid spaces.
Retinoblastoma
In patients with retinoblastoma (Chapters 44 and 45), OCT can detect
recurrent retinal tumors that are not visible clinically, as well as
retinoblastoma-associated epiretinal deformation, retinal edema, subretinal
fluid, and optic nerve and vitreous changes (59). It is also useful in evaluating
diffuse infiltrating retinoblastoma and monitoring the response to treatment
(60).
Trisomy 21
OCT appears useful to study subclinical features of the macula and correlate
them with visual acuity and vision development in trisomy 21 or Down
syndrome (DS). Inner retinal fusion was noted in only 15% of cases
compared to all cases of matched normal children. Other layers that were
abnormal in children with trisomy 21 include the OPL, the ELM, and the IZ.
Thus far, these OCT findings have not been linked to visual acuity. It is
possible that in the future, OCT will help us understand the anatomical basis
of subnormal vision development in this and other syndromes where the
fundus may appear clinically normal (61,62).
Inherited Retinal Diseases
OCT and OCT-A are especially useful for inherited retinal diseases. As
described in the disease-specific (Chapters 22–37), OCT information is often
diagnostic and can provide important information as to the severity of the
phenotype and the interval disease progression. For example, in albinism,
foveal hypoplasia is visible on a macular volume OCT. Similar to the fovea
in preterm infants and infants treated for ROP in which the IRLs persist at the
foveal center, there is absence of a FAZ (63). In juvenile X-linked
retinoschisis, characteristic foveal schisis occurs first in the INL and then in
OPL/ONL on OCT (12,64). The extent and severity of outer retinal atrophy
and loss of photoreceptors or retinal nerve fiber layer (RNFL) atrophy can be
readily noted on OCT in diseases such as Leber congenital amaurosis, Best
disease, Stargardt disease (Figure 16-12), and other inherited retinal diseases
(8). These features can be useful in documenting anatomic disease severity
and progression.
FIGURE 16-12 Photoreceptor loss in Stargardt disease.

SDOCT illustrates marked loss of outer retinal layers in
the macula/fovea secondary to the inherited retinal
dystrophy.
SUMMARY
Today, OCT is an important diagnostic tool during initial examinations and is
increasingly utilized in monitoring pediatric retinal diseases during follow-up
examinations, especially in older children. As we continue to use OCT and
add OCT-A imaging to examinations in infants and young children, we will
continue to gain new insights from findings not previously recognized by
conventional examination and non-OCT imaging modalities.
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17
Fluorescein Angiography in Pediatric
Retinal Diseases
Nikisha Kothari, Niranjan Manoharan, and Irena Tsui
Fluorescein angiography (FA) is an important tool in retinal vascular

conditions that often guides diagnosis, management, and treatment. The
discovery of FA in 1960 by two medical students was serendipitous, and its
value for clinical use in ophthalmology was not immediately recognized (1).
Modern-day FA imaging has been adapted for infant and pediatric
populations and includes imaging iris vasculature as well.
FLUORESCEIN DYE
Sodium fluorescein is an orange-red crystalline hydrocarbon with a molecular
weight of 376 daltons. It is available as 10% fluorescein dye (Fluorescite,
Alcon Inc., Fort Worth, TX), and pediatric patients are dosed by weight (5 to
7.5 mg/kg, maximum dosage 500 mg) for intravenous use. In a clinic setting,
intravenous access and image acquisition can be challenging in children.
Prior to injection, a clear imaging plan should be communicated between
multiple team members. Typically, one person is positioning the head, the
second person is injecting dye in a standard 6-second bolus or faster fashion,
and a third person is using the camera. Transparent tubing is attached to a 25-
gauge butterfly needle for injection, and blood should be seen to draw back
before injecting. An intravenous line can be preplaced if early phases of the
angiography are important to acquire, but they are often not crucial in
pediatric patients.
Performance of FA requires a fundus camera with an excitation and
barrier filter to selectively image retinal vasculature and detect pathology.
White light passes through an excitation filter to only allow blue light at 465
to 490 nm to illuminate the retina. Eighty percent of fluorescein is protein
bound, mainly to albumin, and not available for fluorescence. The remaining
20% unbound fluorescein molecules absorb blue light, fluoresce, and emit
yellow-green light at 520 to 530 nm. The barrier filter then only allows
yellow-green light to reach the camera. Ideally, images are acquired
immediately after injection and then at varying intervals until 10 to 15
minutes after injection (2).
In the absence of pathology, fluorescein dye stays within the circulation
of the retina. The endothelial cells of retinal blood vessels branching from the
central retinal artery serve as the inner blood–retina barrier. Fluorescein dye
normally passes through the fenestrated walls of the choriocapillaris. The
zonulae occludens between retinal pigment epithelial cells provides the outer
blood–retina barrier, separating the pool of fluorescein in and around the
choriocapillaris from the retina.
Fluorescein dye and its metabolites are eliminated by the liver and
kidney. Skin can have a yellowish tinge hours after a fluorescein injection,
and urine can remain fluorescent for 24 to 36 hours. Large series in adults
have estimated rates of mild adverse events to be 1% to 14% and most
commonly, transient nausea and vomiting (3). In high-risk elderly and
hypertensive adults, the overall adverse reaction rate has been reported to be
11% (4). Other less common side effects include vasovagal syncope,
urticaria, pyrexia, and rarely a more serious reaction, such as bronchospasm
or even anaphylaxis. In newborns, cell-mediated immunity does not develop
until 2 to 3 months of age, so an adverse reaction to fluorescein dye is less
likely (5).
Angiographic phases consist of choroidal flush, arterial phase,
arteriovenous phase, venous phase, and recirculation. Choroidal filling occurs
slightly early at 10 to 12 seconds (2). In the following 10 seconds, laminar
venous flow can be observed. Thirty seconds after injection, fluorescein
begins to empty from the choroidal and retinal vasculature. Typically, after
10 minutes, minimal fluorescein dye is visible in the retinal vasculature;
however, structures that normally stain such as the optic nerve head, Bruch
membrane, and sclera become visible during this time. The macula is dark
due to blockage of choroidal fluorescence by densely packed retinal pigment
epithelial cells and xanthophyll pigment (2).
Abnormal findings in a FA can be categorized as areas of
hypofluorescence or hyperfluorescence. Common examples causing hypo-
and hyperfluorescence are listed in Table 17-1.
Table 17-1 Etiology of abnormal findings on

fluorescein angiography
Hypofluorescence can typically be thought of as blocked fluorescence or

vascular filling defects. Blocked fluorescence occurs due to decreased normal
fluorescence of retinal or choroidal structures due to fluid or tissue barrier to
light anterior to the respective retinal or choroidal circulation. Vascular filling
defects occur due to failure of blood stained with fluorescein to enter
occluded vessels.
Hyperfluorescence can be categorized as a window defect, leaking,
pooling, or staining (Table 17-1). Window defects occur due to increased
transmitted choroidal fluorescence due to absence of pigment in the retinal
pigment epithelium that typically acts as a barrier to choroidal fluorescence.
Leakage is abnormal late hyperfluorescence, which shows gradual
enlargement and blurring of margins. Pooling is leakage of fluorescein
confined to a distinct anatomic space. Staining refers to leakage into tissue
with an increase in fluorescence intensity within constant, discrete borders.
Abnormal retinal and choroidal vasculature can also be visualized with
FA. Examples include anastomosis, aneurysms, neovascularization,
telangiectasia, vessel tortuosity/dilation, and persistent hyaloidal vessels or
tunica vasculosa lentis.
ORAL FLUORESCEIN DYE

There have been several reports demonstrating the use of ingested fluorescein
dye (Fluorescite, Alcon Inc., Fort Worth, TX) with good image quality and
clinical utility (6–8). Fluorescein dye can be diluted to a final 2% solution (20
to 25 mg/kg, max dose of 1,000 mg) using beverages such as orange juice,
apple juice, or milk. Oral FA avoids the difficulty of obtaining intravenous
access in pediatric patients but cooperation is still needed to acquire images.
The first images are visible as early as 5 minutes after the ingestion of the
oral fluorescein (6). Image timings that have been recommended for oral
fluorescein are 5, 10, 15, and 20 minutes after ingestion (7). Images are taken
at a slower pace given the increased absorption time of fluorescein dye
through the gastrointestinal system into circulation with oral ingestion and
extend up to 30 minutes. This limitation of oral fluorescein prolongs the test
but also allows time to work with uncooperative patients. Adverse reactions,
including itching, discomfort, and nausea, have been rarely reported with oral
fluorescein dye. No cases of anaphylaxis have been reported to date (6–8). A
limitation is the inability to capture the choroidal flush, arterial phase, or
early laminar venous phase. However, the arterial phase and early laminar
venous phase are usually not crucial in most pediatric retinal and uveitic
conditions. A recent study comparing oral and intravenous FA using ultra–
wide-field FA (Optos, Dumfernline, MA) showed that there was similar
image quality in diseases, such as Coats disease, familial exudative
vitreoretinopathy (FEVR), and pars planitis, and a masked grader could not
tell the difference in route of fluorescein dye administration (8). Therefore,
substitution of off-label oral fluorescein dye for intravenous injection may
allow for FA imaging of patients, who are fearful of needles in an outpatient
setting.
INDOCYANINE GREEN
ANGIOGRAPHY
Indocyanine Green (ICG; Akorn, Lake Forest, IL) is a 775 dalton water-
soluble dye that is used to image the retina and choroid with an excitation
filter of 805 nm and barrier filter of 500 and 810 nm. ICG is contraindicated
in patients who are allergic to shellfish or iodine. The product comes as a
green powder with aqueous solvent and should be used within 6 hours of
mixing. ICG is 98% protein bound after intravenous injection and therefore
stays within the choriocapillaris, making it advantageous for imaging the
choroidal circulation. In children, ICG angiography may be useful for
evaluating posterior uveitis, choroidal hemangiomas, or other suspected
choroidal pathology. The Spectralis (Heidelberg, Germany) uses a scanning
laser ophthalmoscope (SLO) and performs simultaneous FA and ICG
angiography. In addition, the California (Optos, Dunfermline, MA) can now
provide ultra–wide-field ICG angiography.
FIELD OF VIEW
Traditional fundus cameras capture a single frame 30- to 60-degree field of
view and require patient cooperation and time to focus the camera, and these
limitations make traditional FA challenging in children. Moreover, the time-
sensitive nature of FA adds an additional obstacle to capturing simultaneous
peripheral sweeps and producing montage images using traditional cameras.
It was not until the advent of handheld FA (RetCam, Clarity Medical
Systems, Pleasanton, CA) that allowed access to a wide field of retinal view
in supine infants and ultra–wide-field imaging (Optos, Dumfernline, MA) of
upright young patients using scanning laser technology and decreased image
acquisition time when FA in children began to be routinely performed. The
major ocular factor affecting field of view is pupil size, and, therefore,
dilation is helpful in children regardless of the type of camera used. There are
various definitions of what constitutes wide-field imaging with one definition
being images with a >80 degree field of view while reserving ultra–wide-
field terminology for images that capture >130 degree field of view.
Recently, an International Wide-Field Imaging Study Group has proposed the
term wide field be applied to imaging that captured retinal imaging beyond
the macula but posterior to vortex vein ampulla. They recommended using
the term ultra-wide field to refer to images that capture the retina anterior to
the vortex vein ampullae (9).
CONTACT IMAGING MODALITIES

The first commonly used wide-field imaging modality was the RetCam
(Clarity Medical Systems, Pleasanton, CA), which was developed by Massie
in 1997, and is able to capture a 130 degree view of the fundus. Since its
introduction to the market over 20 years ago, clinical and research
applications of the RetCam machine have focused primarily on telemedicine
screening, diagnosis, and management of ROP and other pediatric retinal
diseases and injury. Moreover, its portability on wheels makes evaluation at
the bedside in a neonatal intensive care unit (NICU) or the operating room
(OR) practical. A laptop version of the RetCam called the shuttle is useful for
transporting the camera between sites and can capture color fundus images
but does not have FA capability. There are a variety of lenses with the
RetCam including 130 degrees, 120 degrees, 80 degrees, 30 degrees, and
Portrait, which is used for external and adnexal images. Most commonly,
pediatric retinal images are taken with the 130 degree lens, which can be
positioned to image up to the ora serrata during an examination under
anesthesia. However, the 80 degree lens is valuable for high-resolution
imaging in the macula.
RetCam is a handheld fiber-optic contact lens system, which allows for
imaging recumbent children and requires rotation of the globe or tilting of the
handpiece to acquire peripheral images. The technology is ideal in infants
who can be swaddled and held down without sedation. When using the
camera in older children, an examination under anesthesia is necessary
because of the bright light and contact with the ocular surface. Typically, an
eyelid speculum and viscous coupling agent such as Gonak (Akron, Lake
Forest, IL) or water-based artificial tears gel is used. The handpiece cord is
held superiorly (12 o’clock) by convention to preserve orientation of fundus
images of macula to the left of the optic nerve for images of the right eye and
the converse for images of the left eye. Imaging should be performed with
minimal pressure from the weight of the handpiece on the globe by resting
either part of a hand or arm on a stable surface; otherwise vascular features
may be altered as a consequence of increased intraocular pressure and venous
congestion. The computer screen is positioned to face the photographer
during image acquisition for maximal ergonomics and speed of acquisition.
Focus can be adjusted with a close range target before placing the camera on
the eye. Adjusting the light intensity, focusing, and image capture can be
performed with a foot pedal or by using the keyboard console with the help
of an assistant. The midperiphery is imaged with relative ease. Scleral
indentation and rotating the eyeball are useful to capture pathology in the far
periphery. Images can be captured in snapshots or by recording a video and
going back to save select snapshots from the video.
The RetCam has proprietary software for image manipulation and review
saved in the default format (MLX), although JPEG, DICOM, and PNG are
also available. The interface is user-friendly and allows for image transfer to
portable media in JPG or PNG format for viewing without proprietary
software. One of the most important features of the RetCam software is the
compare function, whereby images from different dates can be reviewed side
by side. This feature facilitates longitudinal review of subtle and difficult-to-
quantify fundus changes, such as vascular tortuosity and dilation in plus ROP
disease. A note entry feature includes input of additional data, which allow
database search by date, physician, disease, and other parameters.
Performing FA with the RetCam requires three adjustments to the
equipment. The light source, lens filter, and software need to be switched
over to FA function. The addition of the lens filter can be cumbersome as it
requires the lens’ tip to be removed and replaced, which should be done
carefully to avoid damaging the lens’ tip attachment (Figure 17-1) (10).
Limitations of the RetCam include difficulty in acquiring high-quality
images: if there is media opacity, poor dilation, dark fundus, and the
reflection artifact from intraocular lenses.
Figure 17-1 Photograph demonstrating the installation of

the barrier filter on the RetCam.
Fundus images from the RetCam are valuable to document injury and disease
as demonstrated in fundus photographs and FA of ROP (Figures 17-2 and
17-3). The use of FA in ROP posttreatment has increased now that anti–
vascular endothelial growth factor (anti-VEGF) is being used as first-line
therapy (Figure 17-3). In nonaccidental trauma, fundus photographs are
important for medical legal documentation and to record number, extent, and
location of retinal hemorrhages (Figure 17-4), and FA can detect
nonperfusion that may benefit from laser prophylaxis or help counsel
regarding prognosis. In retinoblastoma, tumor infiltration, nonperfusion, and
retinal detachment can be detected with FA (11,12) (Figure 17-5).
Figure 17-2 RetCam fundus photograph images (A, B)
and FA Figure 17-2(Continued ) (C) of a 38-week
postmenstrual-age neonate born at 25 weeks of gestation
with a birth weight of 670 g demonstrate tortuous vessels,
peripheral nonperfusion, and leakage.
Figure 17-3 RetCam fundus images (A, B) and FA Figure
17-3(Continued) (C, D) of a 48-week neonate born at 24
weeks and 5 days of gestation with a birth weight of 540 g
and treated with bevacizumab 0.625 mg in both eyes at 34
weeks demonstrate peripheral nonperfusion and vascular
leakage in both eyes.
Figure 17-4 RetCam fundus images and FA demonstrate
retinal hemorrhages and nonperfusion 5 days after
nonaccidental trauma (A, B) and resolution of
hemorrhages with persistent nonperfusion 5 months after
trauma (C, D). (Photograph courtesy of Audrina Berrocal,
Bascom Palmer Eye Institute, Miami, FL.)
Figure 17-5 RetCam fundus image (A) and FA (B)
demonstrate a sensory retinal detachment (arrowhead),
tumor infiltration (arrow), microvascular changes (arrow
on FA), and nonperfusion (arrowhead on FA) in a patient
with retinoblastoma. (A: Reproduced from Kim JW, Ngai
LK, Sadda S, et al. Retcam fluorescein angiography
findings in eyes with advanced retinoblastoma. Br J
Ophthalmol 2014;98(12):1666–1671. With permission
from BMJ Publishing Group Ltd.; B: Photograph courtesy
of Michael Trese, Associated Retinal Consultants, Royal
Oak, MI.)
Another contact camera for pediatric retina use is the Phoenix ICON
(Pleasanton, CA), which has the ability to perform FA. The Phoenix may
provide optimal color images in deeply pigmented eyes. An example of the
peripheral view can be seen in a Phoenix ICON FA image of FEVR treated
with panretinal photocoagulation (Figure 17-6).
Figure 17-6 Phoenix ICON FA in a patient with FEVR
demonstrates staining of laser scars. (Photograph courtesy
of Michael Trese, Associated Retinal Consultants, Royal
Oak, MI.)
NONCONTACT IMAGING
MODALITIES
Ultra–wide-field fluorescein angiography (UWFFA) with the Optos (Optos,
Dumfernline, MA) has allowed noncontact FA imaging in a clinic setting
since 2005. Optos captures up to 200 degrees of the fundus and does not
require dilation in adults. In children, dilation is recommended to reduce
cooperation requirements by decreasing image acquisition time (5,6). The
technology uses an elliptical mirror and creates a virtual focal point inside the
patient’s eye to image the far periphery. Each picture is 3,900 × 3,072 pixels.
Besides FA, wide-angle color fundus photographs, red-free, infrared,
autofluorescence, and ICG images can be taken with the Optos camera.
Optos FA has been used to evaluate retinal vascular diseases and other
peripheral retinal pathology in adults (13). The device provides several
unique advantages in the pediatric population: wide field of view, complete
fundus imaging at one point in time, rapid acquisition time (0.2
seconds/frame), and use of a confocal SLO platform, which reduces optical
aberrations (14). The lower age limit in cooperative children has generally
been 3 years of age (6,15,16). The advantages of Optos FA over traditional
FA include facile detection of far peripheral pathology such as nonperfusion
(Figure 17-7), neovascularization, and vascular leakage, which are relevant
in pediatric diseases, such as ROP, FEVR, and Coats disease. The camera can
also visualize the retina through moderate media opacities, such as cataract
and vitreous hemorrhage. The Optos viewing software can process images
either manually or automatically to adjust image quality and contrast, and
steering software allows automated montage images.
Figure 17-7 A, B: UWFFA demonstrates peripheral
nonperfusion in both eyes of a 10-year-old child with
history of prematurity.
Several studies have evaluated the utility of Optos for FA in cooperative

children in clinic without the necessity of imaging under general anesthesia
(17,18). Depending on the age of the patient, he or she can sit on a rolling
chair with both feet on the floor, stand on a footstool, or sit in the lap of the
parent. The child’s head is positioned on the chin rest with forehead against
the upper bar. The photographer then encourages the child to look at the
fixation target. The flying baby technique has been described for imaging of
premature infants with ROP (18). In this method, a caregiver holds the
infant’s head on his or her shoulder with the neck slightly extended to bring
the camera aperture close enough to the infant’s eye. A speculum is usually
used, and vestibulo–ocular reflex is used to encourage ocular alignment if the
eye rotates out of primary position. This has even been performed in
ventilated babies with a supported endotracheal tube and full medical
supervision in the NICU (19).
A limitation of Optos is decreased resolution and field of view of the
superior and inferior periphery due to eyelid and eyelash artifact. Lid
retraction using fingers, q-tips, tape, or an eye lid speculum can be used if
tolerated to reduce lash artifacts, but this can dry out the cornea and is
generally poorly tolerated in children. Another limitation of Optos is that
color fundus images are taken with two channels only (633 nm red and 532
nm green) and the absence of blue channel images results in a green-
dominant color aberration. This can limit interpretation of color images
particularly in pediatric uveitic conditions, in which color is important.
Spectralis (Heidelberg Engineering, Germany) is another SLO platform
that is capable of UWFFA using a Staurenghi contact lens, which is generally
not tolerated in children, or by using a noncontact lens attachment, which can
be successfully used in children (20). The Spectralis with Flex module is a
customized device mounted on an armature that allows recumbent imaging
although with a small field of view.
Although FA is useful in the diagnosis and management of a wide range
of pediatric retinal diseases, it is especially valuable in retinal vascular
diseases. Several examples of the use of FA in specific pediatric retinal
conditions are presented below. Details of specific diseases may be found in
other chapters in this book.
Although FA is not a routine part of retinopathy of prematurity (ROP)
screening, it has been used in evaluating ROP infants for 50 years (21). FA
does increase the sensitivity and specificity of staging by helping to identify
neovascularization and the vascular–avascular junction not visible on the
fundus view (Figure 17-2) and to sometimes distinguish treated ROP from
the progressive nature of FEVR in cases of clinical ambiguity (22,23). FA in
FEVR shows irregular spouts of vascularization with progressive ischemia,
whereas eyes with classic ROP have a more homogeneous vascular edge that
resolves after adequate treatment (23). It is important to distinguish ROP
from FEVR and/or recognize when the two entities coexist in the same
patient, coined ROPER, to follow patients at appropriate intervals (Table 17-
2).
Table 17-2 Stages of pediatric retinal diseases with

corresponding clinical and fluorescein
angiography (FA) findings
ROP, retinopathy of prematurity; FEVR, familial exudative vitreoretinopathy; DR, diabetic retinopathy;
SS, sickle cell.
In severe ROP, at the vascular–avascular junction, FA can be used to

document abnormal branching of vessels. Hyperfluorescent findings include
perivascular leakage, retinal neovascularization, and isolated vascular tufts
posterior to the ridge that represent popcorn (24). At the posterior pole, areas
of hypofluorescence can be observed due to retinal and choroidal vascular
filling defects (24). Aggressive posterior ROP (AP-ROP) is a severe subtype
of ROP that was added to the International Classification of ROP in 2005
(25). It clinically appears as zone 1 disease with prominent plus disease and
poorly defined features at the edge of retinal vascularization. FA can be
useful to distinguish AP-ROP through the severe capillary loss throughout
the vascularized retina, shunting within the vascularized retina, abnormal
vascular branching, and a circumferential demarcation line (26,27).
FA has contributed to our understanding of the effects and limitations of
ROP treatments. FA reveals in sharp contrast the extent of retinochoroidal
atrophy after laser photocoagulation of the avascular retina. FA after anti-
VEGF treatments for ROP can reveal irregular retinal branching patterns,
abnormal tangles of vessels, persistent arteriovenous shunting, persistent
leakage at the original site of stage 3 disease, and late reactivation with new
areas of neovascularization (Figure 17-2) (28,29). FA has demonstrated the
effects of anti-VEGF agents to modify the natural course of ROP through
vascular remodeling and lack of complete retinal vascularization to the ora
serrata (29–32). Even in older children, untreated regressed ROP may only be
recognized through residual peripheral nonperfusion visible only with
UWFFA (Figure 17-3).
Coats Disease
Coats disease typically presents in school-age children with strabismus,
leukocoria, or failed eye screening. The disease is idiopathic and affects
young boys. Findings include unilateral vascular telangiectasia, nonperfusion,
and exudation that can even lead to macular fibrosis. Although the exudation
is often visible without FA, noncontact Optos UWFFA provides typical
findings that can aid in differentiating Coats from other diseases and assess
the severity and extent of retinal involvement in very young children in the
clinic setting. In younger or noncooperative children who require an EUA,
and during an EUA at the time of treatment, RetCam FA is a valuable tool for
this same assessment.
The typical appearance of active disease on FA is hyperfluorescence of
telangiectasias, hypofluorescence of exudates, and “light bulb” dilation of the
aneurysmal vessels (Figure 17-8). The vessels often show early and
persistent leakage, which leads to exudation (Table 17-2). In stage 3 disease
with retinal detachment, subretinal exudation, and dilated retinal vasculature,
retinovascular findings on FA can help differentiate between Coats disease
and retinoblastoma.
Figure 17-8 In a 7-year-old boy with Coats disease, UWF
fundus imaging (A) demonstrates exudates in the posterior
pole and foveal center and telangiectatic vessels, and
UWFFA (B) demonstrates early blockage from exudation,
peripheral nonperfusion and leakage, telangiectasias, and
tortuous vessels in the temporal retina.
FA is helpful to guide laser treatment to nonperfused retina and assess

whether telangiectatic vessels are adequately treated (33). Traditionally
thought of as a unilateral disease, recent FA studies have found higher rates
of subclinical nonperfusion and capillary telangiectasia in fellow eyes than
previously reported (34,35). This suggests that the pathogenesis of Coats
disease may be due to a yet unknown genetic or systemic association.
Familial Exudative Vitreoretinopathy

FEVR, although variable, typically presents with temporal
neovascularization, nonperfusion, and dragging and can progress to tractional
retinal detachment. There are an ever-increasing number of genes associated
with FEVR, but the percentage of patients who test positive on genetic testing
is typically <50% (36). Because findings can be subclinical, the extent of
involvement tends to be asymmetrical, and there is variable penetrance within
a family, FA can be valuable in identifying disease in infants and children.
For these reasons and because the most common inheritance pattern of FEVR
is autosomal dominant, FA is also useful to screen the parents. Over 50% of
asymptomatic family members with confirmed genetic diagnosis have been
shown to have early stage disease on FA (37).
FA was instrumental in showing that the pathogenesis of disease was
from incomplete peripheral retinal vascularization followed by
neovascularization and its sequelae (38). FA continues to be an essential tool
for diagnosis, staging, and follow-up of FEVR. Typical retinal vascular
findings include peripheral capillary nonperfusion, peripheral telangiectasias,
microaneurysms, neovascularization, and late-phase angiographic posterior
and peripheral vascular leakage (LAPPEL) (39). Commonly, the temporal
peripheral quadrant is most involved with a V-shaped area of nonperfusion
(40). Early stages can be missed on clinical exam or color fundus
photography alone, and the extent of nonperfusion is most readily appreciated
with FA (Table 17-2). Fluorescein leakage from bulbous vascular endings or
telangiectasias at the vascular–avascular junction indicates active progressive
disease.
As with other diseases that manifest peripheral nonperfusion, FA can
guide targeted laser treatment and avoid under treatment. Given the lifelong
risk for progression, serial FA on follow-up is recommended to look for
ischemia from retinal detachment (41). In general, children and adolescents
who present with disease may have a more serious course, and if there is no
progression or symptoms by 20 years old, it is believed the disease may be
more likely to remain stable.
Incontinentia pigmenti (IP), also known as Bloch-Sulzberger syndrome, is an
X-linked dominant syndrome affecting the skin, hair, teeth, eye, and central
nervous system.
The pathophysiology of IP is elucidated to be dysregulation of
eosinophils causing endothelial cell dysfunction (42). It is presumed that this
mechanism of increased inflammation leads to vaso-occlusion and ischemia
in the retina, which may vary between the eyes due to genetic mosaicism.
Retinal findings, similar to FEVR, include peripheral retinal
neovascularization and nonperfusion, hemorrhages, and exudative and
tractional retinal detachment (43). Avascular retina should be treated with
laser photocoagulation or cryotherapy. FA is used to detect areas of
peripheral retinal nonperfusion, neovascularization, enlargement in the foveal
avascular zone, and arteriovenous shunts. UWFFA imaging in clinic without
sedation not only reveals the extent of avascular retina and neovascularization
(Figure 17-9A), but is useful to monitor the response to treatment (Figure
17-9B).
Figure 17-9 UWFFA demonstrates periphery
nonperfusion and leakage consistent with
neovascularization before (A) and after (B) targeted retinal
photocoagulation in the left eye in a 14-year-old girl with
incontinentia pigmenti syndrome.
Uveitis
Uveitis can be a challenging disease to evaluate and treat in pediatric patients.
Posterior uveitis constitutes 40% of all pediatric uveitis compared to 20% in
adults (44). Etiologies can be categorized as infectious (i.e., toxocariasis,
toxoplasmosis, herpetic, cytomegalovirus, tuberculosis, Lyme) or
noninfectious (i.e., juvenile rheumatoid arthritis, pars planitis, multiple
sclerosis, Blau syndrome, sarcoidosis, Vogt-Koyanagi-Harada (VKH)
syndrome) (see also Chapters 47 and 48). FA helps in demarcating the areas
of capillary nonperfusion seen in tuberculosis. In VKH syndrome, early
choroidal hypofluorescence is a typical finding wherein initial pinpoint
hyperfluorescent dots are seen along with late pooling of the dye in the
subretinal space (45). FA can detect the characteristic “petaloid” pattern of
parafoveal hyperfluorescence in uveitic cystoid macular edema, retinal
vasculitis, periphlebitis, neovascularization, and staining of the optic nerve
head. Up to 90% of patients with intermediate uveitis have peripheral retinal
vasculitis (46). Often the extent of retinal vascular inflammation is difficult to
gauge on clinical examination alone and can be limited by media opacities. In
addition, vascular sheathing can persist longer than active inflammation. FA
has been used to monitor response to treatment and modify systemic, local, or
surgical therapy accordingly.
Diabetic Retinopathy
Diabetic retinopathy (DR) in children with type 1 diabetes mellitus typically
develops during puberty or in the mid-teen years (47). Recently, type 2
diabetes mellitus has become increasingly common in obese teenagers with
poor dietary habits, sedentary lifestyles, and at-risk family histories (48). The
utility of FA in children with DR is similar to its role in the management of
adults with DR. FA is able to stage and reveal the extent of nonperfusion,
neovascularization, microaneurysms, and macular leakage better than clinical
exam or color fundus photography alone (49,50).
Sickle Cell Retinopathy

Sickle cell retinopathy is an ocular manifestation of sickle cell disease, an
inherited group of hemoglobinopathies with several systemic morbidities that
predominately occurs in African Americans. The pathogenesis of sickle cell
retinopathy is decreased vascular flow and occlusions in the macula and/or
periphery due to abnormal red blood cell shape within retinal vessels.
Recently, with the routine use of OCT for screening, there has been more
attention on macular ischemia with subsequent retinal thinning,
predominately in the outer retinal layers, which can occur without symptoms
in sickle cell patients (51). FA is essential in classical peripheral staging of
disease and detecting the extent of peripheral nonperfusion, distinguishing
active neovascularization from involuted sea fans, and guiding treatment
(Table 17-2).
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18
Ultrasonographic Imaging in Infants
and Children
Roger P. Harrie
TECHNOLOGY
The fundamental principle of ultrasonography is based on the ability to
discriminate among captured reflections from a high-frequency sound beam
that is focused at a tissue and reflected from different tissue interfaces.
Ultrasound is generally defined as sound waves generated and reflected at
a frequency beyond the range of human hearing (20 kHz). Impedance is
defined as sound velocity times tissue density (Z = v × d). It is the difference
in impedance between different tissues that is responsible for the strength of
the reflected sound waves.
An extremely thin ceramic crystal membrane in the tip of the probe
undergoes a mechanical vibration when stimulated by an electric current,
which generates a sound beam. This same membrane acts as a receiver of the
reflected sound waves (the piezoelectric effect). These signals are amplified
and displayed onto a screen as either vertical lines in the A-scan or as a
coalescence of bright dots in the B-scan. The A-scan probe generates sound
waves in a linear direction with reflection of the focused beam from a focal
area, whereas the B-scan transducer oscillates about 25 times a second in a
plane and generates a sound beam, which is reflected from a 60-degree sector
of the globe. All B-scan probes have a mark on them to indicate the direction
of the oscillation. The frequencies used most commonly in clinical
ophthalmology are 8 MHz for the diagnostic A-scan and 10 MHz for the B-
scan but can be as high as 60 MHz in clinical ultrasound biomicroscopy. The
resolution of the image is dependent on the frequency of the generated and
reflected sound beam. Generally, higher frequency ultrasound provides
greater resolution of images but less penetration. Therefore, 40 to 50 MHz is
generally used in ophthalmology for anterior segment imaging, whereas 10 to
15 MHz is used for posterior segment imaging. Digital ultrasound images are
commonly uploaded to a server and used as reference for follow-up
examinations. It is important to record the orientation of the probe (see
below) with the image.
CONSIDERATIONS FOR USE OF

ULTRASOUND IN INFANTS AND
CHILDREN
The instrumentation used for echography in children is the same as that used
for adults. However, the examination techniques must be modified for this
specific patient population. Children can often be coaxed through the
examination, so it is important to explain the procedure and reassure the child
that the eyes can be closed during the examination. Since children can be
bothered by the sensation of the methylcellulose on the eyelid, it is helpful to
put a dab of it onto their fingertips to reduce anxiety. The examination is
performed through closed lids with gentle pressure. The room lights are
dimmed, and the child is held on the parent’s lap facing forward while the
exam chair is titled back so the child is horizontal. For infants, the examiner
assumes a position at the side of the chair, and the parent’s arm is placed on
the armrest adjacent to the examiner. The child’s head is rested on the
parent’s arm, while the other arm of the parent is used to secure the child.
Anesthetic drops are instilled in the eye in the event that it opens during the
examination. For uncooperative children, the parent or an assistant can be
asked to restrain the child for a quick screening examination. If this does not
provide sufficient information, then an examination under anesthesia can be
performed. This permits an in-depth ultrasound examination with selected
additional studies such as fundus photography and fluorescein angiography,
electrophysiologic testing, and spectral domain optical coherence
tomography. Immersion echography of the anterior segment is most easily
done while performed under anesthesia but can be done in the clinic through
closed lids with a water-filled cover over the tip of the probe. Methods to
accomplish this include fitting the finger of a sterile glove or a Tono-Pen
cover over the probe. Some companies offer commercial probe covers. In-
office sedation with appropriate monitoring has also been used (1).
Some of the benefits of echography in the clinic include the evaluation of
eyes with leukocoria to screen for serious problems such as retinoblastoma or
retinal detachment. It is also useful in cases of vitreous hemorrhage to
identify a retinal tear and to identify and localize a foreign body in cases of
penetrating trauma.
EXAMINATION WITH
ULTRASONOGRAPHY
The globe is first examined with the probe at a transverse position in which
the mark on the probe is parallel to the limbus (Fig. 18-1). Six quadrants are
evaluated starting at the 6:00 position with the probe directed superiorly. This
allows for evaluation of the superior 60 degrees (lateral to medial) segment of
the fundus. The probe is pointed directly at the posterior pole and angled as
far superiorly as possible. This process is repeated by placing the probe at the
4:00 position to image the superior temporal fundus; then at the 2:00 position
to image the inferior temporal fundus; then at the 12:00 position to image the
inferior fundus; then at the 10:00 position, which images the inferior nasal
fundus; and finally at the 8:00 position to image the superior nasal fundus.
The probe is then placed with the mark perpendicular to the limbus and is
placed at the same six positions on the globe to obtain a longitudinal view as
the transducer sweeps from the anterior to the posterior fundus. This allows a
more peripheral view that extends almost to the ciliary body. This technique
of probe positioning permits visualization of the entire posterior segment
from the optic disc to the pars plana. While ultrasound is useful to document
findings, it is also important to communicate the orientation of the probe
relative to the globe when documenting findings. In the case of measuring
tumor thickness, it is important to use standardized scans.
FIGURE 18-1 B-scan probe positions. Transverse
position with transducer moving parallel to limbus as
indicated by mark on probe directed vertically (left).
Longitudinal position with transducer moving
perpendicular to limbus as indicated by mark on probe
directed horizontally (right).
BENEFITS OF ULTRASONOGRAPHY IN
INFANTS AND CHILDREN
Echography can be very helpful in a number of pediatric vitreoretinal
conditions especially when there are media opacities such as corneal scars,
cataracts, anterior chamber hyphema, and vitreous opacities. It is very useful
in the detection of retinal detachment; characterization of visible fundus
lesions such as retinoblastoma or other tumors; identification of intraocular
foreign bodies; and biometry in congenital glaucoma, anisometropic
amblyopia, and unilateral persistent fetal vasculature (PFV) in which axial
length differences between eyes can be determined. It is also useful in
differentiating causes of microphthalmia, such as PFV versus congenital
microphthalmos with a coloboma or cyst.
Many of the above conditions present with a white pupil or leukocoria,
which is the most common indication for ophthalmic ultrasonography in the
pediatric population (2). The most serious cause of leukocoria is
retinoblastoma, which is potentially life-threatening if not promptly
diagnosed and treated.
UTILITY OF ULTRASOUND IMAGING

IN LEUKOCORIA
Retinoblastoma
Echography can be very useful in the evaluation of retinoblastoma, wherein
95% of these tumors have characteristic ultrasonographic findings, and the
atypical 5% create clinical challenges.
Retinoblastoma should always be suspected in a child with an intraocular
mass lesion. Most of these tumors occur in children under the age of 3, but
rare cases occur even in adults (see also Chapters 44 and 45).
Retinoblastomas can occur in an endophytic (growth into the vitreous) or an
exophytic form (growth under the retina), but some tumors display elements
of both growth patterns. A majority (90% to 95%) of these lesions have
calcium deposits ranging from those seen only microscopically to those
easily detectable on imaging studies. Ultrasound is very sensitive in the
detection of calcium (Fig. 18-2), whereas MRI is not. Fine calcium deposits
can be missed on CT, and there is concern about radiation exposure in
children. A less common form of retinoblastoma is the infiltrating type that
grows within the retina and presents as nonspecific retinal thickening (Fig.
18-3). Diffuse infiltrating retinoblastoma occurs most often in older children
and does not contain calcium (see Chapters 44 and 45). A fundus mass in a
child is always presumed to be retinoblastoma until proven otherwise. Most
retinoblastomas are found in the posterior segment of the eye but can rarely
be found in the peripheral retina. These can mimic a chronic inflammatory
process and be confused with intermediate uveitis or pars planitis (Fig. 18-4).
The anterior location is best visualized using immersion ultrasound. Higher-
resolution probes (20 to 50 MHz) show greater detail than a standard 10-
MHz probe, but the 10-MHz probe permits the display of the entire extent of
the tumor and its relationship to adjacent structures.
FIGURE 18-2 B-scan of retinoblastoma with diffuse fine

calcium deposits (arrow). As the gain on the B-scan is
reduced, the high reflectivity of the calcium will remain
bright.
FIGURE 18-3 B-scan of diffuse retinoblastoma (arrows).
This lesion is not seen on ultrasound as a discrete tumor
mass but is more irregular and extends underneath the
retina. No calcium is detected.
FIGURE 18-4 B-scan of retinoblastoma simulating pars
planitis (arrow). The B-scan probe on the temporal globe
is angled very anteriorly to visualize this lesion at the pars
plana.
When retinoblastomas spread outside the globe, it is generally by invasion of

the optic nerve. Echography is usually unhelpful in this situation, because
microscopic invasion of the optic nerve cannot be detected even though the
retrobulbar optic nerve can be visualized by the B-scan and the thickness can
be measured very accurately by the A-scan. MRI can more reliably detect
invasion of the optic nerve, but false-negative and false-positive results have
been reported (3). Usually both modalities are recommended to rule out optic
nerve involvement. MRI can detect trilateral retinoblastoma in which the
pineal gland is involved (see also Chapters 44 and 45).
PERSISTENT FETAL VASCULATURE

In a series of 227 cases presenting with leukocoria that clinically simulated
retinoblastoma, PFV (see Chapter 65) was most commonly misdiagnosed as
retinoblastoma (4). PFV can appear in several forms. Ultrasonography can be
used to distinguish anterior PFV from posterior PFV, although many patients
can have a combination of both. PFV is unilateral in over 90% of cases but
when bilateral carries a poorer visual prognosis. Axial length is generally
shorter in eyes with PFV than in the normal contralateral eye by 0.5 to 1.0
mm, and this can be a helpful differential finding as measured by A-scan. The
ultrasound can also help to differentiate PFV with stalk from an eye with
colobomatous microphthalmia with or without cyst. Findings in the posterior
variant of PFV include vitreous membranes, a stalk (fibrovascular remnants
in Cloquet canal) extending from the optic nerve to the posterior lens capsule,
optic nerve dysplasia, and retinal folds. Imaging of the stalk by echography
can be difficult and may require different B-scan probe positions at high gain
to demonstrate it. A transverse view with the probe held parallel to the limbus
will cut across the stalk making it appear as a small echodensity. A
longitudinal view with the probe perpendicular to the limbus demonstrates
the stalk as an elongated echodensity as the sound beam is reflected along its
length (Fig. 18-5). Sometimes, vitreous traction on the retina can cause a
focal or total retinal detachment.
FIGURE 18-5 B-scan of PFV with vitreous stalk (arrow).
Vascularity was detected as rapid spontaneous flickering
movements inside the stalk during the examination.
Findings in anterior PFV include a shallow anterior chamber, elongated

ciliary processes, cataract, and a retrolental fibrovascular membrane (Fig. 18-
6) responsible for the appearance of leukocoria (5). These are best
demonstrated with an immersion B-scan and may require a high-frequency
probe. The retrolental tissue tends to increase in volume and may contract
and pull ciliary processes inward with increased age. This process can
simulate a growing mass lesion such as a medulloepithelioma (Fig. 18-7).
Medulloepithelioma (diktyoma) is a rare tumor in early childhood that
usually arises from the medullary epithelium of the ciliary body and is best
seen with an immersion scan (Fig. 18-8). Its echographic appearance is
somewhat variable corresponding to the different pathologic presentations,
which include the nonteratoid type that is a dense proliferation of cells and is,
therefore, low to medium reflective. The teratoid form may have a variety of
tissue elements including skeletal muscle, cartilage, and cysts filled with
hyaluronic acid and result in heterogeneous reflectivity.
FIGURE 18-6 Immersion B-scan of PFV (first arrow

cornea, second arrow iris, third arrow retrolental fibrosis).
The retrolental opacity in this case is dense, whereas in
some cases, it is not evident but the posterior lens capsule
may demonstrate irregularity due to the contraction of a
thin retrolental membrane.
FIGURE 18-7 Immersion B-scan of PFV simulating a
diktyoma (arrow). This lesion was documented to grow
over several months.
FIGURE 18-8 Immersion B-scan of diktyoma of the
ciliary body (first arrow cornea, second arrow tumor).
Coats Disease
Coats disease is the second most common condition to cause diagnostic
confusion with retinoblastoma as compared to the other entities associated
with leukocoria. This typically unilateral condition is most common in
children with an average age of about 7 years and occurs in males versus
females at a ratio of 2.5:1. The subretinal space can be filled with lipid
including cholesterol crystals with leakage of serum from telangiectatic
vessels. In advanced cases, there can be slow convection movement of
subretinal exudate on ultrasound. In cases of Coats with marked subretinal
exudation, ultrasonography can produce an image simulating a
retinoblastoma with diffuse punctate high reflectivity from subretinal
cholesterol clefts (Fig. 18-9).
FIGURE 18-9 B-scan of Coats disease with a mass

appearance and fine calcium-like opacities (arrows). Such
a case is difficult to distinguish from retinoblastoma.
Ocular Toxocariasis
Ocular toxocariasis is the third most common condition incorrectly diagnosed
as retinoblastoma (6). The parasitic roundworms Toxocara canis or Toxocara
cati can migrate into the posterior segment usually in children or young
adults and cause an isolated fundus lesion or diffuse vitreous inflammation
presenting as clinical leukocoria. This process presents with a focal
peripheral retinal granuloma in 50% of cases, a macular lesion in 25%, and
moderate to severe vitreous inflammation in 25% (6). In some cases with a
peripheral lesion, there may be a vitreous membrane extending from the
lesion to the posterior fundus (Fig. 18-10), and there may be a posterior
retinal fold or traction detachment. Echography is useful in demonstrating
these findings in the presence of media opacity from vitreous inflammation or
hemorrhage. These findings may mimic familial exudative vitreoretinopathy
(FEVR) and retinopathy of prematurity (ROP), and, therefore, a history can
be useful.
FIGURE 18-10 B-scan of toxocara with a vitreous

membrane attaching at a granuloma of the fundus (arrow).
ROP is another entity in the differential diagnosis of leukocoria (7).
Ultrasound is useful in detecting a detached retina particularly when
leukocoria is present. The peripheral fibrovascular tissue can contract and
detach the peripheral retina with the resultant echographic appearance of
loops of retina (Fig. 18-11) or in total retinal detachment as a tight funnel
(Fig. 18-12). In the later stages of ROP (4A, 4B, and 5) with partial or
complete tractional retinal detachment, ultrasonography is useful in assessing
the diameter of the detached retinal funnel which may be beneficial in
surgical planning (8).
FIGURE 18-11 B-scan of ROP with anterior loops of

detached retina (arrows). This is a very peripheral view
with the probe placed on the temporal sclera and angled
very anteriorly to view the anterior traction retinal
detachment.
FIGURE 18-12 B-scan of total retinal detachment in a
tightly closed funnel (arrow).
Retinal Detachment
Retinal detachment may itself cause leukocoria, or it can accompany other
causes of a white pupil. The echographic criteria for the diagnosis of a
detached retina include a highly reflective membrane attaching at the optic
nerve head (Fig. 18-13), a membrane demonstrating high reflectivity in the
periphery, and B-scan mobility characteristics of somewhat stiff and tethered
after movements compared to the loose and wider excursions of a vitreous
membrane. Therefore, dynamic echography is important in the diagnosis.
Retinal detachments are generally more highly reflective on A-scan than
vitreous membranes (Fig. 18-14).
FIGURE 18-13 B-scan of retinal detachment (arrows).
The highly reflective membrane with an attachment at the
optic nerve head is usually diagnostic of a retinal
detachment, but a vitreous membrane can occasionally
have this appearance.
FIGURE 18-14 A-scan of retinal detachment (second
arrow). The reflectivity of the retinal spike is at the same
height as the initial spike (first arrow) and the scleral spike
(third arrow) as opposed to vitreous membranes, which
are not as highly reflective.
The disease processes discussed to this point account for almost 70% of the
conditions reported to simulate retinoblastoma (4,9). Other less common
causes of leukocoria include Norrie disease (see Chapter 35) and FEVR (see
Chapter 66), which may have the echographic findings of radial retinal folds
and/or an exudative retinal detachment (Fig. 18-15).
FIGURE 18-15 B-scan of FEVR showing marked
subretinal exudate with a detached retina (arrows).
Utility of Ultrasound in the Evaluation after Ocular

Trauma
After leukocoria, ocular trauma is the next most common indication for the
performance of ultrasound in children. It is estimated that 2.4 million eye
injuries occur each year in the United States, and 35% of these occur in
persons 17 years old or less (10) (see also Chapter 69). Blunt trauma is the
most common type of injury in this group. Ultrasound often plays a critical
role in the evaluation of these children when a view of the posterior segment
is obscured by media opacities such as cataract, hyphema, or vitreous
hemorrhage. While it is especially important to determine if the globe has
been ruptured prior to surgical procedures, ultrasound is often not appropriate
in the presence of a ruptured globe due to the risk for applying pressure
through the lids and onto the open globe. However, many ruptures are occult
and not recognized clinically as the intraocular pressure may be normal.
Echographic findings of choroidal or subretinal hemorrhage, focal scleral
irregularity, or vitreous membranes inserting at a specific location in the
posterior segment (Fig. 18-16) may indicate a rupture. The presence of an
intraocular foreign body should always be suspected when a child presents
with unexplained intraocular inflammation even if a history suggestive of a
projectile possibly striking the eye is absent (Fig. 18-17). It is not uncommon
for a child to have an intraocular foreign body and deny any kind of activity
that may have caused it (11). The clinician should have a low threshold of
suspicion.
FIGURE 18-16 Scleral rupture with vitreous incarceration

(arrow). The actual site of the scleral rupture is often not
detectable on echography since the adjacent choroidal and
scleral edema will seal the rupture, but the insertion of
vitreous membranes can “point” to the site as in this case.
FIGURE 18-17 Intraocular foreign body (arrow). The
high reflectivity of the foreign body will be maintained
even as the gain is reduced and the probe is angled
obliquely to the foreign body.
Choroidal Effusion
Ultrasonography is also useful in revealing fluid in the suprachoroidal space
and in differentiating posttraumatic choroidal hemorrhage or serous fluid
seen in choroidal effusion (12)
Limitations
Limitations of echography include replication of the scan. Because
ultrasound is captured in two-dimensional scans, this cross-sectional view (as
opposed to a three-dimensional view) may lead to misinterpretation of the
image. It may also be difficult in complex vitreous hemorrhage, to distinguish
blood or organized membranes from retinal detachment. Shadowing from
anterior structures may also limit a deeper view. Because the probe may
transmit pressure to the globe, it has limitations for use in trauma when there
is a possible open globe. The value of the information obtained from
echography is a function of the experience of the examiner. Subtle diagnostic
cues can be missed by the less experienced clinician.
CONCLUSION
Despite the limitations, echography is a useful ancillary test in the pediatric
population. Many types of intraocular pathology can be detected and often be
accurately diagnosed by ultrasound.
REFERENCES
1. Karaoui M, Varadarai V, Munoz B, et al. Chloral hydrate administered by a dedicated sedation
service can be used safely and effectively for pediatric ophthalmic examination. Am J
Ophthalmol 2018;192:39–46.
2. Harrie RP. Clinical ocular ultrasonography: a case study approach. New York: Springer, 2008.
3. Brisse HJ, Guesmi M, Aerts I, et al. Relevance of CT and MRI in retinoblastoma for the
diagnosis of postlaminar invasion with normal-size optic nerve: a retrospective study of 150
patients with histological comparison. Pediatr Radiol 2007;7(7):649–956.
4. Shields JA, Parsons HM, Shields CL, et al. Lesions simulating retinoblastoma. J Pediatr
Ophthalmol Strabismus 1991;28:338–340.
5. Castillo M, Wallace DK, Suresh MK. Persistent hyperplastic primary vitreous involving the
anterior eye. Am J Neuroradiol 1997;18:1526–1528.
6. Stewart JM, Cubillan LDP, Cunningham ET. Prevalence, clinical features and causes of visual
loss among patients with ocular toxocariasis. Retina 2005;25:1005–1013.
7. Fiedler AR, Shaw DE, Robinson J, et al. Natural history of retinopathy of prematurity; a
prospective study. Eye 1996; 6(Pt3):233–242.
8. Jabbour NM, Eller AE, Hirose T, et al. Stage 5 retinopathy of prematurity. Prognostic value of
morphologic findings. Ophthalmology 1987;94(12):1640–1646.
9. Chuah CT, Lim MC, Seah Y, et al. Pseudoretinoblastoma in enucleated eyes of Asian patients.
Singapore Med J 2006;47(7):617–620.
10. Brophy M, Sinclair SA, Hosteler SG, et al. Pediatric eye injury-related hospitalizations in the
united states. Pediatrics 2006;117(6):1263–1271.
11. Yeh S, Colyer MH, Weichel ED. Current trends in the management of intraocular foreign
bodies. Curr Opin Ophthalmol 2008;19(3):225–233.
12. Wirold J, Orlowski-Szczypinski J. Ultrasonography of the subretinal space in rhegmatogeneous
retinal detachment. Mod Probl Ophthalmol 1976;18:40.
19
Imaging Analysis in Infants and
Children
Sang Jin Kim, John Peter Campbell, and Michael F. Chiang
Advances in retinal image analysis techniques have followed the dramatic

rise in retinal imaging in adults and children over the last 20 years. From
measuring area of geographic atrophy in age-related macular degeneration to
quantifying nonperfusion in patients with retinal vascular disease to
automated detection of diabetic retinopathy using deep learning, several
motivating factors have spurred research into computer-based image analysis.
Automated image analysis technology can be used autonomously, in lieu of
physicians, or as a diagnostic assistive device providing the following
potential advantages to subjective interpretation of retinal images. First,
automated methods can provide quantitative metrics of disease severity that
can be used to screen, diagnose, and monitor disease progression. As
ophthalmology historically has been an image-based specialty with diagnosis
based on subjective pattern recognition of anatomic disease features for some
diseases, automated image analysis techniques can be employed to
complement the clinical exam and measure disease severity objectively and
quantitatively. Second, the automated methods can also improve efficiency of
image interpretation in clinical practice, telemedicine, and reading centers.
For example, given the sheer volume of images that need to be read in a
telemedicine system, automated image analysis could provide a preliminary
diagnosis and triage those with more severe disease for urgent clinical
evaluation. Third, deep learning has demonstrated the potential to detect
relevant clinical and demographic information from retinal images that is not
apparent to human graders. This has the potential to improve our
understanding of disease physiology and improve risk modeling, thereby
providing a better understanding of the relationship between retinal and
systemic disease. Image analysis techniques have been applied to a number
of different retinal imaging techniques (fundus photos, fluorescein
angiography [FA], optical coherence tomography [OCT], and OCT
angiography). In pediatric retina, most of the work has focused on the
problem of quantifying the vascular changes in retinopathy of prematurity
(ROP) (1–6). This chapter will focus on the application of image analysis to
plus disease in ROP as a case study of the potential role image analysis can
play in improving care for children with retinal disease.
CHANGING PARADIGMS IN
DIAGNOSIS
Traditionally, diagnosis of ROP is made by ophthalmoscopy in the neonatal
intensive care unit (NICU), although a number of factors contribute to the
rising use of telemedicine for primary ROP screening (7). More than 80% of
infants in an ROP screening program in the United States have less than
severe ROP and could safely have deferred ophthalmoscopic examinations
(8,9). The number of clinicians who are trained and willing to manage
patients with ROP is decreasing in the United States for a number of reasons,
including high medicolegal liability, inadequate training, and poor
reimbursement (10,11). ROP is increasingly becoming a disease of low- and
middle-income countries where the population at risk is higher, and the
human resources are becoming more scarce (12).
Screening guidelines now recognize telemedical interpretation of wide-
field ROP images as reasonable with appropriate safeguards to ensure quality
control and follow-up (7). In both ophthalmoscopic examination and
telemedicine, clinical diagnosis is based on parameters defined by the
international classification of ROP (ICROP): zone, stage, clock hour extent,
and plus disease (13,14). The NIH-sponsored multicenter cryotherapy for
ROP (CRYO-ROP) (9,15) and early treatment for ROP (ETROP) trials
determined that the presence of plus disease, defined as venous dilatation and
arteriolar tortuosity in central retinal vessels greater than or equal to that of a
standard published photograph, is the most important factor in identifying
infants with severe treatment-requiring disease at risk for blindness (14).
Therefore, it is critical to accurately diagnose plus disease to determine the
need for treatment in ROP.
CHALLENGES IN PLUS DISEASE

DIAGNOSIS
There are several lines of evidence that suggest the diagnosis of plus disease
may be made inconsistently in the real world. Multiple investigators have
demonstrated variability in plus disease even in NIH-funded multicenter trials
(4,16–20). For example, even as far back as the CRYO-ROP protocol,
unmasked clinical collaborators disagreed on the diagnosis of threshold ROP
12% of the time. Since threshold disease required accurate diagnosis of stage
3 ROP and plus disease, some of the disagreement may have been from
diagnosing either parameter (20). In the e-ROP study, nearly 25% of
examinations by certified study graders required adjudication because the
graders disagreed on one of three criteria for clinically significant ROP: zone
I ROP, stage 3 or worse ROP or plus disease (16). Furthermore, the
diagnostic reasoning used by examiners has been variable and often deviated
from the traditional ICROP definition of plus disease (4,21). The difference
in the magnification and field of view of indirect ophthalmoscopy and wide-
field imaging compared to the published standard photograph for plus disease
from the 1980s has been shown to cause bias and inconsistency in diagnosis
(3,22–24). There is also evidence of variation in the diagnosis of plus disease
based on geographic location throughout the world (19) and the change in
clinical diagnosis of plus disease over time (25). Historically, plus disease
was a binary disease characteristic (present or not), but the revised ICROP
“preplus category” entered another parameter that began to suggest some of
the difficulty with a discrete binary diagnosis of plus disease including
continuing evolution in vascular tortuosity and dilation from normal vessels
to those in severe ROP. The multicenter Supplemental Therapeutic Oxygen
for Prethreshold ROP (STOP-ROP) study added specificity to plus disease in
that vascular changes had to be present in at least 2 quadrants, and this
definition was incorporated into the 2005 revised ICROP in 2005 (14).
However, there is variability in how this definition is interpreted and
evidence that this variability may lead to clinically significant differences in
diagnosis (26). Studies add to significant levels of variability in the diagnosis
of preplus disease among experts participating in research (Fig. 19-1) (27).
Research suggests that diagnostic discrepancy results from individual
clinicians having different cut points (e.g., “is this plus or preplus disease”),
despite having better agreement on relative disease severity (e.g., “which
retina looks worse”) (5,27). Given the above challenges, there is a strong
suggestion that the diagnosis of plus disease and treatment recommendations
may vary widely in the real world despite international consensus on the
classification of ROP and evidence from multiple NIH-funded clinical trials
(19).
FIGURE 19-1 Representative examples of eyes with no

plus disease, preplus disease, and plus disease
demonstrating the range of severity that is captured in the
three-level scale. As plus disease runs a continuum from
normal to severe plus, there is evidence that differences in
“cut points” between the categories may be a key
underlying cause of interobserver differences in plus
disease diagnosis. (Figure courtesy of James Brown, PhD
and Jayashree Kalpathy-Cramer, PhD.)
INITIAL COMPUTER BASED IMAGING

ANALYSIS APPROACHES
Early attempts to use computers for quantitative image analysis of plus
disease used manual or semiautomated measurement specific operator-
defined features, such as tortuosity and dilation, from wide-angle fundus
photos (RetCam; Natus Medical Incorporated, Pleasanton, CA) (1). Three
such systems have been developed and validated for wide-angle RetCam
images: ROPTool (2), Retinal Image multiScale Analysis (RISA) (28), and
Computer-Assisted Image Analysis of the Retina (CAIAR) (29). These
systems have been evaluated against expert diagnostic performance but have
not had real-world application because of limitations, such as being
semiautomated (e.g., requiring manual identification of optic disc or key
vascular segments) or having limited correlation using two-level expert
diagnosis (e.g., plus disease vs. not plus). As an example, ROPTool has been
the most widely used of these methods clinically and is a semiautomated
program that calculates both dilation and tortuosity automatically after the
user identifies the optic nerve and a few key vessels (2,30). Compared to a
severity score based on expert gradings of vessel appearances, the area under
the operating characteristic curve (AUC) was 0.91 for ROPTool detection of
cases with some vascular abnormalities or greater versus controls (defined as
a score of 0) (30). The ROPTool dilation measurement has not been found to
correlate as closely with clinical disease (31,32). The ROPTool system is
currently being used in a commercially available screening software platform
(FocusROP; Trumbauersville, PA).
MACHINE LEARNING
Machine learning uses the processing power of a computer to “learn” patterns
from operator-defined features that best solve a particular problem (Fig. 19-
2). The Imaging and Informatics in ROP (I-ROP) consortium published the
results of the “i-ROP ASSIST” program in 2016 (3,33). Manually segmented
images from a trained grader, who manually traced vessels from raw images,
were used as inputs, and a machine learning strategy used support vector
machines and gaussian mixed models as methods to quantify and classify
features. The i-ROP ASSIST program used a vascular metric termed
“acceleration” that was found to have the best diagnostic performance in a 6-
disc-diameter circular crop of wide-angle RetCam images when all retinal
vessels combined were considered. With the real-world application of this
system being limited by the requirement of manual segmentation, the
performance was on par with clinical experts (3,33).
FIGURE 19-2 Schematic representation of machine
learning model. Traditional severity score models, for
example, the Early Treatment for Diabetic Retinopathy
Scale (ETDRS), utilize differentially weighted, operator-
defined features, such as microaneurysms and intraretinal
hemorrhages, to develop a scoring system that has some
prognostic significance. In a machine learning approach,
the features may be defined a priori, and the weights are
learned by the computer. In a deep learning program using
a convolutional neural network, no features are defined a
priori. (Figure courtesy of James Brown, PhD and
Jayashree Kalpathy-Cramer, PhD.)
This analysis further demonstrated that computer-based imaging analysis

(CBIA) has the ability to help us better understand the diagnostic reasoning
of clinicians, since it “reverse engineers” the problem by starting with the
“answer” (e.g., having the gold standard diagnosis of plus disease according
to experts) and identifying the features (e.g., tortuosity and dilation in various
locations of the retina and combinations of these individual quantitative
retinal features) that can best predict expert diagnosis. In this case, it
demonstrated that expert diagnosis can be best modeled when two
nonstandard features are incorporated into the calculation: venous tortuosity
and vascular information from outside the most posterior retina, which
suggested that experts may consciously or unconsciously use these features in
their diagnostic decision making (33). This is especially significant, because
if experts are asked what factors they use to make decisions, they often
disagree (21).
DEEP LEARNING
Recent advances in deep learning take this one step farther by removing any
defined features. Deep convolutional neural networks (CNN) are a form of
supervised machine learning that use only the input data, usually an image,
and the output, or the ground truth about the problem being solved, such as
the status of plus disease, and then iteratively learn the features that predict
the output without exploiting defined features through the parallel processing
power of the graphics processing unit (GPU) (34). Using raw images from
the i-ROP cohort study as the input, the “i-ROP DL” program was able to
classify plus disease with an AUC of 0.98, outperforming 7/8 experts (6). The
i-ROP DL system works by using two CNNs together; the first one produces
a vascular segmentation automatically (Fig. 19-3), and the second operates
only on the segmented image to classify disease.
FIGURE 19-3 Example of a vascular segmentation of a
retinal fundus photograph. This image was produced in the
first step of the Imaging and Informatics in ROP Deep
Learning (i-ROP DL) program that automatically
diagnosed plus disease based on fundus images. The
image was classified as plus by clinical experts.
CHALLENGES
There are several challenges to the application of CBIA in ROP: Humans
often disagree on ROP diagnosis (16–18,27) making the “gold standard” used
to train the CNN less robust. This generally raises the number of images
required to fully train a network. Operator-defined features or datasets may
be biased, and these biases may be conveyed through to CBIA. CNNs require
large amounts of data to train, potentially limiting the application of CNN-
based CBIA techniques in less common pediatric retinal diseases. The image
quality and field of view necessary for diagnosis are not defined. Therefore,
subtle differences in an input image, which have been shown to affect CNN
performance, need to be defined for clinical use. Similarly, it remains to be
proven that CNN outputs can be independent of the camera used to obtain an
image. Whereas CNNs are often quite powerful for image classification tasks,
the layers of the CNN are a “black box” in the sense that it is not easy to
unpack the layers of the CNN to explain how it is working. This lack of
transparency is an obstacle for some clinicians and regulatory bodies to
approve software programs for clinical use and is an area of active research
(35). One approach to unpacking the black box utilizes “heatmaps,” which
evaluate how much each part of an image contributes to the overall
classification of the image (Fig. 19-4).
FIGURE 19-4 Example of a heatmap image. The color

shadings reflect the contribution of each 8 × 8 pixel patch
to the overall classification of the convolutional neural
network for this image from very little contribution,
indicated by lighter values, to significant contribution,
indicated by darker values. (Figure courtesy of James
Brown, PhD and Jayashree Kalpathy-Cramer, PhD.)
FUTURE DIRECTIONS
Though the applications of CBIA in plus disease diagnosis have been the
most well studied, there is the potential to apply similar technology to the
diagnosis of other pediatric retinal diseases such as Coats disease,
incontinentia pigmenti (IP)-associated retinopathy, familial exudative
vitreoretinopathy, inherited retinal degenerations, retinoblastoma, and others.
In addition, though color fundus photographs have primarily been used for
studies of plus disease, CBIA of other images, such as OCT and FA, has also
been investigated (36). CBIA would bring the similar potential advantages,
including quantification of disease severity, automated and objective
diagnosis, standardization of diagnosis between clinicians, and incorporation
in tele-screening programs, to these other applications and modalities as it
does to fundus photographs in ROP. Deep learning may be applied to the
increasing number of approaches to newborn fundus screening, which has not
yet become standard of care in the United States but is being used
increasingly in other parts of the world. Finally, at this time, the numbers of
papers published on CBIA outweighs the numbers of patients who have truly
benefited from the implementation of these technologies into clinical
practice. More work is needed to translate the theoretical advantages of these
technologies into real-world impact for patients.
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20
Image Storage and Retrieval and
Telemedicine
Marco H. Ji, Darius M. Moshfeghi, and Antonio Capone Jr
Telemedicine is defined as the use of electronic information and

communications technologies to provide and support health care when
distance separates the participants (1). During the past decades, advances in
computing technologies have entirely revolutionized our lives, including
medicine. Advances in telecommunication and retinal imaging technologies
have allowed the development of retinal telemedicine that aims to facilitate
the evaluation, diagnosis, and management of the patients remotely. Since
2000, an increasing number of studies have demonstrated the feasibility of
telemedicine in retinopathy of prematurity (ROP), and clinical telemedicine
screening for ROP has increased (2).
ROP remains one of the leading causes of childhood blindness in the
United States and the rest of the developed world (3–6), despite the
availability of effective treatments (7–16). Most ROP cases are mild and tend
to regress, but around 1,100 to 1,500 babies progress to type 1 ROP requiring
treatment in the United States each year, with some progressing to blindness
despite treatment (17). In the United States, screening guidelines are updated
approximately every 5 years based upon a consensus statement from the
American Academy of Ophthalmology (AAO), the American Association for
Pediatric Ophthalmology and Strabismus (AAPOS), the American Academy
of Pediatrics (AAP), and the American Association of Certified Orthoptists
(AACO) (18–20). These screening guidelines are predicated upon the
International Classification of ROP (ICROP) (21–23). (See also Chapter 52)
Current guidelines are extremely inefficient because for the large screened
population, the risk of developing significant ROP is low for most of the
infants while missed cases of more advanced disease can lead to blindness
from a potentially preventable disease.
Evidence from malpractice cases shows that screening or referral failure
contributes to the number of premature infants who go blind from ROP. A
2009 analysis of 12 closed malpractice claims in ROP identified that the care
issues involved failure of transfer of care, inappropriate length of follow-up,
failure to supervise residents, and failure of outpatient referral (24). Most of
the medical care issues, in 13 ROP malpractice cases reviewed in 2007, were
also related to problems of screening (25). In neither series was negligent
treatment a finding or proximate cause of blindness (24,25). The neonatal
intensive care units (NICUs) face the challenge of arranging for screening to
occur, arranging for follow-up at back-transfer NICUs, and discharging
patients to the community; these challenges are made more difficult by
limited available local ROP screening (26,27). Therefore, it seems apparent
that efforts to minimize blindness from ROP should, in addition to new
therapy development, be focused upon effective screening (28).
There is a lack of efficacy data for ROP screening with binocular indirect
ophthalmoscopy (BIO) with a 28- or 30-diopter lens, although this has been
considered the gold standard a priori. Because the examination technique
does not provide a recorded objective measure, when there is disagreement,
one cannot determine which examiner is correct. For example, in the
Cryotherapy for Retinopathy of Prematurity (CRYO-ROP) study, 12% of
eyes that were initially thought to be threshold disease were overruled on
subsequent examination within 3 days by a separate examiner (29). The
authors noted that at least one of the examiners had to be in error, although
they speculated that the time delay between examinations could have
conceivably allowed for regression of disease, particularly in “borderline
cases” (29). Similarly, the consistent 4% interobserver disagreement rate in
the classification of zone III without ROP in the CRYO-ROP and LIGHT-
ROP trials indicates that at least one examiner was incorrect in these cases
(29). In the cases of disagreement in these studies, there was no objective
reference. Inter- and intraobserver variability is recognized and common in
ophthalmology in the setting of photographic interpretation (30–40), and
there are many reasons to believe that it is only exacerbated in the setting of
examination of a live patient (36,40). Although there are no data, it seems
probable that episodes of desaturation, bradycardia, and other issues during
difficult examinations of preterm infants would add further to the complexity
of the examination and increase inter- and intraobserver disagreement.
With respect to training ROP examiners, no formalized ROP training is
specified in the United States, nor is it clear who should be performing the
examinations (41–44). The Accreditation Council for Graduate Medical
Education (ACGME) has not stated a required number of ROP examinations
or treatments during training (45), and in 2012, 15% of pediatric
ophthalmology programs in the United States provided limited or no
exposure to ROP (43). Additionally, for training, there are inherent
limitations to the number of bedside BIO examinations that can be performed
in the preterm infant, and it can be quite difficult to achieve adequate
visualization for a trainee or other observer with the presently available video
binocular indirect ophthalmoscopes (43).
There also appears to be a potential shortage of ROP examiners. In an
AAO survey of self-identified pediatric ophthalmologists or retina specialists,
only 76% of those providing ROP screening were planning to continue
screening moving forward mostly due to financial reasons, with medical
liability, the higher cost of insurance for ophthalmologists who provide ROP
care, the complexity of scheduling care for children, and poor reimbursement
as factors (46). Notably, this has been occurring in the face of an increase in
the total ROP screening population of approximately 33% (47,48). Another
significant limitation of the current disease screening strategy is the extensive
travel and logistical coordination requirements for ophthalmologists and
neonatologists that make it extremely time-consuming and inefficient (49),
and that challenge is even more arduous in rural and underserved areas,
especially in developing countries (50). Thus, for the multiple reasons
described above, there is a very strong impetus for alternate means to screen
for ROP.
REMOTE DIGITAL FUNDUS IMAGING

Experimental Analysis of RDFI
The alternate ROP screening approach that has achieved greatest traction is
telemedicine using wide-angle remote digital fundus imaging (RDFI).
Originally proposed in 1999 (51), proof of concept was demonstrated in 2000
for use in ROP (52). The basic premise is a store-and-forward, hub-and-spoke
model of image management. Images are obtained locally in the NICU,
stored in the camera, uploaded to the Internet in a Health Insurance
Portability and Accountability Act (HIPAA)-compliant fashion, and then
received, interpreted, and adjudicated by the reader. The reader serves as the
hub, with each individual NICU representing a spoke in the RDFI network.
Unlike BIO, extensive evaluation of RDFI in ROP is possible and has been
performed (49,51–74). Studies have been grouped into the following areas:
(a) feasibility assessment (52,73), (b) retrospective comparisons, (c) retinal
disease component evaluation (e.g., Plus disease) (61,63,65,70,75), (d) reader
variation (56,60,67), (e) timing of evaluations (53,55,58), (f) retrospective
review of historical photographs (57,68), (g) contemporaneous prospective
studies (54,62,71,72,74), and (h) real-world experience (59,64,66,69).
In the first demonstration of the potential for RDFI screening in ROP,
expert review of a single 120-degree RetCam image was compared to
contemporaneous BIO for recommendations with respect to follow-up.
Expert reviewers of RDFI identified Plus disease in 95% of eyes described as
such at the bedside (52). The RDFI reviewers were also more conservative in
their interpretation of disease severity than were bedside examiners, making a
diagnosis of prethreshold in multiple eyes that bedside examiners had
diagnosed as threshold (52). Later, the important concept of “clinical
significance” was established, assigning this label to zone I or zone II ROP or
Plus disease (73). The investigators recognized the difficulty in identifying
ROP in zone III on RDFI but also noted that all disease there spontaneously
regressed, buttressing their concept of clinical significance (73). Highlighting
the hazards of lack of study surrounding BIO’s efficacy for ROP screening,
the remote RDFI readers identified zone II, stage 2 ROP eyes that had
initially been missed by BIO but were later confirmed at examination (73).
RDFI images were also shown to be safely and reproducibly obtained by a
trained neonatal nurse even though peripheral visualization was limited (53).
As in earlier studies (52), nonsequential screening with limited fundus views
proved to be an insufficient screening strategy (53).
Ells et al. introduced two important concepts: the first was referral-
warranted ROP (RW-ROP) defined as any ROP in zone I or presence of Plus
disease (≤2 quadrants) or any stage 3 ROP, and the second was standardized
fundus imaging using a series of five standard images per eye with respect to
the optic nerve—centered, superior, nasal, inferior, and temporal (54). RW-
ROP recognized disease that merited serious consideration for treatment as
opposed to being any ROP within a specified area. Using a prospective,
longitudinal study design with bedside BIO examinations for comparison,
RDFI had 100% sensitivity in identifying RW-ROP (54). Equally important,
remote RDFI readers often identified RW-ROP earlier than did examiners
using BIO: 1 week prior to BIO confirmatory examinations in 43% of
patients (10 of 23 patients), at the same time as BIO in 43% of patients, and
after BIO in 13% of patients (3 of 23 patients) (54). For the RDFI-identified
RW-ROP patients prior to BIO, 60% went on to receive laser, as opposed to
the 0% (all who spontaneously regressed) of BIO-identified RW-ROP prior
to RDFI. This observation suggested that the RDFI had greater ability for
accurate longitudinal comparisons in contrast to the BIO that required
remembered perceptions of three-dimensional viewing and visualization of
zone III retina (54). A subsequent study also demonstrated that RDFI had
100% sensitivity in identifying prethreshold and threshold ROP when
compared to contemporaneous bedside BIO (72).
In the PHOTO-ROP trial, six standard photographs were introduced: one
external iris photograph and five fundus views previously described by Ells
(54,71,76). The iris view allowed for visualization of any media opacity, iris
neovascularization, and/or adequate visualization that may hamper RDFI.
The PHOTO-ROP trial also demonstrated that BIO and RDFI were
essentially equivalent at identifying clinically significant ROP (CS-ROP)
(71,76). Notably, PHOTO-ROP researchers pointed out the fault in using the
examiner findings as the reference standard: “The examiner’s interpretation
of fundus findings is presumed to be correct without opportunity for review”
(71).
Chiang et al. (55) demonstrated that even inexperienced ROP readers,
albeit board-certified ophthalmologists, could achieve high levels of
sensitivity and specificity using RDFI after minimal training by an
experienced pediatric ophthalmologist (55). RDFI has been found to have
high sensitivity and specificity for identification of ROP requiring treatment
(58). Chiang et al. have expended significant efforts to assess the
interobserver and intraobserver differences with RDFI compared to
independent BIO examinations using an atlas of ROP images compared with
historical BIO examinations and demonstrated high sensitivity and specificity
in detecting Plus disease (55–58,60,61,75). Dhaliwal et al. (74) demonstrated
increased sensitivity and specificity with RDFI screening when the outcome
was increasingly raised from detection of any ROP to detection of ROP
requiring BIO confirmation and finally to ROP requiring treatment.
The Telemedicine Approaches to Evaluating Acute-Phase ROP (e-ROP)
study is the largest study to date that compared RDFI to the clinical
examination (77). It was conducted in 13 NICUs in North America to
evaluate the use of RDFI in infants in order to detect RW-ROP by non–
physician-trained readers. From the 5,520 pairs of image sets analyzed for
1,257 infants, RDFI using remote nonphysician readers had a sensitivity of
82% and specificity of 90% to detect RW-ROP in an eye at a single session
compared against a “study-trained ophthalmologist, rigorously trained in
ROP diagnostic examinations.” When both eyes were considered as a pair, as
in real-life setting, sensitivity reached 90% and specificity 87% (77). Despite
being the study with the highest level of evidence so far, the e-ROP trial
perpetuated a limitation of preceding prospective comparison studies
(54,71–74). Absent a double-masked evaluation of each and every BIO exam,
there remained an oversampling bias against RDFI in favor of an unproven,
unstudied gold standard that is, by default, assumed to be correct. Similarly,
with respect to feasibility, the e-ROP trial sought to evaluate how many
infants could be successfully imaged but did not have a similar comparator
for BIO, such as how many infants had clear media that allowed for certain
delineation of the extent of the retina.
Using data gathered from the e-ROP trial, Kemper compared RDFI with
bedside serial examinations using a microsimulation approach of a
hypothetical cohort of patients. All cases of treatment-requiring ROP were
detected using RDFI in combination with a BIO examination at discharge
because of the inability of RDFI to classify retina as mature. Using this
approach, more total interventions, considering both RDFI and BIO
examinations, for infants were needed but fewer BIO examinations required
(78).
Safety of RDFI
Clinical examination using BIO is very stressful for preterm infants and can
elicit a considerable amount of discomfort and pain, inducing changes in
blood pressure, apnea, bradycardia or tachycardia, emesis, and epistaxis
(79–84). Also, paralytic ileus, necrotizing enterocolitis, and cardiopulmonary
arrest have been reported but in the sickest infants who required respiratory
support and who had neurologic injuries or infections (85–87). A wide
variety of causes have been proposed including the mechanical effect of the
speculum or the scleral depressor, the oculocardiac reflex, the systemic
effects of anticholinergic and alpha-adrenergic agents used for mydriasis, the
bright light of the camera, and the supine position required for the exam
(79–83). RDFI exposes the preterm infants to the same mydriatic
medications, similar eye manipulation, and bright light and, in addition to
these, the camera in contact with a gel applied to the eye. RDFI causes
comparable discomfort and pain with similar physiologic changes compared
to BIO (83–88), although BIO may produce fewer events (8% BIO vs. 15%
RDFI) (84). This spectrum of physiologic changes and adverse events occurs
principally in the sickest and more fragile infants who are also the ones in
greatest need of ROP screening and timely treatment for ROP (84).
Limitations of RDFI
Despite many advantages, telemedicine can present some caveats. The
accuracy of RDFI is lower for the milder forms of ROP due to the difficulty
to visualize the periphery that can prevent a valid evaluation of zone III
disease, although it is unlikely to be clinically significant in the absence of
Plus disease (89). The lack of perception of depth could reduce the accuracy
of recognition of the fibrovascular proliferation and hence stage 3 disease
(90). Telemedicine performs worse in terms of accuracy, reliability, and
image quality in infants at younger PMAs because of many factors including
more corneal or vitreous opacity and narrow palpebral fissures (57). The BIO
examination allows real-time evaluation of the infants, and in the case of
telemedicine, protocols need to define that the results of the image
assessment should be delivered to the ROP specialist promptly (<24 hours)
(91).
American Academy of Ophthalmology on RDFI

An AAO Ophthalmic Technology Assessment report of RDFI for ROP in
2012 identified five studies with at least level I evidence supported RDFI
with “high accuracy for detection of CS-ROP” and multiple level III studies
that also reported high accuracy without known complications (92). Noted
exceptions included studies with low sensitivity for detection of stage 3
disease (57%) or for detection of stage 2 or greater ROP (76% and 77%) (92).
A joint technical report by the AAP, AAO, and AACO in 2015 noted
additional level I evidence and reconfirmed the high accuracy of RDFI for the
detection of CS-ROP (93). However, they concluded that there was a need
for systemic studies evaluating the cost, benefit, and trade-offs from the
different perspectives including patients, providers, payers, and
policymakers. A 2013 joint policy statement on “Screening Examination of
Premature Infants for ROP” recognized the growing role for digital imaging
in ROP to allow for remote ROP screening and objective documentation of
retinal findings (19). The 2018 revision to the joint policy statement, in the
light of the results of these studies, recognized the value of telemedicine for
preliminary ROP screening or as an adjunct to BIO examination and stated
that in cases of ambiguous images or development of treatment-requiring
ROP, the infant should have access to timely bedside examinations and
treatment (20).
Tele-Education for ROP

Image-based training in contrast to bedside training with BIO has overcome
some barriers such as the significant amount of stress induced by multiple
BIO examinations (94) and the lack of standardized contents and validated
tools to assure quality of training (95). Global Education Network for ROP
(GEN-ROP) tele-education system has been proven to increase the accuracy
of the diagnosis of zone, stage, Plus disease, category, and aggressive
posterior ROP (APROP) and intergrader agreement in diagnosis of ROP (95).
Multiple additional tele-education tools have been released including the
AAO Web-based educational tool to improve the competency of
ophthalmologists-in-training to diagnose and manage ROP
(www.aao.org/interactive-tool/retinopathy-of-prematurity-case-based-
training), the Focus ROP online educational platform and certification
program for RDFI in ROP (www.focusrop.com), and the Wide-field Imaging
of Infants for Screening and Education for Retinopathy of Prematurity
(WISE-ROP) developed in India to train and certify ROP technicians (96).
While tele-education does not have to replace the traditional training based
on bedside BIO exams, it is a valuable additional synergistic teaching
modality.
Imaging and Informatics in Retinopathy of Prematurity

Deep learning (DL) is the subfield of artificial intelligence (AI) dedicated to
developing deep neural networks (DNNs). A DNN is a computing system
that is modeled after the human brain with processing nodes that act as
neurons and the neuron layers behave as segments of the brain (97).
The Imaging and Informatics in Retinopathy of Prematurity (i-ROP)
consortium trained a DNN to detect Plus disease in ROP using a dataset of
5,511 color fundus photos. When the algorithm was tested using a reference
standard diagnosis (RSD) based on the consensus of image grading by three
experts and a clinical diagnosis by one expert, the areas under the receiver
operating characteristic curves were 0.94 for the diagnosis of normal and 0.98
for the diagnosis of Plus disease. Using an independent set of 100 retinal
images, the algorithm reached the sensitivity of 93% with 94% specificity for
the diagnosis of Plus disease (98). The DL algorithm was also tested against
the “i-ROP Plus score,” a grading system to assess posterior pole vascular
abnormalities in ROP on a 1 to 9 scale. Using a hypothetical threshold i-ROP
Plus score of 3, the algorithm had a sensitivity of 94% and specificity of 79%
to detect type 1 ROP (99). While this DL system has been validated with
many good-quality images, further studies will be needed to test its
generalizability (100).
Real-World Application of RDFI Screening for ROP

The Early Treatment for Retinopathy of Prematurity (ETROP) trial reported
that disease treatment is almost entirely predicated upon two determinants:
zone I disease or Plus disease (13) and, to a lesser extent, stage 3 disease that
enters as a deciding factor (13). Accounting for differences in ocular size,
crystalline lens sizes, and aberrations induced by the visual axis, a centered
optic nerve RetCam fundus image provides at least a 100-degree field in
every infant, which is 50-degree radius around the optic nerve. This can be
compared to ICROP zone I, which is a 30-degree radius, and zone II, a 60-
degree radius around the optic nerve (21,23). Although there are no defined
criteria as to what represents posterior zone II, one might reasonably assume
that a 50-degree radius encompasses all of posterior zone II. Thus, a good-
quality, well-centered image could provide an objective method for
ascertaining the key determinants of treatment-warranted ROP (TW-ROP).
Some may argue about the validity of Plus interpretation using RDFI
(75,101–107); however, these arguments generally fail when the longitudinal
comparison power of RDFI is leveraged (54), which is something that is not
feasible with BIO (71). Real-world RDFI networks have proliferated around
the world with much success, such as the Stanford University Network for
Diagnosis of Retinopathy of Prematurity (SUNDROP) in the United States
(108), Karnataka Internet Assisted Diagnosis of Retinopathy of Prematurity
(KIDROP) in India (109), TROPIC in Canada (54) and in Germany (62),
ARTROP in New Zealand (68), and PCA-PERP in Hungary (110).
SUNDROP RDFI Network in the United States

In 2005, the SUNDROP was deployed as a community outreach RDFI
screening program in the San Francisco Bay Area to provide access to
quaternary ROP experts while maintaining the infant in their home NICU.
The SUNDROP program incorporated screening for a defined clinical end
point, utilization of trained nursing staff, sequential imaging, mandatory
fundus and external views, HIPAA-compliant image transfer, clear
delineation of responsibilities, a well-defined ROP safety net, and contractual
obligations with respect to duties for all parties. The SUNDROP telemedicine
outreach program differed from the previously published studies in that RDFI
was the primary means of screening—BIO was only employed if RW-ROP
was reached or upon discharge from the hospital (59,64). All babies were
discharged to one quaternary ROP screener who performed BIO up until such
point that the infant reached criteria for termination of acute retinal screening
examinations (called termination criteria) (19).
Initially, RW-ROP was the trigger for an intervention by the ROP
specialist (13,59,64,112). Eighteen months later, the end point was raised to
TW-ROP, defined as type 1 ETROP (13) and/or threshold ROP (111),
because it became increasingly clear that RDFI was highly successful at
identifying all ROP that needed treatment. Sensitivity and specificity have
increased with each published report (66,69,109,112). In its 14th year of
operation as a store-and-forward, hub-and-spoke RDFI screening program for
ROP (Fig. 20-1), the sensitivity for TW-ROP is 100% with specificity
exceeding 99.8%. More than 1,800 infants have been screened; none of them
has been lost to follow-up, suffered adverse events from the screening
process, or gone on to develop blindness or retinal detachment. The success
of SUNDROP is due to all parties being 100% committed to the program, the
ability for longitudinal comparison, and the capability for outside expert
consultation.
FIGURE 20-1 Schematic representation of the

SUNDROP hub-and-spoke RDFI screening program for
ROP.
KIDROP RDFI Network in India

The KIDROP program is the world’s largest ROP telemedicine network. It
was initiated in the state of Karnataka, India, in 2007 and has expanded to
provide ROP screening through an indigenously developed telemedicine
platform that includes 126 NICUs in 18 districts (3 zones of 6 districts each)
utilizing 20 teams trained in RDFI ROP interpretation from throughout India.
This program addressed the problem of unscreened rural and semirural
premature infants for ROP using a novel telemedicine platform. KIDROP
uses a single RetCam Shuttle, the portable version of RetCam, and recently
they adopted 3nethra neo (Forus Health Pvt. Ltd., Bangalore, India), a low-
cost portable camera developed in India. Dedicated teams of trained
technicians transport the device between NICUs situated within rural and
semiurban neonatal centers on a fixed schedule every week. Image capture,
processing, storage, analysis, and reporting are performed on site by the
technicians using a triage-based algorithm based on the principles of pattern
recognition (Fig. 20-2). Images are also remotely viewed and reported on
smartphones or personal computers by ROP experts from Bangalore where
KIDROP is headquartered, providing real-time diagnosis to the rural infants.
The reports are transmitted to the rural technicians using the cellular network
or the Internet via secure data server on Tele-ROP platform (i2i Tele-solution
Ltd, Bangalore, India) with a digital signature before the child can be handed
over to the mother in most cases. Treatment is also provided at the rural
center itself by the KIDROP team of experts or locally trained
ophthalmologists. In this program, the grading by the highest-level
technicians had a sensitivity of 96% and specificity of 92% to detect
treatment-requiring ROP with a negative predictive value of 99% (110). The
program has now evolved in training resident nurses of selected NICUs to
screen for their admitted infants.
FIGURE 20-2 Schematic representation of the KIDROP
program. Image capture, processing, storage, analysis, and
reporting are performed on site by the technicians using a
triage-based algorithm based on the principles of pattern
recognition. ICROP, International Classification of
Retinopathy of Prematurity.
KIDROP demonstrates the potential of coordinated public–private

partnerships to expand access to care to prevent a worldwide leading cause of
childhood blindness. The KIDROP screeners utilize an integrated RDFI
platform employing automated binary compression, a patented technology to
provide live, lossless image transfer (lossless compression reduces a file’s
size without loss of quality), and live reporting in conjunction with
smartphone or tablet applications. Furthermore, the program has expanded to
provide universal screening in some zones for healthy full-term infants. Two
unique features of the KIDROP program are the widespread implementation
of nonphysician readers to grade and analyze the images as the first point of
contact and the lack of any mandated BIO examination at any point to
terminate screening. It remains to be seen if either of these innovations is
effective: the first in regard to sensitivity and specificity and both in regard to
whether they are generalizable to highly developed countries as defined by
the United Nations Development Program Human Development Index (113).
Furthermore, particularly with respect to the United States, use of
nonphysician readers introduces medicolegal uncertainty (24,25). The
challenges that face KIDROP are (a) barriers to adopt to new technology; (b)
lack of awareness of the disease even among the medical community; (c)
Internet speeds that are variable and often unreliable in the most backward
areas, which hamper live image transfer; and (d) relatively large capital costs
of equipment during initiation of the program. Despite these issues, in the 12
years since inception, around 150,000 imaging sessions have been performed,
over 40,000 unique infants have been enrolled, and over 2,500 infants have
been identified with type 1 ROP. The KIDROP program is now poised for
nationwide expansion in India.
IMPLEMENTATION OF AN RDFI
SCREENING PROGRAM FOR ROP
Personnel
The RDFI team includes the RDFI reader/treater, the nurse photographers,
the bedside nurse, the NICU team (physician, social work, nurse
practitioners), the front desk schedulers, the legal consultants, the hospital
administration, and the family members. Each group has a role to play to
ensure a smooth transition from birth, identification of need for screening,
screening, implementation of reader recommendations, discharge, handoff
and communication to outside screen, and completion of screening
termination criteria. The easiest method for delineating roles is through the
ROP screening contract. In addition to outlining each team member’s
responsibilities, the contract can serve to head off disputes about what to do
when a photographer is unavailable, when an infant needs treatment, when
there is an equipment malfunction, and the like.
Photographic Technique
The photographic screening protocol most widely employed is derived from
the PHOTO-ROP study (Fig. 20-3) using a RetCam (Fig. 20-4). Images are
taken by nonphysician nurses or other trained personnel who have received
training and, ideally, certification in the use of the camera and imaging
technique, such as through Focus ROP or Natus medical online training
program. In the SUNDROP program, the nurse photographers utilized such
training and then have additional annual training with the RDFI reader.
Dilation is key to good photographs. Feedings should be held for at least an
hour around an imaging session to reduce the risk of apnea, bradycardia, and
aspiration. We recommend that the bedside nurse be present throughout the
photographic session. A speculum is an absolute requirement, and the
Alphonso speculum appears well suited for the 130-degree lens of the
RetCam. We do not advocate for routine usage of scleral depressors. With
respect to imaging frequency, RDFI images are most useful when there is a
longitudinal record, and, therefore, a screening frequency of at least weekly is
most likely to ensure continuity of care, instill in the team members the
importance of the enterprise, as well as capture rapidly advancing disease.
FIGURE 20-3 PHOTO-ROP standard image set for a left

eye, consisting of an image of the iris and the optic disc,
followed by images of the temporal, nasal, superior, and
inferior retinal fields.
FIGURE 20-4 Image of the RetCam® 3 camera (Image
provided courtesy of Natus Medical Incorporated).
RetCam remains the most commonly used camera for neonatal ophthalmic
imaging so far in both clinical and research settings, but several fundus
cameras designed for neonatal use have been developed over the years. The
ICON (Phoenix, Pleasanton, CA) is a wide-field contact camera coupled a
handheld probe with direct annular illumination that provides a 100-degree
field of view and a resolution of 8 MP. The PanoCam LT, PRO, and Solo
(Visunex, Fremont, CA) are wide-field wireless cameras that render a 130-
degree field of view and a resolution, respectively, of 8, 13 and 13/8 MP. The
3nethra neo (Forus Health Pvt. Ltd., Bangalore, India) is a lightweight wide-
field portable contact camera, which provides a 120-degree field of view with
a handheld probe, uses warm LED light as the illumination source, and can
capture images with 4 MP resolution. The Pictor Plus (Volk Optical,
Cleveland, OH) and Nidek NM200-D (Nidek Inc., Gamagori, Japan) are two
portable noncontact digital cameras that provide images with, respectively, 5
and 1.5 MP resolution but with a 45- and 30-degree field of view. The Optos
Optomap (Optos PLC, Dunfermline, UK) is a noncontact ultra–wide-field
imaging system that can capture retinal images up to 200 degrees, but it is
table mounted; the only way to examine a newborn with this machine is
utilizing the “flying baby technique,” and because of that, it is inappropriate
for routine ROP screening. The Heidelberg Spectralis (Heidelberg
Engineering, Heidelberg, Germany) became available recently as modified
version “Flex Module” that allows examination of patients in supine position.
The latter is a noncontact camera that offers wide-field 55-degree fundus and
OCT imaging.
Image Transfer and Storage

Once the six images for each eye have been obtained and assessed for quality
and completeness, the images need to be transferred to the RDFI reader in a
HIPAA-compliant fashion using a file transfer protocol, automated
synchronization with the server, or secure email.
To verify receipt, the RDFI reader should be notified independently via
email, text, or phone message that images on X number of infants have been
sent. With absent confirmation of receipt, the nurse photographer should
contact the reader to verify and clarify. With the present camera, 12 images
will occupy approximately 10 MB. These images need to be stored either
with the reader on a secure server or locally with the camera or both. For
larger networks, it will be economical to have redundant off-site storage,
managed by the home institution’s information technology service or similar.
Bear in mind that images may need to be stored until the infant reaches
maturity (21 years of age), due to medicolegal concerns.
RDFI Report
The report of the images taken at each examination will become a permanent
part of the medical record and require general documentation, documentation
per eye and a summary of the interpretation, a recommendation as to future
examinations or treatment, as well as an educational handout for the family
members explaining to them the disease grading and recommendations (24)
(Table 20-1). If the reader finds that the images are unacceptable for grading,
for example, lack sufficient quality, have incomplete image sets or have
artifacts, and are mislabeled, then a request should be made to repeat the
evaluation within 24 hours. Reports should be created and returned no later
than 24 hours after receipt of images, unless other arrangements have been
made in advance such as communicating the results in person or by phone.
These reports serve many functions in addition to their medical support role,
such as education of the NICU team, education of family members, creation
of RDFI team solidarity, and use in institutional review board (for human
studies)-approved studies.
TABLE 20-1 Report of the remote digital fundus

imaging (RDFI) for each ROP examination
RDFI Graders
Although it is understood that every member of an RDFI team is
interdependent upon each other and essential to its success, the RDFI
physician grader holds a leadership role for the team. Even experienced ROP
screeners may feel uncomfortable the first time they begin grading images.
Early in the start-up of the screening program or when a new grader joins the
team, the grader may wish to examine the patients with BIO prior to the
photographs being obtained and then compare the images to the examination
findings.
Presently, there are excellent mentoring opportunities in a live setting or
through online certification courses. It is important for the entire team to have
confidence in the ability of the grader, which means that the grader has to
have confidence in the images. This can mean that the grader has to become
comfortable grading images of suboptimal quality—that is, those with
incomplete sets, poor lighting/contrast, artifact, or defocus—without
compromising care. This confidence can be accomplished by shadowing
when examinations of infants with BIO are being conducted during the
initiation period. Access to BIO examination on site or through timely
transfer should be guaranteed in case of ambiguous or suboptimal images
(20). It is important to keep in mind that the goal of RDFI is not to identify
all of the ROP in the eye but rather to identify the infants at high risk for
blindness to ensure that they receive adequate care. By demonstrating
competence, compassion, and diligence, the grader can inspire the other team
members to excel at their own competencies.
Discharge
It is rare that any baby will meet the criteria for termination of ROP screening
(19) prior to discharge from the NICU, and, therefore, ROP screening with
BIO is very likely to be performed after discharge. The handoff to
postdischarge follow-up is essential, and it is most important that the transfer
of care is successful and that the accepting ROP screening physician sees the
patient. Typically, a premature infant will be discharged around 40 weeks’
PMA and, at this point, is a poor candidate for RDFI with the current
generation of cameras due to size and difficulty with examination. In light of
the difficulty of camera examination in these older infants, in the case of
ambiguous or suboptimal images (20), there may be a need for a BIO
evaluation for ROP screening (20) prior to discharge from the hospital. It is
imperative that an ROP physician continues timely screening examinations
until criteria to terminate ROP screening have been met.
FUTURE DIRECTIONS
Telemedicine opens the doors to a multitude of future directions. The
Newborn Eye Screen Testing (NEST) and its substudy the Global Universal
Eye Screen Testing (GUEST) are currently investigating the impact of the
universal eye screening in newborns; this could help to detect many
abnormalities that are missed at the neonatologic physical examination and
also collect data about early life physiologic development (114,115). Besides
the recent hardware improvements, the software developments have been
progressing even faster, and it has involved every field of medicine. One of
the most exciting aspects of telemedicine is the computer-based image
analysis software platforms that can objectively analyze and quantify retinal
features. Many systems have been developed to date, such as Retinal Image
multiScale Analysis (RISA), and its successor Computer-Aided Image
Analysis of the Retina (CAIAR) developed at Imperial College London,
ROPtool developed at Duke University, VesselMap (IM-EDOS GmbH,
Weimar, Germany), and i-ROP with promising results. In the near future, as
has already happened in other medical fields, AI image analysis systems will
be able to predict outcomes and prognosis with a further improvement of the
management of this disease (116).
SUMMARY
There is a large and growing body of evidence to support the use of RDFI
screening for the identification of referral and TW-ROP. The exact methods
of screening continue to be refined, but the systematic approach championed
by the PHOTO-ROP trial and the SUNDROP telemedicine outreach
programs that utilize standard image sets and trained nurse photographers,
experts in ROP evaluation, and mentored readers with integrated ROP safety
nets has demonstrated that with such approaches, all TW-ROP can be
identified in a timely fashion to prevent blindness. The major trade-off with
the use of RDFI is that of not identifying every bit of ROP disease in the eye
that BIO may potentially do and instead only identifying ROP that requires
treatment to prevent blindness. RDFI programs have the potential to improve
accessibility, quality, and cost of ROP care in both developed and developing
countries and should be initiated with clear contractual understanding of each
team members’ roles and responsibilities predicated further on each member
receiving adequate training and certification.
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SECTION IV
Genetics and Developmental
Disorders in Pediatric Retina
21
Genetic Counseling for Pediatric
Retinal Diseases
Karmen M. Trzupek and Briana L. Sawyer
INTRODUCTION
The completion of the Human Genome Project laid the foundation for a
seemingly infinite number of research programs with goals as audacious as
they are impactful: cataloguing the breadth of common human genetic
variation, identifying and characterizing every functional component of the
genome, and discovering and understanding rare genetic variants associated
with human disease. The success of the latter programs—associating rare
genetic variants with Mendelian genetic diseases—has led to a tremendous
surge in clinically relevant genetic information. This information has the
potential to significantly impact medical care and decision-making for not
only the patient but also their family.
Genetic information can also have emotional, reproductive, social, legal,
and ethical implications; therefore, health care providers involved in ordering
genetic testing or treating patients with genetic diseases should provide their
patients with resources to help explain the meaning of genetic information.
Ophthalmologists managing these patients should have a basic understanding
of the genetics of that condition, the impact of genetic test results on medical
management, and potential implications for the patient and family. A more
detailed discussion of inheritance, family/social implications, recurrence
risks, and options for genetic testing may necessitate a comprehensive
genetics consultation.
Many physicians bridge this need by partnering with genetic counselors,
who can collect detailed family histories, provide genetic and prognostic
counseling, and review molecular testing options with patients. Genetic
counselors carefully review genetic variants in test reports and research their
potential contribution to disease. We all have many variants from the
standard sequence in our genes, as our diverse traits are directed by different
versions of the same genes. A variant is often called a pathogenic mutation if
it is disease-causing. Whether or not a specific variant is disease causing or
benign requires complex analysis of the change in protein structure and
analysis of the literature. Variants may be classified as benign, likely
pathologic, pathologic or uncertain. The American College of Medical
Genetics (ACMG) concludes that both pathogenic and likely pathogenic
variants should be interpreted as disease causing and actionable. For the
purposes of this chapter, the word mutation will be used to signify pathologic
and likely pathologic variants.Following genetic testing, the genetic
counselor may coordinate additional testing of family members, enroll the
patient in research or patient registries, and discuss clinical trials. Genetic
counselors also provide support for the emotional impacts genetic
information and testing results can have on a patient and their family.
GENETIC COMPLEXITY OF
INHERITED RETINAL DISEASES
In 1984, Shom Shamker Bhattacharya, PhD, and colleagues mapped the RP2
gene associated with X-linked retinitis pigmentosa (RP) (1). The ensuing 30
plus years have seen tremendous advances: today, nearly 300 inherited retinal
disease genes have been mapped, and more than 250 of those genes have
been cloned (2).
Retinal disease genetics have proven to be a model of genetic complexity,
displaying not only allelic heterogeneity, where many different disease-
causing mutations are found within a particular gene, but also:
Genetic heterogeneity: Mutations in different genes may cause the same

disease.
Phenotypic heterogeneity: Different mutations within the same gene
may produce different clinical phenotypes.
Clinical heterogeneity: The same mutation in different individuals, even
within the same family, may produce different clinical consequences.
This feels abrupt, I wonder if a transition something such as RP is
particularly complex and shows each of these types of heterogeneity. RP is
particularly complex. RP can be inherited as an autosomal dominant,
autosomal recessive, X-linked, or mitochondrial condition. Rare digenic
forms have also been documented, in which mutations occurring in two
different genes act together to cause a retinal disorder. In addition, whereas
inherited retinal diseases frequently occur as an isolated condition, they may
also occur as just the presenting symptom of a larger recognizable genetic
syndrome or metabolic disease.
For example, some children with Leber congenital amaurosis (LCA),
presenting in infancy or early childhood with apparently isolated retinal
disease, may in fact be affected with an underlying diagnosis of a syndromic
condition such as Joubert syndrome or Senior Loken syndrome (3). Genetic
testing is typically used to guide surveillance.
Conversely, some genes historically associated with syndromic forms of
retinal dystrophy can in fact cause isolated RP or LCA. This has significant
implications for the interpretation of genetic testing data. While USH2A is the
gene most commonly known to be associated with Usher syndrome type 2, it
is also now known to be the most common cause of nonsyndromic RP (4).
The CLN3 gene has long been known to be associated with neuronal
ceroid lipofuscinosis (NCL, or Batten disease), a universally fatal disease that
typically begins with childhood onset severe retinal dystrophy, followed by
seizures and rapid psychomotor deterioration. Recently though, mutations in
CLN3 have been described in multiple unrelated families with nonsyndromic
RP (5). Genes previously believed to cause a very narrow syndromic
phenotype may in fact be associated with a high degree of variability in
symptom type, onset, severity, and progression (6).
Due to the wide degree of phenotypic and clinical heterogeneity known
to occur in some genetic diseases (including but not limited to inherited
retinal diseases), a significant shift is underway in medical genetics to begin
naming diseases according to their underlying molecular cause.
Mutations in RPE65 are known to be associated with not only LCA but
also severe early childhood onset retinal dystrophy and juvenile onset RP
(7,8). CRB1 mutations have been associated with LCA, RP, cone-rod
dystrophy, and RP with preserved para-arteriolar RPE (PPRPE). Gene-based
treatments should benefit patients with underlying mutations in those genes
irrespective of their narrowly defined clinical diagnosis.
CLASSIC INHERITANCE PATTERNS

Mendelian (single gene) disorders are classified as autosomal dominant,
autosomal recessive, or X-linked conditions. They are the result of genetic
mutations in the nuclear DNA. An autosomal dominant disorder is one in
which the disease phenotype is observed in heterozygotes (individuals with
one copy of the genetic mutation). Both genders are equally likely to be
affected, and the mutation is transmitted from generation to generation,
demonstrating a vertical transmission pattern as shown in Figure 21-1A. If
one parent has the disease, the probability that each child will inherit the
mutation is 50%. Best disease, also called Best vitelliform macular dystrophy
(BVMD), is an example of an autosomal dominant condition.
FIGURE 21-1 A–D:Inheritance patterns for genetic
conditions with Mendelian inheritance. (This image is a
work of the National Institutes of Health, part of the
United States Department of Health and Human Services.
As a work of the US Federal Government, the image is in
the public domain.)
Most individuals who are diagnosed with Best disease have a parent who is
also affected, although a wide degree of variability in age of onset and
severity of vision loss is not uncommon. Even within the same family, the
age of onset of symptoms can range from childhood or adolescence to middle
adulthood.
Autosomal recessive disorders are those in which an individual must have
mutations on both copies of the disease-related gene in order to be affected
(see Fig. 21-1B). We all carry multiple single copy recessive mutations
without any effect, and for two unrelated parents, the chances of each
carrying a mutation in the same gene is low but varies based on the
prevalence of the disease. Approximately 1 in 70 people carries a mutation in
an albinism causing gene, for example, whereas the carrier frequency is much
lower for LCA. The affected individual may be compound heterozygous
(having a different mutation on each copy of the disease gene) or
homozygous (having two copies of the same mutation: 1 inherited from each
parent). Homozygosity occurs more frequently when the patient’s parents are
related by blood or have ancestors from the same small ethnic community.
Carriers have only one copy of the mutation and are usually
asymptomatic and phenotypically unaffected. Typically, both parents of a
child affected with an autosomal recessive condition are carriers, and each of
their offspring has a 25% chance of inheriting the disorder. Multiple family
members in a single generation (e.g., siblings) may be affected, as shown in
Figure 21-1B. All children born to a parent affected with an autosomal
recessive condition will at least be carriers of the condition. If a child inherits
a second genetic mutation from the other parent, they too will be affected.
For a rare genetic disorder, this occurs very uncommonly. However, the
probability is increased if the parents are related or from an ethnic
background with a relatively high prevalence of the disorder. (See discussion
of pseudodominance below.) Stargardt disease and Usher syndrome are
conditions that are inherited in an autosomal recessive manner.An X-linked
disorder is one in which the causative genetic mutation is located on the X
chromosome. Because males have only one X chromosome, all males who
inherit a mutation are expected to develop the disorder. Females have two X
chromosomes; thus, if they inherit one copy of the mutation, they are carriers
(see Fig. 21-1C). For a classic X-linked recessive disorder, female carriers are
typically not affected, although they may display some symptoms due to
skewed X inactivation. Generally, these symptoms are milder than those seen
in affected males. Because fathers do not pass an X chromosome to their
sons, the lack of male-to-male transmission is a characteristic feature of X-
linked inheritance. All daughters of a man with an X-linked recessive
condition will be carriers. When the mother is a carrier, the risks to her
children depend on their gender: sons of carrier females have a 50% risk of
being affected, whereas daughters of carriers have a 50% chance of being
carriers as shown in Figure 21.1C. Juvenile retinoschisis caused by mutations
in the RS1 gene is a condition that follows a classic X-linked recessive
inheritance pattern. Female carriers of an RS1 mutation rarely express fundus
abnormalities, although clinical findings have been reported in a few cases
(9). On the other hand, female carriers of mutations in the RPGR gene,
associated with X-linked RP, frequently do develop retinal disease, and may
be diagnosed with “typical” RP (10). Women in some X-linked RP families
may be affected to the degree that the inheritance appears dominant.
While rare, some conditions display X-linked dominant inheritance, in
which females with one copy of the mutation are affected. X-linked dominant
conditions are often embryonically lethal in males. An important example of
an X-linked dominant condition is incontinentia pigmenti (IP), which has
both skin and ocular manifestations (discussed in Chapter 36). Girls with this
condition classically develop the triad of ectodermal dysplasia, CNS
abnormalities, and retinal neovascularization, predisposing to retinal
detachment (11). Women with IP may report multiple miscarriages, generally
around the 3rd or 4th month of gestation, presumably of affected male
fetuses. Some cases of males with IP have been reported due to mosaicism or
a karyotype of 47,XXY providing an additional X chromosome, which
allows for viability.
Genetic conditions can also result from mutations in mitochondrial DNA
(12). Mitochondria are almost exclusively inherited from the mother, and
family histories with a pattern demonstrating mitochondrial inheritance will
often reveal an affected mother with affected children of both genders, as
shown in Figure 21-1D. Extremely rare examples of paternal transmission of
mitochondria have been recently reported, however (13). The presence of
equally affected females and males can help distinguish this inheritance
pattern from X-linked inheritance. Complicating mitochondrial inheritance is
the presence of heteroplasmy, which occurs when cells have some normal
and some have mutated copies of their mitochondrial DNA. In general, an
individual will only show symptoms of a mitochondrial condition if the
proportion of mutated mitochondrial DNA reaches a certain threshold.
Therefore, the disease could be clinically absent in a woman but present in
both her mother and her children. A classic example of mitochondrial
inheritance in the pediatric ophthalmology clinic is Leber hereditary optic
neuropathy (LHON), which causes painless, subacute vision loss, most
frequently in males aged 15 to 30. Although men and women are equally
likely to inherit the mitochondrial mutation, males are four to five times more
likely to lose vision. Less commonly, individuals who inherit an “LHON
mutation” will develop a multiple sclerosis-like illness; females are more
likely than males to develop the neurologic phenotype (14). The reasons for
the gender differences in this mitochondrial condition are unknown.
Another important example of mitochondrial inheritance in the retina
clinic is Kearns-Sayre syndrome, which is associated with pigmentary
retinopathy, progressive external ophthalmoplegia, ptosis, neurologic and
neuromuscular dysfunction, and variable cognitive impairment. However,
nearly all patients with Kearns-Sayre syndrome have deletions of the
mitochondrial DNA that arose sporadically and were not inherited from the
mother. The mother of a proband with a mtDNA deletion syndrome is usually
unaffected and does not have mtDNA deletions in her tissues; therefore, the
risk to the sibs of a proband is low, but not zero, due to the possibility of
germ line mosaicism. Empirically, the recurrence risk is estimated to be 1%
to 4% (15).
COMPLEX INHERITANCE TYPES

A disease is described as simplex if the affected individual has no other
family members with the disorder. Despite the absence of affected family
members, simplex retinal diseases can be the result of dominant, recessive,
X-linked, or mitochondrial inheritance. Others may have environmental
causes. Genetic testing is vital in these cases for accurate counseling
regarding inheritance and recurrence risks. The inheritance of retinal
dystrophies in particular can be difficult to predict based on family history
alone. The Inherited Retinal Disease Clinic at the University of Michigan
performed a retrospective study of patients diagnosed with RP, comparing the
presumed inheritance type derived from clinical diagnosis and family history
with the results of genetic testing (16). In that study, which collected a
detailed family history going back three or four generations for every patient,
genetic testing identified a different inheritance pattern than pedigree analysis
alone in 10% of patients.
These discrepancies in underlying inheritance can result from a number
of genetic factors, including de novo dominant mutations, X-linked disease in
families with manifesting female carriers, dominant disease with reduced
penetrance, and pseudodominance. These complex genetic factors, often
dismissed as too rare for consideration in the typical practice, collectively
account for a significant percentage of the underlying genetics of inherited
retinal disease.
In the pediatric retina practice, it has long been recognized that a small
percentage of children with LCA and severe childhood onset retinal
dystrophy have dominant mutations in the CRX gene. These mutations
usually arise as a de novo event in the child but can be passed down to their
children. Today, de novo dominant mutations associated with LCA are
currently known to occur in three genes: CRX, IMPDH1, and OTX2 (17).
Identifying these cases is vital to the provision of accurate genetic
counseling, both for affected individuals and parents of newly diagnosed
children.
In Stargardt macular dystrophy, the child of an affected individual will
occasionally also be affected with Stargardt macular dystrophy, as a result of
inheriting a mutation in the ABCA4 gene from his or her affected parent and
from an unaffected parent who is a carrier of the condition. This is called
pseudodominance, because on pedigree analysis the inheritance appears to be
dominant, but is actually recessive (18). The carrier frequency of ABCA4
mutations in the general population was previously estimated to be 1/50, but
genetic testing data now illustrates that the true carrier frequency of
mutations in the ABCA4 gene is approximately 1/30. As a result, up to 1 in 60
individuals with Stargardt macular dystrophy or ABCA4-related disease
would be expected to have at least one child who is also affected with the
disease.
Determining the pattern of inheritance can be complicated by incomplete
penetrance and variable expressivity. Penetrance is an on or off phenomenon,
someone has disease or does not. This can be compared with variable
expressivity which is a gradient of clinical features.
Penetrance has historically been thought to be complete in recessive
conditions, but more recent studies have proven that it does not always hold
true. Reduced penetrance is well known in some autosomal dominant
disorders, meaning that some individuals who inherit the disease-causing
genetic mutation do not develop symptoms. Variable expressivity, on the
other hand, is a descriptive term for the variation in phenotype among
individuals with the abnormal genotype (19).
There is usually greater phenotypic variation in autosomal dominant
conditions than in autosomal recessive or X-linked disorders. Dominant RP
resulting from mutations in the PRPF8 and PRPF31 genes is well known to
be associated with reduced penetrance. Similarly, familial exudative
vitreoretinopathy (FEVR) can be associated with both a wide degree of
variability and apparent reduced penetrance. Young children with FEVR may
present with severe vision loss resulting from retinal ischemia and failure of
peripheral retinal vascularization. This is one of only a few inherited retinal
diseases in which expression can be markedly asymmetric. Parents and more
distant family members may be clinically unaffected, though on close
examination, one parent nearly always has a region of avascularity in the
peripheral retina (20).
Many conditions are influenced by environmental factors as well as
genetic mutations, such that risk of disease is increased with exposure to the
environment. A trait or condition is said to have a multifactorial inheritance
pattern if both genetic and environmental factors contribute to the phenotype.
Diabetes is a common example of a multifactorial disease. Many ophthalmic
conditions are multifactorial. For example, age-related macular degeneration
(AMD) has both genetic and environmental factors that contribute to disease
development, progression, and severity. In the pediatric clinic, typical
juvenile myopia is another good example. (In contrast, some types of
pediatric onset high myopia are associated with Mendelian genetic diseases,
such as Stickler syndrome or congenital stationary night blindness [CSNB].)
Genetic testing for these multifactorial diseases is not yet clinically useful,
but with an increasing understanding of the role of genetic susceptibility
factors and the development of precision medicine therapies, this will likely
change in the not-too-distant future.
Family History
The family history is an important tool for determining differential diagnoses,
risk assessment, and the most appropriate genetic testing. Additionally, the
family history allows the genetic counselor to assess patient education needs
and psychosocial influences. A family history is obtained in virtually all
genetic counseling sessions, generally includes at least three generations, and
often assesses a variety of health concerns. For this reason, the family history
is arguably one of the most important tools in both screening and prevention
of common diseases (21,22).
Targeted questions regarding the status of affected as well as unaffected
individuals can help the genetic counselor determine the most likely
diagnosis as well as risk to other family members.
Inheritance patterns and disease-specific risk assessments can be
generated from family history alone in many cases.
The family history and/or pedigree should be an integral part of the
medical record. The term pedigree refers to the way the family history is
visualized. Pedigrees should be drawn in a standard fashion to allow for easy
interpretation by other health care providers. Guidelines for pedigree
documentation have been developed by the National Society of Genetic
Counselors (23).
Figure 21-2 provides an overview of the most commonly used pedigree
symbols. Full names, dates of birth, or any other potentially identifying
information about family members should be excluded from the pedigree in
order to maintain the confidentiality of family members. How to best capture
family history information in the electronic medical record (EMR) is the
subject of a great deal of research. For the past several years, a variety of
institutions and researchers have worked to create a pedigree tool that is
compatible with the EMR; however, clinics and research institutions vary
according to practice.
FIGURE 21-2 Common symbols used in pedigree
construction. (From Bennett RL, French KS, Resta RG, et
al. Standardized human pedigree nomenclature: update
and assessment of the recommendations of the National
Society of Genetic Counselors. J Genet Couns
2008;17:424–433. Copyright © 2008 National Society of
Genetic Counselors, Inc. Adapted by permission of John
Wiley & Sons, Inc.)
Parents should be asked specifically if they are both the biologic parents of
the affected child before undertaking a complex pedigree drawing. Although
this may seem obvious, there are many types of families, and adoptive
parents may not realize that only the medical conditions of biologic relatives
are important to a genetic pedigree. In addition, some families may not have
shared family of origin information with their child; parents should be given
the opportunity to provide additional family information at a later time if they
request that.
Risk Assessment
In most genetic counseling sessions, some discussion of risk will occur. This
may be the risk of an individual developing symptoms, the risk for family
members to carry a genetic mutation, or the risk of progression to more
severe disease. Those present may include the patient and other family
members who wish to understand the risk to themselves and their children (or
future children). To best tailor the counseling session to the needs of the
patient, it is important to identify the patient’s motivation for requesting risk
information. Regardless of the type of risk discussion taking place, the risk
assessment is a crucial part of the genetic counseling session and involves
detailed assessment of the family’s medical history.
Determining the likely inheritance pattern of the condition is the first step
in estimating a specific risk for the patient. This necessitates the
determination of (even mildly) affected family members. In some cases, the
inheritance type can be inferred directly from pedigree analysis or clinical
diagnosis. For example, Best macular dystrophy in a young boy with an
affected father is a dominant disease. Usher syndrome type 1, on the other
hand, is a recessive condition.
Genetic testing may still be valuable for these patients for further risk
stratification within the family but is not critical to determine the risk of
recurrence since autosomal recessive inheritance carries a 25% risk for each
child. However, for many other diseases— particularly RP and LCA—
genetic testing is critical to the determination of the correct inheritance type,
because these diseases may have different inheritance patterns (AR, AD, or in
the case of RP, XL).
For Mendelian diseases, risks are calculated based on the proven
inheritance pattern. In the absence of an identified genetic mutation,
calculations may require more complex statistical methods. For example,
Bayesian analysis can be used to estimate residual risk when an individual is
suspected to have a Mendelian condition such as RP, but genetic test results
are negative.
Genetic Testing
Families and physicians have a variety of motivations for choosing to pursue
genetic testing for inherited eye diseases (24). Historically, these reasons
include confirmation of clinical diagnosis, medical management guidance,
and clarification of inheritance and resulting risk to additional family
members. Today, the most frequent reason patients and families cite for
pursuing genetic testing is potential qualification for gene-specific therapies
and research studies trials. The American Academy of Ophthalmology
supports genetic testing for all patients with a presumed or suspected
inherited, or Mendelian, retinal disease (25).
Clinical Diagnosis
While many inherited retinal conditions have distinct ophthalmologic features
that enable reliable clinical diagnosis, others have overlapping features that
make the diagnosis much more challenging. In these cases, genetic testing
may elucidate the diagnosis and reduce the need for additional
electrophysiology and/or serologic testing frequently used to narrow down
the differential diagnoses. In a young child, this may eliminate the need for
additional sedation or time spent waiting for features to develop.
In the pediatric ophthalmology or retina clinic, determining whether an
ocular condition is isolated or part of an underlying syndromic disease is
critical to providing appropriate care and prognostic counseling. For example,
some children with LCA will develop intellectual disability and/or kidney
disease, whereas most will not. Prior to the availability of comprehensive
genetic testing, many inherited retinal disease specialists recommended
baseline brain MRI testing and periodic renal function studies in all LCA
patients. With genetic testing, only those with mutations in genes that cause
these additional features need this surveillance. Similarly, some children who
present with cone dystrophy or achromatopsia have a syndromic disease
called Alström syndrome, which also predisposes to obesity and liver and
cardiac dysfunction. Children with retinal diseases due to mitochondrial
disorders may require cardiac monitoring; patients with Bardet-Biedl
syndrome may develop diabetes and renal failure. When the molecular cause
of disease can be identified, patients with high risk can be followed
appropriately, but if the mutated gene is known to be associated with
nonsyndromic disease, costly and unnecessary medical screening can be
avoided.
When genetic testing is ordered to rule in or rule out syndromic disease,
thoughtful pretest genetic counseling is critical. Families should be prepared
to face the possibility that their child could have a more extensive disease—
and much more significant health and cognitive consequences—than they
would with retinal disease alone. The choice of genetic test should also be
considered through this lens; genes associated with potentially fatal
conditions such as infantile or late infantile NCL (or Batten disease) are
frequently included on large genetic testing panels but should not be tested
without adequate pretest consent and discussion.
Medical Management
Genetic testing can also assist physicians and patients in pursuing appropriate
medical management. For example, if a patient has a clinical presentation of
cone dysfunction and genetic testing identifies causative mutations in the
ABCA4 gene, then supplementation with vitamin A would be contraindicated
(26). Genetic testing can also identify patients with childhood onset retinal
dystrophy who are at risk for choroidal neovascularization or cystoid macular
edema, as genetic mutations in certain genes, such as CRB1 and BEST1, are
known to be associated with those conditions (27).
With advances in understanding of genotype–phenotype correlations,
genetic testing continues to provide critical information for determining
effective medical management as well as clues to underlying disease
pathology.
Gene-Based Therapies
Recently, demand for genetic testing for inherited retinal diseases has
dramatically increased in response to several significant factors: the
development of gene-based therapies, improvements in genetic testing
capabilities, and reduced cost (with resulting increased access).
In 2017, the FDA approved Voretigene neparvovec–now known as
Luxturna® for the treatment of RPE65-associated retinal dystrophy as the
first US-approved gene therapy for the treatment of a genetic disease.
Therapies for other retinal diseases promise to follow, with gene-based
clinical trials in progress for some types of Usher syndrome, LCA, RP,
Stargardt macular dystrophy, and choroideremia (www.clinicaltrials.gov).
Genetic testing is a vital first step in the identification of patients as
candidates for gene-based therapies and clinical trials. Of course, many
patients will receive positive molecular test results but not qualify for a
current clinical therapy or trial. However, these patients and families can still
contribute to the development of future trials by enrolling in a patient registry
and/or engaging with a clinical research center. As the available database of
patients with known molecular diagnoses grows, clinical trial enrollment
becomes more efficient, and the likelihood of trial success increases.
Improvements in genetic testing methodologies have not only increased
the detection rate of these tests but have done so while decreasing cost. As a
result, patients with pediatric onset retinal diseases now have a very high
likelihood of receiving a positive molecular test result, identifying the cause
of their disease. In fact, in late 2018, the detection rate of genetic testing for
patients with a diagnosis of LCA was approximately 80%, compared with
<50% 5 years earlier (internal communications with clinical genetic testing
labs).
Inheritance and Risk
When genetic testing is successful in identifying the cause of disease, the
clinician can provide accurate risk estimates not only to family members but
also to couples seeking guidance about family planning decisions.
For many families affected with inherited retinal diseases, a genetic
diagnosis does not change their plans for future children. However, other
families choose to pursue genetic testing either before or during pregnancy.
Families who wish to have biologic children without the genetic condition
have two options for genetic testing: in vitro fertilization (IVF) with
preimplantation genetic diagnosis (PGD) or prenatal diagnosis through either
chorionic villus sampling (CVS) or amniocentesis. IVF with PGD allows a
couple to prevent an affected pregnancy by only implanting unaffected
embryos. To date, this option is still expensive and unavailable to many
couples. It also requires a molecular genetic diagnosis before beginning IVF.
If a couple chooses to pursue pregnancy naturally, testing is available in the
first trimester through CVS or after 15 weeks through amniocentesis. While
more affordable than IVF with PGD, both procedures have a small risk of
miscarriage, and the couple may be faced with deciding between terminating
an affected pregnancy or continuing the pregnancy with the knowledge that
the child will be affected. In all cases in which families are weighing
decisions about family planning, a prenatal genetic counselor should be
involved.
After the identification of the underlying genetic mutation(s), at-risk
family members may be interested in being tested. Presymptomatic genetic
diagnosis is appropriate in some cases but should be approached with caution
when testing at-risk family members for conditions where reduced penetrance
or a wide degree of variability in clinical symptoms is known to occur. For
some dominant or X-linked conditions, such as dominant RP or
choroideremia, parents may request genetic testing for an at-risk child. The
American College of Medical Genetics and the American Academy of
Pediatrics explicitly discourage routinely testing at-risk minors for adult-
onset conditions with no available therapy (28). Occasionally though, even
after a thoughtful discussion of this guideline, the family (or child) has strong
reasons for wanting to be tested presymptomatically. In these circumstances,
the consultation of a hospital ethics board may be appropriate. It is important
to note that this guideline for avoiding presymptomatic genetic testing in
minors applies only to conditions that are adult-onset and untreatable. Testing
for Usher syndrome, for example, in a newborn with a sibling who has Usher
syndrome may help guide surveillance and care of the child.
Selecting a Genetic Test

Genetic testing is now available for thousands of genetic diseases, and the
vast majority of this testing can be obtained through CLIA-certified clinical
laboratories. With an abundance of tests as well as laboratories, genetic
counselors can help select the most appropriate test for a patient.
Inappropriate test orders have accounted for millions of dollars in
misappropriated health care spending (29) and, in many cases, an
unnecessary extension of time invested for patients to obtain an accurate
diagnosis. As part of the discussion with the patient about family history, it is
important to determine if any other family members have previously
undergone genetic testing, since both positive and negative genetic test
results can help in choosing the most appropriate test. Genetic testing in
families becomes more straightforward after a mutation has been identified,
as testing can then be targeted to one or two genetic mutations. Mutation-
specific testing typically has a shorter turnaround time and is less expensive
than comprehensive genetic testing.
The process of selecting the most appropriate genetic test must
incorporate a variety of information. In diseases known to be caused by a
single gene, such as X-linked retinoschisis, testing may involve the
evaluation of a single gene. However, most inherited retinal diseases are
known to be associated with many different potentially causative genes. A
multicenter clinical working group convened by the American Academy of
Ophthalmology concluded in its guideline “Recommendations on Clinical
Assessment of Inherited Retinal Degenerations” that:
Multi-gene testing is typically necessary for the successful molecular diagnosis of a

disease such as retinitis pigmentosa, where >100 causative genes are known
(complete gene list available at sph.uth.edu/retnet/). Testing should include genes
known to be associated with syndromic forms of retinal disease, since some
patients presenting initially with only retinal disease may actually be affected with
an underlying syndromic condition, such as Batten disease. Other types of testing,
including single gene analyses, may be more appropriate for certain conditions. As
these technologies continue to evolve, clinicians are encouraged to work with
geneticists and/or genetic counselors to ensure appropriate genetic testing for their
patients (25).
“Multi-gene testing” typically refers to panel-based testing, which is usually

based on either next-generation sequencing or whole exome or whole-
genome technologies, but with interpretation of only the genes well known to
be associated with the disease of interest. Whole exome and whole genome
sequencing with interpretation of the entire exome/genome are not routinely
performed in clinical practice for the diagnosis of inherited retinal diseases,
although these technologies are becoming more pervasive and may become
the standard of care in the future.
Complexity of Genetic Test Results

A cornerstone of the growing precision medicine movement is the
incorporation of genetic testing data into medical decision-making. But how
can that be done if the results are unclear or the consequence of the identified
genetic variant is unknown? When ordering genetic testing, providers may
expect one of two simple results: positive or negative. As the number of
genes tested rises, however, the likelihood of obtaining this simple report
decreases significantly.
Although advances in genetic testing have dramatically improved our
understanding of these conditions and their underlying genetic causes, the
breadth of genes tested has also led to a high rate of reported “variants of
uncertain significance.” The American College of Medical Genetics has
established strict guidelines for determining the significance of a sequence
variant as either pathogenic (disease-causing) or benign; if the available
evidence is lacking or conflicting, the variant defaults to “uncertain
significance” (30). In 2020, most patients tested using a multigene “panel
test” of >100 genes will have multiple variants of uncertain significance
(VUS) identified. Additional complexity of genetic test results lies in the
identification of pathogenic variants not associated with the patient’s disease,
or incidental findings. Comprehensive genetic testing can identify incidental
pathogenic variants that may indicate a patient is a carrier for another retinal
disease or that they have a syndrome involving ocular disease; in these cases,
patients may require evaluation for additional medical concerns. It is possible
that pathogenic variants can be reported in several different genes, including
genes that can be associated with either dominant or recessive ocular
diseases. Because management-altering incidental findings can be reported
with large panel testing for retinal disease, pre-test counseling is of the
utmost importance so patients can understand the possibility of this result
type. When even broader tests, such as whole exome or whole genome
sequencing, are ordered, secondary findings can be reported in genes
completely unrelated to a patient’s ocular condition. Most laboratories allow
patients to choose to have medically actionable mutations reported alongside
the results of testing. The ACMG has generated a list of genes which should
be considered for reporting, even if mutations are found that are not
associated with the patient’s indication for testing; as of early 2020, this list
included 59 genes, referred to as the “ACMG 59.” Genes on this list mainly
cause cancer or cardiovascular phenotypes and have management guidelines
associated with them. For example, mutations in the breast cancer
predisposition gene, BRCA1, may be identified and could require further
screening and evaluation, although a patient was originally tested due to
ocular disease. Laboratories may vary on genes included in secondary
findings reporting, so each lab should be reviewed for their specific reporting
processes and testing options prior to testing. Patients should be encouraged
to consider how this pre-symptomatic test result may impact them – whether
the information would be helpful to them or might induce unnecessary stress
or anxiety. The discussion of secondary findings and whether they will be
reported in a patient’s test should be discussed extensively with the patient
prior to genetic testing. If patients choose to receive information on incidental
findings in genes not suspected to cause their condition, an appointment for
genetic counseling specific to this should be scheduled prior to testing.
These results can be very confusing to both the patient and ordering
provider. The AAO recommends that “genetic tests should be interpreted and
disclosed to the patient and family by a physician or genetic counselor who is
knowledgeable about inherited retinal disease, and who has the time to
discuss potentially sensitive and complex findings.” Incorporating a genetics
expert is in the patient’s and treating provider’s best interest, so that both can
receive the most accurate information from complex testing results.
Various strategies exist for evaluating uncertain genetic variants, in an
attempt to resolve their clinical significance and determine the likelihood that
they may be benign or pathogenic. Some of these strategies are employed
almost exclusively by diagnostic and research laboratories, such as
computational variant-effect prediction and functional assays. However, other
methods used to help evaluate the potential effect of a variant necessitate the
involvement of someone working directly with the family. Genetic
counselors frequently work with patients to obtain additional family member
samples for genetic testing, to establish phase and segregation. Researching
genes and variants, explaining complex results to patients, determining the
need for family testing, and following up with the family and the lab take
time that few ophthalmologists can devote in their clinical practice.
Genetic Privacy
As health care continues to expand and a variety of health care providers
incorporate genetic testing into their practice, some patients worry that this
increased use of genetic information leaves them more vulnerable. Many
patients fear that their genetic information or status will not be maintained
confidentially. To address privacy concerns, legislation has been enacted in
both state and federal arenas over the past 20 years. In 1999, President
Clinton signed an executive order prohibiting any federal agency from using
genetic information in any hiring, firing, or promotion action. Parts of the
Americans with Disabilities Act of 1990, the Health Insurance Portability and
Accountability Act of 1996, and Title VII of the Civil Rights Act of 1964
have also been interpreted to include some protection from genetic
discrimination. Several prominent medical and genetics societies supported
these orders, including the American Medical Association, American College
of Medical Genetics, National Society of Genetic Counselors, and the
Genetic Alliance.
Despite the best intentions of lawmakers, this patchwork of state laws left
many individuals unprotected from perceived, and sometimes real, threats of
discrimination. Clearly, federal legislation was needed to give the public
greater protection and security. Individuals felt that state laws were not
adequate in addressing genetic information specifically, such as genetic test
results or family history information. This was especially true for privately
purchased health insurance plans. In 2008, President Bush signed the Genetic
Information Nondiscrimination Act (GINA) into law (31). GINA provides
additional protection against genetic discrimination: This act more
specifically addresses the definition of genetic information and offers
protection against discrimination by private insurance companies. As such, it
prohibits health insurance plans from determining eligibility or adjusting
premiums based on genetic information. Still, GINA does not extend to life
insurance and long-term disability insurance. With the rise in direct-to-
consumer genetic testing, conversations about genetic privacy and data
ownership have once again emerged on a national stage. Clinicians who order
genetic testing would be wise to follow these discussions in order to address
patient questions.
GENETIC COUNSELORS IN
OPHTHALMOLOGY
With the increasing availability of gene-based treatments and clinical trials, a
larger number of ophthalmologists have become interested in genetic testing.
Even motivated specialists, eager to screen patients as candidates for gene-
based treatment and clinical trials, struggle to meet the demands of
incorporating genetic testing and genetic counseling into routine medical
practice. Comprehensive family history collection, risk assessment, and
discussion of genetic testing options and results require a significant amount
of time, which many clinicians do not have in their regular patient care
setting. Genetic counselors are specifically trained to address both the
educational and social needs of patients and their families and are, therefore,
becoming a more common addition to the clinical team in ophthalmology
either on site or through the use of telemedicine services.
A major component of the genetic counseling process is patient
education, which is crucial for empowering patients to understand their health
care as well as to become their own health advocates. In a 2018 study,
researchers examined the impact of genetic counseling on patients with
inherited heart disease. Using five different measures, they sought to
characterize how genetic counseling affected patients’ empowerment,
knowledge, and anxiety, as well as their relationship with their counselor and
satisfaction with their service. According to the authors, “Genetic counseling
was associated with significantly increased empowerment” (32).
Genetic counselors offer patients in-depth education about the inherited
condition, including an explanation of genes and inheritance patterns. This
often leads to a discussion of additional family members who could be at risk
for the genetic condition. Timing is key with patient education, as patients
may or may not be emotionally ready for complex genetic information at the
time of diagnosis; some would like immediate information, and others may
need weeks to months to process the news of their diagnosis and be ready to
take additional steps. When patients are connected with a genetic counselor,
the counselor can assess patient readiness and pursue patient education when
the timing is most appropriate.
Patients and families nearly always ask questions about long-term
prognosis. These questions are frequently complicated by disappointment
related to how the disease may impact previously held hopes and plans for
the future. Genetic counseling provides patients with both anticipatory
guidance and support in the process of adjusting to changes in expectations
due to a confirmed genetic diagnosis. Genetic counselors are uniquely trained
in short-term counseling related to illness and disability and excel in their
ability to provide patients with psychosocial support.
Many counseling issues can arise in a consultation with the family of a
patient diagnosed with a pediatric retinal condition, including feelings of loss
of independence for both children and parents. Many parents need assistance
in learning to adjust to caring for a child with visual impairment. Parents may
also need assistance in knowing how to discuss vision loss with their affected
child as well as other family members, care providers, and educators. Vision
loss and the psychosocial issues that can accompany this diagnosis are unique
and require a health care provider who is familiar with these issues and has
expertise in counseling patients and families with these conditions.
Genetic counselors also provide patients with contacts for appropriate
local and national support groups and organizations and offer patients support
materials for educating family and community members. Over time, genetic
counselors frequently check in with families to adjust support resources as
patient and family desires and coping abilities change. Additionally, genetic
counselors may utilize services of a social worker or other support person
who can offer longer-term counseling and support resources to the patient
and family.
There are psychosocial risks associated with the diagnosis of a genetic
disease and the disclosure of genetic test results. Patients and families
sometimes describe an emotional burden from learning that their family
members may also develop a genetic disease. Others experience intense
disappointment or anger when genetic testing does not indicate that the
patient will qualify for a clinical trial or therapy. Patients undergoing
presymptomatic genetic testing have reported that receiving abnormal test
results has changed their family structure; they often feel emotionally closer
to family members who have also received abnormal results or the same
diagnosis. Patients who receive normal test results have reported feeling
isolated, especially if they will be the primary caregiver for affected family
members (33). Patients should be supported through these complex emotions
and encouraged to consider and prepare for possible emotional reactions to
the results of testing.
In addition to the familial implications, patients who encounter a
diagnosis of a genetic condition may find themselves facing various ethical
challenges in assimilating the diagnosis into their lives. Perhaps the most
striking example of these challenges is presented in the decision to have, or
not to have, children. Quality of life issues and concerns may present when
examining the risks of having a child, whether it is a patient deciding about
having his or her first child or a couple deciding whether to have future
children. Ethical issues also arise when other family members at risk for the
same condition either do not want to know whether they have a genetic
mutation or simply do not want to share this information with others. In these
situations, the health care provider can be faced with the dilemma of
maintaining patient confidentiality when other family members may wish to
know their own risk. Careful, compassionate, and supportive genetic
counseling can help address the apprehensions of the individual while
ensuring he or she understands the full implications of nondisclosure.
For all of these reasons, patients receiving genetic test results deserve the
time of a trained professional who can explain the significance of the
findings, validate emotional responses, and identify individuals in need of
additional support. Clinical genetic professionals can be identified through
the National Society of Genetic Counselors, the American College of
Medical Genetics, or the American Society of Human Genetics.
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2. RetNet: summaries of genes and loci causing retinal disease. https://sph.uth.edu/RetNet/sum-
dis.htm#D-graph. Updated January 24, 2017. Accessed February 3, 2017.
3. Traboulsi EI, Koenekoop R, Stone EM. Lumpers or splitters? The role of molecular diagnosis in
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22
Anomalies of the Optic Nerve
Chrysanthi Basdekidou and George Caputo
TILTED DISC
Introduction
Tilted optic discs have been considered to be relatively small optic nerve
heads with an oblique or horizontal orientation and oval disc shape that occur
in eyes without high myopia (1–4).
A tilted appearance of the optic disc is a relatively frequent finding
during the course of an ophthalmic examination and forms one component of
the tilted disc syndrome, which includes the following signs (Figure 22-1):
FIGURE 22-1 A, right eye and B, left eye.Bilateral nasal
tilted disc syndrome, associated with inferonasal
staphyloma in a 20-year-old patient. Vision is 20/20 with a
corrected astigmatism.
1. Subnormal vision
2. Myopic astigmatism with oblique cylinders
3. Transposition or tilting of the optic disc downward and nasally
4. Inferior or inferonasal crescent, also called congenital conus
5. Nasal or inferonasal ectasia associated with thinning of the retinal
pigment epithelium and choroid
6. Bilateral temporal loss of visual field (5–8)
A number of reports (6,9–12) stated that more women than men had tilted
discs. Although many studies reported a high frequency of bilateral
involvement (9–12), there are a number of cases that remain misdiagnosed.
Vongphanit et al. (4) found that the tilted disc appearance was bilateral in
37.5% of cases, whereas Riise (10) reports a 74% rate of bilateral
involvement but he included “slighter cases…which would have been
described as normal in the usual ophthalmoscopy.”
Prevalence
In the majority of cases (80%), the tilted disc syndrome is bilateral. There
have been very few past reports of the prevalence of tilted discs. In a report
from a Norwegian practice series of 300 participants aged 7 years or older,
Riise reported that the tilted disc syndrome was found in five (1.7%) (10). An
identical figure was found for tilted discs in an Italian community of 300
participants, reported by Casteldacchia (13). In a German report from 1922
that included tilted disc illustrations, von Szily reported 3.4% of 32,500 eyes
from a clinic had a nontemporal (“heterotypical”) distribution of peripapillary
atrophy (14) described by some authors as the quintessential feature of the
tilted discs (10,15).
The prevalence in an Australian population was 1.6% and is comparable
to the rate of 1.7% reported by both Giuffre (11) and Riise (10). Given that
the tilted disc appearance varies considerably even between eyes within an
individual (10), the use of dysversion alone may lead to an underestimation
of its prevalence and bilaterality. Dysversion of the optic nerve head has also
been referred to as segmental hypoplasia and is characterized by central
retinal vessels emerging temporal to the vertical midline of the disc and being
directed nasally or the nerve head tilting in a vertical direction resulting in a
downward or oblique tilting of the discs with the blood vessels emerging at
the superior or inferior disc rim.
Tilted optic discs occur in about 4 out of 1,000 nonhighly myopic eyes of
adult Chinese (16). As in Caucasians (1,3), tilted discs were associated with
moderate myopia, astigmatism, inferiorly located beta zone of peripapillary
atrophy, decreased visual acuity, and visual field defects. One may infer that
a visual field defect in an eye with a tilted disc is not proof of an acquired
optic nerve disease, such as glaucoma.
Etiology
It is thought that the spectrum of signs present in any given eye is dependent
on the severity of the embryonic optic fissure malclosure at the 6-week stage
of gestation (8). The 6th week shows the incipient differentiation of the inner
layer of the optic cup into a sensory retina, the formation of the secondary
vitreous, the transformation of the posterior cells of the lens vesicle into
primary lens fibers, the development of the periocular vasculature, and the
appearance of the first eyelid folds and of the anlage of the nasolacrimal duct.
The spectrum ranges from a minimal defect as a tilted disc to more severe
fundus involvement, including inferonasal ectasia and frank coloboma (5).
However, some believe this assumption does not explain adequately the
origin of all ocular anomalies found in the syndrome like refractive errors due
to myopia, hypermetropia or corneal astigmatism, and various types of visual
field defects. Fewer retinal nerve fibers than normal enter the defective side
of the disc (15), whereas the opposite side presents fiber crowding, which
may give an erroneous impression of edema. Therefore, it has been suggested
that atrophy of the ganglion cell fibers of the inferonasal retina might occur
following an anomalous closure of the embryonic cleft. This in turn would
explain the hypoplasia of the inferonasal quadrant, the inferior crescent and
lack of fibers in the inferior optic disc, and the anomalous appearance of the
optic nerve head and of the retinal vessels with shifting of the disc upward
due to the imbalance between the ganglion cell fibers coming from the
superior and inferior retina (11). Furthermore, this shift accounts for the
upper temporal visual field defects mostly found in the syndrome. Kim et al.
(17) studied 107 eyes of 107 patients with tilted discs using coronal views of
swept-source optical coherence tomography (OCT). They were able to
correlate the amount of optic disc torsion with different types of posterior
sclera configuration and postulate that the main pathology was in the
posterior sclera rather than the optic nerve itself.
The heredity of the condition is not yet established perhaps because of the
variability in the phenotype in family members of affected individuals and
may account for the lack of reports of a familial nature of the syndrome.
Single clinical features like inferior crescent or transposition of the optic disc
have been reported as associated with various inheritance patterns (10). On
the other hand, Riise believes that the tilted disc syndrome can be familial
with a polygenic mode of inheritance similar to that of refraction anomalies
(10). Bottoni et al. presented a family with dominant penetrance of a tilted
disc (18). The tilted disc syndrome has also been reported in patients with X-
linked congenital stationary blindness (14). The tilted disc syndrome is
believed to be nonprogressive (5).
Clinical Appearance
The most frequent orientation of tilted discs is inferonasal (60%) (Figure 22-
1), thus tilting to the nasal side of the vertical meridian; however,
inferotemporal tilting has also been reported (28%) (5). The appearance of
tilting arises from the pseudorotation of the superior pole of the optic disc (5),
angulation of the optic cup axis inferonasally, and elevation of the
superotemporal neuroretinal rim (6). The optic disc configuration of tilted
discs can sometimes be confused with optic disc swelling, papilledema (6,8),
or presumed optic disc drusen (9), because the elevated retinal nerve fiber
layer (RNFL) appearance may cause blurring of the disc margin and visual
field changes. Although inferonasal tilting is most frequent, inferior tilting is
the most severe (4).
Best corrected visual acuity (BCVA) is reduced in eyes with tilted discs.
This reduction in BCVA is believed to be due to the Stiles-Crawford effect as
the photoreceptors stand obliquely in relation to the visual axis (10). Several
reports (6,8–10,12,19,20) revealed that a high proportion of people with tilted
discs had myopia and astigmatism with the plus axis parallel to the fundus
ectasia in the inferior quadrant. Corneal topography studies indicate that an
irregular curvature contributes to the associated astigmatism and suggests a
corneal origin of the astigmatism (21). Vongphanit et al. have quantified this
relationship and showed a strong association between the tilted disc
appearance and the level of either astigmatism (93.5%) or spherical refractive
error; myopia was present in 62% of subjects with tilted disc syndrome
compared to individuals with normal optic discs. Oblique astigmatism was
also present in over 50% of the cases (4). In addition, central corneal
thickness and corneal volume were found to be thinner in patients with tilted
disc syndrome than in normals (22).
Reduced visual acuity in association with bitemporal depression of the
visual field that is typically incomplete corresponding to the inferonasal optic
fiber loss (Figure 22-2) can suggest a lesion near the optic chiasm.
Recognition of the fundus abnormalities, possible improvement of the field
defect with proper refractive correction (10), and lack of the vertical
boundary at the midline of the visual field should obviate the need for a
major neurologic workup.
FIGURE 22-2 Nasal tilted disc syndrome, fundus
appearance (A), and OCT scan showing fiber-optic layer
thickening on the nasal inferior and superior portion of the
disc (B).
Many field defects have been described in association with the tilted disc
syndrome. The most frequent sites are superotemporal, temporal, and
superior (6,9,12), but nasal, inferior, and paracentral defects have also been
reported. Some studies have reported that the majority of patients with tilted
discs had a visual field defect, whereas other studies have indicated that field
defects were not consistently found (15,20). In some studies, visual field
defects were associated with myopia >6 diopters (8,23). Repeat perimetry
after addition of a −4 lens often eliminated the visual field abnormality,
confirming the refractive nature of this defect (6).
Visual field defects associated with “scooping out” of the optic disc have
been misdiagnosed as glaucomatous (Figure 22-3) (1,8). Bitemporal
hemianopia associated with tilted discs may also be confused with field
defects resulting from chiasmal compression leading to unnecessary
neuroradiologic investigations (1,10).
FIGURE 22-3 Tilted optic disc with marked central
excavation.
Certain features of the field defect help distinguish defects associated with
the tilted disc syndrome from other pathology. The defects tend to be relative
in nature (9,10) and nonprogressive (6,8) and may resolve with a larger test
stimulus or after optical correction (9,10). Defects are usually surrounded by
normal peripheral visual fields and frequently cross the vertical meridians of
the visual field (6,9).
Although not frequently associated with severe visual impairment, tilted
discs may be associated with choroidal neovascularization (7,11,19,20),
chorioretinal atrophy (19), and neurosensory macular detachment (2). Long-
term follow-up of 48 eyes with tilted disc and serous retinal detachment did
not show a benefit of treatment (24). Posterior staphyloma is commonly
associated (11,13) and may affect vision due to choroidal neovascularization
and chorioretinal atrophy (5). Anomalies at the junction of the staphyloma
may cause serous macular detachments or atrophy over time (Figure 22-4).
FIGURE 22-4 A, right eye and B, left eye.Same patient
as Figure 22-1, 10 years later showing marked pigmented
epithelial changes in the area of the inferior staphyloma
and peripapillary inferonasal atrophy.
Giuffre and Vongphanit in their respective case series described inferonasal

pigmentary accumulation in 11% of older patients with severe disc tilting
(4,11). Eyes with pigmentary accumulation were more likely to be myopic
and to have poorer visual acuity than eyes without pigmentary accumulation.
Although pigmentary accumulation could resemble myopic lacquer cracks
(10), in eyes with tilted discs, it is located in the inferonasal retina with a
coarse appearance presenting a circular pattern around the disc (25).
Conclusion
In children, detection of tilted discs may prompt refractive measurement and
correction to exclude corneal astigmatism and prevent amblyopia.
When tilted disc syndrome is accompanied by a visual field defect that
respects the vertical meridian, neuroimaging is recommended.
OPTIC DISC DRUSEN

Introduction
Optic nerve head drusen (ONHD) are a common, benign, congenital anomaly
of the optic nerve, which can rarely lead to decreased visual acuity (26,27).
ONHD are laminated calcified hyaline bodies arising from the prelaminar
portion of the optic nerve (26,28–30). They were first described clinically in
1868 by Liebrich (31) and histopathologically by Muller in 1858 (32). They
occur almost exclusively in the Caucasian population with a prevalence of
0.3%. They have a reported prevalence of 0.4% in children (33). Seventy
percent of drusen are bilateral and may occur as a dominant trait with
incomplete penetrance. Associations with retinitis pigmentosa and angioid
streaks have been described, but no other associated systemic features are
known. They are not associated with neurologic disease, yet may simulate
optic disc edema (ODE) (26,28–30).
It is thought that the formation of ONHD is caused by abnormalities in
axoplasmic transport because of a small scleral canal and later axonal
degeneration (34). Abnormal metabolism is thought to lead to the deposition
of calcium within optic nerve axonal mitochondria. The axons are disrupted
and mitochondria are extruded. Further calcium deposition in the
extracellular space leads to drusen (35).
Up to 87% of patients with drusen have visual field loss. Visual loss from
drusen can be acute or slowly progressive with nonarteritic anterior ischemic
optic neuropathy (NAION) being the most common cause of acute visual loss
in these patients (36). Peripapillary choroidal neovascularization can also be a
cause of acute visual loss.
Clinical Evaluation
In affected patients, the configuration of the optic nerve head is variable, and
the drusen may be visible on the disc surface or buried within the disc
(Figure 22-5). Differentiation of ODE from ONHD is crucial and can lead to
diagnostic uncertainty. Examination techniques commonly performed are
optic disc autofluorescence, fluorescein angiography, B-scan
ultrasonography, OCT, and CT scanning (26,37).
FIGURE 22-5 A, right eye and B, left eye. Bilateral
optic disc drusen in a 10-year-old girl. The nasal side of
the optic nerve is irregular.
Mustonen and Nieminen (38) reported that fundus autofluorescence of the

nerve head was present in 75% of 180 patients with clinically evident optic
disc drusen (Figure 22-6). In addition, fluorescein uptake with buried drusen
can be seen late in angiograms.
FIGURE 22-6 Color fundus (A) and autofluorescence (B)
in the right eye of a 20-year-old patient.
B-scan ultrasonography may detect drusen in 43% of patients with optic disc
swelling (39) (Figure 22-7). Frisen et al. (40) examined five patients with
high-resolution CT scan and noted calcified drusen had increased attenuation
within the optic nerve head (Figure 22-8). MRI performed in order to
exclude other optic nerve or brain anomaly was found to be a complementary
diagnostic instrument showing a surface disc transparency. However, MRI is
expensive, requires special techniques and considerable patient cooperation,
and is, therefore, unlikely to be the first choice of investigation in patients
with ONHD. B-scan ultrasonography, on the other hand, is rapid,
noninvasive, and inexpensive. Furthermore, B-scan ultrasonography has been
shown to be superior to autofluorescence and CT scanning (41).
FIGURE 22-7 Ultrasound of bilateral optic nerve drusen

appearing hyperreflective in a 7-year-old boy.
FIGURE 22-8 Calcified drusen visible in CT scan, in
patient of Figure 22-1.
Recently, there have been a number of reports using OCT to distinguish optic
nerve edema from ONHD that focused on measurements of the peripapillary
RNFL thickness (27,42–45) and direct visualization of the optic nerve head
as important features (27). RNFL thickness, especially in the nasal quadrant,
has been shown to be decreased in ONHD when compared with optic nerve
edema (27). Lee et al. using SDOCT detected 80.0% sensitivity and 88.9%
specificity for nasal RNFL thickness >78.0 mm (27). Johnson et al. reported
80% specificity and 70% sensitivity for nasal RNFL thickness >86 mm for
the differentiation of ONHD from optic nerve edema (46) (Figure 22-9). In
some patients with ONHD, photoreceptor changes also have been
documented (47). Using OCT, Savini et al. (43) identified the SHYPS,
subretinal hyporeflective space, a hyporeflective space located between the
sensory retina and the retinal pigment epithelium and choriocapillaris in ODE
patients. Johnson et al. (46) found a decrease in the mean SHYPS thickness
in ONHD patients compared with those with optic nerve edema. Thus, they
characterized the OCT appearance of optic nerve edema as an elevated optic
nerve head with a smooth internal contour and a SHYPS thickness under the
optic nerve head with a gradient taper away from the disc: An enlarged
basement membrane opening is observed in optic nerve edema. An irregular
surface of the disc creating a “lumpy bumpy” internal optic nerve contour
and a more abrupt taper of the SHYPS were suggestive of ONHD (47).
ONHD has a signal-poor core, with a hyperreflective margin, and is located
anterior to the lamina cribrosa; it is best seen with EDI-OCT or swept-source
OCT (48).
FIGURE 22-9 OCT scan of the head of the optic nerve

showing the drusen in the optic nerve and the elevated
nerve of patient of Figure 22-2.
Complications
ONHD have been reported to cause myriad local vascular complications,
including central retinal artery and vein occlusion and peripapillary choroidal
neovascularization (49).
The presence of bilateral ONHD causing choroidal neovascularization in
children is a rare occurrence with the earliest reported case in a 3-year-old
child (50). Peripapillary choroidal neovascularization can cause central vision
loss by subfoveal progression of the choroidal neovascularization, serous
macular detachment, or submacular hemorrhage (50). Various treatment
strategies have been advocated. Observation may be a viable option, but there
are insufficient data from studies comparing observation with reported
treatment modalities.
Laser photocoagulation was reported successful at 2 years by Delyfer et
al. (51) as a treatment for two patients with bilateral choroidal neovascular
membranes from ONHD and subretinal hemorrhages extending into the
macula. Photodynamic therapy with verteporfin (Visudyne; Novartis
Ophthalmics, Basel, Switzerland) has also been used. Silva et al. (52) were
the first to report successful treatment of bilateral choroidal
neovascularization associated with ONHD in a 10-year-old girl. After two
sessions of photodynamic therapy, no recurrences were noted after 2 years of
follow-up. Surgery is another treatment option. Mateo et al. (53) surgically
removed choroidal neovascular membranes in four eyes with ONHD, all of
which showed significant visual recovery without any recurrence during 12
to 42 months of follow-up. Antivascular endothelial growth factor (VEGF)
agents are now commonly used to treat adults with choroidal
neovascularization from a variety of causes (54). Their use in children has
been reported in Coats disease (55), Best disease (56), and coloboma-related
choroidal neovascularization (57,58). Knape et al. presented a case of
bilateral choroidal neovascular membranes associated with ONHD in a 5-
year-old boy who was successfully treated with a combination of focal laser
photocoagulation and intravitreal bevacizumab (59). Until the safety of anti-
VEGF therapy has been established in the pediatric population, consideration
of these medications should be reserved for select cases of choroidal
neovascularization threatening central visual acuity.
NAION may occur at an unusually young age in patients with optic disc
drusen; it has been described in a 13-year-old child and in several patients in
their later teens and early twenties (36,60–62). The visual outcomes in these
patients vary, although they are slightly better than for other NAION patients.
Some patients do well clinically (60,61). Proposed mechanisms of vision loss
include distortion of blood vessels by drusen (36). Systemic hypotension,
dehydration, and altitude have been suggested as contributory factors for
NAION in the setting of ONHD and may provide an explanation for why
patients often discover symptoms upon awakening (63). No effective
prevention or treatment for NAION is known. The risk of contralateral
involvement is estimated around 15%.
Conclusion
ONHD, despite being a benign condition, have been reported to cause a
number of local vascular complications, including central retinal artery and
vein occlusion, peripapillary choroidal neovascularization, and NAION.
Patients should be aware of these risks and seek ophthalmologic follow-up.
We recommend that parents of children with optic disc drusen be instructed
about the risk of NAION and potential contributory factors, such as
dehydration and altitude, to help prevent vision loss in their children.
OPTIC DISC PIT

Introduction
An optic disc pit is a congenital anomaly of the optic nerve frequently
associated with macular detachment (64–66). Pits are usually small, well-
demarcated, temporally located depressions in the nerve substance (Figure
22-10). They may, however, be situated in any sector of the optic nerve head
(65). They are sometimes categorized as a subset of atypical optic nerve
colobomas (67). In an Indian population of 6,013 people over 40 years of
age, Bassi reports a prevalence of 0.05% (68). Although typically unilateral,
bilateral optic pits can be seen in 15% of cases (69).
FIGURE 22-10 Optic disc pit in a 4-year-old boy.
Familial cases of optic pits suggest a dominant inheritance pattern (70,71).

They have been associated with iris coloboma (72). Optic nerve pits rarely
portend additional malformations in the central nervous system (CNS) (73).
Acquired pits have been associated with high myopia (74). Their presence
among patients with glaucoma constitutes a risk factor for progressive optic
disc damage and visual field loss (75) as well as an abnormal susceptibility of
the optic nerve to damaging effects of intraocular pressure (76).
Histopathologic analysis of optic disc pits reveals perineural herniation of
poorly differentiated retinal tissue combined with vitreous collagen around
the optic nerve into the subarachnoid space (77).
Visual acuity is usually normal. The most common visual field defect is a
paracentral arcuate scotoma associated with an enlarged blind spot (65).
Acquired visual loss is usually due to the development of maculopathy.
Serous macular elevations have been estimated to develop in 25% to 75% of
eyes with optic pits (65). The risk is higher in eyes with larger and temporally
located pits (Figure 22-11).
FIGURE 22-11 A: Large optic disc pit in a 10-year-old
girl. Persistent maculopathy leaving her a 20/100 vision.
Temporal atrophic and pigmented peripapillary scars of
laser photocoagulation. OCT scan shows schisis
appearance of retinal layers (B); localized serous
detachment is seen on the superior border (C).
The Optic Pit Maculopathy

Optic pit maculopathy is defined by the presence of subretinal fluid (SRF),
intraretinal fluid (IRF) with cystic cavities, and multilayered retinal fluid
(MRF) when present in all retinal layers (78). Visual loss is most severe
when patients present with SRF. When disc pit maculopathy is established,
the natural history tends to be poor with most vision loss occurring in the first
6 months after diagnosis in case of SRF (79), although a recent study by Steel
found functional stability in 64% of the cases with 17% having improvement
on OCT at 1 year. However, after 1 year follow-up, the presence of SRF was
associated with worsening and those without SRF with improvement. The
spontaneous improvement rate dropped to 7% in eyes with SRF versus 30%
without SRF, and progression was observed in 27% of cases with SRF versus
9% without (79). Even if spontaneous resolution of maculopathy does occur
in 17% to 25% of cases (78), vision often remains poor with the development
of outer layer lamellar holes, external retinal changes, and pigment
epitheliopathy. True macular holes are rare (80).
The pathogenesis of optic disc pit maculopathy remains unclear.
Fluorescence of the area of the disc pit occurs late in fluorescein
angiography, and there is no leakage into the submacular fluid. Initially, this
was interpreted that the source of macular fluid was not leakage from vessels
within the pit itself (81). Cerebrospinal fluid (CSF) pressure is normally
lower than intraocular pressure so that flow would normally be expected to
occur from the eye to the CSF down a hydrostatic gradient. However, the
possibility that CSF could transudate across the fibrous capsule into the retina
to cause maculopathy could not be excluded (82). Intracranial pressure and
intraocular pressure both fluctuate, although intraocular pressure is usually
greater than intracranial pressure. When intraocular pressure is greater,
vitreous fluid could move into the pit sac. When intracranial pressure is
greater, depending on the body position, for example, some fluid in the sac
may be pushed back into the eye through the pit. However, some fluid in the
sac also could move into the retina, leading to a retinoschisis- like separation.
Studies in eyes with optic pits using high-resolution OCT pictures
identified a membrane crossing the cup of the optic nerve, which may
represent neuroectodermal or astroglial tissue (82) (Figure 22-12). When
present, the membrane appears to originate from the edges of the optic disc
cup and traverses the cup to join the main vessels at the nasal aspect of the
disc or centrally if the vessels emanate centrally from the disc. The thin
membrane is concave and roughly follows the contours of the cup. A
separation is seen between the membrane and the base of the cup, and fine
trabeculae of tissue can sometimes be seen traversing the space, or the space
may appear to contain some poorly defined matter.
FIGURE 22-12 OCT of optic pit maculopathy: extended
schisis-like appearance in outer nuclear layers.
The protective nature of this membrane was suggested by the presence of an

intact membrane in patients without maculopathy (82). OCT studies have
suggested a communication between the schisis cavity or subretinal space
and the disc pit in patients with maculopathy, whereas the retinoschisis-like
retinal separation was most prominent in the outer nuclear layers and
frequently combined with multiple shallow inner retinal separations (83,84)
(Figure 22-13). It has been suggested that a defect in the membrane
overlying the disc pit allows fluid to enter the retina and cause a schisis-like
separation of the retinal layers. True macular detachment can then occur as a
secondary event but can also occur in the absence of associated retinal
thickening and fluid within the retinal layers (83). Katome et al. observed
with swept-source OCT a case of communication between the optic pit and
the subarachnoid spaces in the optic nerve (85).
FIGURE 22-13 OCT scan of the optic nerve of Figure
22-3 patient showing prepapillary glial material.
A role for vitreous traction has also been postulated from the observation that
maculopathy becomes symptomatic in the fourth decade of life and that it can
resolve after spontaneous posterior vitreous detachment (PVD) (86). Bonnet
previously reported that none of the eyes with macular detachment in their
series had evidence of a PVD and that the two eyes that demonstrated
spontaneous reattachment did so after a PVD developed (86). In addition,
Theodossiadis et al. described vitreous abnormalities, such as vitreomacular
traction and vitreous strands over the optic disc associated with optic disc pit
maculopathy (87). In their study, Hirakata et al. provided equivalent evidence
that vitreoretinal traction is an important factor in the pathogenesis of optic
disc pit maculopathy and that induction of PVD relieving any anteroposterior
traction on the macula is effective in reversing macular elevation without the
need for laser treatment (88). During induction of a PVD, a tight adhesion of
the posterior hyaloid or abnormal membrane, such as anomalous Cloquet
canal, to the margin of the disc pit has been reported (89,90).
When there is posterior hyaloid traction caused by age-related vitreous
liquefaction or trauma, it pulls on the optic disc pit. Because the pit is pulled
up like a tent by vitreous traction, the turbulent flow may have increased
access around the vessels to the intraretinal space. This traction affects the
peripapillary retina, especially around the pit. This further facilitates IRF
accumulation. When a PVD occurs, the anterior traction is released. The pit
decompresses and moves downward, and access to the subretinal space
becomes limited. Any traction on the peripapillary retina also is released.
Treatment
Patients normally seek ophthalmologic evaluation because of diminished
visual acuity or the discovery may be made incidentally during a routine
ophthalmologic assessment. When an optic pit is detected, performing an
OCT can be very useful in order to detect the presence of a PVD, a schisis-
like separation, and/or SRF. In cases with severe maculopathy associated
with vitreomacular traction, vitrectomy allows release of the vitreoretinal
traction and should be routinely performed. The ablation of the fibrotic
prepapillary material, the peeling of the internal limiting membrane, and the
peripapillary photocoagulation can be performed but remain controversial
and haven’t proven anatomical and functional benefit (79). The use of a gas
tamponade agent may not be necessary for success (88) but is generally
associated. In fact, subretinal migration of gas or silicone oil has been
reported (91,92). Theoretical calculations suggest that the pressure
differential required for migration of gas through a small defect in the roof of
a cavitary disc lesion is within the range of expected fluctuations in CSF
pressure. Explanations should be given concerning the nature of this
pathology and the visual acuity outcome.
Conclusion
Optic disc pits are usually unilateral, sporadic, and not associated with
systemic disease. The etiology of the associated serous maculopathy still
remains unclear. It could be suggested that the unusual movement of fluid
between the vitreous cavity and subarachnoid space and the traction around
the posterior hyaloid may contribute to the pathogenesis of this disease.
Vitrectomy with induction of a PVD without a gas tamponade or laser
photocoagulation allows resolution of optic disc pit maculopathy in most
cases.
OPTIC NERVE COLOBOMA
Introduction
Colobomas of the optic nerve result from incomplete or abnormal closure of
the primitive embryonic fissure (93). In certain instances, the defect may
extend to involve the choroid, retina, ciliary body, and iris. The term
“coloboma” derives from Greek and means “defect” (from the adjective
“colobos” that means “mutilated” or “cut short”).
Optic disc coloboma comprises a clearly demarcated bowl-shaped
excavation of the optic disc, which is typically decentered and deeper
inferiorly (94). The coloboma occupies the lower part of the optic nerve head.
The neuroretinal rim is absent inferiorly but is usually identifiable superiorly
(Figure 22-14). In cases in which the adjacent inferior retina and choroid are
deficient, microphthalmia may also be evident (95). Optic disc colobomas
may appear as large excavations of the nerve head up to 25 diopters deep and
involving nearly all of the disc, which may be enlarged (96).
FIGURE 22-14 Optic nerve coloboma in a 16-year-old
patient having lost her left eye by retinal detachment.
Macular pigment is visible on the edge of the coloboma
and vision is 20/100.
Optic nerve colobomas are often unilateral; bilateral colobomas are

frequently asymmetrical (96). Significant refractive error and anisometropia
are common. The only feature that relates to visual outcome is the degree of
foveal involvement by the coloboma (97). The size of the coloboma, the
color of the optic nerve, and the presence of subfoveal pigment change are
not related to visual outcome (Figure 22-15).
FIGURE 22-15 A, B: Bilateral optic nerve coloboma
sparing the macula in both eyes. Vision is 20/20 in this 18-
year-old patient.
Most colobomas are sporadic, although an autosomal dominant hereditary

pattern and, less frequently, an autosomal recessive or X-linked recessive
pattern have been reported (96,98). The condition can occur in association
with multiple congenital abnormalities indicative of an insult to the
developing fetus during the 6th week of gestation (Figure 22-16).
FIGURE 22-16 An 18-month child presenting with
bilateral microphthalmia and coloboma. A: Immature iris
and inferior coloboma. B, C: Microspherophakia with
extended ciliary processes and posterior persistent fetal
vascularization (fibrous stalk) attached to the large
chorioretinal coloboma. D: Chorioretinal coloboma of the
fellow eye.
There is a wide range of associations. These have been reviewed by Brodsky

(95) and include the CHARGE (coloboma of the eye, heart defects, atresia of
the nasal choanae, retardation of growth and/or development, genital and/or
urinary abnormalities, and ear abnormalities and deafness) association,
Walker-Warburg syndrome, Goltz focal dermal hypoplasia, Aicardi
syndrome, Goldenhar syndrome, and linear sebaceous nevus syndrome. More
recently, associations with Dandy-Walker malformation (99) and renal
coloboma syndrome with a mutation of PAX2 transcription (100–103) have
also been described. In the latter, the renal disease may vary significantly
from patient to patient. Associated abnormalities include small dysplastic
kidneys, vesicoureteral reflux, and high-frequency hearing loss. Although the
renal disease may be quite variable, all patients identified with mutations in
the PAX2 gene have been observed to have colobomatous optic nerves. Iris
colobomas have not been documented with PAX2 mutations.
Expression of PAX2 is restricted to cells of astrocytic lineage both during
retinal development and in adulthood. Using immunohistochemistry, it has
been found that adult retinal cells with the antigenic phenotype present in
mature perinatal astrocytes are found only in the region surrounding the optic
nerve head and that astrocyte precursor cells expressing PAX2 are found in a
small region surrounding the optic nerve during early development. It has
been argued that these findings suggest that coloboma formation may be
associated with impaired astrocyte differentiation during development (104).
Patients with optic nerve colobomas due to mutations in PAX6 have also been
reported (105). MRI scanning is recommended for patients with unilateral or
bilateral optic nerve colobomas.
The association of optic nerve coloboma with dysgenesis of the internal
carotid artery and transsphenoidal encephalocele with hypopituitarism has led
to the suggestion that the link between these malformations is abnormal
neural crest cell development (106).
Recently, more details came to light concerning the anatomical aspect of
the coloboma; high-resolution OCT scanning has detected a membrane
overlying the disc coloboma of some patients without maculopathy and
probably represents the neuroectodermal lining of the coloboma (82).
Discontinuities of Bruch membrane and retinal pigment epithelium at the
margin may allow subpigment epithelial and choroidal vessels to enter the
subretinal space at this junction. T1 MRI confirms that the intracranial
portion of the optic nerve is reduced in size (107).
Coloboma-Related Complications
The presence of an optic nerve coloboma implies some possible
complications. A small proportion of cases are associated with cysts arising
from the optic nerve sheath, which communicate with the subarachnoid space
(108). Rarely, such cysts can enlarge and lead to compressive optic
neuropathy (109). Peripapillary choroidal neovascularization has been
described in association with optic nerve coloboma (110,111). Studies have
shown good response of this type of neovascular membrane to treatment with
intravitreal bevacizumab (57).
Retinal detachment occurs in approximately 45% of patients with optic
nerve coloboma (66) (Figure 22-17). The source of the SRF is not known but
could derive from fluid entering the retrobulbar space from the surrounding
orbital tissue, from the choriocapillaris, or from the CSF. In contrast to
retinochoroidal colobomas, rhegmatogenous detachment is probably not a
recognized association (112). The retinal detachments associated with optic
nerve colobomas typically are more extensive and more bullous than those
caused by optic nerve pits, which affect only the macular area and are often
accompanied by schisis. The pathophysiology appears to be similar in both of
these conditions with a defect in the abnormal papillary tissue allowing fluid
to communicate to the subretinal space and detach the retina. In patients with
an optic nerve head coloboma, the extent of abnormal tissue is much greater
than it is in optic pits. This papillary defect along with a poorly differentiated
retina and retinal pigment epithelium, an abnormal lamina cribrosa, and an
enlarged scleral canal potentially allows the vitreous cavity, subretinal space,
subarachnoid space, and orbital space to communicate in these patients (113).
If a defect develops and liquefied vitreous has access to the hole, then the
retina detaches. Circumferential intrascleral smooth muscle has been
observed histologically (114) and may account for the rare observation of
spontaneous contractility of the colobomatous optic disc (115). Spontaneous
reattachment however may occur (116).
FIGURE 22-17 Retinal detachment complicating a large
optic nerve coloboma in a 6-month-old boy. The presence
of folds including the macula and the pigment epithelial
changes favor the hypothesis of a congenital retinal
detachment.
Retinal detachments secondary to optic nerve head colobomas are notoriously

difficult to repair. Numerous surgical approaches have been used, including
(in various combinations) vitrectomy, limited laser to the optic nerve head
margin, long-acting tamponade, peeling of the posterior cortical vitreous, and
scleral buckling. Despite varied surgical approaches, most of these retinal
detachments recur and lead to loss of vision in the affected eye (117).
Treatments, including patching and mannitol, the administration of steroids,
diathermy, xenon, or laser photocoagulation, have been described with
variable results (118). Treatments for macular hole have also been considered
for optic nerve coloboma and retinal detachment. Injection of autologous
platelet concentrate has been used to treat macular holes since 1990, but it has
fallen out of favor in recent years. Agents that have been used to stimulate
glial cell proliferation and promote macular hole closure include transforming
growth factor beta (118,119), autologous serum (120), autologous plasma and
thrombin, and autologous platelet concentrate (121,122). More recently,
platelet concentrate has been deemed an effective long-term treatment for
children with a traumatic macular hole after vitrectomy (123).
Conclusion
Chromosomal analysis should be considered in children with optic disc
colobomas as well as MRI imaging in both unilateral and bilateral cases. A
trial of patching may result in improvement of vision in the child presenting
early in life, and optimal refractive correction may be indicated. OCT
imaging assists with understanding the contributing factors to retinal
detachment in individual cases of colobomatous optic disc anomalies and can
thereby assist with determining the most effective approach to management.
THE MORNING GLORY DISC

ANOMALY
Introduction
Morning glory syndrome is a rare congenital anomaly of the optic disc
manifesting as an enlarged and excavated papilla with an elevated peripheral
pigmented choroid and a spoke-like arrangement of retinal vessels emerging
at the disc margin. First described by Handmann in 1929 (124), this unusual
congenital anomaly of the optic disc was reported in 1970 by Kindler and
was termed “morning glory syndrome” because of its similarity to the
morning glory flower (125). Now called morning glory disc anomaly
(MGDA), it consists of an enlarged, funnel-shaped, excavated disc
surrounded by an annulus of chorioretinal pigmentary disturbance (Figure
22-18).
FIGURE 22-18 Morning glory anomaly in a 3-year-old
girl: vision is limited to hand movements. Converging
retinal folds include the macula, and glial tissue is visible
in the center of the disc. Numerous radial vessels emerge
from the latter.
Usually an isolated anomaly, some cases have also had persistent

hyperplastic vitreous, ciliary body cyst, congenital cataract, lens coloboma,
strabismus, nystagmus, orbital hemangiomas, and midline craniofacial
defects such as basal encephalocele, hypertelorism, cleft lip and palate, or
agenesis of the corpus callosum (126–130). Associations with renal
anomalies, reversible and irreversible carotid artery narrowing, and
hypopituitarism have also been reported (94,131,132).
Some patients with MGDA have been reported to have Moyamoya
disease (94,129,133,134). These reports have stressed the importance of
neuroimaging for patients with MGDA because of the potentially severe
consequences of intracranial vascular anomalies such as Moyamoya disease
(126,128,129,135). The prevalence of cerebrovascular anomalies in children
with MGDA, however, remains unknown.
Etiology
The etiology of the anomaly is unknown, and no hereditary tendency is
evident. The association with midline craniofacial anomalies has been
attributed to a developmental defect in embryogenesis during the second
gestational month (129).
The pathogenesis of this condition is unknown. One hypothesis is that the
abnormality results from failed closure of the fetal fissure, similar to what
occurs with optic nerve coloboma. Another theory suggests that there is an
error in mesenchymal differentiation that results in abnormal closure of the
scleral wall and lamina cribrosa (136). Recently, mutations in the PAX6 gene,
which is involved in ocular morphogenesis, have been identified in patients
with MGDA (105).
Clinical Presentation
MG discs are more common in females and rare in black individuals. Most
cases are unilateral. Visual acuity is usually poor in the affected eye and often
with associated amblyopia, although there are rare reports of cases with good
visual acuity (128,133,137). Several bilateral cases have been reported
(124,138).
The clinical diagnosis of the MGDA is established primarily by
ophthalmoscopic examination. The disc is markedly enlarged; it is orange or
pink and appears to be elevated centrally within the confines of a funnel-
shaped peripapillary excavation (138). A wide annulus of peripapillary
excavation surrounds the disc. The blood vessels appear to be increased in
number and often arise from the periphery of the disc. Close inspection
occasionally reveals the presence of small peripapillary arteriovenous
communications (139) (Figure 22-18). The macula may be incorporated into
the excavation (macular capture) (124).
There are some additional tests that can provide further information, such
as visual field examination, visual evoked potentials, and B-scan
ultrasonography (140).
Baer et al. (141) described the first OCT finding in an atypical MGDA
patient with peripapillary excavation lined by retinal pigment epithelium in
2003 but provided no quantitative details. Srinivasan et al. (142) recently
reported quantitative OCT data of a 25-year-old woman with MGDA. They
provided the evidence of increased RNFL thickness and reduced macular
thickness.
Wu et al. reported the OCT analysis of a case of a 4-year-old boy with
MGDA (140). The optic disc OCT revealed exceedingly high cup/disc ratio
in the affected eye. In addition to the greater average thickness of the RNFL,
the temporal RNFL thickness was much greater than the inferior (196 μm vs.
157 μm). This finding is contrary to the “ISNT” rule in normal optic discs, in
which normal disc thickness becomes thinner from inferior to superior to
nasal to temporal (143). Srinivasan et al. (142) also noted this interesting
finding.
Contractile movements occur in morning glory discs (138). Two main
mechanisms have been proposed to explain this phenomenon: pressure
balance (144) and muscular contraction (138,139,145). The muscular
contraction theory was suggested (139,144,145) by several authors and later
supported by the discovery of heterotopic smooth muscle cells in the sclera of
an eye with an optic disc coloboma (146). This explanation is further
supported by the finding of nonvascular contractile smooth muscle cells in
the posterior choroid and sclera of normal eyes (147,148). These cells have
been colocalized with sympathetic nerve terminals in the choroid and
nitroxidergic parasympathetic terminals in the sclera (147) and have proven
to be functionally contractile on immunohistochemical staining for
smoothelin, which is considered to be present only in contractile smooth
muscle cells (148). In light of this, the actions of such contractile cells might
be clear in the excavated and colobomatous eye but might not be evident in
the eye with normal scleral coverage. In addition, the transient monocular
visual loss observed in patients with good visual acuity (139,144,149) further
supports the action of intermittent muscular contraction, which might
contribute to the intermittent compression around the optic nerve.
Lee et al., using computerized analysis, demonstrated contractile
movement mainly around the optic nerve margin in a horizontal direction
(150). Although these findings do not confirm a definitive mechanism, the
authors support smooth muscle contraction for the pathogenesis of the
contractile movement. This observation might be explained by the
distribution patterns of the nonvascular smooth muscle cells, in which a
group of cells were arranged in a semicircular pattern around the optic disc
and another group of cells were located in the foveal region of the temporal
quadrant (148).
Complications
In morning glory syndrome, the peripapillary retina extends into the
anomalous peripapillary scleral defect. The retinal tissue within the defect has
been observed to be thinner, incompletely developed, and atrophic.
Sometimes, the retina is detached in this area.
The pathogenesis of retinal detachment in morning glory disc and the
origin of SRF from vitreous cavity (151–157) or subarachnoid space
(158–160) remain controversial (152,155). Rhegmatogenous retinal
detachment has been reported in 37% of cases and is often confined to the
peripapillary retina (161) (Figure 22-19).
FIGURE 22-19 Inferior and temporal retinal detachment
complicating morning glory anomaly in a 7-year-old girl.
Pigment epithelial mottling is visible at the surface of the
retina confirming the rhegmatogenous mechanism.
Akiyama and colleagues postulated that the origin of the SRF was from the
anomalous disc and most probably from a break on the disc margin (155). Ho
et al. (162) in a series of five eyes used OCT to demonstrate subtle slit-like
breaks at the margin of excavation. Bartz-Schmidt and Heimann (151)
demonstrated a communication between the subretinal space and vitreous
cavity resulting in rhegmatogenous retinal detachment.
In the absence of a retinal break, spontaneous resolution of retinal
detachment with MGDA can occur and should be considered before planning
surgery (153).
Subretinal neovascularization may occasionally develop within the
circumferential zone of pigmentary disturbance adjacent to a morning glory
disc (163).
Conclusion
Since this anomaly is associated with amblyopia, a trial of occlusion therapy
is warranted in small children. Patients with MGDA should undergo
neuroimaging because of the potentially severe consequences of intracranial
vascular anomalies such as Moyamoya disease. OCT can play a role in the
diagnosis of MGDA and its associated ocular complications (162). It
provides more detailed information of the configuration of this congenital
anomaly, thus offering a better way to understand and follow the
pathophysiologic change.
OPTIC NERVE HYPOPLASIA

Introduction
Optic nerve hypoplasia (ONH) is one of the more common congenital
anomalies causing visual impairment in the Western world (164). The first
description of ONH is ascribed to Briere in 1877 (165), but the first artistic
rendering of the optic disc appearance was by Schwarz in 1915 (166).
ONH is a nonprogressive congenital abnormality of one or both optic
nerves associated with a diminished number of axons in the involved nerve(s)
with normal development of supporting tissues and the retinal vascular
system. It is histologically characterized by a subnormal number of optic
nerve axons and is an unprogressive, nonspecific manifestation of damage at
any site of the visual pathways sustained anytime before its full development
(167). It gives rise to varying degrees of vision loss ranging from minimal
visual impairment with almost any type of visual field defect to total
blindness.
The prevalence of ONH in Sweden quadrupled between 1980 and 1999
to 7.1 per 100,000, while all other causes of childhood blindness declined as
diagnosed in the same major ophthalmic center (164). In 2006, the prevalence
of ONH in England had risen to 10.9 per 100,000 children (168). In 2007, the
Babies Count registry reported ONH as the third most prevalent cause of any
vision impairment in children aged 3 years or younger in the United States
behind cortical vision impairment and retinopathy of prematurity (169). Of
all conditions, ONH was the most likely to cause legal blindness.
Although typically bilateral, it may be unilateral in 15% to 25% cases
(170). Incidence is higher in boys than in girls (171). Prenatal and perinatal
risk factors include maternal diabetes, premature birth, young maternal age,
primiparity, maternal weight loss, early gestational vaginal bleeding, and fetal
alcohol syndrome (171). Several recent reports have demonstrated a high
frequency of associated CNS abnormalities in children with ONH, indicating
the complexity of this diagnosis (172–174). In later reports, bilateral ONH far
exceeded unilateral ONH (175). It was also stated that unilateral ONH was
less commonly associated with other CNS lesions.
Up to 60% of cases may have associated structural brain malformations,
such as absence of septum pellucidum and agenesis or thinning of corpus
callosum as in septo-optic dysplasia (SOD) or de Morsier syndrome, cerebral
hemisphere abnormalities, schizencephaly, leukomalacia, and
encephalomalacia (176). The group of conditions of bilateral ONH, absence
of the septum pellucidum, agenesis of the corpus callosum, dysplasia of the
anterior third ventricle, and hypopituitarism is referred to as SOD. The first
recognition of an association of ONH with agenesis of the septum pellucidum
was by Dr. David Reeves at Children’s Hospital Los Angeles in 1941 (177)
(Figure 22-20).
FIGURE 22-20 A: Orbit MRI scan of patient with left
optic nerve hypoplasia. B: MRI scans of a patient with
right optic nerve hypoplasia and septo-optic dysplasia.
Yellow arrows show hypoplastic optic nerve appearance;
orange arrow shows the absence of pituitary stalk,
posterior pituitary dysplasia, and anterior hypoplasia.
(Courtesy of Dr. Monique Elmaleh.)
Endocrine anomalies have been reported in 6% to 71% of patients with ONH

(178). Growth hormone (GH) is often the earliest, and therefore the most
common, pituitary hormone affected (172). The reported proportion of SOD
and GH insufficiency in association with ONH ranges between 13% and 53%
(179,180). Hypopituitarism has been reported to occur in 75% of patients
with ONH, many of whom have no pituitary abnormalities on neuroimaging.
Central hypothyroidism in these children is of particular importance because
of the developmental impairment that may be associated with this diagnosis;
inadequate cortisol response during acute illness or surgery may be life-
threatening (181). Garcia-Filion et al. reported that 93% of children with
ONH and hypothyroidism have developmental delay (176).
Based on the high incidence of structural brain malformations, which
may be associated with an increased risk of endocrinopathy and
developmental delay, most authors agree that neuroimaging of the brain is
advisable in all patients with ONH (178). When available, MRI of the brain
with thin cuts through the sella turcica should be performed while using
gadolinium contrast. Endocrinologic evaluation is necessary in patients with
anomalous neurohypophysis or clinical evidence of endocrine deficiency, but
controversy exists with regard to testing in asymptomatic children with
normal neuroimaging studies.
Etiology
A lesion that causes a reduction of the total number of retinal ganglion cells
before the supportive tissues are fully developed in the first to second
trimester may result in a small disc because the supportive structures may still
be able to adapt to the subnormal size of the nerve tissue of the optic
disc/nerve. Correspondingly, a “late” lesion in the third trimester might result
in a normal-sized disc with a large cup, because the supportive elements have
reached their full size, creating a normal-sized disc, whereas the degeneration
of nervous tissue creates a loss of substance identified as a large cup (182).
There is interest in the theory that ONH represents an exaggeration of the
normal process of axonal cell death that occurs in the developing visual
pathways. Further investigation of the development of the normal optic nerve
is necessary before some of the puzzles that surround the pathophysiology of
ONH and optic atrophy can be unraveled.
The cause of ONH is probably multifactorial. Differentiation of the
retinal ganglion cells begins at 6 weeks of embryonic life, and it has been
suggested that a failure of this differentiation is the cause of ONH.
Consequently, the lesion would be induced before the seventh week of
gestation (183). However, a recent report demonstrates an important role of
the homeobox gene in the development of SOD, which indicates that ONH
might develop even earlier than previously thought (184). In addition, the
frequent association between ONH and other CNS anomalies suggests that
another explanatory factor for ONH may be a secondary degeneration of
ganglion cells and their fibers (185). Depending on the anatomic location of
the insult, this degeneration could be transsynaptic retrograde, as suggested in
periventricular leukomalacia (PVL), or simply retrograde, as suggested in, for
example, craniopharyngioma (185). In addition, various teratologic agents,
such as alcohol, may cause anterograde ONH (186).
The vast majority of cases of ONH cannot be attributed to genetic
mutations at the current time. Familial cases have been reported and are
usually autosomal recessive. However, <1% of cases of ONH in a large series
were found to have mutations in HESX1, a gene known to cause optic nerve
anomalies, and none were found to have SOX2 mutations (187). PAX6
mutations have been reported to result in a wide variety of congenital optic
nerve abnormalities, including hypoplasia (105,188).
ONH has been associated with both postmaturity and prematurity
(189,190). A Swedish study found an increased risk with young maternal age,
primiparity (first-time pregnancy), and early prenatal smoking exposure
without drug or alcohol exposure (191).
The role of teratogens in some cases of OHN has been more convincingly
established. Maternal ingestion of quinine (192), Dilantin (193), lysergic acid
diethylamide (LSD) (194,195), and phencyclidine (196) has all been reported
to be associated with ONH in infancy. However, the evidence for teratogenic
effects is strongest for the genesis of ONH in the fetal alcohol syndrome
(186,197,198).
As demonstrated by Hoyt et al., patients with congenital hemianopsia had
obvious segmental atrophy of the optic disc but also a reduced horizontal
diameter, suggesting that the optic nerves were also hypoplastic (199). Margo
et al. reported optic atrophy with normal-sized nerves in two similar patients
(200). On the basis of these data, Hoyt and Good suggested that the same
type of lesion could result in either ONH or optic atrophy, depending on the
timing of the lesion (201). It may also be speculated that a primary prenatal
adverse event causing ONH might result in a higher vulnerability for a
perinatal secondary insult. It seems possible that the timing of the lesion
might be an explanatory factor for the varying appearance of the optic disc
noted in children with a clinical diagnosis of ONH.
Clinical Presentation
Poor visual behavior is usually the first sign of ONH. Nystagmus usually
develops at 1 to 3 months of age followed by strabismus—typically esotropia
—in the first year of life. Children with markedly asymmetric or unilateral
ONH may present primarily with strabismus rather than nystagmus. Myopia
and hyperopia have been equally found (182).
More than 80% of bilateral cases are legally blind (202). Most children
experience some improvement in their vision in the first few years of life.
This may be due to optic nerve myelination that occurs in the first 4 years of
life and leads to improved axonal conduction (183).
Observations of developmental delay in association with ONH range
from isolated focal defects to global delay. Garcia-Filion et al. (176) found
developmental delays in 71% of ONH patients using standardized
neuropsychological instruments in a prospective study. Motor delays were
the most common (75%), and communication delays were the least common
(44%). The prevalence of autism appears even higher in children with ONH
among the visually impaired children.
Taylor proposed the following criteria concerning the clinical
presentation of ONH (167):
1. Isolated ONH.
2. Absent septum pellucidum. This anomaly has not been associated with
significant intellectual, behavioral, or neurologic dysfunction (169,171).
3. Posterior pituitary hypoplasia commonly associated with endocrine
dysfunction.
4. Neuronal migrational anomalies in the cerebral hemispheres. This
includes, for example, thinning of the corpus callosum, which is
predictive of neurodevelopmental problems.
The optimal method for diagnosing ONH in a young child is direct or indirect
ophthalmoscopy, according to the child’s age. Magnification may be needed,
and documentation using optic head photos may facilitate the optic nerve
head analysis. First, and most important, is an assessment of the area of the
disc relative to the size of the central retinal vessels overlying it. ONH
classically manifests as a small optic nerve head with the distance between
the center of the disc and fovea exceeding three disc diameters and is
frequently accompanied by a double-ring sign and anomalous retinal vessel
morphology. Although most patients with disc diameter/disc to macula
(DD/DM) ratios <0.35 have generally been described as having ONH, some
with DD/DM ratios of 0.30 to 0.35 have normal vision. Fundus photography
may help in the diagnosis.
Small optic disc(s) and the double-ring sign are the hallmarks of the
severe condition. The double-ring sign indicates that a small nerve is present
within the confines of a wider scleral canal (Figure 22-21). In patients with
ONH, a ring of hypopigmentation or hyperpigmentation often, but not
always, surrounds the disc defining the area of the putative scleral canal. This
is presumably caused by migration of sensory retina and/or retinal pigment
epithelium from their original margin at the edge of the optic canal to a new
position at the border of the hypoplastic optic disc (203). This “double-ring”
sign does not define ONH, because a similar appearance may be present in
other conditions, such as myopia. In less marked cases, the diagnosis is
reached first by clinical suspicion and the subsequent measurement of the
relative and absolute size of the disc (Figure 22-22). Abnormal visual fields
and nerve fiber layer photographs are helpful for the diagnosis (179).
Reduced optic nerve diameter from the normal range of 4 to 4.5 mm
measured by ultrasound has been described, and the diameter of the optic
canal may be reduced (204).
FIGURE 22-21 Mild optic nerve hypoplasia in the right
eye of a 5-year-old girl; vision is 20/80 OD and 20/20 OS.
A small hypopigmented rim surrounds the smaller optic
nerve (A) not seen in the fellow eye (B).
FIGURE 22-22 Severe optic nerve hypoplasia in a 2-year-
old girl. Double-ring sign with hyperpigmentation around
the optic nerve. Vision is limited to fix and follow. The
fellow eye was lost by severe intraocular bleeding due to
thrombocytopenia.
OCT studies show that the ONH is associated with thinner RNFL and
ganglion cell layer; the outer nuclear layer has a more dome-shaped aspect
and the macula presents some degree of foveal hypoplasia. Horizontal disc,
horizontal rim, and horizontal cup diameters are also reduced in ONH (205).
The refractive state of the eye should always be assessed as high
hypermetropia may give a false impression of ONH. Segmental ONH, which
is mostly appreciated in the superior disc, can be identified using OCT (206).
ERG can be useful in severe bilateral cases in which the diagnosis of Leber
amaurosis and achromatopsia must be considered. ONH should have normal
ERG amplitudes.
The vision of young children with ONH should be monitored at least
annually, and any refractive errors should be treated. Patching of the better
eye can result in improvement of vision in the worse eye. However, if the
ONH is very asymmetric, maintenance of improved vision requires
prolonged patching that can be disruptive to development in a child with
many other handicaps. Thus, amblyopia therapy should be reserved for those
cases in which the potential vision in each eye is felt to be fairly good or in
which there are failed attempts previously in children with unilateral or
markedly asymmetric ONH.
Conclusion
All infants with poor visual behavior, strabismus, or nystagmus by 3 months
of age should have an ophthalmoscopic examination to rule out ONH. The
finding of reduced vision in a patient with a small optic disc thus supports the
diagnosis of ONH. In cases with reduced vision but normal to subnormal disc
size, other morphologic signs, including reduced rim area, pallor of the disc,
double ring, and abnormal vessels, may be used as additional diagnostic aids.
Once ONH is confirmed ophthalmoscopically, an MRI of the brain
should be obtained. The MRI can rule out treatable conditions such as
hydrocephalus but can also be used to anticipate developmental delay
associated with corpus callosum hypoplasia or other major malformations.
Parents, pediatricians, neurologists, and endocrinologists should be informed
of the potential risk for endocrine dysfunction in affected children to ensure
close monitoring for clinical signs of hypopituitarism (206).
SUPERIOR SEGMENTAL OPTIC

HYPOPLASIA
Superior segmental optic hypoplasia (SSOH), also termed “topless optic
disc” by Landau et al., is a developmental disorder characterized by a relative
superior entry of the central retinal artery, a superior retinal nerve fiber
deficiency, a superior scleral halo, and a superior disc pallor (207). Patients
with SSOH have good visual acuity and inferior altitudinal or sector-like
visual field defects (208). OCT studies report thinning of RNFL in the
superotemporal and nasal quadrants and retinal pigment epithelial/basement
membrane complex extension over the lamina cribrosa in the nasal portion of
the optic nerve in 100% of the cases (209). The pathogenesis of SSOH
remains unknown. Several reports point to an association with maternal type
1 diabetes mellitus (210).
OPTIC NERVE APLASIA

Introduction
Optic nerve aplasia (ONA) is a rare developmental anomaly characterized by
the congenital absence of the optic nerve, central retinal vessels, and retinal
ganglion cells (211). Despite the fact that AON was described almost 140
years ago by von Graefe (212), the pathogenesis remains speculative.
Unilateral ONA is generally associated with otherwise normal brain
development, while bilateral ONA is usually accompanied by severe and
widespread congenital CNS malformations (199,213). Newman was the first
to clearly describe ONA in humans just a few years after the introduction of
the ophthalmoscope: two blind sisters in a nonconsanguineous family of eight
children had normal-sized eyes but no perception of light; one had roving eye
movements, which made ophthalmoscopy difficult. “No appearance of optic
disc, retinal vessels or yellow spot, but an irregularly star-shaped patch of
dark nonvascular appearance occupies the place of the disc.” He went on to
describe the pupil reactions that have cast doubt on whether this was truly a
description of ONA, but the ophthalmoscopic description is so clear as to be
difficult to doubt (214).
Embryology
The choroidal vasculature is normal in ONA suggesting that eye development
is normal through the 2nd month, but absence of retinal vessels and lacunar
retinal defects in ONA suggest defective retinal development and failure of
retinal angiogenesis in the third to fourth month contribute to the
degeneration of retinal ganglion cells (215). Defective retinal angiogenesis
and retinal dysplasia in ONA could be associated with coloboma of the eyes.
One can look at ONA as either a failure of the mesoderm to enter the
fetal fissure and provide vascularization of the retina and optic nerve or as a
result of absent retinal ganglion cell axons, which inhibit retinal ganglion cell
development and provoke subsequent lack of mesodermal induction. Warfel
suggested that there is an initial development of the eye with subsequent
“degeneration” (216). Oster et al. suggest there is a failure of genetic control
of development; the authors propose that since ONH but not ONA results in
the absence of Netrin-1, a protein involved in axonal growth function in
mice, there are other mechanisms involved that act on axons during
development (217).
Different authors have given their approach concerning ONA
development. Weiter et al. suggest that ventral invagination of the optic
vesicle causes nerve fiber misdirection and secondary atrophy (218). Yanoff
et al. (219) postulate a primary failure of ganglion cells to develop and send
out axons. This failure results in a lack of induction of mesodermal ingrowth
including a lack of retinal blood vessel development. Hotchkiss and Green
(220) agree that failure of mesodermal induction secondary to a neuronal
defect in the ganglion cell layer is probably responsible for the development
of aplasia. The fact that eyes with optic aplasia may be nearly normal in size
and have a normal lens suggests the initial development of the eye proceeds
normally with primitive multipotent retinal ganglion cells with vascular
supply from both the hyaloid artery and the annular vessel (221).
PAX6 and OTX2 mutations have been associated with ONA (105).
Because Pax6/PAX6 is expressed in numerous tissues important for optic
nerve development—including the CNS, optic stalk and retinal progenitors at
an early stage, and retinal ganglion cells at a late stage; it is not surprising
that more variable phenotypes are caused by PAX6 mutations. Meier et al.
reported the only study of an autosomal dominant form of nonsyndromic
unilateral and bilateral ONA. They demonstrated that neuroimaging (MRI)
may have an important diagnostic value for uncovering ONA in
microphthalmic patients while implicating the deletion of the CYP26A1 and
CYP26C1 genes as potential susceptibility factors for ONA (221).
Pregnancies are described as normal, but environmental factors might be
a consideration since clinical histories from case reports mention exposure to
a viral-like episode during the first trimester (222), acetone exposure as in
this case, or smoking during pregnancy (223). Aplasia may be produced
experimentally further implicating the potential role of environmental factors
in ONA.
Diagnostic Criteria
According to Taylor, the diagnostic criteria are summarized as follows (167).
The optic disc is absent or rudimentary. There are no, or few, and abnormal
retinal vessels. The vision is extremely poor or absent. There is sometimes a
coloboma of the fundus or iris, and aniridia may occur (218). There may be a
retinal pigment disturbance especially at the site where the optic disc might
have been. There may be other systemic and eye developmental defects such
as coloboma, microphthalmos, cataracts, and sclerocornea (215). Published
reports have described a spectrum of eye abnormalities on the affected side.
Frequently, these eyes are microphthalmic and enophthalmic, and ptosis of
the affected eye is present. Esotropia of the affected eye is common. The
vitreous is usually clear, but a fibrovascular strand may extend from a
choroidal vessel toward the lens, representing hyperplastic primary vitreous
(218). Retinal dysplasia is frequent in histopathologic cases. Choroidal
neovascularization is an occasional finding. Microphthalmos or
anophthalmos may occur in the fellow eye (167).
The ERG wave may be flat. If present, the A- and B-waves are
diminished. An absent or depressed ERG in eyes with aplasia indicates a
more profound retinal dysfunction than merely reduced ganglion cells and
axons. A defect in the development of all retinal cells, both functionally and
anatomically, seems to occur in aplasia of the optic nerve (224). Visual
evoked potential recordings suggest abnormal crossing with greater-than-
normal contralateral fiber projections of the normal nerve (18).
Conclusion
ONA is an extremely rare condition whose origin still remains unclear. It is
mostly a unilateral condition frequently associated with other malformations
in the affected eye. The typical fundus presentation and the pupillary reflex
are sufficient to establish the diagnosis. In microphthalmic patients,
neuroimaging (MRI) may have an important diagnostic value for uncovering
ONA. In bilateral cases, neuroimaging is essential in order to reveal CNS
malformations, whereas other systemic malformations should be questioned.
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23
Albinism
Katherine Ann Lee, Sylvia R. Kodsi, Joseph Carroll, and C. Gail
Summers
INTRODUCTION
Albinism (albus, Latin, meaning white) is a group of disorders involving
melanin pigment biosynthesis with the specific gene in which mutations
occur defining the type. The phenotypic spectrum can be broad among the
different types of albinism and within a specific type. Most are inherited as
autosomal recessive disorders, but ocular albinism (OA1) is inherited as X-
linked recessive. Some types of albinism are syndromic as they have
associated systemic manifestations that can cause more morbidity than the
ocular features.
Most types of albinism are oculocutaneous, meaning that the abnormality
in melanin production affects the skin, hair, and eyes, although the extent
varies. In ocular albinism, the pigment abnormality is primarily limited to the
eyes. Syndromic albinism is associated with oculocutaneous albinism (OCA).
PREVALENCE
Albinism affects approximately 1 per 17,000 persons worldwide, but there is
substantial geographic variability in the prevalence of individual types (1–3).
To date, seven types of nonsyndromic OCA have been identified.
Approximately 1 in 70 persons carries a mutation for a type of albinism (4,5).
There are no known environmental factors that cause albinism, but
environmental factors can contribute greatly to morbidity from the condition.
For example, people with albinism lack melanin protection in their skin from
the sun’s damaging rays. For affected individuals who live in sunny parts of
the world, near the equator, or at high altitude, the risk of skin cancer
increases markedly compared to those who live in less sunny areas. Access to
sunscreen products and skin protective clothing also affects risk of skin
cancer in these individuals. In parts of Africa with intense sunlight and little
access to sunscreen, virtually 100% of patients with albinism have been
reported to develop skin cancer by their 30s or 40s. For patients with
syndromic albinism who have a bleeding diathesis, predisposition to lung and
liver fibrosis, and susceptibility to infections, living in a safe, controlled
environment with access to excellent medical care may extend life far beyond
that of patients living without these advantages.
GENETICS
OCA1 is caused by mutations of the tyrosinase gene (TYR) (6). Tyrosinase is
critical to the first step of melanin biosynthesis. Historically, OCA1 has been
divided into two categories clinically, OCA1A and OCA1B. OCA1A patients
have white hair and skin at birth and they do not gain pigment with age.
OCA1A is the most severely affected type of OCA. Persons with OCA1B are
also lightly pigmented at birth but gain pigmentation in skin and hair with
time, sometimes showing pigmented nevi and the ability to tan (7). OCA1 is
the most common type of albinism in America and China (1:40,000) (8). In
the era of genetic testing, we now understand that what was historically
called OCA1A represented a complete lack of enzyme activity needed to
produce melanin. The most common gene associated with this is TYR, but
with genetic testing, severe null mutations of other genes in pigment
pathways, for example, OCA2, OCA4, and some Hermansky-Pudlak
syndrome genes, may clinically appear to be OCA1A. Many different genetic
types of albinism may clinically fall into the OCA1B description. Over time,
the subtypes of albinism are being referred to by the gene affected, rather
than by historical names based on amount of pigment present.
OCA2, the most common type of albinism worldwide (1:39,000), is the
most common type of albinism in Africa, with a prevalence up to 1 in 3,900
in areas of Southern Africa (9). Among African Americans, the prevalence of
OCA2 is approximately 1 in 10,000 (10,11). The OCA2 product is important
in melanosome biosynthesis and transport of melanosome-associated
proteins. OCA3 is caused by mutations in tyrosinase-related protein 1
(TYRP1), an abundant melanosome protein with several functions. It has a
prevalence of 1:8,500 in Southern Africa related to a founder effect, but it is
also seen in Pakistani, German, Indian, and Japanese populations (9). OCA4
is caused by mutations in a gene that encodes a melanosome membrane
transporter protein (MATP or SLC45A2). The overall prevalence of OCA4 is
1:100,000, but it accounts for 24% of Japanese OCA (12). OCA5, OCA6, and
OCA7 are much more rare, and their phenotypic characteristics are not fully
described (see Table 23-1).
TABLE 23-1 Types of albinism

NA, Not applicable; MIM, Mendelian inheritance in man number.
Ocular albinism (OA1) affects males with a prevalence of 1:20,000 and is

caused by mutations in GPR143. The product of GPR143 is involved in the
growth and maturation of melanosomes. Obligate carrier females often
demonstrate retinal pigmentary changes and mild transillumination defects.
Cutaneous manifestations are minimal (13).
Hermansky-Pudlak syndrome (HPS) includes OCA; accumulation of
ceroid-like substance in tissues, particularly kidney and lung; and a bleeding
diathesis due to abnormal platelet aggregation (14). The 10 genes known to
associate with this syndrome encode lysosome-related proteins (see Table 23-
1). Some types have immunodeficiency, and more severe types are associated
with pulmonary fibrosis and granulomatous colitis. The syndrome is rare
except in the northwest part of Puerto Rico where it affects up to 1 in 8,000
(15).
Chédiak-Higashi syndrome (CHS) is very rare and is caused by mutations
in the LYST gene with features of OCA, neutropenia, and increased
susceptibility to infection. The LYST gene product is involved in the transport
of proteins into lysosomes. A peripheral blood smear demonstrates giant
inclusions in neutrophils (16).
WORLDWIDE IMPACT
As discussed in the previous section, due to founder effects, different types of
albinism are more prevalent than others in certain geographic regions of the
world and in the diaspora of those communities. Taken together, however,
albinism is found all around the world in every ethnicity and geographic
region. Although, for example, Hermansky-Pudlak syndrome is relatively
more common in Puerto Rico than in other parts of the world, patients from
any ethnicity can have Hermansky-Pudlak syndrome. Although
understanding the increased prevalence of specific types based on ancestry
can be helpful, it can also prevent some patients from receiving a timely
diagnosis if they do not fit the typical description.
PATHOPHYSIOLOGY
Melanocytes are melanin-producing cells found in skin, hair follicles, eyes,
inner ear, heart, and brains of humans. Melanin is produced and packaged
into lysosome-like intracellular melanosomes. In the epidermis, mature
melanosomes are distributed by phagocytosis to surrounding keratocytes and
hair follicles (17). Hypopigmentation in albinism is related to reduced
production of melanin, defects in melanosome function and distribution to
surrounding cells, and defects in proteins integral to multiple organelles
including melanosomes.
Melanin synthesis begins with the rate-limiting conversion of L-tyrosine
to L-DOPA by membrane-bound tyrosinase. In subsequent steps L-DOPA is
converted to brownish, black eumelanin or reddish, yellow pheomelanin
(Figure 23-1). Eumelanin confers ultraviolet (UV) light protection;
pheomelanin does not. OCA1 is caused by a complete or partial absence of
tyrosinase and has a profound effect on skin and hair pigmentation as well as
visual function. The functions of the gene products of the other
nonsyndromic and syndromic albinism types are less well understood but
generally involve maintenance of the structural integrity of melanosomes and
similar intracellular organelles and to the distribution of melanosomes to
surrounding keratocytes.
FIGURE 23-1 Simplified diagram of melanin
biosynthesis within the melanocyte. Tyrosine, combined
with enzymes such as tyrosinase (TYR) and tyrosine-
related protein 1 and 2 (TRP1 and TRP2) changes into
eumelanin and pheomelanin pigments. (Reprinted with
permission from Cichorek M, Wachulska M, Stasiewicz
A, et al. Skin melanocytes: biology and development. Adv
Dermatol Allergol 2013;30:30–41. doi:
10.5114/pdia.2013.33376.)
Melanin-producing retinal pigmentary epithelial (RPE) cells present early in
embryogenesis and are important in visual system development. Melanin in
neuroectoderm-derived RPE cells is important to the development of the
fovea. Animal models demonstrate that tyrosinase activity is necessary for
controlled decussation of retinostriate projections at the chiasm and altered
RPE cell melanin production alters retinal ganglion cell projections. In
albinism, absence of melanin results in foveal hypoplasia as well as excess
decussation of nerve fibers at the chiasm (Figure 23-2) disrupting the normal
layering of cells at the lateral geniculate nucleus and reducing stereopsis
(18–20).
FIGURE 23-2 Normally, ganglion cells nasal to the fovea

are routed to cross at the chiasm (left). In albinism, a
variable number of ganglion cells temporal to the fovea
also cross (right), accounting for the misrouting seen in
albinism (see also e-book Chapter 6). (From Creel D,
Witkop CJ, King RA. Asymmetric visually evoked
potentials in human albinos:evidence for visual system
anomalies. Invest Ophthalmol Vis Sci 1974;13:430–440.).

Persons with albinism display a highly variable phenotype of cutaneous
pigmentation from complete absence of pigmentation (historically called
OCA1A) to yellow, reddish, or near-normal pigmentation in other types of
OCA. In lightly pigmented ethnicities, comparison to other family members
may be necessary to appreciate the cutaneous hypopigmentation, or it may be
undetectable related to others in the family. Because eumelanin confers UV
protection, persons with albinism are at increased risk for solar degenerations
and UV-associated malignancies even if they appear to have some
pigmentation (2).
The primary morbidity of albinism is ocular, and persons with albinism
generally have reduced vision (21). Visual acuity can range from 20/20 to
worse than 20/400, and there is a general positive correlation between
pigmentation and visual acuity (22,23). Color vision is normal. Persons with
no pigment are most visually impaired with best corrected visual acuity
ranging between 20/80 and 20/400, although vision is occasionally better.
Most persons with albinism are photosensitive to a degree that inversely
correlates with their amount of pigmentation. High astigmatic refractive
errors are common, particularly with-the-rule astigmatism (24).
OCULAR FEATURES
Typical ocular features of albinism include nystagmus, reduced iris and RPE
pigmentation, abnormal retinostriate projections, and foveal hypoplasia (25a).
Nystagmus is usually present in albinism and starts in early infancy. It is
most often horizontal and conjugate and damps in the first few years of life.
The null zone is where the amplitude of the nystagmus is diminished and
vision is improved. Pendular, periodic alternating, and rotary nystagmus have
been described in albinism.
Melanin pigment is reduced in the posterior iris epithelium, derived from
neuroectoderm, whereas anterior iris stromal melanocytes are derived from
the neural crest. This leads to mild to marked pigment defects best
appreciated by transillumination of the irises with slit lamp biomicroscopy
and slit lamp photography (Figure 23-3). A decrease in pigmentation of the
retinal epithelial cells gives a yellow-orange hue to the retina and allows a
variable increased view of the choroidal vessels in the macula and retinal
periphery (Figure 23-4). A characteristic feature of albinism is the excessive
decussation of retinostriate nerve projections at the optic chiasm where both
nasal and some temporal retinal ganglion cells project to the contralateral
lateral geniculate nucleus (Figure 23-2) (25b). This excessive crossing can be
demonstrated by flash or pattern visual evoked potential with occipital
recording. The misrouting frequently disrupts stereoacuity. The small number
of patients who have stereoacuity typically has relatively better vision and
less iris transillumination and may show granular melanin pigment in their
maculas compared to those without stereoacuity (26).
FIGURE 23-3 Grading of iris transillumination in

albinism from full (grade 4) at top left to minimal (grade
1) at lower right using slit lamp biomicroscopy.
FIGURE 23-4 Grading of macular transparency in

albinism from full (grade 1) at top left, in which choroidal
vessels are easily visualized in the macula, to opaque
(grade 3) at bottom left in which choroidal vessles cannot
be seen in the macula. Note that all three pictures show
foveal hypoplasia whereas the lower right image shows a
normal eye with pigmented retina with a formed fovea.
(Reprinted from Aswad MI, Tauber J, Baum J.
Plasmapheresis treatment in patients with severe atopic
keratoconjunctivitis. Ophthalmology 1988;95(4):444–447.
Copyright © 1988 American Academy of Ophthalmology,
Inc. With permission.)
Foveal Hypoplasia
Beyond altered pigmentation, perhaps the most striking ocular feature in all
forms of albinism is the absence of a normal foveal pit in the retina (i.e.,
foveal hypoplasia). The human fovea is normally characterized by a lateral
displacement of the inner retinal layers, leaving behind a depression or pit in
the central retina (fovea is Latin for “small pit”) (27). Early histologic studies
of a small number of human eyes reported an absence of foveal
differentiation with persistent inner retinal layers at the location of the
incipient fovea in albinism (28–30), and it was generally thought that this
abnormal foveal morphology gave rise to the reduced visual acuity in
individuals with albinism. However, the ex vivo nature of these studies
obviated confirming possible links between foveal structure and visual
function in albinism. In contrast, optical coherence tomography (OCT) allows
for noninvasive volumetric imaging of the living human retina (31). The
availability of OCT has enabled examination of foveal morphology in a
larger number of subjects. The first OCT studies in patients with albinism
corroborated the earlier histology studies, describing the presence of multiple
inner retinal layers throughout the foveal region (32–34). Out of these studies
came the first direct evidence that there is not a single phenotype of “foveal
hypoplasia” in albinism; rather, there is a spectrum of foveal morphology in
patients with albinism—ranging from a complete absence of inner retinal
excavation to a small central depression with a corresponding annular reflex
with indirect ophthalmoscopy (32–35).
As the use of OCT grew, the analysis of OCT images also advanced.
Various groups have developed ways to quantify the size and shape of the
foveal pit in normal individuals. Different studies vary slightly in the specific
parameters used to describe the fovea (e.g., pit depth, pit diameter, pit
volume, central retinal thickness, pit slope, foveal inner retinal area), but they
all describe pronounced variability in foveal morphology across individuals
with normal vision (Figure 23-5) (35–43). Recent studies have applied
similar analyses to individuals with albinism, seeking to quantify the
individual variation in foveal pit morphology that was qualitatively reported
in earlier studies (44). In contrast to the classic clinical picture of albinism,
some subjects had foveal pit morphology that was within the normal range
(44). This emerged not only as a result of more accurate quantification of pit
morphology but also due to having a more thorough description of what
“normal” morphology encompasses.
FIGURE 23-5 Variability in foveal morphology. Shown

on the left are optical coherence tomography (OCT) B-
scans of the fovea in individuals with normal vision (A, B)
and a patient with albinism (C). On the right are
corresponding OCT angiography images of the foveal
avascular zone (FAZ) demonstrating that the size of the
FAZ correlates with the size of the foveal pit (D, E) and
that the FAZ is generally absent in albinism (F).
There are several other important anatomical specializations associated with

the fovea other than a pit (Figure 23-5). First, the normal fovea has an
avascular zone (Figure 23-5 D and E), lacks inner retinal vasculature, owing
to the absence of the inner retinal layers at the foveal center. Individuals with
albinism have been shown to lack this foveal avascular zone (FAZ) (45–47).
This correlation generally supports the developmental model in which an
FAZ is a prerequisite for the formation of a foveal pit (27). In addition, the
cone photoreceptors at the fovea have longer outer segments and have a
higher packing density than their peripheral counterparts. This manifests on
OCT images as a thickening of the hyporeflective outer nuclear layer and a
relative lengthening of the outer segment band (39,48). As with the pit itself,
individuals with albinism can overlap with individuals with normal vision—
both in outer nuclear layer thickness and in outer segment length (44,49). The
widespread variability and overlap with the normal range of anatomy
resurrects the important question of whether differences in foveal structure
lead to differences in visual acuity in albinism. In particular, it was unclear
which specific foveal specializations may be more critically linked to visual
function. To address this, Thomas et al. introduced a grading system to
characterize the degree of foveal hypoplasia (50). This system separates
patients with albinism into one of four grades of hypoplasia, based on the
presence or absence of a foveal pit, inner retinal layer extrusion, outer
segment lengthening, and outer nuclear layer widening (Figure 23-6). Using
this grading scheme, they found that individuals with more “immature”
foveas had worse visual acuity than did individuals with more well-
developed foveas, although there was substantial variability in visual acuity
within each grade. Lee et al. recently proposed refining these grades to
incorporate quantitative measurements (e.g., outer nuclear thickness and
outer segment length), given the extensive variability of these features (49).
Such measures are not readily available on clinical OCT devices, so
qualitative assessment of the various foveal features remains an important
prognostic indicator of visual function.
FIGURE 23-6 Thomas et al. grading scheme of foveal
hypoplasia (50). Shown in (A) is a normal retina
highlighting the four key features used to grade
hypoplasia. Section (B) shows the four grades used to
describe the fovea in patients with albinism, based on the
presence or absence of the four key features associated
with normal foveal morphology. The “atypical” grade is
not observed in albinism but can be seen in patients with
achromatopsia. (Reprinted from Thomas MG, Kumar A,
Mohammad A, et al. Structural grading of foveal
hypoplasia using spectral-domain optical coherence
tomography: a predictor of visual acuity? Ophthalmology
2011;118(8):1653–1660. Copyright © 2011 American
Academy of Ophthalmology. With permission.)
NONSYNDROMIC ALBINISM
The most common types of albinism are OCA1 and OCA2. Therefore, the
phenotypes are more completely described than the less prevalent types of
albinism, OCA3 and OCA4, or the more recently described types of albinism,
OCA5, OCA6, and OCA7.
Oculocutaneous Albinism, Type 1

OCA1, caused by mutations in TYR, has been termed OCA1A when no
pigment is present. This was previously called tyrosinase-negative albinism
and is considered the most severe type as there is lifelong absence of melanin
pigment in the skin, hair, and eyes. Early in life, the irises may appear pink
due to the absence of melanin pigment in the posterior iris epithelium and the
reduced thickness of the iris stroma. The hair and lashes are white. In older
persons, the hair may appear to have some pigment due to shampoo or
minerals in water. Occasionally, a normally pigmented hair will be identified,
an example of revertant mosaicism (51).
Compared to other types of albinism, best corrected visual acuity in
OCA1A is usually more reduced (52). Glasses are often needed due to
astigmatism that increases in the first decade of life (24). Winsor et al. found
a mean best corrected visual acuity of 20/128 (range 20/60–250) at age at
least 15 years (52), and Schweigert et al. found a mean of 20/104 (range
20/80–200) at age 10 (24). Others have recorded vision in the range of legal
blindness (≤20/200) (53–56). Those few persons with OCA1A and somewhat
better vision are postulated to have initiated glasses at an earlier age, have
different retinal or striate morphology than those with poorer vision, or to
have mutations that encode for better visual acuity (52,57). Since many of
these studies were performed prior to the availability of genetic testing, some
of the severely affected “OCA1A” patients in the series may have had severe
mutations in other genes, accounting for some of the variability.
OCA1B, referred to as yellow OCA in the past, is caused by mutations
that result in some enzyme formation and, therefore, permit a small amount
of melanin pigment. In vitro studies suggest that the enzyme tyrosinase is
variably more stable in OCA1B compared to OCA1A in which it is unstable
and inactive, accounting for the differing phenotypes (58). Individuals with
OCA1B may also be born with white hair that darkens to blond or brown.
Melanin pigment can be identified in the posterior iris epithelium with slit
lamp biomicroscopy at birth. When granular-appearing melanin pigment is
identified in the macula, a relatively better visual prognosis can be offered
(26). Skin may lightly tan with exposure to sun, and eyelashes often darken to
brown (7).
Binocular visual acuity in OCA1B has been reported to be as good as
20/20 (59). Mean best corrected visual acuity for OCA1B was 20/34 (range
20/20 to 16/200) in a series of individuals at least 15 years old (52) and was
20/59 (range 20/25 to 20/125) in a group of 10-year-old patients with
OCA1B (24). Gargiulo et al. have reported monocular visual acuity between
20/25 and 20/600 in OCA1, most of whom appeared to have OCA1B (55).
A subset of OCA1B, minimal pigment OCA, has a heterogeneous
phenotype associated with a small amount of residual enzyme activity, and
visual acuity has been reported to range between 20/50 and 20/200 (60). The
temperature-sensitive variant of OCA1B, in which hair on the cooler areas
(arms, legs) of the body is more pigmented than that on the warmer parts of
the body (scalp, axilla, pubis), has been reported to have visual acuity of
20/200 (61). As with OCA1A, most of the studies of patients with OCA1B
were done prior to the advent of genetic testing, which identified mutations in
other genes including OCA2, OCA3, and OCA4. Inevitably, some patients
reported that OCA1B had mutations in genes other than OCA1, and this adds
to the variability in the reports.
Individuals with OCA2, previously known as brown albinism, are often born
with pigmented hair that is lighter than unaffected family members. Most
often, hair color is blond, but there may be a red tint due to mutations in the
MC1R (melanocortin-1 receptor) gene, which modifies the phenotype by
allowing the production of a pheomelanin (yellow/red hair) (62). This type of
albinism is very common in Africa where skin and hair are light brown.
Mutations in the OCA2 gene, previously referred to as the P gene, interfere
with the normal trafficking of tyrosinase into the melanosome. Very severe
mutations in patients with OCA2 may have little or no apparent pigment at
birth, though some may develop over time. There is a founder mutation in
OCA2 in some parts of Africa (63).
Mean best corrected visual acuity was 20/66 (range 20/30 to 125) in a
group of 10-year-old patients with OCA2 (24) and was 20/59 (range 20/20 to
80) in a group at least 15 years old (52).

OCA3, previously known as rufous albinism, is recognized in the South
African population as being the second most common (18%), following
OCA2 (64). Skin is reddish-brown in color, hair is ginger-red, and irises are
blue or brown. Mutations in TYRP1 (tyrosinase-related protein 1), an enzyme
in melanin biosynthesis, cause OCA3 and result in delayed maturation or
early degradation of tyrosinase (65). The visual system can be minimally
affected. Approximately one-fourth has no nystagmus.
OCA3 has rarely been reported in populations outside of South Africa.
An Asian Indian boy with OCA3 was born with light reddish blond hair that
became medium brown with a reddish tint by age 3; he developed a slight
tan. His irises were brown and he had nystagmus (66). A German boy of
Caucasian origin was born to distantly consanguineous parents. His hair was
blond with a reddish tint, he did not tan, and irises were blue-green. With
correction for a large amount of hyperopia, visual acuity was 0.4; he had
nystagmus (67). A Japanese girl with OCA3 had no nystagmus or
photophobia; her hair was blond, her skin was lighter than others who were
unaffected in her family, and her irises and eyelashes were dark brown (68).
The phenotype of OCA4, caused by mutations in the MATP (membrane-
associated transporter protein, also known as SLC45A2) gene, was first
described in a Turkish individual whose parents were possibly distantly
related (69). His level of generalized hypopigmentation was similar to OCA2,
but he did not have mutations in the OCA2 gene. He had very light skin that
did not appear to tan, very light blond hair, and blue irises, in addition to the
ocular features of albinism. A Korean girl with OCA4 had a similar
phenotype (70). A Moroccan boy with OCA4 had a phenotype similar to
OCA1A with complete absence of melanin pigment (71).
As OCA4 was recognized in other ethnic backgrounds, the phenotype
expanded to include brown-black hair, brown irises, and absence of
nystagmus, indicating the marked heterogeneity of the phenotype and the
need for genetic testing to identify this type of albinism (12,72). Inagaki et al.
found that 24% of 75 unrelated Japanese patients with variation in phenotype
were genetically identified as having OCA4 (12). This was later confirmed at
27% (73). The gene functions as a membrane transporter within the
melanosome.
Among five German individuals of Caucasian background with OCA4,
four had severe hypopigmentation and visual acuity ranged from 20/200 to
20/400. The fifth person with OCA4 was born with blond hair that darkened,
and he developed a slight tan; he had less severe ocular hypopigmentation,
his nystagmus was less, and his visual acuity measured 20/30 in one eye and
20/40 in the other despite having foveal hypoplasia (72). Of the eight
Japanese individuals reported, one who had brown hair and a slight tan had
visual acuity of 20/30 in the right eye and 20/20 in the left eye and no
nystagmus (74). A consanguineous Pakistani family with OCA4 had
hypopigmented hair and skin although they had some pigmentation; all were
photophobic, and vision was variably reduced (75).

OCA5 was first described by Kausar et al. in a consanguineous Pakistani
family with six affected individuals (76). Skin was white, and hair was
golden blond, whereas unaffected members had brown hair. Vision in
affected persons was reduced and they had photophobia, nystagmus, and
foveal hypoplasia. There was no evidence for syndromic albinism.
Linkage analysis mapped OCA5 to chromosome 4q24. The gene has not
yet been identified (77).

Wei et al. reported a Chinese girl who was diagnosed with OCA at age 3
(78). She was born with blond hair that later darkened to brown. She had
mild photophobia and nystagmus. Vision measured 20/100. Irises were
translucent and had a brown tint. The fundi were hypopigmented and an OCT
confirmed foveal hypoplasia. She had no evidence of syndromic albinism.
Testing showed low eumelanin levels in the proband’s hair, and
melanosomes in skin melanocytes were less mature compared to unaffected
family members.
Exome sequencing detected two deleterious mutations in the SLC24A5
gene. The gene has been identified in persons with diverse ethnic
backgrounds. It is hypothesized that the gene protein is important in
melanosome maturation and/or production of melanin in melanosomes.
Recently, an Italian male was reported with two SLC24A5 mutations.
Vision was in the range of 1/10 to 3/10, and he had medium iris
transillumination and foveal hypoplasia (79).

Study of an inbred family and three additional individuals from the Faroe
Islands plus a person from Lithuania identified mutations in the C10orf11
gene as a cause of the OCA of affected individuals (80). The function of
C10orf11 is not well understood but is likely related to melanocyte
development and differentiation.
Skin was lighter than unaffected relatives and hair color varied from
white or light blond to dark brown. The affected individuals were not
particularly photosensitive. Best corrected visual acuity measured between
20/30 and 20/400. Most affected individuals were moderately to severely
hyperopic. Iris transillumination was typically extensive. The fundi were
hypopigmented, and all had foveal hypoplasia. Eight who had visual evoked
potentials showed the crossed asymmetry typical of albinism. The individual
with Lithuanian heritage had dark blond hair and extensive iris
transillumination; vision was 20/100.
Ocular Albinism
OA1, also designated Nettleship-Falls ocular albinism (81,82), is the only
type of albinism that is not inherited as an autosomal recessive disorder.
Males affected with this X-linked disorder typically show only the ocular
features of albinism, but their skin and hair appear to have a normal amount
of melanin, although it may be somewhat lighter than what is typical for their
families. Despite the name, skin biopsies of affected males and carrier
females may show macromelanosomes, indicating OA1 is not solely ocular
(83). A family history of males being affected over several generations is
often obtained. Female carriers are rarely symptomatic (84), but up to 90%
will show the variegated pigmentation of the fundi, which represents random
inactivation of the X chromosome, also known as lyonization or pigmentary
mosaicism (Figure 23-7) (82,85). Many carriers also have iris
transillumination, particularly in families with constitutive light
pigmentation.
FIGURE 23-7 The “mud-splattered” fundus of the
obligate carrier of X-linked OA1 is an example of random
inactivation of the X chromosome, referred to as
lyonization or pigmentary mosaicism. The darker areas
represent expression of the normal X chromosome,
whereas the areas without melanin pigment represent the
expression of the X chromosome carrying the mutation in
GPR143 that causes OA1.
Vision is variably reduced in OA1, with mean best corrected visual acuity of
20/63 (range 20/50 to 80) reported by Winsor et al. (52). Others have noted
vision as poor as 20/200 to 20/400 (28,86–90), but occasionally vision is as
good as 20/40 or better (86,88,89).
When the obligate carrier is affected, the pedigree may suggest autosomal
dominant nystagmus or a pigmented type of AD albinism; however, there
will never be male to male transmission. Examination of the mother’s retina
showing the characteristic pigmentary mosaicism establishes the diagnosis. If
the affected male has a new mutation in GPR143, the family history will be
negative, examination of the mother will be negative, and testing with a
genetic panel will be necessary to establish this type of albinism. Counseling
should include that no male offspring will be affected but all female offspring
will be carriers for OA1. Female carriers have a 50% (1 in 2) chance of
passing the mutation to male offspring who will be affected and to female
offspring who will be carriers.
Some have described an autosomal recessive ocular albinism, but most of
these have been shown to have a mildly pigmenting type of OCA (91–93).
The range of pigmentation present in all genetic types of albinism varies
widely based on the residual protein activity of the defective gene product.
Genetic testing rather than assessment of external pigmentation should be
used to diagnose the subtypes of albinism.
SYNDROMIC ALBINISM
Hermansky-Pudlak Syndrome
HPS, first described in 1959 in two unrelated Czechoslovakians (94), refers
to a heterogeneous group of disorders characterized by OCA, a platelet
storage pool deficiency causing a bleeding diathesis, and in some types,
associated with granulomatous colitis, interstitial lung disease, and/or
immunodeficiency. Electron microscopy of platelets shows absence of dense
bodies (Figure 23-8).
FIGURE 23-8 Electron micrograph of normal control
(Ctrl) platelets left, with dense bodies (arrowheads),
compared to platelets without dense bodies from a patient
with HPS (right). (Reprinted from Carmona-Rivera C,
Golas G, Hess RA, et al. Clinical, molecular, and cellular
features of non-Puerto Rican Hermansky-Pudlak
syndrome patients of Hispanic descent. J Invest Dermatol
2011;131:2394–2400. Copyright © 2011 The Society for
Investigative Dermatology, Inc. With permission.)
The bleeding disorder may be mild or severe due to impaired platelet

secondary aggregation. When mild (e.g., bruising, rare epistaxis), HPS can be
overlooked as the cause of albinism. More severe bleeding can occur
following labor and delivery, trauma, or surgery and, in some cases, can be
life threatening. Pulmonary fibrosis typically develops by the fourth decade
and can be fatal by the fifth decade unless lung transplant is performed.
Granulomatous colitis can present with severe rectal bleeding (95–97).
Because of the phenotypic variability, particularly regarding the
nonocular features of this autosomal recessive syndromic type of albinism,
genetic testing can be performed to determine the type of HPS (see Table 23-
1), to ascertain prognosis, the need for additional testing, and treatment (98).
The genes involved in HPS affect the biogenesis and trafficking of
lysosomal-related organelles, for example, melanosomes in melanocytes and
delta granules (dense bodies) in platelets. The protein products of these genes
belong to the following complexes: BLOC-1 (biogenesis of lysosome-related
organelles complex-1), BLOC-2, BLOC-3, or AP3 (adaptor protein complex-
3). The types of HPS with the same protein complex have similar severity of
comorbidities. Each type of human HPS has a murine model that has
facilitated the understanding of the gene function.
There are additional HPS mouse models without human homologs,
suggesting that there may be more genes that cause human HPS yet to be
identified.
In general, visual acuity has been reported to be reduced to a variable
level, usually between 20/50 and 5/200 in HPS (99–103). Ocular findings are
similar to other types of albinism.
HPS-1
In the northwest part of the island of Puerto Rico, 1 in 1,800 people have a
homozygous 16-base pair (bp) duplication in exon 15 of the HPS-1 gene due
to a founder effect. Other Hispanic individuals who have HPS-1 but who do
not have ancestors from Puerto Rico do not have this duplication (104). HPS-
1 is associated with an increased risk of pulmonary fibrosis. Brantly et al.
found a mean age of onset of pulmonary symptoms at age 35 years (96).
A study of 21 individuals with the 16-bp deletion showed mean visual
acuity of approximately 20/125 and moderate iris transillumination and
macular translucency (103). Another study of 16 persons with HPS-1 showed
mean visual acuity of 20/160-2 (range 20/40 to 500) and significantly more
iris transillumination when compared to a group with HPS-3 (105).
Yousaf et al. described 11 individuals with HPS-1 in 3 Pakistani families
(106). Affected persons had golden, golden-brown, or brown hair, white skin,
and hazel-green to green irises, lighter than unaffected family members. All
were photosensitive but three siblings, ages 10 to 25 years, had no
nystagmus. Of the three who had visual acuity measured, all siblings had
nystagmus, two had acuity of 6/18 in each eye, and the third, who had a large
amount of hyperopic astigmatism, had count fingers vision. Eight had
gastrointestinal problems and the remaining three were 15 years old or
younger.
One study reported the eye findings in 45 Puerto Rican patients with the
16-bp duplication. Median best corrected visual acuity was 20/400 (range
20/40 to 1,300). Most had complete iris transillumination although some
variability was reported. Most had a somewhat translucent macula, allowing
choroidal vessels to be partially visualized, but again, variability was
reported. In addition, most patients were noted to have periodic alternating
nystagmus (107).
Ten Japanese persons with HPS-1 were noted to have blond to brown hair
and blue to brown irises. All but one had nystagmus. One patient, who was
60 years old, had pulmonary fibrosis (108).
HPS-2
HPS-2 is severe and rare. It was first reported in two brothers in a family of
Dutch origin, ages 20 and 25 years, who had a history of congenital
acetabular dysplasia (109). They had bruising and epistaxis in childhood.
Recurrent otitis media and upper respiratory infections, greater than in their
unaffected siblings, were noted. Examination showed dysmetria and poor
tandem gait; the older brother also had a tremor. Magnetic resonance imaging
of the head was normal. Both had mild pulmonary fibrosis and neutropenia.
The brothers were found to have mutations in the β3A subunit, which caused
either degradation or destabilization of all four of the adaptor complex-3
subunits and interference with the trafficking of lysosomal membrane
proteins (110). They had white hair early in life but it eventually turned
blond. Both had nystagmus and almost complete iris transillumination.
Corrected visual acuity measured 20/200 in the right eye and 20/160 in the
left eye for each brother. Radial lens opacities were noted.
Huizing et al. reported a third patient with HPS-2 who was a 5-year-old
Cajun/native American boy (111). He was born with white hair that darkened
to silvery blond to tan at age 5. He had melena associated with a rotavirus
infection but did not bleed excessively with circumcision. He had multiple
episodes of respiratory distress, and at age 20 months, a biopsy showed
nonspecific interstitial pneumonitis. Severe neutropenia was treated with G-
CSF (Neupogen). The patient had short stature and was very delayed in
acquiring developmental skills. He had tricuspid regurgitation and pulmonary
hypertension. Dysmorphic facial features included epicanthus; low-set,
posteriorly rotated ears; broad nasal root; thin upper lip with long philtrum;
and retrognathia. Other findings included a severe pectus deformity and genu
valgus. He had horizontal pendular nystagmus and blue irises with full iris
transillumination. A visual evoked potential was consistent with the
retinostriate misrouting seen in albinism.
More recently, six children with HPS-2 were reported to highlight their
pulmonary disease and how pulmonary disease in HPS-2 can occur at an
earlier age than in HPS-1. Symptoms began an average of 3.3 years prior to
diagnosis. Diagnosis of pulmonary fibrosis was made at a mean age of 8.8
years (range 2 to 15 years). The pulmonary fibrosis was rapidly progressive
and was complicated by recurrent infections exacerbated by neutropenia,
pneumothoraces, and scoliosis.
A pair of siblings with the same mutations was noted to have dissimilar
clinical courses (112).
HPS-3
HPS-3 has been identified in central Puerto Rico where approximately 1 in
16,000 is affected. It is due to a 3,904-bp deletion due to a founder effect
(113). HPS-3 has also been found in those with Ashkenazi Jewish heritage
and in non–Puerto Ricans without the 3,904-bp deletion (114). A common
splice-site mutation removing exon 5 from the gene found among Ashkenazi
Jewish patients is felt to be due to a founder effect. A 4-year-old boy from the
Arabian Peninsula with easy bruising was also identified with HPS-3 (115).
Hypopigmentation is minimal (hair color tan to light brown) and bleeding is
generally mild in HPS-3, manifested as epistaxis, bruising, and menorrhagia.
Pulmonary fibrosis and gastrointestinal issues do not seem to be associated
with this type of HPS. This type of HPS can be easily overlooked and
diagnosed as a type of OCA or OA1.
In a study of eight non–Puerto Ricans with HPS-3, ages 3 to 52 years,
visual acuity ranged from 20/60 to 20/160 (114). A study of 14 individuals
with HPS-3 found mean visual acuity of 20/125 + 2 (range 20/50 to 320).
Visual acuity was significantly better and iris transillumination was less than
that in those with HPS-1 (105).
A study of 19 patients with HPS-3 found median best corrected visual
acuity to be 20/200 (range 20/25 to 400), significantly better when compared
to 45 patients with HPS-1. This study also found that those with HPS-3 had
significantly less iris transillumination and less macular translucency than
those with HPS-1 (107).
HPS-4
Anderson et al. identified seven persons of varied ethnicities with HPS-4,
ages 3 to 61 years. All gave a history of excessive bleeding and one had
granulomatous colitis. Three had restrictive lung disease, with one dying at
age 61. The true prevalence of pulmonary fibrosis in this group with HPS-4 is
unknown as four were under age 20 and were asymptomatic. Hair and skin
pigmentation varied markedly with unaffected family members. Visual acuity
ranged from 10/30 to 20/200 (116).
Two Pakistani siblings in a family of Sindh origin were also reported to
have HPS-4 (106). They had brown hair, pink white skin, and brown irises.
They reported photosensitivity and had nystagmus. They were 34 and 38
years old and appeared to have a milder phenotype, reporting no bleeding
diathesis.
HPS-5
HPS-5 is relatively rare compared with other types of HPS. Individuals with
HPS-5 are mildly hypopigmented and bleeding problems are mild. They do
not seem to have the other systemic manifestations of HPS. Because of this, a
diagnosis of HPS can be missed without molecular analysis.
Huizing et al. reported on four patients with HPS-5 (117). A 10-year-old
boy with a European heritage had bruising and light brown hair. He had
nystagmus and almost full transillumination. Vision was 20/200. A 51-year-
old Swiss woman had bruising, mild epistaxis, and prolonged menstrual
periods, but no postpartum bleeding. She had nystagmus, moderate iris
transillumination, and vision of 20/100 in one eye and 20/125 in the other.
Her hair was blond at birth but darkened to brown. She did not have
pulmonary disease or granulomatous colitis. Her 43-year-old sister had
bruising in addition to some bloody stools, but her gastrointestinal evaluation
was normal; albinism was diagnosed at age 12, and she had almost full iris
transillumination. Vision measured 20/100 in one eye and 20/125 in the
other. A 21-year-old with bruising, epistaxis, and prolonged menses had light
brown hair and almost complete iris transillumination. Vision was 20/160 in
one eye and 20/200 in the other. She did not have evidence of interstitial lung
disease or colitis.
A Venezuelan Cuban boy with HPS-5 had no bleeding with surgery but
had leg bruising and minor epistaxis; his vision was 20/100 in one eye and
20/125 in the other (104).
A series of 11 patients with HPS-5, most of French or Turkish origin,
reported that hair color was typically light brown, irises were blue to brown,
and skin did not tan. All had nystagmus and foveal hypoplasia. All but two
who were just 1 year old had reduced visual acuity (118).
HPS-6
HPS-6 is associated with mild hypopigmentation and mild bleeding
problems. A 39-year-old Belgian woman with epistaxis and bleeding
following dental extractions was diagnosed with HPS-6. She had no
symptoms to suggest pulmonary or gastrointestinal disease. She had OCA.
Her brother had similar findings and was also found to have HPS-6. There
was no history of consanguinity, but both parental families were from a
similar locale and siblings had a homozygous deletion in the HPS-6 gene.
This is an ortholog of the ruby eye mouse (ru) gene. Mutations in ru lead to a
mouse model of HPS. Electron microscopy showed only a rare dense body in
platelets (119).
Yousaf et al. reported two Pakistani siblings with HPS-6 (106). Both had
red hair, pink white skin, and dark brown irides. Vision measured 6/18 in
each eye of the 14-year-old and was 6/60 in the right eye and 6/36 in the left
eye of the 10-year-old. Both were photosensitive but only the younger one
had nystagmus. The younger sibling had a normal bleeding time and a normal
clotting time. Neither had gastrointestinal problems.
Huizing et al. studied four patients with HPS-6 (120). The first was a 36-
year-old woman of Irish and German origin, who was diagnosed with HPS at
age 26 due to bleeding problems and albinism. She had several surgeries
requiring transfusions and had a history of bruising, menorrhagia, and
frequent infections. Evaluation showed no evidence of interstitial lung
disease. Her hair was light brown. Vision was 20/100 in one eye and 20/125
in the other. She had nystagmus, marked iris transillumination, and foveal
hypoplasia. The second patient was a 22-year-old woman of Scottish,
English, and German origin. She had prolonged bleeding and epistaxis early
in life but a diagnosis of HPS was not made until age 16. Chest CT showed
no evidence of pulmonary fibrosis. Her hair was brown. Vision measured
20/80 in the right eye and 20/63 in the left. She had nystagmus, marked iris
transillumination, and foveal hypoplasia. The next patient was a 13-year-old
girl of German and Dutch heritage. She had nearly white eyelashes, brows,
and hair early in life but developed pigment later. A diagnosis of tyrosinase-
positive albinism was made and a diagnosis of HPS wasn’t made until age
16. She had strabismus surgery without bleeding problems. She had
developmental delay and bruised easily. Visual acuity was 20/200 in each
eye. The last patient was a 52-year-old Italian man with rotary nystagmus and
a history of bruising in childhood. HPS was diagnosed at age 44 due to OCA
and gastrointestinal symptoms, which included gastroesophageal reflux,
esophageal dysmotility, hiatal hernia, and dysphagia. High-resolution chest
CT showed no evidence of interstitial lung disease. He had brown hair.
Vision measured 20/160 in one eye and 20/125 in the other. He had moderate
transillumination of his brown irises, and foveal hypoplasia.
Despite these reports of relatively mild disease, a 58-year-old Caucasian
woman, born to consanguineous parents, reported no history of albinism but
did have a history of severe bleeding that included severe menorrhagia
requiring transfusions and bleeding after vaginal delivery and gastric polyp
removal that required treatment for hemodynamic instability. She was found
to have HPS-6 without colitis or pulmonary fibrosis. An eye examination
disclosed vision of 20/25 in the right eye and 20/40 in the left. She had no
nystagmus and very mild iris transillumination (121).
HPS-7
A 48-year-old Portuguese woman, born to consanguineous parents, was
identified with HPS-7 (122). She had bruising and a tendency to bleed, but
pulmonary function and CT of her lungs were normal.
A 6-year-old boy from Paraguay with HPS-7 and delay in his motor and
language development was reported by Bryan et al. (123). The boy had easy
bruising and posttraumatic hemorrhage resulting in pulmonary scarring and
right hemiparesis. He had no evidence of interstitial pulmonary fibrosis,
colitis, or immunodeficiency. His hair was light brown and his skin was
hypopigmented, much lighter than his parents. He had nystagmus, almost full
iris transillumination of his brown irises, and best corrected visual acuity was
reduced to 20/200 in each eye.
HPS-8
A homozygous germline frameshift BLOC1S3 mutation was identified as the
cause of HPS-8 in a 21-year-old Pakistani male, born to consanguineous
parents. His hair was silvery at birth and darkened to gold. His irises were
hazel and his skin was pale and did not tan. Visual acuity was 6/60 in each
eye. He had iris transillumination and foveal hypoplasia, and a visual evoked
potential showed excessive retinostriate decussation. His sister was similarly
affected but she had high myopia and her vision was 6/36. Other family
members were similarly affected with vision ranging from 6/36 to 6/120.
Easy bruising, prolonged bleeding from wounds, epistaxis, and menorrhagia
were common. Pulmonary fibrosis and colitis were not (124).
An Iranian boy with HPS-8, born to consanguineous parents, had
nystagmus noted early in life. He began wearing glasses at 3 months of age.
His skin was lighter than his parents. At age 6 years, his vision measured
20/200 in the right eye and 20/125 in the left eye. He had iris
transillumination, foveal hypoplasia, and an exotropia. He had easy bruising
and gingival bleeding but did not have excessive bleeding with circumcision
(125).
HPS-9
A 17-year-old woman from Northern Italy with absent PLDN protein
expression, indicative of HPS-9, was reported by Badolato et al. (126). She
had nystagmus related to OCA and recurring cutaneous infections.
Leukopenia and immunodeficiency were noted. She had a fever associated
with seizures at age 6. Exam showed leukopenia and thrombocytopenia. She
was found to have homozygous nonsense mutation in the pallidin (PLDN)
gene.
A 4-year-old Pakistani boy has also been reported to have PLDN
mutations (106). He had OCA with photophobia and nystagmus. His hair was
golden white, his skin was pink white, and his irises were light brown. Both
his bleeding time and clotting time were prolonged.
Okamura and colleagues reported a 52-year-old Japanese woman with
HPS-9 and consanguineous parentage. Her hair was blond and her irises were
brown. She had nystagmus and mild visual impairment. There was no history
of interstitial pulmonary disease or colitis (127).
HPS-10
Only one person with HPS-10 has been described to date. This Turkish boy,
born to consanguineous parents, had microcephaly and developed myoclonic
and generalized seizures that became intractable. He had severe
neurodevelopmental delay. Imaging showed a large arachnoid cyst in the
posterior fossa, a small telencephalon, enlarged internal and external
cerebrospinal fluid spaces, and reduced myelination. Otoacoustic potentials
and brainstem evoked audiometry were reduced (128).
The boy had chronic neutropenia and an immunodeficiency with frequent
infections. His immunoglobulin levels were normal but he had impaired NK-
and T-cell degranulation. Bone marrow aspirate showed hypersegmented
neutrophils. The patient had chronic interstitial lung disease and at age 3.5
years, developed pneumonia with sepsis and died (128).
Hair was lightly pigmented and he had intermittent nystagmus and no
ocular fixation. Other features included large, low-set ears, hypotelorism, a
flat philtrum, retrognathia with Pierre Robin sequence, flat acetabula, and
hepatosplenomegaly. The boy did not have a bleeding diathesis, but Ammann
et al. (128) indicated that the patient may have had a subclinical platelet
granule defect. The homozygous truncating mutation in the AP3D1 gene was
identified by exome sequencing, with genetic and phenotypic similarities to
the “mocha” mouse (129).
Chédiak-Higashi Syndrome
CHS is another type of syndromic OCA inherited as an autosomal recessive
disorder. Hair is white to blond to brown and develops a silvery gray sheen.
Frequent bacterial infections and a bleeding disorder are common and over
time, peripheral neuropathy develops. Bacterial infections are due to
neutropenia and dysfunctional natural killer (NK) cells. Bleeding is due to a
storage pool deficiency in platelets; electron microscopy shows irregular and
a reduced number of dense bodies (130). Hematopoietic stem cell
transplantation (HSCT) is preferred prior to the accelerated lymphohistiocytic
phase, which is fatal, typically within the first decade of life when left
untreated (131,132). HSCT successfully treats the hematologic and
immunologic manifestations of CHS. The neuropathy, with both central and
peripheral components and involving both motor and sensory nerves, appears
to progress despite successful transplantation. HSCT also does not affect the
pigment dilution.
CHS is caused by mutations in the CHS1/LYST gene, which regulates the
size and movement of lysosomal organelles. Enlarged granules that are
peroxidase-positive are found in cells due to abnormal lysosomal
enlargement (Figure 23-9). Melanosomes are enlarged in melanocytes and
delta bodies are enlarged in platelets. The cells may be inappropriately
distributed, accounting for the hypopigmentation, bacterial susceptibility, and
neuropathy (133,134).
FIGURE 23-9 Peripheral blood smear showing neutrophil

with giant cytoplasmic granules that are pathognomonic of
Chédiak-Higashi syndrome. The granules can reach a size
of 4 μ. (Reprinted from Antunes H, Pereira A, Cunha I.
Chediak-Higashi syndrome: pathognomonic feature. The
Lancet 2013;382(9903):1514. Copyright © 2013 Elsevier
Ltd. With permission.)
BenEzra and colleagues described a 1½-year-old Arab boy with recurrent

infections, ecchymoses, pancytopenia, an increased bleeding time,
hepatosplenomegaly, and bone marrow with histiocytes with large lysosomal
cytoplasmic inclusions who was diagnosed with CHS (135). The boy was
more lightly pigmented than unaffected family members and had silvery
white hair and nystagmus. He had no iris transillumination of his blue irises
but had minimal pigment in the retinal pigment epithelium and appeared to
have foveal hypoplasia. Electroretinography (ERG) showed progressive
abnormalities.
DIAGNOSTIC STUDIES
The clinical diagnosis of albinism is made by ocular examination with the
findings of foveal hypoplasia, iris transillumination defects, nystagmus,
decreased pigmentation of the fundus, and lighter skin color than other
members of the family. In those patients in whom the diagnosis of albinism is
unclear or not obvious, visual evoked potentials can be used to show
excessive retinostriate decussation of the nerve fiber at the chiasm in
albinism; this test is helpful if abnormal decussation is documented, but not
finding abnormal decussation does not rule out albinism (136). If the patient
is male and thought to possibly have OA1, the mother can be examined to
look for characteristic mosaic patterns of hypopigmentation in the iris and
retina (137). It is important in all patients with albinism to identify any
patient who may be classified as having an HPS phenotype because these
patients may have associated systemic disease. Therefore, all newly
diagnosed patients with albinism should have either molecular genetic testing
with an albinism panel containing all known genes, electron microscopy of
platelet morphology to look for absence of dense bodies, or platelet
aggregation studies. If there is a history of easy bruising or epistaxis, frequent
infections or lung disease, or if surgery is planned, this testing is imperative
(138). If the patient has silvery hair along with OCA, then the diagnosis of
CHS should be considered and along with a prolonged bleeding time, there
will be giant peroxidase-positive lysosomal-like organelles seen in the blood
leukocytes (139). Alternatively, genetic panels can be sent to see if the patient
has any mutations in the genes known to cause HPS or CHS (138).
Albinism genetic testing panels can be performed to identify the exact
genetic mutation present in each patient with albinism. If the genetic mutation
can be identified, the family can be offered preimplantation genetic testing
and in vitro fertilization with unaffected embryos, allowing the family to
conceive a disease-free child (140).
DIFFERENTIAL DIAGNOSIS
OCA with marked iris transillumination, foveal hypoplasia, nystagmus, and
extremely pale skin is not confused with many other disease entities.
However, as described previously, not all patients have the aforementioned
clinical appearance. Foveal hypoplasia, which is seen in essentially all
albinism patients either clinically or subclinically by OCT, can be seen in
many other disorders including aniridia, achromatopsia, isolated foveal
hypoplasia, Aland eye disease/incomplete congenital stationary night
blindness, Stickler syndrome (141), prematurity, and even in normal eyes
(142–146). ERG testing would differentiate Aland eye/incomplete congenital
stationary night blindness and achromatopsia from albinism. Aniridia can be
differentiated from albinism by abnormalities of the iris (which may at times
be subtle). Isolated foveal hypoplasia does not have the iris transillumination
defects seen in albinism. Slit lamp examination of the iris prior to pupillary
dilation is extremely important in differentiating albinism from other diseases
with foveal hypoplasia when nystagmus is present. Iris transillumination
defects in children can also be seen in X-linked megalocornea, trauma, iris
atrophy (after herpes infection or uveitis), and Axenfeld-Rieger spectrum. In
adults, pseudoexfoliation and pigment dispersion syndrome are the most
common causes of iris transillumination (138). Patients with Stickler
syndrome usually have high degrees of myopia from preschool age.
Albinism is one of the most common etiologies of congenital sensory
nystagmus in infancy. In a study of 202 consecutive patients presenting to
pediatric ophthalmology with congenital nystagmus, 19% had a diagnosis of
albinism (147). Other causes of congenital sensory nystagmus with minimal
fundus changes include Leber congenital amaurosis, congenital stationary
night blindness, achromatopsia, and optic nerve hypoplasia. Clinical
examination can be useful to identify the above retinal dystrophies because
they often present with paradoxical pupils and abnormalities of color vision,
which are absent in albinism. A large positive angle kappa is also useful in
helping make the clinical diagnosis of albinism (148). Electroretinography
and genetic testing can also be used to identify congenital retinal dystrophies.
MANAGEMENT AND VISUAL

REHABILITATION
Ophthalmic Issues
High refractive errors are common in albinism and require treatment with
glasses or contact lenses. Correction of significant refractive errors improves
visual and motor outcomes and should be prescribed by 4 to 6 months of age
(149). Spectacles can also reduce abnormal head position (150). Filtering or
photochromic lenses can alleviate the photosensitivity that occurs in patients
with albinism. Glasses with UV protection are also desirable. Low-vision
aids and services should be considered for patients with albinism; however,
electronic technology to enlarge print has frequently replaced the use of
bifocals, magnifiers, and large print worksheets. Braille is not usually
required. Mobility training can be helpful. Some patients are eligible for
restricted driving licenses in the United States. See Chapter 14 for a full
discussion of visual rehabilitation.
A randomized, controlled clinical trial to evaluate if levodopa, a
precursor to dopamine, which is an intermediate product in the biosynthesis
of melanin pigment, would improve visual acuity in patients with albinism,
did not show a benefit for the doses used (151).
Eye muscle surgery can be performed in patients with albinism for
strabismus, torticollis, and nystagmus. Many patients will have all three
issues. Although fine binocular function is lacking in many patients with
albinism, there are patients who demonstrate some degree of stereopsis (26).
Strabismus surgery to align the oculomotor system has been shown in some
patients with albinism to allow for the development of binocular vision and
should be considered for both functional and psychosocial benefits (152). In
many patients with albinism, after strabismus surgery to align the eyes, the
visual acuity and nystagmus have also been reported to improve (153,154).
Eye muscle surgery in patients with HPS can usually be performed without
excessive risk of bleeding, but hematologic consultation should be obtained
prior to surgery.
Torticollis seen in albinism maybe related to either uncorrected refractive
errors or nystagmus with a null point. If the torticollis is secondary to
nystagmus with a null point, the torticollis can be diminished by performing
eye muscle surgery by moving the null point to primary position and in the
direction of the head turn (155). Extraocular muscle surgery can successfully
be performed to eliminate both horizontal and vertical head posture
secondary to nystagmus with a null point (156).
Because the amplitude of nystagmus appears to be inversely correlated
with the visual acuity in albinism (157), attempts have been made both
medically and surgically to decrease nystagmus and thereby improve vision.
Infantile periodic alternating nystagmus can be seen in up to 33% of patients
with albinism, and in this setting, oral baclofen can be used to diminish
nystagmus (158).
The two primary muscle procedures that have been attempted to decrease
the intensity of nystagmus include large four-muscle horizontal recessions
(159) and four-muscle tenotomy (160). Both procedures have been shown to
decrease nystagmus amplitude but not eliminate it and improve vision
slightly, although large four-muscle recession has a higher rate of induced
strabismus (160,161).
Although rhegmatogenous retinal detachments are rare in persons with
albinism, they can be more difficult to treat due to difficulty in visualizing the
break and in delivering the endolaser due to lack of melanin pigment.
(Cryotherapy may offer a better solution.) However, the risk for proliferative
vitreoretinopathy appears to be low and may also be due to lack of pigment
(162).
Dermatologic Issues
Strict sun protection in albinism should begin in infancy and be continued
lifelong because affected persons are at increased risk for all types of skin
cancer (163). This includes avoiding UV light exposure during peak hours of
sunlight, wearing protective clothing, using sunscreen and frequent
reapplication, and avoiding medications that increase photosensitivity. Recent
studies have found that sunscreen ingredients attain blood levels after topical
application (164). No detrimental effects have been shown for these levels,
and the increased risk of skin cancer must be considered, but frequent
dermatologic examinations should begin by the teenage years. Any
suspicious or concerning lesions should be evaluated by a dermatologist.
Systemic Issues
Children with albinism have normal neurologic development despite visual
impairment (165). However, children and adults have a higher prevalence of
attention deficit/hyperactivity disorder (ADHD). The prevalence of ADHD
appears to be unrelated to best corrected visual acuity or type of albinism
(166).
Because sunlight avoidance is recommended in OCA, patients with
albinism may not produce enough vitamin D. However, studies have shown
that vitamin D levels tend to be adequate due to enough incidental UV light
exposure (167). If there is concern about low vitamin D levels, then serum
vitamin D levels can be drawn.
At this writing (May 2019), there are no gene therapy trials for albinism.
However, research is being done in mouse models that may be applicable for
human studies. An AAV vector to supply human tyrosinase to a mouse model
for OCA1 resulted in melanin production in neuroectodermally derived
(RPE) and neural crest–derived (choroid, iris) cells (168). Another study
successfully used lentiviral-mediated transfer of the HPS-1 gene in vitro into
a patient’s dermal melanocytes (169). One mouse model study has shown
that eye and skin pigmentation can be improved in OCA1B mice, which has
residual tyrosinase activity, by treatment with nitisinone, which is an FDA-
approved inhibitor of tyrosine degradation (170). Nitisinone also increased
melanin in melanocytes in a human with OCA1B. A similar effect was not
observed in OCA3 mice in which albinism is caused by mutations in
tyrosinase-related protein 1 (171). A human study in patients with OCA1B is
under way to determine if visual function can be improved with nitisinone.
Patients with either HPS or CHS should avoid the use of aspirin and
aspirin-containing products and nonsteroidal anti-inflammatory agents. Skin
wounds can be treated with gel foam soaked in thrombin. Prophylaxis with
aminocaproic acid or desmopressin can be helpful, but when surgical wounds
are expected to be more extensive, platelet transfusions may be needed to
control bleeding.
Persons with CHS require treatment with antibiotics for infections
although their response to treatment may be slower than normal. Early
treatment with HSCT prior to the accelerated phase treats the immunologic
and hematologic complications of CHS.
Persons with HPS and pulmonary fibrosis require preventative and
supportive care. Influenza and pneumococcal vaccines should be given.
Oxygen can be given as needed and air pollutants, including smoking, should
be avoided (97). Neither steroids nor pirfenidone, an antifibrotic agent, has
been shown to be effective for treatment of pulmonary fibrosis in HPS (172).
The only effective treatment for the advanced lung disease in HPS is lung
transplantation. Three patients have been reported with successful lung
transplant and no evidence of recurrence up to 6 years after transplant.
Avoidance of alloimmunization is recommended for optimal success (173).
Genetic testing and/or platelet testing should be offered for ascertaining
which albinism patients have syndromic, potentially life-threatening forms.
Genetic counseling should be offered for family planning purposes.
FUTURE TREATMENTS
There is hope that medical treatment such as nitisinone, or gene replacement
therapies, may be available in the future for patients with albinism.
RESOURCES
National Organization for Albinism and Hypopigmentation (NOAH)
—www.albinism.org
World Albinism Alliance—worldalbinism.org
Albinism Fellowship UK and Ireland—www.albinism.org.uk
Albinism Trust New Zealand—albinism.org.nz Albinism Fellowship of
Australia—www.albinismaustralia.org HPS Network—HPSnetwork.org
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24
Neuronal Ceroid Lipofuscinoses and
Lysosomal Storage Diseases
Sana Idrees, and Mina Chung
Lysosomal storage diseases (LSDs) are a group of over 70 monogenetic

disorders affecting the function of the lysosome, a cytoplasmic organelle
responsible for the metabolism of macromolecules, which is critical for
cellular homeostasis. Mutations in genes encoding lysosomal enzymes or
integral membrane proteins lead to the accumulation of incompletely
degraded substrates in the lysosome and ultimately cell death. The majority
of LSDs are autosomal recessively inherited with the exception of MPS II
and Fabry disease, which are X-linked. LSDs can be classified based on the
biochemical type of stored material, such as glycan, lipid, protein, or
lipofuscin, and further subdivided by the age of onset and disease severity.
Individually, these conditions are rare, but taken together, LSDs affect
approximately 1 in 8,000 live births (1). Prevalence of certain LSDs can be
higher in select populations due to founder effects.
The clinical features of the various LSDs depend on the specific substrate
accumulated and its systemic distribution. Most LSDs present with
neurodevelopmental regression and organomegaly. Other systemic features
include skeletal deformities and cardiac abnormalities. Ocular manifestations,
including cataract, glaucoma, vitreous degeneration, retinopathy, and optic
nerve swelling or atrophy, are common and can result in severe visual
impairment.
Diagnosis of LSDs is made based on the clinical presentation and can be
confirmed by measurement of elevated substrate levels, enzymatic analysis,
or single gene testing. Because the presenting symptoms are nonspecific and
these conditions are rare, diagnosis may take time to establish particularly in
milder or later-onset cases. LSDs should be considered in children who
initially achieve developmental milestones and then plateau or regress. The
advent of next-generation genetic testing, particularly whole exome
sequencing, may aid in earlier diagnosis in the future.
The management of LSDs requires a multidisciplinary approach as
supportive care can prolong life expectancy and improve quality of life.
Carrier screening, newborn screening, and prenatal screening can help to
prevent the occurrence of LSDs in at-risk populations. Some LSDs are
treatable with enzyme replacement therapy (ERT), substrate reduction
therapy (SRT), gene therapy, or hematopoietic stem cell transplantation
(HSCT) (2).
LSDs with ophthalmic involvement, in particular neuronal ceroid
lipofuscinosis, will be the focus of this chapter. Please also see Chapter 25.
NEURONAL CEROID
LIPOFUSCINOSES
The neuronal ceroid lipofuscinoses (NCLs) are a unique group of inherited
lysosomal storage disorders characterized by retinopathy progressing to
blindness, progressive decline of cognitive and motor capabilities, variable
cerebellar atrophy, myoclonic epilepsy, and premature death. Storage
material is found in various cell types, but the destruction of tissue is largely
confined to the central nervous system. There are 14 genetically distinct
forms of human NCLs. Classically, NCLs were categorized by the age of
onset, but more recently as causative genes have been identified, a genetic
classification has emerged.
Otto Christian Stengel published the first clinical description of NCL in
Norway in 1826. Stengel was a general practitioner who observed four
siblings with unremarkable early development go on to develop blindness,
progressive mental deterioration, loss of speech, and epileptic seizures. Two
of the siblings died by age 20 and 21 years (3). However, Stengel’s
description went unnoticed to the scientific community until the 1950s (4).
In the early 20th century, further studies of familial cases of progressive
vision loss and psychomotor retardation of later onset were described. Late
infantile NCL (LINCL) was described by Jan Janský and Max Bielschowsky,
otherwise known as Janský-Bielschowsky disease (5,6). Frederick Batten,
Wolfgang Spielmeyer, and Heinrich Vogt all described juvenile NCL
(JNCL), which was known as Batten-Spielmeyer-Vogt disease at that time
(7–9). In 1925, Hugo Kufs described adult-onset mental deterioration with
similar intraneuronal storage but without vision loss. This subtype was later
termed adult NCL (ANCL) or Kufs disease (10). Later, Matti Haltia and
Pirkko Santavuori identified a subtype of NCL with early onset that is now
known as infantile NCL (INCL) or Haltia-Santavuori (11–13).
The term Batten disease has been used in the literature to refer to NCLs
as a group but, in particular, JNCL.
Incidence
Collectively, the NCLs are the most common cause of childhood-onset
neurodegeneration (14,15). The incidence of NCLs varies by geographic
area. The estimated incidence in the United States is 1.6 to 2.4/100,000,
whereas in Finland, the incidence is estimated at 4.8/100,000 and in Iceland,
7/100,000 (16,17).
Genetics
At least 14 genes, CLN1 through CLN14, have been implicated in the
development of NCL. Most NCLs are autosomal recessively inherited,
although cases of autosomal dominant NCL have been described (18,19).
Additional genes and mutations are being recognized regularly; below is a
summary of known genes reported in PubMed at this writing.
The underlying defect in CLN1 disease is the lack of activity of the
lysosomal palmitoyl protein thioesterase (PPT1), an enzyme that removes
palmitate residues from proteins (20). The physiologic function of PPT1 is
not well understood but appears to be necessary for the maintenance of
cellular processes, such as apoptosis, endocytosis, vesicular trafficking,
synaptic function, and intracellular signaling (21). To date, 71 mutations have
been described in CLN1 (22).
CLN2 encodes the lysosomal enzyme tripeptidyl peptidase (TPP1) of the
serine carboxyl proteinase family, and mutations in this gene generally cause
LINCL (23). Thus far, 129 mutations in this gene have been found (22).
TPP1 has been associated with the removal of tripeptides from the N-termini
of small polypeptides, such as the subunit c of mitochondrial ATP synthase
(24).
The CLN3 gene associated with the most common subtype, also termed
JNCL or Batten disease, encodes a membrane protein whose function is
unknown. This protein is present predominantly in the endolysosome system,
but it has been reported to localize to membrane lipid rafts in synaptosomes,
Golgi apparatus, cell membrane, and mitochondria (25). Seventy-eight
mutations have been identified in CLN3. In most patients, at least one of their
two mutations is a common ancestral 1-kb deletion founding mutation (26).
The CLN4/DNAJC5 gene encodes cysteine-string protein α (CSPα),
which underlies the autosomal dominant adult form of NCL that is also
known as Parry disease. It has been suggested that CSPα is a substrate of
PPT1, and thus, CLN1 and CLN4 diseases are related, and aberrant protein
palmitoylation may be critical in the pathogenesis of NCL disease (27,28).
CLN5 encodes a soluble protein that is directed toward the lysosome and
reported to interact with the gene products of CLN2 and CLN3, which
suggests that there may be a common molecular pathway or critical
interaction between pathways involved in various NCL subtypes (20). To
date, 37 mutations in this gene have been identified (22).
CLN6 encodes a protein of unknown function with seven transmembrane
domains localized to the endoplasmic reticulum (ER) (29,30). Seventy
mutations have been identified in CLN6 (22).
The CLN7 gene product MFSD8 belongs to the large major facilitator
superfamily (MFS), which transports specific classes of substrates (31).
There are 38 known disease-causing mutations in this gene (22).
CLN8 encodes a polytopic membrane protein that localizes to the ER and
shuttles between the ER and ER–Golgi complex. The exact function of the
protein is unknown. It is known to belong to the TRAM-Lag1p-CLN8 (TLC)
protein family, which has roles in biosynthesis, metabolism, transport, and
detection of lipids (31). Thirty-five mutations in this gene have been
identified (22).
CLN9 has been proposed as a NCL entity, but no specific gene has been
identified (32).
CNL10 encodes for cathepsin D (CTSD), a lysosomal enzyme involved
in neuronal stability. Alterations in the macroautophagy–lysosomal
degradation pathway have been implicated in the mediation of
neurodegeneration in CNL10 disease (33,34). Ten mutations have been
described in this gene (22).
Mutations in the GRN gene have been implicated in the development of
CLN11 disease (35).
CNL12 disease was reported in a single family and resulted from a
mutation in ATP13A2 (36).
CLN13 has been associated with 11 mutations involving cathepsin F
(CTSF) deficiency (22,37).
A single mutation has been associated with CLN14, which encodes for
KCTD7, a potassium channel tetramerization domain-containing protein 7
(38).
Pathophysiology
Molecular genetic studies have identified more than 360 mutations in the 14
genes underlying the various subtypes of human NCL (39). The genes
implicated in NCL are of importance in the normal development and
maintenance of cerebral neurons. Some of the known genes encode for
soluble lysosomal enzymes. Several of the other known genes involved in
NCL encode for transmembrane proteins with largely unknown functions.
The molecular genetic heterogeneity of NCLs is in stark contrast with the
uniform histopathologic features of the group of NCL diseases (40).
All forms of NCL share at least two essential features. First, there is
accumulation in the lysosomes of nerve cells and, to a lesser extent, other
cells of autofluorescent, electron-dense, periodic acid-Schiff and Sudan black
B-positive granules. The storage material in INCL is easily extractable and
autofluorescent. The electron-dense storage cytosomes in LINCL and JNCL
are largely resistant to lipid solvents (41,42). The second essential feature is
that NCL is associated with progressive and selective loss of neurons,
particularly in the cerebral and cerebellar cortex, and less consistently in the
retina. These changes are associated with mental, motor, and visual
deterioration (40,43).
NCLs are considered LSDs as many of the NCL-associated proteins are
present in lysosomes and lipofuscin-like ceroid lipopigments also accumulate
in lysosomes in NCLs. However, unlike classic LSDs, the storage material in
NCLs is not disease specific. The ultrastructural inclusion patterns under
electron microscopy were formerly thought to be unique and characteristic
for each major clinical subtype of NCL but have since been demonstrated to
not correlate absolutely with clinical presentation. A single subtype of NCL
may contain more than one pattern of inclusion (44).
Studies of NCL mouse models have suggested that the pathophysiology
of the disease involves dysregulation of autophagy. Animal models of CLN3,
CLN5, CLN6, and CLN7 disease have shown lysosomal dysfunction in the
brain and defective autophagosome maturation with increased autophagic and
lysosomal compartments and substrates (45–51).
Clinical Signs and Symptoms

NCLs have variable ages of onset and severity. The clinical characteristics of
most childhood forms of NCL include progressive loss of vision, mental and
motor deterioration, epileptic seizures, and premature death.
Classically, infantile CLN1 disease manifests between 6 and 12 months
of age with irritability followed by rapid psychomotor deterioration, central
hypotonia, and deceleration of head growth. Myoclonic jerks and other
seizure types quickly follow this. The disease continues until death in early
childhood. CLN1 disease has the widest range of age of onset (52).
Late infantile CLN2 disease usually presents in the third year of life with
intractable epilepsy and arrest of cognitive development. Early in the disease
course, myoclonus and ataxia are commonly seen. Subsequently, progressive
cognitive and motor decline are observed. Retinopathy is usually not
prominent early in the disease course and can be overlooked as individuals
progress to more severe neurologic deficits, including spasticity, truncal
hypotonia, loss of head control, near-continuous myoclonus, frequent
seizures, and extended vegetative state until death in early adolescence. Death
is often secondary to aspiration pneumonia (53).
Juvenile CLN3 disease is of particular importance to ophthalmologists as
the presenting symptom is vision loss starting between the ages of 4 and 8
years. Initially, the ophthalmic signs can be indistinguishable from Stargardt
disease or cone dystrophy, but the vision loss is then followed by a
progressive neurologic decline. During adolescence, epilepsy and
extrapyramidal or parkinsonian signs, such as rigidity, hypokinesia, shuffling
gait, and impaired balance, become more prominent. Anxiety and aggression
are also common (54). The clinical course can be variable, but the disease is
usually fatal in the second or third decade. CLN3 disease may also present in
adulthood with vision loss that is sometimes followed by heart failure (55).
CLN4 disease, autosomal dominant ANCL, presents at 25 to 45 years of
age with variable symptoms including generalized seizures, dyskinesia,
psychiatric manifestations, progressive dementia, and early death. Vision is
not impaired (28).
The first symptoms of CLN5 disease, known as variant LINCL, typically
present between ages 4 and 7, but cases of presentation through adulthood
have also been reported. The usual course of the disease is motor clumsiness
followed by progressive vision loss and blindness, dementia, motor decline,
myoclonus, and seizures. The rate of progression is variable. Ultimately,
death occurs between ages 14 and 36 years (44).
CLN6 disease has an age of onset ranging from 18 months to 8 years of
age, most between 3 and 5 years. Visual deterioration occurs early in the
disease in half of affected individuals. The most prominent symptoms are
motor impairment, developmental delay, dysarthria, and ataxia. Seizures
usually begin before age 5 and occur in a majority of individuals with CLN6
disease. Progression is rapid, and most children die between age 5 and 12.
(52)
CLN7 disease has an age of onset between 2 and 7 years of age. The
initial presenting signs are psychomotor regression or seizures. Later,
progressive cognitive and motor decline, myoclonus, personality changes,
and blindness develop (56).
CLN8 disease causes LINCL or Northern epilepsy (NE). LINCL
secondary to CLN8 mutation presents with mild developmental delay in late
infancy with progressive myoclonus, seizures, retinopathy with vision loss,
and psychomotor regression beginning at age 3 to 6 years (57). NE has an age
of onset of 5 to 10 years with seizures as the presenting symptom and is
followed by progressive cognitive decline. Patients with NE may survive
until 50 to 60 years of age (58).
CLN9 disease is clinically similar to juvenile CLN3 disease but may have
a more rapid course. (52)
In the congenital form, CLN10 disease is characterized by primary
microcephaly, neonatal epilepsy, respiratory insufficiency, and rigidity.
Death occurs shortly after birth within hours to weeks. A late onset form of
CLN10 disease can be seen in juveniles and adults (34).
LN11 disease is characterized by rapidly progressive vision loss due to
retinal dystrophy, seizures, cerebellar ataxia, and cerebellar atrophy with
onset typically in adulthood (35,59).
CLN12 disease was described in a Belgian family with symptoms of
unsteady gait, myoclonus, and mood disturbance at age 11 to 13, which
progressed to clear extrapyramidal involvement with akinesia, rigidity, and
dysarthric speech. There was no retinal involvement reported (36).
The clinical phenotype of CLN13 disease, also known as Kufs type B, is
characterized by behavioral abnormalities and dementia with adult onset.
Symptoms may include motor dysfunction, ataxia, extrapyramidal signs, and
bulbar signs (37,60–62).
Visual decline is a consistent feature of INCL, LINCL, and JNCL. In
JNCL, rapid visual decline is most commonly the presenting feature of the
disease. Early macular changes, pigmentary abnormalities, vascular
attenuation, and optic pallor are all well-established ophthalmic
manifestations of NCL (63). More recently, cataract and acute secondary
glaucoma development in JNCL have been described (64).
Diagnostic Studies
The approach to diagnosis of NCL should take into account the age of onset
and clinical presentation. With the advent of gene panels for specific clinical
presentations, such as retinal degenerations, and panels of NCL genes,
molecular genetic testing is usually the first diagnostic step. However,
clinical features in combination with electron microscopy demonstrating
characteristic storage material can also be diagnostic of NCL, albeit without
specificity of type. Enzymatic testing can be used to diagnose some subtypes.
For example, epilepsy and microcephaly in a neonate should prompt further
investigation into CLN10 disease as a possible diagnosis, and enzymatic
testing for CTSD can be considered. If enzymatic testing is negative for
CTSD, further or concurrent testing for PPT1 (CLN1) and TPP1 (CLN2)
should be considered. The benefit of performing molecular genetic testing
initially is that if positive, it confirms NCL and gives a granular description
of genetic subtype and mutation. The downside is that because not all genes
or mutations are known, molecular genetic testing is sometimes normal in
patients with NCL. Index of suspicion must remain high, and multimodal
testing including molecular genetics, biopsy, leukocyte assay, and enzyme
levels may be used in concert.
CLN1 and CLN2 diseases are more likely considerations in young
children under the age of 6 with otherwise unexplained epilepsy and
developmental arrest. Even if enzyme testing for PPT1 and TPP1 is negative,
if electron microscopy demonstrates characteristic storage material, genetic
testing for CLN5, CLN6, CLN7, CLN8, and CLN14 should be performed.
School-aged children between ages 4 and 7 presenting with rapid vision
loss should be tested for CLN3 disease. CLN3 is included in many early-
onset retinal degeneration panels, and parents of children being tested with a
broad panel should be counseled ahead of time about the possibility of
neurologic conditions on the panel. Lymphocytes can be assessed for the
presence of vacuoles, and enzymatic testing for PPT1, TPP1, and CTSD can
be performed. A skin biopsy is indicated to determine if characteristic NCL
storage material is present either simultaneously or after other testing is
negative. A mucosal, conjunctival, or skin biopsy will demonstrate the typical
storage material in all the genetic subtypes.
In adults with suspected NCL, the first line of diagnostic investigations
includes enzymatic assays for PPT1, TPP1, CTSD, and CTSF. If the
enzymatic assays are normal, ultrastructural examination for storage material
should be performed. If storage material is present, genetic testing for
autosomal recessive CLN6, CLN11, CLN13, and autosomal dominant CLN4
should be considered. If negative, the remaining genes associated with NCL
should be tested.
In high-risk cases, prenatal testing on fetal cells obtained by chorionic
villus sampling at 10 to 12 weeks of gestation or by amniocentesis between
15 and 18 weeks of gestation can reveal deficient activity of enzymes PPT1,
TPP1, and CTSD or mutations associated with NCL (65). Couples who have
had a child affected with NCL have a 25% risk with each subsequent
pregnancy of another affected child. If genetic testing is performed on the
affected child, parents may choose to use in vitro fertilization with
preimplantation genetic testing for subsequent conceptions to reduce the risk
of having other affected children.
Given that visual decline is among the first symptoms in the most
common subtype, CLN3, detection of retinal degeneration, which occurs
early in the LSD, is critical in diagnosis. Ophthalmoscopic findings may
include cystoid macular edema, bull’s-eye maculopathy, macular–retinal
pigment epithelium (RPE) atrophy, attenuated retinal vessels, peripheral
pigmentary deposition, and widespread atrophy in advanced disease (Figure
24-1A) (66–69).
FIGURE 24-1 Multimodal imaging of an 8-year-old

patient with CLN3 disease. A: Fundus photograph shows
macular RPE atrophy, arteriolar attenuation, and
peripheral depigmentation. B: FAF image demonstrates
central macular hypoautofluorescence with surrounding
annular hyperautofluorescence. C: OCT shows loss of the
foveal ellipsoid zone and outer nuclear layer with
hyperreflective deposits at the level of the RPE. Note
bull’s-eye maculopathy seen on fundus photography and
FAF imaging. Macula OCT shows absent outer nuclear
layer under the fovea and thinning of all retinal layers,
inner and outer, characteristic of a neuroretinopathy.
Fundus autofluorescence (FAF) imaging demonstrates central retinal

hypoautofluorescence with surrounding hyperautofluorescence (Figure 24-
1B). In advanced cases, generalized hypoautofluorescence can be observed
(69). These clinical autofluorescence findings are consistent with
histopathologic reports of hypoautofluorescence of RPE cells, photoreceptor
cell death, and reduced RPE phagocytosis of the retinal outer segment
(70,71).
Optical coherence tomography (OCT) in early JNCL shows thinning of
the outer nuclear and photoreceptor layers, abnormal homogeneity of the
optical reflectivity in the inner retinal layers, and hyperreflective granules in
the RPE (Figure 24-1C) (72). In a study of 11 patients with JNCL, spectral-
domain OCT demonstrated degeneration within the ganglion cell complex
ranging from fine rippling of the inner limiting membrane to extensive loss of
the nerve fiber layer, and confocal scanning laser ophthalmoscopy (cSLO)
showed a radial pattern of striation surrounding the macular area but sparing
the fovea (73). In patients with preserved visual acuity, OCT demonstrates
sparing of the foveal ellipsoid zone but absence of the ellipsoid zone and
thinning of the outer nuclear layer outside the fovea on OCT. Decreased
visual acuity corresponds to moderate to complete ellipsoid zone and outer
nuclear loss. Hyperreflective deposits at the level of the RPE are visible with
advanced disease (69).
Findings from electroretinograms (ERGs) in early cases of INCL and
JNCL include marked loss and increased implicit latency of the scotopic and
photopic b-wave with relative preservation of the a-wave resulting in a
characteristic electronegative ERG (see ERG chapter in Appendix). These
changes suggest relative preservation of the photoreceptor outer segment
function with severely disrupted transmission of signals to bipolar cells. As
the disease progresses, ERG responses become undetectable (63,74,75). In
CLN3-associated isolated retinal degeneration, ERG showed markedly
abnormal scotopic responses with variable photopic responses shortly after
disease onset. Patients with earlier onset of disease showed more severe loss
of both rod and cone responses, and patients with later onset showed
relatively preserved photopic responses (69). In CLN8 disease, the retina may
appear normal early in the course, but ERG will be abnormal and becomes
progressively more so over time (Figure 24-2).
FIGURE 24-2 Full-field ERG (ERG) at two timepoints in

a child with CLN8. At 9 years of age, the light-adapted
(LA) cone ERG (A) is low in amplitude and delayed; the
dark-adapted (DA) rod ERG has very low and delayed dim
flash (B). 30-Hz flicker shows artifact due to flinching at
the flickering light (C). Bright flash (D) amplitudes are
low and delayed, and oscillatory potentials are absent (E).
Of note, the ERG is not electronegative as seen in CLN3
(see ERG Atlas, Chapter 10). At 10 years of age, the ERG
of the same child shows that LA cone ERG (F) is stable
and 30-Hz flicker response is again obscured by artifact
(H). The amplitudes in the DA rod ERG (G, I, J) are now
essentially nonrecordable. The child still had ambulatory
vision at age 10 years.
Histopathologic evaluation of the retina in NCL demonstrates decreased cell

counts and autofluorescent lipofuscin granules in all layers of the retina with
central epiretinal membrane. The peripheral retina remains better preserved
but with shorter photoreceptor outer segments. Immunofluorescence analysis
demonstrates degenerated rods and cones throughout the retina with
improved preservation in the periphery (75,76).
Management
The treatment of NCLs is largely symptomatic and directed to improve the
quality of life. Antiepileptics of choice are valproic acid and lamotrigine
(77–79). Benzodiazepines and sedatives are commonly used for sleep
disturbance, anxiety, and spasticity (80). Melatonin has also been used, but
evidence regarding its efficacy has been inconclusive (81–83).
Trihexyphenidyl improves dystonia and sialorrhea. Delusions and
hallucinations can be managed with atypical neuroleptics, such as risperidone
or olanzapine. Selective serotonin reuptake inhibitors may be useful if
depression is suspected.
Vision Rehabilitation
There is no treatment for retinal degeneration in NCL. As the visual
deterioration progresses, affected individuals can be referred for low vision
aids to optimize their ability to function.
Roles of Other Physicians and Health Care Providers
Patients with NCLs benefit from a team-based management approach. They
are typically followed by a primary care provider, neurologist, and
ophthalmologist. Additionally, they often benefit from therapy by a physical
therapist, occupational therapist, and speech therapist. Palliative care should
be involved early in the course to support families.
Ethical Considerations
When the disease is advanced in NCL, patients often cannot be fed naturally.
The decision to institute or withhold artificial nutrition should be made by the
parents or their representatives who are emotionally and intellectually
prepared to do so (84).
Future Treatments
NCL genes appear to be significant in the normal development and
maintenance of cerebral neurons. Elucidation of their specific functions and
interactions in health and disease is important for the identification of
therapeutic targets. Progress in the understanding of the natural history and
the biochemical and molecular cascade of events involved in the
pathogenesis of the NCLs will be required to achieve significant therapeutic
advances in the future.
Current and future approaches to treatment of NCL involve enzyme
replacement, gene therapy, neural stem cell replacement, immune therapy,
and other pharmacologic approaches (85).
ERT is being actively investigated as a therapeutic option for LSDs,
including NCLs. Numerous animal models of ERT have been studied and
shown reduction in the storage material accumulation in the brain and
improved disease phenotype and pathology (86–93). Cerliponase alfa
(recombinant TPP1) has been FDA approved for treatment of CLN2 disease.
A study of 17 patients receiving intraventricular infusion every 2 weeks over
a period of 2 years compared to matched historical controls showed a slower
rate of decline in motor and language function compared to controls (94).
Overall ERT appears to be a promising therapeutic approach for NCL.
Feasibility may be limited by the need for intraventricular infusion, but
further investigations including the development of compounds that can cross
the blood–brain barrier are necessary.
Gene therapy studies in animal models using adeno-associated virus and
lentivirus vectors have shown promise in the treatment of NCLs (95–102).
Gene therapy clinical trials in humans are underway, and a preliminary report
from study NCT00151216 (ClinicalTrials.gov) showed promise in reducing
the rate or progression in LINCL (103).
HSCT with umbilical cord blood and bone marrow transplantation has
been attempted in patients with NCL. However, the effect has been small
(104–107). Neuronal stem cell therapies have been investigated for LSDs,
(108,109) and a phase I clinical trial studying human neural stem cell
transplantation in INCL and LINCL showed that the therapy was well
tolerated (110). Further studies are required to explore the efficacy of stem
cell therapy in NCLs.
Antioxidant therapy has also been considered for treatment in NCL. This
is based upon the theory that free oxygen radicals cause a peroxidation defect
of polyunsaturated fatty acids, which leads to damage to fat deposits.
However, there is no clear conclusion on the effectivity of antioxidants in the
treatment of NCLs (111,112).
The use of anti-inflammatory medications has been explored based upon
the suggestion that inflammation throughout the central nervous system is a
key component in the pathogenesis of neurodegeneration in NCL. Mouse
model studies have shown decreased neuroinflammation, decreased
deposition of immunoglobulin G in the brain, and neuroprotective effects of
mycophenolate mofetil (113). Mycophenolate has been shown to be well
tolerated in JNCL, but further studies regarding its efficacy are necessary
(114,115).
In INCL disease, the affected enzyme cleaves fatty acid thioesters in
plasma membranes, and it has been suggested that small-molecule
compounds cysteamine and N-acetylcysteine may provide some benefit.
Studies of these two drugs have shown a delay in the development of
isoelectric EEG, depletion of the characteristic storage deposits seen on
electron microscopy known as granular osmiophilic deposits (GRODs), and
subjective benefits, such as decreased irritability as reported by parents and
physicians. However, they have not shown significant benefit in halting
disease progression (116,117).
An ideal treatment approach for NCL would be a drug-based therapy,
which would be less invasive than a gene or enzyme replacement methods
that require surgical access to the central nervous system. However, treatment
with most drugs or with gene or enzyme replacement would all face the
challenge of crossing the blood–brain barrier (118). Work in animal models
has suggested that there may be benefit in presymptomatic treatment (118).
Clinical trial designs must be optimized, and target outcome measures are
selected for these rare conditions (119,120). As new treatments become
available, a heightened awareness among clinicians can facilitate earlier
diagnosis.
SIALIDOSIS
Genetics
Sialidosis type 1 and type 2 are autosomal recessively inherited (121). Both
types are caused by mutations in the NEU1 gene (122).
Pathophysiology
Deficiency of the lysosomal enzyme neuraminidase 1 encoded by NEU1
leads to a disorder of glycoprotein degradation and the lysosomal
accumulation of sialylated glycopeptides (121).

Type 1 sialidosis, a milder form, presents in the second or third decade of life
with visual and neurologic symptoms, including night blindness, color vision
abnormalities, myoclonic epilepsy, ataxia, and dysarthria. On fundus
examination, a cherry-red spot in the macula is commonly seen, for which
sialidosis is also known as “cherry-red spot myoclonus syndrome” (see
Chapter 25 for a photograph of the cherry-red spot). Nystagmus and corneal
or lenticular opacities may also occur (121,123–125).
Type 2 sialidosis has an earlier onset of symptoms and is further
subdivided into congenital, infantile, and juvenile forms. The congenital form
may present with hydrops fetalis. A cherry-red spot is seen on fundus exam
in most but not all affected individuals. Other features characteristic of type 2
sialidosis include hepatosplenomegaly, dysostosis multiplex, developmental
delay, and abnormal facies (121,126).
Diagnostic Studies
The diagnosis of sialidosis is suggested based upon increased urinary bound
sialic acid excretion (121). A definitive diagnosis can be made with genetic
analysis (126).
GALACTOSIALIDOSIS
Genetics
Galactosialidosis is an autosomal recessive condition (127,128) associated
with mutations in the CTSA gene (129).
Pathophysiology
The protective protein/cathepsin A (PPCA) encoded by CTSA forms a
complex with the lysosomal enzymes β-galactosidase and neuraminidase to
digest glycoproteins. Defective PPCA leads to decreased β-galactosidase and
neuraminidase function (127,128).

Early infantile, late infantile, and juvenile/adult forms of galactosialidosis
occur. The early infantile form can present with hydrops,
hepatosplenomegaly, psychomotor delay, coarse facies, and skeletal
dysplasia. Heart failure leads to premature death, typically in early infancy. A
cherry-red spot in the macula can be seen on fundus exam (127–129).
In the late infantile form, characteristic findings include cardiac
involvement, hepatosplenomegaly, psychomotor retardation, corneal
clouding, and macular cherry-red spot. The mean life expectancy is about 15
years (127–129).
The juvenile/adult form is mainly seen in individuals of Japanese descent.
The mean age of onset is 16 years but can be much later. Affected individuals
exhibit myoclonus, ataxia, neurologic deterioration, and macular cherry-red
spot. Age of death varies between 13 and 45 years (127–129).
Diagnostic Studies
Diagnosis is based upon an enzyme assay demonstrating combined
deficiency of β-galactosidase and neuraminidase (127,128).
GAUCHER DISEASE
Prevalence
Gaucher disease is the most common LSD with an estimated prevalence of 1
in 40,000 worldwide and as high as 1:1,000 among Ashkenazi Jews (130).
Genetics
Gaucher disease is an autosomal recessive caused by mutations in the gene
encoding β-glucosidase (GBA) (131).
Pathophysiology
Deficiency in β-glucocerebrosidase results in intracellular accumulation of
glucosylceramide within macrophages, termed “Gaucher cells.” (132).

Gaucher disease is subdivided into three clinical phenotypes which
encompass a spectrum of disease severity. Type 1, comprising 90% of cases,
does not have neurologic involvement and is characterized by
hepatosplenomegaly, anemia, thrombocytopenia, and bone pain. Type 2,
termed acute neuronopathic, presents at birth or early infancy with seizures,
spasticity, pyramidal signs, severe developmental delay, and death by age 2
years. Type 3, a subacute neuronopathic form, includes the visceral and bony
features with oculomotor apraxia, hearing loss, and cognitive decline in
adolescence or early adulthood (133).
Ophthalmic features include white deposits on the corneal endothelium,
anterior chamber angle, vitreous, and retina (134). Progressive macular
atrophy has been described (135).
Diagnostic Studies
The detection of insufficient β-glucosidase enzyme activity in peripheral
leukocytes is diagnostic of Gaucher disease, which can be definitively
confirmed by identification of two pathogenic variants on the GBA gene
(136).
NIEMANN-PICK DISEASE
Genetics
Niemann-Pick disease (NPD) types A and B are caused by mutations in the
sphingomyelin phosphodiesterase-1 gene SPMD1 (137) and type C by
mutations in NPC1 and NPC2 (138). All three types are autosomal
recessively inherited.
Pathophysiology
NPD is characterized by defective lysosomal sphingomyelin metabolism. In
types A and B, defects in acid sphingomyelinase (ASM) encoded by SPMD1
lead to the accumulation of sphingomyelin (139). In type C, defects in NPC
proteins lead to impaired egress of cholesterol from the lysosomes, resulting
in the accumulation of unesterified cholesterol, a major component of
sphingomyelin (138).
Type A NPD, a severe infantile form, presents in early infancy with
hepatosplenomegaly, hypotonia, and weakness, with evidence of
developmental delay at 6 months of age. At age 1 to 2 years, fundus exam
may demonstrate a cherry-red spot in the macula, and ERG is abnormal
(140). Some affected children may demonstrate a gray, granular appearance
of the macula, which is referred to as the macular halo syndrome (141).
Survival beyond 2 years of age is rare (137).
Type B NPD tends to be milder in its phenotype. Clinical presentation is
typically later. In some cases, splenomegaly does not become apparent until
adulthood. Patients with type B NPD may develop respiratory complications
at 15 to 20 years of age secondary to alveolar infiltration. The central nervous
system is usually spared. Some patients with type B NPD may have cherry-
red spots in the macula. The age of onset and rate of progression is variable,
and they frequently live into adulthood (137,139).
Type C NPD classically presents in late childhood with survival into the
second decade; however, a spectrum of phenotypes can occur ranging from
neonatal jaundice, vertical gaze palsy, seizures, and neurodegeneration, to
late onset, slowly progressive disease with intellectual impairment beginning
in adolescence or early adulthood (142).
Diagnostic Studies
The diagnosis of type A or B NPD is confirmed by measurement of
decreased ASM levels from peripheral leukocytes, cultured fibroblasts, or
lymphoblasts (137). Bone marrow biopsy may demonstrate lipid-laden
macrophages termed “sea-blue histiocytes.” In type C NPD, cultured
fibroblasts show impaired cholesterol esterification and positive filipin
staining (142).
Management
Type C NPD is treatable with the SRT miglustat, which can be administered
orally, cross the blood–brain barrier, and slow the progression of neurologic
symptoms. Improved swallowing reduces the incidence of aspiration
pneumonia (143–145).
GM1 GANGLIOSIDOSIS
Genetics
GM1 gangliosidosis is an autosomal recessive disorder caused by mutations
in β-galactosidase-1 (GLB1) (146).
Pathophysiology
Deficiency of β-galactosidase leads to lysosomal ganglioside accumulation
and neurodegeneration. The clinical presentation is variable, with infantile,
late infantile/juvenile, and adult forms (146).

Type I GM1 gangliosidosis, the severe infantile form, presents in early
infancy with arrested development followed by developmental regression.
Exaggerated startle response, rigidity, spasticity, and seizures are common.
Dysmorphic features, skeletal dysplasia, cardiomyopathy, and hepatomegaly
are characteristic with eventual regression to a vegetative state.
Ophthalmologic exam demonstrates corneal clouding, optic atrophy, macular
cherry-red spot, and retinal edema (146).
Type II can be subdivided into late infantile and juvenile forms. The late
infantile form begins between the first and third year of life. The juvenile
form presents between the third and tenth year of life with an insidious
plateauing of cognitive and motor development, which is followed by a slow
regression. Developmental regression leads to early death (146).
Type III is the chronic adult form with onset between 3 and 30 years of
age. It is characterized by skeletal involvement, extrapyramidal signs, gait
and speech disturbances, and features of parkinsonism. Intellectual
impairment occurs late in the disease (146).
Diagnostic Studies
Enzyme assay for β-galactosidase shows markedly reduced or absent enzyme
activity in leukocytes or fibroblasts (147).
GM2 GANGLIOSIDOSIS (TAY-SACHS
DISEASE)
Prevalence
Tay-Sachs disease has a carrier frequency of 1 in 25 among the Ashkenazi
Jewish population. The incidence in unscreened Jewish populations is 1 in
3,900 births. It is rare in most other populations (148).
Genetics
Tay-Sachs disease, an autosomal recessive condition, is caused by mutations
in the α subunit of the hexosaminidase A gene (HEXA). Severe HEXA mutant
alleles have been demonstrated in the Ashkenazi Jewish population, and 96%
of Jewish Tay-Sachs disease carriers have one of three common mutations
(148).
Pathophysiology
Deficiency of β-hexosaminidase α-subunits, which normally participate in the
hydrolysis of ganglioside GM2, results in the accumulation of GM2
ganglioside in the lysosomes of the central nervous system (149).

In the classic infantile form, patients develop some degree of motor weakness
by 4 to 5 months of age. Features of this disease include progressive
psychomotor retardation, megalencephaly, blindness, and premature death by
3 to 5 years of age. Fundus examination shows a cherry-red spot in the
macula (149) (see Figure 24-3).
FIGURE 24-3 Color fundus image of a left eye from a
patient with Tay-Sach disease (GM2 gangliosidosis or
hexosaminidase A deficiency) showing a cherry-red spot.
The accumulation of gangliosides or sphingolipids within
the retinal ganglion cells produce a white appearance
surrounding the fovea, which lacks ganglion cells and can
transmit a natural red appearance from the underlying
choroid. As the ganglion cells die, the cherry-red spot may
disappear, and thus it may not be present in older affected
children or adults.
Diagnostic Studies
The diagnosis is confirmed by assay of β-hexosaminidase α-subunits in
serum, leukocytes, or other tissues. Genetic analysis can confirm the
diagnosis (150).
Management
Although no treatment is currently available, improvements in supportive
care have been shown to increase survival from 1.5 years to 5 to 6 years of
age (151).
Carrier screening for Tay-Sachs disease has been effective in reducing
the incidence of this disease in the at-risk Ashkenazi population (152).
GM2 GANGLIOSIDOSIS (SANDHOFF

DISEASE)
Genetics
Sandhoff disease is an autosomal recessive disease caused by mutation in the
β subunit of hexosaminidase (HEXB) (153).
Pathophysiology
The β subunit of both β-hexosaminidase α (Hex A) and β-hexosaminidase β
(Hex B) is defective, leading to the accumulation of GM2 ganglioside and
oligosaccharides carrying terminal N-acetylhexosamine residues, resulting in
neurologic and systemic manifestations (152). Gene expression profiling has
suggested a role of inflammation as a factor in the neurodegenerative process
(154).

The clinical course is indistinguishable from that of Tay-Sachs disease,
except that organomegaly may occur in Sandhoff disease. Affected
individuals have a cherry-red spot in the macula (153).
Diagnostic Studies
Enzyme assay shows deficiency of both Hex A and Hex B (152). Diagnosis
can be confirmed by genetic mutation analysis (153).
FARBER DISEASE
Genetics
Farber disease is an extremely rare autosomal recessive disorder associated
with mutations in the ASAH1 gene, which encodes acid ceramidase (AC),
also called N-acylsphingosine amidohydrolase (ASAH). Approximately 160
cases have been reported to date in the worldwide literature (155).
Pathophysiology
AC is responsible for the degradation of the glycolipid ceramide, which plays
a central role in sphingomyelin metabolism. AC deficiency leads to
lysosomal accumulation of ceramide (155).

Farber disease classically presents in early infancy with painful swollen
joints, subcutaneous nodules, and progressive hoarseness; however, the
phenotype is variable and may include hepatosplenomegaly (156), rapid
neurologic deterioration (157), pulmonary infiltrates (158), conjunctival
granulomas, and macular cherry-red spot (159).
Diagnostic Studies
The diagnosis of Farber disease is made based upon clinical presentation and
confirmed with an assay of ceramidase activity in leukocytes, plasma, skin
fibroblasts, and other tissues. Histopathology of granulomatous tissue shows
proliferation of connective tissue with hyalinization, cholesterol crystal-like
changes, lipid-laden macrophages, and inclusions known as “Farber bodies.”
(155).
FABRY DISEASE
Genetics
Fabry disease is an X-linked condition primarily affecting males caused by
mutations in the GLA gene, which encodes α-galactosidase A (GLA) (160).
Heterozygous females also manifest multisystem disease and thus should not
be considered carriers (161).
Pathophysiology
GLA deficiency results in the accumulation of the lysosomal
glycosphingolipid globotriaosylceramide (GL-3) in nerves and other organs
(160).

Fabry disease is typically characterized by extremity pain, angiokeratomas,
and hypohidrosis. A range of phenotypic features including vasculopathy,
ischemic stroke, peripheral neuropathy, cardiac dysfunction, and chronic
kidney disease may also occur, with some patients having later onset with
milder features (160). Ocular findings include whorl-like corneal deposits
known as cornea verticillata, lens opacities, conjunctival vessel aneurysmal
dilation, and retinal vascular tortuosity. These changes typically do not
impair vision (162,163).
Diagnostic Studies
The diagnosis is confirmed by enzyme assay demonstrating deficiency of
GLA in leukocytes, plasma, serum, fibroblasts, or other tissues (160).
Management
The clinical management of Fabry disease requires a comprehensive
multidisciplinary approach to address the various organs involved (164). ERT
with intravenous α-galactosidase A has been shown to reduce tissue GL-3
levels (165) and ameliorate cardiomyopathy (166). Oral therapy with
migalastat, a small-molecule chaperone that stabilizes some mutant forms of
the GLA enzyme, can maintain renal function in amenable cases (167).
MUCOPOLYSACCHARIDOSIS TYPE I
(HURLER SYNDROME, HURLER-
SCHEIE SYNDROME, AND SCHEIE
SYNDROME)
Prevalence
Hurler syndrome occurs in 1 in 100,000 births. Male and female children are
equally affected (168).
Genetics
Mucopolysaccharidosis type I is an autosomal recessive disorder caused by
mutations in the gene encoding α-L-iduronidase (IDUA) (168).
Pathophysiology
The α-L-iduronidase enzyme hydrolyzes the glycosaminoglycans heparan
sulfate and dermatan sulfate (168). Deficiency of IDUA leads to the
accumulation of these mucopolysaccharides.

Depending on the severity of the genetic mutation, a range of clinical
phenotypes exists with Hurler syndrome being the most severe form, and
Scheie syndrome the mildest. Hurler-Scheie syndrome is an intermediate
phenotype. See Chapter 25 for further discussion and photographs of typical
facies.
Hurler syndrome is characterized by severe multiorgan involvement due
to accumulation of mucopolysaccharides. During infancy, affected
individuals may develop hepatosplenomegaly, cardiac disease, macroglossia,
coarse facies, and dysostosis multiplex. Developmental delay is evident by 1
year of age. Neurosensory and conductive hearing loss is common. Ocular
manifestations include corneal clouding leading to severe visual impairment.
Retinal degeneration and glaucoma may also occur. If this syndrome is left
untreated, death secondary to heart failure typically occurs in adolescence
(169).
Hurler-Scheie syndrome is typically diagnosed at 2 to 6 years of age.
Facial features are less coarse than in Hurler syndrome. Other characteristics
include skeletal abnormalities leading to weakness or paralysis,
hepatosplenomegaly, respiratory compromise, and mild cognitive
impairment. Ocular findings are similar to Hurler syndrome. Life expectancy
extends into the late teens or early 20s (168,169).
Scheie syndrome has physical symptoms similar to Hurler syndrome and
Hurler-Scheie syndrome. However, patients have normal intelligence. Most
patients die before the age of 30 years (168,169).
Diagnostic Studies
Diagnosis is suspected based upon clinical exam and measurement of urinary
glycosaminoglycan levels. Diagnosis is confirmed with enzyme activity
assays of fibroblasts, leukocytes, plasma, or serum (168).
Management
Hematopoietic stem cell transplant, performed before 2 years of age, provides
an opportunity to improve the neurocognitive outcome, eliminate
hepatosplenomegaly, and reduce airway obstruction (170). ERT with
recombinant α-L-iduronidase (laronidase) has been shown to improve
pulmonary function but is not effective for the orthopedic manifestations
(171).
MUCOPOLYSACCHARIDOSIS TYPE II
(HUNTER SYNDROME)
Genetics
Hunter syndrome is X-linked and primarily affects males, though some
female patients have been described (172). It is associated with mutations in
the gene encoding iduronate-2-sulfatase (IDS).
Pathophysiology
Deficiency of IDS, an enzyme responsible for the metabolism of heparan
sulfate and dermatan sulfate, leads to the accumulation of these
glycosaminoglycans in multiple organ systems (173).

Hunter syndrome has a phenotypic range with mild and severe forms. Early
manifestations include recurrent respiratory infections, coarse facial features,
joint stiffness, otitis media, umbilical/inguinal hernias, cardiomyopathy, and
hepatosplenomegaly. The characteristic short stature becomes evident around
the age of 8 years. Ocular features include nyctalopia and retinal
degeneration. Unlike Hurler syndrome, there is no corneal clouding.
Approximately two-thirds of individuals develop neurologic symptoms,
including carpal tunnel syndrome, communicating hydrocephalus, spinal cord
compression, and hearing loss. Death occurs in the second decade of life in
patients with cognitive impairment. Those without cognitive impairment may
survive into the 60s (173).
Diagnostic Studies
Screening is performed by quantitative assessment of urinary
glycosaminoglycan excretion. Diagnosis is confirmed by enzyme activity
assay in leukocytes, fibroblasts, or plasma. Genetic testing is available for
genetic counseling and carrier detection (172).
Management
ERT with idursulfase, a recombinant form for IDS, is effective at improving
somatic signs and symptoms of the disease. However, it does not cross the
blood–brain barrier, and consequently treatment of the neurologic aspect of
the disease remains a challenge (174).
MUCOPOLYSACCHARIDOSIS TYPE III

(SANFILIPPO SYNDROME)
Genetics
Sanfilippo syndrome is inherited in an autosomal recessive pattern. Four
genes have been associated with Sanfilippo syndrome, each encoding a
different enzyme involved in the degradation of heparan sulfate: Type A,
heparan N-sulfatase (SGSH); Type B, α-N-acetylglucosaminidase (NAGLU);
Type C, heparan acetyl CoA:α-glucosaminide acetyltransferase (HGSNAT);
and Type D, N-acetylglucosamine 6-sulfatase (GNS) (175).
Pathophysiology
Deficiency in one of the four enzymes involved in the catabolism of heparan
sulfate leads to its accumulation, particularly in the central nervous system
(175).

Developmental delay presents between 1 and 4 years of age, followed by
aggressive behavior and mental deterioration. In later stages, the behavioral
features are replaced by motor dysfunction, such as swallowing difficulties
and spasticity. Life expectancy extends to the second or third decade (176).
Retinal degeneration with electroretinographic findings of rod and cone
photoreceptor dysfunction is a prominent feature of the disorder (177–179).
Diagnostic Studies
Diagnostic screening can be performed with urinary glycosaminoglycan
analysis. The diagnosis and differentiation between the four subtypes can be
confirmed with enzyme activity assay of the different enzymes from
leukocytes or fibroblasts or by mutational analysis for SGSH, NAGLU,
HGSNAT, and GNS genes (175).
MUCOPOLYSACCHARIDOSIS TYPE IV
(MORQUIO SYNDROME)
Genetics
Mucopolysaccharidosis IV comes in two forms, both autosomal recessive.
Type IVA, also known as Morquio A syndrome, results from mutations in the
gene encoding galactosamine-6-sulfate sulfatase, GALNS (180). Type IVB, or
Morquio B syndrome, results from mutations in the gene encoding β-
galactosidase, GLB1 (181).
Pathophysiology
In mucopolysaccharidosis IVA, deficiency of GALNS leads to the
accumulation of keratin sulfate and chondroitin-6-sulfate, whereas in
mucopolysaccharidosis IVB, deficiency of β-galactosidase causes
accumulation of only keratin sulfate (180,182).

Morquio A syndrome is associated with skeletal and joint abnormalities,
upper airway obstruction, cardiac valvular disease, spinal cord compression,
odontoid hypoplasia, and corneal clouding (183) (see Figure 24-4).
FIGURE 24-4 Corneal clouding in a 7-year-old child with
Morquio syndrome. Fundus examination was normal.
Corneas became clearer after enzyme replacement therapy
was started.
Morquio B syndrome is characterized by skeletal changes, impaired cardiac

function, and corneal clouding (181).
In children with abnormal cervical vertebrae, care should be taken when
instilling eyedrops. Lying the child on an exam table or reclined chair and
keeping the head in line with the body while placing the drops can help avoid
potentially dangerous manipulations of the neck.
Diagnostic Studies
Urine and blood analysis for keratan sulfate is suggestive of the diagnosis in
Morquio A syndrome, but enzyme activity assay is necessary to confirm the
diagnosis (183).
Urinary excretion of keratan sulfate and oligosaccharides is abnormal in
Morquio B syndrome and suggestive of the diagnosis (182).
Management
Bone marrow transplant has been performed in some patients, and ERT has
shown promising results in Morquio A (183,184).
MUCOPOLYSACCHARIDOSIS TYPE VI
(MAROTEAUX-LAMY SYNDROME)
Genetics
Maroteaux-Lamy syndrome is an autosomal recessive disorder caused by
mutations in the gene encoding N-acetyl-galactosamine 4-sulfatase
(arylsulfatase B), ARSB (185).
Pathophysiology
Deficiency of arylsulfatase B leads to the accumulation of dermatan sulfate
and chondroitin sulfate in multiple tissues (185).

Disease manifestations, age of onset, and progression vary widely among
Maroteaux-Lamy syndrome patients. Classic findings include short stature,
skeletal abnormalities including dysostosis multiplex, joint stiffness and
contractures, respiratory complications, cardiac disease, and hearing loss
(185,186). Ocular findings include corneal clouding, glaucoma, retinopathy,
and optic disc swelling or atrophy (185). Characteristic coarse facies and
macroglossia may be noted (Figure 24-5).
FIGURE 24-5 A child with Maroteaux-Lamy syndrome
(A) lateral and (B) front face view, with typical facies.
Diagnostic Studies
Diagnosis of Maroteaux-Lamy syndrome is made based upon the presence of
clinical features, increased glycosaminoglycan levels in urine or low enzyme
arylsulfatase B activity in dried blood spots, and measurement of enzyme
activity levels in leukocytes or fibroblasts (184).
Management
ERT with galsulfase has demonstrated efficacy, particularly when initiated
early (187).
MUCOPOLYSACCHARIDOSIS VII (SLY

SYNDROME)
Genetics
Sly syndrome is an autosomal recessive condition caused by mutations in the
β-glucuronidase (GUSB) gene (188).
Pathophysiology
The enzyme β-glucuronidase is deficient in Sly syndrome, resulting in the
accumulation of glucuronic acid (189).

There is considerable phenotypic variability in Sly syndrome, including
hydrops fetalis in severe cases, short stature, skeletal dysplasia,
hepatosplenomegaly, hernias, cardiac involvement, pulmonary insufficiency,
and cognitive impairment (190). Ophthalmic features include corneal
clouding (191).
Diagnostic Studies
Diagnosis is made by enzyme activity assay demonstrating a deficiency of β-
glucuronidase in serum, leukocytes, or cultured fibroblasts. Molecular genetic
analysis can confirm the diagnosis (192).
CYSTINOSIS
Prevalence
The incidence of cystinosis is variable worldwide and occurs in
approximately 1 in 100,000 to 200,000 live births in North America (193).
Genetics
Cystinosis is an autosomal recessive disorder caused by mutation in the
CTNS gene (194).
Pathophysiology
Cystinosin, the protein encoded by CTNS, is a transmembrane protein
responsible for the transport of cystine from the lysosome into the cytoplasm.
Defective cystinosin function leads to the accumulation of cystine in cells and
tissues (193).

The foremost characteristic of classic infantile cystinosis is renal
involvement, presenting with Fanconi syndrome (polyuria, polydipsia,
electrolyte disturbances, and dehydration) and ultimately renal failure.
Growth deficiency, hypothyroidism, insulin-dependent diabetes mellitus, and
delayed puberty also occur (193,195). Ocular features include the
accumulation of cystine crystals in the cornea, conjunctiva, uvea, and sclera,
with eventual corneal clouding, pain, and photophobia. Peripheral pigmentary
retinopathy and visual impairment develop with increasing age (196).
Diagnostic Studies
Elevated leukocyte cystine levels within are diagnostic of cystinosis. Prenatal
diagnostic testing can be performed on cultured amniocytes or samples of
chorionic villi. Neonatal diagnosis can be made based upon the measurement
of placental cystine and leukocyte cystine (193).
Management
Cystine-depleting therapy with oral cysteamine (β-mercaptoethylamine) has
been shown to improve renal function and other clinical parameters of the
disease, particularly when instituted early. Diligent cysteamine therapy can
prevent hypothyroidism, enhance growth, deplete cystine in the muscle
parenchyma, and prevent multiple other systemic complications. Oral
cysteamine therapy can also reduce or delay retinopathy. Cysteamine eye
drops can dissolve corneal crystals and relieve photophobia. End-stage renal
failure is treated with hemodialysis and renal transplantation (197–200).
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25
Retinal Manifestations of Metabolic
Disease in Children
Arif O. Khan
The retina is an extracranial extension of the central nervous system with

high metabolic demand. Thus, it is not surprising that metabolic disorders
often have retinal manifestations, particularly pediatric neurometabolic
diseases. This chapter highlights selected metabolic disorders with prominent
retinal findings in childhood. For many, the retinal findings can be useful in
narrowing the differential diagnosis for children who are already under active
medical care. For some, the patient’s first presentation may be to the
ophthalmologist, who could be in a unique position to facilitate earlier
diagnosis and thus optimal care.
PrevalenceIndividually, some of these diseases are very rare, limiting
accuracy of prevalence estimates, which range from 1/8,500 for some
disorders to 1/100,000 for others. However, as a group, they are numerous
with some specific conditions more common in particular endogamous ethnic
groups.
Many pediatric metabolic disorders with retinal manifestations are
monogenic diseases associated with classic clinical features that were defined
from severe cases and before the widespread availably of genetic testing.
However, it is now clear that for virtually all of these diseases, age of onset,
degree of systemic involvement, and clinical severity can be highly variable.
The phenotypic heterogeneity is related to the degree of protein dysfunction
from the particular gene mutation as well as to other genetic and
environmental factors.
Disorder of Ornithine Metabolism (Gyrate Atrophy)
Gyrate atrophy is an autosomal recessive retinal dystrophy caused by
mutations in the gene for ornithine aminotransferase (OAT) (1,2). The rarity
of gyrate atrophy limits prevalence data; many reported cases are from
Finland (3). The lack of OAT, a vitamin B6-dependent mitochondrial
enzyme, disrupts the conversion of ornithine to pyrroline-5-carboxylate and
results in hyperornithinemia, which is the presumed mechanism of disease.
Gyrate atrophy of the retina is exceptional as an inborn error of
metabolism in that the systemic manifestations are primarily in the retina in
childhood. In adults, neuromuscular and hair findings occur (4–6) as well as
central nervous system changes (5). The diagnosis can be confirmed by
metabolic or genetic testing. A low-arginine diet and protein restriction can
slow disease progression (7,8). Dietary supplementation with pyridoxine
(vitamin B6), a cofactor for OAT, may provide additional benefit (9).
Affected children typically present with nyctalopia and loss of peripheral
vision as teenagers. Some present earlier, typically because of progressive
myopia. Early cataracts can occur. Retinal examination reveals characteristic
peripheral circular areas of chorioretinal atrophy and scalloped retinal
pigment epithelium (1,3). The macula is typically preserved in childhood, but
over decades, macular changes, optic atrophy, and arteriolar attenuation occur
as the degeneration is progressive. Some patients develop cystoid macular
degeneration early or later in the course of disease. Dietary modification may
resolve the cystoid macular degeneration that occurs in some patients (10).
Older patients show nonspecific end-stage chorioretinal atrophy, but
sometimes, the outline of former peripheral scalloped edges can be
appreciated (Figure 25-1).
Figure 25-1 (Gyrate atrophy): A male in his late 20s
presented with long-standing poor vision since childhood,
but he could not provide details. He had had cataract
surgery both eyes in his early 20s with no improvement in
vision. Vision was 20/200 either eye. Examination
revealed severe chorioretinal atrophy and cystoid macular
degeneration in the central macula in both eyes. The
outlines of old scalloped lesions can be appreciated in the
midperiphery of both eyes (A, B). The macula looks
abnormally dark in both eyes; however, this is because it is
an island of viable retina and is actually the only relatively
normal retina in each eye. In the left eye (B), the preserved
macular pigment is irregular in shape due to the scalloped
configuration of encroaching degeneration typical of
gyrate atrophy. Electroretinography was nonrecordable.
Blood ornithine levels were elevated and genetic testing
confirmed homozygous OAT mutation. The right (A) and
left (B) eyes were similar as is typical in genetic eye
disease.
Disorders of Cobalamin (Vitamin B12) Metabolism

Cobalamin (Cbl; vitamin B12) is a cobalt-containing water-soluble vitamin
that humans must ingest to obtain. Once ingested, Cbl needs to be absorbed,
to be transported, and to undergo multiple intracellular conversions. As
methylcobalamin, it is a cofactor of the cytoplasmic enzyme methionine
synthase, which converts homocysteine to methionine (11). As
adenosylcobalamin, it is a cofactor of the mitochondrial enzyme
methylmalonyl-coenzyme A mutase, which converts L-methylmalonyl-CoA
to succinyl-CoA, an intermediate of the Krebs cycle (11). Among the various
disorders that disrupt different Cbl pathways, Cbl-C–type methylmalonic
aciduria with homocystinuria is the most common and includes retinopathy.
According to newborn screening data from the state of New York from 2005
to 2008, the incidence was 1 in 100,000 (12). The disorder is caused by
biallelic mutations in the gene methylmalonic aciduria and homocystinuria
type C (MMACHC) and results in impaired intracellular synthesis of both
adenosylcobalamin and methylcobalamin (11). Increased plasma total
homocysteine, low to normal plasma methionine, homocystinuria, and
methylmalonic aciduria are the biochemical hallmarks of this disease.
Interestingly, although classic homocystinuria is associated with lens
subluxation, Cbl-C–type methylmalonic aciduria with homocystinuria is not
(13).
The phenotypic spectrum of Cbl-C–type methylmalonic aciduria with
homocystinuria is wide, varying from severe prenatal to mild adulthood-onset
disease (12). The infantile form is the most frequently recognized form (14).
Neonates show nonspecific symptoms of intrauterine growth retardation,
failure to thrive, hypotonia, lethargy, poor feeding, and vomiting.
Hematologic abnormalities include megaloblastic anemia and macrocytosis.
Neurologic findings include seizures, cerebral atrophy, white matter
abnormalities, and enlarged ventricles; in some cases, subacute degeneration
of the spinal cord is a feature. A variety of congenital malformations have
been reported in some patients, including mild facial dysmorphology,
structural heart defects, microcephaly, and hydrocephalus. Metabolic
decompensation, metabolic acidosis, and multiple organ failure may be
precipitated by stress or high protein intake. “Late-onset” disease is the term
used for patients who have overt symptoms after 4 years of age. Such patients
can present in any decade of life with neurologic regression, neuropsychiatric
symptoms, progressive encephalopathy, subacute combined degeneration of
the spinal cord, hematologic manifestations, and/or thromboembolic
complications (12,14). The diagnosis can be confirmed by metabolic or
genetic testing. Treatment for Cbl-C–type methylmalonic aciduria with
homocystinuria includes dietary protein restriction with appropriate dietary
supplementation (12,13).
Maculopathy has been documented as early as 3.5 months of age, tends to
progress rapidly, and sometimes evolves into a pseudocoloboma appearance
(15,16). Some cases with macular pseudocoloboma may represent a
developmental defect rather than a degenerative process (13). Pigmentary
retinopathy and generalized retinal dysfunction with optic nerve head pallor
are often seen with time. In the late-onset form of the disease, retinal
degeneration is less frequent. Metabolic control does not seem to impact the
ocular phenotype, which is more correlated with age of onset of the disease
and genotype (12,13). The degree of retinal dysfunction does not seem to be
strictly correlated to the degree of metabolic control (13). Retinal findings are
most common in the infantile form and can be helpful in making the
diagnosis for an infant with failure to thrive (Figure 25-2A and B).
Figure 25-2 (Disorders of cobalamin) (A, B): Cobalamin-
C–type methylmalonic aciduria with homocystinuria. An
infant with hydrocephalus, hypotonia, and developmental
delay was noted to have poor vision. At 1 year old, the
appearance of the retina was normal. At 2 years old,
abnormal dark pigment in the central macula could be
appreciated in both right (A) and left (B) eyes.
Biochemical and genetic testing confirmed cobalamin-C–
type methylmalonic aciduria with homocystinuria and
homozygous MMACHC mutation. C, D: Cobalamin-G–
type homocystinuria. An infant presented with congenital
nystagmus and failure to thrive. Genetic testing revealed
biallelic MTR mutations. At age 12 years old, diffuse
white dots were seen on fundus exam and optical
coherence tomography showed abnormal lamination and
deposits. The right eye is shown in C and D. (Photographs
C, D: Courtesy of Arlene V. Drack, MD, University of
Iowa.)
Cobalamin G deficiency (Cbl-G) is even rarer and also causes a retinal

dystrophy (17). Infants may present with nystagmus and in the first decades
of life develop a maculopathy consisting of whitish dots in the posterior pole
with corresponding foveal abnormalities on OCT (Figures 25-2C and D).
Cbl-G–type homocystinuria is caused by biallelic mutations in the gene for
methionine synthase (MTR). Megaloblastic anemia is typical, methylmalonic
aciduria is not present, and ectopia lentis is not a feature. In all cobalamin
deficiency disorders, developmental delay, thrombotic episodes, and vision
complications are risks necessitating careful follow-up with metabolic
specialists. Although medical and dietary control is important, these measures
do not completely prevent systemic complications (13,18).
DISORDER OF LIPOPROTEIN
METABOLISM
(ABETALIPOPROTEINEMIA)
Abetalipoproteinemia (Bassen-Kornzweig syndrome) is a malabsorption
syndrome caused by lack of plasma apolipoprotein B-containing lipoproteins,
that is, chylomicrons, very low–density lipoprotein, and low-density
lipoprotein (19). These are carriers of fat and fat-soluble vitamins (A, E, K),
and, therefore, affected children develop manifestations of impaired fat and
fat-soluble vitamin absorption. Biallelic mutations in the 97 kDa subunit of
microsomal triglyceride transfer protein (MTP) cause the disease (19).
Abetalipoproteinemia is rare in the general population and estimated to occur
in one in a million.
The earliest signs of deficient fat and fat-soluble vitamin absorption
include diarrhea, steatorrhea, and failure to thrive. Neurologic impairment,
particularly spinocerebellar degeneration and axonal neuropathy, occur
during early or later childhood along with muscle weakness (20). Vitamin K
deficiency leads to a bleeding diathesis. Laboratory testing reveals low levels
of cholesterol, plasma chylomicrons, very low–density lipoproteins, and low-
density lipoproteins; in addition, acanthocytosis is characteristic and may
result in anemia (20). The diagnosis can be confirmed in a patient with
suspected signs by laboratory or genetic testing. Because
abetalipoproteinemia is most amenable to treatment by vitamin
supplementation at an early stage, it is important to diagnosis as early as
possible. A peripheral blood smear for acanthocytosis is a simple readably
accessible test that can be helpful in quickly confirming a suspected
diagnosis. Affected children for whom supplementation is initiated prior to 2
years of age tend not to develop the neurologic complications usually
associated with untreated abetalipoproteinemia (21). Reduced intake of
dietary fats relieves gastrointestinal symptoms.
Retinal degeneration is part of the phenotype and often begins during
early or later childhood. Children present with central visual loss and an
atypical retinopathy although, for some, nyctalopia is the initial visual
complaint. Yellowish dots and mottling are often seen in the posterior pole
and midperiphery. A gliosis can be appreciated over the central macula (20).
With time, the entire retina is eventually affected by pigmentary retinopathy.
Angioid streaks have been reported (22), as has helicoid peripapillary
degeneration (23). Early vitamin supplementation attenuates retinal
degeneration but retinal changes can still progress (21,24). The diagnosis
should be suspected in a child with retinal dystrophy in the setting of
neurologic impairment and muscle weakness (Figure 25-3).
Figure 25-3 (Abetalipoproteinemia): A 16-year-old boy

with retinal dystrophy in the setting of developmental
delay, motor weakness, and intolerance to fatty food was
diagnosed with abetalipoproteinemia and confirmed to
harbor the homozygous MTP mutation. In both the right
and left eyes, there is a dystrophic appearance to the retina
with choroidal show and macular atrophy (Figure top, left
and right). Optical coherence tomography of the right eye
shows outer retinal thinning, particularly in the central
macula (Figure bottom, left). The left eye was similar
(not shown). A peripheral blood smear showed spiculated
red cells with surface projections of varying size
(acanthocytosis) (Figure bottom, right). (Khan AO,
Basamh O, Alkatan HM. Ophthalmic diagnosis and optical
coherence tomography of abetalipoproteinemia, a treatable
form of pediatric retinal dystrophy. J AAPOS 2019. pii:
S1091-8531(19)30019-9. doi:
10.1016/j.jaapos.2019.01.005.)
DISORDERS OF LIPID METABOLISM

Sjögren-Larsson Syndrome
Sjögren-Larsson syndrome is a deficiency of fatty aldehyde dehydrogenase
caused by biallelic mutations in the gene aldehyde dehydrogenase family 3
subfamily A member 2 (ALDH3A2) (25). It is very rare, with most reported
cases from Sweden. The accumulation of aldehydes has structural
consequences for cell membrane integrity, particularly in the skin, brain, and
retina (26). Most patients have erythema at birth with rapid progression to
ichthyosis and concurrent pruritus. Soon thereafter, spastic diplegia and
severe learning difficulties are characteristic. Diagnosis can be confirmed by
metabolic or genetic testing (25). Treatment is supportive.
Photophobia, myopia, astigmatism, and subnormal vision are common
(27). Glistening yellow-white retinal dots can be appreciated in the macula of
affected children as early as 1 year of age, and the number of dots seems to
increase with age (27). The extent of the macular abnormality does not
correlate with the severity of the ichthyosis or with the severity of the
neurologic abnormalities. Ocular coherence tomography can show a cystoid
foveal degeneration and reveals the crystals to be in the inner retina (28).
Macular pigment levels are abnormally low (29). The diagnosis should be
suspected in a child with pruritic ichthyosis, emerging neurologic features,
and crystalline maculopathy.
LCHAD Deficiency
Lack of the mitochondrial enzyme long-chain 3-hydroxyacyl-CoA
dehydrogenase (LCHAD) causes LCHAD deficiency, a disorder of fatty acid
breakdown. LCHAD is part of the mitochondrial trifunctional protein, an
enzyme complex consisting of four α and four β subunits. The α subunit
contains LCHAD; biallelic mutations in the gene hydroxyacyl-CoA
dehydrogenase/3-ketoacyl-CoA thiolase/enoyl-CoA hydratase α subunit
(HADHA) cause LCHAD deficiency (30). LCHAD deficiency is more
common in Northern Europe (1/100,000 in Sweden), and many European
patients have the same homozygous HADHA mutation (30,31). An affected
child does not produce sufficient ketone bodies during fasting, leading to a
severe, life-threatening hypoketotic hypoglycemia and potential sudden
infantile death syndrome. Other infantile features include hypertrophic
cardiomyopathy, hepatomegaly, polyneuropathy, and intermittent
rhabdomyolysis. The diagnosis can be confirmed by metabolic or genetic
testing. Newborn screening for LCHAD deficiency and other trifunctional
protein deficiency is available via acylcarnitine profiling of dried blood spots.
Early recognition and dietary modification through avoidance of long-chain
triglycerides increases life expectancy.
LCHAD deficiency is unique among disorders of fatty acid metabolism
in that it commonly affects the retina. Macular pigmentary changes can be
seen within the first 2 years of life (32) and may be accompanied by
photophobia. In those patients who survive to older ages, macular
chorioretinal atrophy occurs, pigmentary changes develop in the periphery
and eventually become atrophic as well, and electroretinography readings
progressively decline to eventually become nonrecordable (32).
Developmental supranuclear flake-like lens opacities and progressive myopia
can be seen in older patients (32). Dietary modification and supplementation
can help preserve retinal function and may improve visual acuity in some
children (33). Fasting-induced hypoketotic hypoglycemia in an infant with
central chorioretinopathy strongly suggests the diagnosis of LCHAD
deficiency (Figure 25-4).
Figure 25-4 A 5-year-old girl who originally presented
with liver failure at 9 months old was confirmed to have
LCHAD deficiency and was referred for ophthalmic
evaluation. Dilated fundus examination revealed retinal
pigment epithelium mottling and clumping just outside the
vascular arcades and increased pigment in the central
macula in both eyes (A,B). (Courtesy Scott E. Brodie,
MD, PhD).
LYSOSOMAL STORAGE DISEASE
Lysosomes are membrane-enclosed organelles responsible for the
degradation of a variety of biomolecules. Two major classes of biomolecules
that require regular degradation are complex carbohydrates such as
mucopolysaccharides and complex lipids such as sphingolipids. Gene
mutations that impair enzymes responsible for lysosomal degradation of these
particular biomolecules are a common mechanism for lysosomal storage
disease that often affects the retina. A rarer mechanism for lysosomal storage
disease is defective small molecule transport. This is the mechanism for
cystinosis, which has retinal manifestations. For some forms of lysosomal
storage disease, the genetic basis is known, but the associated protein
function has not been fully elucidated. One such example is CLN3-related
neuronal ceroid lipofuscinosis, which typically first manifests as retinal
degeneration.
Mucopolysaccharidoses
Mucopolysaccharidoses are a group of lysosomal diseases due to deficiency
of enzymes that degrade glycosaminoglycans (i.e., mucopolysaccharides).
Although individually mucopolysaccharidosis (MPS) is rare, as a group, the
estimated prevalence is 1/7,700 in Australia (34). The resultant buildup of
glycosaminoglycans in various tissues causes a spectrum of systemic clinical
features. These can include course facies, short stature, skeletal and joint
deformities, respiratory compromise, neurologic impairment, and cardiac
disease (35). In the eye, virtually all tissues can be affected by the
glycosaminoglycan deposition (36). Retinal degeneration can occur, and
electroretinography findings may be more prominent than ophthalmoscopic
findings (37). Axial shortening with hyperopia is common, and congested
retinal vessels can be seen if axial lengths are significantly shortened from
scleral glycosaminoglycan deposition. Other common ocular manifestations
include corneal haze, optic nerve swelling and atrophy, iris cysts, ocular
hypertension, and glaucoma (34). Depending on the specific enzyme
deficiency and its severity, different forms have characteristic clinical
features and range from being obvious and fatal in infancy to being subtle
and compatible with a normal life span. All are autosomal recessive except
for Hunter syndrome, which is X-linked. Table 25-1 summarizes the major
forms. The diagnosis can be confirmed by laboratory and genetic testing, and
urinalysis is a useful screening tool (38). Treatment modalities, such as bone
marrow transplantation and enzyme replacement therapy, are available for
several subtypes (35). Thus, it is important to make the systemic diagnosis as
early as possible.
Table 25-1 Major forms of mucopolysaccharidoses
Individuals with MPS are often seen by many different specialists before (or
without) the diagnosis being made (39). It may be the astute ophthalmologist
who first suspects the diagnosis, particularly in early-onset cases (Figure 25-
5) or in milder cases. The diagnosis should be suspected when an individual
has retinal findings or corneal haze in the context of clinical features
compatible with systemic glycosaminoglycan deposition (Figure 25-6).
Figure 25-5 (Mucopolysaccharidosis): Course facial
features can be appreciated in this infant child with
undiagnosed mucopolysaccharidosis. He was referred for
potential glaucoma because of cloudy corneas. Review of
systems revealed recurrent upper respiratory injections and
multiple joint laxity. There was no glaucoma; the corneal
haze was from glycosaminoglycan deposition.
Figure 25-6 (Mucopolysaccharidosis): A 30-year-old male
was referred for dry eye. He had short stature. Review of
symptoms revealed recurrent upper respiratory infections,
bilateral carpal tunnel syndrome, and sleep apnea.
Ophthalmic examination was significant for hyperopia,
bilateral iridociliary cysts, and retinal vascular congestion.
The ocular findings in the context of the review of systems
are virtually pathognomonic for systemic
glycosaminoglycan deposition as seen in
mucopolysaccharidosis. Images of the ciliary body cysts
and retinal vascular congestion with dilated, engorged
veins from the right eye are shown in (A) and (B),
respectively.
Sphingolipidoses
Sphingolipids are glycolipids important for cellular membrane function,
particularly in the brain and nervous tissue (40). Sphingolipidoses vary in
severity and age of onset depending upon the degree of the specific enzyme
dysfunction. With a few notable exceptions (e.g., Gaucher disease and Fabry
disease), sphingolipidoses are characterized by childhood neurodegenerative
disease; when this occurs, there is often accompanying retinal degeneration.
Regional loss of retinal transparency occurs where undigested biomolecules
accumulate in the thick ganglion cell layer that surrounds the fovea. Since the
fovea lacks ganglion cells, the resultant contrast in color between the
perifovea and fovea results in a central foveal “cherry-red spot” (Figure 25-
7). Often the cherry-red spot is a sign of significant systemic disease. It is
characteristic for Tay-Sachs and Sandhoff disease and can sometimes be seen
in Niemann-Pick disease, GM1 gangliosidosis, Farber disease, and
metachromatic leukodystrophy (41). Because it can sometimes disappear
over time as the intumescent ganglion cells die and optic atrophy develops,
the absence of a cherry-red spot should not be used to rule out a diagnosis,
particularly for older children (42).
Figure 25-7 (Cherry-red spot): A 14-year-old boy with
sialidosis, a rare lysosomal storage disease, had macular
cherry-red spots (right eye shown). He also had snowflake
cataracts. Genetic testing confirmed biallelic NEU1
mutations.
Sphingolipidoses characterized by early neurodegenerative disease are

summarized in Table 25-2. Some are historically more common in Ashkenazi
Jews (e.g., Tay-Sachs and Niemann-Pick type A), but this has changed with
genetic counseling (43). Long-term sphingolipid deposition in long-term
survivors can result in atherosclerotic disease as well as conjunctival and
retinal vessel tortuosity (Figure 25-8).
Table 25-2 Sphingolipidoses with early

neurodegeneration
Figure 25-8 (Sphingolipidosis): A 34-year-old male with
Niemann-Pick type B had severe atherosclerotic
cardiovascular disease as well as a history of liver
transplantation. He had bilateral conjunctival vascular
tortuosity with aneurysm dilatations (right eye shown in A,
arrowhead), bilateral retinal arteriolar and venous
tortuosity (right eye shown in B, arrowheads), and
bilateral cherry-red spots with surrounding macular halos
(right eye shown in B, double arrowheads). (Fundus
image reprinted with permission from Rudich DS, Curcio
CA, Wasserstein M, Brodie SE. Inner macular
hyperreflectivity demonstrated by optical coherence
tomography in Niemann-Pick disease. JAMA Ophthalmol
2013;131(9):1244–1246; Courtesy Danielle Rudich MD
and Scott E. Brodie MD, PhD.)
Fabry disease, one of the more common storage diseases (1/117,000 in

Australia (34)), is exceptional among the sphingolipidoses. It is X-lined,
rather than autosomal recessive, and slowly progressive into adulthood, rather
than being typically pediatric onset (44). Unlike most sphingolipidoses,
enzyme replacement therapy is available rather than only supportive
treatment. Fabry disease is due to hemizygous mutation in galactosidase
alpha (GLA), and lack of the enzyme results in glycolipid deposition (44).
The age of onset and severity of the phenotype is related to the degree of
residual enzyme function. Glycolipid deposition leads to microvascular
pathology and cellular dysfunction associated with a spectrum of systemic
features, most characteristically skin lesions (angiokeratomas), painful
neuropathy, decreased sweat, early stroke, cardiomyopathy, and renal
dysfunction (45). The ophthalmic manifestations of Fabry disease often do
not significantly impact vision but can be very useful in making the diagnosis
for a patient with systemic features that suggest the disease. Retinal and
conjunctival vessel tortuosity can be seen and may be associated with a more
rapidly progressive form of the disease (46). Cornea verticillata is the most
frequently reported eye abnormality. Anterior capsular or subcapsular
cataract or posterior radial subcapsular cataract can occur. The generalized
predisposition to vascular occlusion can occasionally cause posterior segment
vascular disease, such as retinal artery occlusion (47). Fabry disease can be
confirmed in a patient with clinical suspicion of the diagnosis by laboratory
or genetic testing. Early diagnosis is crucial, because enzyme replacement
treatment is available and is most effective when initiated early (44).
Gaucher disease is considered the most common lysosomal storage
disease, estimated to be 1/40,000 in the United States and even more
common in Ashkenazi Jews (48). Biallelic mutations in β-glucocerebrosidase
(GBA) lead to accumulation of glucocerebride (48). Classic clinical features
are hepatosplenomegaly, anemia, thrombocytopenia, and bone pain and
fractures. The diagnosis can be confirmed by laboratory or genetic testing.
Enzyme replacement treatment is available and, thus, early diagnosis is
important. Although ocular features are not a classic feature of Gaucher
disease, those who have juvenile or adulthood forms of the disease can
develop preretinal white dots, vitreous opacities, and retinal vascular
abnormalities (49,50). Those with the rarer central nervous systemic form of
the disease can develop limited ocular motility.
Mucolipidoses
Mucolipidoses are rare conditions that share combined features of
mucopolysaccharidoses and sphingolipidoses. Sialidosis from biallelic
mutations in NEU1 and galactosialidosis from biallelic mutations in CTSA
can include cherry-red spots (Figure 25-7) (51,52).
Neuronal Ceroid Lipofuscinoses

Neuronal ceroid lipofuscinoses are a group of childhood neurodegenerative
disorders characterized by progressive neuronal loss in the cerebrum,
cerebellum, and retina. Histology shows accumulation of autofluorescent
lipopigments that resemble ceroid and lipofuscin in neurons (53,54). This
group of disorders was originally classified according to onset of neurologic
symptoms and cellular localization of the storage deposits but are now
defined genetically (CLN1-CLN14). In the United States, the prevalence is
estimated to be between 1:56,000 and 1:67,000 (53). Recurrent clinical
features include mental and motor deterioration, epileptic seizures,
progressive retinal degeneration, and premature death. Although 14
associated proteins have been identified to date, not all have had their
function elucidated (53). The term Batten disease was originally used to
describe CLN3-related disease but now is also used to describe the various
subtypes collectively as a group (54,55). Subtypes are summarized in Table
25-3.
Table 25-3 Subtypes of neuronal ceroid

lipofuscinoses
The most common form of NCL is juvenile neuronal ceroid lipofuscinosis,
the classic form of Batten disease due to biallelic mutations in CLN3 (56).
Because CNL3-related disease typically presents with visual loss before
psychomotor regression and epilepsy, it is also the most relevant form to
ophthalmologists (Figure 25-9). Biallelic mutations in CLN5 or CLN11 can
sometimes initially present with visual loss, but this is rarer (57,58) (Figure
25-10). Juvenile neuronal ceroid lipofuscinosis has an estimated incidence of
1/100,000 (55) and is relatively more common in Scandinavia, where the
incidence ranges from 2 to 7/100,000 (59). Noticeable vision impairment
from retinal degeneration typically occurs at 5 to 6 years of age. For unclear
reasons, boys tend to have visual symptoms earlier than girls, but girls tend to
experience a more severe disease course (60). Once the retinal degeneration
is diagnosed, visual loss rapidly progresses. A bulls-eye maculopathy can be
seen but is not always present. Electroretinography shows depressed and
delayed waveforms and, characteristically, an electronegative waveform.
With time, electroretinography becomes nonrecordable. When CLN3-related
disease is suspected from the ophthalmic presentation, targeted questioning
may identify psychosocial changes that occurred before the visual
complaints, such as moody behavior and nightmares.
Figure 25-9 (CLN3): A 5-year-old boy from Scandinavia
was noted to have decreased vision several months prior to
when he started school. Best-corrected visual acuity was
20/60, 20/100, and bulls-eye maculopathy was noted
(A,B) with abnormal macular autofluorescence (C,D).
Electroretinography revealed a nonrecordable scotopic rod
response, a depressed combined flash response with an
electronegative waveform, and depressed and delayed
photopic responses. Review of systems revealed moody
behavior and recurrent nightmares. Sequencing of CLN3 in
the child and his parents confirmed compound
heterozygosity for mutations in the child.
Figure 25-10 (CLN5): A 5-year-old girl was noted to be
clumsy for the 2 years prior to presentation and over the
same time period was increasingly failing to follow
commands. Best-corrected visual acuity was 20/200.
Retinal examination confirmed faint bulls-eye
maculopathy with blunting of foveal reflex (A,B) and
mildly abnormal macular autofluorescence with a faint
ring of hyperfluorescence around the fovea (C,D).
Neuroimaging revealed cerebellar hypoplasia. Genetic
testing confirmed homozygous mutation in CLN5. Retinal
examination in such cases may initially appear grossly
normal, causing some children to be suspected of
malingering.
Neurologic functions decline quickly after significant visual loss in CLN3-
related disease. There is progressive dementia, ataxia, extrapyramidal signs,
seizures, and loss of independent function. Cardiac involvement results in
electrophysiologic dysfunction and ventricular hypertrophy. Premature death
usually occurs in the second or third decade of life (61).
Diagnosis can be confirmed by genetic testing. Many patients have a
recurrent 1.02 kb deletion (55). Although currently untreatable, early
diagnosis is important to institute appropriate counseling and support, and
ophthalmologists are uniquely positioned to facilitate this. The diagnosis
should be suspected in a child with rapidly progressive juvenile retinal
dystrophy, particularly when in the setting of psychosocial changes.
Certain CLN3 mutations have been associated with isolated rod–cone
dystrophy usually diagnosed after (but sometimes before) the second decade
of life (62,63). The retinal dystrophy in these isolated cases neither presents
as early nor progresses as rapidly as is seen in CLN3-related disease. In one
case, neurologic decline occurred during adulthood (64). Nonsyndromic
retinal dystrophy has also been recognized in association with certain
mutations in CLN7 (55).
Systemic gene therapy and/or enzyme therapy are in clinical trials at this
writing for many forms of Batten disease. As this is a rapidly changing
landscape, www.clinicaltrials.gov should be consulted each time patients
with any of the neuronal ceroid lipofuscinoses are being seen, and parents
should be given the Web site address to follow on their own. This Web site
lists most clinical trials being conducted worldwide; however, entries are not
vetted; therefore, encouraging patients to discuss trials with a physician is
recommended.
Cystinosis
Cystinosis is an autosomal recessive lysosomal storage disease of cystine due
to biallelic mutations in cystinosin (CTNS), which encodes a lysosomal
cystine carrier protein (65). It is rare, with the infantile form estimated to
occur in 1/325,000 births in France (66). Proximal renal tubular defect
(Fanconi-like), hypothyroidism, and crystalline corneal deposits are the main
clinical features. Diagnosis can be confirmed by genetic testing in a patient
with suspicious clinical features. The mainstay of treatment is the cystine-
depleting agent cysteamine, both orally and topically. Early recognition and
treatment optimize outcomes.
Retinal changes are part of the phenotype, although corneal haze and
photophobia can limit detailed examination of the retina. The most common
retinal finding is depigmentation of the peripheral retina with pigment
epithelial mottling, which has been observed as early as 6 months of age (67).
With time, pigment migration into the retina can cause a retinitis pigmentosa
appearance. Electroretinography of older cystinosis patients shows decreased
cone and rod function (67). Intraretinal crystals can sometimes be appreciated
on fundus examination or by optical coherence tomography (68,69). Oral
treatment seems to improve the retinal findings (67). The diagnosis should be
suspected in a child with nephropathy and crystals in the cornea or retina.
PEROXISOMAL DISORDERS
Peroxisomes are cellular organelles that participate in multiple catabolic and
synthetic biochemical pathways, the majority of which involve intermediary
lipid metabolism (70). The well-characterized pathways of very long-chain
fatty acid β-oxidation, α-oxidation of phytanic acid, and plasmalogen
biosynthesis are used routinely for the biochemical diagnosis of peroxisome
biogenesis disorders (71). Disorders that were previously considered separate
conditions—Zellweger syndrome, neonatal adrenoleukodystrophy, Refsum
disease, and Heimler syndrome—are now recognized as a continuum of
peroxisomal disease phenotypes, known as Zellweger spectrum disorders,
and retinal dystrophy is a major recurrent feature (72,73). Biallelic mutations
in 14 different peroxisomal genes (PEX genes) are responsible for the
majority of cases (73). Peroxisomal disorders often cause deafness as well as
retinal degeneration, and patients may be misdiagnosed as having Usher
syndrome. It has been estimated that peroxisomal biogenesis disorders occur
in 1/50,000 births in North American, but the prevalence is likely higher (73).
Rhizomelic chondrodysplasia punctata (PEX7 mutations) is a related
peroxisomal disorder but with an associated ocular phenotype of pediatric
cataract rather than retinal dystrophy, although there is one case report for
which retinitis pigmentosa developed (74). Primary hyperoxaluria is a
peroxisomal disorder that can include a crystalline retinopathy.
Zellweger Spectrum Disorders
Children with the severe congenital phenotype of Zellweger spectrum
disorder (cerebrohepatorenal syndrome; classic Zellweger syndrome) are
hypotonic, feed poorly, and fail to show developmental progression (73,75).
They have seizures, liver cysts with hepatic dysfunction, and often a recurrent
facies (round face, high forehead, large anterior fontanel, hypertelorism,
epicanthic folds, high-arched palate, and micrognathia). Bony stippling
(chondrodysplasia punctata) of the knees and hips can be seen by X-ray.
Those who survive develop sensorineural hearing loss and retinal dystrophy
(73,75). Cataracts and glaucoma can also occur. Less severe phenotypes in
this spectrum show a wide range of presentations and variable involvement of
the potentially affected organ syndromes. With the identification of additional
peroxisomal genes and advances in genetic testing, phenotypes that would
have been previously thought to be too mild for peroxisomal disease have
been found to be due to PEX mutations. For example, Heimler syndrome—
sensorineural hearing loss, amelogenesis imperfecta, nail abnormalities, and
juvenile or adult-onset retinal dystrophy—can be caused by biallelic
mutations in PEX7 (72). For a child with retinal dystrophy in the setting of
multiple organ disease, peroxisomal disorder is part of the differential
diagnosis (Figure 25-11).
Figure 25-11 (Peroxisomal facies): This 2-year-old boy
with hypotonia and developmental delay was seen because
of poor visual behavior. The retinal appearance was
grossly normal but electroretinography was nonrecordable.
Genetic testing revealed homozygous mutations in the
recently recognized peroxisomal gene ACBD5. His facial
features were consistent with peroxisomal disease: round
face, high forehead, apparent hypertelorism, and
prominent epicanthal folds.
Primary Hyperoxaluria
Primary hyperoxaluria type I is a disorder of progressive calcium oxalate
urolithiasis and nephrocalcinosis caused by biallelic mutations in the
peroxisomal enzyme alanine-glyoxylate aminotransferase (AGT) (76). In the
absence of AGT, glyoxylate is converted to oxalate, which forms insoluble
calcium oxalate crystals that accumulate in the kidney and other organs.
Treatment is supportive and includes dietary modification with high fluid
intake (77). Primary hyperoxaluria type I is very rare in the general
population, with an estimated prevalence of one in a million.
Oxalate-induced crystalline retinopathy seems to be correlated with the
severity of the disease. It is characterized by numerous discrete yellow flecks
scattered throughout all layers of the retina and retinal pigment epithelium
starting in the macula but extending throughout the retina. With time, there
are areas of irregular chorioretinal atrophy, retinal pigmentary changes,
confluent patches of retinal pigment epithelium hyperplasia, and patches of
fibrosis (78).
CONGENITAL DISORDERS OF
GLYCOSYLATION
Glycosylation is a process used by eucaryotic cells to enhance specialized
functions for biomolecules and cellular processes (79). Congenital disorders
of glycosylation are an increasingly recognized group of disorders secondary
to genetic defects in the synthesis and attachment of glycoprotein and
glycolipid glycans. Each is individually rare, but as a group, more than 125
such disorders have been recognized (79). The phenotypic and genetic
spectrum is wide and continues to be defined with presentations ranging from
severe neonatal lethal to mild adult onset. Neurologic impairment is the most
common unifying feature. Retinal findings been clearly documented for
several forms of the disease, as summarized in Table 25-4 (Figure 25-12).
The prevalence of congenital disorders of glycosylation in Europe has been
estimated at one in a million (80).
TABLE 25-4 Congenital disorders of glycosylation

(CDG) with retinal findings
Figure 25-12 (Congenital disorder of glycosylation
[CDG]): A 13-year-old girl was referred for evaluation of
low vision since soon after birth. Clinical examination
confirmed retinal dystrophy (right eye shown in A,B) with
ring of hyperautofluorescence around the macula and
stippling of the periphery. She also had developmental
disability and chronic dermatitis (C). Genetic testing
uncovered homozygosity for a SRDA3 deletion and thus
confirmed SRDA3-CDG. (Reprinted with permission from
Khan AO. Early-onset retinal dystrophy and chronic
dermatitis in a girl with an undiagnosed congenital
disorder of glycosylation (SRD5A3-CDG). Ophthalmic
Genet 2018:1–3.)
MITOCHONDRIAL DISORDERS
Through oxidative phosphorylation and ATP production, mitochondria are
the major energy source for the cell. The number of mitochondria within a
specific cell is related to the cell’s energy requirements. Mitochondria are
organelles with their own semiautonomous genome that is maternally
inherited and can vary in sequence among different tissues or even in the
same cell, a characteristic known as heteroplasmy (81). Mutations of the
mitochondrial genome can cause mitochondrial disease that is associated with
retinopathy. The severity of the phenotype for a patient carrying a given
mitochondrial mutation is related not only to the mutation itself but also to
the degree of mitochondrial heteroplasmy. Diagnosis in a suspected case can
be confirmed by mitochondrial DNA genome analysis of blood or tissue.
Because most proteins required for mitochondrial function are encoded in
nuclear DNA, mitochondrial disease can follow Mendelian rather than
maternal inheritance (78). One such example is LCHAD deficiency
(discussed under Disorders of Lipid Metabolism). Mitochondrial diseases are
likely more common than currently recognized with an estimated prevalence
of 1/8,500 (82). Mutations of the mitochondrial genome should be suspected
when a patient presents with atrophic macular retinal pigment epitheliopathy
in association with ocular motility disorders, deafness, or generalized
weakness.
Kearns-Sayre Syndrome
Kearns-Sayre syndrome is a form of chronic progressive external
ophthalmoplegia with pigmentary retinopathy that begins in childhood or
young adulthood, by definition before 20 years of age (83). Cardiac block is a
common feature, and ptosis often accompanies the chronic progressive
external ophthalmoplegia. Other systemic features can include short stature,
cerebellar ataxia, muscle weakness, endocrine abnormalities, and progressive
hearing loss. (80). The majority of cases are sporadic, from new
mitochondrial DNA deletions (84,85). The retinopathy shows widespread
retinal pigmentary changes and a “salt and pepper–like” appearance. Age of
onset and progression is variable, but most patients do not progress to severe
visual loss with nonrecordable electroretinography. A patient with retinal
dystrophy in the setting of chronic external ophthalmoplegia should be
investigated for the possibility of cardiac conduction defects (Figure 25-13).
Figure 25-13 A 16-year-old boy presented with a 5-year
history of slowly progressive ptosis and ophthalmoplegia
(A). Fundus examination confirmed pigmentary
retinopathy with mottled peripheral pigment and irregular
pigmentation of the macula. A darkly pigmented precursor
to a typical bone spicule–like lesion can also be seen (B,C:
arrowhead).
Mitochondrial Variants A3243G (MELAS/MIDD) and

T8993G (Leigh Syndrome/NARP)
Mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes
(MELAS) is a multisystem disorder with typical onset between 2 and 10
years of age (86). Initial signs are often exercise intolerance or proximal
muscle weakness. Short stature is common. Recurrent headaches and
episodes of vomiting, hearing loss, diabetes, and seizures are classic features.
The seizures can be associated with stroke-like episodes of transient
hemiparesis or cortical blindness and altered consciousness. Cardiac
conduction defects can occur as well. The mitochondrial A3243G mutation is
the most common pathologic variant associated with MELAS. Pigmentary
retinopathy is part of the phenotype. There is wide spectrum of age and
severity of presentation, but it often starts as macular retinal pigment
epitheliopathy (87).
The same mitochondrial A3243G mutation can be associated with the
phenotype maternally inherited diabetes and deafness (MIDD) (87). MIDD-
associated retinal dystrophy is typically an adult-onset macular dystrophy that
does not significantly affect visual acuity and shows mottled patchy
hyperpigmentation in the macula that can evolve into a characteristic
perifoveal discontinuous crescentic chorioretinal atrophy (88–90). Pattern
dystrophy (91) is another presentation, and corneal endothelial
polymegathism may be seen (92).
Another specific mitochondrial variant known to be associated with
retinal dystrophy is T8993G (93). The retinal phenotype can range from
infantile (94) to adult-onset (95) retinal dystrophy, the severity generally
correlating with the systemic phenotype. The systemic phenotype can range
from the severe lethal infantile Leigh disease (infantile necrotizing
encephalopathy) to adult-onset neuropathy–ataxia–retinitis pigmentosa
(NARP) (96).
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26
Generalized Retinal and Choroidal
Diseases
Meghan M. Brown, and Kean T. Oh
INTRODUCTION
In 1990, evidence that rhodopsin mutations caused retinitis pigmentosa (RP)
heralded a new perception of the pathophysiology and an evolving approach
to retinal dystrophies. Since then, an ever-increasing number of genes have
been identified as causative in retinal and choroidal dystrophies. With the
identification of disease-causing genes, ophthalmologists have elucidated the
pathophysiology of ocular diseases and have suggested therapies for
previously untreatable diseases. For example, mutations in RPE65 cause
Leber congenital amaurosis (LCA) or rod–cone dystrophy in adults. Gene
therapy using viral vectors to introduce working copies of the gene has been
validated by animal models and clinical trials. It is now available for RPE65-
related LCA (1,2) (see also Chapters 27 and 28).
The advent of treatments for genetic diseases will likely be specific for
certain gene products. Therefore, clinical diagnoses will be important to
identify patients with phenotypes who would benefit from specific therapies.
Furthermore, understanding and characterizing the natural history of disease
will be necessary to accurately interpret future clinical trials.
The goal of this chapter is to provide the clinical descriptions, prevalence,
genetics, differential diagnoses, and pathophysiologies of inherited retinal
and choroidal disorders that may present in infancy and early childhood.
Ancillary studies, such as electroretinography (ERG) and perimetry, and
molecular pathophysiology are reviewed for specific diseases. In the course
of writing this chapter, the Web site, Online Mendelian Inheritance in Man,
was used to review these conditions from the molecular genetic etiology
standpoint (http://www.ncbi.nlm.nih.gov/omim). Diagnostic studies, gene
therapy, and visual rehabilitation are covered in greater depth elsewhere in
the text.
GENERALIZED RETINAL AND

CHOROIDAL DYSTROPHIES
Retinitis Pigmentosa
RP describes a heterogeneous set of conditions that share progressive rod loss
initially followed by cone loss in most cases. The loss of photoreceptors
results in characteristic symptoms and fundus findings. RP constitutes the
archetypal generalized retinal dystrophy. It is the most common retinal
dystrophy and is broadly subdivided into nonsyndromic and syndromic forms
of disease. Because of their significance, some syndromic forms have their
own chapters in this textbook (see Chapters 32 and 33).
Cone–Rod Dystrophy
The definition of the group of cone–rod dystrophies is based primarily on
electrophysiologic characteristics. It now includes a number of phenotypic
expressions once believed to represent entirely separate diseases. Cone–rod
dystrophies are generalized photoreceptor disorders characterized on ERG by
a greater loss of cone function than rod function and consequently have
common symptomatic presentations (3). Like RP, cone–rod dystrophies are
heterogeneous with a wide range of presentations, disease courses, and
severity.
Clinical Presentation and Disease Course

Most cone–rod dystrophies (Table 26-1) present late in the second decade of
life to early adulthood; for example, RPGR-ORF15–associated X-linked
cone–rod dystrophy typically presents by the age of 45 years (4,5). However,
some cone–rod dystrophies present earlier in life. Autosomal dominant cone–
rod dystrophy caused by mutations in the CRX gene may present in the first
decade of life with symptoms of decreased central vision and
dyschromatopsia. The CRX gene also has been associated with both LCA and
a late onset of RP in which patients remain asymptomatic until the sixth
decade of life (6). Autosomal recessive cone–rod dystrophy can be caused by
the ABCA4 gene and present in conjunction with, or in the absence of, classic
findings associated with Stargardt disease/fundus flavimaculatus in childhood
(7–9).
TABLE 26-1 Genes associated with cone–rod

dystrophies
cGMP, cyclic guanosine monophosphate; RPGR, retinitis pigmentosa GTPase regulator; PDE,
phosphodiesterase enzyme; ATP, adenosine triphosphate.
Presenting symptoms of cone–rod dystrophy include decreased central vision,

dyschromatopsia, and light sensitivity (10). Patients typically report that they
are most comfortable in low ambient light settings such as dawn or twilight.
Early in the course, patients may not have nyctalopia, but with progression of
disease, they will develop nyctalopia and peripheral vision loss. Typically,
fundus changes are similar to RP with bone spicules and retinal pigment
epithelium (RPE) atrophy. However, fundus changes tend to occur in the
macula and posterior pole before the retinal periphery. Macular changes
include geographic atrophy, a beaten bronze appearance, a bull’s-eye
maculopathy, and a tapetal sheen (Figure 26-1).
FIGURE 26-1 Cone–rod dystrophy. Patient with an X-

linked cone–rod dystrophy. There is a subtle tapetal sheen
temporally, and the patient has a geographic area of central
macular atrophy.
The characteristic ERG demonstrates greater involvement of the cone system

than the rod system with full-field testing. One form of recessive cone–rod
dystrophy caused by mutations in the KCNV2 gene is characterized by the
typical reduction in cone function and a concomitant supranormal rod
response (11). Further attempts have been made to characterize cone–rod
dystrophies based on their dark-adapted perimetry characteristics (12).
Finally, the multifocal ERG is diffusely abnormal and may be nonrecordable
in patients with cone–rod dystrophies.
Patients with focal macular changes, such as seen in Stargardt disease,
will have greater central loss (Figure 26-2).
FIGURE 26-2 Stargardt disease. A:Note the classic
appearance of pisciform flecks in the macula and beaten
bronze appearance centrally. B:Multifocal ERG. The top
image represents trace arrays for the central 40 degrees of
the patient’s field of vision. Note the loss of amplitude
centrally. This is depicted in a graphic format in the
bottom image as a response density plot. The central
depression represents the loss of function due to Stargardt
disease. The peripheral, shallower depression represents
the blind spot.
Clinical diseases once believed to be separate from cone–rod dystrophies are

now included within the broad group because of shared electrophysiologic
characteristics. Foremost among these is Stargardt disease. Disorders caused
by the ABCA4 gene constitute a large proportion of previously identified
autosomal recessive CRD (8,9). The presence of cone dysfunction has been
noted to be mild to severe and may present at any point in the course of a
classic clinical appearance of Stargardt disease/fundus flavimaculatus.
Although derangements in ERG function are more common in late stages of
this disease, patients have presented with minimal fundus changes and severe
cone dysfunction early in the course of the disease. Lois et al. (7) suggested
that the presence of advanced ERG derangements might be an indicator of a
poor long-term outcome. Itabashi et al. (13) reported that patients with a
clinical phenotype of fundus flavimaculatus, that is, pisciform changes
involving the midperipheral and posterior retina rather than disease limited to
the macula, tended to present with worse ERG dysfunction. Further details
regarding Stargardt disease are dealt with in detail elsewhere in this textbook
(Chapter 30).
Another condition with severe cone dysfunction at presentation is fundus
albipunctatus. Although fundus albipunctatus is classically considered to be
nonprogressive, some patients will also develop cone dysfunction
accompanied by macular changes in midadulthood (14) (see Chapter 30).
Systemic disease associations with cone–rod dystrophies are rare.
Spinocerebellar ataxia has been associated with cone–rod dystrophies (15).
There are two reported cases of thiamine-responsive megaloblastic anemia
and cone–rod dystrophy (16) and a reported case of chondroplasia with cone–
rod dystrophy (17). Alström syndrome also demonstrates a cone–rod
dystrophy phenotype with cardiomyopathy and type 2 diabetes mellitus (18).
The CNNM4 gene is associated with Jalili syndrome, which includes
autosomal recessive cone–rod dystrophy, amelogenesis imperfecta, and
neurofibromatosis type 1 (19). Finally, Bardet-Biedl syndrome causes a
pigmentary retinopathy and photoreceptor degeneration that may have a
cone–rod ERG phenotype because of associated central macular atrophy (see
also Chapter 32).
Spondylometaphyseal dysplasia with cone–rod dystrophy (SMDCRD) is
characterized by skeletal abnormalities, bowed lower limbs and pigmentary
maculopathy with evidence of cone–rod dysfunction (20). Mutations in the
CEP78 gene result in cone–rod dystrophy and hearing loss (CRDHL)—a
syndrome that presents with sensorineural hearing loss and a severe
deficiency of central vision due to the higher rate of cone photoreceptor
degeneration compared to rod photoreceptor degeneration (21).
Molecular Genetics of Cone–Rod Dystrophies

Like RP, cone–rod dystrophies can be inherited in an autosomal dominant,
autosomal recessive, or X-linked recessive manner (Table 26-1). There are
reports of potential associations with mitochondrial disease and cone
dystrophies as well (22). Twenty or more genes have been identified to be
associated with cone–rod dystrophies, while multiple other loci have been
mapped.
Kohl et al. (23) determined that 50% of autosomal dominant cone–rod
dystrophy was caused by mutations in the CRX, PRPH2, retinal guanylate
cyclase (GUCY2D), guanylate cyclase activator 1A (GUCA1A), and PROM1
genes. Other genes playing a lesser role in the etiology of autosomal
dominant cone–rod dystrophy include GUCA1A (GCAP1), GUCA1B
(GCAP2), Rab3A-interacting molecule (RIMS1), AIPL1, UNC119, and
PITPNM3 (3,24–27).
Genes associated with autosomal recessive cone–rod dystrophies are
ABCA4, CNNM4, CACNA2D4, KCNV2, PDE6C, PDE6H, RPGRIP, RAXL1,
CDHR1, ADAM9, SEMA4A, and RDH5 (14,19,28–30). Most of these genes
have been identified in only isolated pedigrees. However, some groups
suggest that in up to 35% of patients with autosomal recessive cone–rod
dystrophy, the cause may be mutations in the ABCA4 gene (31). It is entirely
possible that these groups are identifying patients who would otherwise have
been characterized as having Stargardt disease or fundus flavimaculatus at
different stages in the evolution of their clinical disease rather than ABCA4-
related cone–rod dystrophy. The distinction between these diseases given
their clinical features and molecular genetics remains to be clarified (see
Chapter 30).
Regardless, electrophysiologic characteristics consistent with cone–rod
dystrophy are likely a frequent component of ABCA4-related disease and
associated with the entire range of its clinical appearances.
The genes thus far identified to cause X-linked cone–rod dystrophy
include RPGR, with the mutations occurring specifically in its ORF15 exon
(CORDX1) (5). This exon is characterized by a highly repetitive sequence,
which may lead it to be a mutation hotspot in the RPGR gene. The CACNA1F
gene (CORDX3) was identified as a cause of cone–rod dystrophies in
addition to X-linked congenital stationary night blindness (CSNB). One other
locus for cone–rod dystrophy (CORDX2) has been linked to Xq27, with a
suggested locus between markers DXS292 and DXS1113, but the gene has
not yet been identified (32).
Several genes associated with cone–rod dystrophies cause other retinal or
macular dystrophies. These include pattern dystrophies of the macula
(PRPH2), LCA (CRX, GUCY2D), RP (CRX, peripherin/RDS, ABCA4,
RPGR), macular degeneration without full-field ERG abnormalities (RPGR)
(33), CSNB (RDH5 and CACNA1F), complete achromatopsia (GNAT2), and
Stargardt disease (ABCA4). Cone–rod dystrophies, like RP, demonstrate that
mutations in the same gene can result in different clinical phenotypes
previously considered to be entirely separate disease entities.
Choroideremia
Choroideremia is an X-linked disorder resulting in choroidal degeneration
with subsequent loss of retinal function. It was first described in 1871 by
Mauthner. The gene (CHM) was identified in 1992. Late-stage choroideremia
has an unmistakable appearance of widespread loss of RPE, choriocapillaris,
and larger choroidal vessels with occasional patches of pigment and sparing
of a small patch of RPE and retina in the macula. When choroideremia
presents in childhood and adolescence, its symptoms and clinical appearance
can be very similar to those of RP.
Clinical Presentation and Disease Course

Choroideremia typically presents in males in the second to third decade of
life with the primary symptom of nyctalopia (34). On occasion, patients
report symptoms as early as the first decade of life. Early in the course of the
disease, nummular patches of RPE atrophy appear in the midperipheral
retina. These areas become confluent with progressive loss of choriocapillaris
initially and larger choroidal vessels subsequently (Figure 26-3). The end-
stage results in bare sclera extending from the posterior pole to the periphery.
In one study, Roberts et al. (35) found that 84% of patients younger than 60
years had 20/40 or better central vision, whereas 33% of patients older than
60 years had 20/200 or worse vision. Coussa et al. noted that visual acuity
remained very stable until 50 years of age after which there was a rapid
decline in central visual acuity. Thus, patients often maintain central vision
until late in the course of disease but lose peripheral fields steadily from the
time of diagnosis onward. As expected, the ERG shows progressive loss of
retinal function throughout the disease course, eventually becoming
nonrecordable in the late stages. The female carriers of mutations in the CHM
gene, in this study, had good stable vision (36).
FIGURE 26-3 Choroideremia. Left fundus image of a 28-
year-old man with choroideremia. He has severely
constricted visual fields, and his central vision is 20/80 OD
and finger-counting vision OS. Note the absence of RPE
and choriocapillaris throughout the fundus. Large-caliber
choroidal vessels are easily visible.
Female carriers of choroideremia demonstrate a spectrum of clinical

manifestations ranging from essentially no findings to symptomatic loss of
retinal function and apparent retinal degeneration (34). This spectrum likely
is dependent on the degree of mosaicism and X inactivation. In general,
carriers of mutations in the CHM gene demonstrate a moth-eaten appearance
of RPE, with RPE clumping and atrophy. Many carriers also will demonstrate
drusenoid deposition and peripapillary atrophy. However, some individuals
will have a completely normal fundus. The degree of dysfunction on ancillary
tests corresponds with the apparent clinical involvement.
Differential Diagnosis
Choroideremia can closely mimic the presentation of X-linked RP in its early
stages. Both conditions have nearly identical clinical manifestations and
symptoms and present at approximately the same age (i.e., second decade of
life). Examination of the carriers (preferably, the mother of the patient) is
helpful because the carrier state of X-linked RP can manifest with a tapetal
macular sheen or with patches of retina characteristic of RP. Gyrate atrophy
can also resemble choroideremia in its early stages; however, gyrate atrophy
has an autosomal recessive inheritance pattern, hyperornithinemia, and a
milder course than choroideremia.
Molecular Findings
The CHM gene was identified in 1993 by Sankila and further characterized
by Seabra (37–39). The CHM gene codes for a Rab escort protein (REP-1)
for the enzyme geranylgeranyl transferase, which is involved in adding a
geranylgeranyl moiety to the end of Rab proteins, in a process known as
prenylation (38–40). Rab proteins are used for intracellular signaling in
photoreceptors and for breakdown and clearance of outer segment disc
membranes in RPE cells (41). Thus, the major mechanism of retinal cell
death is thought to be a result of the buildup of unprenylated Rab proteins
(42). Nearly all disease-causing changes in the CHM gene were found to
prevent translation or result in significant truncation of the protein product.
There was one reported case of a missense mutation resulting in disease (43),
although in a review of 57 families with choroideremia, missense mutations
did not account for a significant fraction of disease-causing changes.
Transitions and transversions in the CHM gene accounted for more than 40%
of identified changes, small deletions (<5 base pairs [bp]) an additional 28%
of the mutations, partial gene deletions 9%, and complete deletions 4% (44).
At this time, the severity of the mutation does not appear to correlate with
either disease severity or carrier state manifestations.
Thus far, CHM gene replacement therapy has proven effective in CHM
mice using AAV2 and in cells of patients with CHM mutations in vitro (42).
These advances have led to two clinical trials beginning in 2015 to assess the
safety and efficacy of gene vector treatment in CHM patients (42,45). Initial
findings support improvement of rod and cone function after treatment with
retinal gene therapy (46).
Achromatopsia
Achromatopsia is a congenital inability to discriminate color. It is a rare
condition estimated to have a prevalence of 3 in 100,000. However, the
prevalence of achromatopsia on the island of Pingelap in Micronesia is 4% to
10% due to a sharp reduction in the population after a typhoon that struck in
the late 1700s. All affected individuals trace their lineage back to 1 of about
20 survivors, who was a carrier of the gene mutation (47). Huddart first
described in 1,777 a patient with achromatopsia from a family with two
affected brothers and three unaffected siblings. However, Daubeney is
credited with the first overall description of achromatopsia in 1684 (48).
Achromatopsia is divided into two groups: complete achromatopsia (rod
monochromacy) and incomplete achromatopsia (blue cone monochromacy).
Complete achromatopsia is autosomal recessive and results in loss of all cone
function, whereas incomplete achromatopsia is X-linked recessive and results
in loss of red and green opsins only. Several other cone disorders may result
in deficient color vision and overlap in phenotype with achromatopsia and
blue cone monochromacy.
The most common cone dysfunction syndromes include complete and
incomplete achromatopsia, oligocone trichromacy, bradyopsia, and Bornholm
eye disease (49) (Table 26-2).
TABLE 26-2 Genes associated with achromatopsia

and bradyopsia
XL, X-linked; AR, autosomal recessive; AD, autosomal dominant; cGMP, cyclic guanosine
monophosphate.
Clinical Characteristics of Achromatopsia

Both complete and incomplete achromatopsia are marked by poor color
discrimination from birth, photosensitivity or photoaversion, decreased
vision, and relative clinical stability. Infants may stare at lights; however,
when an object of interest is presented in room light, they squint and appear
photophobic.
Nystagmus occurs in both forms of achromatopsia. However, by the time
a patient presents for evaluation in late childhood or early adulthood,
nystagmus may be barely noticeable. Patients with either form of
achromatopsia tend to be myopic, sometimes with oblique astigmatism.
The two forms of achromatopsia are distinguished by the apparent
inheritance pattern, although patients with incomplete achromatopsia can
have better visual acuity (as good as 20/50 (50)); some color discrimination
on certain clinical tests (51), such as the Sloan achromatopsia test (10); and
macular changes with progression late in the course of the disease. In
contrast, patients with complete achromatopsia tend to have visual acuity of
20/200 or worse.
Fundus findings in achromatopsia are relatively normal, with a reduced
foveal light reflex in some cases. Some report macular atrophy in both forms
of achromatopsia late in the course of the disease (50). Nathans et al. (52)
have documented progression of disease in incomplete achromatopsia with
macular atrophy. Nevertheless, both conditions have remarkably normal-
appearing retinas and vessels. Electrophysiologic testing demonstrates loss of
cone responses in the presence of essentially normal rod responses. In
incomplete achromatopsia, minimal residual cone function can be detected
with specialized testing, but under standard ERG testing, cone function
typically is nonrecordable (Figure 26-4).
FIGURE 26-4 Full-field ERG of achromatopsia and
cCSNB. ERGs of a 12-year-old girl with congenital
achromatopsia, a 4-year-old patient with autosomal
recessive cCSNB, and a normal for comparison.
(Reproduced by permission of Taylor and Francis Group,
LLC, from Weleber RG. Infantile and childhood retinal
blindness: a molecular perspective [The Franceschetti
Lecture]. Ophthalmic Genet 2002;23:71–97, with
permission.)
The differential diagnosis for a child with decreased vision and nystagmus
includes albinism, LCA, achromatopsia, CSNB, chorioretinal coloboma,
peroxisomal disorders, optic nerve hypoplasia, aniridia/PAX6 variants, and
central nervous system causes. ERG characteristics and a patient’s presenting
symptoms are useful in differentiating among conditions. A study by Lambert
et al. (53) documented the misdiagnosis of LCA in 40% of patients. The most
common misdiagnoses were CSNB, Joubert syndrome (retinal dystrophy
with hypoplasia of the cerebellar vermis), RP, and achromatopsia. Other
conditions that have been misdiagnosed as LCA include infantile Refsum
disease, Zellweger syndrome, Senior-Löken syndrome (nephrophthisis and
retinal dystrophy), cone–rod dystrophy, and Alström syndrome (53,54). An
algorithm for the evaluation of patients with congenital nystagmus is
therefore useful; a version developed after evaluating the cause of congenital
nystagmus in 202 consecutive patients has been reported (55) and is
discussed extensively in Chapter 12.
Molecular Genetics of Achromatopsia

Complete achromatopsia (rod monochromacy) is an autosomal recessive
condition caused by defects in the cone photoreceptor cGMP-gated cation
channel. The channel complex is composed of two α-subunits and two β-
subunits. Mutations in the genes that code for each subunit have been
implicated in the pathogenesis of rod monochromacy. The gene for the α-
subunit (CNGA3) was the first gene identified and is located on chromosome
2q11 (56,57). Complete achromatopsia caused by changes in the CNGA3
gene is called complete achromatopsia type 2 and accounts for up to 25% of
all patients with complete achromatopsia. The β-subunit is coded for by the
gene CNGB3 and is believed to cause 40% to 50% of complete
achromatopsia (56). CNGB3 is found on chromosome 8q21 and is called
achromatopsia type 3. The inhabitants of the island of Pingelap carry a serine
to phenylalanine missense change (p.Ser322Phe) that results in their disease
(47). Another gene implicated in complete achromatopsia is GNAT2 located
on chromosome 1p13. GNAT2 codes for the transducin α-subunit in cone
photoreceptors. (GNAT1 codes for the α-subunit of transducin in rods.) Kohl
identified five families with complete achromatopsia caused by homozygous
changes in GNAT2 (50,58). Finally, PDE6C on chromosome 10q has also
been reported in association with complete achromatopsia.
Studies have shown successful gene supplementation therapy with
recombinant adeno-associated virus vectors (rAAV) for animal models with
CNGA3, CNGB3, and GNAT2 achromatopsia (59). Clinical trials are
currently underway to determine the safety and efficacy of similar gene
therapy treatments in patients with the more common forms of CNGA3 and
CNGB3 mutations (60).
Incomplete achromatopsia (blue cone monochromacy) is caused by loss
of the medium (green) and long (red) wavelength cone opsins encoded by
genes that are found together on the X chromosome. Three mechanisms have
been identified as causing incomplete achromatopsia. First, a locus control
region (LCR) was identified 4.02-kb upstream from the start of the red opsin
gene and more than 42-kb upstream from the start of the green opsin gene.
The LCR region is responsible for unwinding the DNA prior to transcription
regulated by the separate promoters of each respective opsin gene. A 579-bp
deletion in the LCR has been identified in 18.2% of patients with incomplete
achromatopsia studied; this mutation prevents transcription of either the red
or green opsin genes (52). The second mechanism is related to unequal
recombination of a red opsin gene with a mutant green opsin gene resulting in
nonfunctional proteins. The final mechanism resulted from deletion of the
green opsin gene with a missense mutation in the remaining red opsin gene
(61). The first mechanism demonstrates that a change in the coding sequence
4,000-bp upstream from the 5′ initiation site of the gene can still result in
disease, in this case incomplete achromatopsia.
Oligocone Trichromacy
Oligocone trichromacy was first reported by van Lith in 1973 with additional
patients subsequently being reported in the literature. This condition is
extremely rare and related to achromatopsia. The clinical presentation of
oligocone trichromacy differs from achromatopsia with normal or minimally
affected color discrimination. Otherwise, patients can present with subnormal
visual acuity, congenital nystagmus, a normal fundus, and photophobia. The
ancillary studies also mirror achromatopsia with normal visual fields and an
absence of cone function on ERG. Several achromatopsia genes, such as
CNGB3, CNGA3, PDE6C, and GNAT2, have been implicated, and Anderson
et al. (62) suggest that this condition is a mild version of achromatopsia with
preferential loss of peripheral cones and reduced number of foveal cones
(49).
Bradyopsia
Bradyopsia is a condition characterized by stationary cone dysfunction. It
was first described in 1991 in a series of three unrelated Dutch patients (63).
These patients presented with stationary subnormal vision, photophobia, and
prolonged electroretinal response suppression. In 2004, Nishiguchi et al.
identified mutations in the RGS9 or RGS9BP (R9AP) genes. The proteins
encoded by these genes interact with transducin to turn off the light-evoked
response. Patients with bradyopsia present with mildly reduced visual acuity,
photosensitivity, no dyschromatopsia, and normal or near-normal color
discrimination. Visual field testing shows mildly reduced general sensitivity
possibly with more pronounced central loss of function, and full-field ERG
testing demonstrates markedly abnormal cone responses. However, diagnosis
in young children can be difficult due to normal structural ophthalmic
examinations and improvement of visual acuity with pinhole (64–66). This
condition has only been reported in a very limited number of patients and is
considered to be a stationary cone dysfunction condition (67). Literature
notes varied mutations in patients of Dutch, Pakistani, Afghani, Guatemalan,
Chinese, British, Saudi, and Japanese descent—with a founder effect on the
Arabian Peninsula (65).
Bradyopsia shares many clinical characteristics with oligocone
trichromacy, making it difficult to discern the two conditions on clinical
findings alone (68). Differences are found at a cellular (photoreceptor) level
—with bradyopsia demonstrating an intact and normal photoreceptor
mosaicism compared to a sparse mosaic found in oligocone trichromacy (68).
Strauss et al. (68) highlighted the importance of cellular imaging to
distinguish these two conditions as well as to determine potential therapeutic
options.
Enhanced S-Cone Syndrome/Goldmann-Favre Disease

The enhanced S-cone syndrome (ESCS) was first described by Marmor et al.
(69) based on specific electrophysiologic findings. Patients with ESCS
presented with nyctalopia at an early age, cystoid changes in the macula, and
an annular pigmentary retinopathy involving the region around and just
outside the temporal vascular arcades (Figure 26-5). Visual acuity could be
as good as 20/20, but with the onset of cystoid changes of the macula, it
could be as severe as 20/200.
FIGURE 26-5 Fundus image of a patient with ESCS.
Patient has cystic changes involving the fovea with
midperipheral RPE changes characteristic of Goldmann-
Favre/ESCS. This patient underwent ERG testing,
including spectral ERGs, which demonstrated the
characteristic findings of ESCS. (Courtesy of Richard G.
Weleber, M.D.)
Color vision generally is unaffected, and the ERG shows a characteristic

appearance. Rod-isolated responses are nonrecordable. However, the
maximum combined response and photopic responses closely mirror each
other in amplitude and waveform morphology. Spectral testing of the cones
under photopic conditions demonstrates decreased function of long (L; red)
and medium (M; green) wavelength sensitive cones and enhancement of
short (S; blue) wavelength sensitive cones (69). Several patients with
Goldmann-Favre syndrome were also noted to have these ERG
characteristics. The gene for ESCS was identified as the NR2E3 gene in 2000
(70). This gene codes for a retinal orphan nuclear receptor, a transcription
factor that plays a role in determining cone photoreceptor phenotype in
embryogenesis. Without this receptor, cones remain as S-cones without
differentiating into L/M–cones. However, some cones are still able to
differentiate despite the loss of the NR2E3 gene product, which accounts for
the normal color vision and relatively good visual acuity that can be exhibited
by these patients. NR2E3 also is associated with a retinal degeneration
resembling Goldmann-Favre syndrome described in Crypto-Jews from
Portugal (71). These individuals demonstrate nyctalopia and constricted
visual fields. However, these patients were not studied with spectral ERGs to
determine whether they also had ESCS (50). Goldmann-Favre syndrome and
ESCS are now known to be different clinical manifestations of mutations in
the same gene, NR2E3 (72).
MISCELLANEOUS MACULAR
DYSTROPHIES
Pattern Dystrophy
Pattern dystrophy encompasses a heterogeneous group of macular
dystrophies with a wide range of phenotypic presentations. It is characterized
by a buildup of lipofuscin causing variable patterns of damage to the retinal
tissue. It is frequently caused by mutations in the peripherin/RDS gene,
renamed PRPH2 (73). A recent study also implicates OTX2 mutations in the
development of pattern dystrophy in two families (74). Pattern dystrophy
should be included in the differential diagnosis of macular dystrophies but is
not dealt with in detail in this chapter because of its later onset of
presentation. Typical patients with pattern dystrophies are identified in
adulthood. Even in pedigrees with known disease, macular changes are
typically noted in the third decade of life.
Best Disease
Best disease was first described by Adams in 1883. However, it was named
for Frederick Best, who described eight members of one family with the
classic macular changes 1905. The mode of inheritance is autosomal
dominant with incomplete penetrance and variable expressivity as some
patients have the abnormal allele and a normal-appearing fundus.
Consequently, the true prevalence of this condition is unknown. Still, it is not
a common disease; it has a reported prevalence of <1 in 10,000 in Iowa (75).
Clinical Findings
The clinical appearance of Best disease is highly variable and includes a
normal-appearing fundus (5% to 32%), macular RPE atrophy, or a gliotic
scar from choroidal neovascularization (76,77). The classic ophthalmoscopic
appearance of Best disease is symmetrical, bilateral, yellow-orange
vitelliform lesions (Figure 26-6) (76). The vitelliform lesion represents
lipofuscin pigment and has been reported to develop as early as shortly after
birth and as late as age 64 years (78). The vitelliform appearance seems to be
the least stable of the range of fundus manifestations seen in Best disease.
Vitelliform lesions evolve into one of several appearances depending on the
resorption of lipofuscin and include (a) a well-circumscribed gliotic scar; (b)
a pseudohypopyon; (c) a “vitelleruptive” scrambled, scattered distribution of
lipofuscin; and (d) simple macular atrophy. Attempts had been made to
classify the various findings into stages (77). Staging, however, implies a
chronologic progression from one appearance to another that is not
necessarily true in Best disease. Eccentric vitelliform lesions (ectopic
vitelliform lesions) and midperipheral fleck-like deposits also can be seen in
these patients (Figure 26-7) (75). Multifocal Best disease may occur as a
variant or represent a completely different condition (76). Gliotic nodules
were previously thought to be preceded by choroidal neovascularization
because classic choroidal neovascularization has been documented in Best
disease (78,79). However, choroidal neovascularization does not necessarily
precede gliotic lesions and is not required to be present when hemorrhage is
noted clinically. Incidental trauma has been documented to cause subretinal
hemorrhage in the absence of choroidal neovascularization in patients with
vitelliform lesions. Many of these patients will spontaneously recover vision,
depending on the amount of hemorrhage and degree of vision loss. However,
demonstration of a definite choroidal neovascular complex or severe vision
loss associated with hemorrhage may be an indication for intervention to
surgically evacuate the hemorrhage and extract the choroidal neovascular
complex (Figure 26-8) (79).
FIGURE 26-6 Best disease. Left macula of a 19-year-old
man with Best disease. Note the vitelliform lesion in the
macula. The other eye demonstrates a smaller vitelliform
lesion than the left eye. Visual acuity is 20/30 OD and
20/40 OS.
FIGURE 26-7 Flecks present in Best disease. Right eye of
a 54-year-old male with Best disease is shown using a
photographic montage. Note midperipheral lipofuscin
deposits. Visual acuity was 20/40 OD. (Reproduced with
permission from Mullins RF, et al. Late development of
vitelliform lesions and flecks in a patient with best disease:
clinicopathologic correlation. Arch Ophthalmol
Medical Association.)
FIGURE 26-8 Best disease with choroidal
neovascularization. A:Right eye demonstrates RPE
elevation and subretinal hemorrhage. B:Fundus
fluorescein angiography in the late phases shows leakage
consistent with choroidal neovascularization. Early phases
clearly demonstrated lacy choroidal neovascularization.
(Reprinted with permission from Chung M, et al. Visual
outcome following subretinal hemorrhage in Best’s
disease. Retina 2001;21:575–580, with permission.)
The visual prognosis for patients with Best disease is good; 75% of patients
maintain 20/40 or better vision in one eye through age 50 years (80). From a
series in Iowa, the average visual acuity was 20/25 for patients younger than
30 years, 20/30 for patients aged 30 to 60 years, and 20/40 for patients older
than 60 years (81). Even in the presence of a large vitelliform lesion, patients
typically retained relatively good visual acuity.
Lipofuscin blocks fluorescence on fluorescein angiography. Thus,
vitelliform lesions will block background fluorescence through most of the
angiogram. With evolution of the lesion, hyperfluorescence can be seen as a
window defect from RPE atrophy or as leakage from a choroidal neovascular
membrane (76). The ERG is normal in patients with Best disease, whereas
the electrooculogram (EOG), which measures the standing electrical potential
across the RPE, is markedly abnormal. The EOG is reported as a ratio
between the highest amplitude measured in a light-adapted state and the
lowest amplitude measured in the dark-adapted state. The EOG normally is
>1.7, but in patients with Best disease, it is below 1.5 and often 1.0 (76).
The differential diagnosis relates to the variable clinical appearances of Best
disease. Pattern dystrophy can present with a small vitelliform lesion or
atrophy. This phenotypic presentation of pattern dystrophy is often described
as adult vitelliform macular dystrophy. However, pattern dystrophy rarely
presents with visual symptoms or clinical signs in childhood. Stargardt
disease, North Carolina macular dystrophy (NCMD), and Best disease can all
have macular atrophy.
Family history and examination of family members can help to
differentiate these conditions. Multifocal Best disease (multifocal vitelliform
dystrophy) has also been described as a distinct autosomal dominant disease
with classic vitelliform lesions and a normal EOG.
Pathophysiology
Best disease was linked to chromosome 11q13 in 1992 (82), and the gene
was identified by Marquardt et al. in 1998. The gene product was called
bestrophin (83). Marmorstein et al. (84) localized bestrophin to the
basolateral plasma membrane of the RPE. A recent study also noted a
potential disruption of the polarized conductance due to a mutation in BEST1,
leading to a mislocalization of bestrophin-1 from the basolateral membrane to
the apical membrane (85). Sun et al. (86) and Gomez et al. (87) hypothesized
that bestrophin functions as a chloride channel or a putative ion exchanger.
This gene product now is recognized as a calcium-activated chloride channel
involved in regulating voltage-dependent calcium channels. Several
mutations have been implicated in defective calcium-activated chloride
channel export, as well as disruption of interactions between the N and C
terminus, resulting in altered chloride channel function (85). This gene also
plays a role in ocular development causing a distinct disease, autosomal
dominant vitreoretinochoroidopathy related to that function (88). This
condition is characterized by peripheral retinal and choroidal changes,
presenile cataracts, glaucoma, iris dysgenesis, and nanophthalmos (89,90).
Genetics
The presence of a family history of macular disease in a patient suspected of
having Best disease was a strong predictor of identifying a disease-causing
sequence variation in the gene (91).
Meunier et al. systematically screened patients with vitelliform lesions
for disease-causing changes in both the bestrophin gene (BEST1) and the
peripherin gene (PRPH2). They concluded that age of onset (<40 years) was
the major criterion to distinguish between Best disease and pattern dystrophy.
A disease-causing mutation in BEST1 was detected in 83% of patients
suspected of having Best disease, based on a family history and an abnormal
EOG (92). Boon et al. evaluated patients with multifocal vitelliform
dystrophy. They concluded that this group of patients was both clinically and
genetically heterogeneous. In this study, changes were found in the BEST1
gene (9/15 subjects) and the PRPH2 gene (1/15 subjects) (93).
Finally, the BEST1 gene is involved in causation of an autosomal
recessive bestrophinopathy (ARB). This disease is characterized by
lipofuscin deposition that is often multifocal in the retina, central vision loss,
and a markedly abnormal EOG. It is associated with many different biallelic
missense and nonsense mutations in BEST1. The abnormal gene products
from the identified mutations did not impair formation of wild-type channels
in a heterozygous state. The authors proposed that patients with ARB
represent the null phenotype for bestrophin disease in humans (94).
North Carolina Macular Dystrophy
First described in 1971 by Lefler et al. (95) and later in 1984 by Hermsen and
Judisch (96) as central areolar pigment epithelial dystrophy, NCMD was
renamed by Gass in 1987. Each of the families described in these initial
publications was related to the same family in the mountains of North
Carolina. The family was traced back to 1715 and now consists of more than
5,000 individuals throughout the United States. NCMD was the first macular
dystrophy demonstrating a clear inheritance pattern, but the causative gene
was elusive for many years (97). The gene was finally identified to be a
retinal transcription factor, PDRM13, in 2016 by Small et al. (98). Both
mutations and duplications have been reported to result in NCMD (98). This
disease was previously considered rare but has now been identified in
pedigrees in South Korea, Britain, Germany, and France. Additional families
in the United States that could not trace their ancestry to the original family
have also been identified (99–106).
Clinical Findings
NCMD is an autosomal dominant congenital macular disorder believed to be
completely penetrant with variable expressivity. Patients typically have much
better vision than would be predicted by their clinical appearances, with
visual acuities ranging from 20/20 to 20/400 and a median visual acuity of
20/60 (106). The fundus appearance can vary from a few scattered macular
drusen (Figure 26-9) to a large excavated macular staphyloma-like lesion
(Figure 26-10). Small (107) established a grading system for the fundus
characteristics. Grade I described scattered fine hard drusen, while grade II
described patients with confluent drusen in the central macula and grade III
described patients with a macular staphyloma-like lesion. The staphyloma
frequently has associated gliotic tissue involved in or at its rim (99,108–111)
and has been documented in a patient as young as age 3 months. Khurana et
al. assessed 13 members of a family with NCMD using spectral domain
ocular coherence tomography and ultrasonography. The classic lesion,
previously described as a staphyloma or coloboma, was found to not involve
the sclera but rather to represent deep chorioretinal excavations. They
suggested that these pathognomonic lesions for NCMD be termed “macular
caldera” (112). In general, NCMD is considered nonprogressive or, at worst,
minimally progressive. The congenital nature of this condition allows the
development of eccentric fixation and relatively good vision, better than
would be predicted, despite grade III changes (100). Choroidal
neovascularization has been documented and appears to be most common in
patients who present with extensive drusen in the central macular area. The
presence of choroidal neovascularization may result in severe vision loss
(108–111).
FIGURE 26-9 North Carolina macular dystrophy. Right

eye of a man from a family with NCMD. He demonstrates
relatively mild findings with drusen in the maculae of both
eyes. Visual acuity is 20/20 OU.
FIGURE 26-10 North Carolina macular dystrophy. Left
macula of a patient with a staphyloma-like lesion due to
NCMD. The patient demonstrates grade III fundus
characteristics with a pathognomonic macular caldera.
Also note the associated gliotic rim of tissue surrounding
the macular staphyloma.
For patients without the large bilateral macular staphylomata, the differential
diagnosis can include other macular dystrophies, such as Stargardt disease or
Best disease. Cone dystrophies may also present with macular changes but
will have characteristic symptoms and a diagnostic ERG. Congenital
toxoplasmosis and potentially other congenital infections can also present
with large excavated lesions and present in infancy and childhood.
Molecular Genetics
NCMD was first linked to chromosome 6 in 1991 with subsequent refinement
of its locus (MCDR1) to 6q16–21 (108–111). Since the first description of
MCDR1, families in France and Belize have been described with very close
linkage to the MCDR1 locus and a clinical appearance consistent with that of
the originally described family. There are 10 genes in the MCDR1 locus, and
in the original NCMD family, it was determined that a variant lies upstream
of the PRDM13 gene (98). This gene encodes a retinal transcription factor
and has been found to be the only gene in the MCDR1 region that is solely
expressed in the neural retina (98). As a member of a family of “helix–loop–
helix” DNA-binding proteins, PRDM13 plays a role in regulating gene
expression during development (98). Other families with a clinical
appearance resembling NCMD have been excluded from the MCDR1 locus
(112–115). Two groups studying families in Britain and Denmark,
respectively, subsequently identified a locus on chromosome 5 (5p13–p15)
associated with an autosomal dominant macular dystrophy—now known as
MCDR3—closely resembling NCMD (98,114,116). It was later determined
that a tandem duplication in the MCDR3 locus, downstream of transcription
factor IRX1, is likely the disease-causing mutation (98,117). Pattern, spatial
dosage, and timing of expression of IRX1 may be important in macular
development—as it was found to be highly expressed in the fetal macula at
19 to 20 weeks of gestation (117). Between the two loci, there is some degree
of genetic heterogeneity for patients who demonstrate the clinical features
and course of NCMD.
Autosomal Dominant Stargardt-Like Macular

Dystrophy
Autosomal dominant Stargardt-like macular dystrophy (STGD3) is an early-
onset degenerative disorder that affects the macula with a progressive
pathology (118). Genetic and genealogic studies have indicated that the
majority of cases in North America can be traced to a single large family of
over 2,000 individuals and indicates a common ancestor or founder (119).
Clinical Characteristics and Disease Course

STGD3 presents clinically in the early stages—during the first or second
decade of life—as visual loss and macular changes with or without yellowish
flecks (Figure 26-11) (120). A progressive degeneration of the macula
follows, eventually resulting in subsequent peripheral retina degeneration and
legal blindness.
FIGURE 26-11 Autosomal dominant Stargardt-like

macular dystrophy. Left eye of an ELOVL4-related
disease patient. Note the central retinal pigment epithelium
atrophy, macular degeneration, and yellowish flecks.
These pisciform flecks resemble similar lipofuscin flecks
in ABCA4-related disease such as Stargardt disease and
fundus flavimaculatus (working on provenance).
(Reproduced with permission of Arlene Drack, MD, UIHC
Dept. of Ophthalmology.)
In terms of therapeutic options, there are currently no existing definitive

treatments for STGD3 in humans. Animal models show conflicting results
when given DHA supplementation (121,122). However, MacDonald et al.
(123) demonstrated improvement in visual function both subjectively and
objectively in a 15-year-old patient after 2 months of DHA supplementation
(123). Thus, dietary supplementation may serve as a possible treatment
option. Choi et al. (124) also recommend fish oil supplementation although
no direct benefits have been implicated in the clinical course of STGD3
(124).
Differential Diagnoses
In the early stages, a fundus phenotype of atrophic macular degeneration with
or without yellowish flecks is shared with several other retinal conditions
(120). These conditions include fundus flavimaculatus, progressive dominant
cone dystrophy, pattern dystrophy, peripherin/RDS mutations, toxic
retinopathies, as well as autosomal recessive Stargardt macular dystrophy.
The progressive pathology, loss of central vision, atrophy of the RPE, and
accumulation of lipofuscin are also characteristic of age-related macular
degeneration—the difference being that STGD3 presents with an early onset
(118). The progressive nature of the disease also distinguishes it from
NCMD, whose gene is also located on the long arm of chromosome 6 (125).
Molecular Genetics of Autosomal Dominant Stargardt-

Like Macular Dystrophy
A genetic locus for STGD3 was first identified in 1994 by Stone et al. (125)
on chromosome 6q and further specified to the gene locus on chromosome
6q16 by Donoso et al. (119) in 2001.
Later studies determined that STGD3 results from mutations in the
ELOVL4 gene—whose protein is involved in the rate-limiting step of very
long-chain fatty acid (VLC-PUFA) synthesis (121). Several studies have
indicated varying numbers of base pair deletions in ELOVL4, all resulting in
a defective gene product (121,126,127). Other mutations occurring
downstream of ELOVL4 result in a truncated protein and mislocalization to
the Golgi, as well as the generation of toxic products from the aberrant
enzymatic activity of the truncated protein (121). Although the active site is
preserved in the truncated protein, it lacks innate enzymatic activity (128).
Additionally, the buildup of toxic intermediates may cause an additive
effect in decreasing the number of VLC-PUFA products—implicating direct
supplementation with VLC-PUFA as a possible treatment option (126).
Sorsby Fundus Dystrophy
Originally described by Sorsby et al. (129) in 1949, Sorsby fundus dystrophy
(SFD)—also called Sorsby pseudoinflammatory macular dystrophy—is an
autosomal dominant retinal dystrophy (130). The average age of
manifestation is usually in the fourth or fifth decade of life; however, since its
initial discovery, an early-onset form has also been identified. In a Finnish
family with consanguineous parents, each of the eight children were affected
—with the most extreme case showing symptoms at age 13 (131).
SFD is characterized by the loss of central vision in the early stages as a
result of macular disease and choroidal neovascularization. Color vision
defects and complaints of nyctalopia may also be present. The disease then
progresses to peripheral loss later in life. In the original report, Sorsby et al.
(129) noted bilateral macular hemorrhage, edema and exudates, as well as
exposure and sclerosis of choroidal vessels. The predominant histopathologic
feature continues to be amorphous, drusen-like deposits between the RPE and
the inner collagenous layer of Bruch membrane (132). In 1994, Weber et al.
(131) demonstrated linkage to markers on chromosome 22q12 in a single
large family with SFD. It was determined that the tissue inhibitor of
metalloproteinases-3 (TIMP3) gene is implicated in the pathogenesis of SFD
through its role in extracellular matrix remodeling (131). Most SFD-
associated variants have involved a gain or loss of a cysteine residue (133).
However, differing clinical pictures and interfamilial phenotypic variations
noted in several studies implicate the possibility of genetic heterogeneity
(130,134–136).
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27
Leber Congenital Amaurosis
Razek Georges Coussa, Robert K. Koenekoop, and Elias I.
Traboulsi
PREVALENCE
In 1869 Theodore Leber first described a very early-onset form of severe
vision loss that manifested itself before age 1 year and in which patients had
nystagmus and sluggishly reactive pupils. Franceschetti subsequently
demonstrated that these patients had markedly reduced or nonrecordable
electroretinograms (1). The spectrum of clinical inherited retinal diseases that
is now referred to as Leber congenital amaurosis (LCA) has become broader
and includes patients with vision that ranges from no perception of light
(NLP) to fairly preserved central vision. To complicate this further, we now
recognize many systemic disorders that have a retinal degeneration that
simulates LCA. In 1985, Foxman proposed subdividing patients into four
groups based on the onset of their retinal disease, and any associated systemic
findings. Group 1 is composed of uncomplicated LCA with ophthalmic
findings but no systemic disease. Group 2 includes patients with retinal
disease and systemic findings such as Senior-Löken syndrome (SLSN) or
cerebrohepatorenal syndrome of Zellweger. Groups 3 and 4 consist of
children with initially normal vision but with very early onset of signs and
symptoms consistent with retinitis pigmentosa (RP) (2). Most important in
the definition of LCA is the absence of another specific retinal or multisystem
disorder (3). Today, LCA is recognized as a heterogeneous group of inherited
retinal disorders that together account for approximately 5% of all retinal
dystrophies and approximately 10% to 20% of all cases of congenital
blindness (4,5). The estimated prevalence of this clinical group of diseases
ranges from 1:33,000 (6) to 1:81,000 (7).
There are no environmental factors that contribute to LCA at this writing.
GENETICS
Currently, 20 genes associated with LCA account for more than 70% of cases
of nonsyndromic LCA in the Western world (8). These genes, which are all
related to retinal function or structure are GUCY2D (LCA1), RPE65 (LCA2),
SPATA7 (LCA3), AIPL1 (LCA4), LCA5 (LCA5), RPGRIP1 (LCA6), CRX
(LCA7), CRB1 (LCA8), NMNAT1 (LCA9), CEP290/NPHP6 (LCA10),
IMPDH1 (LCA11), RD3 (LCA12), RDH12 (LCA13), LRAT (LCA14),
TULP1 (LCA15), IQCB1/NPHP5, CLUAP1, PRPH2, KCNJ13, and IFT140.
Interestingly, the 20 causal LCA genes can be categorized into seven
distinct molecular pathways, each affecting a particular aspect of retinal
function and/or structure and, thus, potentially causing specific phenotypic
manifestations (Table 27-1).
TABLE 27-1 Molecular pathways causing LCA
WORLDWIDE IMPACT
The contribution of each gene to the totality of cases in any one population
varies according to geographic location possibly due to founder effect–related
predominance. For example, the NPHP6 (CEP290) gene accounts for 15% to
20% of LCA in Northwestern Europe and as much as 30% of LCA in North
America (7). However, mutations in this gene are rare causes of LCA in other
geographic regions, such as southern India (9). Mutations in RPE65 account
for <10% of cases in North America but about 85% in Costa Rica (10).
PATHOPHYSIOLOGY
The wide range of clinical phenotypes and severity of vision loss in patients
with LCA can probably be explained on the basis of the underlying genetic
heterogeneity as well as gene–gene interactions and epigenetic factors (7).
Interestingly, some of the LCA genes have been associated with later-onset
retinal degeneration as well as with congenital onset (Table 27-2).
TULP1 has been a cause of early-onset RP rather than LCA (4,11,12).
CRX (95) and GUCY2D (96) have been associated with both autosomal
dominant cone–rod dystrophies, and with congenital night blindness (13).
CRB1 has been reported to cause RP12 with the particular phenotype of
preservation of periarteriolar RPE (14,15). RPE65 has also been associated
with later-onset retinal dystrophies (16). Mutations in another uncommon
gene causing autosomal recessive LCA, RDH12, has resulted in autosomal
dominant RP phenotype (17). Table 27-2 summarizes the different types of
LCA, designated by their underlying genetic etiology, and their sometimes
unique clinical characteristics (2,18,19).
TABLE 27-2 Types of LCA by underlying

causative genes
Clinical phenotypic manifestations and other associated findings are also given. Phenotypes and
associated findings are not present in every patient.

Given the genetic heterogeneity of LCA, its presentation and course can be
highly variable. In general, the disease manifests itself in the first 6 to 12
months of life with visual inattention, absence of fixation and tracking, as
well as pendular nystagmus (104). Some patients demonstrate paradoxical
pupillary responses to light (Flynn pupils) and may exhibit persistent and
vigorous eye pressing (oculodigital sign) (104).
Interestingly, children with GUCY2D-related LCA typically exhibit
significant photophobia, whereas those with RPE65-related LCA present with
nyctalopia and better vision in mesopic illumination. Photophobia has also
been reported in some children with CEP290- or RPGRIP1-associated LCA.
The presenting visual acuity of children with LCA may vary between
20/200 or better, especially in patients with RPE65 or CRB1 mutations, and
occasionally in some patients with CEP290 mutations, to no light perception
(NLP). The natural history of vision loss in LCA is complex but can be
divided into three categories: In the majority of LCA patients (71% to 75%),
VA remains stable, while 15% and 10% show VA deterioration and
improvement, respectively (6,104–107). In their longitudinal study of 27
LCA patients, Fulton et al. reported a VA ranging between 20/500 and NLP
in one-third of their patients (105).
There is reasonable evidence for genotype–phenotype correlations among
LCA patients (108). Children with LCA caused by mutations in the CRB1,
RPE65, LRAT, and CEP290 genes may have VA better than 20/50 at
presentation (105,109), whereas those with mutations in GUCY2D and CRX
genes tend to have more severe visual impairment (110,111). This wide
variability in VA at presentation could potentially result in misdiagnosing the
child with other early childhood–onset retinal dystrophies (ECORD, defined
as VA better than legal visual blindness before 10 years of age (111)) or
severe early childhood–onset retinal dystrophy (SECORD, defined as legal
blindness before 10 years of age (112)) or even infantile “motor” nystagmus
rather than LCA. Patients are occasionally diagnosed with bilateral
amblyopia or with delayed visual maturation. What is common, however, in
many LCA patients is a certain stability in central VA despite a noticeable
progression in retinal pigmentation over the course of the disease. LCA
patients with macular atrophic changes may also lose central VA over time
(106).
Due to the genetic heterogeneity of LCA, there appears to be a
corresponding phenotypic heterogeneity in ophthalmoscopic findings. Retinal
findings range from a normal-appearing fundus to more severe macular
changes such as pseudocolobomas and/or chorioretinal atrophy. In fact, it is
estimated that about 50% of LCA patients have a normal-appearing fundus in
the first year of life (Figure 27-1) (106). The remaining patients could have
typical blood vessel attenuation, bone spicule RPE pigmentation (Figures 27-
4E–F, 27-5E–F, and 27-6C–D), or nummular pigmentation (Figure 27-3).
FIGURE 27-1 A 21-year-old woman with mutation in
GUCY2D. A, B: OD and OS color photos show normal-
appearing macula and optic nerves. C, D: Normal wide-
field color photos of OD and OS. E, F: Normal wide-field
FAF photos of OD and OS. G, H: Grossly normal foveal
SDOCT OD and OS.
FIGURE 27-2 A 2-year-old girl with mutation in RPE65.
A: OD color photo shows foveal RPE nummular
pigmentation at 2 years of age. B, C: OD and OS foveal
SDOCT show a compact retina with a speckled pattern at
the level of IS/OS junction at 3 years of age. D, E: OD and
OS color photos showing stable foveal RPE nummular
pigmentation at 4.5 years of age.
FIGURE 27-3 A 16-year-old girl with mutation in CRX.
A: OD color photo shows foveal RPE nummular
pigmentation. B: OS color photo shows foveal RPE
nummular pigmentation and inferonasal RPE
depigmentation. C: OD color photo shows inferonasal
RPE depigmentation. D: OS color photo shows RPE
depigmentation extending into the inferior midperiphery.
FIGURE 27-4 A 13-year-old girl with mutation in CRB1.
A, B: OD and OS color photos show foveal RPE
depigmentation, bone spicules, and preservation of the
para-arteriolar RPE. C, D: OD and OS FAF photos reveal
macular and peripapillary hypoautofluorescence with
hyperautofluorescence along the para-arteriolar RPE. E, F:
OD and OS wide-field color photos demonstrate foveal
RPE depigmentation, bone spicules, and preservation of
the para-arteriolar RPE. G, H: OD and OS FAF wide-field
photos show macular, peripapillary, and midequatorial
hypoautofluorescence with hyperautofluorescence along
the para-arteriolar RPE. I, J: Foveal SDOCT OD and OS
reveal thickened fovea with abnormally laminated retina.
FIGURE 27-5 A 23-year-old man with mutation in
CEP290. A, B: OD and OS color photos show foveal RPE
depigmentation and blunted foveal reflex. C, D: OD and
OS FAF photos demonstrate bull’s-eye foveal hypo-
/hyperautofluorescence and inferonasal patchy
hypoautofluorescence. E, F: OD and OS color photos
show midperipheral and peripheral nummular RPE
pigmentation with bone spicules. G, H: OD and OS FAF
photos show bull’s-eye foveal hypo-
/hyperautofluorescence pattern and peripheral patchy
hypoautofluorescence more prominent nasally. I, J: OD
and OS foveal SDOCT reveal perifoveal loss of IS/OS
junction.
FIGURE 27-6 A 13-year-old boy with mutation in
RDH12. A, B: OD and OS color photos show foveal RPE
pigmentation and atrophic changes, blood vessel
attenuation, and scant bone spicules. C, D: OD and OS
wide-field color photos reveal nummular RPE
pigmentation and scant bone spicules. E, F: OD and OS
wide-field FAF photos demonstrate macular
hypoautofluorescence. G, H: OD and OS foveal SDOCT
show abnormal retinal architecture and diffuse loss of the
IS/OS junction.
Macular pseudocolobomas are associated with LCA5- and NMNAT1-related

disease. Bull’s-eye macular changes have also been reported in TULP1 and
RD3 mutations (Figure 27-7) (12). Peripheral retinal findings also vary from
bone spicule pigmentation, which can occur as early as 9 months of age, to
diffuse RPE atrophic changes (Figures 27-4E–F and 27-6C–D) (3).
Interestingly, patients with mutations in CRB1 have very characteristic
preservation of para-arteriolar RPE (Figure 27-4A–H) and thickened retina
with macular cystic changes on optical coherence tomography (OCT)
(Figure 27-8I–J). These phenotypic–genotypic correlations are all
summarized in Table 27-1.
FIGURE 27-7 An 8-year-old girl with mutation in
TULP1. A, B: OD and OS color photos show foveal RPE
atrophy. C, D: OD and OS wide-field color photos
demonstrate nummular RPE pigmentation. E, F: OD and
OS wide-field FAF photos show foveal
hyperautofluorescence with peripheral granular
hypoautofluorescence. G, H: OD and OS foveal SDOCT
reveal a staphyloma-like foveal contour with perifoveal
loss of IS/OS junction.
FIGURE 27-8 A 16-year-old woman with mutation in

CRB1. A, B: OD and OS foveal SDOCT show thickened
retina with macular cystic changes.
Other associated clinical findings include oculodigital sign, paradoxical

and/or poorly reactive pupils, keratoconus (reported in CRX-, GUCY2D-, and
AIPL1-related LCA), high hyperopia in most types or myopia (TULP1-
related LCA), and cataracts (reported in CRX-, GUCY2D-related LCA) (12)
and CRB1.
DIAGNOSTIC STUDIES
Due to the extensive phenotypic heterogeneity and the potential overlap with
other conditions associated with nystagmus and decreased vision early in life,
clinical testing with electroretinography, OCT, fundus autofluorescence
(FAF), and Goldmann visual field when possible are all useful and
complementary (Table 27-1).
Any child who presents with infantile nystagmus, inability to track visually
early in life, and/or decreased vision should have an exhaustive ophthalmic
exam. The differential diagnosis for infantile nystagmus, which is the most
common sign for infantile and early-onset inherited retinal dystrophies, is
broad and includes primary sensory retinal abnormalities (LCA,
achromatopsia, blue cone monochromacy, and congenital stationary night
blindness [CSNB]), vitreoretinal abnormalities (retinopathy of prematurity,
Norrie disease, Familial exudative vitreoretinopathy), foveal hypoplasia
(albinism, aniridia), optic nerve disorders (hypoplasia, coloboma), congenital
infectious retinochoroidopathies (syphilis, rubella, TB, toxoplasmosis),
congenital glaucoma, bilateral amblyogenic cataracts, as well as severe
corneal opacifications due to congenital malformations or systemic diseases
causing corneal depositions. One negative test should never be definitive as
the end of a workup. Rather a decision tree that directs attention to alternate
tests after a negative finding is helpful in the workup of infantile nystagmus
(113).
In most cases, LCA presents typically as an isolated condition without
syndromic manifestations. Occasionally, similar retinal findings are
associated with systemic diseases (syndromes) that may carry significant
morbidity and potentially mortality for the infant. Hence, it is crucial not to
overlook medical conditions for which additional systemic care by other
medical specialty services may be necessary as the correct diagnosis would
allow genetic counseling and family planning. Systemic abnormalities that
should raise red flags include deafness, dilated infantile cardiomyopathy
(Alström syndrome) nephropathies, neurodevelopmental delay,
microcephaly, and CNS malformations as well as skeletal abnormalities. In
many of these situations, associated systemic findings, such as seizures,
failure to thrive, and other neurologic or systemic signs, will lead the
diagnosis away from isolated LCA, but the potential still exists for the
ophthalmologist to assist in making the definitive syndromic diagnosis for the
patient and his/her family. Imaging studies, metabolic/biochemical studies,
and a pediatric and medical genetic evaluation are indicated in these cases
(114).
In one study, the rate of LCA misdiagnosis was 40% (3). The most
common misdiagnoses were CSNB, RP, achromatopsia, and Joubert
syndrome (early-onset LCA-like retinal dystrophy, hypoplasia of the
cerebellar vermis, hypotonia, developmental delay, cystic nephronophthisis).
Other conditions that have been misdiagnosed as LCA include Refsum
disease (cerebellar ataxia, peripheral neuropathy, elevated cerebrospinal fluid
protein levels, progressive deafness, anosmia, cardiac dysfunction and RP),
Zellweger syndrome (neurologic dysfunction, craniofacial abnormalities,
hepatomegaly, renal cysts, hypotonia, and RP), Alström syndrome (childhood
obesity, sensorineural hearing loss, hyperlipidemia, type 2 diabetes mellitus,
dilated cardiomyopathy, renal and hepatic dysfunction, alopecia, and RP) (3).
In Senior-Löken syndrome, a typical LCA presentation may be followed in
later life by nephronophthisis and renal failure; all of the systemic
manifestations of syndromes may not be present in infancy; therefore, a
review of systems and reassessment as the child ages are important in follow-
up.
Distinguishing between LCA and early-onset RP may be more of a
semantic issue, especially when mutations in some of the genes implicated in
LCA (e.g., RPE65, CRX, TULP1, RDH12) may also be a cause of later-onset
RP. LCA is almost exclusively autosomal recessive (CRX mutations cause an
autosomal dominant form), while RP has varied forms of inheritance (113).
Patients with early-onset RP will frequently lack searching nystagmus and
have a slightly older age of onset as well as better central vision (4,12). For
the overwhelming majority of patients with LCA, the ERG is essentially
nonrecordable for all stimuli, while that in early-onset RP may relatively
spare the photopic component. Fazzi et al. suggested that the visual
impairment in LCA is more stable compared to RP that results in a more
progressive visual decline (113). Koenekoop suggested that the genetic
changes associated with LCA may affect normal retinal development (6,114).
CLASSIFICATION
There are four possible classification schemes for LCA based on (a) VA and
natural progression, (b) the disease molecular pathway (Table 27-2), (c)
foveal OCT findings (Table 27-3) (42), and (d) histopathology of the retina
from enucleated eyes of patients with LCA (Table 27-3) (6).
TABLE 27-3 LCA classification patterns and their

respective associated genes
These classification schemes help to channel the genetic workup toward

specific causal genes and potentially help in disease prognostication and
patient counseling; furthermore, they are key to understand disease structural
and functional expression on a cellular level in order to guide emerging
medical and surgical treatment strategies.
MANAGEMENT
The first step in managing children suspected of having LCA is early and
accurate clinical and molecular genetic diagnosis. The latter would then allow
the proper course of clinical interventions that impact the child’s visual and
functional development, education, as well as future family planning. It is
imperative to rule out systemic diseases with an LCA retinal phenotype.
Currently most gene panels for LCA will include genes that cause many if
not all such syndromes, including Alström syndrome, Joubert syndrome, as
well as several other ciliopathies. It is important to review all of the genes
included in a panel when ordering to avoid either omission of possible
syndromic genes of interest or inclusion of genes that should be discussed
ahead of time, such as those that cause fatal illnesses like Batten disease. One
of the challenges is the attribution of any developmental delay to the child’s
blindness, when in fact it could be a sign of CNS dysfunction. Many
clinicians routinely obtain renal function tests in patients with LCA; others
order brain MRIs looking for evidence of Joubert syndrome. Appropriate
molecular genetic testing and the identification of the responsible gene is the
single most important diagnostic step in the management of patients with
LCA and other retinal dystrophies, as accurate molecular genetic diagnosis
helps guide additional medical workup and surveillance.
Families are always devastated as they learn about their child’s disease
and need counseling and support. There are numerous family support groups,
many centered on a particular LCA gene. Families can be directed to join
such groups and discuss potential challenges with others, but they should be
cautioned about the variability in the severity of the diseases and their natural
course even within the same underlying gene group.
In December 2017, the Food and Drug Association (FDA) approved, for
the first time, the use of subretinal injections of adeno-associated virus
(AAV) vectors carrying RPE65 cDNA for the purpose of gene replacement
and augmentation in patients with RPE65-associated inherited retinal
dystrophies (19). The treatment resulted in significant improvement in the
ability of patients to navigate under dim lighting conditions; this
improvement was maintained for at least 3 years posttreatment (18,115).
In December 2018, a new form of treatment was successfully tested and
showed promising results. Cideciyan et al. treated 10 LCA patients with
pathologic mutations in CEP290, at least one of which was 2991+1655A>G,
which alters splicing, with intravitreal injections of antisense oligonucleotide.
The authors reported improvements in visual acuity or retinal sensitivity in
four patients, which were retained at the 6-month follow-up.
Most eyes tolerated treatment well; however, some lenticular opacities
were reported (116). More detailed information on LCA treatment is
discussed in Chapters 28 and 68.
When an infant is diagnosed with vision loss, an early intervention plan must
be put in place immediately to assure that the child begins to learn by
nonvisual means and that parents know how best to support this. Local public
school systems usually have a developmental team in place to assess and
support visually impaired children even before school age, and families
should be encouraged to contact their local school system. All children and
adults with LCA should be referred to a low-vision clinic upon diagnosis.
Low-vision specialists follow these patients throughout life and can
recommend appropriate accommodations as needs and goals change (see
Chapter 14). School accommodations are especially important. The
Americans with Disabilities Act states that by law, public schools must
provide disabled students with all necessary accommodations, but these must
be delineated and advocated for in each student’s case. Technology has
revolutionized academic and personal access to visual information for the
visually impaired. Children and parents should be connected to resources to
educate them about these devices and applications.
ROLE OF OTHER PHYSICIANS AND

Ophthalmologists must communicate with the primary care providers of
patients with LCA at diagnosis to make them aware of the potential for
systemic issues or to inform of known associations once a molecular genetic
diagnosis is confirmed. In a blind child, even temporary hearing loss can be
severely disabling, so ear infections must be treated promptly. Low-vision
specialists can advise about important activities such as driving, and
assessment should be scheduled near the age of driving permit eligibility (see
Chapter 14).
With the advent of an FDA-approved gene therapy for one type of LCA,
there is a heightened moral obligation for clinicians to arrive at an accurate
molecular genetic diagnosis in early life for children with infantile nystagmus
and/or poor vision. Because genetic results may also impact family planning,
physicians should consider referring families to a genetic counselor or
medical geneticist for an in-depth discussion of recurrence risk for other
children and procedures such as IVF with preimplantation genetic testing to
reduce the risk of having other affected children.
FUTURE TREATMENTS
Gene therapy is already a reality for one form of LCA. Although other
genetic types may not be as easy to correct as RPE65 with gene
replacement/augmentation, there is hope that gene therapy or cell therapy will
improve vision for many patients with LCA in the future.
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28
Gene Therapy
Tomas S. Aleman, Albert M. Maguire, and Jean Bennett
INTRODUCTION
Status of Gene Therapy for Genetic Disease
The progress in identifying the structure of DNA, deciphering the genetic
code, and generation of tools with which to manipulate DNA in the 1950s-
1960s led many to realize that it might be possible to manipulate and deliver
genes to humans in order to ameliorate disease (1). The first human gene
transfer experiment was done in 1970 by Stanfield Rogers, an American
doctor at the Oak Ridge National Laboratory in Tennessee. Rogers, along
with a German physician, attempted to treat two sisters (18 months and 5
years old) with seizures and mental impairment due to argininemia (2). They
used the Shope papilloma virus to try to reduce the patients’ arginine levels,
thinking (erroneously) that delivery of a gene that regulated arginine
expression would ameliorate the disease. While their efforts may have been
well intentioned, they did not have a well-devised study design and the study
results were never published. The event did attract the attention of other
scientists, however. Concern about the conduct of this gene therapy
experiment led to establishment of a Recombinant DNA Advisory Committee
(RAC; established 1974) whose mission was to regulate recombinant DNA
research.
When another physician, Martin J. Cline, MD, went to Italy in 1980 and
delivered bone marrow cells exposed to recombinant DNA to 2 patients with
β-thalassemia, there was general outrage (3). The actions were in direct
opposition to NIH gene therapy guidelines and university guidelines.
Fortunately, no one was harmed, and as a result of his action, Dr. Cline’s
research career came to an abrupt end. The first approved human gene
transfer study was initiated in September 1990 and that targeted adenosine
deaminase (ADA) deficiency, a form of severe combined immune deficiency.
The intervention involved intravenous delivery of white blood cells infected
ex vivo with an ADA-containing retrovirus (4). The patient, who was treated
during childhood, is now in her 30s and remains healthy. This study and
further developments in technology for cloning genes, for generating
transgene cassettes, and for developing recombinant viral vectors provided
further impetus to the field.
While the gene therapy field had significant setbacks around the turn of
the century, technologic developments and sophisticated study designs are
now beginning to pay off. The first country to approve a commercial gene
therapy product, an oncolytic adenovirus, was China in 2004 (5). Glybera,
which targets lipoprotein lipase deficiency, was the first gene therapy to be
approved in Europe (in 2012). The first gene therapy drug to be approved by
the Food and Drug Administration (FDA) in the United States and recently
the European Medicines Agency (EMA) is Luxturna (voretigene neparvovec-
rzyl), an agent designed to ameliorate a pediatric onset inherited retinal
dystrophy (see below) (https://seekingalpha.com/news/3411817-novartis-
lands-luxturna-approval). A number of combination cell and ex vivo gene
therapy reagents have also been approved in the United States, including
Kymriah and Yescarta, reagents targeting B-cell acute lymphoblastic
leukemia and lymphoma, respectively.
These successes have fueled interest and investment in further
development of gene therapies, including those that target pediatric retinal
disease.
General Principles of Gene Therapy

Gene therapy is a pharmacologic approach whereby exogenous genes or
nucleic acids are introduced into host cells to effect local production of a
therapeutic protein (gene augmentation therapy), to halt expression of a
disease-associated protein (gene suppression therapy), or to repair or edit a
transcript. In gene augmentation therapy, the product can be a protein
(enzyme, structural protein, etc.) or RNA.
Gene correction can also be achieved using delivery of nucleic acids
designed to correct a specific mutation (Figure 28-1).
FIGURE 28-1 Retinal genes targeted in gene
augmentation therapy clinical trials to date (May 2019).
They are indicated according to the year in which each
gene was first identified as disease causing. Modified from
RetNet (https://sph.uth.edu/retnet/sum-dis.htm#D-graph).
ND4 (6), CHM (7), PDE6β (8–9), MYO7A (10), RPGR (11),
ABCA4, (ABCR) (12), RPE65 (13,14), RS1 (15), RLBP1
(CRALBP, CPK) (16), CNGB3 (17), MERTK (15,18),
CNGA3 (19), CEP290 (20). In many cases, multiple
groups nearly simultaneously identified disease-causing
genes. Due to space limitations, only one or two references
are listed. A gene therapy product for RPE65 deficiency
(Luxturna) was the first to be approved in the United
States and the European Union. Phase III clinical trials are
in progress for choroideremia (CHM) and Leber’s
Hereditary Optic Neuropathy caused by ND4 mutations.
Gene therapy can take advantage of the normal transcriptional and

translational apparatus of the host cells to produce the therapeutic molecule
encoded by the exogenous genetic material. Gene therapy therefore requires
two events: (a) uptake of the template genetic material and transport to the
target cell nucleus and (b) expression of the exogenous gene by the host
tissue. It can be used to replace an essential gene product in a disease that is
caused by an inherited gene mutation. It can also be used to modulate the
production of a substance to positively affect the outcome of a disease that
has a complicated pathogenesis, that is, a combination of genetic and
environmental insults. An attractive feature of gene therapy is that it may
more specifically target affected cells or tissues and, with prolonged
expression, minimize the number of drug administrations.
Although the spectrum of therapeutic molecules that can be produced
through gene therapy is limited to peptides, proteins, and nucleic acids, this
does not limit the types of diseases that can be treated. In much the same way
that vascular endothelial growth factor inhibitors are presently used to treat
neovascular complications of diabetes, vascular occlusive disease, and age-
related macular degeneration, genes can produce growth factors or
neurotrophic factors that have a nonspecific effect applicable to a variety of
disease processes. For example, in animal studies, rod-derived cone viability
factor (RdCVF) has a potent cone photoreceptor survival effect in several
animal models of early-onset inherited retinal degeneration (IRD) (21–24).
The gene encoding RdCVF, nucleoredoxin-like (Nxnl) also encodes a second
product that has enzymatic properties that serve to protect against oxidative
damage (22). Thus, RdCVF may be useful for treating a variety of disorders,
including retinitis pigmentosa (RP), Leber congenital amaurosis (LCA), and
macular degeneration, and may also be useful in enhancing the success of
other therapeutic strategies, such as stem cell transplantation (Figure 28-1)
(see also e-book, chapters 7 and 8) (23).
*Note that since so many diseases are discussed in this chapter, in lieu of a separate outline describing
each disease, information on the genetics, diagnosis, symptoms, etc. relevant to the diseases, is included
in Chapters ?23, 27 and 29-36.
Therapy can be highly specific, with replacement or augmentation of

defective gene products in genetic diseases caused by lack-of-function gene
mutations. For example, gene augmentation therapy using an RPE65-
encoding cDNA can provide specific treatment for a type of LCA caused by
mutations resulting in lack of a functional RPE65 gene product (Figure 28-2;
Table 28-1) (46–50).
FIGURE 28-2 Intraocular gene delivery approaches.
Cartoons of the structure of the healthy retina are
contrasted with those at different stages of degeneration.
In the diseased retina, if photoreceptor cells persist, it may
be possible to reverse vision loss through delivery of the
wild-type version of the disease-causing gene (i.e., gene
augmentation therapy). Alternatively, it may be possible to
delivering oligonucleotides, which mediate correct
splicing of the transcript or to correct the gene/transcript
using CRISPR-Cas or other editing approaches. It may
also be possible to maintain the health of neural retina
through delivery of genes encoding neurotrophic or
metabolic support factors. If photoreceptor cells have
degenerated, transplanted cells (potentially treated with
gene therapy reagents ex vivo) may provide some visual
function. Finally, viral vectors can be employed that
render other cells in the retina light sensitive through
delivery of optogenetic molecules.
TABLE 28-1 Gene therapy clinical trials for

retinal diseases using recombinant viruses as of
May 2019
aPhotoreceptor-specific promoter, but details not disclosed.
bNot yet recruiting.
SR, subretinal; IVT, intravitreal; NA, not available; AAV, adeno-associated virus; lenti, lentivirus;
CBA, chicken beta actin; CMV, cytomegalovirus, FO, follow on; Shaded panels: enrollment and
follow-up completed.
Treatment of pediatric retinal diseases presents special issues not encountered

with adult disease. The biology of many pediatric retinal conditions interfaces
with the development of the retina and the visual system. Some conditions
are expressed prenatally and are manifest at birth, for example, the lack of
foveal development in oculocutaneous albinism. Treatment performed after
birth would be expected to have limited impact on these conditions because
the biologic effects may be irreversible. In utero ocular gene therapy has been
demonstrated experimentally and would provide an avenue to correct the
defects before tissues and organ systems are established by aberrant
developmental pathways (51). There are other blinding conditions that are
detectable early in life but in which the cellular anatomy is roughly intact. In
such diseases that develop pathologic changes postnatally, such as
retinopathy of prematurity (ROP) or familiar exudative vitreoretinopathy
(FEVR), there may be a window of time for intervention after birth to prevent
permanent damage of foveal structure and impairment of central visual
pathways from amblyopia.
Although the technical demands of in utero and early postnatal
intervention would present major challenges, there would be distinct
advantages. Prenatal application of an exogenous gene would help induce
immune tolerance to the foreign antigen. Transduction efficiency would
likely be enhanced because a larger area of the outer retina could be targeted
with one injection. This is not only because of the greater number of cells
present per unit area in the fetal retina but also because there are no
attachments between the neural retina and the retinal pigment epithelium
(RPE) early in development. This permits free diffusion of the subretinally
administered drug. Correction of the defect at an early stage would also
reduce the occurrence of later tissue damage. This is important in early-onset
IRDs because photoreceptors, which die in these diseases, are terminally
differentiated cells after birth and cannot regenerate. Finally, early
application of the therapeutic gene would take advantage of the plasticity of
the developing retinal to central nervous system (CNS) axonal connections
present early in life. If treatment were performed too late in development,
higher order CNS neurons might already have established synapses from
other sensory systems, and cortical areas for vision would no longer exist.
Several retinal diseases exhibit a short, well-defined period of biologic
activity in the early postnatal period and early childhood. Intervention would
need to occur during or before infancy in order to arrest the disease process
and salvage retinal function. For example, threshold ROP occurs on average
about 37 weeks postgestational age. Retinal detachment as stage 4 ROP
typically occurs about the time of what would have been a full-term delivery,
40 weeks gestation. The ideal window of opportunity for treatment in ROP is
thus limited to a period of a few weeks. In inherited retinoblastoma, tumor
appearance occurs within the first year of life and sometimes presents at
birth. Effective treatment for retinoblastoma would require intervention
within a narrow time period of active tumorigenesis. In LCA, visual deficits
are present at birth or in early childhood. The retina undergoes progressive
degeneration through childhood and adult life. Intervention with gene therapy
should ideally take place within a defined time period before histologic
degeneration of photoreceptors and retina develops. Similarly, in other
diseases that are first symptomatic in childhood (choroideremia, Stargardt
disease, early-onset RP), intervention early in the course of the disease may
ultimately provide the most benefit.
There are other important considerations in treating pediatric retinal
diseases. One question is how an intervention would impact photoreceptors
that are still undergoing differentiation? This is a potential issue for foveal
delivery in infants <6 months of age and was considered in the labeling of the
recently approved drug for RPE65 disease (Luxturna). In addition, the
currently available techniques for subretinal injection have utilized standard
pars plana vitrectomy methods. This is problematic in infant eyes due to the
incomplete development of the pars plana and the smaller size of the ocular
structures. Not until 2 to 3 years of age does the eye reach approximately
90% of adult anatomic dimensions. Although alternative techniques have
been developed to perform vitreoretinal procedures in eyes beginning near 40
weeks gestational age, the surgical risks are considerable.
GENE TRANSFER AGENTS

Gene transfer can be achieved by several different methods. Ultimately, the
optimal technique will achieve the gene expression or modification profile
required for the specific application with the minimum of toxicity. Naked
DNA does not penetrate cell membranes readily; however, transient
expression with modest transduction efficiency can be achieved by a variety
of physicochemical techniques. The advantage of these techniques is there
are no size limitations to the DNA, there is no exposure to potentially
immunogenic virus capsid proteins, and it is possible to produce these vectors
in large quantities. Cationic lipids that interact with cell membranes can
enhance transfer efficiency (52,53). With many of these physicochemical
techniques, efficiency of transfer to many cell types and duration of gene
expression are quite limited. One promising approach is an RNA antisense
molecule that can bind and correct aberrant splicing (see ProQR, below,
Table 28-2).
TABLE 28-2 Gene therapy clinical trials for

retinal diseases using nucleic acids
IVT, intravitreal; #, not yet recruiting; NA, not available.
Viruses are used to take advantage of their ability to transfer exogenous

nucleic acids into host cells. Viral vectors have been considered ideally suited
for treatment of many diseases because of their superior gene transfer
efficiency and stability of gene expression. Using recombinant DNA
engineering techniques, viruses are designed so that their ability to replicate
is eliminated and their pathogenic properties are minimized. Different viruses
demonstrate distinctly different characteristics with regard to cargo capacity,
cell tropism, immune response, and stability of transgene expression.
Recombinant adenovirus is highly efficient at transducing RPE cells and
Müller cells (55,56). Onset of gene expression is rapid, occurring within 24 to
48 hours of vector administration. However, transduction efficiency with
adenovirus of retinal cell types such as photoreceptors is poor. In addition,
the duration of gene expression is limited by the lack of integration of the
transgene and the host immune response to the vector (57,58). For this
reason, adenoviruses are predominantly used now where immune response
may be desirable. Thus, selection of this vector years ago for use in a phase 1
clinical trial for retinoblastoma was appropriate (59). Although the phase 1
results were promising, those studies were not continued.
Retroviruses have long been available for use in gene therapy
applications. Many retroviral vectors appear to have limited application in
retinal gene therapy due to their inability to efficiently target nonreplicating,
terminally differentiated cells, such as those in the neural retina. Retroviral
vectors have prolonged expression in part, due to the fact that this vector
integrates into the host genome. Although this might be an advantage, it also
poses a risk of insertional mutagenesis. This risk was realized in a human
gene therapy trial for X-linked severe combined immune deficiency (SCID),
in which children whose disease had been “cured” by retroviral treatment
developed a leukemia-like syndrome (60).
There is one class of retrovirus, lentivirus, which shows favorable
somatic transduction profiles. The majority of the recombinant lentiviruses
that have been evaluated are derived from the human immunodeficiency virus
(HIV). Multiple safeguards are in place to minimize the risk of producing a
wild-type (replication competent) virus in development of recombinant
lentiviral vectors. Recombinant lentiviral vectors have also been generated
using nonhuman parental strains, such as those derived from the cow, cat, and
horse. A phase I/IIa study using an equine-derived lentiviral vector to deliver
the ABCA4 cDNA for Stargardt disease is in process (61). A phase I/IIa study
using a similar vector for treatment of the RP in Usher syndrome (type 1B;
MYO7A gene) is also in process (Figure 28-2; Table 28-1).
In recent years, there has been a large increase in interest in the
development of adeno-associated virus (AAV) as a vector for ophthalmic
gene therapy. AAV has been used in approximately 3 dozen different ocular
protocols to date at more than 30 different clinical trial sites in multiple
different continents (Table 28-1). More than 850 different people have had an
ocular injection of a recombinant virus and that number is anticipated to
exceed 1,150 in mid-2019. The majority of these have received recombinant
AAV (3 trials have employed lentivirus). The recombinant viruses have been
delivered by subretinal or intravitreal injection and the safety record has been
excellent. While the majority of AAV trials to date have been early-mid
phase studies, there are currently six different phase 3 (registrational) trials
active for two different disease targets—choroideremia and Leber hereditary
optic neuropathy (Leber hereditary optic neuropathy [LHON]; Figure 28-2;
Table 28-1) (25).
AAV is a parvovirus, which has minimal pathogenic effects in humans.
Although wild-type AAV can integrate into genomic DNA, integration has
not been reported after delivery of recombinant AAV to the retina. Instead,
the AAV-delivered transgenes appear to remain as episomes in the host
nucleus and provide stable gene expression in the retinal cells over time,
because they do not divide. AAV has expression profiles that are particularly
favorable for applications to retinal disease. Dozens of different AAV
serotypes have been identified or engineered, and many of these transduce
photoreceptor cells with extremely high efficiency (62–67). Most of the
different AAV serotypes have a strong tropism for the RPE. When injected
into the vitreous, some AAV vectors are capable of transducing retinal
ganglion cells (68). Generation of novel AAV serotypes through evolutionary
design or through application of in silico approaches has further expanded the
toolkit (69). Through evolutionary design, AAVs have been developed,
which are trophic to bipolar cells or to Müller cells or which have an
enhanced ability to penetrate the retina from the vitreal aspect (70–73).
Although AAV does induce an immune response, the response is
generally relatively mild even after injection into the vitreous (46,74). When
inflammatory responses are seen, these are typically dose dependent and
found after intravitreal (not subretinal) injection. As further evidence of the
safety profile of AAV, it is possible to safely readminister AAV serotype 2
(AAV2.hRPE65v2 = Luxturna) to the contralateral eye in humans with
RPE65 deficiency previously treated with subretinal administration and
thereby to improve visual responses (48,75–77). It has not yet been possible
to elicit a therapeutic response when AAV is readministered in other organ
systems. This phenomenon is a unique attribute of the retina as a target organ.
The most significant limitation to the use of AAV is the small cargo
capacity of the vector. Whereas lentiviral vectors can accommodate
transgenes up to 7 kb in size, AAV can package a maximum of 4.8 kb
(30,36). Use of dual AAV vectors may be able to overcome this limitation to
some extent (26,34).
Current Path for Testing Gene Therapy in Clinical

Trials
The prerequisite for developing a therapy specific to an inherited disorder is
to identify the disease-causing gene (Figure 28-2). To date, mutations in at
least 269 different genes have been found to result in visual deficits in
humans. Many of these also result in vision deficits in animal models. The
translational path typically requires laboratory evidence that delivery of a
particular reagent results in improved retinal or visual function and/or
prolongs the health of retinal cells. The test(s) used depend on the species,
disease pathophysiology/biochemistry, and the amount of degeneration at the
time of intervention. In the case of puppies born blind due to RPE65
deficiency, proof-of-concept studies included comparison of before and after
results of noninvasive tests such as electroretinography, pupillometry, ability
to navigate an obstacle course, and measures of amplitude and frequency of
nystagmus (27,40,78–80). The results were further related to evidence of
production of RPE65 protein at the site of injection and biochemical evidence
demonstrating that this protein was functional and allowed it to cleave the
substrate, retinal esters, to the retinoid 11-cis-retinal necessary for completion
of the visual cycle. Effects of route of administration (intravitreal vs.
subretinal), age at intervention, and dose were also evaluated. A benefit of
studies involving the retina is that this tissue can be assessed noninvasively
and over time by ophthalmoscopic examination in order to assess safety. In
most species, it is also possible to assess details of the retinal structure over
time noninvasively through fundus photography, autofluorescence
measurements, and optical coherence tomography (OCT). Additional safety
data were obtained from these animals included both ocular and systemic
histopathology, results of local and systemic humoral and cellular
immunologic responses, and biodistribution (i.e., whether or not the reagent
escaped the eye and spread to other organ systems).
In studies in which the model organism is a mouse, tests are used that are
relevant to this species. Standard electroretinography and pupillometry can be
used. However, navigational tasks typically require these normally nocturnal
animals to be placed in an environment where they are forced to use visual
cues to escape (e.g., a water bath with a visually detectable escape platform
(41)). Visual acuity can be measured in a mouse using optokinetic
nystagmus, analogous to the testing employed in pre/nonverbal humans.
Similar outcome measures as those used in dogs and mice have been
applied to many other animal models of blinding disease, including zebrafish,
cats, rats, chicks, pigs, and monkeys. There are some diseases where there is
no available animal model or the animal model may lack a phenotype.
Alternatively, its phenotype may be irrelevant to the human condition.
Examples of this latter situation include animal models of the RP found in
Usher syndrome. While there are a number of spontaneous mouse mutants of
the various forms of Usher syndrome, these models manifest deafness but a
very mild, if any, retinal phenotype. Other examples include the X-linked
retinal degeneration, choroideremia. In this disease, affected males are
typically legally blind in young adulthood. Studies in mice are challenging
because (unlike the human disease) lack of function of the choroideremia
gene is lethal to mouse embryos. Thus, conditional choroideremia mouse
mutants have been generated in which only heterozygous female mice
manifest retinal disease (42).
In conditions in which animal models are lacking or of questionable
relevance, it is possible to submit efficacy data for cell-based models prior to
moving to safety studies (see below). This is a strategy that was used to move
forward with a clinical trial for choroideremia. There, the lead clinical
reagent was used to demonstrate amelioration of the biochemical deficit (Rab
rab geranylgeranyl transfer activity) and of protein trafficking deficits in
human patient-derived induced pluripotent cell (iPSC) models (81). The
ability to differentiate iPSC cells into retinal models that can be used to test
gene therapy approaches (82) provides a critical resource for preclinical
investigations of the many diseases where animal models are not available.
After generating proof-of-concept data, preclinical safety and toxicity
studies are typically conducted in unaffected animals under “Good
Laboratory Practices” (GLP) conditions using reagents prepared under the
same fastidious conditions as the ultimate reagent to be tested in people. The
study is monitored closely and individuals who do not have competing
interests report data. Typically, dosing includes a dose that is at least five
times higher than the planned highest dose to be used in humans. Local and
systemic effects are evaluated and the results help guide the ultimate clinical
trial.
Investigators must apply to the appropriate institutional (Institutional
Review Board, Institutional Bioethics Committee, etc.) and federal bodies,
for example, the FDA or EMA in order to comply with guidelines to conduct
a gene therapy clinical trial. In this process, special attention is given to the
ethical justification rationale for enrolling children, due to their status as
“vulnerable subjects.”
Prior to the first retinal gene therapy studies, gene therapy trials for
nonlethal diseases had only enrolled adults. Enrollment of children in a gene
therapy study for a nonlethal disease was first discussed at the NIH
Recombinant DNA Advisory Committee (RAC) in December 2005. The
RAC approved enrollment of children based on the assessment of reasonable
prospect of benefit and the diminishing chance of improvement with
advancing age (i.e., favorable risk: benefit ratio). This marked the first
approved enrollment of children for gene therapy studies of a nonlethal
disease (RPE65 deficiency).
Once approvals have been granted by the institutional and federal
agencies and funding is in hand, the trial can be initiated. Funding is not a
trivial issue even with a limited study with few patients—a phase 1 trial can
cost several million dollars. Phase 3 (registrational) trials easily cost an order
of magnitude more than phase 1 trials. In the early stages of a clinical
investigation, adults are typically enrolled prior to children, even in early-
onset diseases, in order to de-risk the intervention first in adults (“order of
preference”). A Data Safety Monitoring Board (DSMB) is established in
order to evaluate safety signals and to determine whether it is appropriate to
progress to higher doses. In the United States, it is mandated that subjects in
gene therapy trials to be followed for 10 years (recently reduced from 15
years) after administration of the test product.
CURRENT STATUS OF GENE

THERAPY FOR INHERITED
PEDIATRIC-ONSET RETINAL
DISEASES
Studies Using Viral Vectors
RPE65 Deficiency
The first gene therapy clinical trials targeting LCA due to RPE65 mutations
were initiated more than 11 years ago (2007; Table 28-1). LCA, one of the
most severe forms of RP because it is manifest in infancy or early childhood,
is rare and can be caused by mutations in any of at least 20 different genes.
The severely impaired vision early in life slowly deteriorates due to
progressive degeneration of retinal photoreceptors (Figure 28-3A and B). A
lack of RPE65 protein impairs the visual transduction (retinoid) cycle,
making it nearly impossible for rod photoreceptors to respond to light (83).
The first proof-of-concept that gene augmentation therapy could ameliorate
this condition was gathered in a spontaneous RPE65 mutant dog, the Swedish
Briard. The data demonstrating that blindness in this dog model could be
reversed by gene therapy (40,78,79), supported near-simultaneous initiation
of three different dose escalation clinical trials testing AAV2-mediated gene
augmentation therapy for LCA due to RPE65 mutations. Additional groups
initiated RPE65 clinical trials thereafter, and the current tally of RPE65 gene
therapy clinical trials is 9 (Table 28-1).
FIGURE 28-3 En-face imaging in retinal degenerations.
A:Fundus photography and photoreceptor thickness
topography in RPE65-LCA. Fundus image is rather
normal, but there is severe photoreceptor layer (ONL)
thinning with a central to superior region of better
preservation mapped in shades of green. B:Macular
changes in association with an overt pigmentary
retinopathy with areas of preservation in peripapillary
retina and nasal midperiphery in RDH12-IRD.
C:Unimpressive macular appearance on fundus exam of a
young patient with ABCA4-STGD but obvious fast
progressing maculopathy on near-infrared fundus
autofluorescence (NIR-FAF).
The initial three studies and most of the subsequent studies used an AAV
serotype 2 vector (AAV2) delivering the wild-type human RPE65 cDNA
subretinally to the RPE in one eye. One later trial used AAV4 (84) and
another is using AAV5 (Table 28-1). There are numerous other variables that
differ between the groups, including inclusion criteria, type of promoter,
dose, location of injection, immunomodulation protocols, and outcome
measures. Reports from all of the trials revealed a high degree of safety
(43,77,84–86). Safety seemed to relate predominantly to the surgical
procedure and most of the adverse events were reversible. The second subject
in one of the early trials developed a macular hole after injection, and surgical
procedures were modified thereafter so as to minimize the possibility of that
outcome. (Nevertheless, that individual showed strong gains in the retinal
function and vision from his treated eye.) (54) The outcome measures
reported to be improved in one or more of the early trials included increase in
light sensitivity, improved visual acuity and visual fields, improved pupillary
light reflex (PLR), and improved mobility (47,50,87). More than 150
different eyes have now been injected in various clinical trials with AAV
carrying the wild-type RPE65 cDNA (Table 28-1), and there is an excellent
safety record. There has been no evidence of vertical transmission—an initial
concern of the U.S. FDA. Several of the subjects conceived and delivered
children who are healthy and thriving.
Further, there were generally large improvements in retinal/visual
function (43,76,85,88). Two of the phase 1 studies reported a decline in
retinal and visual function (in 3 of the 16 patients injected) after the 3-year
timepoint. However, in one of these studies, the level of retinal function
continued to be substantially higher beyond the 3-year timepoint than at
baseline (88). There was also a concern that the degenerative process
continued although, again, improvements in visual function persisted. The
second group that reported decline in response had used a cell-specific (RPE)
promoter to drive expression of the hRPE65 cDNA. That team reported a
decrease in sensitivity after 6 to 12 months in 6 of 12 participants (43).
However, the subjects had not responded as dramatically in that study as in
other studies. It is possible that the lower response was due to use of the
weaker promoter or to other variables that differed from the other studies.
One variable that differed considerably from the other studies was the volume
injected under the retina during AAV delivery. In any case, it is impossible to
make a direct comparison of the results between the various phase 1 dose
escalation studies because there were so many variables that differed between
the studies, including vector design and methods of purification, whether or
not surfactant was used to minimize adherence of vector to inert surfaces,
whether systemic steroids were administered, the exact dosing, surgical
delivery procedures, and outcome measures used to evaluate treatment effect.
After consulting with the FDA about plans for developing a phase 3
clinical trial, the team at Children’s Hospital of Philadelphia (CHOP)
designed a follow-on study for the phase 1 dose escalation trial that aimed to
deliver the gene therapy reagent to the contralateral retina. The FDA had
stated that if the reagent was approved, doctors would attempt to deliver it to
both eyes, and so the team should test safety and efficacy of both eyes being
treated. The subjects welcomed this guidance, as they had been requesting
(unbeknownst to the FDA) injection of their second eyes.
The contralateral eye readministration studies were preceded by large
animal studies aiming to be sure that this would be safe. The concern was that
an immune response to the vector delivered in the first eye could impair the
outcome in the second eye. An immune response could also potentially block
binding of the AAV and prevent improvement of the second eye. The large
animal studies did indeed provide reassuring safety and efficacy data (75).
Individuals who had enrolled in the phase 1 dose escalation study were
invited to participate in the contralateral eye “follow-on” study, in some cases
years after the injection in their first eye. The follow-on study data
demonstrated both safety and robust improvements in retinal and visual
function (76,89). The majority of the subjects gained function as assessed by
multiple outcome measures, including light sensitivity, PLR, visual fields,
and ability to navigate an obstacle course under low light conditions. The
results also mark the first successful readministration of a gene therapy for a
genetic disease. A phase III (registration) trial was then implemented (Spark
Therapeutics, Sponsor) in which AAV-hRPE65v2 vector was delivered by
bilateral subretinal delivery (48,76) (Tables 28-1 and 28-2). This was a
randomized, multicenter, controlled trial and the primary endpoint was
devised to measure a “clinically meaningful” difference in visual behavior.
This test was based on the ability to ambulate through a mobility course
illuminated at set light levels—the “multi-luminance mobility test (MLMT).”
Subjects, who were 4 years of age or older, were randomized 2:1 to
intervention versus no treatment. The individuals who did not receive
treatment were followed over the course of a year and then crossed over to
the intervention group. The intervention group received bilateral injections of
the AAV-hRPE65v2 within a 2-week interval. The primary endpoint was
improved navigation on the MLMT. Secondary endpoints included full-field
light sensitivity, visual fields, and visual acuity.
The phase 3 trial met its primary and secondary endpoints (77). An FDA
Advisory Committee voted unanimously to approve AAV2-hRPE65v2
(voretigene neparvovec-rzyl; Luxturna) in October 2017, and Luxturna was
formally approved as a drug by the FDA on December 19, 2017. This is the
first gene therapy reagent targeting a genetic disease to be approved drug
status by the U.S. FDA. It is also the first gene therapy for a retinal disease to
be approved worldwide. A surprise was that the drug was approved for
individuals age 1 year of age and older (even though the clinical trials had not
enrolled individuals that young).
A challenge for Spark Therapeutics was to establish Centers of
Excellence where assurance could be obtained not only about the eligibility
of each patient (i.e., presence of bi-allelic RPE65 disease, presence of
sufficient retinal cells, etc.) but also that the accuracy of the subretinal
injection (Figure 28-4) could be assured. The first three centers reported
near-simultaneous injections on March 20, 2018. Recently, the Committee
for Medicinal Products for Human Use (CHMP) at the EMA, the European
equivalent to the U.S. FDA, recommended approval of Luxturna. The
European Commission authorized EU-wide marketing on November 23,
2018. Decisions about price and reimbursement will take place in each
European country. Availability will be further expanded through Novartis
(90).
FIGURE 28-4 Still photos from subretinal injection of

Luxturna in a patient with LCA due to bi-allelic RPE65
mutations. The injection results in a localized retinal
detachment that resolves within a day. (A) Prior to
injection; (B) during injection; and (C) cannula withdrawn
from bleb. Optic disk is visible at one of the borders of the
bleb. (Surgeon, A.M. Maguire, MD.)
Choroideremia
Choroideremia (CHM) is a slowly progressive, XL recessive retinal
degeneration. CHM gene therapy studies were first initiated at University of
Oxford in 2013, and now there are six different CHM trials (enrolling >75
adult male patients) in progress (Tables 28-1 and 28-2). The initial goal in
CHM gene therapy is to preserve central vision (and to halt further
progression of the disease). This is because as the disease progresses, few
viable (resuscitatable) cells remain in the periphery (Figures 28-5 and 28-6).
Because of the slow progression of this disease, it may take several years to
obtain a complete efficacy assessment. The reports to date from the Oxford
group indicate that there is a high degree of safety associated with the
subretinal injection procedure targeting the fovea and that visual acuity can
improve (and the improvement can persist) in a subset of participants (91,92).
An earlier report from another Center in Alberta, Canada, using the same
AAV2.REP1 vector as a center of a NightStarX-sponsored trial, showed no
effect of intervention on the rate of decline of functional RPE in the treated
eye versus the untreated eye after 2 years of follow-up (93). A Spark
Therapeutics–sponsored clinical trial with centers at the University of
Pennsylvania and Harvard University as well as NightStarX-sponsored trials
with centers in Miami, Oxford, and Tubingen have confirmed this
observation (93–96). As of November of 2018, adverse events consisting of
permanent changes in the outer retina after the subfoveal injections have been
reported by both the Nightstar Therapeutics–sponsored trials (Alberta = 1/6
treated eyes, Miami = 2/6, Oxford = 2/12 eyes, Tubingen = 1/6 eyes) and a
center of the Spark Therapeutics trial in Philadelphia (1/9 eyes) (93–96). Six-
month data were reported by Aleman et al. for nine subjects injected
subretinally with a similar vector showing no significant differences in visual
acuity or retinal sensitivity in the treated versus the untreated eye and
individual vulnerability to subfoveal injection (97). Longer-term follow-up of
these studies is pending. Roche is planning a new clinical trial for
choroideremia using intravitreal injection of an AAV7m8, generated by
evolutionary design (Table 28-1). Biogen is planning a new trial using
subretinal injection of AAV8. One question facing the CHM investigators is
whether it may be more effective to treat the peripheral retina of younger
individuals affected with CHM.
FIGURE 28-5 Colocalized structure and function
correlations in choroideremia. Images are 9-mm long, SD-
OCT cross sections along the horizontal meridian through
the fovea in patients with the earliest central abnormalities
(top 2 panels) compared with patients (bottom 2 panels)
with later stage abnormalities. Nuclear layers are labeled.
Bars above the scans show psychophysically determined
rod (blue bar: dark-adapted, 500-nm stimulus) and cone
(gray bar: light-adapted, white stimulus) sensitivities.
Lines above bars define lower limit (mean − 2 standard
deviations) of sensitivity for the dark-adapted (dashed
lines) and light-adapted (dotted lines) conditions in normal
subjects. Calibration bar to the bottom left.
FIGURE 28-6 Choroideremia as an example of earliest

structural change and new potential for structural
dissociation at the level of the photoreceptor outer segment
(POS). A: Color fundus photography (left), near-infrared
reflectance (NIR-REF) imaging (middle), and horizontal
9-mm spectral-domain optical coherence tomography (SD
OCT) cross sections through the fovea in two young
patients with CHM exemplifying earliest abnormalities.
Vertical arrows on SD-OCT images point to locations
outside of which increased back-scattering from retinal
pigment epithelium (RPE) demelanization can be
appreciated. B: Area of apparent normal pigmentation of
the fundus by NIR-REF plotted against age (left). An
exponential decay function (dark gray trace) describes the
data. C: The ellipsoid zone (EZ) lateral extent plotted as a
function of the lateral extent of RPE with apparently
preserved melanization. D: The outer nuclear layer (ONL)
thickness as a function of EZ to RPE distance. Values are
specified as a fraction of the mean normal value for each
retinal location. Dashed lines are the lower limit (normal
mean − 2 standard deviations) for each parameter.
Achromatopsia
This rare condition causes extreme photophobia, poor visual acuity, and poor
color vision. Four different achromatopsia (ACHM) gene therapy studies
have been initiated: two targeting the CNGB3 form of the disease and two
targeting the CNGA3 form (Table 28-1). These studies are using
photoreceptor-specific promoters and AAVs (AAV8, AAV2tYF) that
transduce photoreceptors more efficiently than AAV2. Preliminary data have
been published through the 3-month timepoint for the first three subjects in
the RD-CURE CNGA3 trial (98). These subjects had received 1 × 1010 vg
AAV8.CNGA3 subretinally. None of them showed clinically evident ocular
inflammation; however, one patient had asymptomatic hyperreflective spots
visualized by OCT in the treated retina peaking 2 weeks after surgery. These
resolved with systemic steroid treatment.
XL Retinoschisis
XL retinoschisis results in splits (schisis) between the different layers of the
retina. When these splits occur in the macula, they can impair visual acuity.
The condition is caused by a lack of a secreted protein (retinoschisin). Two
different trials enrolled a total of 51 affected males (Table 28-1). Both studies
used intravitreal injection of AAV delivering the RS1 gene. Intravitreal
delivery may be effective in this disease due to the diffusability of the
secreted protein through the retina. The intravitreal route of delivery also
minimizes the possibility that an injection itself might lead to an additional
split in these vulnerable retinas. Results from one of the trials, the dose
escalation phase ½ trial run at NEI/NIH, were recently reported. There was
dose-related inflammation and immune response, some of which decreased
visual function, but this ultimately returned to baseline. Retinal cavities
closed transiently in one participant in the high-dose (1e11 vg) group
although there was moderate inflammation and no improvement in visual
acuity or ERGs were noted (99).
Retinitis Pigmentosa
Six different forms of RP are targets in clinical trials using different AAVs,
delivery routes, and strategies (Table 28-1). MERTK deficiency, a disease
manifest in RPE cells, was the first to be addressed. The initial results using
subretinal injection show a high degree of safety. However, it was not clear
whether there were long-term improvements (100). A second RP study
delivers optogenetic therapy to retinas of patients with end-stage
degeneration. The goal is to render other cells in the degenerated retinas
(ganglion cells) light sensitive. Another optogenetic therapy trial recently
enrolled its first subject using both delivery of a modified channelopsin as
well as use of a device (https://www.gensight-
biologics.com/2018/10/26/gensight-biologics-enrolls-first-subject-in-first-in-
man-pioneer-phase-i-ii-clinical-trial-of-gs030-combining-gene-therapy-and-
optogenetics-for-the-treatment-of-retinitis-pigmentosa/). A third RP trial
targets the retinal component of Usher syndrome due to MYO7A mutations.
The MYO7A cDNA is too large to fit within the AAV confines and so a
lentiviral (EIAV) vector was used. The fourth form of RP to be targeted is X-
linked and due to mutations in RPGR. Three different groups are conducting
RPGR clinical trials that differ in AAV serotype (AAV2-tyrosine modified,
AAV5, or AAV2), promoter (hRK, GRK1, or CBA), and age of subjects.
Two other rare forms of RP are also under investigation in phase I/I studies: a
form due to PDE6β-deficiency and one due to RLBP1 (also known as
CPK80) deficiency (Table 28-1).
Stargardt Disease
Mutations in the ABCA4 gene, expressed in photoreceptors, result in
autosomal recessive Stargardt disease (Figures 28-3C and 28-7). ABCA4
mutations can result in a spectrum of disease, including cone–rod dystrophy,
fundus flavimaculatus, RP, and possibly AMD. Similar to the MY07A trial,
the Stargardt trial in progress uses a lentiviral vector in order to accommodate
the large size of the ABCA4 cDNA (Table 28-1).
Leber Hereditary Optic Neuropathy

Inherited mitochondrial mutations can cause LHON, a disease in which there
is sudden onset of blindness due to defective function of ganglion cells. There
are several trials in progress targeting ND4 mitochondrial mutations (Table
28-1). A mitochondrial targeting signal is incorporated in order to target the
ND4 gene to the mitochondria. In each of the trials, an AAV2 vector,
delivered intravitreally, is used to target the ganglion cells (68). A phase III
study for LHON is underway and includes an arm aiming to reverse vision
loss in the initially treated eye (REVERSE) as well as an arm (RESCUE)
aiming to prevent vision loss in the second eye after symptoms initially
manifest in the first eye. Early reports suggested that there was significant
improvement in visual acuity of treated eyes at week 72 and continuous
bilateral improvement of both visual acuity and contrast sensitivity
(https://www.gensight-biologics.com/2018/10/18/gensight-biologics-reports-
positive-72-week-data-from-reverse-phase-iii-clinical-trial-of-gs010-for-the-
treatment-of-leber-hereditary-optic-neuropathy-lhon/).
Studies Using Nonviral Vectors
CEP290 (ProQR)
On September 5, 2018, ProQR Therapeutics announced positive interim
results from a phase I/II clinical trial of an antisense RNA, QR-110, in
patients heterozygous or homozygous for the common p.Cys998X mutation
found in patients with CEP290 mutations (also known as LCA10; Table 28-
1) and published 3-month interim results shortly thereafter (101). The plan
was to administer repeated doses (a total of 4 doses over 1 year) of low,
medium, and high titers of this reagent since the expectation was that it
would have a limited half-life.
The total number of individuals treated in the phase 1/2 trial was
ultimately 10, with an interim report presenting data from the first 8 patients,
including one “super responder” who improved from LP to 20/400 vision.
Subjects ranged in age from 8 to 44 years of age at the time of treatment. All
eight subjects were followed through 3 months after receiving low and
medium doses and 4 of the patients had been followed at least 6 months.
The doses were administered through unilateral intravitreal injection of
an RNA antisense oligonucleotide. The investigators reported improvement
in visual function observed starting at 2 months after treatment, which was
maximal by 3 months. Visual function was assessed through best-corrected
visual acuity (BCVA), which showed a mean improvement of −0.67
logMAR, with 62.5% of individuals showing an improvement of more than
−0.3 logMAR compared to baseline (i.e., more than 15 letters or 3 lines on
the eye chart). There were negligible changes in the contralateral eye (0.02
logMAR, one letter).
There were improvements in additional outcome measures, which had
been previously tested in LCA10 (and other) subjects (102). These included
red and blue light sensitivity as judged by full-field stimulus testing. Those
were −0.74 and −0.91 log cd/m2, respectively. Further, the individuals
improved several light levels in performing the mobility test. There was
comparable activity observed in the first two dose levels, with improvements
in visual acuity and ability to navigate a mobility course at the 3-month
timepoint. Adverse events included cataracts and cystoid macular edema.
This is the first human study targeting a disease originating in
photoreceptors to evaluate the safety and efficacy of an RNA-based
therapeutic and we must await publication of the longer term results to fully
assess their impact. However, because of the positive interim results, ProQR
is planning a pivotal phase 2/3 trial.
Pipeline for Gene Therapy for Pediatric Retinal

Diseases
Approval of the first gene therapy product in the United States and
progression of several studies to phase 3 clinical trials have encouraged both
small and large pharma to enter the retinal gene therapy field. Many have
recognized that progress in retinal disease targets may be a stepping stone to
developing gene-based treatments for more common diseases. So far, there
are gene therapy clinical trials in different phases for more than a dozen
different genetic targets (Figure 28-1; Tables 28-1 and 28-2). Further, there
is a robust pipeline that continues to enlarge which will bring additional
genetic forms of IRD to clinical trial in the near future, including gene defects
associated with LCA (LCA5, RDH12 ([Figures 28-3B, 28-8, and 28-9]),
CRB1, AIPL1, GUCY2D), RP (rhodopsin, PDE6α, SPATA7, PRPF31),
syndromic IRDs (Bardet-Biedl syndrome and Usher syndrome), macular
dystrophies and cone dystrophies (RDS/PRPH2, blue cone monochromatism,
achromatopsia), and chorioretinal dystrophies (choroideremia and gyrate
atrophy). Several groups have initiated informal as well as formal natural
history studies, prerequisite for selecting outcome measures to be used in
these trials (Table 28-3), for example, references (103–106).
TABLE 28-3 Natural history studies prerequisite

to pediatric onset gene therapy clinical trials
The progress in genome editing using CRISPR-Cas systems further expands
the opportunity to intervene with specific genetic diseases (Figure 28-1).
There are no human clinical trials in progress using CRISPR-Cas9 for retinal
disease; however, the first proof-of-concept studies have been published. In
vivo CRISPR-Cas9 corrections have been demonstrated in rat models of arRP
(Mertk, Pde6β) and of AdRP (rhodopsin) and a mouse model of ARRP
(Pde6β deficiency) (107–110). One of the challenges of CRISPR-Cas9 is that
mutations are often scattered across genes and so one reagent that may be
useful in one family may not be useful in another. There is a mutation in
CEP290 (the IVS26 mutation), which causes a high percentage of LCA, and
this is a promising target for CRISPR-Cas9 engineering (111). There are
many safety and efficiency concerns that must be satisfied before this
approach can be tested.
There is also great interest in the potential to develop gene therapy
platforms capable of ameliorating retinal disease regardless of the genetic
cause of the condition. Multiple different strategies that have been described,
including delivery of neurotrophic factors (RdCVF, XIAP, etc.), genes which
support metabolism of these cells (NRF2) and genes encoding molecules with
antioxidant effect (23,112–116). The ability to use one reagent to prolong the
life and function of cells regardless of genetic defect would be very useful.
Such reagents could potentially be used to ameliorate disease caused by
acquired conditions as well.
Current Challenges for Pediatric Retinal Gene Therapy

There are a number of challenges at present facing development of pediatric
retinal gene therapy. For many diseases, there is a lack of relevant animal
models (see above). Thanks to developments in iPSC technology, it is now
possible to generate models in a dish. Labs around the world are now able to
develop cell models of RPE cells that reflect all of the morphologic and
functional features of RPE in vivo. It has taken longer to develop a similar
technology for generating mature photoreceptors; however, there are now
several groups that have achieved this goal. Availability of these models and
transfer of the know-how for these new technologies will expedite
development of appropriate therapies.
The field continues to struggle with accessibility of genotyping
procedures. While these may be available at some tertiary academic centers
and CLIA-approved testing is offered by a number of companies, many
children are left behind with respect to genetic testing. A key impediment is
economic—gene screening in the United States is sometimes not covered by
insurance companies based on the reasoning that results do not impact care
for diseases which have no treatment. Even when tests are covered, health
systems may not be able to commit the personnel, time, and resources needed
to go through complex processes involved in securing insurance approval;
testing requests may end in a denial and the end of the search for a molecular
cause of these diseases. One benefit of the emergence of social media over
the past decade is that families with specific inherited conditions are sharing
information and pressing for attention from health care providers and
insurers. Grass root organizations have also served as a resource for
physicians evaluating natural history and recruiting for clinical trials (Table
28-3).
Policies designed to protect children as vulnerable subjects may delay
enrollment of pediatric patients for a clinical trial although they might be the
ideal target population based on biologic considerations. One example is
choroideremia, where clinical trials have targeted adults, who typically have
only a small island of central retina that is viable. In adults with
choroideremia, there is little that can be done to test either the ability to halt
progression of the disease or to reverse the deficits. However, if it were
possible/ethical to enroll children, it would be possible to treat the rapidly
degenerating peripheral retina, preserving useful vision as well as obtaining a
rapid readout of treatment efficacy. Enrollment of children could eliminate
the need for subfoveal injections, also—a procedure that carries some risk
(see above).
OUTCOME MEASURES,
THERAPEUTIC WINDOWS, AND NEW
TARGETS
Outcome measure research is at the center of clinical trial design for IRDs.
The heterogeneous nature of the primary abnormalities at the molecular and
histologic level in these diseases, the diversity of the potential cellular targets
for treatments, and the wide spectrum of disease severity present an equally
heterogeneous structural and functional landscape to explore. The ideal
outcome measure would be easy to implement and deliver, sensitive, specific,
reproducible and able to provide a read out in the short term.
Outcome measures of safety and efficacy must be tailored not only to the
specific disease but also to the disease stage and age of the patient. Whereas
early-onset retinal degenerations (EORDs) constitute attractive platforms
where the risk/benefit ratio may be stacked in favor of visual gain,
implementing outcome measures in the pediatric population is far from
simple. For some EORDs, there is evidence that there will be no option but to
treat at a younger age and regulatory bodies will be expected to require
outcome measures that are similar or equivalent to those used in earlier gene
therapy clinical trials in the adult population, adding complexity to clinical
trial design. Whereas evaluating vision in children older than about 8 to 10
years of age may be possible using the same outcome measures used in
adults, the situation is very different for younger children (103). Irrespective
of the age, an effort should always be made to obtain measures of vision in
young pediatric patients. Techniques that rely on preferential looking may be
used to obtain measures of visual acuity, for example, with the use of Teller
acuity cards. Later in infancy and childhood, a transition can be made to age-
appropriate measures of acuity. Full-field stimulus or sensitivity testing
(FST) techniques were developed in preparation for the earliest gene therapy
trials in an effort to overcome the difficulties in determining visual
sensitivities in patients with severe vision loss that exceeds the dynamic
range of conventional perimetry and in whom poor fixation and nystagmus is
expected to complicate the interpretation of the perimetric tests, often
preventing investigators from using visual fields altogether (117,118).
Although FST technology has been successfully used in a number of gene
therapy trials, there is paucity of information on its reproducibility
particularly in pediatric populations (93,102,118–120). Commercially
available instruments and software allow for customization of FST testing
protocols to accommodate testing of children (Figure 28-8). Protocols should
be as brief as feasibly possible with short interstimulus intervals, limited
number of repeats and stimulation conditions, and starting stimulus
luminances that are appropriate to the expected sensitivity losses of the
specific condition and/or patient (104). Binocular testing and participation of
the tester and family with encouragement during testing may be considered.
A reassuring recent report suggests that these general guidelines can yield
results that are as reproducible in children as in adult populations (103).
An added advantage of this form of testing is the possibility of
determining photoreceptor mediation of vision and the depth of the visual
deficit, which are important diagnostic features of these disorders and may be
part of eligibility criteria for treatment trials. Certain molecular forms of
IRDs, for example, are expected to show mostly residual cone-mediated
vision (e.g., CEP290, RPE65), whereas others (e.g., CRB1, RDH12, AIPL1,
GUCY2D) may show substantial rod vision (Figure 28-3)
(102,103,121–125). The information obtained can provide a more specific
clinical diagnosis, guide and confirm molecular results, and serve as
measures of safety and efficacy in clinical trials.
Complementary objective measures of vision in pediatric IRDs are
desirable giving the intrinsic uncertainty of measuring vision
psychophysically in this group of patients. Although there is a role for
electrophysiologic tests of vision in early-onset cone
dystrophies/dysfunctions, gold standard, diagnostic tests for retina-wide rod
and cone retinal degenerations, such as the electroretinogram (ERG), may not
be informative as outcome measures due to the limited signal available for
detection as a result of early-onset abnormalities. The quantitation of the PLR
by pupillometry is a promising alternative already proven effective in
confirming treatment efficacy during early RPE65-LCA trials
(47,49,126,127). There is scarcity, however, of information on the
reproducibility of the measures, particularly in children, which will be needed
to document subtle changes in vision following the interventions. Like FST,
the transient PRL (TPLR) provides objective information about the depth of
the sensitivity losses and photoreceptor mediation of vision (Figure 28-9)
(103,126). Evidence that TPLR thresholds and photoreceptor mediation of
the TPLR responses correspond well with psychophysically determined
measures of vision by FST has been recently provided supporting the role of
this testing modality in the retinal degeneration clinic and research.
The retinal appearance in pediatric IRDs ranges from deceivingly benign
to the presence of an unmistakable pigmentary retinopathy (Figure 28-3).
Certain findings in isolation or in the context of syndromes may serve as
diagnostic hints of certain molecular subtypes. Examples include the
refractive state of the eye (CRB1) (122), the presence of atrophic
maculopathies and macular pseudo-colobomas (IDH3A, RDH12, AIPL1,
MMACHC, NMNAT1, ABCA4, CRX, DHX38, PRDM13, CNNM4, CNGA3,
PDE6C, ATF) (103,128–138) or true
colobomas/microphthalmia/anophthalmia (SOX2, OTX2, PAX6, STRA6,
ALDH1A3, RARB, VSX2, RAX, FOXE3, BMP4, BMP7, GDF3, GDF6,
ABCB6, ATOH7, C12orf57, TENM3,ODZ3, VAX1, SALL2, YAP1, CRB1)
(139,140), optic nerve drusen (RDH12, MFRP, CRB1) (140–142), and of
early and peculiar pigmentary changes (NR2E3, NRL, CRB1) (143–146).
Once established, however, these nonpathognomonic qualitative signs are of
limited use as outcome measures in clinical trials.
En-face retinal imaging with scanning laser ophthalmoscopy and cross-
sectional retinal imaging of the structural detail of the retina with spectral
domain optical coherence tomography (SD-OCT) have offered new
quantitative possibilities to the study of retinal degenerations. Measures of
photoreceptor-specific vision colocalized to the underlying retinal structure
imaged with SD-OCT constitutes a powerful, yet not widely used tool to
explore these conditions (Figure 28-5), and a proven outcome measure of
efficacy of IRDs in RPE65-LCA (49). The premise for gene therapy for outer
retinal degenerations is the demonstration of treatable photoreceptors, which
can be identified with SD-OCT (147). Confirmation of vision loss in excess
of that expected for the photoreceptor loss has been deemed the ideal setting
to treat with gene augmentation (102,147). In such situation, the predominant
dysfunction may be acutely improved after the correction of the underlying
defect such as in RPE65-LCA.
This scenario of structural–functional dissociation, however, is not the
most frequently encountered. The more common picture in IRDs is that of
pure photoreceptor degeneration in which there is proportionality between the
loss of photoreceptors and the associated vision loss (147). In this common
situation, outcomes that measure progression or modification of the natural
history of the disease after treatment are needed. Detection of fast progressing
windows of structural and functional changes that indicate where to intervene
may be desirable (Figures 28-6 and 28-7) (103,129). Careful quantitation of
the structural changes at the level of the outer retina, specifically at the level
of the photoreceptor outer segments (POSs), however, may provide a new
level of structural–functional situation that may be corrected with gene
therapy and that can provide acute evidence of the efficacy of the
interventions (102). That is, vision loss that is purely associated with POS
abnormalities may exist in the presence of a normal photoreceptor layer
(outer nuclear layer, ONL) thickness, providing a new setting of structural–
functional dissociation that may be improved after the physiology of the POS
is restored with treatment (102,104).
FIGURE 28-7 Short-term follow-up of an ABCA4-STGD
patient with SD-OCT demonstrates progression detectable
only by imaging. Shown are 7.6-mm horizontal cross
sections through the fovea at baseline and a year later.
Nuclear layers and outer retinal bands are labeled: GCL,
ganglion cell layer; INL, inner nuclear layer; ONL, outer
nuclear layer; ELM, external limiting membrane; IS/OS,
outer segment/inner segment interface band; IZ,
interdigitation of the photoreceptor outer segment with
apical RPE; RPE/BrM, RPE/Bruch’s membrane.
Horizontal arrow shows sequence of loss of the bands in
the outer retina: IZ →IS/OS → ELM → foveal ONL
thinning. Vertical arrows signal place interruption or loss
of the IS/OS that corresponds with the loss of the POS;
arrows are offset horizontally at baseline compared to a
year later due to the centripetal progression of the
abnormalities.
FIGURE 28-8 Full-field sensitivity (FST) testing in

RDH12-LCA. Left panel: FST sensitivity estimates to
dark-adapted spectral stimuli (blue, 467 nm; red, 637 nm)
in 18 patients (ages 4 to 17) with RDH12-LCA. Dashed
line is the lower limit (mean − 2 SD) of the sensitivity to
the short wavelength 467 nm stimulus in normal subjects.
There is measurable rod vision with an average 2 log units
of sensitivity loss. Right panel:Interocular difference of
the dark-adapted FST estimates for each stimulus
condition (blue, 467 nm; red, 637 nm). Diagonal line is
the equality line. Sensitivity estimates are remarkably
symmetric despite interocular asymmetries in visual acuity
and severe central retinal abnormalities.
FIGURE 28-9 Objective measures of vision in
LCA/EORDs; transient pupillary light reflex in RHD12.
A:Families of dark-adapted TPLR waveforms elicited with
brief (100 ms), blue (467 nm) stimuli delivered over an 8
log units of intensity range on the same stimulator unit
used for FST measurements. Traces plot the horizontal
pupil diameter as a function of time in a normal subject
compared to a representative patient with RDH12-LCA.
Overlapping thick red traces near TPLR threshold are
responses elicited by red (637 nm) stimuli scotopically
matched to the blue stimulus that supports rod mediation
in RDH12-LCA. Stimulus monitor shown as a vertical bar
at bottom left. B:FST sensitivity loss plotted against TPLR
sensitivity loss for each eye of patients who were able to
complete both tests (n = 18). Unfilled symbols = right eye;
gray symbols = left eyes. Diagonal line is the equality
line. TPLR threshold is rod mediated for all patients and
shows about one log unit less sensitivity loss than
psychophysically estimated thresholds.
Metabolic agents can dramatically, albeit transiently, improve vision across
the entire retina in RPE65-LCA independent of the topography of the residual
photoreceptors (88,148,149). To date, however, the most efficient and
frequently used method of delivery of long-lasting treatment effect is through
gene therapy subretinal injections. The photoreceptor topography determined
by SD-OCT in retinal degenerations is thus needed to determine eligibility
and in planning of the locations to be targeted by these subretinal injections
(147,150). Reports of macular thinning following initial subretinal injections
dictated caution and often avoidance of the foveal region in the initial phases
of the RPE65-LCA gene therapy trials (87). The natural history of some
forms of IRDs leave no choice of location to treat but the fovea, as this
important region often has the only residual tissue in later stage disease, such
as in choroideremia and forms of RP, or because the predominant
abnormalities are located within the central retina, such as in Stargardt
disease (STGD), cone–rod dystrophies, and cone dystrophies, such as
achromatopsia (151). Accumulated experience serves to outline the safety
profile of the subfoveal injections. There is transient (<6 months) shortening
of the foveal cone outer segments but for the most part no major disruptions
of the foveal anatomy in most cases (97). There have been reports of thinning
and macular hole formation in a number of patients after macular subretinal
injections (92–94,96,97,152,153). The risk factors that predispose to this
outcome are unknown, but data from the Spark Therapeutics–sponsored
choroideremia trial suggests that there can be acute loss (<48 hours) of foveal
photoreceptors postinjection pointing to a possible mechanical damage of the
POS by the fluid wave created by the subretinal injection. Abnormalities at
the level of the outer retina detected on SD-OCT in the CHM trials have been
interpreted as representing an inflammatory response, which can provide an
alternative mechanism.
FUTURE CHALLENGES
New Disease Targets

Presently, the potential disease targets for EORDs remain focused on
conditions with dissociations between the retinal structure and function.
Examples are RPE65-LCA and CEP290-LCA (154,155). It remains to be
determined if gene augmentation treatments can be efficacious in forms of
LCA/EORD that do not follow such a pattern. There is also a need to treat
diseases caused by defects in photoreceptor proteins (e.g., LCA5-LCA,
CEP290-LCA, RDH12-LCA), to move the target of interventions from the
RPE, the primary site of diseases such as RPE65-LCA, into the
photoreceptor, and from simpler mechanisms of disease to complex causes of
vision loss in IRD. There is also the fact that the severity of the photoreceptor
degeneration for some of the molecular subtypes may leave very little room
for treatment beyond the earliest disease stages and youngest ages adding
complexity to treatments and their outcomes for safety and efficacy.
Examples are diseases such as CRB1-IRD, RDH12-IRD and PRPF31-IRD
where epigenetic modifiers of disease expression may play an important role
for triggering onset or progression of the postnatal disease.
Today, for even those pediatric retinal diseases that we think we know
well, such as RPE65-LCA, CEP290-LCA, and achromatopsia, there remain a
number of questions in need of further exploration and which may be best
addressed by studying young patients. For example, what is the impact that
early diseases and treatments early in life have on the structural and
functional development of the retina and visual brain? How early in life
should we treat? How can we expand therapeutic windows as we wait for a
cure for a specific molecular subtype? It is by moving the target of our
research to younger populations of patients that we will be able to answer
these questions and prevent the life-long consequences of irreversible, early-
onset severe vision loss.
Treatment Choices for End-Stage Disease and

Developmental Disorders
Gene therapy may be an option for disease in which the photoreceptor or
ganglion cells are present, even if sickly. However, once the cells have died,
or come to a point where they cannot be resuscitated, gene therapy is no
longer an option. If the cells are no longer viable, there are two options at
present: optogenetic therapy or cell transplantation (Figure 28-1).
The first optogenetic gene therapy trials are in progress (see above; Table
28-1) and involve delivery of a molecule that renders the cell light sensitive
to a remaining cell type in the retina (in the current trials, these are ganglion
cells). While optogenetic therapy has proven effective in animal models,
there are many variables that will determine its efficacy in humans. These
include (a) the cellular target. Ganglion cells may not be ideal since targeting
of these cells would override the functions of other cells in the retina, many
of which enhance vision. (b) The nature of the optogenetic molecule. The
channel protein used in the GenSight Biologics S.A. trial is used in
conjunction with goggles that focus the stimulus appropriately. Undoubtedly,
new optogenetic molecules will be engineered in the future to be able to
harness the wavelengths and intensities of light that are used in everyday life.
An alternative strategy is to delivery stem cells treated ex vivo with gene
therapy reagents into the retina with the goal of having them take on the
function of the cells that have died due to disease progression. This approach
is most likely to be effective if the diseased cells are RPE cells, as those can
likely be delivered safely in a monolayer. It will be more challenging for
photoreceptors or other neural retina cells because these cells will need to
develop synaptic contacts with other cells in the retina in order to be able to
function properly.
A number of early-onset IRDs involve retinal/photoreceptor
maldevelopment with secondary degenerations. Examples are NR2E3-, NRL-,
and CRB1-IRDs (122,156,157). In these diseases, the hope until now was to
treat with gene therapy to prevent secondary degeneration of photoreceptors
through interventions aimed at epigenetic events (158,159). Until recently,
modification of the basic developmental phenotype in these disorders was
considered impossible. Recent demonstration of the possibility of
reprogramming rod photoreceptors in mature murine retina by CRISPR-Cas9
(160) raises the possibility that some of the developmental phenotypes may
become amenable to treatment in the future (Figure 28-1).
A challenge to the field is how to make gene therapy drugs that are
approved available to those who could benefit. Insurance companies are now
making Luxturna, the first approved gene therapy drug, available. National
health systems in the European Union will soon contribute. Hopefully, as
more gene therapy drugs are approved, the costs will also decrease.
SUMMARY
The recent approval of the first retinal gene therapy product, Luxturna, for
RPE65-related retinal dystrophy by U.S. and European regulators has fueled
the development of gene therapy for additional pediatric-onset retinal disease
targets. Gene therapy clinical trials are either in progress or are being planned
for many of these diseases. Additional strategies and novel vectors and
delivery systems are being developed for others. With the continued pace of
progress with ocular gene therapy, gene-based approaches will likely soon
become standard of care for additional, currently untreatable, early-onset
blinding diseases.
ACKNOWLEDGMENTS
We gratefully acknowledge support by the Foundation Fighting Blindness–
sponsored CHOP-PENN Pediatric Center for Retinal Degenerations, the
Brenda and Matthew Shapiro Stewardship and the Robert and Susan
Heidenberg Investigative Research Fund for Ocular Gene Therapy, RDH12
Fund for Sight, Research to Prevent Blindness, the Paul and Evanina Mackall
Foundation Trust, the Center for Advanced Retinal and Ocular Therapeutics,
and the F.M. Kirby Foundation. Most importantly, we acknowledge the
pioneering efforts of patients in participating in clinical research studies.
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29
Congenital Stationary Night Blindness
(CSNB)
Christina Zeitz, Juliette Varin, and Isabelle Audo
PREVALENCE
Congenital stationary night blindness (CSNB) is a clinically and genetically
heterogeneous inherited retinal disorder. To our knowledge, the frequency of
CSNB in the general population has not been documented. Together from our
and published data, we estimate that at least 700 index patients worldwide
have a genetically confirmed diagnosis of CSNB (1). From our worldwide
cohort, approximately 170 French patients have a genetically confirmed
diagnosis of CSNB (1,2). Thus, the prevalence in France with 67.19 million
people would be at least approximately 1:400,000. This is very rare, but we
speculate that many cases are undiagnosed. Only by using specific clinical
examinations and/or molecular genetic testing can CSNB be correctly
diagnosed. In respect to the collection of our large European cohort with
inherited retinal disorders, including approximately 5,000 index cases, 2% of
those present with CSNB.

Clinical signs are present from birth. The name “CSNB” is somehow
confusing, since not all patients are displaying night blindness and for
specific subtypes of CSNB progression has also been noted. Often diurnal
vision is also affected: reduced visual acuity, light sensitivity, high myopia,
nystagmus, and strabismus may be present. Fundus examination and full-field
electroretinogram (ffERG) recordings incorporating the International Society
for Clinical Electrophysiology of Vision (ISCEV) standards (3) can precisely
clinically diagnose patients. Furthermore, documentation of the mode of
inheritance is important for the proper classification of CSNB. Together,
these data help to rapidly identify the known genetic subtypes. Table 29-1
summarizes the main clinical features of the different forms of CSNB.
TABLE 29-1 Summary of general clinical

characteristics and mode of inheritance of most
CSNB
Modified from Vincent A, Audo I, Tavares E, et al. Biallelic mutations in GNB3 cause a unique form
of autosomal-recessive congenital stationary night blindness. Am J Hum Genet. 2016;98:1011–1019;
Zeitz C, Friedburg C, Preising MN, et al. [Overview of congenital stationary night blindness with
predominantly normal fundus appearance]. Klin Monbl Augenheilkd 2018;235:281–289.
Riggs Type of Congenital Stationary Night Blindness

—A Form of Night Blindness With Largely Normal
Fundus
The Riggs type of CSNB (4) corresponds to a rod- photoreceptor
dysfunction. The ffERG reflects severely reduced scotopic responses. The b-
wave is severely reduced or absent at low light intensities (dark adaptation
[DA] 0.01). At a bright flash, the b-wave and the a-wave are reduced (DA
10.0) due to primary rod dysfunction. In contrast, photopic ERGs (light-
adapted [LA] 3.0 and LA 3.0 30 Hz) are normal due to normal cone function
(Figure 29-1, example of Riggs form of CSNB). This form of CSNB has
been reported in autosomal dominant and autosomal recessive pedigrees with
specific mutations in genes coding for proteins implicated in the rod
phototransduction cascade. The phenotype is relatively mild including night
blindness, no nystagmus, and normal photopic visual acuity with only a few
cases showing myopia (1,6,7). This relatively mild phenotype may explain
why to date only a small number of cases with this type of CSNB were
described. This form of CSNB was first described in two autosomal dominant
families: the Nougaret family, coming from South France (8–12), and another
family reported by Rambusch (13,14). In the meanwhile, a similar phenotype
has also been reported in some cases with autosomal recessive CSNB.
However, in the latter cases especially the photopic responses are less
consistent with the classic Riggs type of CSNB (15–18). All patients in the
reports above showed largely normal fundi.
FIGURE 29-1 Schematic drawing of the ERG phenotype
found in CSNB patients along with the affected cell type
and the associated gene. Patients presenting a Riggs type
of CSNB have a rod (purple) dysfunction visible in dark-
adapted condition (DA), and normal cone function as
shown by light-adapted ERG (LA), due to mutations in
either RHO, GNAT1, PDE6B, or SLC24A1 and in RDH5
for the particular phenotype of fundus albipunctatus and
SAG or GRK1 for Oguchi disease. In icCSNB, the b-wave
in dark-adapted conditions is reduced due to mutations
coding for proteins at the photoreceptor synapse
(CACNA1F, CABP4, and CACNA2D4). The complete
form of CSNB is characterized by an absence of the b-
wave on the ERG as a consequence of mutations in genes
coding for proteins at the dendritic tips of cone ON-bipolar
cells (ON-cone BC, light blue) or rod bipolar cells (rod
BC, dark blue) which are GRM6, GPR179, NYX, TRPM1,
or LRIT3. To be noted, for LRIT3 also photoreceptor
expression was suggested (5). See also Chapter 10.
Fundus Albipunctatus—A Form of Night Blindness

With Fundus Abnormalities
Fundus albipunctatus (FA), indirectly, represents also a rod-photoreceptor
dysfunction. Although the expression of the gene defect underlying this
disease is restricted to the retinal pigmented epithelium, the mutant form
leads to disturbance of rhodopsin recycling, which is specifically expressed in
rod photoreceptors. Therefore, patients cannot recover from light activation
and are effectively “bleached” most of the time. The diagnosis cannot be
made solely by ISCEV standard ERGs as recovery following extended DA, a
condition specific to FA, needs to be confirmed (1). Patients with FA show
similar scotopic ERGs to those found in patients with the Riggs type of
CSNB. But, in most patients with FA, unlike Riggs type of CSNB, a
significant or complete recovery of rod-mediated ERG amplitudes can be
observed following prolonged dark adaptation although there is phenotypic
variability (19). Photopic ERGs are mildly abnormal in about half of the
cases and often show flicker ERG delay (1). In addition patients with FA
present normal visual acuity, color vision, and visual fields but still display
night blindness. Strikingly, patients with FA have specific fundus
abnormalities. They often show small white dots in the posterior pole and
midperiphery with sparing of the macular region. With time, the fundus
appearance may change from flecks in childhood to fine dots with age that
may fade or increase over the years (1,20–22). Although FA does not present
a progressive rod–cone dystrophy showing optic nerve pallor, retinal blood
vessel attenuation, or pigmentary bone spicule-like migration in the
periphery, in some patients a phenotypic variability leading to more
progressive phenotypes has been described (19,23). FA has been described as
an autosomal recessive trait. FA is a relatively frequent cause of CSNB, due
to founder mutations in one underlying gene (RDH5) (1).
Oguchi Disease—A Form of Night Blindness With

Fundus Abnormalities
Oguchi disease (OD) is also characterized by rod-photoreceptor dysfunction.
Again, patients with OD show similar scotopic ERGs as found in patients
with the Riggs form of CSNB. In the dark-adapted condition, at low light
intensities (DA 0.01), the b-wave is severely reduced or absent while in
response to a bright flash in addition to the b-wave reduction, the a-wave is
also reduced (DA 10.0), which reflects primary rod dysfunction. After
prolonged dark adaptation, rod sensitivity recovers leading to nearly normal
a- and b-waves on the ERG in response to a single flash (23). However,
unlike FA, the ERG response to a subsequent single bright flash is markedly
attenuated and similar to that recorded after short dark adaptation (20
minutes). The abnormal desensitization of the rod system to a repeated bright
flash is caused by continued activation of the phototransduction cascade by
rhodopsin molecules. This continues until all the chromophore is recycled,
requiring a further extended period of DA (1,24). Photopic recordings are
usually normal (25). Similar to patients with FA, OD patients are
congenitally night blind but have normal visual acuity, color vision, and
visual fields (1). Also similar to FA, patients with OD show specific fundus
abnormalities. These are described by the Mizuo-Nakamura phenomenon: the
fundus has a golden-yellow discoloration that disappears after prolonged dark
adaptation (26,27). Although Oguchi disease is considered to be a stationary
and relatively mild disease, some cases show more severe phenotypes and
disease progression (28–33). The disease is inherited in an autosomal
recessive mode of inheritance, with only a few cases described (GRK1 and
SAG).
Schubert-Bornschein Type of Congenital Stationary

Night Blindness—A Form of “Night Blindness” With
Largely Normal Fundus
The Schubert-Bornschein type of CSNB represents a signaling defect from
photoreceptors to ON-bipolar cells. Similar to the Riggs type of CSNB, the
ffERG shows severely reduced scotopic responses. At low light intensities
(DA 0.01) the b-wave is reduced or absent. Nevertheless, unlike in the Riggs
type of CSNB, after a bright-flash stimulation only the b-wave is reduced
while the a-wave is normal (DA 10.0), resulting in an electronegative
waveform (34). Among the types of CSNB with largely normal fundi, the
Schubert-Bornschein type of CSNB is the most common form. It can be
further subdivided into an incomplete (ic) and complete (c) form of CSNB
(Figure 29-1). This classification is based on ffERG characteristics (35,36)
but correlates as well with the localization of the proteins implicated in
CSNB (1).
Incomplete Congenital Stationary Night Blindness

The incomplete form of CSNB (icCSNB) is characterized by both ON- and
OFF-bipolar cell dysfunction. The ffERG shows reduced but present scotopic
responses to a dim flash. This is why this form was called incomplete CSNB
(1). The b-wave is reduced but present at low light intensities (DA 0.01), and
under a bright flash, only the b-wave is still reduced, while the a-wave is
normal (DA 10.0), which confirms normal rod phototransduction. This
pattern results in the previously mentioned electronegative ERG waveform
(35). The photopic responses are severely affected: the LA 3.0 30 Hz ERG is
severely reduced and delayed with most having a distinct bifid peak. The
single-flash cone ERG (LA 3.0) is also markedly subnormal with a
profoundly reduced b/a ratio such that the a- and b-wave are usually of
similar size (1). ON and OFF responses are abnormal as showed by long
duration stimulation (37). Incomplete CSNB might be confused with cone
dystrophy due to profound photopic alteration, due to the genesis of the ERG
responses at the inner retinal level, but in the case of icCNSB, the macula is
usually normal (1,38). However, in some cases, disease progression and more
severe phenotypes were noted (39–43). The incomplete form is a common
form of CSNB and has been mainly reported in X-linked and in a few
autosomal recessive cases with mutations in genes coding for proteins present
at the synapse of photoreceptors. The phenotype of icCSNB is more
heterogeneous than the one observed of cCSNB (please see below) with
patients presenting little night vision disturbances (36,44–46). However,
photophobia is more common in icCSNB (45). In addition, icCSNB patients
may have myopia, hyperopia, nystagmus, strabismus, reduced visual acuity,
and color vision defects (45).
Complete Congenital Stationary Night Blindness

The complete form of CSNB (cCSNB) is characterized by selective ON-
bipolar cell dysfunction. The ffERG shows severely reduced or absent
scotopic responses to a dim flash, thus the term complete CSNB (1). At low
light intensities (DA 0.01), the b-wave is absent. At a bright flash, only the b-
wave is reduced, while the a-wave is normal (DA 10.0), which is consistent
with normal rod function and results in the previously mentioned
electronegative ERG waveform (35). Compared to icCSNB, the photopic
responses are less altered: the LA 3.0 30 Hz ERG is often of normal
amplitude, but it has a pathognomonic flattened trough and mild implicit time
shift. The single-flash cone ERG (LA 3.0) displays a normal amplitude of the
a-wave with a broadened trough; the wave form has a sharply rising b-wave
with no oscillatory potentials and a reduced b/a ratio (35,47). Long duration
stimulation shows selective abnormalities of the ON responses (37).
Similarly to icCSNB, the complete form of CSNB is also a common form of
CSNB with reported X-linked and autosomal recessive cases with mutations
in genes coding for proteins present at the dendritic tips of ON-bipolar cells.
Patients with cCSNB are indeed congenitally night blind, although they may
not report this in early childhood. They have variably decreased visual acuity
and often show high myopia, nystagmus, and strabismus (1,45). Nystagmus
may disappear with increasing age. Disease progression is rare (48).
GNB3-CSNB
Recently a novel gene defect underlying CSNB was identified (49,50). The
phenotype cannot further be classified (51). Only a few cases have been
described so far, and the phenotypes seem to be variable even in those. At
low light intensities (DA 0.01), the b-wave is reduced. At a bright flash, only
the b-wave is reduced, while the a-wave is normal (DA 10.0), confirming
normal rod phototransduction. The photopic responses are very variable: the
LA 30 Hz ERG can be reduced and delayed. In the single-flash cone ERG
(LA 3.0), the a-wave is normal but can be delayed, and the b-wave is reduced
and delayed. Long duration stimulation shows abnormalities of the ON but
not of the OFF responses. Patients with mutations in GNB3 may be night
blind, showing myopia and nystagmus. More patients with the same gene
defect to be identified in the future may help to better classify this novel form
of CSNB.
GENETICS AND PATHOPHYSIOLOGY

Table 29-2 summarizes the major gene defects underlying CSNB.
TABLE 29-2 Gene defects of CSNB

aCRSD, congenital nonprogressive rod–cone synaptic disorder.
bPatient with this gene defect was previously diagnosed with icCSNB.
From Zeitz C, Friedburg C, Preising MN, et al. [Overview of congenital stationary night blindness with
predominantly normal fundus appearance]. Klin Monbl Augenheilkd 2018;235:281–289.
Gene Defects Implicated in Congenital Stationary

Night Blindness
Inherited retinal disorders (IRD) are genetically and clinically heterogeneous.
Although often it is difficult to deliver clear genotype–phenotype correlations
in IRD, for CSNB it is possible. Mutations in genes important for the rod
phototransduction cascade can lead to isolated rod-photoreceptor dysfunction
as found in the Riggs form of CSNB, in FA and OD (Figure 29-1). In
contrast, mutations in genes important for the signaling from photoreceptors
to bipolar cells or in genes important for the uptake of this signal lead to
incomplete and complete CSNB, respectively. In vitro and in vivo models are
helpful to dissect retinal signaling and the pathogenic mechanisms implicated
in CSNB (1). Table 29-2 summarizes the different gene defects underlying
CSNB, their chromosomal localization, the mode of inheritance, and the link
to OMIM. Figure 29-1 shows the retinal localization of the molecules
implicated in CSNB in a schematic drawing.
Gene Defects Underlying the Riggs Type of

Congenital Stationary Night Blindness, Fundus
Albipunctatus, and Oguchi Disease
Specific mutations in genes coding for proteins important for the rod-
phototransduction cascade, such as RHO coding for rhodopsin; GNAT1,
coding for the α-subunit of transducin; PDE6B, coding for the β-subunit of
the phosphodiesterase; and SLC24A1, coding for the solute carrier family 24
member 1, have been identified in autosomal dominant and autosomal
recessive patients with the Riggs type of CSNB (1). The Nougaret family
from the South of France had the p.Gly38Asp mutation in GNAT1 (12).
Concurrently, two other GNAT1 missense mutations were found in two
autosomal dominant families (7,52) and a homozygous GNAT1 missense
mutation in one autosomal recessive family (17). On the other hand, the
Rambusch family had the p.His258Asn mutation in PDE6B (53). To date,
only a second autosomal dominant family with a mutation in PDE6B was
found (54). Similarly, only a few autosomal dominant families revealed
mutations in RHO (55–59) and a few autosomal recessive families revealed
mutations in SLC24A1 (15,16). The exact pathogenic mechanism of these
mutations in genes coding for proteins of the phototransduction cascade
remains to be elucidated.
Specific mutations in genes coding for proteins important for the rod-
phototransduction cascade, including RDH5, coding for the retinol
dehydrogenase, SAG coding for arrestin, and GRK1 coding for the rhodopsin
kinase, have been identified in patients with autosomal recessive FA (RDH5)
and OD (SAG and GRK1) showing some similarities with patients with the
Riggs form of CSNB but having additional fundus abnormalities. As
mentioned before specific phenotypes can be recovered after extended DA.
This correlates with the function of the affected proteins. Indeed, RDH5 is
responsible for converting 11-cis-retinol into 11-cis-retinal in the retinal
pigment epithelium (RPE) and is thus involved in the recycling of rhodopsin.
Thus, rhodopsin regeneration is delayed, FA patients are effectively
“bleached,” but after long DA, rhodopsin levels can be normalized and thus
the ERG recovers (1). OD patients have mutations in SAG and GRK1, both
genes encoding proteins involved in the deactivation process of the
phototransduction cascade (60,61). The phenotype represents basically no
shutoff of the phototransduction cascade. After extended DA the ERG and
fundus phenotype can be restored.
Gene Defects Underlying Incomplete Congenital

Stationary Night Blindness
Mutations in CACNA1F, coding for the α1-subunit (Cav1.4) of an L-type
voltage-dependent calcium channel, CABP4 coding for the calcium-binding
protein 4, and CACNA2D4 coding for the calcium channel, voltage-
dependent, α-2/δ subunit 4 lead to icCSNB or related rod–cone dystrophies
with some overlapping phenotypes (1,62–65). The mutation spectrum
comprises all kind of mutations. More recently we showed that intronic and
synonymous variants in CACNA1F can also lead to a splice defect causing
icCSNB (2). The respective proteins are important downstream of the
phototransduction cascade, by transmitting signals from the photoreceptors to
the adjacent bipolar cells. Indeed, they localize in a horse-shoe-shaped
manner in rod and cone photoreceptor synapse active zone within the outer
plexiform layer (OPL) (1,65–67). Together these molecules are important for
the correct functioning of the calcium channel. During darkness calcium ions
are taken up by this channel, leading to glutamate release at the synaptic cleft
(1). Together, Cav1.4, CABP4, and CACNA2D4 form the pore and are
important to correctly target the channel to the synaptic membrane, to
modulate calcium currents, and to bind calcium ions (1,68–74). Mutations in
CACNA1F, CABP4, and CACNA2D4 can be associated with loss or gain of
function with insufficiently expressed genes resulting in an altered or
nonfunctional calcium channel activity disturbing the regulation of the
glutamate at the synaptic cleft. Different pathogenic mechanisms have been
associated with the different mutations in these genes, which may explain the
phenotypic variability. Both rods and cones make synaptic contacts with
bipolar cells. There are two types of bipolar cells: ON- and OFF-bipolar cells
expressing different glutamate receptors and responding differently to light.
ON-bipolar cells express the metabotropic glutamate receptor 6
(GRM6/mGluR6) (75–77) and depolarize in response to light (78–80), while
OFF-bipolar cells express ionotropic glutamate receptors and hyperpolarize at
light offset (81–83). ON-bipolar cells make synaptic contacts with both rod
and cone photoreceptors, while OFF-bipolar cells only contact with cone
photoreceptors (84). Since molecules implicated in icCSNB localize in
synaptic terminals of both, rod and cones, as a consequence ON and OFF
responses in those patients are altered as shown in the ERG by long duration
stimulation.
Gene Defects Underlying Complete Congenital

Stationary Night Blindness
Mutations in GRM6, coding for metabotropic glutamate receptor 6; GPR179,
coding for the G protein-coupled receptor 179; LRIT3 coding for the leucine-
rich repeat, Ig-like and transmembrane domains 3 protein; NYX, coding for
nyctalopin; and TRPM1, coding for the transient receptor potential cation
channel subfamily M member 1, lead to cCSNB (85–92). The mutation
spectrum comprises every kind of mutation (1). The respective proteins play
their role in ON-bipolar cells by receiving the signals transmitted from the
synaptic cleft. Indeed, they localize mainly at the dendritic tips of ON-bipolar
cells within the OPL and are important for the depolarization of ON-bipolar
cells at light stimulation, leading to glutamate decrease and TRPM1 channel
opening at the end of this cascade (78,80,87,93–98). Mutations in these
molecules lead to absence of the b-wave and of ON responses as shown in
the ERG by long duration stimulation.
GNB3-Gene Defect
As mentioned above, the GNB3 gene defect cannot be strictly classified in the
different subforms of CSNB. Thus, we did not include the protein
localization of GNB3 in Figure 29-1. GNB3 coding the β subunit of the G
protein heterotrimer (Gαβγ) is known to be expressed in cones and ON-
bipolar cells and modulates ON-bipolar cell signaling and cone transducin
function in mice (99). Due to its expression in cones as well in ON-bipolar
cells, the dual phenotype associated with GNB3 mutations maybe explained
(49).
DIAGNOSTIC STUDIES
Genetic testing of CSNB patients is important for genetic counseling of
patients and their families to distinguish from progressive retinal dystrophies
with similar phenotypic features (1). For example, night blindness is one of
the first presenting signs of progressive rod–cone dystrophy, also called
retinitis pigmentosa. At a young age, patients may initially show normal or
near-normal fundus appearance. Therefore, in addition to accurate
phenotyping, molecular confirmation of CSNB helps to correctly diagnose
and counsel patients. CSNB patients with largely normal fundus, a Riggs-
ERG and autosomal dominant or autosomal recessive inheritance should be
screened for mutations in RHO = GNAT1 > PDE6B and SLC24A1 > GNAT1,
respectively. For patients with autosomal recessive mode of inheritance and
FA, RDH5 should be targeted, while patients with autosomal recessive CSNB
and a phenotype suggestive of OD should be screened for mutations in GRK1
and SAG (1). Patients and especially male patients with the Schubert-
Bornschein type of CSNB should be first screened in CACNA1F and NYX
(1). Both genes are located on the X chromosome and represent the major
causes of this form of CSNB. If a clinical discrimination of icCSNB versus
cCSNB is made, only CACNA1F or NYX needs to be investigated. Our
experience showed that at least 80% of these cases show mutations in one of
those genes. Females and excluded male patients with icCSNB could be
screened in CABP4 and CACNA2D4, especially if they present with high
hyperopia and photophobia. Cases of cCSNB should be screened for defects
in TRPM1 > GRM6 > GPR179 > LRIT3. In cases where no difference
between icCSNB and cCSNB is made, the following mutation detection
strategy should be applied
CACNA1F > NYX > TRPM1 > GRM6 > GPR179 > CABP4 > LRIT3 >
CACNA2D4.
We developed this strategy, based on the prevalence of the specific gene
defects (1). Our recent experience showed that intronic variants and
synonymous variants may be also disease causing and should not be
overlooked (2). In case only preliminary clinical phenotyping data are
available, unbiased microarray analysis and targeted next generation
sequencing (NGS) could be applied (Genetic Testing Registry at
https://www.ncbi.nlm.nih.gov/gtr) (100–102). The prior method is based on
allele-specific primer extension analysis, which allows the detection of
known mutations. The array is regularly updated with new mutations in
known genes and mutations that will be identified in novel gene defects.
However, since there are only a few mutation hotspots and founder mutations
in CSNB and their implicated genes, targeted NGS approaches seem to be
more appropriate. Albeit, initially GC-rich and repetitive regions were less
well covered by the latter methods, more recent techniques seem to overcome
these challenges. After exclusion of mutations by the above-mentioned
method, targeted whole genome sequencing or whole exome or whole
genome sequencing should be applied to identify the disease causing
mutation.
REFERRAL TO LOW VISION

SPECIALISTS
Patients with Schubert-Bornshein type CSNB display abnormal visual acuity
of variable severity and should be appropriately managed for that. These
patients become severely handicapped in dim light and therefore require
referral specifically to plan strategies for safe navigation at school and work,
appropriate driving restrictions, and other accommodations. If best corrected
visual acuity is below normal, all usual accommodations for low vision are
needed in addition to special considerations for night vision (see Chapter 14).
SUMMARY
Inherited retinal disorders are very heterogeneous and can be deciphered
depending on the congenital or progressive course of the disease or by the
type of retinal cell that are involved. Herein we describe the genetic and
phenotypic characterization of CSNB. Depending on the mutated gene,
CSNB patients can present a rod-photoreceptor defect (Riggs type of CSNB)
with or without fundus abnormalities (Oguchi disease, fundus albipunctatus)
or a transmission defect from photoreceptor to bipolar cells (Schubert-
Bornschein type). The incomplete form of Schubert-Bornschein type of
CSNB is due to a defect of proteins localized at the photoreceptor synapse,
while the complete form results from a defect of proteins localized in ON-
bipolar cells. Together with other clinical symptoms, clear genotype–
phenotype correlations can be made as described herein.
FUTURE TREATMENTS
To date, CSNB is still an incurable disease. Gene therapy seems to be the
more suitable approach to treat CSNB as it is largely a stationary disorder and
the gene defect underlying the disease has been diagnosed in about every
patient. This kind of treatment has already showed its efficacy in inherited
retinal disorders as for LCA due to mutations in RPE65 (103,104) using
adeno-associated virus (AAV) vector (105–107). Concerning CSNB, a gene
therapy approach for X-linked cCSNB has been studied (108). Intravitreal
injection of the wild-type copy of Nyx using an AAV vector in mice lacking
this gene led to a partial rescue of the b-wave in scotopic conditions and
gating of the cation channel TRPM1. The partial rescue was obtained in
extremely young mice, while the treatment was inefficient in adult mice.
Nevertheless, the improvement following the treatment was mild, and further
investigations on the restoration of the visual acuity were not documented.
Such treatment has not been tested in nonhuman primates so far. More
investigations concerning this therapeutic strategy for other CSNB need also
to be done.
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30
Stargardt Macular Dystrophy/Fundus
Flavimaculatus: From Disease and
Gene Discovery to Identification of
Disease Pathways to the Development
of Therapies
Robert K. Koenekoop and Thomas M. Aaberg Sr
INTRODUCTION AND HISTORICAL

PERSPECTIVES
Stargardt macular dystrophy (STGD1, OMIM 248200) was described by
Professor Karl Stargardt in 1909 and represents the most common inherited
macular dystrophy of childhood, often with an onset before age 10 years (1).
STGD1 frequently leads to legal blindness due to central retinal pigment
epithelium (RPE) atrophy and photoreceptor degeneration, with clinical
evidence of macular atrophy and subretinal yellow flecks (see Figure 30-1).
FIGURE 30-1 Fundus photo of the left eye. STGD1 with
macular atrophy and subretinal yellow flecks. (Reprinted
by permission from Springer: Tsang SH, et al. Atlas of
inherited retinal diseases. Advances in experimental
medicine and biology. Vol. 1085. Cham: Springer; 2018.
Copyright © 2018 Springer International Publishing AG,
part of Springer Nature.)
The flecks of STGD1 represent autofluorescent lipofuscin deposits, which

contain bisretinoid fluorophores. Bisretinoids are toxic to RPE cells. Despite
two colored drawings in the original Stargardt 1909 report showing
perimacular yellow–white flecks, little or no attention was given to the fleck
component of the disorder until Professor Adolph Franceschetti, in
Switzerland, began discussing them in 1963 and deemed them separate from
STGD1. He first used the term “Fundus Flavimaculatus” (FF) (2). In 1980,
Eagle et al. were the first to report the accumulation of a lipofuscin-like
substance in the RPE in two post-mortem eyes from a 24-year-old man with
FF (3). This focused attention on the potential role of lipofuscin accumulation
in the pathogenesis of STGD1. Delori et al., using spectrophotometry, also
demonstrated an abnormally high level of “lipofuscin-like material” in the
RPE in STGD1 patients (4), beginning the understanding of its
pathophysiology.
In 1997, Professor Rando Allikmets discovered that STGD1 is caused by
mutations in ABCA4, a retinal gene that encodes an important retinal “pump”
in the visual cycle (aka the vitamin A or retinoid cycle) (5). ABCA4 acts as a
photoreceptor outer segment (OS) membrane “flippase” and translocates
retinaldehyde conjugated to phosphatidylethanolamine across photoreceptor
OS disc membranes (generated by the photoexcitation of 11-cis-retinal) to the
cytoplasmic side of the photoreceptors. ABCA4 is expressed in rods (6),
cones (7), and RPE cells (8). Deficient ABCA4, due to biallelic mutations,
leads to the accumulation of the bisretinoids, clinically showing retinal flecks
with hypofluorescence alternating with hyperfluorescence on fundus
autofluorescence imaging (FAF) of the STGD1 retinas (see Figure 30-2).
FIGURE 30-2 Fundus photos, fundus autofluorescence

(FAF), and optical coherence tomography (OCT) of the
central retina of the right (top) and left eye (bottom).
STGD1 disease showing an atrophic macular lesion
(beaten metal), hypofluorescence in the macula and
hyperfluorescence in the surrounding retina (no flecks),
and marked loss of the ellipsoid zones in the fovea and
perifoveas l regions. (Reprinted by permission from
Springer: Tsang SH, et al. Atlas of inherited retinal
diseases. Advances in experimental medicine and biology.
Vol. 1085. Cham: Springer; 2018. Copyright © 2018
Springer International Publishing AG, part of Springer
Nature.)
After fluorescein angiography (FA) was developed (6), a dark (“silent”)

choroid was noted in over 70% of STGD1 patients due to the blockage of
fluorescein by the lipofuscin laden cells (see Figure 30-3).
FIGURE 30-3 Fundus fluorescent angiography. STGD1

disease with dark choroid, hiding the choroidal flush due
to massive accumulation of lipofuscin the overlying layer,
the RPE.
Professor Deutman, in The Netherlands (9), was the first to use the term
“Stargardt flavimaculatus,” reopening the discussions of STGD1 and FF
being separate or the same disease. Professor Aaberg, in the United States
(10), reported that both STGD1 and FF patients can coexist within the same
families. He stressed that STGD1 patients present with atrophic macular
changes and subtle or obvious subretinal yellow flecks in childhood or young
adulthood (see Figure 30-4).
FIGURE 30-4 Fundus photo and FAF of the right and left
eyes. STGD1 with significant macular atrophy and flecks
with a significant macular hypofluorescence and
surrounding hyperfluorescent flecks.
The debate about whether STGD1 and FF are the same or different entities
has been settled as genetic studies have demonstrated a firm molecular
relationship between the two diseases (11–13) following discovery of ABCA4
mutations in both clinical disorders. The designation STGD1 in this chapter
refers to both entities. STGD1 can be severe and early onset but develops
after normal early visual development, normal foveation, and proper, early
development of the fixation reflex, which may provide opportunities for
meaningful therapeutic interventions.
How STGD1 evolved from disease description and naming in 1909, to
gene and protein discovery more than 100 years later in 1997, and to
potential treatments today represents an exciting scientific story. This history
serves as a model for understanding and advancing research into all inherited
retinal degenerations (IRDs).
GENE DISCOVERY
ABCA4 gene discovery started with linkage analyses of large multiplex
STGD1 families, followed by gene identification through positional cloning,
culminating in protein identification and characterization. Professor Josseline
Kaplan, in Paris, performed linkage analysis on eight French multiplex
STGD1 families and assigned the STGD1 disease locus for the first time to
chromosome 1p21-p13 (14).
Gerber et al. in 1995, then reported mapping of the FF disease locus to
chromosome 1p13-p21 as well, in the genetic interval near locus D1S435 in
four multiplex FF families (15).
At about this time, Professor Rando Allikmets was characterizing the
protein superfamily of mammalian ABC transporters and mapping and
correlating (assigning) its members as possible candidates to various human
disease phenotypes. The ABC superfamily includes genes whose products are
transmembrane proteins involved in energy-dependent transport of a wide
spectrum of substrates across membranes (5). One of the studied ABC
transporters, ABCR (now called ABCA4), mapped to the human chromosome
1p13-p21 region close to microsatellite markers D1S236 and D1S188.
Northern blot analyses showed that ABCA4 expression is uniquely retina
specific and not found in other human tissue. Mutational analysis of the
ABCA4 gene in 48 STGD1 families revealed a total of 19 different mutations,
the majority representing missense mutations in conserved amino acid
positions (5).
Clinical studies, together with more recent molecular genetics studies,
demonstrate that STGD1 and FF are variable expressions of the same
disorder (16); other disorders have joined this list to reveal an expanding
spectrum of disease types and severities including autosomal recessive
ABCA4 mutations in retinitis pigmentosa (RP), Stargardt maculopathy (see
Figure 30-5), and cone–rod dystrophy (see Figure 30-6). Heterozygous
ABCA4 mutations have been postulated by some authors and refuted by
others to play a role in age-related macular degeneration (AMD) (17,18).
FIGURE 30-5 Right and left eye with retina photography

and FAF. Severe macular disease in a patient with STGD1
and ABCA4 mutations with enlarging macular atrophy,
retinal fibrosis, and beaten metal changes. FAF shows
more extensive changes with hyper and hypofluorescent
changes alternating. (Reprinted by permission from
Springer: Tsang SH, et al. Atlas of inherited retinal
diseases. Advances in experimental medicine and biology.
Vol. 1085. Cham: Springer; 2018. Copyright © 2018
Springer International Publishing AG, part of Springer
Nature.)
FIGURE 30-6 Right and left eyes with fundus

photography and FAF. Severe diffuse disease in a patient
with CRD and ABCA4 mutations, showing advanced
retinal degeneration, with choroidal show and choroidal
sclerosis, vascular narrowing, and extensive macular
pigmentary clumping. FAF shows a markedly abnormal
pattern of lipofuscin, almost devoid of signals. (Reprinted
by permission from Springer: Tsang SH, Sharma T. Atlas
of inherited retinal diseases. Advances in experimental
medicine and biology. Vol. 1085. Cham: Springer; 2018.
Copyright © 2018 Springer International Publishing AG,
part of Springer Nature.)
PREVALENCE
Although STGD1 is one of the most common causes of early-onset macular
degeneration, it is still a rare disease having an estimated prevalence of
1:10,000 (19).
Because the accumulation of abnormal catabolic debris in the RPE is
believed to lead to oxidative damage at the cellular level, and the fact that
oxidative damage from light exposure and smoking can influence other
macular diseases, such as age-related macular degeneration, it is possible that
these environmental stressors may be cofactors in disease progression in
STGD1. It has also been suggested that mutations in ABCA4 may make
patients more susceptible to retinal toxic medications (20). Because by-
products of vitamin A generated after light bleaching are part of the abnormal
accumulations in STGD1, excess vitamin A in the diet may worsen retinal
function and should be avoided. Vitamin supplements containing vitamin A
are contraindicated. Similarly, bright light exposure may hasten vision loss; a
study of patching one eye in a STDG1 patient resulted in better vision in that
eye (21).
Weng and associates reported a substantial deposition of A2E in the
mouse RPE of the knockout mouse model (abca4−/−) for STGD1. The mice
also developed delayed dark adaptation (DA) (22). If these mice were
sheltered from bright light, the retinal degeneration was attenuated (23). Radu
and coworkers demonstrated that vitamin A supplementation significantly
increased lipofuscin pigment in the RPE of the Abca4 knockout mouse model
of Stargardt disease (24). Studies like these support the concept that lifestyle
modifications may delay the progression of disease and perhaps preserve
useful vision.
PATHOPHYSIOLOGY
Some of the questions surrounding STGD1 are about how ABCA4 mutations
cause retinal cell death, which cell populations die first and in what sequence.
Also, how fast does the maculopathy expand and does it correlate to RPE
loss? In addition, is the expansion of atrophy (measured by the ellipsoid zone
(EZ line) on optical coherence tomography (OCT) a good clinical outcome
measure to study the possible efficacy and safety of new experimental
treatments?
To answer these questions, the pathogenic sequence of ABCA4-associated
disease must first be determined. Does the disease commence in RPE (and
then affect PR death) or in photoreceptors first (and then spread to RPE) or
are both cells affected concurrently?
Hypothesis 1: STGD1 Commences in the RPE Cells
Evidence that STGD1 is initiated in the RPE cells came from multiple
mouse and human histopathologic studies and was the first disease sequence
hypothesis to be proposed. Eagle et al. found the accumulation of lipofuscin
in RPE on histopathology studies of cadaver eyes and postulated that STGD1
commences in the RPE. Clinical studies led to a proposed disease model of a
pathogenic sequence of lipofuscin accumulation leading to RPE cell damage
and then to photoreceptor loss (25).
In ABCA4-associated diseases, one of the most noticeable retinal
changes is the presence of autofluorescent foci (yellowish subretinal fundus
flecks) captured by short wave length (SW-AF) signals and imaging. For
many years, it had been assumed that flecks represented lipofuscin-laden
RPE cells. Flecks are dynamic features of the STGD1 fundus; therefore,
Paavo et al. analyzed flecks in a precise sequence of short wave length (SW-
AF), near-infrared autofluorescence (NIR-AF), and infrared reflectance (IR-
R) imaging, followed by SD OCT and by quantitative AF (qAF) (26). They
related these multimodal findings to structural information acquired by SD-
OCT. They then studied images from 12 STGD1 patients (6 females and 6
males; age range from 9 to 61 years old) who had been recruited after
obtaining a clinical diagnosis of STGD1 and positive ABCA4 genetic testing.
They found that isolated flecks in SW-AF images are often situated
within hypo autofluorescent (hypo AF) zones in NIR-AF images. Hypo AF in
NIR indicates RPE atrophy. They also found that the flecks visualized in SW-
AF images corresponded to hyperreflective aberrations extending radially
within photoreceptor-attributable bands (EZ) on the SD-OCT scans. These
observations suggested that fundus flecks are a manifestation of degenerating
photoreceptor cells in STGD1, not lipofuscin-laden RPE cells (26).
Hypothesis 2: STGD1 commences in photoreceptors. Some authors have
documented by histopathology and in vivo imaging that photoreceptor (PR)
loss occurs earlier than RPE loss in the disease pathophysiology. In these
studies, photoreceptors have shorter OSs even when the apposing RPE still
appears to be organized, whereas adaptive optics studies showed increased
photoreceptor spacing with normal RPE spacing (27,28). In a 2-year study of
16 STGD1 patients studying loss of the EZ on SD-OCT imaging, Cai et al.
demonstrated that a gradual expansion in the total area of EZ loss in STGD1
occurs at around 0.31 mm2/year, similar to the expansion of the RPE loss on
FAF at 0.33 mm2/year (29). The expansion is nonuniform and is not seen
equally at all edges of an existing lesion (29) (see Figure 30-7). These
studies, however, do not provide definitive proof that PR loss precedes RPE
loss, as each study has limitations.
FIGURE 30-7 OCT images of EZ zone and measurements
in STGD, year 1 and year 2. Note the unequal, but
measurable expansion of the macular atrophy. (Reprinted
from Cai C, Light J, Handa J. Quantifying the rate of
ellipsoid zone loss in Stargardt disease. Am J Ophthalmol
2018;186:1–9. Copyright © 2017 Elsevier. With
permission.)
To investigate early STGD1, Song et al. (27) used AOSLO to evaluate two
brothers with macular atrophy, a mild phenotype, and their unaffected
parents. One son (more severe phenotype) was found to have FAF indicating
central hypo AF, surrounding hyper AF at 0.7 mm, and uniform AF at 1.7
mm. The other son (less severe phenotype) had a subtle bull’s-eye
maculopathy with no peripheral flecks. OCT showed foveal preservation of
the OS with a thickened external limiting membrane, perifoveal atrophy of
the outer retina and RPE, and normal layers at 1.7 mm. Fundus AF indicated
a bull’s-eye with central hyper AF, and surrounding annular hypo AF, then
hyper AF at 0.7 mm. Autofluorescence was uniform at 1.7 mm. Although
both affected individuals manifested macular atrophy without flecks,
photoreceptor spacing was significantly increased peripheral to the clinically
detectable lesions. FAF was homogeneous at these locations, suggesting that
a decline in photoreceptors precedes lipofuscin accumulation in RPE in the
macular atrophy phenotype. Thus, this second hypothesis suggests that the
disease sequence in ABCA4-related STGD1 commences in rods and cones
and proceeds to the RPE, opposite of the hypothesis of Cideciyan (25). A
third hypothesis is that the disease sequence commences in the RPE and
photoreceptors at the same time.
Cukras et al. examined the temporal–spatial patterns of FAF with
excitation at both 488 nm (standard FAF) and at 795 nm (near-infrared
autofluorescence, NIR) (Figure 30-8) in a longitudinal case series involving
8 eyes of 4 patients (range of follow-up, 11 to 57 months; mean, 39 months)
(30).
FIGURE 30-8 STGD1 patient retinal photo STGD1
patient retinal photo (A), standard FAF (B), and NIR (C)
studies. The photo of the left eye shows the macular
atrophy and yellow subretinal flecks. The standard FAF
shows the central hypo FAF, and the surrounding hyper
FAF, corresponding to the clinical flecks. NIR shows more
extensive hypo FAF in the center, and alternating hypo
and hyper FAF of the clinical flecks. (Reproduced with
permission from Cukras CA, Wong WT, Caruso R, et al.
Centrifugal expansion of fundus autofluorescence patterns
in Stargardt disease over time. Arch Ophthalmol
Medical Association.)
The goal of this study was to elucidate molecular and cellular mechanisms
underlying STGD1 by studying longitudinal hyper and hypofluorescent
patterns both with NIR and regular FAF imaging. In addition, they evaluated
if these findings may constitute potential outcome measures in clinical trials.
NIR evaluates melanin-containing RPE cells, while standard FAF evaluates
photoreceptor health. With aging in normal eyes, melanin, unlike lipofuscin,
decreases in the RPE; melanin granules in RPE cells decrease in number as
those containing lipofuscins accumulate. These trends have led to the
hypothesis that melanin in RPE cells may regulate lipofuscin accumulation
and/or the photo-oxidation of lipofuscin-related compounds. Research has
demonstrated that lipofuscin-related compounds can act as photosensitizers,
increasing the production of reactive oxygen species. Chemical modification
of proteins and DNA could then induce RPE cell death. The study by Cukras
et al. (30) explores these melanin–lipofuscin interactions and how they
contribute to the accelerated course of lipofuscin accumulation in STGD1
(Figure 30-9).
FIGURE 30-9 Progression of FAF and NIR lesions in two

eyes (A) and (B) over time. (Reproduced with permission
from Cukras CA, Wong WT, Caruso R, et al. Centrifugal
expansion of fundus autofluorescence patterns in Stargardt
disease over time. Arch Ophthalmol 2012;130(2):171–179.
Copyright © 2012 American Medical Association.)
They discovered and identified a remarkable pattern of centrifugal

progression of patterns of fleck creation and then fleck dissolution in STGD1
patients followed up to 57 months (almost 5 years) (Figure 30-10).
FIGURE 30-10 Centrifugal and radial movement of the
subretinal flecks and disappearance of flecks in 3 different
eyes with Stargardt disease. (Reproduced with permission
Cukras CA, Wong WT, Caruso R, et al. Centrifugal
expansion of fundus autofluorescence patterns in Stargardt
disease over time. Arch Ophthalmol 2012;130(2):171–179.
Copyright © 2012 American Medical Association.)
The ellipsoid zone (EZ) represents the inner segment/outer segment junction
(IS/OS jct) of photoreceptor cells. In the center of the fovea, this represents
the IS/OS jct of the cones. Tanna et al. explored EZ parameters in 41 patients
and found a mean annual rate of transverse EZ loss (± SD) was 279.5 ± 259.9
μm/year. The mean rate of area of EZ loss (± SD) was 1.20 ± 1.29 mm2/year.
This highlights the reliability of SD-OCT in measuring EZ loss, which
appears to be a robust measurement of photoreceptor integrity (31).
To elucidate the cellular origin of AF abnormalities in STGD1, Song et
al. utilized fluorescence adaptive optics (FAO) scanning light
ophthalmoscopy to provide autofluorescence (AF) images in vivo with
microscopic resolution. They found that AO images of cones, rods, and RPE
cells at the “leading edge” of the macular atrophy show AF signals that
appear to colocalize with photoreceptors. Their observation is consistent with
reports of fluorescence arising from photoreceptors in STGD1 (32).
There is extensive variability in both phenotypic and gene expression
associated with ABCA4 mutations. This variability manifests itself in the
wide spectrum of disease “types” (e.g., STGD1, FF, CRD, RP, possibly
AMD, etc.). Variability is also found in the severity spectrum associated with
STGD1 itself (i.e., ranging from severe, early onset to late onset with foveal
sparing). Added to this phenotypic diversity is the variability linked to the
diversity of expression patterns of ABCA4 in both rods and cones and in RPE
cells. Documenting and predicting the spatial and temporal progression of
ABCA4-associated diseases is, therefore, critical for the development of
potential therapies. Therapeutic windows, disease stages, disease
classification, and severity ranges must be established to consider what,
when, where, and how to deliver the agents now being tested (drugs, genes,
and/or cells) for treatments, either to slow, halt, or reverse the disease course.
In an attempt to explain the diversity of phenotypes associated with
ABCA4, ranging from FF and STGD1 to severe autosomal recessive (ar)RP
and autosomal recessive cone–rod dystrophy (arCRD), a genotype–
phenotype model was proposed (33). According to this model, the severity of
the functional consequences of the combination of ABCA4 mutations
correlates with the severity of the clinical phenotype (33,34). Patients with a
severe and a mild variant or two moderately severe variants are expected to
present with STGD1, patients with a severe and a moderately severe mutation
develop CRD, and individuals with two severe mutations develop a severe
form of CRD. The CRD phenotype has been associated with an early age at
onset (<10 years) (35,36) and in a later stage often resembles RP. The amount
of remaining ABCA4 protein function is thus inversely correlated with the
severity of the resulting disease phenotype under this hypothesis, which has
been validated with later studies (37).
According to the above model, two “mild” or “hypomorphic” variants are
expected not to give rise to a disease phenotype (34). This model explains
how there can be an unusually high carrier frequency of mild variants in the
general population, which can be as high as 5.5% (e.g., c.2588G>C; p.
Gly863Ala, Gly863del) in Sweden (38) and 11.3% (c.5882G>A; p.
(Gly1961Glu) in Somalia (Figure 30-11) (39,40) when the disease itself is
rare. Some nonexomic and synonymous variants in ABCA4 have been shown
to be disease causing in combination with more severe alleles (41).
FIGURE. 30-11 p. G1961E ABCA4 phenotype. Color

fundus photo, FAF, and OCT of a STGD1 patient with the
p. G1961E ABCA4 genotype, illustrating the mild bull’s-
eye maculopathy and the “optical gap” in the fovea.

STGD1 characteristically presents as central visual loss in a child, with subtle
retinal changes consisting of macular atrophy with or without yellowish
subretinal flecks. However, this presentation does not always occur; Bax et
al. showed that of 280 STGD1 patients, more than 11% presented with
normal appearing retinas. Common misdiagnoses were amblyopia, myopia,
and psychiatric conditions, such as malingering (42). Overlap exists between
the STGD1 phenotype caused by ABCA4 and other early macular dystrophies
caused by other retinal genes. Wolock et al. demonstrated that part of the
missing heritability of STGD1 (15%) is caused by dominant mutations in
PRPH2 and CRX, recessive mutations in CERKL, and several other genes
(40).
Although the most common onset of Stargardt disease is in the preteen to
teenage years, a proportion of patients can exhibit onset in early adulthood
(“adult-onset STGD1”) or in late adulthood (“late-onset STGD1”).
Interestingly, late-onset ABCA4-associated retinal dystrophy tends to be
associated with parafoveal atrophy (not foveal atrophy) and relative foveal
sparing with preservation of visual acuity (see Figure 30-12). Identifying the
genetic and molecular correlates of this important (foveal sparing) phenotype
will likely point to a separate ABCA4-induced pathophysiologic mechanism,
in turn providing insight into a treatment option.
FIGURE 30-12 Fundus photo and FAF of the right eye of

a patient with adult-onset STGD1 with foveal sparing.
This patient has 20/20 visual acuity at age 45 years old and
harbors the ABCA4 p.N1868I (c.5603A>T) mutation.
Adult- and late-onset Stargardt disease phenotypes are often associated with
milder missense mutations, which may retain partial ABCA4 protein
function. This is very much unlike the null mutations in ABCA4, which are
associated with early-onset STGD1, arRP, and arCRD. Many late-onset
Stargardt disease cases were found to have only one mutant allele while the
second allele could not be found. A report by Zernant et al. demonstrated that
the second allele in many of these cases harbored a single ABCA4 variant,
p.N1868I (c.5603A>T) (43) (Figure 30-12). Importantly, this variant was
previously considered not to be disease causing, because the frequency of the
p.Asn1868Ile allele, 7%, is unusually high in the general population for a
disease-causing allele. Deep intronic variants and deletions account for other
cases of “single allele” Stargardt (44).
Stargardt macular dystrophy is now believed to be a spectrum of
maculopathies with overlapping and diverging phenotypic characteristics, all
caused by a spectrum of ABCA4 mutations. Early onset (juvenile STGD1),
early adulthood, late onset, foveal sparing, and FF are five broad clinical
groups of diseases. In the past 20 years, attempts have been made to stage
and group STGD1 further by classifying STGD1 clinically using
electrophysiologic, autofluorescence, and acuity measures. Here follows three
commonly used staging classifications for STGD1 disease.
Staging System No. 1.

Ophthalmoscopic/Electrophysiologic/Psychophysical
Classification of STGD1
In 1976, one of the first and most frequently used classification systems was
developed and reported by Professor Gerry Fishman (45), who classified
STGD1 into four stages: Stage 1: presence of variable pigmentary changes in
the macula from initial faint and irregular pigment mottling to a beaten-metal
or “snail-slime” appearance to an eventual atrophy of the RPE and
choriocapillaris. A ring of flecks often circumscribes an area within 1 disc
diameter on all sides of the fovea. Initially, relative and eventually absolute
central or paracentral scotomas are seen. Electroretinogram (ERG) and
electrooculogram (EOG) measurements are usually normal. Stage 2: presence
of fundus flecks beyond 1 disc diameter of the margin from the fovea. The
flecks extend beyond the vascular arcades and often nasal to the optic disc.
Partial resorption of the flecks may be observed in this stage. Peripheral
visual fields are normal, a relative central scotoma may be observed in
patients with macular involvement. ERG amplitudes and EOG ratios are most
often normal, but subnormal cone and rod responses may be observed. Some
patients take longer to reach normal scotopic ERG amplitudes.
These patients not infrequently manifest a prolonged period for DA.
Stage 3: presence of diffusely resorbed flecks and an exposed choriocapillaris
with atrophy within the macula. EOG testing shows subnormal ratios for light
peak to dark trough with subnormal cone or cone and rod ERG amplitudes.
DA shows a prolonged cone–rod break time and a prolonged period to reach
normal or elevated rod final thresholds. Central field defects are similar to
those in stage 2; however, a degree of peripheral or mid-peripheral field
impairment may be evident. Stage 4: presence of diffusely resorbed flecks
and extensive choriocapillaris and retinal pigment epithelial cell atrophy
throughout the fundus. Peripheral fields are moderately to extensively
restricted. ERG show reduced cone and rod amplitudes. Elevated cone and
rod thresholds are seen on DA testing.
Staging System No. 2. ERG Classification of STGD1

Lois et al. established a classification of STGD1 based on electrophysiologic
findings. Group 1: severe pattern ERG (PERG) abnormality (indicating
macular dysfunction) and normal full-field (ff) ERGs. Group 2: abnormal
PERG with an additional diffuse loss of cone function on ffERG. Group 3:
abnormal PERG with additional diffuse loss of both cone and rod function on
ffERG (46). These groups have prognostic implications as group 1 has the
best prognosis, group 3 the worst, and group 2 intermediate in follow-up
studies (47). Clinically significant ERG deterioration was found to be
divergent in the three phenotypic groups as 22% of group 1, 65% of group 2,
and 100% of group 3 progressed (47).
Staging System No. 3. Autofluorescence Classification

of STGD1
FAF is a qualitative, noninvasive imaging modality that maps the retinal
lipofuscin deposits by hypo- and hyperfluorescence alterations. FAF was
developed and introduced by Delori in the 1980s (4). Fluorophores, naturally
occurring molecules that absorb and emit light of specified wavelengths,
make up the majority of lipofuscin. Excessive accumulation of lipofuscin
granules in the lysosomal compartment of RPE cells represents a common
downstream pathogenic pathway in various hereditary and complex retinal
diseases, including STGD1. The major lipofuscin fluorophore is A2E, a by-
product of the retinoid cycle, as a result of the production of all-trans retinal
after light capture of 11-cis-retinal. A2E exerts diverse toxic effects on the
RPE. To produce autofluorescence, a fluorophore absorbs a photon of the
excitation wavelength, which elevates an electron to an excited, higher
energy state, then dissipates energy through molecular collisions, and finally
emits a quantum of light at a lower energy and longer wavelength
transitioning back to the ground state. FAF utilizes blue-light excitation
(SWI). FAF may also use other excitation wavelengths to detect melanin with
near-infrared (NIR) autofluorescence. Lipofuscin absorbs blue light with a
peak excitation wavelength of 470 nm and emits yellow–green light at a peak
wavelength of 600 to 610 nm. In contrast to lipofuscin, melanin in the RPE
cells has a peak excitation at 787 nm and is the primary fluorophore in near-
infrared autofluorescence. RPE melanin acts as an antioxidant, protecting
against free radicals, redox-reactive heavy metals, photo-oxidation, and
lipofuscin accumulation. Fujinami et al. classified 68 STGD1 patients into
three FAF subtypes at baseline (48). Type 1: localized low FAF signals in the
fovea surrounded by a homogeneous background (n = 19). Type 2: localized
low signal at the macula but surrounded by a heterogeneous FAF background
with numerous foci of abnormal signals (n = 41). Type 3: multiple low-signal
areas at the posterior pole with a heterogeneous background (n = 8). When
the areas of reduced FAF signal were measured, and the rate of atrophy
enlargement (RAE) was calculated over a mean interval of 9.1 years,
important differences in progression were found. The RAE (mm2/year) was
0.06 mm2/year for type 1, 0.67 mm2/year in type 2, and 4.37 mm2/year in
type 3. As the disease type is more severe, so is the rate of progression of the
FAF lesions. There was a significant association between FAF subtype and
genotype. Milder ABCA4 sequence variants (including missense mutations)
were found in type 1 while more deleterious sequence variants (including
nonsense mutations) were identified in type 3 (48).
Subtypes of STGD1
An increasing number of ABCA4 mutations are being correlated to various
STGD1 phenotypes. The p. G1961E (p.G1961E) ABCA4 mutation
phenotypic subtype of STGD1 is a common, mild type of STGD1,
characterized by bull’s-eye maculopathy and mild progression (see Figure
30-11). There is lower accumulation of lipofuscin (autofluorescence [AF])
and often long-term preservation of cone and rod function.
Another STGD1 phenotype is ROC, rapid-onset chorioretinopathy (49).
Characteristically, there is a homogenous ring distribution of hyper
autofluorescence (AF) apparent at an early-disease stage with nascent fleck
development in the mid-periphery. A dense, homogeneous distribution of
autofluorescence concentrated across the macula subsequently progresses to
macular atrophy. This spatially corresponds to reduced signal on NIR-AF
indicating a widespread loss of underlying RPE in this region while flecks
develop in the periphery over time. Most ABCA4 mutations causing ROC are
null alleles, with no expected ABCA4 protein (24). The ROC phenotype is
characterized by early onset (<10 years of age) and a rapid progression to
advanced disease. Rapid accumulation of bisretinoid lipofuscin in
photoreceptor cells, secondary deposition in the RPE, followed by
widespread RPE death in the macula, thinning of the choriocapillaris and CS
layer, diffuse, multifocal atrophy, and generalized electrophysiologic
dysfunction lead to significant visual decline at a much younger age than in
other cases of ABCA4 disease (24).
PATHOLOGIC STUDIES OF STGD1

Lipofuscin-laden RPE cells from a patient with STGD1/FF demonstrate
tremendous distension and loss of hexagonal architecture. These
hypertrophied RPE cells show abundant periodic acid–Schiff–positive
material in the apical compartment. This accumulation appears to be
responsible for RPE dysfunction with eventual photoreceptor death and
severe vision loss in STGD1 patients (50). Progressive accumulation of
lipofuscin likely alters other components of RPE cells, such as melanin (20).
The apical surface of the RPE cell extends microvilli to encompass the rod
and cone OSs. The apex of the RPE cell is responsible for the phagocytosis of
shed photoreceptor OS discs forming phagosomes in the apical RPE cell
cytoplasm. The eventual fate of the phagosome is to be incorporated into the
lysosomal system for degradation and partial recycling. Cathepsin D is an
important protease involved in digestion of the rhodopsin-rich disk
membranes in the phagocytic process. Perturbation of cathepsin D, or other
catabolic enzyme systems, such as ubiquitination, can lead to the buildup of
intracellular and extracellular debris (51).
Lipofuscin normally (in normal controls) accumulates in the RPE in an
apparent biphasic pattern with one peak occurring between 10 and 20 years
of age and the second peak occurring around 50 years of age (52). Lipofuscin
buildup is greatest in the posterior pole, especially in the macula, but spares
the fovea (53). Macular pigments composed of lutein and zeaxanthin may
influence (protect) the accumulation of lipofuscin in the fovea (54).
Alternatively, specialized cones that reside in the fovea may have cellular
membrane properties or different visual pigments that preclude the formation
of lipofuscin in this region. These hypotheses have not yet been tested.
Lipofuscin is contained within granules of relatively uniform size. It is a
lipid–protein aggregate that auto-fluoresces when excited by SW light. The
composition of RPE lipofuscin is controversial, but most believe that partially
degraded OS disks and autophagy processes contribute to the bulk of
lipofuscin (55). There is a growing body of evidence that lipofuscin may
actually induce oxidative damage by acting as a photosensitizing agent
generating reactive oxygen species (56). As a design of function, the macula
is exposed to a lifetime of light irradiation, including blue light wavelengths
that have been shown to induce reactive oxygen intermediates (57). In
addition, biochemical properties of lipofuscin may actually interfere with the
enzymatic pathways of degradation by influencing lysosomal pH (58). Thus,
as lipofuscin accumulates, the cellular catabolic machinery may be damaged
or inhibited leading to more accumulation. This cycle would theoretically
continue throughout life until the RPE cell is overwhelmed and cellular death
and atrophy occur. Disease in the photoreceptors or in the RPE may result in
this cycle.
The ABCA4 gene encodes a protein that is an ABC transporter. These
transporters are involved in the transport of all-trans retinal from the
photoreceptor OS disc membrane to the OS cytoplasm. Knockout mouse
(Abcr−/−) studies strongly suggest that ABCA4 functions as an export
flippase for retinyl derivatives (22). This transport reaction is part of a larger
cycle that replenishes the supply of the critical light-sensing form of vitamin
A, 11-cis-retinal, in the photoreceptor OSs. The process of light absorption in
the photoreceptors converts 11-cis-retinal to a derivative called all-trans
retinal, which is released into the disk membrane and then chemically
modified and transported to an RPE cell. Defects in the ABCA4 protein,
however, slow the chemical modification of all-trans retinal, resulting in a
buildup of this compound. All-trans retinal spontaneously reacts at a low rate
with membrane lipids to form a substance known as A2E. Sparrow et al. have
provided evidence that A2E impairs RPE viability by sensitizing the cells to
light damage (59).
Earlier work suggested that ABCA4 produced a retina-specific
transporter protein exclusively confined to the disk membrane of rod OSs (3).
Yet clinically, STGD1 can severely impact central macular function. In the
foveal region, cone photoreceptor density approximates 200,000 cells/mm2.
These densely packed neuronal cells are primarily cones but have
specialized architecture that resembles rods (53). Molday et al. have
suggested that the ABCA4 protein is present in both rod and cone
photoreceptors (7). They conclude that the loss in central vision experienced
by many STGD1 patients arises directly from ABCA4-mediated foveal cone
degeneration. Scholl et al. showed altered electrophysiology in both the rod
and cone systems in patients with genetically confirmed STGD1 (60).
Genetics
STGD1 is an autosomal recessive disease caused by biallelic mutations in the
ABCA4 gene. However, in multiple international cohort studies, complete
sequencing of the ABCA4 locus in STGD1 patients identifies two expected
disease-causing alleles in approximately 75% of patients and only one
mutation in approximately 15% of patients. Thus, approximately 10% of
ABCA4 mutations remain to be determined. One of the hallmarks of ABCA4
and STGD1 is the aforementioned extensive phenotypic variability; in age of
onset, rate of progression, disease types, attained severity and remaining
vision. Phenotypic variability is caused in large part by the extensive ABCA4
disease-associated genetic variation, as more than 1,000 definitely or possibly
disease-causing variants have been determined in the coding sequences and
splice sites of the ABCA4 gene.
It is important to identify the remaining 25% of disease causing ABCA4
mutations. Recently, the understanding of disease-causing genetic variations
in ABCA4 improved as a result of two major advances (44). First, many
noncoding, (outside of the exons) disease-associated ABCA4 alleles have
been identified and proven to be pathogenic, mostly by affecting splicing.
Splicing is the molecular mechanism that removes introns and joins together
exons as the DNA is transcribed into messenger RNA. Second, it was
recently determined that some ABCA4 variants, which had been considered
benign because of high minor allele frequency (MAF) in the normal, general
population, are in fact very mild conditional alleles. These mild mutations
result in STGD1 disease only when in trans (trans means in an association
with a mutation on the other allele) with a deleterious mutation. These mild
mutations are now called “extreme hypomorphs” (41). Deep intronic and
hypomorphic alleles have added to our understanding of ABCA4 and allowed
an increased number of patients to be designated “definitely caused by
ABCA4,” important for entry into studies and therapeutic trials. Even in 2018,
21 years after ABCA4 discovery, new ABCA4 mutations are still identified,
likely because there are important geographical isolates. Nassisi et al. (61)
collected DNA samples of 397 index subjects from France and analyzed the
exons and flanking intronic regions of ABCA4 (NM_000350.2) by microarray
analysis and direct Sanger sequencing. At the end of the screening, at least
two likely pathogenic mutations were found in 302 patients (76.1%) while 95
remained unsolved: 40 (10.1%) with no variants identified, 52 (13.1%) with
one heterozygous mutation, and 3 (0.7%) with at least one variant of
uncertain significance (VUS). Sixty-three novel variants were identified in
the cohort. Three of them were variants of uncertain significance. The other
60 mutations were classified as likely pathogenic or pathogenic and were
identified in 61 patients (15.4%). The majority of those were missense (55%)
followed by frameshift and nonsense variants (30%), intronic (11.7%)
variants, and in-frame deletions (3.3%). Clinical evaluation of each subject
confirms the tendency that truncating mutations lead to a more severe
phenotype than missense mutations with more severe electroretinographic
(ERG) impairment (P = 0.002) and an earlier age of onset (P = 0.037) (61).
Out of 36 STGD1 probands with one or no ABCA4 variants found, in 24
(67%), deep-intronic variants were identified (62). Five novel variants
resulted in pseudo-inclusions of messenger RNA due to strengthening of
cryptic splice sites or by disrupting a splicing silencer motif, analyzed, and
demonstrated by using in vitro splice assays in HEK293T cells and patient-
derived fibroblasts. Remarkably, antisense oligonucleotides targeting the
aberrant splice processes resulted in (partial) correction of all splicing
defects.
GENETIC TESTING FOR PATIENTS
WITH STGD1
Commercially available Stargardt genetic testing now utilizes both
microarray testing and sequence analysis of the entire coding region of the
ABCA4 gene, and more than 1,000 mutations have been identified. Many
gene testing panels include phenocopies of Stargardt such as pattern
dystrophy, cone–rod dystrophy, and genes for other types of early-onset
maculopathy.
ABCA4 is highly polymorphic, making interpretation of genetic results
complex. There are five possible types of genetic variants that may be
discovered:
1. Pathogenic, when the variant is truncating a protein product.

2. Likely pathogenic, when the variant is nontruncating and enriched in the
ABCA4—Leiden Open Variation Database (LOVD) dataset compared
to the nFE ExAC control group.
3. Uncertain significance, when the variant is nontruncating, more frequent
in the ABCA4-LOVD dataset than in then FE ExAC control group, but
not significantly enriched.
4. Likely benign, when the variant has a higher frequency in the nFE ExAC
population than in the ABCA4-LOVD dataset.
5. Benign, when the variant has a frequency >0.005 in the nFE ExAC
population and is not one of the known mild variants p.(Gly863Ala,
Gly863del) or p.(Gly1961Glu).
Cornelis suggests categorization of pathogenicity into the following four

groups: (a) Pathogenic, very strong criterion (PVS), for example, truncating.
(b) Pathogenic, strong criterion (PS), for example, another variant leading to
the same amino acid change has been associated with disease. (c) Pathogenic,
moderate criterion (PM), for example, the variant is located in a well-
established functional domain. (d) Pathogenic, supporting criterion (PP), for
example, all in silico prediction programs predict pathogenicity (63).
WORLDWIDE IMPACT
STGD1/FF affects males and females equally and spans all ethnic
populations. Various ethnic populations have been found to harbor specific
allelic variations in the ABCA4 gene that are disease causing and more
common in one ethnic group than another. Nondisease causing
polymorphisms may be overrepresented in some ethnic groups as well, so
great care must be taken to compare to normal controls of the same
geographic/ancestral background when estimating the pathogenic probability
of novel alleles.
MANAGEMENT OF STGD1 PATIENTS

AND VISUAL REHABILITATION
School-Age Issues
In children and teens with STGD1, it is very important to employ adequate
low-vision care and modify behavioral and environmental factors to
maximize visual performance. For example, these students may have light
and glare control problems in their classrooms. Adaptations, such as sitting
away from the window or shutting the curtains, may be necessary. Moreover,
these students should have permission from school administrators to wear
special sun filters (sunglasses or brimmed caps) in classrooms and on campus
when needed to decrease glare and light sensitivity.
Because the vision loss may progress over time, each student should have
a teacher of the visually impaired (TVI). A TVI can assess the classroom and
educational plan for the child. With the low-vision specialist’s
recommendations, the TVI makes adaptations including low-vision devices,
large-print materials, closed circuit TV, iPad linked to Smartboard, and other
special services the child will need each year in school. Some students may
require extended time on reading assignments and tests. Bifocals may be
useful for near work. Computer software is also available to convert standard
programs into more user-friendly formats for people with low vision.
Mobility is usually minimally affected but should be evaluated periodically
by an orientation and mobility specialist.
Eccentric Fixation: Eccentric Fixation and Preferred

Retinal Locus in STGD1
The preferred retinal locus from the fovea (PRL) is defined as the distance in
degrees between the center of gravity of all recorded fixation events and the
foveal center (see Figure 30-13).
FIGURE 30-13 Three images of the right eye of a young

woman with STGD1; the left photo has the preferred
retinal locus (PRL), where fixation occurred, mapped by
the MP-1 microperimeter and marked in blue. In the center
photo a grid is placed around the PRL to measure distance
from the fovea. Note that the preferred fixation is superior
to the anatomic fovea. On the fundus autofluorescent
image on the far right, the PRL is marked with a red cross
and the area of foveal atrophy can be more clearly seen.
((Reprinted from Schonbach and ProgStar Study Group et
al. Longitudinal changes of fixation location and stability
within 12 months in Stargardt disease: ProgStar report no.
12. Am J Ophthalmol 2018;193:54–61. Copyright © 2018
Elsevier. With permission.)
One of the most characteristic primate reflexes in the visual system is the
fixation reflex at around week 6 after birth. Without the development of this
reflex, nystagmus ensues. Foveal fixation is made possible by several waves
of poorly understood events, including centrifugal migration of the retinal
inner layers (including the NFL, ganglion layer, inner plexiform, inner retinal
layer, and outer plexiform layers) and foveal avascular zone development and
cone packing. STGD1 patients develop this reflex but lose the foveal cones
early in the disease process. The foveal reflex mechanism learned in infancy
forces the STGD1 patient to continue to put the visual image on the atrophic
fovea, a deep central scotoma. Many patients slowly, automatically learn to
diverge from this learned fixation location and develop a PRL outside the
atrophic lesion. Many patients have a superior PRL, for reasons still
unknown. Some STGD1 patients need help in discovering their PRL. Low
vision specialists can assist with learning the PRL. Reinhard et al. show that
the majority (70%) of STGD1 patients, who fixated eccentrically, used a PRL
that was located above the lesion on the retina, which is below the scotoma in
the visual space (64).
Bivariate Contour Ellipse Area

The bivariate contour ellipse area (BCEA) is the smallest ellipse that
encompasses 1, 2, or 3 standard deviations of all fixation events. The new
microperimeters (MAIA, MP-1) can fully automatically draw this ellipse
around the recorded fixation points and calculate the area as shown in the
image. A smaller BCEA corresponds to more stable fixation (see Figure 30-
14).
FIGURE 30-14 The BCEA is calculated and represents

the ellipsoid area of PRL with SD, showing that the
distance of the BCEA from the center of the fovea can
vary over time in tehe same patient.
(Reprinted from Schonbach and ProgStar Study Group, et al. Longitudinal changes of fixation location
and stability within 12 months in Stargardt disease: ProgStar report no. 12. Am J Ophthalmol
2018;193:54–61. Copyright © 2018 Elsevier. With permission.)
Schonbach et al. evaluated and measured longitudinal aspects of PRL and

BCEA in a large group of STGD1 patients. Surprisingly, the measures were
not stable and showed transient movement around the atrophic foveas (65).
Role of other physicians and healthcare providers
Patients with STGD1/FF are advised to avoid excess intake of vitamin A;
therefore, they may benefit from a dietician consult or discussion with their
primary care doctor who can educate about vitamin A rich foods and monitor
so vitamin A deficiency does not develop. Some clinical trials for Stargardt
disease involve the use of medications that are teratogenic and, therefore,
require the use of birth control while on the medication. Communication with
other healthcare providers about these and other issues is necessary for the
overall well-being of patients. Many patients with STGD1 are able to obtain a
driver’s license; this is discussed further in Chapter 14.
Because this is an AR disorder, parents of an affected child have a 25% risk
with each subsequent child that the child may be affected. Formal genetic
counseling detailing the risks should be offered. IVF with preimplantation
genetic testing of the embryos before implantation can be considered if there
is a molecular genetic diagnosis prior to initiation of pregnancy. Affected
patients are obligate carriers and will always pass on one mutant allele to all
of their offspring but cannot pass on the disease itself; this will only happen if
their partner is also a carrier. Because AR STGD1 is caused by only one
gene, carrier testing of partners can be accomplished and can be used to
provide accurate information to patients and their partners. It is the ethical
responsibility of physicians caring for patients with STGD1 to explain their
genetic risks and/or to refer them for in depth genetic counseling. There is
also an obligation to inform patients about the possibility of enrolling in
clinical trials. Providing patients with the address for www.clinicaltrials.gov,
and offering referral to a genetic eye disorder center, gives patients the ability
to learn more about their options www.clinicaltrials.gov does not vet clinical
trials, however, so it is wise to advise patients to discuss trials they are
considering with a knowledgeable physician.
FUTURE TREATMENTS FOR STGD1

AND ABCA4 PATIENTS
There are at this time (2019), no commercially available treatments for
STGD1, but many promising treatment regimens have been tested
successfully in animals and are currently undergoing efficacy and safety
studies in humans. There are currently nine separate, active therapeutic
studies in humans for STGD1 in various stages of intervention and
development, that is, seven pharmacologic interventions, one gene
replacement, and one stem cell intervention as of this writing (May 2019)
(www.clinicaltrials.gov). It is unimaginable that Professor Karl Stargardt
could have foreseen the flurry and depth of activities surrounding the disease
named after him.
Animal Therapeutic Studies

In 2003, Radu and coworkers showed that in the Abca4 knockout mouse,
increasing ambient light exposure resulted in an increase in toxic-oxidized
A2E within the retinal pigment epithelial cells (66). This accumulation of
oxidized A2E was strongly suppressed by treating the mice with isotretinoin,
an inhibitor of rhodopsin regeneration. The same group of investigators
showed that the use of fenretinide, by binding to retinol binding protein,
reduced serum vitamin A levels and arrested the accumulation of A2E and
lipofuscin autofluorescence in retinal pigment epithelial cells in the knockout
mouse model for Stargardt disease (24).
Isotretinoin has been shown to slow the synthesis of 11-cis-retinaldehyde
and regeneration of rhodopsin by inhibiting 11-cis-retinol dehydrogenase in
the visual cycle. Recently, the effects of isotretinoin on lipofuscin
accumulation in Abcr−/− knockout mice were studied. Isotretinoin appeared
to block the formation of A2E and the accumulation of lipofuscin pigments,
and ERG parameters remained stable. Isotretinoin also blocked the slower,
age-dependent accumulation of lipofuscin in wild-type mice. These results
suggest that treatment with isotretinoin may inhibit lipofuscin accumulation
and thus delay the onset of visual loss in STGD1/FF patients (66).
Observational and experimental data suggest that antioxidant and/or zinc
supplements may delay progression of ARMD and vision loss, but this has
not been proven for Stargardt (67). Biochemical studies on the mechanism of
oxidative damage to the RPE cell may lead to other therapeutic interventions
(68).
The eye is a unique immune-privileged organ that is well suited for gene
therapy. The positive results of clinical trials of gene therapy for RPE65-
associated Leber congenital amaurosis show potential for gene therapy as an
effective treatment for other hereditary retinal degenerations. The currently
preferred method for nucleic acid delivery is a viral vector. When considering
STGD1, the ABCA4 coding region (cDNA) is very large, which limits its
ability to be packaged into an adeno-associated virus (AAV). Two vectors
that have the capacity to package ABCA4 are gutted adenovirus and
lentivirus; another possible approach is dividing the cDNA into two pieces,
carried by dual AAV vectors. Studies in animal models of STGD1 have
shown efficacy and safety of gene therapy and have begun to set the stage for
clinical trials of gene therapy in humans with STGD1 (69). Gene therapy
trials for STGD1 may be identified on www.clinicaltrials.gov.
Stem cell therapy may offer a future treatment for STGD1. Human
embryonic stem cells (ESC) have been used to create RPE cells. Preliminary
experimentation of pars plana vitrectomy with injection of these cells into the
subretinal space of the macula along with systemic immunosuppression
showed survival and assimilation of the stem cell–derived cells into animal
models (70,71). These techniques offer future hope with greater research.
There is an excellent review of all currently (as of May 2018) available
experimental treatments for STGD1 summarized by the group of Professor
Byron Lam (72).
Currently, six therapeutic approaches to altering the disease course of
STGD1 are being investigated to slow the disease course, halt the disease
course, or improve visual function.
Approach 1: Decrease the Toxic By-products of

ABCA4 Disease
Stoichiometrically, replacing the C20 hydrogen atoms of vitamin A with
deuterium atoms (i.e., C20-D3-vitamin A) predicts that this bond may be
harder to cleave and may impede vitamin A dimerization, preventing toxic
bisretinoid formation (73). Slowing the intrinsic reactivity of vitamin A to
dimerize could slow lipofuscin formation in the RPE. ALK-001 (C20-D3-
vitamin A) administered to Abca4−/− mice showed an 80% reduction in A2E,
a 95% reduction in ATR-dimer, and a 70% decrease in FAF at 3 months of
age. After 6 months, the treated group showed fewer lipofuscin granules, and
at 12 months, they showed improved ERG (74). These encouraging data have
prompted a clinical trial in humans.
Approach 2: Modulation of the Visual Cycle

Vision Medicine’s VM200 molecule for STGD1 is an oral aldehyde trap that
sequesters the toxic compound, all-trans retinal, to potentially prevent retinal
cell death by making it unable to form A2E. VM200 preserves retinal
structure in Abca4−/− Rdh8−/− mice, as measured by in vivo retinal
architecture by OCT. VM200 may also preserve retinal function, as an
increased concentration of 11-cis-retinal was found in mice (75). A human
trial is in the works.
Accutane Oral Treatment Studies
Isotretinoin (Accutane) inhibits 11-cis-RDH in the visual cycle, thus
slowing the synthesis of 11-cis-retinal and thereby slowing the regeneration
of rhodopsin. Abca4−/− mice were injected with isotretinoin and responded by
decreased production of A2E, along with less formation of lipofuscin
granules in the retina compared to controls (24). The hypothesis is that
isotretinoin would delay visual loss in young patients with STGD1, but this
has not yet been tested in humans.
Inhibition of RPE65 (Emixustat)

Emixustat (also known as ACU-4429, developed by Acucela Inc) is a small
nonretinoid that inhibits retinoid isomerohydrolase, the enzyme encoded by
RPE65. This inhibition reduces the conversion of all-trans-retinyl ester to 11-
cis-retinol and prevents the accumulation of A2E. In a phase 2/3 trial testing
geographic atrophy growth rate changes and visual acuity in AMD patients,
emixustat was not significantly different from placebo and did not alter the
disease course. A new trial is planned to study STGD1 patients (76).
Retinol-Binding Protein (RBP4) Antagonists:

Fenretinide (a Retinoid)
The receptor RBP4 is required for influx of serum retinol from the circulation
(after meals) into the RPE to participate in the visual cycle and the ultimate
formation of 11-cis-retinal for visual capture of photons in both rods and
cones. Formation of the tertiary retinol-binding protein 4 (RBP4)–
transthyretin–retinol complex in the serum is required for this influx to occur
successfully. Fenretinide (from Sirion Therapeutics) is an oral synthetic
retinoid derivative that competes with all-trans retinol (derived from the
capture of light by 11-cis-retinal) for its binding to the retinol-binding protein
4 (RBP4), thereby preventing transport into and lowering the levels of retinol
in the RPE cells.
Fenretinide has been shown to reduce formation of A2E in a mouse
model of STGD1 (48). However, fenretinide may induce apoptosis and may
be teratogenic. Despite this, a study was performed on AMD patients with
geographic lesions. Fenretinide appeared to slow down the annual growth
rate of GA, but the differences with controls were not significant. Also the
incidence of neovascularization was lower in the fenretinide group but not
significant, and there was no change in VA. At least twenty percent of
patients experienced adverse effects (77).
Retinal-Binding Protein-4 Antagonist: BPN-14136 (A

Nonretinoid)
BPN-14136 is another, novel RBP4 antagonist and is a nonretinoid, without
the apoptotic and teratogenic potential of fenretinide (78). Eyecup extracts of
12-week-old light- or dark-adapted BALB/cJ mice that had received BPN-
14136 were tested for RBP4 reduction, followed by retinoid cycle reduction.
The 2-week BPN-14136 treatment induced a 90% serum RBP4 reduction and
a 40% to 50% decrease in visual cycle retinoids (retinyl ester, 11-cis-retinal,
and all-trans-retinal), suggesting an excellent efficacy profile in mice. As
excessive production of lipofuscin bisretinoids is likely the major
biochemical trigger of monogenic Stargardt disease caused by recessive
mutations in the ABCA4 gene, BPN-14136 was tested in the ABCA4−/− mouse
model. Visual cycle retinoids, such as all-trans- and 11-cis-retinaldehyde,
serve as precursors in the biosynthesis of cytotoxic bisretinoids; thus, BPN-
14136 was tested to evaluate its ability to lower both groups of molecules.
BPN-14136 administration elicited the desired 40% to 50% partial reduction
in total concentration of visual cycle retinoids in dark-adapted mice, as well
as in light-adapted animals. BPN-14136 induced a sustained serum RBP4
reduction over the treatment period, which translated into the inhibition of
A2E formation by 50% in the treated animals. Parallel studies by biochemical
analyses of A2E, with quantification of lipofuscin autofluorescence
established that BPN-14136 also induced a 75% reduction of lipofuscin
fluorophores in the treated Abca4−/− mice.
Finally, in the Abca4−/− mouse model, enhanced accumulation of
lipofuscin bisretinoids induces dysregulation of the complement system in the
retina and complement-mediated inflammation in the RPE. BPN-14136
treatment normalized bisretinoid-induced up-regulation of complement factor
D (CFD) and C3 in the mouse Abca4−/− retina. Similarly, levels of C-reactive
protein up-regulated in the Abca4−/− mouse retina were reduced. Expression
of complement factor H (CFH), which was partially suppressed in the
Abca4−/− mouse retina, was restored following administration of BPN-
14136.
Approach 3: Modulation of the Complement Pathways

(i.e., Complement Inhibition)
Abca4−/− (STGD1) mice exhibit buildup of bisretinoid-containing lipofuscin
pigments in the RPE and increased oxidative stress, augmented complement
activation, and slow degeneration of photoreceptors (79). A reduction in
complement negative regulatory proteins (CRPs), possibly due to bisretinoid
accumulation, may be responsible for the increased complement activation
seen in the RPE of STGD1 mice. They found that CRPs prevent attack on
host cells by the complement system, and complement receptor 1-like protein
y (CRRY) is an important CRP in mice. CRRY expression was increased in
the RPE of mice by gene therapy. A sustained, several-fold increased
expression of CRRY in the RPE was documented, which significantly
reduced the complement factors C3/C3b in the RPE. It also showed slower
photoreceptor degeneration (79). This suggests that complement attack on the
RPE is an important etiologic factor in STGD1.
Approach 4: Nutritional Studies

MacDonald et al. studied DHA supplementation in a small cohort of STGD1
patients over a period of 12 months. MfERG, VA, HVF, and color confusion
were tested, while microperimetry of the macula, and in vivo OCT of the
macular structure and fundus auto fluorescence (FAF) were not. In the
primary endpoint for this study, mfERG, no changes in peak amplitude were
observed between periods of placebo and DHA supplementation. The other
secondary outcomes, such as visual acuity, visual field, and color confusion,
also led to the conclusion that macular function was not modified (80).
MfERG may not be an ideal endpoint for this study, as it is variable and
subject to micromovements of the eye and retina not registered by the
machine.
Approach 5: Stem Cell Studies

ESC are a potentially valuable source of donor cells for therapeutic repair and
regeneration of many atrophic retinal degenerations, including STGD1 and
AMD. ESCs’ pluripotency and innate capacity for self-renewal offer a
virtually unlimited supply of highly specialized retinal cells for therapeutic
transplantation. Caution is warranted, as there is also present a potential risk
of harm from uncontrolled proliferation and unintended differentiation.
ESCs’ impact in experimental models has been investigated extensively, but
evidence of their safety and potential efficacy in humans is limited. Mehat et
al. performed a phase 1/2 open-label multicenter dose-escalation trial in 12
adults (age range: 34 to 53 years old) with STGD1 and biallelic ABCA4
mutations to evaluate the safety and tolerability of subretinal transplantation
of hESC-derived RPE cells (70). They administered subretinal hESC-RPE
cells to a target area extending centrifugally from relatively well-preserved
functional retina across a transitional zone of progressive degeneration to an
area of atrophic nonfunctional retina. This would determine the safety of the
transplanted cells in relatively healthy retina would provide data on potential
benefit for function and survival of overlying photoreceptor cells in the
degenerating and atrophic areas. Retinal structure and function were
evaluated using microperimetry and SD OCT. No significant benefit to
retinal function was noted at 12 months.
Approach 6: Gene Replacement of ABCA4 (Gene

Therapy for STGD1)
The ABCA4 gene (molecular size 6.8 kb) far exceeds the 4.5 to 5.0 kb
capacity of the AAV vector, leading to utilization of an equine infectious
anemia lentivirus (EIAV) for gene transfer.
Subretinal injection of lenti-ABCA4 by Kong et al. in 2008 was found to
be effective in the knock out Abca4−/− mouse model, in which treated eyes
had a significant reduction of A2E concentrations compared to untreated and
mock-treated (lenti-null vector) control eyes. Treated eyes of Abca4−/− mice
accumulated 8 to 12 pmol per eye of A2E 1 year after treatment, amounts
similar to wild-type controls, whereas mock-treated or untreated eyes had 3 to
5 times more A2E (27 to 39 pmol per eye, P = 0.001 to 0.005) (81).
Oxford Biomedica, in coordination with the Sanofi group, are sponsoring
an escalating dose phase I/II clinical trial of SAR422459, formerly known as
StarGen (NCT 01367444). Moore et al. (82), are investigating safety and
preliminary signs of efficacy in STGD1 over a 48-week follow-up period.
The trial, which plans to enroll an estimated 46 patients, began in June 2011
and has an expected completion date in November 2019. Another study
(NCT01736592) is assessing the long-term safety and tolerability for the
original SAR422459 trial, in which these patients are followed without
further intervention for 15 years (82).
McClements et al. showed that the efficacy of recombination of dual
vectors is dependent on the length of DNA overlap between two transgenes.
With optimized recombination, full-length ABCA4 protein is expressed in
the photoreceptor OSs of Abca4−/− mice at levels sufficient to reduce
bisretinoid formation and correct the autofluorescent phenotype (83).
More detailed mechanistic studies and therapeutic investigations are now
possible, because of the discovery of a canine ABCA4 model (84), more
closely representing the human disease, and the recent development of
induced pluripotent stem cells (iPSCs) containing 2 ABCA4 mutations from
reprogrammed human fibroblast cells using the Yamanaka technique (85).
SUMMARY
In summary, STGD1 is a major childhood and adult hereditary retinal
degeneration. The causal ABCA4 gene is involved in retinal health,
maintenance, and physiology and when mutated causes a spectrum of retinal
degenerations. The story of the description of this retinal degeneration, the
subsequent discovery of the causative gene and disease pathway, and the
progress toward treatments are representative of the strides being made for
patients with blinding retinal diseases in the molecular genetic era.
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31
Mitochondrial, Peroxisomal,
Glycosylation Disorders
Samer Khateb, Ann Saada, Patrick Yu-Wai-Man, John B. Kerrison,
and Itay Chowers
MITOCHONDRIA
Mitochondria are intracellular, dynamic organelles present in all nucleated
cells that are essential for maintaining cellular metabolism and homeostasis.
They regulate a number of metabolic processes involving steroid and heme
biosynthesis; pyruvate, urea, nucleotide, and sulfur metabolism; and fatty
acid and amino acid degradation. Mitochondria also play critical roles in
calcium homeostasis, caspase-dependent apoptosis, the cellular stress
response, and protein degradation. However, their central function is the
production of cellular energy in the form of adenosine triphosphate (ATP).
This process of oxidative phosphorylation (OXPHOS) is carried out by the
five multisubunit respiratory chain complexes that are embedded within the
mitochondrial inner membrane (IM). Complexes I to IV transfer electrons
with oxygen as the terminal electron acceptor, and the energy released is used
to establish an electrochemical proton gradient across the IM, which, in turn,
is utilized by complex V (ATP synthase) to phosphorylate ADP to ATP.
A unique feature of mitochondria is that they harbor their own genetic
code in the form or a small, circular DNA molecule (16.5 kb), which is a
legacy from the organelle’s endosymbiotic origin. The number of
mitochondria in a cell is dictated by its energetic demand, and each
mitochondrion contains several mitochondrial DNA (mtDNA) molecules,
resulting in a high copy number. As a result, there can be a combination of
both wild-type and mutant mtDNA, resulting in a situation known as
heteroplasmy. In some cases, the level of the mutant mtDNA species
correlates with the severity of the resulting clinical phenotype. The
mitochondrial genome encodes for 13 OXPHOS proteins, 2 ribosomal RNAs,
and 22 transfer RNAs involved in mitochondrial protein synthesis. The
remaining subunits (~70) are encoded by the nuclear genome, including all
components of complex II. The majority of proteins (~1,500) that constitute
the mitochondrial machinery are nuclear-encoded, and following import into
mitochondria, these mediate a host of function including mtDNA replication,
transcription, and translation. The mitochondrial genome is maternally
inherited, whereas nuclear-encoded mitochondrial genes will be inherited in a
Mendelian pattern, which can complicate molecular diagnosis and genetic
counseling (1–3).
Mitochondrial Disorders
Mitochondrial disorders are genetically and phenotypically highly variable,
with patients presenting with either isolated or multisystem organ
involvement (4). About half of all patients will manifest neuro-
ophthalmologic features, in particular involvement of the optic nerve, outer
retina, and extraocular muscles (5,6). Leber hereditary optic neuropathy
(LHON; OMIM #535000) is a classical mitochondrial optic neuropathy
caused by mtDNA point mutations. The importance of mitochondria for
normal optic nerve function is further highlighted by autosomal dominant
optic atrophy (ADOA; OMIM #165300 and #165500) caused by mutations in
nuclear genes encoding for the OPA1 and OPA3 mitochondrial proteins.
Progressive degeneration of the outer retina is seen in a number of
mitochondrial syndromes, such as (a) the Kearns-Sayre syndrome (KSS;
OMIM #530000) (7–9) in combination with chronic progressive external
ophthalmoplegia (CPEO; OMIM #258450, #609286) (10–13); (b)
mitochondrial myopathies without external ophthalmoplegia (14,15); (c) the
neuropathy, ataxia, and retinitis pigmentosa (NARP; OMIM #551500)
syndrome (16–18); and (d) Leigh syndrome marked by involvement of the
basal ganglia and brainstem (19,20). The retinopathy seen in patients with
mitochondrial disease can be generalized with primary involvement of the
peripheral retina or localized to the macula. On fundus examination, the
pigmentary disturbance can resemble the typical bone spicule appearance
seen in retinitis pigmentosa with choriocapillaris atrophy, or it can be more
variable with areas of retinal hyperpigmentation and/or hypopigmentation
(14,21–23).
LEBER HEREDITARY OPTIC
NEUROPATHY
Prevalence and Worldwide Impact
LHON is the most common primary mtDNA disorder in the general
population with an estimated prevalence of 1 in 25,000 to 1 in 40,000 in
northern Europe (21,24). This phenotype was noted initially by Albrecht von
Graefe (1858) (25) and then characterized further by Theodor Leber (1840–
1917) as a familial neuro-ophthalmologic disease (26).
Environmental Factors and Genetics

The majority (~90%) of LHON cases are due to one of three mtDNA point
mutations, namely, m.3460G>A (MT-ND1), m.11778G>A (MT-ND4), and
m.14484T>C (MT-ND6), with the m.11778G>A being the most common
mutation worldwide (60% to 90%) (27–29). The remaining cases are caused
by rarer mutations that mostly occur in mutational hotspots within mtDNA
genes encoding for complex I (NAD–coenzyme Q reductase) subunits (30)
(http://www.mitomap.org/bin/view.pl/MITOMAP/MutationsLHON). It is not
unusual for patients to fail to report a family history of LHON. The
percentage of probands reporting other affected family members is 78%,
43%, and 65% for the m.3460G>A, m.11778G>A, and m.14484T>C
mutations, respectively (31,32). Strikingly, LHON shows incomplete
penetrance and a marked sex bias with male carriers being much more likely
to lose vision compared with female carriers. Although there is considerable
variability both between and within families, overall, the penetrance of
LHON is about 50% for male carriers and 10% for female carriers. The
presence of the mtDNA mutation is necessary, but not sufficient on its own to
trigger visual loss, implying the involvement of additional genetic and
environmental factors as disease modifiers (29,33,34).
Although a genetic modifier on the X chromosome is an attractive
hypothesis, linkage analysis has failed to identify an X-linked nuclear
susceptibility gene to account for the male predominance seen in LHON
(35–37). Hormonal factors could also contribute to this sex bias with
estrogens having a neuroprotective effect in the LHON cell models studied
(38,39). The activation of mitochondrial biogenesis has been invoked as a
compensatory mechanism based on the fact that unaffected LHON carriers
have an increased mtDNA copy number compared with affected patients
(40). In addition to nuclear genetic factors, the mtDNA background defined
by specific haplogroups is also thought to influence the risk of vision loss in
this mitochondrial disorder (41). The strongest risk factors associated with an
increased risk of vision loss among LHON carriers are smoking and to a
lesser extent excessive alcohol consumption (32,42,43).
Pathophysiology
Central to the pathophysiology of LHON is OXPHOS dysfunction with
impaired assembly and functioning of complex I of the mitochondrial
respiratory chain (44). Studies performed on various tissues from patients
with LHON, including lymphocytes, platelets, fibroblasts, and muscle
biopsies, have confirmed reduced complex I–driven mitochondrial respiration
with reduced ATP production, increased levels of reactive oxygen species
(ROS), and glutamate excitotoxicity (5,30,34,45,46). The generation of
retinal ganglion cells (RGCs) from induced pluripotent stem cells and the
development of more faithful animal models, which have proven challenging,
will provide further insight into the complex pathophysiology of LHON (47).

LHON is characterized by bilateral, painless, subacute vision loss with both
eyes being affected simultaneously in about 25% of cases. If only one eye is
affected initially, the fellow eye usually becomes involved within the next 2
to 4 months. The peak age of onset is in the second and third decade of life,
but disease conversion can occur at any age from early childhood to late
adulthood (31,48). The prognosis is poor with visual acuities usually worse
than 20/200 and bilateral dense central or centrocecal scotomas that have a
major impact on visual functioning and quality of life (30,32,49). The final
visual outcome is influenced by the underlying mtDNA mutation with the
m.14484T>C mutation carrying the best visual prognosis (50–52).
Spontaneous recovery occurs in some patients and typically happens within
the first year of disease onset if it is going to occur. The likelihood of
clinically relevant recovery is greatest with the m.14484T>C mutation (37%
to 58%), least with the m.11778G>A mutation (4% to 25%), and intermediate
with the m.3460G>A mutation (22% to 25%) (27,51,53,54). The prognosis is
also better for patients with onset of vision loss in childhood before the age of
12 years old (55).
In the acute phase, the optic nerve typically looks hyperemic with
peripapillary telangiectasia, swelling of the retinal nerve fiber layer, and
tortuosity of the central retinal vessels (56) (Figure 31-1). In about 20% of
cases, the optic nerve looks entirely normal, which can lead to diagnostic
delays and initial labeling as functional visual loss. As the disease progresses,
the swelling of the retinal nerve fiber layer starts to subside, and there is
development of optic atrophy with pallor of the neuroretinal rim and
nonglaucomatous cupping in a proportion of cases (57). Interestingly, some
unaffected LHON carriers can show fluctuating areas of segmental retinal
nerve fiber layer swelling and subclinical psychophysical abnormalities,
namely, loss of color vision affecting mostly the red-green system, reduced
contrast sensitivity, and subnormal electroretinogram (ERG) and visual
evoked potential (VEP) recordings (58).
Figure 31-1 A 26-year-old woman presented with acute
painless visual loss in her right eye. She had a known
family history of LHON secondary to the m.11778G>A
mtDNA mutation. Her best-corrected visual acuities were
20/400 in the right eye and 20/20 in the left
(asymptomatic) eye at baseline (top images). The right
fundus showed the classical changes seen in acute LHON
with optic disc hyperemia, peripapillary telangiectasias,
and tortuosity of the central retinal vessels (A). The left
fundus was normal (B). Humphrey perimetry showed a
cecocentral scotoma in the right eye (C) and a normal
visual field in the left eye (E). On the optic disc OCT
profile, RNFL thickening was observed in both eyes
(D,F). The left eye became clinically affected 1 month
later. Two years later (bottom images), the patient’s best-
corrected visual acuities were hand movement in both
eyes, and both optic nerves were atrophic with global
pallor (A,B). Humphrey perimetry showed bilateral dense
central scotomas (C,E) and there was significant thinning
of the peripapillary RNFL (D,F).
Systemic Manifestations
The majority of patients with LHON will develop isolated optic atrophy due
to the preferential susceptibility of RGCs within the inner retina. However,
extraocular neuromuscular features have been reported in a small subgroup of
patients with LHON, including cerebellar ataxia, movement disorders,
myopathy, and peripheral neuropathy (59,60). Dystonia and myoclonus are
two movement disorders that have been consistently associated with the three
primary mtDNA LHON mutations (61). Another well-characterized
syndromic association is the development of a multiple sclerosis (MS)-like
illness in patients with LHON. This LHON-MS overlap syndrome, or
Harding disease, was first reported in women carrying the m.11778G>A
mtDNA mutation, but it was subsequently described with the m.3460G>A
and m.14484T>C mtDNA mutations as well (62). Patients with the LHON-
MS overlap syndrome have a better visual prognosis compared with classical
LHON, but they usually have recurrent episodes of vision loss that can be
accompanied with ocular pain similar to demyelinating optic neuritis (63). It
should be stressed that establishing a true causal association has not always
been possible in so-called LHON-plus phenotypes, for example, the link
between LHON and an increased risk of cardiac arrhythmias is still uncertain
(57,61,64).
Diagnostic Studies
The differential diagnosis includes demyelinating, ischemic, compressive,
infiltrative, infectious, and neoplastic causes of a bilateral optic neuropathy.
A definitive diagnosis is made upon the identification of a pathogenic
mtDNA mutation known to cause LHON with molecular genetic testing.
Brain MRI, VEPs, flicker fusion, color vision testing, and visual field testing
in addition to visual acuity and intraocular pressure measurement should be
part of the workup of patients with optic atrophy. A careful personal and
family history, as well as nutritional history, is warranted.
Management
No established treatments are available at the present time to prevent vision
loss, arrest the progression of vision loss, or restore vision. However,
significant progress was made in recent years toward the development of
targeted treatments for LHON. Ubiquinone (CoQ10) is an electron essential
carrier within the inner mitochondrial membrane ensuring efficient transfer of
electrons along the mitochondrial respiratory chain from complexes I and II
to complex III. Idebenone is a short-chain ubiquinone analogue that crosses
the blood–brain barrier, and its safety and efficacy were investigated as part
of a randomized controlled trial (RHODOS) that recruited 85 patients
carrying the m.3460G>A, m.11778G>A, and m.14484T>C mtDNA
mutations (65). Although RHODOS failed its primary outcome measure, a
subgroup of treated patients was found to benefit, and this observation was
confirmed in a larger retrospective case series of 103 patients treated with
variable doses of idebenone (66). An expert panel has recommended that
idebenone should be started as soon as possible (at a dose of 300 mg three
times per day) for patients with LHON with visual loss of <1 year (67). This
regimen should be continued for at least 1 year to determine whether there is
a treatment effect, and if a clinically relevant response is observed, the
treatment should be continued for 1 year after the plateau phase has been
reached. The expert panel concluded that there was not enough evidence to
recommend treatment for chronic LHON cases. An open-label clinical trial
on five patients with LHON suggested that EPI-743, an orally absorbed small
molecule, which targets the enzyme NADPH quinone oxidoreductase 1, may
be beneficial in arresting visual loss in LHON (68,69). Another potential
future treatment technique for LHON is gene therapy. A gene-replacement
strategy based on allotopic expression has been described to correct the
defect in complex I function in cells harboring the m.11778G>A mutation
(70). The in vivo feasibility, efficacy, and safety of such a technique are
currently being evaluated in a number of clinical trials (39,71).
AUTOSOMAL DOMINANT OPTIC

ATROPHY
Autosomal dominant optic atrophy (ADOA), also known as Kjer disease, is
the most common inherited optic neuropathy in the population with an
estimated minimum prevalence of about 1:25,000 individuals based on
clinical and molecular genetic data (72).
Genetics and Pathophysiology

Most ADOA patients harbor mutations in the OPA1 gene, whereas fewer
cases of OPA3 mutations have been reported. The OPA1 gene consists of 31
exons spread over 100 kb of genomic DNA, and alternative splicing results in
8 isoforms. OPA1 encodes a dynamin-like mitochondrial GTPase, and many
of the over 250 pathogenic mutations reported cluster in the GTPase domain
and the dynamin central region involved in oligomerization of this protein
into cylindrical tubular structures and catalytic activation. Single base pair
mutations are the most common, followed by deletions and insertions. OPA1
testing by sequencing is useful, but some large-scale rearrangements may be
missed by this technique (24,73–76). OPA1 mutations are autosomal
dominant, that is, a mutation on just one allele confers disease and can be
passed directly from parent to child. However, compound heterozygous
OPA1 mutations have been reported in a small number of patients presenting
with a Behr-like phenotype (77). Notably, homozygous severe OPA1
mutations are probably not compatible with life or are severely deleterious
with multiple systemic associations as recently reported (78).
Pathophysiology
The OPA1 protein is most highly expressed in the retina but is also expressed
in the brain and muscle. It associates with mitochondrial IM phospholipids
such as cardiolipin and plays a pivotal role in the maintenance of
mitochondrial morphology. Specifically, OPA1 protein is involved in
mitochondrial fusion and the maintenance and modulation of the
mitochondrial cristae allowing adaptation to metabolic demands. OPA1 is
important for preserving the mitochondrial membrane structure and integrity
(79) and is also involved in mtDNA maintenance. Loss of OPA1 will,
therefore, lead to decreased mtDNA stability and decreased ATP production
leading to cell damage and apoptosis.
Studies suggest that patients with ADOA are born with fewer optic nerve
axons and smaller optic discs, supporting the hypothesis that subsequent
vision loss depends on further age-related loss of fibers, which also occurs in
control subjects. It may also suggest OPA1 involvement in eye development
(80). Interestingly, some mutations in DNML1, a protein involved in
mitochondrial fission (i.e., opposite function of OPA1), also result in ADOA
(81).
The pathologic mechanism of OPA3 mutations associated with the
autosomal recessive and dominant (less common) forms of 3-
methylglutaconic aciduria type III (Costeff syndrome) seems to be similar to
that of OPA1 mutations, as mutant OPA3 mice also showed disrupted
mitochondrial morphology in the retina confirming the importance of the
mitochondrial network in retinal function (74,82–84). Notably, other diseases
linked to abnormal mitochondrial morphology involving mitochondrial
membrane and mitochondria-associated ER membranes (MAMs), such as
Charcot-Marie-Tooth type 2A (MFN2), hereditary motor and sensory
neuropathy type VI (DST), hereditary spastic paraplegia (SPG7), and
Wolfram syndromes (WFS1 and CIDS2), also exhibit ocular manifestations
(5,85–88).
Clinical Symptoms and Signs

The classical presentation of autosomal dominant optic neuropathy includes
insidious childhood onset (between ages 4 and 6 years) presenting with
progressive bilateral vision loss, temporal pallor of the optic disc, and central
vision loss caused by selective loss of RGCs resulting in optic nerve
degeneration. The phenotype is generally milder than LHON, but the
expression is variable, including within families, ranging from subclinical
manifestations to legal blindness and mean visual acuities of 6/18 to 6/60
(89). Patients may show generalized dyschromatopsia, and isolated tritanopia
is uncommon (90). Visual field defects in ADOA reflect the tendency of the
disease for involvement of the papillomacular bundle, manifesting central,
cecocentral, or paracentral scotomata with preserved peripheral fields.
Similar to LHON, the pupil response to light is relatively preserved (24,91).
Potentially, OPA1 variants are also associated with the risk for developing
normal tension glaucoma in adulthood (92). Thickness mapping of retinal
layers using spectral domain optical coherence tomography (SD-OCT)
demonstrated significant thinning of perifoveal RNFL, GCL, inner plexiform
layer (IPL), and peripapillary RNFL in ADOA patients compared to healthy
subjects (93).
ADOA is characterized by the preferential loss of RGCs. However, OPA1 is
expressed also in the inner ear, and the most frequent extraocular
manifestations of OPA1 mutations are sensorineural deafness present in about
two-thirds of the cases (ADOAD). Additionally, up to 20% of patients with
OPA1 mutations will develop a more severe disease variant (ADOA “plus”)
with additional neuromuscular features including ataxia, myopathy,
peripheral neuropathy, CPEO, or MS-like disorders. Muscle pathology
resembles classical mitochondrial diseases with cytochrome c oxidase–
deficient fibers and ragged red fibers. Patients also harbor mtDNA deletions
in the affected muscle tissue. Notably, individuals with mutations that affect
the OPA1 GTPase domain are more likely to develop multisystem neurologic
disease (24,73,92,94). A recent study showed that classic ADOA patients
were more likely to harbor mutations in exons 8 and 9 and were more
commonly maternally inherited while ADOA-plus patients had mutations in
exons 14, 15, and 17 with being mostly paternally inherited (79).
Diagnostic Studies
The differential diagnosis includes all common causes of optic neuropathy:
LHON and other inherited optic neuropathies such as Leigh syndrome,
Wolfram syndrome, compressive, inflammatory, ischemic, and toxic; these
may be differentiated from ADOA by clinical examination; systemic
manifestations; family history; specialized testing, such as VEP; and, most
importantly, molecular genetic testing.
Management
Current management includes genetic counseling. A small retrospective case
series has explored the use of idebenone in patients with DOA carrying
confirmed pathogenic OPA1 mutations. The evidence presented is limited,
and a larger study is needed to define the therapeutic role of idebenone in this
group of patients (95). Future treatment may include better targeted oral
neuroprotective strategies and gene therapy (96).
KEARNS-SAYRE SYNDROME AND

CHRONIC PROGRESSIVE EXTERNAL
OPHTHALMOPLEGIA
While the exact prevalence of this condition is unknown, the prevalence is
estimated to be approximately 1 to 3 per 100,000 individuals. There is no
ethnic or gender predisposition to the disease (97).

A mitochondrial defect was suspected to underlie the KSS based on the
inheritance pattern observed in several affected families (98). In
mitochondrial inheritance, the pathologic variant is passed through the
maternal lineage to both males and females. Males cannot pass the mutation
on. Mitochondrial mutations or deletions are often sporadic, with no family
history and no mutation in the mother of the proband; however, the mutation
may be present in low levels in the mother or only in her oocytes. It was later
found that KSS, CPEO, and Pearson syndrome (99) are most commonly
caused by deletion of a part of the mtDNA (9,100,101). The deletions
typically are 1.3 to 7.6 kb in length, but there is an increased prevalence of
the 4.9-kb deletion. Interestingly, identical mtDNA deletions can lead to
either CPEO or KSS (with its additional systemic manifestations), and this
may at least in part depend on the distribution and extent of mtDNA deletions
in different tissues. The finding of sideroblastic anemia and pancreatic
dysfunction in an infant, followed by hearing loss, corneal endothelial
dysfunction, and retinal degeneration in those patients who survive infancy,
is a rare variant of KSS known as Pearson syndrome.
CPEO with multiple mtDNA deletions is also associated with mutations
in the nuclear genome and can be transmitted in autosomal dominant or
autosomal recessive modes of inheritance. However, sporadic, single large de
novo mtDNA deletions are typically not transmitted to the offspring.
Mutations in at least seven genes encoded by nuclear DNA cause autosomal
dominant CPEO, including POLG1, POLG2, ANT1, Twinkle (C10orf2),
RRM2B, DNA2, and OPA1 with POLG1 being the most common. POLG1
encodes the catalytic subunit of the mtDNA polymerase (polymerase
gamma), whereas the accessory subunit is encoded by the POLG2 gene. Over
150 POLG1 mutations have been associated with a wide range of
mitochondrial diseases presenting with a continuous spectrum of overlapping
symptoms and signs, ranging from fatal infantile disease to CPEO presenting
in adulthood (102,103) (http://tools.niehs.nih.gov/polg/), whereas mutations
in POLG2 are less common (104).
Most of the abovementioned genes are associated with mtDNA
replication and maintenance; thus, alterations in their function might lead to
mtDNA deletions (105). Genetic analysis in these patients usually finds
multiple concurrent mitochondrial mutations, suggesting that the nuclear
DNA mutations induce instability of mtDNA replication (12,106). Notably,
external ophthalmoplegia and other ocular manifestations are also found in
the autosomal recessive mtDNA depletion syndromes, caused by mutations
in genes encoding proteins of importance for mtDNA maintenance (TYMP,
DGUOK, TK2, MGM1, and RNASEH1) (107).

KSS is characterized by the triad of CPEO, pigmentary retinopathy, and heart
block (7,108). Onset is in the first or second decade of life. CPEO is
characterized by a bilateral symmetric ptosis associated with ophthalmoplegia
and orbicularis oculi weakness. In addition, KSS patients manifest an atypical
pigmentary retinopathy in which the macula is often the first part of the retina
to be affected followed by the peripheral retina. In some patients, all parts of
the retina are affected simultaneously. Macular lesion resembling adult-onset
vitelliform macular dystrophy and chronic subretinal fluid were also reported
in KSS cases previously (109,110). Histology of a retina from KSS patients
showed atrophy of the RPE and outer retina that was most marked posteriorly
(8). In addition to the different retinal involvement patterns, KSS also differs
from classic RP in that there is usually no blood vessel attenuation and no
waxy optic nerve appearance accompanying the pigmentary retinopathy.
Furthermore, visual acuity, visual fields, and ERGs are usually only mildly
affected. Of note, corneal edema due to endothelial dysfunction was reported
in KSS patients also (111). Pearson syndrome classically has corneal edema
as well.
Neurologic manifestations in KSS may include cerebellar ataxia, hearing
loss, dementia, and weakness of facial, pharyngeal, trunk, and extremity
muscles. Heart block is a characteristic finding (7,112,113). POLG mutations
have an overlapping spectrum of symptoms involving the nervous system as
well as liver and muscle. One of the most severe manifestations is fatal
hepatic failure, the Alpers-Huttenlocher syndrome, which is caused by over
50 different POLG mutations. Other mtDNA deletion syndromes may also
present with extraocular manifestations including hearing loss, muscle
weakness, and neuropathy (103) or juvenile-onset sensory-ataxia neuropathy,
dysarthria, and ophthalmoplegia (SANDO), combination of spinocerebellar
ataxia and epilepsy (SCAE) with or without external ophthalmoplegia, and
mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) caused by
TYMP mutations is characterized by prominent gastrointestinal symptoms
(11,102,114). Diabetes mellitus may be present in Pearson syndrome.
Diagnostic Studies
KSS is suspected when the typical ocular, neurologic, and cardiac
manifestations occur. Ancillary tests include blood and spinal fluid lactic acid
levels and spinal tap for increased protein level (>1 g/L; the underlying cause
of which is still unknown). Electrocardiography is useful to detect and
evaluate the severity of the heart block, and genetic testing can establish the
diagnosis in cases of doubt. Muscle biopsy can demonstrate characteristic
ragged red fibers, electron microscopy may show large numbers of abnormal
mitochondria, and enzymatic assays show decreased activities of complexes
I, III, and IV with intact II. In some KSS, patient’s serum creatine kinase,
gamma globulin, and pyruvate may be elevated in the blood. The
identification of a disease-causing mutation confirms the diagnosis and
facilitates genetic counseling. CT scan may identify calcium accumulation in
specific areas of the brain.
Management
As there is no cure, management is symptomatic and supportive. Careful
correction of ptosis or strabismus may be beneficial in selected cases. The
cardiac conduction defect may be life threatening and is treated with a
pacemaker and antiarrhythmic drug therapy. Hormonal replacement therapy
is administered for endocrinopathies, and cochlear implants can be used for
patients with sensorineural hearing loss. CoQ10 (ubiquinone) is used in KSS
patients as a mitochondrial antioxidant.
Tetracycline treatment has been evaluated in CPEO; however, the results
did not formally support its effectiveness (115). At present, only
mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) has been
successfully treated by allogeneic hematopoietic stem cell transplantation
(116). Genetic therapies are being developed for KSS (117). Heteroplasmy
shifts through selective degradation of mutant DNA is a promising emerging
technology using heterodimeric mitochondrially targeted zinc finger
nucleases (mtZFNs) (118).
NEUROPATHY, ATAXIA, AND

RETINITIS PIGMENTOSA (NARP) AND
LEIGH SYNDROMES
NARP and Leigh syndromes are two inherited mitochondrial diseases
associated with the same mtDNA mutation at nucleotide position 8993. Leigh
syndrome is also associated with other mtDNA mutations
(http://www.mitomap.org/bin/view.pl/MITOMAP/ClinicalPhenotypesPolypeptide
and mutations in several nuclear genes.

The exact prevalence of NARP is unknown but may be estimated at 0.8 to
1:100,000, while Leigh syndrome is estimated at approximately 1 in 36,000
(Based on Orphanet Web site: https://www.orpha.net/).

The T>G point mutation at nucleotide 8993 of mtDNA (MT-ATP6) causes a
substitution of a highly conserved leucine by an arginine residue in the
ATPase 6 protein, a subunit of mitochondrial respiratory chain complex V
(18). mtDNA with both the T and the G nucleotides is present in the same
subject (heteroplasmy), but the proportion of the mutant mtDNA differs
among patients. Usually, a high proportion of mutant mtDNA is associated
with more severe clinical manifestations (119–122). Two clinical syndromes
were described in association with the 8993 mtDNA mutation. The first,
subacute necrotizing encephalomyopathy or Leigh syndrome, is a severe
disease, with patients showing developmental delay, ataxia, psychomotor
regression, seizures, peripheral neuropathy, and optic atrophy. Onset is
usually during infancy with progression to death within months to years. The
8993 mutation was found to be a common cause of Leigh syndrome, and
most of the patients had more than 90% of mutant mtDNA (121). The second
entity, NARP syndrome, first reported by Holt et al. in 1990 (18), is usually
milder than Leigh syndrome. Patients show a variety of clinical
manifestations such as migraine, sensory neuropathy, proximal muscle
weakness, ataxia, seizures, dementia, and pigmentary retinopathy
(18,21,120). Patients with the NARP syndrome usually carry around 80% of
mutant mtDNA. At levels below 75%, carriers may show pigmentary
retinopathy, suffer from migraines, or be totally asymptomatic depending on
the mutation (119,120). Leigh disease has also been associated with
mutations in other mtDNA genes mostly in mitochondrial genes ND3 and
ND5 (123,124). A recent Japanese study showed that of 106 individuals with
Leigh or Leigh-like syndrome, 10 of the 30 individuals with mtDNA
pathogenic variants had variants in MT-ATP6 (125).
In several other patients with Leigh syndrome, mutations were reported
in other mitochondrial genes (mainly encoding complex I subunits but also in
complexes II to IV). Additionally nuclear genes include SURF1, POLG and
other cytochrome oxidase assembly genes (126,127), as well as pyruvate
dehydrogenase complex (PDHc) genes mainly PDH1A (128). Interestingly,
age of onset, clinical symptoms, and prognosis are similar in Leigh syndrome
patients with mtDNA or nuclear DNA mutations (124).

Ocular manifestations of patients harboring mtDNA 8993 mutation can vary
from early salt-and-pepper retinopathy to retinitis pigmentosa or as a
pigmentary retinopathy (18,119,120). In addition, sluggish pupils, nystagmus,
and ophthalmoplegia were also reported (Figure 31-2). Puddu et al. (129)
described three patients with NARP syndrome, two of whom had ocular
manifestations. The fundus examination showed bone spicule pigmentation in
the midperiphery, and despite being described as RP, the ERG findings were
normal except for subnormal photopic responses. In another report, Ortiz et
al. (21) described eight patients from two families with the 8993 mutation in
whom the ocular manifestations ranged from a mild salt-and-pepper
retinopathy to severe RP-like changes with maculopathy. Four of these
patients underwent ERG examination. One showed normal results, and three
revealed rod–cone-type dysfunction. In five patients who had perimetry, four
had normal visual fields and one had a paracentral scotoma.
Figure 31-2 Color fundus photographs of two patients
with the NARP phenotype demonstrating the variable
ocular manifestations that can occur. A:A 16-year-old
male patient with patches of pigment and multiple small
retinal scars, narrowing of the blood vessels, and
pigmentary changes within the fovea. The optic disc was
normal. This patient had reduced visual acuity and
nyctalopia. Fluorescein angiography showed a bull’s-eye
maculopathy in both eyes. Visual perimetry revealed
central and paracentral scotomas. The full-field ERG
showed decreased cone- and rod-derived amplitudes.
B:The sister of the male patient was 12 years old at that
time. She manifested bilateral bull’s-eye maculopathy with
temporal optic disc pallor. No pigmentary changes were
apparent. The sister had mild reduction of visual acuity,
episodic ataxia, and mild mental retardation. The full-field
ERG showed decreased cone-derived, but normal rod-
derived, amplitudes.
Chowers et al. (16) described three members from a family harboring the
8993 mtDNA mutation in whom great variability in the clinical
manifestations of the disease was observed. Two of the three patients had
ocular abnormalities that were different from classic RP. The clinical
findings, visual fields, and ERG were typical of cone–rod dystrophy in one of
the patients and progressive cone dystrophy in another patient. Other retinal
findings in NARP include a bull’s-eye maculopathy and rod–cone type of
retinal dystrophy (14).
As discussed, there is great variability in the ocular and systemic
manifestations among patients carrying the mtDNA 8993 mutation. Several
factors could account for the spectrum of abnormalities observed. First, the
burden of mutant mtDNA in a particular tissue may vary thus influencing
phenotype, with a higher proportion of mutant mtDNA in blood lymphocytes
being a marker for more severe clinical manifestations. However, Chowers et
al. (16) observed that patients with a lower percentage of mutant mtDNA in
the blood lymphocytes can have a more severe ocular disease compared with
patients with higher proportion of mutant mtDNA. This can be partially
explained if the proportion of mutant mtDNA in the retinas differs from that
in the blood or other tissues, especially the central nervous system (CNS), but
such differences have not yet been demonstrated (119). Differences among
patients may also depend on age. Ortiz et al. (21) described two patients,
aged 12 and 14 years, who suffered from NARP syndrome and had a rod–
cone type of ERG dysfunction. The retinopathy of these patients was much
less advanced than that of patients reported by Chowers et al. who were of a
similar age but with a predominantly cone dysfunction. Thus, patients with
the NARP syndrome may manifest either rod- or cone-dominant retinal
dystrophy at a similar age. It is likely that other factors, not yet understood,
modulate the phenotypic expression of the 8993 mtDNA mutation. In support
of this, two of the three reported NARP pedigrees undergoing ERG had
predominantly cone dysfunction, while the third pedigree had predominantly
rod dysfunction. This finding could not be explained by the differences in
age, stages of the disease, or percentage of mutant mtDNA among families.
Diagnostic Studies
Diagnosis is based on identification of the typical combination of ocular and
systemic manifestations along with the mitochondrial inheritance pattern.
Ancillary tests may include ERG, color vision, and fluorescein angiography.
Lactate levels are constantly elevated in spinal tap and sometimes also in
blood tests. Brain imaging shows certain topology of the brainstem and basal
ganglia lesions, often in association with leukodystrophy and cerebral
atrophy. Definitive diagnosis is made by genetic workup.
Management
No treatment has been shown to be of benefit in arresting the progression of
retinal dystrophy in patients carrying the mtDNA 8993 mutation. In addition,
patients should be referred to a clinical geneticist for evaluation of medical
and family history because although the 8993 mutation is transmitted from
mother to each of her offspring, the proportion of mutant mtDNA in each
offspring can differ considerably from that of the mother and with it the
clinical manifestations. New therapeutic approaches include sodium pyruvate
to improve the redox state and increase ATP production via glycolysis (130).
A single-arm clinical trial using EPI-743, a derivative of coenzyme Q10,
included 10 consecutive children aged 1 to 13 years, reversed the progression
of the diseases (69,131). Idebenone or gene therapy may be beneficial for
NARP and Leigh syndrome patients (132,133).
MITOCHONDRIAL
ENCEPHALOPATHY, LACTIC
ACIDOSIS, AND STROKE-LIKE
EPISODE (MELAS) SYNDROME
While vision loss in mitochondrial genetic disorders more commonly occurs
from retinal degeneration or optic nerve disease, it can occur also from
damage to the retrochiasmal visual pathways as in MELAS (134,135), a
condition that is mostly caused by a mutation at mtDNA nucleotide position
3243 in the leucine tRNA gene. Other features included diabetes mellitus and
deafness. Of note, the m.3243A>G mutation may also be associated with a
pigmentary retinopathy or optic neuropathy as well as CPEO (136). Other
mitochondrial diseases, caused by mitochondrial and nuclear mutations,
frequently present with lactic acidosis and encephalomyopathy. These
include isolated complex I deficiency and mutations in c12of65 (137), and
the majority of these patients also manifest visual impairment (138). A
retrospective study of patients with molecularly defined mitochondrial
disease detected that nearly half of them had some form of visual impairment
(3).

Patients harboring the A3243G mutation present intrafamilial and
interfamilial variable expressivity (139) and can have cortical blindness,
ptosis, external ophthalmoplegia (140), iris atrophy, posterior subcapsular
cataract, pigmentary retinopathy with maculopathy, RPE and choroidal
atrophy (141), transient optic disc edema (142), and optic atrophy as proven
by SD-OCT (23,143–146). Rummelt et al. have demonstrated histologic
findings corroborating the clinical manifestations in two eyes of a patient
with the MELAS syndrome and the mtDNA 3243 mutation, including
posterior subcapsular cataract, photoreceptor degeneration, and RPE atrophy
and hyperpigmentation. In addition, multiple cell types showed enlarged and
abnormal mitochondria (144). There are contradictory reports regarding the
correlation between severity of RPE abnormalities and systemic phenotype as
a consequence of heteroplasmy (147,148).
Diagnostic Studies
The diagnosis is based on clinical findings and molecular analysis, which is
often preceded by enzymatic analysis of muscle biopsy, especially in the
cases of disorders caused by mutations in nuclear genes.
Management
Although no treatment is established, novel therapies such as oral L-arginine
(149), succinate (150), EPI-743 (69), and idebenone (151) or gene therapies
are being investigated in several clinical trials and may be beneficial for
MELAS. Similar to other mitochondrial diseases, MELAS patients should
avoid mitochondrial toxins such as smoking, aminoglycoside antibiotics,
dichloroacetate, linezolid, and alcohol. Valproic acid should be also avoided
in the treatment of seizures.
THE PEROXISOMES
Peroxisomes are single membrane–enclosed cytoplasmic organelles derived
from the endoplasmic reticulum. There are several hundred peroxisomes per
cell, but unlike mitochondria, they do not contain DNA even though they
replicate by division. Numerous catabolic and anabolic processes take place
within the peroxisomes and involve more than 50 enzymes. Major functions
of the peroxisomes are beta-oxidation of very long (>C22) fatty acids
(VLCFA) and alpha-oxidation of 3-methyl–branched fatty acids such as
phytanic acid. Peroxisomes also perform biosynthetic functions and are
involved in the synthesis of plasmalogens and docosahexaenoic acid (DHA),
which are critical for cellular membrane function and bile acids. They are
also involved in detoxification of glyoxylate and in D-amino acid and pipe
colic acid oxidation. Some of abovementioned reactions generate hydrogen
peroxide (hence the name peroxisome), which is used for oxidation of
additional molecules or detoxified by conversion to water and oxygen by the
enzyme catalase.
Proteins required for peroxisome assembly are targeted to the peroxisome
by either carboxy (peroxisome targeting signal 1 or PTS1) or amino (PTS2)
terminus targeting signals. Specific translocation complexes then mediate
their transport into the peroxisome. Sixteen proteins that control peroxisome
assembly, division, and inheritance, which are encoded by PEX genes, are
named peroxins, and they are essential for normal development (152–154).
Peroxisomes are not isolated entities but interact with most organelles mainly
mitochondria and endoplasmic reticulum; and peroxisomal proteins are
sometimes even shared with other cellular compartments.
Peroxisomal Disorders
Peroxisomal disorders are a group of rare diseases presenting with
overlapping clinical manifestations and affecting multiple tissues including
the eye. They are mostly detected by measuring saturated very long-chain
fatty acids (VLCFA), phytanic and pristanic acids in plasma, and decreased
erythrocyte plasmalogens; however, since many genes are involved, the final
diagnosis is molecular.
Peroxisomal diseases are usually classified into two main categories:
peroxisome biogenesis disorders (PBD) and single enzyme/transporter (PED)
defects. Several PBDs commonly affect the retina: Zellweger syndrome (ZS;
OMIM #214100), the most severe form, and the intermediate and milder
forms, previously termed neonatal adrenoleukodystrophy (NALD), and
infantile Refsum disease (IRD) are parts of the Zellweger spectrum disorders
(ZSDs). ZSDs are associated with mutations in a large number of different
PEX genes. At the cellular level, defects causing ZSD lead to decreased
numbers of enlarged but dysfunctional peroxisomes, called “ghosts.” A
distinct PDB is rhizomelic chondrodysplasia punctata type I (RCDP type I;
OMIM #215100), which is mainly associated with cataract and mutated
PEX7 (155).
The second group of peroxisomal disorders comprises those for which the
function of the peroxisome is abnormal, but its assembly is not affected. This
group includes entities such as X-linked adrenoleukodystrophy (OMIM
#300100), primary hyperoxaluria type 1 (PHT1) (OMIM #259900), classical
Refsum disease (OMIM #266500), and RCDP types II and III. X-linked
adrenoleukodystrophy is associated with mutations in the ABCD1 gene,
resulting in a defect in peroxisomal beta-oxidation. This leads to the
accumulation of VLCFA in all tissues of the body, with clinical
manifestations that are primarily related to the adrenal cortex, CNS myelin
(including visual disturbance), and Leydig cell dysfunction (154,156). In
recent years, a number of newly identified peroxisomal disorders have been
described caused by mutations in genes coding for peroxisomal proteins,
exemplified by HAO1, PMP70, ACBD5, and ACOX2 (154).

Peroxisomal disorders are rare with approximate prevalence of ZS, NALD,
and IRD of 1:30,000 births. Shimozawa estimated a prevalence of PBD in
Japan to be 0.22 in 100,000 births (157). ALD is considered the most
common: 1:200,000 to 1.6:100,000 live births (158–160). Moser (161)
recorded the relative prevalence of the different peroxisomal disorders in
1,345 patients. In his series, X-linked adrenoleukodystrophy was the most
common disorder (1,184 cases), followed by ZS (105 cases) and NALD (54
cases). Refsum disease and several other rare peroxisomal disorders were
diagnosed in the remaining patients. In Japan, NALD is estimated to be
between 1:30,000 and 1:50,000, relatively more prevalent compared to the
other subtypes (157,162).
The following paragraphs will first consider the three PBDs affecting the
retina, followed by descriptions of other peroxisomal disorders affecting the
retina.
PEROXISOMAL BIOGENESIS
DISORDERS AFFECTING THE RETINA
More than 20 genes related to peroxisome biogenesis have been identified.
PBDs have been associated with mutations in 14 of these genes (PEX1,
PEX2, PEX3, PEX5, PEX6, PEX7, PEX10, PEX11b, PEX12, PEX13, PEX14,
PEX16, PEX19, PEX26). Frequently, mutations in the same peroxin gene can
lead to any of the PBD phenotypes, although defects in some peroxins are
only associated with the more severe ZS phenotype (163). In general, disease
severity correlates with the patient age at onset of symptoms and the
predicted consequence on peroxin function, capacity for matrix protein
import, residual enzyme functions, and peroxisome numbers. The most
severe form of ZS presents at birth with a classic malformation syndrome
described by its characteristic features as cerebro-hepato-renal syndrome
including severe hypotonia, seizures, and characteristic craniofacial
dysmorphisms. Eye disease includes cataracts, congenital glaucoma, retinal
dystrophy, and optic atrophy (153).
The generation of mouse models for PBD has provided interesting
insights into the pathogenesis of these diseases. Disruption of PEX2, PEX5,
and PEX11b in mouse models can frequently result in findings similar to
PBD in humans, including neuronal migration defects, hypotonia,
developmental delay, and neonatal lethality. However, in mice with PEX11b
disruption, there is no detectable defect in peroxisomal protein import, and
only mild defects in peroxisomal fatty acid beta-oxidation and peroxisomal
ether lipid biosynthesis are present (164). These findings are intriguing in that
features of the PBD phenotype that are present in these PEX11b-deficient
mice, but they do not have the biochemical and cellular abnormalities usually
associated with PEX mutations, as they have apparently an intact peroxisomal
biogenesis. A more recent mouse recapitulating mild human ZSD (PEX1-
G843D knockin) developed a retinopathy similar to that observed in human
patients, with evidence of cone photoreceptor cell death, and will be useful
for evaluating therapies (165).

Patients with ZS show a variety of ocular manifestations including corneal
clouding, posterior embryotoxon, glaucoma, cataract, RPE clumping,
attenuation of retinal blood vessels, optic disc pallor or hypoplasia, and
extinguished ERG (166). In addition, pendular nystagmus is almost always
present (167). Myopia has been reported in some patients. In IRD and
NALD, the predominant findings are in the posterior segment where there are
arteriolar attenuation, pigment clumping, and optic atrophy (166). The natural
history of the disease is rapid progression to total blindness.
The histopathology in ZS, NALD, and IRD shows loss of photoreceptors
and other retinal cells and infiltration of macrophages laden with cytoplasmic
inclusion bodies. Similar findings were detected in other areas of the CNS.
ZS (cerebro-hepato-renal) presents in the neonatal period, but other ZSDs can
manifest any time from infancy and during the first decade of life. All PBDs
are associated with a variety of systemic abnormalities, additionally to the
ocular manifestations. Death in ZS occurs within a few months of birth, while
milder ZSDs survive for longer periods.
ZS patients present with skeletal and dysmorphic features including
talipes equinovarus, limb contractures, high forehead, epicanthal folds,
hypoplastic supraorbital ridge, micrognathia, high-arched palate, and
hypertelorism. Dysmorphism can be intermediate or mild or even absent in
other ZSDs. Less affected patients have adrenal and cortical atrophy but do
not develop adrenal insufficiency as ZS. Neurologic manifestations include
seizures, psychomotor retardation, hypotonia, cerebellar ataxia, and reduced
hearing. Additional abnormalities include hepatomegaly, jaundice, congenital
heart defects, leukodystrophy, osteopenia, and peripheral neuropathy
(163,168). Generally, clinical features vary with age of onset (153).
Diagnostic Studies
The differential diagnosis includes a long list of other inborn errors of
metabolism sharing the systemic manifestations such as Prader-Willi
syndrome, spinal muscular atrophy, congenital myotonic dystrophy type 1,
and other congenital myopathies. ZSDs may be confused with other causes of
sensorineural hearing loss and decreased vision in infancy including Usher
syndromes 1 and 2, Cockayne syndrome, Leber congenital amaurosis, and
neuronal ceroid lipofuscinosis.
The diagnosis is based on identifying the combination of typical systemic
and ocular manifestations mentioned previously together with genetic
workup. Ancillary tests may include an ERG test, which is severely reduced
or extinguished in all three diseases. Plasma VLCFA, pipecolic acid, bile
acid, and phytanic and pristanic acids are typically increased, whereas the
plasmalogen level in red blood cells is reduced. Of note, phytanic and
pristanic acids are diet dependent and could remain normal in young infants.
Genetic testing and identification of the characteristic mutations can establish
the diagnosis.
Management
Management is mainly supportive such as feeding and nutrition, hearing aids,
cataract removal, antiepileptic drugs, and cholic acid–replacement therapy.
Recently, allogeneic hematopoietic cell transplantation (HCT) has been
shown to provide long-term disease stabilization and survival (5-year
survival: 55% for untreated and 78% for treated). The success rate is largely
dependent on the pre-HCT neurologic status and the age it is given (169,170).
Gene therapy was already tested on two NALD patients, who showed
cerebral demyelination arrest (171). Large-scale screening has identified
small molecules, which may rescue peroxisome function in ZSDs. The in
vivo effect of such compounds is still unknown (172). Additional substances
that stimulate peroxisomal biogenesis are being investigated currently
(173,174).
CLASSICAL REFSUM DISEASE

Environmental Factors, Genetics, and Pathophysiology
Classical or adult Refsum disease (ARD) is another disorder affecting
peroxisomal biogenesis but has a distinct phenotype. It is less severe and
occurs in late childhood and adults often associated with retinitis pigmentosa,
anosmia, deafness, ataxia, and polyneuropathy. ARD is an autosomal
recessive disorder affecting peroxisomal alpha-oxidation, associated with
mutations in either phytanoyl–coenzyme A hydroxylase (PAHX) or peroxin-7
(PEX7) genes (175–177). The resulting biochemical defects lead to the
accumulation of the branched-chain fatty acid phytanic acid (3,7,11,15-
tetramethyl-hexadecanoic acid) by perturbation of its normal degradation
pathway while VLCFA levels remain normal. Apart from its main alpha-
oxidation degradation pathway, phytanic acid can also be degraded by
omega-oxidization; thus, elevated levels of urinary 3-methyladipic acid can
be found in Refsum patients. Precursors of phytanic acid mainly phytol (a
constituent of chlorophyll) present in leaves and plants are degraded to
phytanic acid by bacteria in the digestive system of ruminal animals. Thus,
the source of the main phytanic acid in humans originates from meat and
milk products from ruminant animals and varies with diet (178).

Ocular manifestations are characterized by RP-like changes including
nyctalopia, progressively constricted visual fields, granular type of
pigmentary retinopathy, and attenuated blood vessels (166,179–181). Of
note, ARD patients often lack bone spicule–like pigmentations seen in
classical RP patients. Retinal histology shows atrophy of the retinal RPE and
lipid-laden RPE cells in areas where RPE has not yet atrophied. Lipid
deposits have been found in the pigment epithelium of the iris, ciliary body,
and retina as well as other tissues leading to degeneration (182,183). The
outer nuclear and plexiform layers are also atrophic, the inner nuclear layer is
thinned, and the ganglion cell number is decreased (181,182,184). Cataract is
found in about 50% of Refsum disease patients (185). Additional ocular
manifestations include miotic unreactive pupils and nystagmus (186).
Refsum disease usually manifests between the first and third decades.
Multiple systems are involved. Neurologic manifestations include peripheral
polyneuropathy and cerebellar signs such as ataxia, anosmia, and decreased
hearing. Additional manifestations are cardiac arrhythmia (that can be life
threatening), skeletal malformations (mostly epiphyseal dysplasia), and
ichthyosis-like skin changes (177,179,180,184,187).
Diagnostic Studies
The diagnosis is based on typical clinical findings. Ancillary tests may
include kinetic perimetry, which is useful in documenting visual field loss
and in following its progression, and ERG, which shows decreased photopic
and scotopic responses. Skeletal radiography and brain MRI may be helpful
in demonstrating structural abnormalities. Blood levels of phytanic acid are
typically elevated (>10 to 50 mg/dL), and low-density lipoprotein (LDL) and
high-density lipoprotein (HDL) cholesterol levels are decreased and increased
protein concentration in cerebrospinal fluid (CSF) (156,161,188). Mutation
detection confirms the final diagnosis.
Management
Refsum disease is a rare case of an RP-like disease that is amenable to
treatment. As phytanic acid cannot be synthesized in humans, its
accumulation in Refsum disease patients depends on exogenous intake.
Accordingly, a diet free of foods containing phytol, phytanic acid, or their
precursors leads to reduced phytanic acid levels in the blood and to clinical
improvement. Plasmapheresis to remove phytanic acid can be an alternative
effective approach to diet in preventing the clinical manifestations of Refsum
disease and in severe or rapidly worsening cases (189–191). It is thus
imperative that Refsum disease diagnosis be made as early as possible (192).
Novel potential treatments include CYP4, which was shown to be effective in
eliminating phytanic acid (193).
PRIMARY HYPEROXALURIA TYPE 1

PHT1 is a rare peroxisomal autosomal recessive disease with a defect in the
gene, AGXT, which encodes the enzyme alanine–glyoxylate
aminotransferase, a hepatic peroxisomal enzyme that converts glyoxylate to
glycine (194). This defect leads to progressive overproduction of oxalate and
formation of calcium-oxalate crystals. Urologic manifestations first
predominate with high urinary oxalate and glycolic acid excretion and
nephrolithiasis, which are followed by progressive nephrocalcinosis and renal
failure. Additional cardiomyopathies, osteopathies and synovitis are known
complications. Deposition of calcium-oxalate crystals includes the eye
leading to visual impairment. Untreated, the disease leads to death from renal
failure in childhood or early adulthood.
Retinal oxalosis can be variable from subtle perifoveal crystals without
any implications to vision up to subfoveal fibrosis associated with macular
RPE hyperplasia, proliferative retinopathy, or optic atrophy (195–199).
Derveaux et al. published four unrelated cases with PHT1 presented with four
distinct retinal phenotypes summarizing the possible funduscopic findings in
this disease (200). This included normal-appearing fundus together with
normal ERG; numerous deep yellowish flecks in the posterior pole and focal
perifoveal RPE hyperplasia; and diffuse deep flecks with confluent black
ringlets of RPE hyperplasia in the macular area and foveal scarring due to
subretinal fibrosis in the most severe cases. Histopathologic findings in the
eye demonstrated oxalate birefringent crystal deposition in the conjunctiva,
episclera, sclera, ciliary body, ciliary processes, neuroretina, and RPE
(201,202). Specifically, they were found in larger extent in the inner and
outer retina and in lesser extent in the RPE and sub-RPE space
(199,202,203). In addition, extensive distribution of oxalate deposits along
choroidal and retinal arteriolar vessel walls has been reported (199,202).
Typically, these deposits are distributed in the posterior pole, decreasing in
size and number toward the periphery (204). One additional large series of
patients presented a wide range of retinal findings from multiple small
parafoveal subretinal black rings with a white center in mild cases to a large
black geographic shape lesion underlying the macula area with a whitish
subretinal area in more severe cases (205). These black areas represent
hyperplastic and hypertrophic RPE reaction to oxalate deposition, while the
white center may represent choroidal neovascularization (202,206,207).
Optic disc pallor was also reported in patients with PHT1 and may be
associated with papilledema or vascular decompensation secondary to oxalate
deposition. In cases involving the optic nerve, visual acuity was markedly
reduced (205). Longitudinal follow-up of some cases showed reversal of
arteriolar and neuroretinal oxalate deposition after kidney transplantation or
increased frequency of dialysis (197,208).
Management
Decreasing oxalate load in the body is one approach, although kidney
transplantation is associated with a high recurrence rate. Pyridoxine is useful
in 10% to 30% of cases to decrease the body’s production of oxalate. Liver
transplantation, when successful, is a definitive treatment for this disease
(201,209).
CONGENITAL DISORDERS OF
GLYCOSYLATION (CDG)
Environmental Factors, Genetics, and Pathophysiology
Over half of all intracellular proteins are posttranslationally glycosylated
(glycans) either by N-glycosylation of asparagine or O-glycosylated to serine
or threonine. The synthesis of N-glycans is complex and involves three
compartments, cytosol, endoplasmic reticulum, and Golgi apparatus, while
O-glycosylation mainly occurs in the Golgi. A CDG is a rapidly expanding
group of inherited disorders with abnormal glycan metabolism leading to
impaired glycosylation of proteins and lipids. The nomenclature has varied,
but currently there are four different biochemical groups involving defects in
protein N-linked glycosylation, protein O-linked glycosylation, glycolipid,
and GPI anchor glycosylation and a group with defects in multiple
glycosylation pathways. The first described and most frequent CDG, which
was reported in 1984 by Jaeken et al. (210), is phosphomannose mutase
deficiency PMM2-CDG (previously CDG1a) affecting protein N-
glycosylation. Since then, over 130 human genetic disorders have been
associated with abnormal glycosylation, and the number is still increasing
(211).

O-glycosylation is crucial for normal eye embryogenesis and physiology,
leading to eye disorder in 60% of pure O-glycosylation defects. Up to date,
370 CDG subjects presented with eye involvement (212). B3GALTL-CDG,
LARGE-CDG, B3GAT3-CDG, and XYLT2-CDG are the most frequent
CDG to have ocular manifestation as 98% of B3GALTL-CDG were reported
to suffer from eye involvement. PMM2-CDG patients, most commonly CDG,
show a high prevalence of myopia, which has been suggested to reflect
altered proteoglycan synthesis in sclera and progressive pigmentary
retinopathy (213,214). Patients with SRD5A3-CDA dolichol phosphate
synthesis defect (formerly CDG-1x) display several eye defects including
microphthalmia, iris and chorioretinal colobomas, optic atrophy/hypoplasia,
cataract, and glaucoma in both adult- and early-onset forms (215–220).
Typically, SRDA3-CDG patients present with retinitis pigmentosa combined
with bone spicule pigmentation and attenuated blood vessels (215,221,222).
The systemic manifestations of the different CDGs are extremely variable
depending on the specific gene and mutation involved. PMM2-CDG patients
typically present with dysmorphic facial features, strabismus, inverted
nipples, abnormal fat distribution, endocrine abnormalities, and coagulation
defects; and MPI-CDG (formerly CDG1b) patients present with liver disease,
coagulopathy, hyperinsulinemic hypoglycemia, and gastrointestinal
symptoms (protein-losing enteropathy). Other glycosylation diseases are
much more difficult to classify by phenotype. Presentations include a
pleiotropy of symptoms including neurologic deficits, facial dysmorphism,
skeletal dysplasia, muscular dystrophy, joint and skin laxity, congenital
malformations, and developmental delay as recently reviewed by Ferreira et
al. (211,223).
Diagnostic Studies
As the clinical manifestations of CDGs are so varied, they frequently overlap
with other neurologic and mitochondrial disorders (211,223), and with a few
exceptions, biochemical and molecular workup is imperative to reach a
diagnosis. Many CDGs are linked to deficient sialylation of serum
transferrin, and thus, transferrin isoelectric focusing (TIEF) or transferrin
capillary electrophoresis is widely used to screen for N-glycosylation CDGs.
Notably abnormal transferrin pattern can also be secondary to alcoholism,
liver disease, galactosemia, and fructosemia. Advancements in diagnostic
techniques performed in specialized laboratories now include the analysis of
whole serum N-glycans by MALDI-TOF mass spectrometry and transferrin
analysis by high-resolution QTOF mass spectrometry. O-glycans are
analyzed by isoelectric focusing and by mass spectrometry analysis of apoC-
III (224). As many genes are involved in CDG, whole exome and genome
analysis might be performed in parallel or even prior to biochemical
techniques to obtain a final diagnosis.
Management
To date, management of CDGs is largely symptomatic. Only a few specific
CDGs benefit from oral supplementation in the form of mannose (MPI-CDG)
or galactose (PGM1-CDG, TMEM165-CDG), and correct diagnosis is
imperative before starting treatment. Thus, novel treatment strategies are
urgently needed (211).
Optic atrophy from any cause may result in central scotomas that decrease
acuity and prevent good vision for reading and other important functions.
Patients should be referred to a low-vision specialist for evaluation and
rehabilitation. See Chapter 14 for a full discussion of vision rehabilitation.

Mitochondrial disorders may affect most organ systems of the body. Once a
mitochondrial disorder is suspected or confirmed, a general examination
including electroencephalogram, hearing test, and routine laboratory studies
should be performed.
Mitochondrial disorders are inherited through the maternal lineage, and some
have more severe expression in males than females. A detailed discussion
about recurrence risks for the patient and other family members is essential.
This may ideally be accomplished through referral to a medical geneticist or
genetic counselor.
FUTURE TREATMENTS
Human gene therapy trials are underway for one mitochondrial disorder
causing blindness, LHON. In the future, there will likely be more. Patients
should be offered a molecular genetic diagnosis in order to be able to take
advantage of advances in treatment. Patients can be referred to
www.clinicaltrials.gov for information on clinical trials. At this writing,
www.clinicaltrials.gov is only lightly curated, so the validity and
appropriateness of a trial for each patient must be discussed with a physician
knowledgeable in the field.
SUMMARY
Mitochondrial, peroxisomal, and glycosylation disorders have been the
subject of intense research in recent years. Multiple genes associated with
these disorders have been identified, and significant insight into the genetics
has been gained, especially due to the significant progress in high-throughput
sequencing techniques, including whole genome and exome sequencing,
leading to new classifications and to development of diagnostic genetic tests
for many of these disorders. On the other hand, the pathogenesis of the
diseases described in this chapter is still unclear in many instances despite the
discovery of the causative gene. The variable disease course in patients with
the same mutations suggests that additional yet unrecognized modifying
factors playing an important role in these disorders. Of note, the significant
advancement in ophthalmology imaging techniques contributes largely to the
precise phenotyping of these similar diseases in terms of the ocular findings
leading to novel subclassifications and better diagnosis, follow-up, and
prognostic prediction.
Furthermore, the complex and unique inheritance mode of mitochondrial
disorders, heteroplasmy in particular, hampers the potential for prenatal
diagnosis in many instances. For example, although all offspring of a mother
carrying the NARP-causing mtDNA 8993 point mutation will also carry the
same defect, the phenotype may vary considerably between offspring; some
may be asymptomatic, and others may be severely affected. In this instance,
determining the fraction of mtDNA with the mutation by prenatal diagnostic
testing will not necessarily reliably reflect the fraction in the target tissue for
this disorder such as the retina and brain. Thus, basing decisions regarding
pregnancy on such diagnostic tests raises complex ethical and moral issues.
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32
Bardet-Biedl Syndrome
Seongjin Seo, and Arlene V. Drack
BARDET-BIEDL SYNDROME
Prevalence
How many genes in the human body, when mutated, can cause the specific
constellation of extra fingers and toes, retinitis pigmentosa (RP), obesity,
renal and gonadal dysfunction, and developmental/behavioral issues?
Surprisingly, the number is 21 and counting for the autosomal recessive
disorder known as Bardet-Biedl syndrome (BBS), a ciliopathy that is
teaching us much about this class of disease and the roles of cilia in health
and disease (1). BBS is diagnosed clinically when a patient has four primary
features or three primary and two secondary features from a list of
characteristics originally described by Beales et al. in 1999 (2) and updated
by Forsythe and Beales in 2013 (3).
Primary and secondary features include the following.
Primary
Cone–rod dystrophy
Polydactyly (typically postaxial)
Truncal obesity
Learning difficulties
Genital anomalies
Renal anomalies
Secondary
Speech delay
Developmental delay
Diabetes mellitus
Dental anomalies
Congenital heart disease
Brachydactyly/syndactyly
Ataxia/poor coordination
Anosmia/hyposmia
BBS is estimated to occur in approximately 1/160,000 people in outbred

European populations (4), but this may be underestimated due to the absence
of the cardinal features in some patients. In some populations, the incidence
is significantly higher. It is reported to be approximately 1/18,000 in
Newfoundland, Canada (5,6). Newfoundland has a relatively isolated
population founded by approximately 30,000 initial inhabitants suggesting a
founder effect; however, the BBS patients from this region carry multiple
mutations in multiple genes. BBS shows no racial, gender, or ethnic
predisposition. Consanguinity does increase the risk of BBS; therefore,
regions in which consanguinity is common have a higher prevalence, such as
the Bedouins of Kuwait with an estimated prevalence of 1/13,500 (4).
Environmental Factors
There are no known environmental factors impacting the development of
BBS, other than consanguinity. Obesity is a feature of BBS and may be
amenable to environmental intervention in the form of reduced calorie diets;
however, management is difficult, especially in young children. Experimental
evidence suggests that the obesity seen in BBS is due to resistance to leptin, a
hormone that signals satiety to the brain (7,8).
Genetics
Between the World Wars, Georges Bardet (in Paris) and Albert Biedl (at the
German University of Prague) each described patients with features that
would become cardinal phenotypes of BBS (9–12) and postulated on the
familial nature of these features. Subsequent reports of small families showed
that BBS is inherited as an autosomal recessive disorder.
The identification of the genes that cause rare autosomal recessive
diseases is difficult in outbred populations, in which mating between
unrelated individuals leads to increased genetic variation. Inbred populations
that are genetically isolated were proposed to be especially useful for
identifying rare disease-causing genes that act through an autosomal
recessive pattern of inheritance, since the frequency of homozygous
mutations is so much higher (about 6.25% of the alleles in the offspring of
first cousins are homozygous). Studying inbred populations has been fruitful
in mapping and discovering the genes responsible for BBS. The Bedouin-
Arab population of the Negev region of Israel shows a high degree of
traditional consanguineous marriages, in addition to generally having
relatively large families (which increases the likelihood of identifying
multiple affected members in a generation when on average only one in four
offspring is affected).
Linkage analysis initially mapped BBS in one Bedouin kindred to
chromosome 16 (13). The unusual set of features and rarity of the disease
might suggest that BBS is likely due to the action of one mutation, or
mutations in one gene. However, subsequent screening to determine if this
region of the genome segregated with BBS in a second Bedouin population
showed that BBS exhibits locus heterogeneity (13). Later studies that
identified loci and genes responsible for BBS revealed that disruption of any
of several genes can lead to essentially the same phenotypes (14).
As of the date of this writing, Online Mendelian Inheritance in Man
reports 21 BBS genes (http://www.omim.org/entry/209900) (Table 32-1).
With some exceptions, these genes did not initially show obvious homology
to genes with known functions. The mechanism(s) by which homozygous or
compound heterozygous mutations in any of these disparate genes results in
the same clinical findings was elusive for several years, but then biochemical
data provided a compelling explanation: the protein products of many of
these genes interact in multiprotein complexes and are intimately related in
function (see next section of this chapter and Figure 32-1).
TABLE 32-1 Genes associated with Bardet-Biedl
syndrome
FIGURE 32-1 Current model for BBS protein functions.

Protein trafficking in normal cilia (Left). The BBS/CCT
chaperonin complex facilitates BBSome assembly. The
BBSome acts as an adapter between IFT particles and
certain ciliary cargo proteins to transport them in and out
of cilia. BBS17 mediates the BBSome–IFT interaction to
facilitate BBSome exit from cilia. Protein trafficking in the
absence of BBS proteins (Right). Inactivating mutations in
BBSome components or BBS/CCT chaperonin complex
components result in a lack of functional BBSomes. This
causes inefficient trafficking of certain ciliary proteins as
well as accumulation of nonciliary proteins in the ciliary
compartment.
Like most genetic disorders, BBS has been reported as having variable
expressivity, which is notable in the retinal pathology as well as other
features of the disease. About 90% of patients exhibit retinal degeneration
with lower percentages of patients having the other manifestations (1,3). BBS
has been suggested to show triallelic inheritance, a condition in which two
mutations in one gene are not sufficient to cause disease but require an
additional mutation at a second locus (15). It is undoubtedly the case that
genetic background influences the phenotype(s) associated with BBS, and
variations in other BBS genes are good candidates for these influences
especially given what is now known about the interactions of BBS proteins
(described below). Generalizations about a disorder caused by multiple
genes, including many that probably remain to be discovered, are precarious;
the majority of familial BBS behaves as an autosomal recessive disorder
(16–18). It is estimated that up to 10% of BBS families harbor mutations in
more than one BBS gene, but the role this plays in disease is still uncertain
(4). In mouse models of BBS, mutations in two alleles at one BBS locus are
sufficient to cause BBS phenotypes (19–22); however, heterozygous
mutations of CEP290 have been shown to modify the phenotype (23,24).
Digenic inheritance has not been reported. Webb et al. screened BBS
patients in Newfoundland for known BBS mutations and found three
members of a large BBS1 family who carried one BBS1 mutation in addition
to a BBS3 mutation that would be predicted to cause disease if homozygous
(25). None of these individuals exhibited any features of BBS, which argues
against digenic inheritance, at least for these two genes. These same authors
also noted no increase in incidence of renal disease in heterozygous carriers
of a disease-causing mutation (25). Fan et al. found that a heterozygous
mutation in BBS3 modified the phenotype in homozygous M390R BBS1
(26).
BBS has been conflated with Lawrence-Moon syndrome (LMS) (OMIM
# 245800); however, the latter condition has not been consistently shown to
exhibit polydactyly or obesity, cardinal features of BBS, and the relationship
between these syndromes is not entirely clear. One study found that two
patients who clinically fit the phenotype of LMS both had mutations in
known BBS genes (BBS2 and BBS6) (6). It seems probable that BBS and
LMS are different manifestations of overlapping phenotypes caused by the
interactions of related genes.
At this writing, there are 21 genes reported to cause classic BBS or a
BBS-like syndrome: BBS1, BBS2, ARL6, BBS4, BBS5, MKKS, BBS7, TTC8,
BBS9, BBS10, TRIM32, BBS12, MKS1, CEP290, WDPCP, SDCCAG8,
LZTFL1, BBIP1, IFT27, IFT74, and C8ORF37 (Table 32-1). There are likely
to be more discovered. In turn, this will broaden the clinical diagnosis of BBS
as more patients with partial forms are discovered to harbor mutations in
known BBS genes. The nomenclature of ophthalmology, and all of medicine,
is changing to reflect the new level of knowledge that comes from molecular
diagnoses.
Worldwide Impact for Childhood Blindness Including

Public Health Issues
BBS causes a rapidly progressive retinal degeneration leading to legal
blindness in the majority of patients by the age of 20 to 30 years (4). Since
these patients are often multiply handicapped, including renal failure, this is
especially debilitating. Areas of the world in which consanguinity are more
common have a higher burden of BBS.
Pathophysiology
Studies during the past two decades have uncovered that BBS gene products
localize to primary cilia, and loss of BBS gene functions causes ciliary
dysfunction in multiple tissues (27,28) (and references therein). Primary cilia
are small sensory organelles that protrude from the cell surface and receive
signals from the surrounding environment (29) (and references therein). As
sensory organelles, primary cilia harbor various signaling molecules
including receptors and maintain a unique protein composition compared to
the rest of the cell body. Certain molecules enter and exit cilia upon
activation or inactivation of signaling pathways. BBS proteins are involved in
protein trafficking in and out of cilia and ciliary protein homeostasis (30,31)
(and references therein).
Insights into the pathophysiology of BBS have been gained through the
use of animal models in which specific BBS genes are disrupted. Inactivation
of BBS genes in mice results in lack of flagella in spermatozoa as well as
abnormalities in cilia in brain ependymal cells, airway epithelial cells, and
olfactory neurons (19–22,32–35). Inactivation of BBS genes also causes
photoreceptor degeneration in the retina (19–22,32,34–39). The
photoreceptor outer segment is a highly specialized form of primary cilia,
filled with proteins necessary for light perception and phototransduction. In
contrast, proteins involved in other cellular activities such as protein
synthesis, energy production, or synaptic transmission are excluded from this
compartment. Recent studies uncovered that loss of BBS gene functions leads
to accumulation of non–outer segment proteins in the outer segment and
outer segment malformation (37–39). Combined with studies in
Chlamydomonas reinhardtii BBS mutants (40,41), these findings suggest that
the primary function of BBS proteins is to export nonciliary proteins out of
cilia (Figure 32-1). Further investigation is needed to understand how these
abnormalities induce photoreceptor cell death. Besides retinal degeneration,
human BBS patients often exhibit polydactyly and obesity. Although precise
mechanisms are not sufficiently understood, polydactyly is thought to be
related to a defect in the sonic hedgehog (SHH) signaling pathway during
limb development (42,43), and obesity may be due to defects in leptin, NPY,
and MCHR1 signaling pathways in the hypothalamus (7,8,44,45).
Specific roles of individual BBS proteins are also emerging (Figure 32-
1). Of the 21 BBS proteins identified thus far, eight (BBS1, BBS2, BBS4,
BBS5, BBS7, BBS8, BBS9, and BBS18 [aka BBIP1]) form a stable complex,
known as the BBSome (46,47). This complex is thought to mediate protein
trafficking in and out of cilia by interacting with intraflagellar transport (IFT)
particles, which transport ciliary proteins along the axonemal microtubules
(40,46,48–52). BBS3 (aka ARL6) is a member of the Ras superfamily of
small GTPases and controls BBSome recruitment to the plasma or ciliary
membranes (50,53). BBS17 (aka LZTFL1) is currently thought to mediate
the interaction between the BBSome and IFT particles and facilitate ciliary
exit of the BBSome (43,50,52). BBS6, BBS10, and BBS12 have sequence
homology to the CCT/TRiC family of group II chaperonins (54–57). These
proteins form another complex, the BBS/CCT chaperonin complex together
with six CCT chaperonins (CCT1, CCT2, CCT3, CCT4, CCT5, and CCT8),
and facilitate BBSome assembly (58). Consistent with their closely related
molecular functions around the BBSome, null mutations in these 13 BBS
genes result in typical BBS phenotypes described earlier, and no obvious
genotype–phenotype correlation has been observed.
The rest of the BBS proteins are more remotely linked to the BBSome.
BBS13 (aka MKS1), BBS14 (aka CEP290), BBS15 (aka WDPCP), and
BBS16 (aka SDCCAG8) localize to the transition zone at the ciliary base (59)
(and references therein). These proteins are part of a protein network that
builds the ciliary gate and controls entry and exit of ciliary proteins. BBS19
(IFT27) and BBS20 (IFT172) are components of the IFT complex. Null
mutations in these genes result in phenotypically overlapping but distinct
ciliopathies such as Meckel-Gruber syndrome (MKS), Joubert syndrome
(JBTS), Senior-Löken syndrome (SLSN), and Jeune syndrome (27,28) (and
references therein). In contrast, mutations in these genes found in patients
with a clinical diagnosis of BBS are usually hypomorphic alleles, indicating
that partial loss of these protein functions results in BBS-like phenotypes.
Although the precise molecular function of BBS21 (C8ORF37) is currently
unknown, inactivation of BBS21 also results in OS morphogenesis defects
(60).

The age of onset of BBS is at conception when a fetus inherits two abnormal
copies of one of the BBS genes. Typically, one mutant allele is inherited from
each parent, although uniparental disomy for an abnormal BBS gene or
inheritance of one abnormal gene and spontaneous mutation of the other
allele are also theoretical possibilities. Although age of onset is at birth, age
at diagnosis is often delayed as many features are protean until the child is
several years of age.
Polydactyly
The diagnosis of BBS may be suspected at birth if polydactyly is present.
This may be manifested as extra fingers, toes, or both. The supernumerary
digits are typically extra-axial near the little finger or toe, although there is a
report of an extra finger between the third and fourth digits (61). Digits may
be rudimentary or fully formed. An unrelated syndrome of isolated autosomal
dominant polydactyly also exists and is relatively common, so extra digits
may be ligated and removed shortly after birth leaving only tiny scars.
Parents may not mention this on history, and patients may be unaware that
extra digits were present at birth. For this reason, specific questions about
extra digits should be asked, and hands and feet examined if BBS is
suspected. Sixty-three to eighty-one percent of BBS patients have
polydactyly, which may involve all four limbs or any combination of upper
or lower (3).
Retinal Degeneration
In the absence of polydactyly, the first symptom of BBS is often decreased
vision in dim light, typically noticed around the age of 5 to 7 years. The
electroretinogram (ERG) may be abnormal before this; in a large cohort of
families with BBS, all patients with genetically proven BBS who were older
than 3 years had some evidence of retinal degeneration (62). Cone–rod
dystrophy has been reported in 93% of BBS patients (3). What begins as
night blindness progresses to visual field loss and RP. Visual acuity is often
good in the early stages of retinopathy, and since young children have limited
independent activities in dim light at this age, specific questions must be
asked to elicit symptoms. Questions such as “Can he/she find a seat in a dark
movie theater without assistance?” and “Can he/she see stars at night?” are
often illustrative. Parents may report noticing that the child has difficulty
ambulating when camping at night or trick-or-treating on Halloween. In
bright light, the child may appear to see perfectly normally. Nystagmus has
been reported in 59% of patients (62) but is not considered a typical finding
in BBS. 43.7% of patients in one study developed cataracts (62). Most
patients in this study were legally blind by late teens or early adulthood, but
no patient lost all light perception. At a young age, the fundus appears
normal. Many patients develop a “bull’s-eye maculopathy” appearance,
which may begin as a blunted fovea (Figure 32-2). The peripheral vision
decreases steadily, but central vision, while imperfect, is retained longer.
When central vision does decrease, it may be attributable to cystoid macular
edema (CME) and/or epiretinal membranes. Over time, atrophy of the macula
occurs (Figure 32-3). On slit lamp examination of the anterior vitreous in
childhood, a significant cellular reaction can be seen (63) (Figure 32-4). This
may precede a decline in vision (Drack, unpublished observations). The cells
seen in the vitreous may be an inflammatory or autoimmune overlay related
to the retinal degeneration but may also be debris related to cell death.
Posterior subcapsular cataracts may develop over time. Legal blindness often
develops by the time patients are in their 20s or 30s due to severely decreased
visual field, maculopathy causing poor acuity, or both. A typical RP picture
with waxy pale disc, thinned arteriole, and bone spicule–like pigmentation
eventually occurs (Figure 32-5). In mouse models of BBS, a striking loss of
outer retinal layers is seen on histology (Figure 32-6).
FIGURE 32-2 Left fundus of a 10-year-old with BBS1.
Visual acuity was 20/70 right and 20/40 left. The child had
been followed since age 3 years for nystagmus, esotropia,
and amblyopia as well as obesity. Referral was made when
nyctalopia was noted.
FIGURE 32-3 Red free and OCT of the right fundus
shows subtle bull’s-eye appearance and thinning of outer
retinal layers. Note the relatively preserved outer nuclear
layer just beneath the foveal depression. Outer segment
lengthening has been lost.
FIGURE 32-4 Anterior vitreous cellular reaction in the

right eye of the same patient as in Figure 32-3.
FIGURE 32-5 Bone spicule–like pigmentation in the
retinal periphery of a 15-year-old with BBS.
FIGURE 32-6 Summary of BBS mouse model ocular
phenotypes characterized by our research group. Images
from mutant mice (B,D,F,G,I) are compared to normal
wild-type (WT) controls (A,C,E,H); (A) shows a WT
mouse retinal fundus; (B) shows fundus abnormalities
including arteriolar constriction, scalloped hypo- and
hyperpigmentation, and pale optic nerve in a Bbs3-/-
mouse; (C) shows a normal ERG response in a wild-type
mouse; (D) demonstrates loss of ERG waveform in a 5-
month-old BBS mouse; (E) shows normal histology of the
retina in a WT mouse and progressive retinal degeneration
(F,G) in BBS mice retinas at two points in time; (H,I)
show ultrastructural abnormalities in BBS mouse outer
segments (I) compared to control retina (H) on electron
microscopy.
Obesity
Obesity may precede vision loss but is infrequently suspected to be due to
BBS unless an astute clinician searches for other features. The obesity is
truncal. While infants with BBS are usually of normal weight, many children
are noticeably obese by 1 to 5 years of age. The obesity that accompanies
BBS is especially difficult to manage. Studies in mice and humans have
demonstrated that it is due to leptin resistance (7,64). Leptin is a hormone
that triggers a feeling of satiety, so resistance renders those afflicted with
BBS not feeling full after eating a sufficient quantity of food. In addition,
animal studies have shown that even when eating the same amount of food as
unaffected littermates, BBS mice gain more weight (7), suggesting that
energy expenditure also plays a role. Obesity has been reported in 72% to
92% of patients (3). Adolescents and adults with BBS often are able to better
control food intake than young children who cannot yet understand that they
must stop eating before satiety.
Renal and Genital Anomalies
Kidney disorders occur in 53% of patients (3) and may be progressive. One
study suggests that BBS10 patients may have the most severe renal disease
(62). Renal ultrasound evaluation should be recommended early in the course
and followed by a kidney specialist if abnormal. Urine concentrating ability
may be diminished leading to copious urine production and difficulty with
bed wetting. Genital anomalies have been reported in up to 98% of patients
(3). Although BBS mice models do not have significant renal impairment,
spermatogenesis is impaired and males are infertile (19–22).
Developmental Disability
Developmental delay and psychiatric disorders are overrepresented in BBS
patients but are not universal. In one study, patients with BBS due to
mutations in BBS1 or in BBS12 appeared to have milder cognitive disorders;
many BBS patients have university degrees (62). About 61% of BBS patients
have been reported to have learning difficulties and 50% to 91% have been
reported to have developmental delays (3), but poor vision may play a role in
testing. In a behavioral study of 21 BBS patients, repetitive behaviors,
obsessions, and autistic features were noted, but none had diagnosable
autism. Anxiety, depression, and difficulty with social interactions and with
attention were also noted (65). BBS mice exhibit a defect in social dominance
phenotype (21).
Other Features
Later-occurring complications include diabetes mellitus and hypertension, but
these can present in childhood especially in the setting of obesity. Some
patients have heart defects. Anosmia has been reported in 60% of patients
(3), although patients may not be aware of it since it is present from early life;
BBS2 mice have anosmia (21,66). Hearing loss and “glue ear” (serous otitis
media) have been reported in some BBS patients. Surveillance for these
disorders should commence in childhood, and the importance of close follow-
up should be discussed.
Diagnostic Studies
When BBS is suspected based on ocular history or physical findings, workup
begins with a history and full eye examination. Careful inspection of the
hands and feet for surgical scars or remnants of extra digits should be
performed. The retina may appear normal early in the course even though the
ERG is not; therefore, a prudent first step is an ERG. This is a challenging
test for many children to perform, especially if they have developmental or
behavioral issues as is often the case with BBS. Using DTL electrodes rather
than contact lens electrodes may make an otherwise unobtainable
examination possible. If an ERG is not possible while the child is awake,
ERG with sedation or anesthesia should be considered keeping in mind that
ERG amplitudes may be variably reduced by up to 50% by anesthetic agents.
Sedation and anesthesia have a small but significant risk of complications.
Therefore, if the clinical picture is typical and ERG cannot be performed
easily, doing genetic testing before ERG is advisable. The ERG in BBS early
in the course is usually a rod-cone dystrophy, but cone-rod pattern has also
been reported (67). Later the ERG eventually becomes nonrecordable.
Optical coherence tomography (OCT) may be easier for children to cooperate
with and shows thinning of the outer retina, but this may not be detectable
very early in the disease. Kidney ultrasonography should be recommended
once the diagnosis is made.
Genetic Testing
Approximately 80% of patients will have a molecular diagnosis on genetic
testing (4). Certainty of diagnosis is important to rule out other causes of
retinal degeneration in children and to put into place the proper surveillance
for systemic disorders. In addition, since this is an autosomal recessive
disorder, genetic counseling for recurrence risk is important for families.
Clinical laboratories testing for BBS can be found at www.genetests.org.
Many patients have novel mutations, so complete sequencing of all of the
known BBS genes is often necessary. A certified genetic counselor or
medical geneticist may be consulted to help patients and families understand
the implications of genetic testing results and to counsel about recurrence
risk. It is important to remember that at this time, a negative genetic test
never rules out a disorder due to genes yet to be found and limitations of
testing. Identification of rare variants predicted to change the protein product
of a gene is often, but not always, diagnostic. Every person has a large
number of variations from the norm in their genome, and most are not
pathologic. When a mutation is found in a BBS gene on genetic testing, it
must be evaluated for pathogenicity by determining whether there are
mutations on both of the patient’s alleles (often achieved by testing parental
samples), whether the change has been reported in other patients, whether
and how often it is found in normal controls, and whether or not a conserved
amino acid would be changed in the final protein. Some genetic changes
cannot be classified definitively as disease causing versus benign at this time.
Most genetic testing laboratories provide this data in the test report; it is
outside of the expertise of most clinicians to assess which molecular changes
are likely to be pathologic, and even for experts the knowledge used to make
these determinations is being continuously acquired.
All patients who can cooperate should have a slit lamp biomicroscopic
examination to look for cells in the anterior vitreous and subsequent posterior
subcapsular cataract. If central acuity is diminished, OCT is helpful to
diagnose CME.
Several disorders overlap clinically with BBS despite being caused by genes
that do not cause typical BBS. In other cases, mutated genes, which cause the
clinical phenotype of disorders such as Leber congenital amaurosis (LCA)
can also cause the clinical phenotype known as BBS. Our nomenclature is in
flux as our knowledge expands. In addition, there may be phenocopies of
BBS such as autosomal dominant polydactyly plus retinitis pigmentosa or
obesity plus retinitis pigmentosa. Molecular genetic diagnosis is especially
important to differentiate these overlap syndromes.
When a child presents in the 5- to 7-year-old age range with either
nyctalopia or decreased vision, with or without nystagmus, BBS should be in
the differential diagnosis. If visual acuity is poor and nystagmus is present,
LCA should also be considered. Two types of LCA, caused by mutations in
CEP290 and NPHP5, may overlap BBS with early-onset retinal degeneration
and renal and/or neurologic disease. Patients with CEP290-associated LCA
may also have anosmia. If a bull’s-eye maculopathy is present, Batten disease
should be considered. This is a progressive neurodegenerative disorder that
begins with rapid retinal degeneration (see Chapter 24). Stargardt disease (see
Chapter 30) presents with a similar macular appearance and age of onset but
is an isolated retinopathy. If Stargardt disease is suspected, genetic testing for
ABCA4 and ELOVL4 should be considered. If obesity is present, BBS and
Alström rise to the top of the list. Alström syndrome shares obesity, retinal
degeneration, hearing loss, “glue ear,” and some other features of BBS but is
caused by mutations in the ALMS1 gene (68). Alström syndrome is also a
ciliopathy, but presents with photophobia in most cases. Orofacial–digital
syndrome (OFDS or Mohr syndrome) may be in the differential diagnosis
due to digit anomalies but can be differentiated clinically by a bifid thumb
and tachypnea/hypoventilation episodes. Like BBS, OFDS encompasses
multiple genes and ciliopathy syndromes; however, unlike BBS, only one
type includes a progressive retinal dystrophy (69). Other disorders in the
differential diagnosis include McKusick-Kaufman syndrome, JBTS,
Biemond syndrome, and obesity plus congenital stationary night blindness.
Management
The only prevention for BBS itself is carrier testing and subsequent family
planning for identified carriers. If carrier state is known and genotyped,
preimplantation genetic testing combined with in vitro fertilization can be
considered to reduce the risk of an affected child. Unfortunately, at this
writing there is no way to prevent the retinal degeneration from occurring in
patients with biallelic BBS mutations, even though the retina is normal or
very mildly affected for the first several years of life. This long span of time
before retinal degeneration occurs should afford time for a preventive
treatment, and significant research is ongoing in this endeavor.
If CME is present, a systemic or topical carbonic anhydrase inhibitor
(CAI) such as dorzolamide may be prescribed. This has been shown to
improve both anatomy and acuity in some patients with different types of RP
(70). Topical brinzolamide may be better tolerated by children because it
does not sting upon installation (Drack, unpublished data).
It is unknown whether treating the vitreous cellular reaction in RP, and
specifically in BBS, is beneficial. While the disease is not primarily
inflammatory, just as the inflammatory response to a disease or infection may
cause secondary damage in other parts of the body, the same may be true in
the eye. If the vitritis is inflammatory, steroids, nonsteroidal anti-
inflammatories or biologics may have a role in treatment. Side effects of
steroids can be significant, particularly elevated blood glucose in susceptible
individuals such as those with BBS. More research is necessary into this
aspect of genetic retinal disorders (63).
Experimental approaches to intervene in the retinal degeneration that
occurs in BBS are similar to those of other retinal dystrophies. For the
purpose of brevity, we will discuss three areas of intervention: light
modulation, antiapoptotic agents, and gene transfer.
Modulation of light exposure may be of benefit in a variety of retinal
degenerative diseases, as there is evidence from animal models that increased
light exposure is harmful and avoidance of light exposure may be beneficial
(71). Blue light (wavelength 400 to 480 nm) appears to be especially
damaging through a variety of mechanisms (72,73). The efficacy of light
reduction in RP has not been proven. There are anecdotal reports of slowed
progression when wearing tinted contact lenses; however, there is also a
published report of a patient with RP and unilateral cataract who did not have
any apparent preservation of retina in the eye with limited light exposure
(71). In a mouse model of BBS1 homozygous for the p.M390R mutation,
little preservation of retinal function was seen with darkrearing, and operating
microscope light exposure did not increase the rate of retinal degeneration
(74).
Tauroursodeoxycholic acid (TUDCA) is a bile acid found in bear bile,
which is used in traditional Chinese medicine. In using this substance to treat
liver disorders, it was noted that in addition to the bile acid effect, TUDCA
also appeared to decrease apoptosis, the programmed cell death that leads to
organ failure. Studies in animal models of neuro and retinal degeneration
showed promising results (75,76). In the Bbs1M390R/M390R mouse model, we
found that administration of TUDCA significantly slowed the retinal
degeneration as evidenced by both ERG and histologic examination (77). In
addition, Bbs1 mice that received TUDCA had significantly lower body
weight than untreated mice (77). In humans, TUDCA has been found to
improve insulin utilization (78). These findings make TUDCA a prime
candidate for human clinical trials of treatment for BBS, and possibly
Alström syndrome. Since TUDCA is not FDA approved, patients should not
attempt to procure it and treat themselves. The optimal dose for treatment in
humans is not known. UDCA is a related compound, which is FDA
approved, but the taurine moiety it lacks may be important to the desired
effect.
Subretinal gene therapy in animals and humans can improve retinal
function in at least one type of genetic retinal degeneration, RPE65-
associated LCA (79,80). Subretinal gene therapy for this indication is now
FDA approved for children as young as 1 year of age.
Since the genetic defect resides in the RPE, the layer under the
photoreceptors, it is an ideal candidate for this type of therapy. The
ciliopathies may present challenges to gene replacement since the gene
products of several genes must act in concert and establishing the appropriate
ratio of BBSome components may be important.
Some success in improving retinal health over small areas of retina has
been reported with subretinal gene replacement therapy in the mouse model
of BBS4 (81). Our group has shown small improvements in ERG in Bbs1 and
Bbs2 mice as well as biochemical rescue of the BBSome (82); however,
overexpression toxicity has also been noted (83). Subretinal gene therapy in
the Bbs10-null mouse has shown early promising results (84).
Attempts to prevent obesity may be successful if eating habits are
carefully taught and monitored. Likewise, diabetes, hypertension, and renal
failure may be ameliorated by early dietary intervention and close monitoring
with medication as indicated.
A low-vision specialist should advise the school about methods to improve
education. Enlarging print materials through the use of a CCTV and the use
of auditory and other technology can be life changing for patients with low
vision. Patients should be advised to carry a flashlight at all times in case they
find themselves inadvertently in dimly lit surroundings. Some patients may
have vision adequate for driving in the teenage years, but since the retinal
degeneration is progressive, this should be monitored closely. See also
Chapter 14.

Since BBS is a multisystem disorder, it is best managed through a
coordinated effort of dedicated specialists who can address each of the
separate aspects of the disease while working together for the good of the
patient. Communication with geneticists and primary care doctors is
imperative since these patients will need lifelong follow-up for systemic
complications. Evaluation for kidney disease should be performed at
diagnosis. Referral to nutritionists and educational specialists is
recommended. An endocrinologic evaluation around puberty is beneficial.
Primary care doctors should be informed of the underlying syndrome and the
need for specialized surveillance.
As in all genetic eye disorders, physicians have an ethical obligation to
inform patients/parents that the condition may be genetic and that accurate
genetic testing is available. Consultation with medical geneticists, genetic
counselors, and/or reproductive care geneticists should be offered as in vitro
fertilization with preimplantation genetic testing may be desired by some
families to reduce the risk of having future affected offspring. Genetic
information should be offered in a nonjudgmental manner, referring
patients/parents to other specialists as needed. There is also an obligation to
inform patients/parents that there is a plethora of ongoing research that may
be applicable to them now or in the future and to offer websites such as
www.clinicaltrials.gov, referral to other specialists, and other resources to
help manage their condition.
Future Treatments
Ongoing research into gene therapy for retinal degenerative conditions in
which many cells are still viable offers hope of gene replacement therapy in
the future for BBS. The success of in vivo correction of genetic mutations
using oligonucleotides or CRISPR is also encouraging (85,86). For patients
in whom too many retinal cells have been lost, stem cell replacement of the
photoreceptors is under active investigation (87).
CONCLUSION
BBS has been considered an extremely rare disorder that few
ophthalmologists will see in practice. We now know it is far more
pleiomorphic, and likely more common, than was thought. The interesting
finding of multiple genes leading to the same striking phenotype has led to
knowledge about how cilia affect every organ system in the body and to the
understanding of the interaction of multiple genetic protein products in a new
way. Finally, BBS is a prime example of how not only our understanding but
also our naming of clinical entities is evolving in the genetic era. Whereas
BBS was initially confidently diagnosed as a combination of obesity, RP, and
polydactyly with LMS separate due to lack of polydactyly and presence of
spastic gait, we now know that there are myriad overlapping disorders due to
mutations in genes that contribute to the same protein complexes and cilia
functioning that can cause parts or all or additions to what was initially called
BBS. The nomenclature is now a combination of the name of the gene
affected (e.g., BBS1, BBS2, etc.), and the clinical features, for example,
patients with mutations in CEP290, may be called either BBS or LCA
depending on the patient’s clinical manifestations. While this can be
confusing, the primary goal remains the same: to give patients and families
an accurate diagnosis in order to help them manage their disorder and give
them hope that continuing research will find treatments and eventually cures.
ACKNOWLEDGMENTS
The authors wish to acknowledge Dr. Val Sheffield for generation of mouse
models and his pioneering work on Bardet-Biedl Syndrome. Supported in
part by the Ronald Keech Professorship (Drack).
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33
Usher Syndrome
Alaa Koleilat, Raymond Iezzi Jr, and Lisa A. Schimmenti
PREVALENCE/WORLDWIDE IMPACT
Usher syndrome (USH) is the most common genetic cause of deafness and
blindness worldwide. In the general population, USH impacts 4 to 17 people
per 100,000 but can be diagnosed in 3% to 6% of all deaf and hard-of-hearing
children (1–3). USH is considered a rare disease in the general population. In
the United States, a rare disease is defined as a condition that affects fewer
than 200,000 Americans. However, among the deaf/hard-of-hearing
population, it is a common condition, supporting the need for greater
awareness of this condition.
As hearing loss precedes vision loss in USH, the diagnosis may not be
made until the second decade as the loss of night and peripheral vision
impacts activities such as driving. The only way to make a definitive
diagnosis of USH prior to the loss of vision is through genetic testing;
however, electroretinogram (ERG) may be suggestive since the ERG often
becomes abnormal before major symptoms are noticed. USH is a public
health concern and, for deaf/blind adults who are visual communicators, can
result in social isolation, loneliness, and depression (4); therefore, early
identification and diagnosis can impact decision-making for communication
and, in time, may identify individuals who could benefit from emerging
therapies.
Usher syndrome is a genetic condition, and there is no current literature that
indicates any environmental factors contributing to the disease.
GENETICS
Usher syndrome is a genetic disorder that has an autosomal recessive
inheritance pattern. This means that an affected individual would need to
inherit mutations in the same Usher gene that can be different from one
another: one of which comes from the biologic mother and one from the
biologic father.
There is no X-linked nor sex limitation to USH; females and males are
equally likely to inherit the disease. Ten genes have been identified to cause
Usher syndrome as well as one modifier gene (Table 33-1). If each parent
carries one copy of the same gene with a pathogenic variant, then each child
has a 25% chance of inheriting both abnormal copies and will be born with
Usher syndrome. A parent that is a carrier for a gene that causes Usher
syndrome will not present with the disease, and may or may not have a
family history.
TABLE 33-1 List of genes identified to cause

Usher syndrome
PATHOPHYSIOLOGY
Usher Proteins in the Hair Cell
The overall process of hearing is the translation of a mechanical signal to a
chemical signal in the cochlea. It has been estimated that the cochlea has over
20,000 hair cells. Each hair cell has a collection of small actin-filled
projections at the apical end of the cell called stereocilia (25); they are
typically ordered from shortest to tallest. When a hair cell is deflected toward
the tallest stereocilia due to sound, tip links composed of PCDH15 and
CDH23 cause the mechanotransduction (MET) channels—located at the top
of the stereocilia or lower tip link density—to open, allowing positively
charged ions (K+ and Ca2+) to flow into the cell (25–30). The increase in
cations in the hair cell changes the membrane potential necessary for
depolarization. L-type voltage-gated calcium channels (Cav1.3) open in
response to depolarization and increase intracellular calcium (31). Calcium is
critical in the hair cell as it mediates the release of glutamatergic vesicles
tethered to the ribbon synapse, a structure not only important for transporting
neurotransmitters but also necessary for clustering of Cav1.3 channels at the
presynapse and for stabilizing contact with afferent neurons. Once vesicles
from the ribbon synapse fuse with the plasma membrane, their contents are
released into the synaptic cleft and bind onto AMPA receptors for synaptic
transmission (32,33) (Figure 33-1). Variants in many genes involved in this
process can cause hearing loss; however, variants in genes involved
specifically in the mechanotransduction process can cause Usher syndrome
and prevent the mechanical signal (sound deflection) from being converted to
a chemical signal (neurotransmitter release).
FIGURE 33-1 This illustration of a typical human hair
cell depicts localization of Usher proteins myosin7A, sans,
harmonin, cadherin 23, and protocadherin 15 (inset) and
their role in the process of stereocilia deflection and tip
link structure.
Usher Proteins in the Photoreceptor

Patients with Usher syndrome can experience retinitis pigmentosa as early as
the first decade of life. It is a progressive degeneration of photoreceptors that
initially leads to loss of peripheral vision and poor night vision. Although the
degeneration is mostly due to rod dysfunction, cones can degenerate later in
the progression of the disease leading to reduced central vision as well. Usher
proteins form a complex involved in vesicle trafficking near the
photoreceptor synapse but also in trafficking at the connecting cilia (34).
Based on studies in the mouse model (sh-1), myosin7a—an unconventional
motor protein—has been implicated in the transport of opsin from the inner
segment to the outer segment of the photoreceptors (35,36). Additionally,
myosin7a has been detected to play a role in the phagocytosis of distal outer
segment disks as well as melanosome transport in the pigmented retinal
epithelium (37,38). There are many USH1 and USH2 proteins that are
involved in the ciliary region of photoreceptors. Specifically, protocadherin
15, an USH1 protein, has been implicated to be involved in alignment of
outer segment disks (37). David S. Papermaster in 2002 (39) reported that
usherin, VLGR1b, and sans, all USH2 proteins, are associated with the
periciliary ridge complex. Additionally, the extracellular domains of usherin
and VLGR1b are part of the connecting cilium between the membranes of the
inner segment, and the intracellular domains of these proteins anchor to
whirlin in the cytoplasm (Figure 33-2). One of whirlin’s main roles is to link
to sans and myosin7a (40). Whirlin has been shown to localize to the
periciliary complex of the photoreceptor (41).
FIGURE 33-2 Illustration of the photoreceptor and
localization of usherin and myosin7A proteins. Usherin
localizes to the connecting cilium complex, while
myosin7A localizes to the periciliary complex as well as
pigmented retinal epithelium.
Usher syndrome is subdivided into three clinical types depending upon the
causal gene, severity of hearing loss, and the onset of retinitis pigmentosa.
This classification was originally established by Davenport and Omenn
(42,43). Patients with Usher syndrome type 1 (USH1) have severe to
profound hearing loss (deafness) present at birth; demonstrate abnormal
balance, typically walking after 18 months of age; and experience decreased
night vision within the first decade of life progressing to severe vision loss.
Patients with Usher syndrome type 2 (USH2) have moderate to severe
hearing loss present at birth and decreased night vision by adolescence
progressing to severe vision loss. They do not have balance issues or delayed
walking. Those with Usher syndrome type 3 (USH3) present with progressive
hearing loss in childhood to early teens; these patients also vary in the
severity and age of onset of the vision problems and typically have
normal/near-normal balance in childhood, but this may change later in life
(Figure 33-3).
FIGURE 33-3 Representative audiograms for each type of

Usher syndrome. Audiograms for USH1 and USH2 were
generated using data from patients with a confirmed
genetic diagnosis.
(Data used to generate USH3 audiogram was from the best hearing ear from Ness SL, Ben-Yosef T,
Bar-Lev A, et al. Genetic homogeneity and phenotypic variability among Ashkenazi Jews with Usher
syndrome type III. J Med Genet 2003;40(10):767–772, Ref. (44).)
DIAGNOSTIC STUDIES
Newborn hearing screening occurs in nearly all developed nations, and it has
been a topic of discussion since the late 1960s. Newborn hearing screening is
now part of the standard of care for newborns that identifies children who are
at risk of having hearing loss and require additional testing. It also has been
instrumental in identifying children who have medical conditions that can
cause late-onset hearing loss. In 2016, over 94.8% of babies born in the
United States were screened for hearing loss before 1 month of age (CDC,
2016), and over 6,100 infants were identified and diagnosed with hearing
loss. If an infant fails the newborn hearing screening, they will be referred to
an audiologist for further testing. A failed hearing assessment does not
automatically suggest hearing loss; a baby may have other reasons for not
passing the newborn hearing screen such as fluid in the ear canal. However, if
an audiologist confirms hearing loss, one of the first actions is to determine
the type of hearing loss. More than 50% of hearing loss in newborns has a
genetic basis (43,45); therefore, genetic testing is typically the next step in
determining a diagnosis. Arun Sharma and colleagues reported that about
20% of children with sensorineural hearing loss also have vision problems;
therefore it is important to detect genetic hearing loss as early as possible,
because patients may be identified presymptomatically for vision loss (46).
This is particularly important for patients with Usher syndrome, because
onset of vision loss is different depending on the type of Usher syndrome and
will, therefore, affect the timeline for treatment and management of the vision
loss. In addition, since all patients with hearing loss depend to some degree
on watching lip movements to help understand speech, easily correctable
problems such as refractive error that might interfere with lip reading can be
diagnosed and treated.
The clinical evaluation of a patient with retinitis pigmentosa includes a
full medical history, nutritional history, and history of vision symptoms,
focused on nyctalopia, decreased visual acuity, and glare. Further, a family
history is critical. Ideally, a comprehensive eye examination should include
funduscopy, optical coherence tomography (OCT), visual field testing, and
full-field electroretinography, documenting A-wave and B-wave amplitudes
as well as implicit time. Examinations must be tailored to the age of the
patient, and, if necessary, some aspects must be performed under anesthesia.
With the advent of genetic testing, diagnosis can often be made with
molecular genetics rather than by subjecting a child to general anesthesia for
testing. Examination findings of retinitis pigmentosa may include peripheral
bone spicules, optic nerve pallor, attenuation of the retinal vasculature,
cataract, and in some cases cystoid macular edema (CME), low-grade ocular
inflammation, and epiretinal membrane.
Edwards et al. (47) made the observation that visual field area and visual
acuity are more impaired in USH1 patients than USH2; however, a more
recent publication reports that visual field area, visual acuity, and the
incidence of macular lesions did not differ between USH1 and USH2 patients
(48). Seelinger et al. used electroretinography testing to show that although
patients with USH 1 and USH2 have similar loss of amplitude, there are
significant differences in implicit times or peak times (49). Patients with
USH2 had marked delay in implicit times, whereas USH1 patients had
normal to moderate delays. In 2002, Tsilou et al. (48) also reported that
electroretinographic amplitude did not differ between USH1 and USH2
patients. Testa et al. assessed the distribution of macular abnormalities in
USH patients using OCT (50). They observed that USH1 and USH2 patients
have a high prevalence of CME and epiretinal membrane. Consequently,
OCT screening in USH patients plays an important role in their management
and treatment. Lastly, videonystagmography is used to detect involuntary eye
movements that might indicate balance abnormalities.
MANAGEMENT
One of the most important aspects of managing USH is early identification.
At this time, nearly all USH1 and USH2 patients will be diagnosed to be deaf
or hard of hearing, but genetic testing can determine presymptomatically if
that child will go on to have vision loss later in childhood or teenage years.
Christine Yoshinaga-Itano and her colleagues reported that there is
significantly better language development when children are identified to be
deaf or hard of hearing before 6 months of age (51), thus providing evidence
for the importance of newborn hearing screening. Additionally, early
identification of children who are deaf and may go on to have vision
impairment allows families and intervention specialists time to develop the
support and acquire the resources necessary to address both developmental
and educational needs. Some patients with USH2 may pass the newborn
hearing screen; if there is a family history of USH2, early genetic testing may
be appropriate. Proper management and treatment for patients with USH
requires an interdisciplinary team approach once they are identified.
The treatment plan for affected individuals depends on the type of USH
they are diagnosed with; for example, patients with USH1 are profoundly
deaf and typically may not experience benefit from hearing aids. Cochlear
implantation would be of benefit for children diagnosed with USH1, and
more specifically early implantation at or before 1 year of age will provide
the most benefit for developing oral language in anticipation of the fact that
vision loss may become severe enough to make American Sign Language
difficult. Although tactile sign language is a possibility, the rarity of
interpreters with expertise in tactile sign language limits its utility to allow
ready access to communication.
Patients with USH1 typically suffer from vestibular abnormalities. Thus,
early identification through newborn hearing screening and genetic testing
can be of benefit for planning physical therapy needs. For children and adults
with Usher syndrome type 1, balance problems related to vestibular
anomalies pose significant issues. In childhood, walking is delayed past 18
months of age. In older children, difficulty with activities that require balance
such as bicycling or skating may limit the ability of the child to participate
fully. Adults may have difficulty with balance and may experience falls.
Physical therapy to help with balance and fall risk can be of benefit. Many
patients make remarkable adaptations and learn to participate in many
activities requiring balance over time.
At the present time, no FDA-approved treatments for USH related to
retinitis pigmentosa are available. In 1993, a large randomized clinical trial
was conducted over a 6-year time span in which adults with retinitis
pigmentosa—including USH2 patients—ingested 15,000 IU vitamin A
palmitate supplements daily. This study showed that, on average, patients
taking vitamin A palmitate experienced a 20% slower annual decline of ERG
amplitude loss. This study did not demonstrate preservation of visual field,
however. Therefore, it is unclear whether supplementation with vitamin A
palmitate is of therapeutic benefit to patients with USH associated retinitis
pigmentosa (52). High doses of vitamin A can be toxic and medical
monitoring is required for long-term use. Additionally, patients with USH2
may experience CME. This is often associated with mild to moderate
decreases in visual acuity.
Treatment with topical carbonic anhydrase inhibitors may be effective in
up to 30% of patients (53). If the CME does not respond to topical treatment,
oral carbonic anhydrase inhibitors may be effective. Systemic treatment
requires monitoring of liver enzymes, electrolytes, white blood count, and
maintenance of hydration. Side effects may include dose-related tingling in
the hands and feet that may improve over time or with dosage adjustment.
Additional side effects include fatigue, alteration of taste, gastrointestinal
upset, and nephrolithiasis. Patients should be informed of these issues prior to
starting treatment and monitored over the course of treatment. For those
patients who cannot tolerate systemic carbonic anhydrase inhibitors,
sustained-release intraocular dexamethasone implants may induce temporary
resolution of CME. Long-term repeat injections may be required to treat the
CME; however, this may cause steroid-induced ocular hypertension (54).
The care of individuals with Usher syndrome is best provided through a
team approach. This team includes experts in Audiology, Ophthalmolgy,
Otorhinolaryngology, Genetics, Pediatrics, Speech and Language Pathology,
and Rehabilitation Medicine. As patients will present with hearing loss first,
particularly if hearing loss is picked up by newborn hearing screening or by
language delay in the first 2 years of life, the early years of management will
be primarily through Audiology, Otorhinolaryngology, and Speech and
Language services. Although many deaf/hard-of-hearing children will be
referred for vision examinations, a normal eye examination in early
childhood does not rule out Usher syndrome. Genetic evaluations and testing
are part of the care for all deaf/hard-of-hearing children; genetic testing
panels that include all known genes for Usher syndrome as well as other
genes associated with early-onset hearing loss are recommended. Positive
genetic testing for Usher syndrome will provide guidance for early
management in anticipation of onset of vision loss in later years.
VISUAL REHABILITATION
Low vision services that include support for reading, daily activities, and use
of a cane for mobility would be similar to patients affected with retinitis
pigmentosa of other causes; however, tactile interventions may be necessary
for older patients with USH who have not had cochlear implants (see also
Chapter 14, low vision).

Social issues for individuals with hearing loss followed by lost vision are
significant. Deafness/hearing loss poses its own set of challenges in terms of
social isolation particularly within the hearing community, but for deaf/blind
individuals particularly those with Usher syndrome type 1 who are
profoundly deaf and then start losing vision, there may be barriers to full
employment and other social activities. Active effort is required on the part of
employers and schools to support dual sensory impairments that include
strategies for full inclusion under the Americans with Disabilities Act (55).
Deaf/blind individuals are often eligible for drivers and home aide workers
free of charge due to their disabilities, so referral to social services is
important.
Educational issues for individuals with Usher syndrome change
throughout the lifetime and are impacted by communication modality, oral,
or sign language. Early on, children with Usher syndrome may receive
school-based services based on their hearing impairment. As children enter
their teen years, and the first significant vision impairments occur, services
are needed to support both vision and hearing issues for classroom success. It
is not unusual for a teen with Usher syndrome to first be diagnosed when
they do not pass the peripheral vision testing required for licensure when
applying for their driver’s license for the first time. Issues surrounding
independence related to driving need to be addressed in a supportive manner
with referral to other professionals as needed.
With the increased access and ease of genetic testing, ethical dilemmas arise
related to the implications genetic variants have on families. Although
genetic testing is extremely beneficial to diagnosing disease and determining
prognoses for patients, the interpretation of results can be complex and
should be provided in the context of genetic counseling. Additionally, as
Usher syndrome is an autosomal recessive condition, genetic testing results
will have implications for future offspring; thus, genetic counseling is
warranted.
Many USH patients may define themselves as being culturally Deaf,
meaning that they communicate with American Sign Language and are part
of the Deaf (capital “D”) Community, Deaf individuals do not view deafness
as a disability or medical condition, but rather another aspect of their life that
can be managed and does not inhibit their ability to function in their daily
lives. This should be taken into consideration especially if one or both parents
are deaf or consider themselves to be culturally Deaf; they may not choose to
support medically treating the deafness or supporting oral language for their
child. Thus, information about language choices should be presented in a
nonjudgmental manner allowing parents to make choices for their children
while providing full information about future vision loss, which may limit
visual language ability.
FUTURE TREATMENTS
Since the completion of the Human Genome Project, there have been
significant improvements in genetic sequencing technology that increase
speed and lower costs. Currently in 2019, patients can have their entire
genome sequenced for a few thousand dollars. With over 110 chromosomal
loci and more than 65 genes identified to cause hearing loss, genetic
sequencing is critical to identify a diagnosis as well as proactively plan for
the vision loss in USH patients (56). Genetic sequencing technology has
contributed greatly to the increasing literature on specific pathogenic variants
prevalent in different populations that cause USH (1,57–60). Identification of
these pathogenic variants is critical in understanding the molecular
mechanisms and functions of proteins encoded by these genes. This has led to
a more complete and deeper understanding of the visual and auditory
systems.
Although there is no cure for the hearing loss or vision loss due to USH,
there are many treatments currently being pursued, specifically for the
retinopathy associated with USH. Zinc finger nucleases are DNA binding
proteins that target and edit a region of the DNA by inducing a double-
stranded break. This technology was shown to induce gene repair
mechanisms in USH1C, at p.R31X, resulting in recovery of protein
expression (61). About 10% of pathogenic variants that cause USH lead to an
early stop codon. One approach to treat USH is to use pharmacologic
compounds to force the cell to translate beyond the stop codon, thereby
resulting in a full-length protein. This method was investigated by Goldmann
et al. in which a drug, PTC124, was used in cell culture and on mice to treat
the retinal USH phenotype. They found that PTC124 was able to read-
through the stop codon and restore protein function (62).
With over 2,500 clinical trials currently in process worldwide, gene
therapy has been a groundbreaking advancement in medicine and has shown
promise to treat human disease (63). Gene therapy has been used to treat the
retinopathy in animal models with USH1 or USH2 with some success. Mice
with USH do not develop vision loss, even though they lack the normal
protein associated with the defective gene in the retina. Thus, biochemical
rather than functional vision assays are needed. Hashimoto et al. used a
CMV-MYO7A chimeric promoter and found that in vivo abnormal
accumulation of opsin was rectified in the connecting cilium of the
photoreceptors, and melanosomes were localized to their normal location—
indicating a functional myosin7a protein through the use of lentiviral gene
replacement therapy (64). In 2011, another study used the most common gene
delivery system—the adeno-associated virus (AAV)–to transport mouse
whirlin cDNA driven by the human rhodopsin kinase promoter. This vector
was injected under the retina of whirlin deficient mice, and they concluded
that whirlin was successfully restored, and the protein complex whirlin forms
with usherin and GPR98 was produced for up to 6 months after the injection
(65).
There are a two clinical trials taking place, at the time of writing, which
use gene therapy in patients with retinitis pigmentosa due to USH1B: they
included the UshStat trial (NCT01505062) and the STELLAR trial
(NCT03780257). The UshStat trial is currently recruiting for a phase I/IIa
safety study of subretinal injection of UshStat—a recombinant EIAV-based
lentiviral vector expressing MYO7A that showed efficacy in the mouse
model of USH1b (66). The STELLAR trial is conducted by ProQR, a biotech
company based in the Netherlands, which seeks to develop therapies for rare
diseases. This Phase I/II trial will investigate the intravitreal injection of a
drug called QR-421a to treat retinitis pigmentosa in patients with USH2A
specifically due to variants in exon 13. This drug is designed to skip exon 13
during the synthesis of the protein resulting in a truncated yet functional
usherin protein. ProQR is also developing an additional therapy to target a
specific USH2A pathogenic variant, c.7595-2144A>G.
Few studies have been conducted using gene therapy for the treatment of
the hearing loss associated with USH. One study assessed the efficacy of an
AA-Clrn1-UTR vector in Clrn-1 deficient mice. They identified that one
injection of this vector preserved the hair bundle structure and hearing of
mice through their adult life (67). This same group also identified a small
molecule that reduces hearing loss in the mouse model of USH3 (68) and
published the first study to explore the use of pharmacologic agents to treat
hearing loss in a mouse model of USH.
CONCLUDING REMARKS
There has been considerable progress in the last decade in understanding the
clinical findings in Usher syndrome as well as discovery of the genetic bases
of Usher syndrome and their consequences at the molecular and cellular
levels in both vision and hearing. Cost-effective and accurate diagnostic tests
are available and currently implemented in clinical practice including specific
sequencing panels for suspected genetic hearing loss. Lastly, there are
multiple studies both in humans and animals underway using gene therapy as
a treatment for the retinal degeneration for all three types of USH that show
promise. We are enthusiastic about potentially new treatments for USH
patients and to further understanding the mechanisms behind this life-altering
disease.
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34
X-Linked Retinoschisis
Catherine A. Cukras, Laryssa A. Huryn, Michael T. Trese and Paul
A. Sieving
PREVALENCE
X-linked retinoschisis (XLRS) is a congenital vitreoretinal dystrophy seen in
males with an estimated prevalence of 1:5,000 to 1:25,000 (1). Affected
patients typically present at an early stage with vision loss due to macular
changes with characteristic cystic maculopathy (Figure 34-1).
FIGURE 34-1 Fundus photographs showing the typical
presentation of XLRS. A: Affected male with spoke–
wheel pattern of foveal cysts. B: Affected male with
visible inner leaf schisis margin with adjacent white
dendriform retinal vessels in the inferotemporal quadrant.
Patients also manifest a selective reduction of b-wave amplitude in the

scotopic (dark-adapted) electroretinogram (ERG) (Figure 34-2). The
peripheral retina is involved in about half of the affected males, with splitting
of the inner retina causing large elevations (Figures 34-1 and 34-3) leading
to (a) a substantial decrease in visual field sensitivity; (b) a predisposition to
hemorrhaging into the vitreous; and (c) the possibility of full-thickness retinal
detachments in severe cases. Many subjects manifest peripheral vitreous
“veils” and vascular sheathing (Figure 34-1B). The trait is passed genetically
to affected males by their mothers. XLRS female carriers cannot be identified
clinically. Their fundus examination is normal, and there are only a few
reports that show minimal ERG abnormalities (2,3). DNA diagnostic testing
is available and helpful to identify mutations in the RS1 gene and
consequently to identify women who carry the trait for XLRS.
FIGURE 34-2 Scotopic full-field ERG traces from an

affected XLRS male (45 years old), harboring a c354del1-
ins18 mutation of the retinoschisis (RS1) gene. XLRS
affected male shows typical ERG with b-wave reduction
but a-wave preservation in dark-adapted recordings,
compared with a normal individual. The arrow indicates
the characteristic “electronegative” waveform with b-wave
smaller than the a-wave.
FIGURE 34-3 Infrared images (left side) and OCT (right
side) scans from a boy (12 years old) with XLRS. Top
panel: Macular scan, showing foveal schisis and
parafoveal inner retina cavities. Lower panel: Vertical
scan showing the wide delamination of the retina. Images
were taken by Spectralis OCT (Heidelberg Engineering).
Environmental and genetic factors, including modifier genes, are predicted to
account for some of the phenotypic variation in an XLRS mouse model (4)
and, hence, are possible contributors to the wide phenotypic heterogeneity
found across the ages of members of XLRS families (5,6).
However, modifier genes have not been described for human XLRS.
Affected males lack a fully functional retinoschisin protein that normally
participates in holding retinal layers together and that contributes to the
integrity of the retinal architecture through its adhesive properties (7–9).
Hence, XLRS disease is believed to leave the retina vulnerable to mechanical
shock and injury. Contusive or acceleration–deceleration trauma can
exacerbate vision loss in XLRS patients. Prudent medical advice to XLRS
males includes avoiding excessive body contact sports and wearing sports
goggles for all contact sports if they do choose to participate. Environmental
factors are not currently suspected of contributing to the etiology or
progression of the condition.
GENETICS
The RS1 gene for XLRS was identified on the X chromosome by positional
cloning in 1997 (10). The RS1 gene contains 6 exons that encode a 224-
amino acid protein called retinoschisin. More than 150 mutations in RS1 have
been reported (11), and a comprehensive list of allelic variants (sequence
polymorphisms) is maintained through the Retinoschisis Consortium
database (www.dmd.nl/rs/). Most variants are missense mutations that are
clustered in the discoidin domain (exons 4 to 6) and lead to misfolded
protein. Founder effects are recognized, as 95% of XLRS patients with
Finnish ancestry have one of three mutations in the RS1 gene: p.E72K
(c.214G>A), p.G74V (c.221G>T), and p.G109R (c.325G>C) (12). Exons 1,
2, and 3 of the RS1 gene are susceptible to major deletions, splice site
mutations, and introduction of a stop codon, all of which would cause a
truncated or absolute loss of protein. We observed that less severe mutations
tend to occur in exons 4 to 6, and the more severe mutations involve exons 1
to 3 (5,13–15), the exception being alterations involving the addition or
subtraction of a cysteine residue, which is believed to confer a severe
mutation, as it would potentially disrupt a disulfide bond important for
tertiary folding that is predominantly found to occur in exons 1 to 3 (15).
XLRS is inherited in classical X-linked recessive manner and is not a
syndromic condition. Women carriers have a 50% chance of transmitting the
disease-causing mutation in each pregnancy. Males who harbor the mutation
are affected, and they transmit the mutation to all their daughters and none of
their sons. Females who inherit the mutation will be carriers and nearly
always are asymptomatic but can transmit the trait to their sons.
Sporadic cases of bilateral symmetrical maculopathy without known
family history of other affected males can be particularly challenging to the
clinician who is trying to establish a diagnosis and prognosis for the patient
and family. Electrophysiologic testing and supplemental molecular genetic
analysis are key diagnostic aids. Mutation analysis can identify mutations in
the RS1 gene in at least 90% of affected males (16). The value of
supplemental molecular genetic analysis is exemplified by the report of a
patient with the referring diagnosis of bull’s-eye maculopathy and selective
reduction in the b-wave of the ERG (17). Clinical findings suggested the
possibility of XLRS, and the patient was subsequently shown to have a novel
mutation (p.A100P) in the RS1 gene (17).
The methods to grade clinical severity of XLRS disease are not
standardized and vary considerably (2,18–23), which makes it difficult to
harmonize definitive genotype–phenotype correlations across different
reports. As the disease affects both central and peripheral function, the
utilization of combinations of visual acuity as a central functional measure
and full-field ERGs as a measure of whole-retina and synaptic function have
been utilized to grade severity. Despite these caveats, in general, disease
severity and clinical characteristics associated with missense mutations are
thought to present with relatively similar and with modest clinical severity
(19,23). Our recent reports (14,15,24) indicate that RS1 mutations that disrupt
the formation of retinoschisin protein and give a null-protein phenotype,
along with mutations that cause loss of a cysteine residue with consequent
anomalous protein folding, cause more severe disease with major reduction
of b-wave amplitudes and b/a ratios and with delayed a-wave implicit time.
By contrast, RS1 mutations that cause conservative amino acid substitutions
tend to be less severe. Many examples have been published of severe
phenotypes in families with RS1 deletions or splice site mutations that
produce premature truncated or no retinoschisin protein; severe disease
included retinal detachment and extensive macular lesions with low visual
acuity and subnormal or undetectable b-wave and reduced a-wave amplitudes
(1,15,22). By contrast, patients with conservative RS1 missense mutations
likely to alter protein structure minimally frequently had normal or
measurable ERG b-waves (18).
Even with these general patterns, gene-to-clinical correlation remains
difficult, as even family members with the same mutation or mutation type
can exhibit considerable variation (5,6,18,25,26).
Severe XLRS disease, including low vision, vitreous hemorrhage, and
varying extent of peripheral schisis, was reported within a Chinese family
with a conservative missense mutation (6), and three adult XLRS male
patients with a cysteine residue mutation (typically causing severe effects on
protein structure) had mild clinical disease and well-preserved ERGs (25).
Cysteine residue mutations disrupt disulphide bonds that are important in
stabilizing the protein configuration, generally with catastrophic
consequence. Phenotypic variability was found even between identical twin
boys presenting with nystagmus in early infancy and similar fundus
involvement limited to foveal schisis. Neither twin subsequently experienced
severe anatomical complications of the disease. The refraction of one twin at
age 2.3 years was +5.00 +2.50 ×105 (OD) and +6.00 +2.25 ×90 (OS).
Although his vision remained stable throughout his early childhood, he had
only 20/200 acuity when last tested at 22 years of age. His identical twin
brother was less hyperopic, measured at +3.75 OU, and always had less
severe nystagmus and has maintained approximately 20/40 acuity in both
eyes.
PATHOPHYSIOLOGY
Photoreceptors are an abundant primary site of retinoschisin synthesis (27).
Retinoschisin was also found in the inner retina, which suggests that the
protein is secreted by or transported to sites beyond the photoreceptors where
it is produced. Retinoschisin can be secreted by differentiated retinoblastoma
(Weri-Rb1) cells, implying that the protein is produced by photoreceptors and
then secreted (27). Antibody to retinoschisin binds to photoreceptors and
bipolar cells but not to Müller cells (28); this challenges the long-held
hypothesis that retinoschisin was a Müller cell defect. We observed that
retinoschisin is expressed by nearly all retinal neurons during development
and in the adult retina of mice (29).
The retinoschisin protein has a single discoidin domain that is thought to
help maintain cell adhesion to extracellular matrix proteins and mediate cell–
cell interactions (7,30). This may be particularly important in maintaining the
photoreceptor synapse with the bipolar cell (8).
Retinoschisin may also help regulate the fluid balance between
photoreceptor compartments (31). This hypothesis is based on the co-
localization of retinoschisin with the NaK-ATPase, a retinal cationic channel
(7,32).
Three mouse models of XLRS have been described that result in a
deficiency of endogenous murine retinoschisin (8,33,34). The scotopic ERG
of the male Rs1-KO mice mimics that of XLRS patients and has selective b-
wave reduction. One mouse model also shows an additional severe effect in
the cone ERG (8). The retina of the Rs1-KO mouse has a disorganized
architecture with areas of schisis in the inner nuclear layer, photoreceptor
loss, and shorter outer segments (8,33,35). These findings of the murine
XLRS retina support a functional role of retinoschisin in maintaining cell–
cell adhesion.
More recently (36), study of the synapse in the Rs1-KO mouse has
revealed abnormally low concentrations of intracellular Ca2+ in photoreceptor
terminals, indicating that the presynaptic function is disrupted by the absence
of Rs1 protein expression. The postsynaptic terminal also demonstrated
abnormalities in the Rs1-KO mouse with greatly diminished TRPM1 staining
at the dendritic tips with relatively normal localization of mGluR6 receptors.
Interestingly, months after AAV8-RS1 application, intracellular Ca2+
concentrations in photoreceptor presynaptic terminals were restored to near
wild type and TRPM1 receptors on postsynaptic dendritic tips also increased.
These studies revealed a role for RS1 in addition to its structural role of
maintaining cell -cell adhesion in the synapse of photoreceptor–bipolar cell
signaling. This synaptic role would be in line with the changes to the ERG b-
wave that is observed in both the human disease as well as the Rs1-KO mice.

Clinical findings in XLRS vary broadly and, especially in less overt cases,
and can mistakenly be diagnosed as “amblyopia” in young affected boys.
Family history indicating an X-linked inheritance of visual abnormality
should help in the differential diagnosis.
Affected males have reduced central visual acuity, which typically comes
to parental and medical attention during early school-age years when children
manifest difficulties in reading.
Visual acuity typically is in the range of 20/40 to 20/120 with an
increased prevalence of hyperopia. Fundus examination reveals bilateral
foveal schisis in virtually all patients at some point in their clinical history
and may be the only physical finding of the disease. The foveal schisis
appears in a spoke–wheel pattern with areas of retinal folds surrounding the
microcysts (Figures 34-1 and 34-3). The human fovea is incompletely
developed at birth and does not reach adult conformation until age 15 months
(37). Hence, one can anticipate that the “spoke–wheel” intraretinal parafoveal
cysts of XLRS may not be fully clinically apparent at birth.
The classic foveal schisis seen early in life may collapse and develop into
an atrophic stage of the disease (2). Hirose et al. noted that individuals with
XLRS over the age of 45 showed macular atrophy on optical coherence
tomography (OCT) and others have noted age-related foveal thinning (38,39).
To our knowledge, there is only one published report of longitudinal, 15-year
natural history data on a single patient demonstrating cyst evolution on OCT
(40).
Peripheral retinoschisis involves the inferotemporal region in
approximately half of patients, leading to reduced visual field sensitivity
(19,41). The schisis may progress toward the central retina or become flat
and leave only retinal pigmented epithelium (RPE) abnormalities or retinal
scars (Figure 34-1B). Peripheral chorioretinal scars may mimic resolved
chorioretinitis. Macular scars can also occur. Fewer than half of XLRS cases
have “vitreous veils” consisting of partial-thickness retinal layers
delaminating into the vitreous, along with overlying retinal vessels, which
can bleed and cause vitreous hemorrhage. The frequency of complications
such as vitreous hemorrhage has been reported to range from 3% to 21%,
while retinal detachment has ranged from 5% to 40% in the literature with
approximately 11% for both in our clinical practices (21,40–43).
Interestingly, there has been reported a case of a boy with recurrent vitreous
hemorrhage in both eyes with an RS1 mutation but without foveal schisis
(45).
Optic nerve pallor, dragged disc appearance, or optic nerve head
neovascularization (46,47) has been described. XLRS patients can have an
unusual whitish fundus appearance when the dark-adapted retina is exposed
to light, termed the Mizuo phenomenon (48,49), which is believed to result
from abnormal potassium processing in the retina. This phenomenon
disappears after vitrectomy, presumably as a result of removing the interface
between the retina and the vitreous (50). Other unusual presentations of
XLRS include a macular hole (51,52), pseudomacular hole (5), strabismus
(53), macular dragging and pigmentary changes (54), macular multiple fine
white dots (55), bilateral macular detachment (56), vitreous hemorrhage (57),
foveal ectopia (58), and retinal white flecks (59) that may simulate fundus
albipunctatus (60).
DIAGNOSTIC STUDIES
Visual acuity is nearly always subnormal and can reach 20/200 or less in
some patients. Lesch et al. (2) described no substantial differences of mean
best-corrected visual acuity of “cystic” (<25 years old) versus “atrophic”
(>25 years old) XLRS groups, in agreement with previous reports (21,22).
However, a trend of decreased visual acuity with increasing age has been
reported (40). Peripheral visual field is affected by peripheral schisis, which
can produce absolute scotomas.
Color vision is generally unaffected, and patients do not complain of
night blindness.
OCT has revealed schisis lesions that frequently lie deeper in the retina
than previously thought, often in the midretinal layers (61) (Figure 34-3).
Areas appearing clinically intact can show shallow schisis across the macula,
termed flat or lamellar schisis (20) found in the macular area between the
arcades and associated with an abnormal ERG. Gerth et al. (62) showed using
OCT images, evidence of extrafoveal schisis occurring within the outer and
inner nuclear and ganglion cell layers, and evidence of photoreceptor outer
and inner segment layer disruption. Also Gregori et al. (63) identified cystoid
spaces accounting for retinal splitting in the inner nuclear layer and outer
plexiform layer. Small cysts, separate from those involved in foveal splitting,
can be found in the outer nuclear layer and in the ganglion cell layer and/or
nerve fiber layer using OCT.
We find that OCT changes may be associated with surgical outcomes. In
one case of a patient who had bilateral surgical intervention, one eye
recovered good of visual function in the eye which macular schisis resolved
on OCT. However, in the fellow eye that failed to recover visual function, the
macular schisis remained. Gupta et al. report a case of foveal schisis
resolution after vitrectomy for retinal detachment involving the macula (64).
The OCT combined with clinical appearance of the retina on funduscopy
has been used as one classification system for XLRS (Table 34-1). In a report
by Prenner et al. (20), patients were classified into four types based on the
presence of foveal cysts only (type 1), foveal cysts by clinical examination
plus lamellar macular schisis detected by OCT (type 2), findings of type 2
XLRS plus peripheral schisis (type 3), or foveal cysts and peripheral schisis
(type 4) (20). Lesch et al. used OCT in their large cohort of XLRS subjects to
understand the progression of disease across the lifespan and the effect of age
on the phenotype and genotype in XLRS (2). The authors found correlations
between younger age and thicker foveas and between younger age and larger
b-wave amplitudes. Based on these findings, the authors proposed the
definition of a (a) cystic form of XLRS found before 26 years of age or (b) an
atrophic form of XLRS found 26 years of age or older. In the younger group,
there were OCT findings of (a) increased foveomacular thickness due to
intraretinal cysts, and in the older group of patients, there was (b) central
thinning from macular atrophy. Our data have also corroborated this finding
of older individuals having age-related thinning with foveal atrophy being the
predominant finding in patients over 45 years of age (40). Interestingly, we
did not find a statistically significant correlation between OCT thinning and
visual acuity. We have proposed that functional worsening of ERG responses
with age by cross-sectional study of families with XLRS suggests the
progressive nature of XLRS disease (5). However, longitudinal study of
patients over a mean time of 6.8 years demonstrated no statistically
significant change in the ERG b/a ratio within eyes over this length of time,
indicating that potential progressive changes are slow moving (40).
TABLE 34-1 Classification system for XLRS

XLRS, X-linked retinoschisis; OCT, optical coherence tomography (20).
Young patients with XLRS usually have a normal retinal fluorescein

angiogram, but older patients may have macular window defects from RPE
changes. Peripheral vascular changes can be noted with angiography, with
areas of peripheral retinal nonperfusion and leakage of dye from abnormal
retinal capillaries (65) and leakage into vitreous veils (46). Wide-field
imaging with color photographs and fluorescein angiography using a wide-
field cameras has enabled more detailed evaluation of both “exudative” and
“nonexudative” lesions, defined by the presence or absence of lipid (66). In
performing a fluorescein angiogram with a wide-field camera, the presence of
increased vascular permeability can be appreciated as a potential source for
the fluid and exudate accumulation and give insight into the etiology of this
finding (66).
The ERG is a diagnostically useful test for XLRS. The dark-adapted ERG
shows reduced b-wave amplitude, while the a-wave frequently remains
normal (15,44). This ERG configuration is termed an electronegative
response, because the positive-going b-wave fails to return to above baseline
from the negative-going a-wave (Figure 34-2). Sixty percent of XLRS eyes
had the “electronegative waveform” in a cohort of 68 affected men (15). As
the b-wave originates from activity of depolarizing bipolar cells that lie
postsynaptic to the rods (67–69), this implies primary involvement of the
inner retina in XLRS.
Although an electronegative ERG response is not pathognomonic
exclusively of XLRS, only a few other retinal dystrophies result in this
waveform configuration, and finding this ERG type in a male with retinal
schisis is clinically confirmatory of XLRS. A nonelectronegative ERG does
not, however, rule out the possibility of XLRS, because patients with a
confirmed mutation in the RS1 gene may present with a near-normal scotopic
ERG b-wave (15,16,70) even in the presence of typical fundus findings of
XLRS (5). Although there is a large spectrum of the degree to which the b-
wave is reduced relative to the a-wave across patients, the responses between
the two eyes of a patient with XLRS are highly correlated with similar
reductions in their b-wave relative to a-wave (40).
Although some patients with RS1 missense mutations have reduced a-
wave amplitudes, the a-wave amplitude in XLRS generally remains normal,
which has raised the question whether photoreceptors are affected in XLRS
(15), as retinoschisin protein is heavily expressed in photoreceptor inner
segments and in the photoreceptor–bipolar cell synapse (28,71). Carriers of
XLRS cannot be identified by electrophysiologic analysis, because the a- and
b-waves of the ERG are normal. Final thresholds of the dark adaptation curve
and the phases of cone and rod adaptation occur at the appropriate time. The
Arden ratio of the electrooculogram is normal (44).
Alterations in the multifocal ERG (mfERG) have been found with both
the XLRS mouse model and in affected men (72,73). Although the relative
sensitivity is unknown compared to the full-field ERG, mfERG may be
useful to study the severity of macular involvement and to identify some
carriers of XLRS. Kim et al. (3) reported abnormal mfERG in two of nine
female obligate carriers of the disease with normal-appearing fundus
examinations.
The differential diagnosis of the electronegative ERG includes congenital
stationary night blindness, Batten disease, BRAT1 mutations, cancer-
associated retinopathy, and vascular occlusions among others. Careful
history, age of onset, and especially molecular genetic testing are useful to
differentiate from other causes of reduced b-wave on the ERG.
Isolated cases of XLRS offer no opportunity to examine other family
members and not infrequently are mislabeled as some disorder that may
mimic retinoschisis (Table 34-2). Amblyopia, retinal dialysis, congenital
infection, and retinitis pigmentosa (RP) as the initial referring diagnoses were
later diagnosed as XLRS in a cohort of 56 males in 16 British families (42).
An isolated case of a male XLRS does not imply a new mutation, because the
female carrier mother essentially never exhibits clinical signs of the trait and
has normal vision. De novo mutations have been reported for XLRS (74), but
such instances are exceedingly uncommon.
TABLE 34-2 Differential diagnosis of retinoschisis

aKnown also as Goldmann-Favre syndrome.
ERG, electroretinogram; OCT, optical coherence tomography; RP, retinitis pigmentosa.
The maculopathy in cases of rod–cone dystrophy or Stargardt disease can

mimic XLRS, particularly because the dark choroid sign is not always
present in Stargardt disease or in the autosomal dominant form from ELOVL4
gene mutations (75). The identification of X-linked segregating pattern of
inheritance differentiates XLRS from these conditions. Additionally,
Stargardt disease and rod–cone dystrophy do not exhibit selective reduction
in the ERG b-wave amplitude.
Enhanced S-cone (Goldmann-Favre) syndrome is an autosomal recessive
vitreoretinopathy with mutations in the photoreceptor-specific nuclear
receptor, NR2E3, that causes poor vision in early infancy. These patients can
also have increased sensitivity to blue light. Affected individuals may present
with foveal retinoschisis and peripheral lattice degeneration that could be
confused with XLRS. There are no vitreous veils as seen in XLRS. Unlike
XLRS, Goldmann-Favre patients complain of night blindness and have a
quite different ERG with markedly reduced a- and b-waves (76).
Wagner vitreoretinal dystrophy is an autosomal dominant disease
characterized by myopia, vitreous syneresis, and pigmentary clumping in the
macula (77). Unlike retinoschisis, cataracts commonly develop during the
teenage years in Wagner dystrophy and progress significantly by the fourth
decade. Retinal detachment is a frequent complication and might be confused
with XLRS. Although the ERG may be abnormal in patients with Wagner
disease, selective b-wave reduction is not seen.
Another possible misdiagnosis is acquired retinoschisis, which affects
both males and females, is usually bilateral, and can be observed in
asymptomatic patients after age 50 on routine ophthalmic examination.
Degenerative retinoschisis generally is limited to the middle retinal layers and
is geographically limited, whereas XLRS involves the full thickness of the
retina and occurs across a broad extent of it (62).
The severe presentation of XLRS with early-onset retinal detachment
may be confused with retinoblastoma suspected for the presence of
leukocoria, X-linked Norrie syndrome, in which bilateral retinal detachment
may be present at birth or infancy (but it can also manifest with mental
retardation and deafness), and autosomal dominant Stickler syndrome, which
has retinal detachment in the first decade of life, reduced ERG, chorioretinal
atrophy, perivascular pigmentary degeneration, and possibly joint, hearing,
and facial abnormalities.
In later age when XLRS patients progress to the “atrophic” stage of the
disease, confusion may come from a misdiagnosis of age-related macular
degeneration, which rarely has peripheral retinoschisis, vascular sheathing, or
extensive peripheral pigment epithelial changes that are found in juvenile
retinoschisis.
MEDICAL MANAGEMENT
No proven treatment is currently available for XLRS; however, there are
reports of the utilization of carbonic anhydrase inhibitors (CAIs) to decrease
macular cysts. The proposed therapeutic mechanism of CAI use in XLRS is
thought to involve an increase in RPE cells’ ability to transport fluid from the
apical side through the basement membrane with subsequent transport into
the choroid. Apushkin et al. were the first to report on the effect of topical
CAIs (Dorzolamide 2%) on cystic macular cavities in XLRS. They described
a reduction in macular thickness as measured with OCT as well as
improvement in visual acuity, defined as 7 or more letter increase in BCVA
on ETDRS in 5 of 8 patients within 2 months of medication administration
(78). Since then, multiple case reports have been published utilizing both
topical and oral CAIs with variable degrees of structural change—ranging
from decrease in cysts to worsening of cysts—and variable, but most often
little, effect on visual acuity (79–85). Abalem et al. noted significant variation
of central foveal thickness throughout a day of measurements, proposing this
as an explanation for inconsistent responses to CAIs (86). In serial
observations of seven XLRS patients, we observed fluctuations of macular
schisis cavities and OCT thickness over weeks to months without any
intervention. Pennesi et al. reported on a prospective evaluation of 51 XLRS
patients treated with topical and/or oral CAIs (87). They noted variable
changes in retinal cyst cavity volume over time but no significant change in
visual acuity. The topical treatment is well tolerated for the most part and has
a favorable safety profile; however, larger studies with longer duration are
needed to evaluate efficacy. Long-term safety of systemic CAIs is unknown.
None of these CAI approaches are FDA approved for XLRS treatment.
Management of Amblyopia and Strabismus in X-

Linked Retinoschisis
George et al. (42) noted a bimodal age presentation of XLRS in childhood.
The youngest presented with strabismus and nystagmus at <2 years of age,
and others presented with reduced acuity at 5 to 7 years of age when they
entered school. As a consequence, the ophthalmic care of a child with XLRS
should carefully evaluate and treat refractive errors, strabismus, and
amblyopia.
Roesch et al. (21) reported that almost 20% of children with XLRS had
significant vision loss in childhood and that half of this vision loss in
childhood was due to complications of the disorder: retinal detachment,
vitreous hemorrhage, and neovascular glaucoma. Cataract was found in 11
eyes of the children and adolescents, representing 6% of the entire cohort,
and was associated with severe retinopathy.
At the National Eye Institute, a review of 120 patients evaluated with
XLRS revealed that 23% of patients had a >15 letter difference in visual
acuity between their two eyes with 43% of this subset demonstrating
evidence of strabismus (40). The proportion of cases associated with
strabismus was estimated as 30% of the studied cohort in studies by George
et al. and Roesch et al. (21,42). Esotropia and exotropia occurred in the same
proportion, although poor vision (<20/200) and severe schisis were highest in
the group with exotropia. Hyperopia is reported more frequently (21,88). Any
structural or functional disparity between eyes has the potential to lead to
amblyopia and the contribution of developmental visual loss is impossible to
separate completely from any biologic disparity. Our surveillance of these
patients throughout the period of visual development in childhood should be
frequent addressing the correction of refractive errors and strabismus
management as an early intervention for amblyopia. Chapter 11 deals with
amblyopia in more detail.
SURGICAL TREATMENT OF RETINAL

DETACHMENT WITH CONGENITAL
RETINOSCHISIS
Rhegmatogenous retinal detachment in XLRS can be challenging. Usually,
the rhegmatogenous component involves an outer- and inner-wall hole in an
area of peripheral schisis. The inner-wall holes are often easy to find and can
be very large. By themselves, these inner-wall holes do not lead to full-
thickness retinal detachment and do not require treatment. The search for
outer-wall holes can be more difficult, and these holes must be closed to
successfully repair the retinal detachment. When the outer-wall hole is not
obvious, the most common site is at the posterior border of the peripheral
schisis cavity and the normal retina. These eyes have often been repaired with
a scleral buckle encompassing the outer- and often inner-wall holes. The
advantage of a scleral buckle is that it is less invasive than vitrectomy. In
addition, a disadvantage of vitrectomy in infant eyes with XLRS is that the
hyaloid cannot be easily peeled from the retina. It seems that peeling of the
posterior vitreous may be damaging, particularly in type 2 and 3 eyes, which
have shallow, flat macular schisis that may not be clinically apparent. It is the
authors’ experience that type 2 and 3 XLRS are clinically more complex and
difficult to handle. Enzymatic degradation of the vitreoretinal junction with
ocriplasmin may give a less traumatic separation and allow more complete
relief of vitreoretinal traction. However, concerns of ocriplasmin on retinal
function exist (see Chapter 4), and more study of this is needed. If buckling
fails, we perform a vitrectomy, because it allows us to treat the outer-wall
break with laser after internal drainage and flattening the retina. Care is taken
to avoid removing the inner wall of the schisis cavity. At this time, we have
no way to reconnect the anterior to the posterior leaflet of retina after they
split. Currently, we divide eyes with a need of vitrectomy into several
categories. First, eyes are divided by the composition of the inner-wall schisis
cavity: a schisis cavity that has a largely intact inner wall or those in which
the inner wall is largely thin radial bands often containing vessels and very
large inner-wall holes. This latter type seems to rarely lead to retinal
detachment. Second are eyes in which the schisis cavity overhangs or rises
from below to cover the macula. This physical finding should be
accompanied by a noticeable change in vision. Third is the combined schisis–
retinal detachment; depending on the amount of schisis fluid and the size of
retinal holes, these may be treated with scleral buckles as described above or
treated by vitrectomy techniques.
Vitrectomy Techniques for X-Linked Retinoschisis

Our current vitrectomy technique is to use ocriplasmin injection if possible in
older children 1 week before surgery. Then, a 2-port vitrectomy is performed
with removal of the posterior hyaloid, followed by flattening the schisis
cavity by internal drainage with a small 39-gauge cannula through a small
inner-wall retinotomy. If no retinal detachment is present, the schisis cavity
fluid is not viscous and can be easily drained through a small cannula. This is
generally done under air. An 80% silicone oil exchange is then performed
with the intent of leaving the silicone oil in the eye permanently. In eyes with
combined schisis–retinal detachment, the fluid of the schisis–retinal
detachment cavity is more viscous and requires a larger bore cannula. In
these eyes, silicone oil is also used and laser treatment for outer-wall breaks
is performed. It is hopeful that in the future by collapsing schisis cavities and
using a synthetic retinoschisin, we may be able to reconnect the inner and
outer leaflets of the schisis cavity.
Tractional retinal detachment can also occur in XLRS, in which the
retinal detachment occurs posterior to the peripheral schisis cavity and
involves the fovea with a shallow macular detachment. This can be difficult
to determine clinically, but it may be detectable by OCT testing. The visual
acuity in these eyes can improve after vitrectomy in which the hyaloid is
carefully attempted to be removed from the macular area. In one report,
visual acuity improved from 20/200 to 20/50 (61). These eyes are best
operated in type 4 XLRS, but if the child has type 2 or type 3 XLRS and
requires vitrectomy, no longer do we perform large inner-wall retinectomies.
In eyes with type 2 or 3 XLRS that have undergone retinectomy,
proliferation can form along the anterior surface of the anterior leaflet of the
schisis. As this tissue contracts, the anterior schisis leaflet can scroll up,
further splitting the retina. Although both gas and silicone oil have been used
as a postoperative tamponade, we tend to use silicone oil as described above.
This also has the advantage of supporting the retina in younger children who
cannot position well. We use an 80% fill and have the child position
facedown at night and briefly during the day to maintain lens clarity. By
using this technique, we have had lenses remain clear for over a decade.
Other therapeutic options for XLRS may conceivably be possible in the
future to prevent progression in XLRS, including (a) reattachment of the
inner and outer leaflet with a combination of mechanical techniques and
biologic interventions and (b) enzymatic vitreolysis to eliminate foveal
traction (89).
VISION REHABILITATION AND LOW-

VISION AIDS
Males affected with retinoschisis may benefit from visual aids. Acuity can
fluctuate to a modest extent from XLRS, but acuity typically remains
between 20/60 and 20/120 during teenage and middle-aged years. Such levels
of vision are amenable to help with low-vision aids, including handheld
spectacle magnifiers and telescopes. These aids can be provided by low-
vision specialists. The primary visual deficit occurs in central vision,
although peripheral vision fields can be impaired to some degree but typically
far less than from progressive retinal dystrophies, such as RP. XLRS
normally impairs dark adaptation to a minimal degree and typically does not
cause significant night blindness nor does it retard the speed of adapting to
darkness. However, on general principles, visual function of individuals
affected by reduced acuity becomes more difficult in dim lighting.
Fortunately, there are no known systemic associations with XLRS. Primary
care providers should be kept informed of the patient’s vision and the
medications they are taking. Referral to a medical geneticist and/or genetic
counselor to discuss recurrence risk is necessary.
Ethical Considerations and Genetic Counseling

Genetic counseling is advised for families with XLRS In keeping with the
nature of an X-linked recessive trait, this condition only affects males.
Female carriers cannot be identified by clinical examination, as they
experience no visual effects, either on retinal structure or visual function, and
they are identified only by pedigree analysis or molecular testing of the RS1
gene. Directed counseling is advised for females who have a brother with
XLRS, because these women have a 50% chance of having inherited the RS1
gene carrier state from their mother, and if they did inherit the affected RS1
gene, they would then transmit the condition to any male offspring with 50%
risk. The RS1 gene has been identified, and molecular genetic diagnosis is
available to determine whether or not these women are carriers. Ethical
questions are raised by molecular diagnostic testing of young females who
are at risk for the carrier state but who have not yet reached the age of
consent. Although such testing may be useful for planning by other family
relatives, the information would not directly affect medical care of the girl,
which should be explained to the family members. Each request for carrier
testing should be considered on its own merit. If the result does not affect the
child’s reproductive choice, then it is recommended that testing be deferred
until the child is at an age of consent and is able to understand the issues (90).
FUTURE TREATMENTS BASED ON

RESEARCH
Currently, there are no proven treatments for XLRS. Various experimental
treatment strategies have been considered, including the use of CAIs,
dorzolamide and acetazolamide, as described above and more recently, gene
transfer.
The retinoschisin protein acts biologically as a cell adhesion factor
through properties of the discoidin domain as well as having a role in
presynaptic and postsynaptic organization of the photoreceptor and bipolar
cell synapse. In the absence of this protein, or from a functional deficit due to
misfolding of the protein, or from defective secretion to the extracellular
matrix (91), retinal structural integrity is impaired and subject to lamellar
separation or even retinal detachment.
Therapeutic intervention theoretically may be possible by restoring the
protein function to stabilize the retina. To this end, gene therapy studies
recently have shown that delivery of the RS1 gene to adult XLRS mice can
restore the ERG b-wave and rescue physiologic function (33). RS1 gene
replacement therapy improved both retinal ERG function and morphology in
affected mice (Figure 34-4), and the rate and extent of degeneration were
less severe in the treated eye compared with the untreated eye of the same
animal (92). RS1 gene therapy also reduces the retinoschisis intraretinal
cavities on OCT examination, indicating that structural rescue can occur in
young mice within weeks after providing the normal gene (unpublished data
from our laboratory).
FIGURE 34-4 Successful treatment of Rs1-knockout

mouse retina using an AAV (adeno-associated virus)-Rs1
gene vector with human Rs1 promoter. Evaluation was 12
weeks after intravitreal injection in one eye at age 31 days,
with fellow eye not treated. OCT shows successful closure
of intraretinal schisis cavities and functional improvement
of the ERG response with recovery of the b-wave This is
very similar to but not the identical presentation as in
Preclinical Dose-Escalation Study of Intravitreal AAV-
RS1 Gene Therapy in a Mouse Model of X-linked
Retinoschisis: Dose-Dependent Expression and Improved
Retinal Structure and Function. (Bush RA, Zeng Y, Colosi
P, et al. Preclinical dose-escalation study of intravitreal
AAV-RS1 gene therapy in a mouse model of X-linked
retinoschisis: dose-dependent expression and improved
retinal structure and function. Hum Gene Ther
2016;27(5):376–389. doi: 10.1089/hum.2015.142.)
At least two human XLRS gene therapy trials have been mounted. One
recently published 18-month results evaluated the safety and tolerability of an
RS1 adeno-associated virus (AAV8) in a dose escalation study
(ClinicalTrials.Gov NCT02317887) (93). Nine participants were assigned to
three dosage groups (1e9 vector genomes (vg)/eye, 1e10 vg/eye and 1e11
vg/eye) administered by intravitreal injection. Intravitreal delivery of a viral
vector offers advantages for the potential to reach a larger area of the retina,
and this route of administration would be particularly suited to XLRS disease
in which there is structural change to the retina. The RS1-AAV8 vector was
well tolerated in all but one individual who developed significant intraocular
inflammation leading to subsequent tractional sequelae. Systemic antibodies
to AAV8 were observed in a dose-related manner, but no systemic antibodies
were observed to the RS1 protein. One subject showed collapse of the
macular cystic cavities after receiving the higher dose of 3e11 vg/eye, which
provided a signal of possible efficacy, and this study is continuing as of this
time.
In the future, gene delivery may afford an opportunity to use protein
replacement as a treatment strategy to stabilize the retina of affected males
with XLRS. However, the lack of evidence of functional improvement for
any participant in this first reported clinical trial means that further studies are
needed. Both the window of opportunity (94) as well as the appropriate dose
and immunosuppressive regimen will have to be considered in future trials.
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35
Norrie Disease
Kimberly A. Drenser
PREVALENCE
The incidence and prevalence of Norrie disease are not known. Norrie disease
is a rare X-linked recessive disorder with clinical manifestations extensively
characterized by Warburg (1–6). The first reports were in Denmark, and since
then, it has been reported across various ethnicities. Norrie disease represents
the most severe phenotype in the spectrum of Wnt-associated
vitreoretinopathies, in which the ligand (norrin) is a functional knock-out.
More minor mutations affecting norrin lead to suboptimal binding to the
receptor complex and result in X-linked familial exudative vitreoretinopathy
(FEVR) (7). There is a complex relationship among FEVR, Norrie,
retinopathy of prematurity, and Coats disease that is still not fully understood.
Although there are several different genetic and environmental factors that
influence this range of phenotypes, this chapter will primarily consider Norrie
disease.
None known.
GENETICS
Ocular manifestations are severe and often the presenting finding at the time
of diagnosis. Additionally, approximately one-third of patients develop
hearing loss and two-thirds mental retardation. Boys with Norrie disease are
typically blind, lacking light perception (LP) from birth or shortly thereafter,
most often by 3 months of age secondary to severely dysplastic retinae.
Findings in infancy include leukocoria, iris atrophy, retrolental fibroplasia,
vitreous hemorrhage, dysplastic retinae with gray or grayish-yellow
pseudoglioma appearances, retinal folds, and retinal detachments that are
often hemorrhagic. These eyes subsequently develop cataracts and opaque
corneas and become phthisical within the first decade of life. Historically, no
treatment has been offered other than management of a blind, painful eye.
Norrie disease is associated with mutations in the NDP gene, which is
located on the short arm of chromosome X at position 11.4. The gene
product, norrin, is a small secreted protein with a cysteine-knot motif (7–10).
Norrin is a member of the mucin-like subgroup of 10-membered cysteine-
knot proteins. The cysteine-knot motif is highly conserved in many growth
factors, including transforming growth factor-β, human chorionic
gonadotropin, nerve growth factor, and platelet-derived growth factor. Norrin
has two primary domains, a signal peptide that directs localization of the
molecule and a cysteine-knot, which provides the structural conformation
required for receptor binding and activation of signal transduction. Norrin
acts as a nontraditional ligand in a Wnt receptor: β-catenin signal
transduction pathway, which plays a regulatory role in retinal development
and regression of hyaloid vessels in the eye (8,10,11). Frizzled receptors are
coupled to the β-catenin canonical and noncanonical signaling pathway,
which results in activation of Wnt target genes. A Fzd4 knockout mouse
demonstrates and highlights the importance of this pathway in vasculogenesis
and normal retinal development. Lack of Fzd4 results in abnormal vessel
growth with anomalous capillary maturation and increased areas of avascular
retina. A mouse model lacking norrin expression results in impaired retinal
capillary beds, specifically the intermediate and deep plexi (8,11). Computer
modeling of norrin supports the importance of the cysteine residues and their
disulfide bonds in the structural conformation of norrin and, presumably, in
norrin function (13). Correlations between genotype and phenotype have
confirmed that interfering with the cysteine knot-motif significantly
compromises norrin’s ability to bind its receptor and activate the signal
transduction pathway (7). Mutations not affecting cysteine residues may alter
protein folding and compromise pathway activation to varying degrees, but
their effect on norrin structure and function is not as clear and appears to
result in partial pathway activation, resulting in a less pathologic heritable
phenotype as seen in FEVR. Untranslated regions within NDP have
regulatory functions that control the expression and stability of norrin.
Mutations in these regions have also been associated with Norrie disease
(13–15).
The Norrie disease protein (NDP), originally named EVR2, was
described over a decade ago, giving way to the classification of a number of
congenital retinopathies as “NDP-related.” These include persistent fetal
vasculature syndrome (PFVS), retinopathy of prematurity (ROP), Coats
disease, and X-linked FEVR (13,16,17). These vitreoretinopathies, however,
may involve mutations in genes other than NDP, such as FZD4, LRP5,
TSPAN12, ZNF408, and ATOH7 (18). Severe presentations of these diseases,
as well as stage 5 ROP, can be indistinguishable from Norrie disease by
examination alone (Figure 35.1) making a differential diagnosis with careful
attention to excluding other causes important.
FIGURE 35.1 The retrolental fibrosis and contraction of

the primary vitreous may look similar to stage 5 ROP.
WORLDWIDE IMPACT
Unknown.
PATHOPHYSIOLOGY
The norrin protein is a ligand specific for development and maintenance of
the retina, inner ear, and parts of the CNS. Wnt 7 may also act in the ear and
CNS to activate the Fzd4 and Lrp5 receptors, but it does not exist in the
retina, which may explain why mutations that eliminate norrin function have
an absolute impact on the eye and more variable impact on hearing and
cognition (19).

Norrie disease is an X-linked recessive neurodevelopmental disorder
characterized by incomplete retinal vascularization and incomplete regression
of primary hyaloid structures. Diagnosis is based on birth history, family
history, and clinical examination, supported by genetic mutations in the NDP
gene. Norrie disease typically presents at birth or shortly thereafter with
leukocoria and visual loss and is usually bilateral and symmetric, with 30% to
50% of patients also manifesting sensorineural deafness and CNS
disturbances (1,4). Ocular findings include retinal detachment or fold,
persistent fetal vasculature, vitreous hemorrhage, iris atrophy, and corneal
opacities (16). The eyes are of average size (corneal diameter 9 to 10 mm)
initially but will become smaller with time as the traction tethering the
posterior lens capsule and posterior retina limits eye growth. Incomplete
maturity of the iris vessels is common with elongation of the ciliary processes
due to traction (Figure 35.2). There is a stalk (unregressed hyaloid
vasculature) that attaches to the posterior aspect of the lens with variable-
sized footplates. The stalk attaches to a sphere of dysplastic retina with an
exudative appearance and a yellow color. Unbranched retinal vessels can be
seen coursing through the tissue. Continuing toward the periphery, there is a
variable area of avascular attached retina with underlying areas of pigment
change (Figure 35.3). Untreated, this process will continue to contract
resulting in a retrolental plaque with pseudoglioma and eventual phthisis.
FIGURE 35.2 The iris may demonstrate anomalous
vasculature with poor dilation and varying degrees of
posterior capsular plaque (A). The posterior pole of the
same eye shows elongation of the ciliary processes and
persistent hyaloid structures (stalk) demonstrating the
anteroposterior changes affecting the eye (B).
FIGURE 35.3 The retinal vasculature shows primitive
development with a small area of vascularization,
subretinal exudate, and retinal pigment changes under the
peripheral avascular retina (A). Fluorescein angiography
clearly shows the abnormal pattern of retinal vasculature
(B).
DIAGNOSTIC STUDIES
Norrie disease remains a clinical diagnosis based initially on eye findings. An
examination under anesthesia is often required to evaluate the retina for
bilateral stalks with detached and poorly developed retina in the setting of an
infant with poor vision and abnormal red reflex. A differential diagnosis
should include ROP, FEVR, and PFV. Genetic testing using a panel, which
includes the NDP gene is key. Hearing tests, monitored over the lifespan of
the child, and neurodevelopmental evaluation are recommended. Family
history may be helpful, as affected male family members generally have
severe vision loss documented at or near birth. Abnormal hearing and/or
cognitive issues are seen in both Norrie disease and extreme prematurity.
Birth history and clinical course will help distinguish between these two
entities. Retinoblastoma must also be ruled out. Often this can be done easily
by clinical examination alone, but occasionally computed tomographic
scanning, brain and orbit magnetic resonance imaging, and/or ultrasound are
necessary when leukocoria is the presenting sign.
MANAGEMENT
Historically only supportive measures have been available for patients.
Warburg’s extensive study of Norrie disease only revealed rare cases with
vision beyond infancy. Out of 24 patients in her series, all had no LP except
for one boy who could count fingers until age 12 when LP was lost and
another boy who could perceive light (4). In Warburg’s review of the
literature (1966), she identified an additional 106 patients with Norrie
disease. Of these, only six patients were noted to have LP or pupillary
reactivity after 3 months of age. Jacklin et al. (20) reported on a patient who
underwent lensectomy and vitrectomy in one eye and found that the operated
eye avoided phthisis bulbi at 2 years of age unlike the unoperated eye. With
early surgical intervention, we found that 70% of patients maintained at least
LP vision in one eye after vitrectomy. Additionally, only 8% of eyes became
phthisical (21). Therefore, we believe that surgical intervention at the earliest
time point may be beneficial for these patients.
We have found in all of our Norrie disease patients that there is residual
stalk tissue (hyaloidal vessel remnant) connecting the posterior lens to the
dysplastic retina. Transection of this stalk tissue releases significant traction
on the retina. It is our belief that this release of the anteroposterior traction on
the retina is likely to, at least in part, explain the benefit of vitrectomy for
Norrie disease. Release of this traction not only allows the retina to settle
posteriorly but also eliminates an anteroposterior tether that likely restricts
normal ocular development and progressive damage to the lens and ciliary
body in addition to worsening the tractional retinal detachment. In cases in
which there is a dense retrolental plaque with total retinal detachment,
meticulous peeling away of this tissue from the retinal surface and ciliary
processes not only affords the opportunity for the retina to gradually reattach
but also decreases the likelihood of hypotony secondary to ciliary body
traction. Given the natural history of the disease (loss of LP by 3 months of
age in the vast majority of cases), we perform vitrectomy as early as possible.
If there is significant lens opacity obscuring our view of the fundus or
significant retrolental fibroplasia intimately apposed to the lens, then we
often remove the lens in addition to performing a vitrectomy (21).
Interestingly, despite the presence of large areas of avascular retina
(which we have not treated with laser photocoagulation, cryotherapy, or anti-
VEGF agents), we have not noted any neovascularization in our patients.
Perhaps the avascular retina is so dysgenic that it does not mount a significant
up-regulation in vascular endothelial growth factor or other angiogenic
factors.
When patients develop phthisis, enucleation is reasonable if the eye
becomes painful.
Patients with Norrie disease may have some ambulatory vision, although
most retain LP only after surgery. Working with a low vision specialist is
recommended. Chapter 14 discusses low vision services.
Roles of other physicians and health care providersNorrie disease is a
neurodevelopmental syndrome, and as such requires multiple specialists to be
involved in the care of the child. A pediatric neurologist will be the most
important caregiver. Patients will also need counseling from a geneticist and
will require repeated hearing tests.
REFERENCES
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of pediatric patients expressing mutations in the Norrie disease gene. Arch 8. Ophthalmol
2007;125(2):225–230.
8. Xu Q, Wang Y, Dabdoub A, et al. Vascular development in the retina and inner ear: control by
Norrin and Frizzled-4, a high-affinity ligand-receptor pair. Cell 2004;116:883–895.
9. Black G, Redmond RM. The molecular biology of Norrie disease. Eye 1994;8:491–496.
10. Berger W. Molecular dissection of Norrie disease. Acta Anat 1998;16:95–100.
11. Robitaille J, MacDonald ML, Kaykas A, et al. Mutant frizzled-4 disrupts retinal angiogenesis in
familial exudative vitreoretinopathy. Nat Genet 2002;32:326–330.
12. Ohlmann AV, Adamek E, Ohlmann A, Lutjen-Drecoll E. Norrie gene product is necessary for
regression of hyaloid vessels. Invest Ophthalmol Vis Sci 2004;45:2384–2390.
13. NDP Related Retinopathies [database online]. Bethesda, MD: National Institute of Health;
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expression. Gene 1999;227:181–188.
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sequences. Genome Res 2000;10: 1807–1816.
16. Drenser KA, Fecko A, Dailey W, Trese MT. Characteristic phenotypic retinal appearance in
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21. Schulman J, Jampol LM, Schwartz H. Peripheral proliferative vitreoretinopathy in a full-term
infant. Am J Ophthalmol 1980;90:509–514.
17. Kondo H. Complex genetics of familial exudative vitreoretinopathy and related pediatric retinal
detachments. Taiwan J Ophthalmol 2015;5(2):56–62.
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in blood-brain barrier and blood-retina barrier development and maintenance. Proc Natl Acad
Sci U S A 2018;115(50):E11827–E11836.
19. Jacklin HN. Falciform fold, retinal detachment and Norrie disease. Am J Ophthalmol
1980;90(1):76–80.
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Arch Ophthalmol 2010;128(4):456–460.
36
T. Y. Alvin Liu, and Morton F. Goldberg
INTRODUCTION
Incontinentia pigmenti (IP), also known as Bloch-Sulzberger syndrome, is a
rare inherited disease in which a generalized abnormality of ectodermal
structures manifests as characteristic skin lesions (Figure 36-1) and variable
degrees of retinal and central nervous system (CNS) disease due to a
progressive vasculopathy.
FIGURE 36-1 A newborn infant with IP and stage 1
dermatologic manifestations. The vesicles and erythema
are distributed linearly along the trunk and arms, sparing
the face.
PREVALENCE AND ENVIRONMENTAL

FACTORS
IP is a rare condition with approximately 1,000 cases reported in the literature
(1). It does not appear to have an ethnic predilection; nor have any
environmental factors been identified.
Ophthalmic manifestations are reported in approximately 35% of patients
with IP, although underdiagnosis of milder ophthalmologic manifestations
may lead to an underestimation of the prevalence of ophthalmologic
involvement (2). Approximately one in five IP patients will experience
vision-threatening disease. When blindness occurs, it is often due to retinal
detachment, which occurs in 3% of cases (2–4), but can be seen in up to 22%
of eyes with extended follow-up (5). Marked asymmetry of the retinal
vascular disease is common.
WORLDWIDE IMPACT
IP has been reported in patients around the world. Although the total number
of patients affected is small, untreated severe cases can cause blindness,
increasing the burden of vision impairment.
GENETICS
Pedigrees of IP reflect classic mendelian inheritance in an X-linked dominant
pattern. Because homozygosity is lethal in utero, only women inherit the
disease. Although highly penetrant, the expressivity of the disease is quite
variable presumably due to female X chromosome inactivation (lyonization).
De novo mutations appear to be common, as an affected pedigree is identified
in only half of new cases (2). Affected women in such pedigrees often had
male miscarriages. Extended pedigree analysis has demonstrated a 2:1 ratio
of daughters to sons among affected mothers. Most reports of males with IP
have no family history of the disease, suggesting somatic mosaicism (6). Rare
reports of males with inherited IP can be explained by a Klinefelter karyotype
(47,XXY), thus being exceptions that prove the rule (7). All males with the
disease should have their karyotypes evaluated for this reason. Appropriate
genetic counseling should be offered to all affected families.
DISEASE PATHOGENESIS
In 2000, the International Incontinentia Pigmenti Consortium identified
mutations in the gene encoding for nuclear factor kappa B (NF-kB) essential
modulator protein (NEMO) at locus Xq28 (8) as the cause of the disease (9).
This small protein acts as a regulatory subunit for activation of NF-kB, a
transcription factor that plays a critical role in the expression of genes crucial
to cell survival and proliferation. NF-kB regulates genes involved in critical
developmental processes, innate and adaptive immune responses, cell
adhesion, and apoptosis. It also mediates inflammatory cascades (10). A
genomic deletion accounts for 80% to 90% of the disease and suggests that
the disease is caused by loss of function.
Indeed, targeted deletion of the gene encoding NEMO in mice
recapitulates the human disease. NEMO-deficient male mice die in utero. The
female mice develop patchy skin lesions characterized by granulocyte
infiltration and hyperproliferation of keratocytes, similar to the histology of
patients with IP (11).
Histopathologic studies of the eyes of NEMO-deficient mice have helped
elucidate the pathogenesis of the retinal disease. Their retinal vasculature is
characterized by increased arteriolar tortuosity. The arteriolar lumens are
narrowed due to endothelial hypertrophy and basement membrane thickening
(12). Complete retinal vasoocclusion is commonly observed in human
histopathology of IP but was not seen in these mice (13). It seems plausible
that the luminal narrowing represents an earlier stage of the disease evolution.
Interestingly, the inflammatory changes and perivascular eosinophilic
infiltration seen in dermatologic histology were not observed in the retinas of
these mice.
The peripheral retinal perfusion of many patients with IP is abnormally
reduced and appears to be responsible for many of the blinding complications
of the disease. Fluorescein angiography (FA) of the disease reveals many
similarities between the retinopathy of IP and retinopathy of prematurity
(ROP). In both cases, vascular tortuosity is prominent. A rather abrupt
transition occurs between mature vascular central retina and an avascular
periphery (Figure 36-2). In severe cases, later fibrovascular proliferation,
exudation, and retinal detachment occur in a manner quite similar to ROP.
The advent of ultra–wide-field (UWF) retinal imaging, defined by the
Diabetic Retinopathy Clinical Research Network (DRCR.net) as a field of
view of 100 degrees or more (14), has permitted better characterization of
vascular abnormalities in the far peripheral retina in various retinal vascular
diseases, including IP (15). UWF FA findings in IP include abnormal
vascular anastomoses, pathologically straightened retinal vessels, varying
degrees of nonperfusion, and microaneurysms (Figure 36-3) (16).
FIGURE 36-2 A fluorescein angiogram of a young girl

with IP reveals peripheral nonperfusion, vascular loops,
and preretinal neovascularization.
FIGURE 36-3 Abnormalities on ultra–wide-field
fluorescein angiography (reproduced with permission from
JAMA Ophthalmology). Vascular anastomotic loops in the
far peripheral retina and areas of nonperfusion (arrow) in
(A) and (B).
Several important findings distinguish the retinopathy of IP from ROP.

Whereas the ischemia of ROP is entirely attributable to incomplete
vascularization, retinal vasoocclusion of previously formed vessels appears to
play a central role in the pathogenesis of IP. Fluorescein angiograms of the
latter can reveal patchy areas of capillary dropout more reminiscent of
diabetic retinopathy or sickle cell retinopathy than the halted angiogenesis of
ROP. Acute central retinal arterial occlusions with blindness have been
reported in IP (17). Foveal hypoplasia appears common in children affected
by ROP, whereas the maculae of children with IP are characterized by an
irregularly enlarged foveal avascular zone. The perifoveal capillary closure
that characterizes this process appears to be a dynamic process. In some
cases, abnormal vessels cross the ordinarily avascular zone of the fovea (17).

Children with IP can lose vision from disease of the retina, optic nerve,
and/or brain (18). The severe vision loss of IP is often due to vitreoretinal
complications of an avascular peripheral retina that can have a striking
similarity to ROP. Vascular remodeling and collaterals demarcate an abrupt
transition from vascular central retina to avascular peripheral retina. Often,
epiretinal neovascularization develops at this border. In severe cases, this
fibrovascular proliferation can lead to vitreous hemorrhage. It can also
contract, leading to detachment of the neurosensory retina. The most severe
cases develop a funnel-shaped retinal detachment indistinguishable from
stage 4 and 5 ROP. Less severe cases can produce epiretinal
neovascularization, temporal dragging, and macular heterotopias. Rarely,
initial ophthalmic manifestation of IP can present as anterior segment
neovascularization. In our practice, a 6-week-old infant girl first presented
with heterochromia due to extensive neovascularization of the iris in her left
eye. FA of her left eye showed extensive retinal nonperfusion for 360 degrees
without retinal neovascularization (Liu and Handa, unpublished data, 2017).
Macular pathology is another important cause of decreased vision among
patients with IP. Studies with FA have revealed an absent or anomalous
foveal avascular zone (Figure 36-4). This has been termed “foveal
hypoplasia.” Initially thought to reflect a developmental aberration, it more
likely represents vascular remodeling after vasoocclusive events in the
macula. An intriguing report of a transient classic “cherry-red spot”
appearance, observed in a 12-day-old infant with IP, supports this hypothesis
(19). The inventions of optical coherence tomography (OCT) and optical
coherence tomography angiography (OCTA) have revolutionized macular
evaluations of various retinal vascular diseases, including IP. OCT uses low-
coherence interferometry and measures time-of-flight delay of reflected light
to produce cross-sectional images of biologic tissues (20), and OCTA uses a
technique called split-spectrum amplitude-decorrelation (21) to image both
macular structure and blood flow simultaneously. Both imaging modalities
are noninvasive. Various structural abnormalities on OCT have been reported
in IP patients, including inner retinal thinning (16,22–24), irregularities of the
outer plexiform layer (16,24), and intraretinal cystoid changes (Figure 36-5)
(25). In addition, flow loss (23) on OCTA, specifically in the superficial
capillary plexus and deep capillary plexus (16), has been described in IP
patients together with decreased quantitative vascular density (Figure 36-6)
(16).
FIGURE 36-4 A fluorescein angiogram of a young girl
with IP reveals multiple retinal arteriolar occlusions and
capillary dropout, leading to a markedly enlarged foveal
avascular zone.
FIGURE 36-5 Structural macular abnormalities on
spectral-domain optical coherence tomography
(reproduced with permission from JAMA Ophthalmology).
Outer plexiform layer irregularity (A, B) and inner retinal
thinning (B, C).
FIGURE 36-6 Abnormalities on optical coherence
tomography angiography (reproduced with permission
from JAMA Ophthalmology). Macular vascular flow loss
in the superficial capillary plexus (outlined by yellow dots
in A) and in the deep capillary plexus (outlined by yellow
dots in B). Note images A and B were obtained from
different eyes.
Patients with IP can manifest abnormalities of the retinal pigment epithelium.

These findings include increased macular pigmentation, subretinal
pigmentary clumping, and mottling, sometimes associated with extensive
epiretinal proliferation (proliferative vitreoretinopathy) (4,26). When these
pigmentary changes are present, they are usually noted in the setting of
significant retinal vascular disease, often complicated by vitreoretinal
sequelae. These retinal pigment abnormalities likely reflect a secondary
disease process (27), rather than a primary abnormality in the retinal pigment
epithelium.
Microphthalmos and congenital cataract have been described in 10% to
20% of large case series (28,29). Optic nerve atrophy also has been reported
in these children. This atrophy likely reflects the extensive retinal damage
that usually accompanies the finding, although some cases may be caused by
intracranial pathology or possibly a primary vascular insult to the optic nerve.
Subepithelial corneal opacities can occur in children with IP, though there is
only one report of such opacity appearing visually significant. In adults, the
spectrum of anterior segment pathology that has been observed (cataract, iris
atrophy, band keratopathy, and phthisis) is likely secondary to the underlying
vitreoretinal pathology, especially retinal detachment.
Strabismus is common among patients with IP. Although many of these
cases can be attributed to sensory deprivation due to retinal or lens disease,
intracranial pathology, anisometropia, and a positive kappa angle may be the
cause in specific individuals.
SYSTEMIC MANIFESTATIONS OF
DISEASE
IP was originally recognized as a dermatologic disease. The widely adopted
clinical criteria for diagnosing IP emphasize the dermatologic manifestations
and do not account for the common CNS manifestations of the disease (Table
36-1) (30). These criteria were developed before identification of the genetic
defect but still prove useful for establishing a clinical diagnosis (1,28).
However, genetically confirmed cases of IP without clinical dermatologic
manifestations have been reported (30).
TABLE 36-1 Diagnostic criteria for IP

At least one major criterion is necessary to make a firm diagnosis of sporadic IP. The minor criteria, if
present, will support the diagnosis, but because of their high incidence, complete absence should induce
a degree of uncertainty.IP, incontinentia pigmenti.
Reproduced from Landy SJ, Donnai D. Incontinentia pigmenti (Bloch-Sulzberger syndrome). J Med
Genet 1993;30(1):53–59. With permission from BMJ Publishing Group Ltd.
The dermatologic disease can be described in distinct stages; however, in any

particular individual, the stages may overlap or be skipped (Figure 36-1):
Stage 1: Diffuse blisters and erythema with relative sparing of the face.
Stage 2: Hyperkeratotic papules develop on the distal limbs and scalp.
Stage 3: Linear hyperpigmented streaks and whorls along the lines of
Blaschko.
Stage 4: Pale, hairless patches often seen on the posterior calves.
A transient mild dystrophy of the nails is common. Rarely, painful keratotic

subungual tumors can develop (31). Mild alopecia is common, and the hair is
often wiry and coarse (32).
Dental abnormalities are common (~80%). The enamel and overall tooth
quality are normal, but delayed eruption, hypodontia, and malformation lead
to a typical dental appearance that is important to recognize and manage (31).
Breast abnormalities, including unilateral aplasia, hypoplasia, and
supernumerary nipples, appear more common among patients with IP (30).
CENTRAL NERVOUS SYSTEM

INVOLVEMENT
CNS abnormalities, together with retinal disease, account for most of the
morbidity of IP. Large case series report that approximately 30% of patients
with IP have clinically evident CNS disease (2). It appears that the CNS
involvement is most common among those with prominent skin disease (33).
Thus, the true incidence of CNS disease may be significantly lower if patients
with more mild skin findings are included (28,30).
CNS disease in IP may manifest as an isolated seizure, epilepsy, ischemic
stroke, microcephaly, encephalopathy, or encephalomyelitis. Affected
children can develop hemiplegias, cerebellar ataxias, psychomotor delay, and
intellectual disability. Ischemic stroke usually occurs in the first 2 months of
life but has been reported in a 4-year-old (34). Reports have described strokes
in the distribution of large cerebral arteries (35–37), but angiographic
findings support a microvascular occlusive etiology that predominantly
involves the deep white matter (36,38). Seizures are the most common CNS
manifestation of IP (34). They usually present within the first week of life
and almost always before age 4. The rare reports of first seizures in later
childhood occur in the setting of new CNS disease such as stroke (37). MR
imaging of children with seizures due to IP almost invariably demonstrates
vascular insufficiency due to ischemia and necrosis (34). There are also two
interesting reports of children with IP who developed acute disseminated
encephalomyelitis at ages 6 and 7 months. Both had laboratory and radiologic
evidence of demyelinating disease and recovered good function (39,40).
Neurologic imaging of patients with IP commonly shows periventricular
leukomalacia and subcortical white matter lesions. Occasional reports of
white matter cavities and cysts, as well as ventricular dilation and cerebral
atrophy, suggest tissue loss from vasoocclusive events. Intracranial bleeding
is often detected. This is usually present in the setting of hemorrhagic
necrosis and encephalomalacia consistent with ischemic events. Patterns of
cerebellar hemisphere and vermis atrophy are likewise thought to be
secondary to intrauterine cerebrovascular damage (41,42). IP can include
polymicrogyria, a developmental brain disorder due to second trimester
insult, demonstrating antenatal CNS disease (43).
Only limited neuropathology reports are available. A 7-week-old infant
was found to have focal areas of cerebral and cerebellar necrosis and
neuronal loss consistent with vascular insufficiency, without demonstrable
pathologic changes of the blood vessels (44). A 3-month-old was found to
have similar evidence of vascular insufficiency, but perivascular cuffs of
lymphocytes, histiocytes, and eosinophils, as well as mononuclear nodules,
were also described suggesting that inflammation plays a role in the
pathogenesis of vaso-occlusion (45).
DIAGNOSTIC STUDIES
Genetic sequence analysis is available to detect both the genomic deletion
that accounts for 80% of the disease and smaller mutations (usually in exon
10) that account for another 8.6% of the disease. For rare cases in which
genetic testing does not establish a diagnosis, peripheral eosinophilia and skin
biopsy findings of eosinophilic infiltration and extracellular melanin can
support the diagnosis (46).
FA is an invaluable tool for evaluating the retinal vasculature. For
patients with IP, it can reveal the extent of peripheral nonperfusion and
subclinical neovascularization and guide peripheral ablation by
photocoagulation (47,48). A large series of infants examined by FA
demonstrated irregular and distorted foveal avascular zones in all eyes (19).
Careful sequential examination of the macula with FA revealed abnormal
macular perfusion and dynamic remodeling of the fovea as well as a macular
infarct.
Recent advancements in retina imaging, including UWF FA, OCT, and
OCTA, have permitted detection of structural and vascular abnormalities that
may not be apparent on standard ophthalmoscopy in IP patients (15,22).
UWF FA can capture pathologies that are peripheral to standard views of 30-
degree FA even with peripheral sweeps (16). Simultaneous multimodal
imaging shows that IP patients can have severe peripheral vascular
pathologies with near-normal macular structure and vasculature, suggesting a
potential difference in susceptibility of the central and peripheral retina to
vascular insult. Areas of flow loss on OCTA are associated with inner retinal
thinning on structural OCT in the same locations, although the exact temporal
relationship between pathologies seen on these two imaging modalities needs
further investigation (16).
When vitreous hemorrhage, cataract, and retrolental fibroplasia prevent
an adequate retinal exam, ocular ultrasonography can characterize the extent
of disease and narrow the differential diagnosis by excluding calcification
and/or mass lesions. The electroretinographic recordings of a 13-month-old
girl with IP have been reported (49). Both photopic and scotopic responses
were markedly diminished. The decreased flicker response to 30 Hz and
abnormal oscillatory potentials indicate cone function compromise consistent
with the foveal abnormality observed in this patient.
IP has a highly characteristic dermatologic appearance. When the skin lesions
are present, peripheral retinal nonperfusion and the associated vitreoretinal
sequelae are best explained by this diagnosis. Occasionally, the skin lesions
are subtle or subclinical, and alternative ophthalmologic diagnosis must be
entertained. ROP, familial exudative vitreoretinopathy, Norrie disease, Eales
disease, and sickle cell retinopathy are all characterized by peripheral retinal
nonperfusion and secondary neovascularization. Extensive peripheral
nonperfusion on FA has been shown in children who sustained nonaccidental
trauma (50,51). Epiretinal neovascularization has been observed in this
setting (52). If no view of the posterior pole is possible, the differential
diagnosis must be expanded to include other causes of leukokoria.
Screening
Of all the various manifestations of IP, the peripheral retinal vascular disease
is most amenable to treatment. Like many diseases of the peripheral retinal
vasculature, prompt, thorough ablative therapy with laser photocoagulation,
preferably, or cryotherapy may prevent and/or halt the vision-threatening
sequelae of retinal neovascularization, exudation, hemorrhage, and retinal
detachment. Because the incidence of vasoocclusive events peaks in the early
postpartum period, early diagnosis of the disease and recognition of the need
for ophthalmic monitoring are critical. Children with IP should be evaluated
soon after diagnosis by an ophthalmologist skilled in examination of the
pediatric peripheral retina and strong consideration should be given to
performing a UWF FA to detect areas of nonperfused retina that might be
missed on funduscopic examination (15). When no retinal pathology is
evident, the interval of follow-up depends on the age of the child and the
extent to which the periphery can be evaluated. When the diagnosis is
suspected at birth, due to either characteristic skin lesions or family history,
retinal examination within 1 or 2 weeks of birth is optimal. Even very subtle
vascular abnormalities warrant close follow-up, as the retinal vascular disease
can progress rapidly over the first few weeks of life (44). Generally, we
believe that if ablative treatment is delivered for nonperfused retina and/or no
significant pathology develops by 6 months of age, then the interval of
follow-up can be extended. Examination under anesthesia is often required.
More studies are required to better understand the long-term risk in these
patients.
MANAGEMENT
Peripheral Ablation
Some children with IP and peripheral retinal avascularity develop profound
visual loss due to retinal detachment. Initial reports of peripheral ablation
with cryotherapy and laser photocoagulation described prompt control of the
neovascular response and prevention of retinal detachment (53–55).
Subsequent reports of treatment for milder degrees of neovascularization and
for extensive nonperfusion without neovascularization support peripheral
ablation as an effective therapy for stabilizing the disease (56–60). However,
the criteria for treatment of these milder cases are not clear, as the
proliferative retinopathy of IP often stabilizes without intervention and
peripheral ablation can be associated with an increased risk of cicatricial
pathology such as epiretinal membranes and proliferative vitreoretinopathy.
One report describes a girl, with peripheral nonperfusion, neovascularization,
and macular ectopia, who maintained vision over 13 years of follow-up
without treatment (61). In our practice, five children with mild
neovascularization have been monitored without treatments and remained
stable with follow-up ranging from 1 to 6 years. Nonetheless, some adults
present with tractional detachments related to IP and nonperfused retina that
had not been previously treated, suggesting a reason for early treatment of
nonperfused retina.
Vitreoretinal Surgery
Repair of retinal detachment due to IP is possible, although anatomic success
rates appear low in total retinal detachment and may be similar to eyes with
stage 5 ROP (27). Often, a combination of vitrectomy and scleral buckling is
employed (see also Chapters 60 and 61). Preservation of vision may depend
on the extent of the retinal detachment and of ischemic retina that might
affect the macula (22). In early studies, prior to UWF FA and indirect laser
management, at least one child retained fixation behavior and limited
ambulatory vision (27). Similar to ROP, central involvement is associated
with poor anatomic outcomes (62). In cases of localized neurosensory
detachment, observation without surgery occasionally may be reasonable, as
there is a report of a patient with retinal pigment epithelium
hypopigmentation and retinoschisis in a pattern suggestive of a spontaneously
resolved localized retinal detachment (63).
Intravitreal Antivascular Endothelial Growth Factor

Therapy
Intravitreal antivascular endothelial growth factor (VEGF) injections are
controversial in IP. There are two cases in which ranibizumab or
bevacizumab was reported to be efficacious adjuvant therapy for proliferative
retinopathy (64,65). However, the use of intravitreal anti-VEGF therapies in
premature infants reduces systemic VEGF levels (66). Thus, anticipated
benefit from its use should be carefully weighed against potentially serious
systemic complications, especially given the higher risk of ischemic stroke in
IP patients and the lack of strong evidence that anti-VEGF treatment is
beneficial in IP.
Media Opacity
Children with IP can develop cataract and vitreous hemorrhage, which should
be addressed promptly to minimize the risk of amblyopia (29). The corneal
subepithelial opacities are generally not visually significant.
Strabismus
Strabismus in children with IP often presents in infancy, is usually attributed
to sensory deprivation due to retinal pathology, and can be addressed for
cosmesis in later childhood (29). Pediatric eyes with significant macular
pathology can occasionally have surprisingly good visual acuity. However,
and if the child has good vision in both eyes, ocular alignment surgery should
be considered in the first years of life to maximize the chance for stereopsis.
Amblyopia therapy may be appropriate for many children with ocular
manifestations of IP. In the setting of a normal-appearing retina,
neuroimaging may be considered.
Enucleation
In contrast to ROP, the vitreoretinal disease of IP is often markedly
asymmetric. Good visual function is often present in the fellow eye. Thus,
aggressive attempts at surgical repair of very advanced cases are sometimes
not performed because of the possibility of phthisis and intractable pain.
Enucleation in the setting of advanced IP has been reported, although painful
amaurosis fortunately appears rare (29).
Adult Vitreoretinal Sequelae

Patients with IP require lifelong vitreoretinal monitoring. In our practice, a
young girl with IP developed a visually significant epiretinal membrane at 7
years of age, after undergoing laser ablation and cryotherapy as an infant. Her
vision improved from 20/200 to 20/40 after vitrectomy with membrane peel.
She was stable until she was 13 years old, when she developed a localized
exudative retinal detachment with intraretinal lipid and fluid in the temporal
macula, requiring scleral buckle placement and additional laser and
cryotherapy (Baranano, Goldberg, and Wenick, unpublished data, 2018). An
untreated, avascular retina puts patients at increased risk for retinal tears at
the time that they develop a spontaneous posterior vitreous detachment (67).
Our extended follow-up study shows that up to 22% of IP eyes can develop
retinal detachment over long observational periods. There is a bimodal
distribution of retinal detachments with most tractional retinal detachments
occurring by age 2.5 years and most rhegmatogenous retinal detachments
happening in adults at a median age of 31.5 years (5).
Visual Rehabilitation
Some patients with IP develop low vision or legal blindness despite the best
ophthalmologic care. These patients should be referred at the earliest possible
time to a low vision service for guidance and assistance in procuring
accommodations for home, work, and school.
Roles of Other Providers

The care of children and families with IP often requires the coordinated care
of multiple specialties, including dermatology, pediatrics, neurology,
dentistry, and genetic counseling as well as ophthalmology. The extent of
involvement of each specialty is determined by the particular pattern of the
individual patient’s disease and needs. As the skin lesions typically improve
spontaneously, dermatologic management usually needs only to be
supportive. Laser treatment of skin hyperpigmentation is discouraged because
of a report of this therapy triggering an extensive vesiculobullous eruption
(68). In the absence of clinical neurologic disease, extensive neurologic
imaging can be deferred. However, because of the high concordance of
neurologic and ophthalmologic findings (33), neuroimaging should be
considered for children with retinal vascular disease. Likewise, once a
detailed ophthalmic exam reveals a normal retinal vasculature beyond
infancy, routine ophthalmic monitoring is probably sufficient by itself unless
symptoms of CNS disease occur.
All affected women, regardless of the severity of their phenotype, should
understand the importance of timely neonatal eye exams for their daughters
and the likelihood of lethality in males. Genetic counseling should be
provided. If a woman has a molecular genetic diagnosis, in vitro fertilization
with preimplantation genetic testing may be possible to reduce the chance of
an affected offspring. Families should be counseled that spontaneous
mutations do occur, so not having a family history does not mean the
condition is not genetic. In addition, due to extremely variable expressivity, it
is important for families to understand that future affected generations may
be more or less severely affected than the current generation.
FUTURE TREATMENTS
Therapies to replace the defective NEMO protein may be a possibility for
patients in the future. Continued evolution of surgical techniques for
alleviating retinal traction and detachment may improve the outcome for
patients who have severe disease.
ACKNOWLEDGMENT
We would like to acknowledge and thank Dr. David Baranano for his
invaluable contribution to the previous edition’s chapter on incontinentia
pigmenti.
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37
High Myopia and Vitreoretinopathies
Sherveen S. Salek, and G. Baker Hubbard III
OVERVIEW, PREVALENCE, AND

WORLDWIDE IMPACT
Myopia remains a growing public health challenge worldwide with increased
incidence and prevalence over the past several decades especially in younger
children (1). Estimates predict that by the year 2050, myopia and high
myopia will affect 5 billion and 1 billion people, respectively, worldwide (2).
High myopia, defined as >6 D of negative refractive error, is associated with
increased risk of pathologic complications, including retinal detachment
(RD), primary open-angle glaucoma, and choroidal neovascularization in
adults. Juvenile myopia has also grown in incidence, increasing the need for
earlier identification and correction of refractive error (3). Furthermore,
juvenile myopia has several important syndromic associations with additional
ocular and systemic comorbidities that require identification and treatment.
The most important of these, Stickler, Wagner, Marfan, Knobloch, and
Ehlers-Danlos syndromes, will be discussed below.
PATHOPHYSIOLOGY AND
High myopia is characterized by elongation of axial length followed by
stretching of the posterior eye wall, leading to complications such as
choroidal thinning, staphyloma, macular schisis, macular hole, and changes
in optic nerve morphology (4).
Both environmental and genetic factors are thought to play a role in axial
length elongation, although the exact etiology and mechanisms remain
unknown. Time spent outdoors in childhood is thought to play a role in
preventing onset of myopia, although the correlation of time spent outdoors
with progression of myopia remains less clearly established (5). A correlation
between increased levels of schooling or professional education and myopia
has also been found (6). Recent studies have found an increase in prevalence
of myopia, up to 80%, in Asian schoolchildren. The rate of high myopia in
these children has increased to 20% (7). In East and Southeast Asian
children, high myopia significantly increases in prevalence around the age of
11, with affected children appearing to become myopic initially around the
age of 6. Time spent outdoors appears to affect the progression of these
children to high myopia, and several interventions have been proposed to
increase time outdoors for schoolchildren with the aim of addressing this
public health challenge. There are increasing numbers of prospective studies
evaluating the role of time spent outdoors in development and progression of
myopia, which have tried various methods of standardization to ensure that
time spent outdoors is accurately measured (8).
GENETICS
The continuing epidemic of nonsyndromic myopia in recent decades has
raised the question of whether genetic or environmental factors are more
relevant. Numerous genes have been associated with nonsyndromic myopia.
One review of recent literature listed 21 candidate myopia genes, some of
them identified in earlier genome-wide association studies, which are
involved in cellular and biochemical processes as diverse as mannosylation,
glycosylation, lens development, gliogenesis, and Schwann cell
differentiation (9). Many of these processes are involved in connective tissue
formation. The goal of future studies is to identify the pathogenesis of these
mutations and study potential interventions for nonsyndromic myopia based
on this knowledge.
Stickler syndrome is a hereditary connective tissue disorder with multiple
ocular and extraocular phenotypes caused by a broad array of genetic
mutations. Abnormalities of the vitreous gel, commonly termed as the
pathognomonic “membranous vitreous,” constitute type I Stickler syndrome
and constitute the majority of patients (10,11). This phenotype correlates with
mutations in the COL2A1 gene. Type II Stickler syndrome consists of the
beaded and irregular strands of vitreous scattered sparsely throughout the
vitreous cavity and has been shown to correlate with COL11A1 mutations
(12). More recent investigations, however, have found patients with COL2A1
mutations with a hypoplastic vitreous phenotype with sparse irregular
lamellae.
The early onset of high myopia with vitreous abnormalities is thought to
be a marker for Stickler syndrome and should warrant further investigation,
including a detailed family history, retinal examination of both eyes, skeletal
survey, and consideration of genetic testing (13). Genetic testing is especially
useful in patients with early-onset high myopia (defined as greater than minus
6 D of refractive error prior to age 7) and whose evaluations do not reveal
additional systemic abnormalities or family history. Examination of children
with early-onset high myopia with mutations in COL2A1 and COL11A1 may
reveal the presence of posterior vitreous detachment, foveal hypoplasia,
hypermobility of the elbow joint, and vitreous abnormalities (14).
Wagner syndrome is caused by mutations in the VCAN gene (also known
as Versican or CSPG2), resulting in haploinsufficiency and an autosomal
dominant phenotype. Like Stickler syndrome, Wagner syndrome can also
have extraocular manifestations. A case report by Ankala and colleagues
found that an 11.7 kilobase (kb) deletion encompassing exon 8 of VCAN was
correlated with distinctive facial features and gastrointestinal symptoms (15).
Mosaicism in VCAN mutations is also correlated with a milder Wagner
phenotype, demonstrating asymmetry between each eye (16).
Marfan syndrome is caused by mutations in FBN1, which encodes the
fibrillin-1 protein. Fibrillin microfibrils are found in association with elastic
fibers and are essential components in many extracellular matrix tissues (17).
Histologic examination of aborted human fetal eyes from 5 to 11 weeks of
gestational age reveals increased expression of fibrillin-1 protein in vascular
structures, bridging the ciliary body and the developing lens, hyaloid, and
retina, which suggests a scaffolding role for this vasculature in the embryonic
development of these structures (18).
Initial genetic mapping in 1996 localized the causative mutation in
Knobloch syndrome to the 21q22.3 locus and consequently identified
COL18A1 as the causative gene (19). The encoded protein has a role in
determination of retinal architecture and is also thought to affect
embryogenesis and neural tube closure. A recent review of 12 patients from 7
families found presence of anterior segment abnormalities in Knobloch
syndrome, including cataract, lens subluxation, absent iris crypts, and
transillumination defects (TIDs) in the iris (20). The presence of TIDs was
associated with glaucoma in two patients in this series. Fundus characteristics
in this autosomal recessive condition include a collapsed vitreous appearance,
macular atrophy, and mottling of the fundus. Interestingly, COL18A1
mutations have been linked with iridocorneal angle closure in adults without
any other ocular or systemic features of Knobloch syndrome (21). This
finding lends further support to the role of the collagen-encoding gene in the
formation of angle structure, in addition to the retinal architecture.
CLINICAL FEATURES
Myopia can be diagnosed on the basis of the spherical equivalent of the
negative refractive error. Retinal findings in myopia include chorioretinal
atrophy, lattice degeneration, pigmentary degeneration, lacquer cracks,
posterior staphyloma, Fuchs spot, macular degeneration, as well as retinal
breaks and RD (22). Nonsyndromic myopia refers to the absence of any other
extraocular manifestations.
Stickler syndrome, also called hereditary arthro-ophthalmopathy or
hereditary arthro-ophthalmo-dystrophy, is an inherited disorder of collagen
formation. Besides its ocular manifestations, patients also have
musculoskeletal, orofacial, and auditory abnormalities. The condition remains
rare with an incidence of approximately 1 in 7,500 to 9,000 live births (23).
Stickler syndrome is characterized by the variable presence of a small
mandible, flat nasal bridge, upturned nasal tip, bifid uvula, cleft palate,
vitreous abnormalities, cataract, and increased rate of RD (24) (Figure 37-1).
Stickler syndrome is divided into the type I, type II, and nonocular type III
phenotypes. As discussed previously, type I patients have mutations in
COL2A1 and have the distinct optically empty or, by some descriptions,
membranous vitreous appearance, whereas type II Stickler patients have
strands of beaded vitreous present with mutations in the COL11A2 gene (25).
Cortical cataracts, often wedge-shaped, can be present in Stickler syndrome,
and increased incidence of infantile-onset glaucoma is also observed (26).
The fundus examination can reveal both radial perivascular and
circumferential lattice degeneration. RDs can occur in 50% of patients with
Stickler syndrome and result from giant retinal tears, usually located
anteriorly, as well as posterior breaks in the retina. Careful examination of the
retinal periphery is warranted in all patients with Stickler syndrome or with a
suspected family history.
Extraocular manifestations of Stickler syndrome that can aid in the
diagnosis include the above-mentioned craniofacial anomalies, in particular
the flat nasal bridge. Additional findings include thoracolumbar spinal
anomalies. One series reported 34% of patients with scoliosis, 74% endplate
abnormalities, 64% Schmorl nodes, 43% platyspondylia, and 43%
Scheuermann-like kyphosis (27). Of note, only 1 patient in this series of 53
patients had absence of any spinal abnormalities. Although the findings were
usually associated with symptoms of back pain, they were generally self-
limited and did not require spinal surgery. Stickler syndrome is also
associated with sensorineural hearing loss. Type I patients have
predominantly higher-frequency hearing loss on audiometric testing, and type
II patients have more moderate hearing loss across low and high frequencies
(28). Conductive hearing loss has also been reported in patients with Stickler
syndrome who have cleft palate defects. Patients with mutations in COL11A1
and COL11A2 (Type II phenotype) appear to have a higher association with
clinical hearing impairment than those with COL2A1 mutations (Type I) (29).
Wagner syndrome is an autosomal dominant inherited vitreoretinopathy
caused by a mutation in the VCAN gene. The prevalence is unknown, and
only 300 cases have been described worldwide with approximately half in the
Netherlands (30). Patients with Wagner syndrome present with ocular
findings consisting of empty vitreous, lattice degeneration, chorioretinal
atrophy, and mild myopia. Peripheral tractional RDs are observed in patients
with Wagner syndrome and usually do not have extraocular manifestations
(31) unlike patients with Stickler syndrome. Dark adaptation responses are
usually preserved. However, unlike Stickler syndrome, Wagner syndrome
can be associated with diminished B-wave amplitudes on both light-adapted
and dark-adapted electroretinography (ERG). Furthermore, multifocal ERG
and visual fields can also be severely restricted in these patients (32).
Marfan syndrome is an autosomal dominant connective tissue disorder
present in approximately 1 in 5,000 people (33). Marfan syndrome is caused
by mutations in the fibrillin gene, FBN1, which encodes an extracellular
protein, fibrillin-1, and is associated with numerous ocular (including
vitreoretinal), musculoskeletal, and cardiac abnormalities associated with
significant morbidity and mortality (34). Previous investigators have noted an
incidence of RD of 3.5% to 11% in patients with Marfan syndrome (35).
Patients with ectopia lentis or prior lens extraction have a higher incidence of
RD with one study reporting RD in 38% (36).
Knobloch syndrome is a rare autosomal recessive condition characterized
by high myopia and, in some cases, occipital encephalocele. The prevalence
is unknown (37). Patients also have increased risk of RD, vitreoretinal
degeneration, lens subluxation, cataract, smooth irides, and persistent fetal
vasculature (20).
Ehlers-Danlos syndrome is a heterogeneous group of connective tissue
disorders most commonly characterized by joint hypermobility and laxity and
is associated with an increased risk of RD (38). Categorized into various
subtypes, the condition is present in approximately 1 in 5,000 individuals
(39). However, the prevalence of retinal pathology in these patients is
unknown.
DIAGNOSTIC STUDIES
The diagnosis of high myopia is based on meeting the threshold refractive
error of a spherical equivalent of at least minus 6 D. Several investigators
have sought to determine if there are factors that can predict nonsyndromic
pediatric high myopia. A large observational cohort study found that the
presence of low hyperopia or moderate myopia on spherical equivalent
refraction was associated with later development of juvenile myopia and
subsequent progression (40). Lens power, axial length, and accommodative
convergence to accommodation (AC/A) ratio also had a statistically
significant association, but these measurements are difficult to ascertain in a
clinical setting. Closer reading distance, in addition to a more myopic
baseline refraction, has been associated with faster progression of myopia
over a 1-year period in a study of second grade schoolchildren in Taiwan
(41).
Slit lamp and dilated fundus examination form the cornerstone of the
diagnosis of both juvenile myopia and the inherited vitreoretinopathies. A
careful family history can alert one as to whether additional workup of family
members or more advanced diagnostics are necessary. In patients suspected
of having Stickler syndrome, genetic testing may be warranted and can help
with counseling of other family members. Radiographic studies, audiometry,
and electrocardiography can also alert the clinician to the presence of
extraocular manifestations that require monitoring and potential treatment.
Similarly, patients with Marfan syndrome may benefit from genetic testing
and further consultation with pediatrics and cardiology for monitoring of
potentially life-threatening aortic vascular pathology (42).
MANAGEMENT
Myopia and vitreoretinopathies may result in RDs that can warrant surgical
interventions (see also Chapters 60 and 61). This section discusses several
key areas of management of juvenile myopia and inherited
vitreoretinopathies that do not involve intraocular surgery or scleral buckling.
Prophylactic Retinopexy in Stickler Syndrome
Equipment
Cryopexy system or indirect laser.
Supporting Evidence and Cryopexy and Laser

Techniques
Given the substantially elevated risk of rhegmatogenous RD in patients with
Stickler syndrome, it is reasonable to consider prophylactic therapy to reduce
the risk of RD. Type I Stickler with COL2A1 mutations is believed to be
particularly amenable to prophylactic treatment in the peripheral retina,
because a high proportion of causative retinal breaks in type I Stickler occur
at or near the ora serrata. A retrospective comparative case series of 487
patients with type 1 Stickler syndrome found that patients with both unilateral
and bilateral prophylactic treatment had 8.4-fold and 5.0-fold reduction,
respectively, in the risk of RD in comparison to matched untreated control
patients (43). In the study, 229 underwent bilateral and 64 unilateral
prophylaxis, and 194 received no prophylactic intervention. Of the 487
patients, 426 were tested for COL2A1 mutations and 96.9% of these were
found to test positive. The technique utilized in this study consisted of a
single row of contiguous transconjunctival cryopexy of the anterior retina just
posterior to the ora serrata for 360 degrees. This technique, as described by
Snead and colleagues, has become known as the Cambridge prophylactic
cryotherapy protocol (43,44).
Outcome Expectations
Of note, about 9.0% of patients failed prophylactic therapy. Failure of
prophylactic therapy was attributed to both untreated posterior breaks as well
as anterior breaks that extended through the cryotherapy barrier (43,44).
Although the published Cambridge protocol from 2014 is the most robust
data available to support prophylactic cryotherapy in patients with Stickler
syndrome to reduce the risk of RD, there are no randomized clinical trials yet
available. Furthermore, many ophthalmologists find it more facile to treat
with laser retinopexy as opposed to cryotherapy and extrapolate the results of
the Cambridge protocol to the former treatment (Figure 37-2). Case series of
successful prophylactic laser for Stickler syndrome have been reported in the
literature, although the reports are on a smaller scale (45). The optimal laser,
location, and extent of treatment remain unknown (46). Even with successful
treatment, retinal detachment can occur from breaks i the posterior retina
(Figure 37-3).
Complications
The authors note that only a single patient had an RD that was attributed to a
new break that was formed as a result of contraction at the posterior border of
the cryotherapy treatment. No serious side effects (choroidal hemorrhage,
macular pucker, or unexplained visual loss) were reported in the study, and
the most common side effect was eyelid or conjunctival inflammation.
Accommodative insufficiency and anisocoria/mydriasis were reported in
9.6% and 2.0% of patients, respectively. Most accommodative insufficiency
resolves after about a month in children (43,44).
Outdoor Time and Myopia
Outdoor time has been hypothesized to prevent onset and progression of
myopia. One of the challenges of large studies to evaluate outdoor time and
light exposure has been ensuring standardization and appropriate
measurement of light exposure and activity both during and after school
hours. Most studies on this subject depend on questionnaires of children,
parents, and teachers. However, these assessments can be subject to recall
bias, especially given that they depend mostly on parents’ completion (47). A
combination of accelerometers and global positioning system (GPS) devices
has been used to track indoor versus outdoor times for children, but even
these methods can sometimes fail to distinguish between these two settings.
The devices may also not accurately measure activity, although they are
continuing to improve in terms of accuracy and cost and are being used more
often in myopia studies. Sensors can be incorporated into custom-designed
wearable vests in order to ensure adherence and proper measurement (48).
Portable light meters have also been used to measure light exposure, which is
the direct variable that has been hypothesized to affect myopia onset and
progression through the light-dopamine pathway (49).
A randomized controlled trial in Taiwan examined the effectiveness of
increased outdoor time in schoolchildren aged 6 and 7 and controlled for light
intensity in outdoor activities (50). Participants in the intervention group were
encouraged to have 11 or more hours of outdoor activity each week and wore
light meters during school hours in order to record light exposure accurately.
Outside of school, participants kept logs to record outdoor exposure. At the
end of 1 year, patients in the intervention arm with increased outdoor
exposure had reduced rates of onset of myopia, progression of refractive
myopia, as well as reduced axial elongation (0.28 mm vs. 0.33 mm).
Although these results were statistically significant, their clinical significance
is still being studied.
An additional study of 382 first graders from Beijing found less time
spent outdoors, more time spent indoors, and parental myopia to be
associated with axial elongation during a 4-year follow-up. This study was
based on a questionnaire of recall for outdoor exposure. Paternal education,
family income, gender, and region of residence were also associated with
axial length elongation and progression of myopic refractive error but only in
univariate analysis (51).
Atropine
Topical atropine has gained increasing utilization for prevention of myopia
and progression with multiple randomized controlled trials, demonstrating its
safety and efficacy (52). In Taiwan, 49.5% of children diagnosed with
myopia were prescribed topical atropine in 2007 (53). Global surveys have
shown that this trend has gone worldwide with an increasing number of
pediatric ophthalmologists initiating treatment with topical atropine for
children with progression of myopia (54). Additional studies in China have
demonstrated reduced axial length elongation in children 7 to 12 years of age
who receive atropine (55). Increasingly lower concentrations of atropine are
being used due to studies concluding equal effectiveness with reduce adverse
effects. Atropine arrives in a standard 1% concentration, but diluted 0.01%
atropine has been reported to slow myopic progression over 1 year of follow-
up. Children with low baseline refractive error (less than minus 1 D) have
been reported to respond especially well to diluted atropine in comparison
with controls (56).
A large meta-analysis of atropine for treatment of myopia examined
3,137 children from 19 unique studies (both randomized controlled trials and
cohort studies). Weighted mean differences in myopic progression between
treatment and control groups were 0.50 D per year for low-dose atropine,
0.57 D per year for moderate-dose atropine, and 0.62 D per year for high-
dose atropine (57). The authors concluded that all doses were equally
beneficial with respect to myopia progression. High-dose atropine was,
however, associated with more photophobia (43.1%) than low-dose atropine
(6.3%) in the meta-analysis. Interestingly, there were differences in axial
length elongation between the various concentrations of atropine, although
the reduction of spherical equivalent refractive error was similar (58). In
addition to photophobia, other common adverse effects of atropine include
poor near visual acuity and allergy (57). Due to the potential for systemic
absorption and its antimuscarinic effects (including irritability, fever, and
delirium), atropine topical drops are not recommended under the age of 3
months and are limited to one drop per eye per day in children under 3 years
of age. In addition, there are no data on efficacy of atropine in syndromic
myopia and, therefore, it is not recommended.
Refractive Surgery
Although excimer laser refractive surgery is FDA-approved and widely used
for adults age 18 and older, there is less data for its use in children. Use in
children should be restricted to cases of anisometropic amblyopia or bilateral
high refractive error with poor compliance with refractive correction (59).
There are several challenges to its use in a pediatric population that include
the need for sedation or general anesthesia in younger children; the need to
transport heavy and expensive excimer laser equipment from the clinic to a
hospital or surgery center where pediatric anesthesiology is available; and the
difficulty in preventing children from eye rubbing in the postprocedure
period (60). Changing spherical equivalent refraction and axial length during
growth and development are additional challenges to pediatric refractive
surgery. However, laser in situ keratomileusis (LASIK), photorefractive
keratectomy (PRK), and phakic intraocular lens (pIOL) implantation have all
gained increasing use in the pediatric population for a broad range of
refractive errors. PRK has been used in children as young as 1 year old,
whereas LASIK has been reported for children as young as 5 years of age
(61). Parents should be counseled that, in addition to potential procedural
complications, additional correction with glasses or contact lenses might still
be necessary after LASIK or PRK in children. More frequent postoperative
checks may be necessary, and ophthalmologists should emphasize the need to
avoid eye rubbing to parents and children who have undergone refractive
surgery. Potential LASIK complications include flap dislocation and potential
destabilization of the corneal stroma. PRK complications can include corneal
haze and substantial postoperative pain. Potential complications of pIOL
include pupillary block, pigment dispersion, ocular hypertension, endothelial
cell loss, RD, and endophthalmitis (60).
Orthokeratology
Orthokeratology refers to the reshaping of the corneal surface through contact
lens wear and is effective for treating myopic spherical error up to minus 5.0
D and up to 1.5 D of astigmatism (62). The normal cornea has a prolate
curvature, meaning that it is steep centrally and flat in the periphery. Reverse
geometry gas permeable lenses have a flat secondary curvature and are worn
in order to produce a flattening of the central cornea, allowing for treatment
of myopia (63). The lens “pushes” against the central cornea and “pulls” on
the periphery, creating a flatter, more plateau-shaped architecture. Several
randomized controlled trials have shown a benefit to gas permeable
orthokeratology lenses at slowing myopic progression and reducing axial
length elongation. However, the effects on axial length elongation do fade
after 5 years (64). Lens hygiene should be emphasized, as there is a risk of
microbial keratitis associated with orthokeratology lenses. Several factors
have resulted in an improved safety profile for orthokeratology lenses in
clinical practice over the last few years. These include advances in lens
material and design with good oxygen permeability, coupled with rigorous
education of parents, children, and providers, which includes specific
certification required in order to fit these lenses. Despite these improvements,
the risk of microbial keratitis remains. Patients should be encouraged to
discontinue lenses and to have an evaluation promptly for any discomfort.
Common organisms from culture results include Acanthamoeba and
Pseudomonas aeruginosa (65).
Refractive error and amblyopia are common indications for low vision
evaluation and services in various global settings, especially in children age 6
and above (66,67). Visual disability secondary to refractive error in children
is common, is often uncorrected, and poses a significant public health
challenge for providers in almost any educational setting (68). Correction of
refractive error secondary to myopia and hereditary vitreoretinopathies
should remain the priority for the clinician in order to maximize vision
potential and prevent amblyopia in the growing child. There remains a dearth
of data from clinical trials evaluating the efficacy of low vision aids
specifically for myopia in children, and such future investigations would
yield insight as to which interventions would be most helpful for this
population (69). In particular, further study of assistive technologies for low
vision is necessary (70). This may assist patients with macular pathology
whose vision does not improve with refraction.
Roles of Other Physician and Health Care Providers

Several of the above-mentioned disorders have systemic manifestations that
require close collaboration between the ophthalmologist and health care
providers in other specialties. In particular, the pediatrician serves a critical
role for initial identification and triage of extraocular findings and appropriate
referral for diagnostic testing, genetic counseling, and pediatric subspecialty
care (71). Communication of these findings with family members is
paramount at all stages of care.
Marfan syndrome, especially its cardiovascular effects, requires close
monitoring and management. In recent years, it has become evident that the
cardiovascular manifestations of Marfan syndrome are not simply a
byproduct of premature aging but are actually present and pathologic during
childhood (72). Aortic disease, the most life-threatening of the manifestations
of Marfan syndrome, most commonly results from progressive ascending
aortic aneurysm. Proper diagnosis of Marfan syndrome is critical, as several
other entities, such as Loeys-Dietz syndrome and Ehler-Danlos syndrome,
can mimic the findings of Marfan syndrome (73). The revised Ghent criteria
for diagnosis of Marfan syndrome place a greater emphasis on the phenotype
of ascending aortic aneurysm (along with ectopia lentis) (74). All patients
with a diagnosis of Marfan syndrome should be referred to a pediatric
cardiologist. While the mainstay of treatment for ascending aborting
aneurysm has traditionally been surgical repair, recent data has shown a
benefit for angiotensin receptor blockers (ARBs) as medical therapy for
slowing the rate of aneurysmal dilatation (75). There are increasing data that
the use of ARBs, even in young children <5 years of age with Marfan
syndrome, can slow the rate of progressive aortic dilatation and reduce the
risk of aneurysm formation later in life (76). This underscores the importance
of early diagnosis of Marfan syndrome in young children, because this
therapy can be initiated at an earlier stage (77). Diagnosis at an early age can
be difficult based on the examination alone. Genetic testing is considered for
patients in whom there is clinical suspicion based on exam findings or family
history to allow for early diagnosis and potential treatment.
Pediatricians can also be alerted to several cardinal features of Stickler
syndrome that can aid in the diagnosis. The musculoskeletal features of this
condition may be most easily seen in clinic—scoliosis, spondylolisthesis, or
Scheuermann-like kyphotic deformity, as well as femoral head failure
resulting from slipped epiphysis, along with radiographic presence of
osteoarthritis (before age 40) (78,79). Furthermore, pediatricians can be
alerted to orofacial features, including cleft palate, malar hypoplasia, broad or
flat nasal bridge, and micro-/retrognathia) (Figure 37-1). Sensorineural or
conductive hearing loss should arouse suspicion of Stickler syndrome when
in combination with the above findings. Also, in patients who are suspected
of having Stickler syndrome, hearing should be tested, because the
combination of hearing and vision impairment can be devastating to
childhood interaction, activities, and development (80). Referrals to pediatric
otolaryngology, orthopedics, genetics, and audiology should be strongly
considered in patients with Stickler syndrome. In addition, airway
management during mask ventilation and endotracheal intubation can be
challenging in Stickler patients, specifically those with mandibular
hypoplasia, and the surgeon should ensure that appropriate preoperative
evaluation is performed beforehand and that anesthesiology is aware and able
to manage the airway for surgery (81). Close interdisciplinary care and
coordination is critical between these specialties and ophthalmology.
Genetic testing for syndromic hereditary vitreoretinopathies causing myopia
continues to unlock additional information about the pathophysiology and
potential treatments of these conditions. Family members can be alerted of
important screening for sight-threatening conditions, and both the patient and
family can receive counseling on the need to evaluate for other systemic
associations with the condition based on the specific gene testing result (see
also Chapter 21). As discussed previously, the effects of each gene can vary
greatly even within the same condition. Proper training of physicians and
ancillary care personnel is becoming essential, along with referral to genetic
counseling to help interpret testing results specifically for the patient and
family and gauge their understanding of the results to guide further
communication (82).
Direct-to-consumer commercial genetic testing has become increasingly
available and accessible to the lay public over the last 5 years and in some
cases promises patients information about their susceptibility to an increasing
array of ocular conditions. The American Academy of Ophthalmology
continues to recommend against the inclusion of such genes in these kits, as
the ramifications of specific results may require expertise in interpretation for
each individual patient (83). With the rapid advances in this field, the impetus
is on ophthalmologists to be prepared to guide patients to appropriate avenues
for genetic testing and counseling, including those with the above conditions
(see also Chapter 21).
OTHER TREATMENTS
Atropine has been discussed above as a treatment for myopia, specifically to
reduce myopic progression. Given its widespread use in East Asian countries,
an interesting question is whether treatment with dilute atropine in premyopic
children can prevent the onset of myopia. A retrospective cohort study from
Taiwan examined the use of 0.025% atropine versus placebo in children 6 to
12 years of age and found a clinically and statistically significant reduction in
onset of myopia at 1 year of follow-up (21% vs 54%, p = 0.0002) (84). A
current phase 3 randomized controlled trial is enrolling patients to study the
effect of atropine 0.01% for prevention of myopia (85). The results should
provide important data as to a potential way to help slow the onslaught of this
public health epidemic.
Scleral collagen cross-linking is another promising treatment for myopic
progression. Animal studies in rabbit sclera have elucidated important
findings about the biomechanics of this condition and treatment (86).
Additional studies have shown reduced expression of a cross-linking enzyme,
lysyl oxidase (LOX), in guinea pig sclera with form deprivation myopia,
presenting a possible pharmacologic target for future cross-linking
interventions (87).
Figure 37-1 Orofacial findings in Stickler syndrome.
Prominent features include flat nasal bridge, large myopic
eyes (blocked for anonymity), upturned nasal tip, and
small mandible (micrognathia). (Photos courtesy G. Baker
Hubbard III, MD; Emory Eye Center, Atlanta, GA.)
Figure 37-2 Right eye of patient with Stickler syndrome
treated with prophylactic laser. (Photo courtesy G. Baker
Figure 37-3 Posterior retinal break causing
rhegmatogenous RD in a patient with Stickler syndrome.
Prophylactic laser barricade in the periphery would not
have prevented this detachment due to the posterior
location of the retinal break. (Photo courtesy G. Baker
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SECTION V
Pathophysiology In Retinal Diseases
38
Mechanisms of VEGF Signaling in
Developmental and Pathologic
Angiogenesis Related to Retinopathy
of Prematurity
M. Elizabeth Hartnett, Haibo Wang, Aaron B. Simmons, Colin A.
Bretz, Eric Kunz, and Aniket Ramshekar
ROP EVOLUTION AND DIFFERENCES

WORLDWIDE AND RELATIONSHIP TO
OXYGEN
High oxygen at birth damages newly developed retinal vessels and was found
to be a major contributor to retinopathy of prematurity (ROP) when it was
first described by Terry as retrolental fibroplasia (RLF) in the United States
in 1942 (1,2). At that time, incubators were much less sophisticated than
today, and there was little understanding of the effects of high oxygen on
preterm infant retinal vascular development. Therefore, infants ended up
being exposed to constant high oxygen to support overall survival. With
advancements in neonatal care and the ability to monitor and regulate oxygen
levels to infants, RLF nearly disappeared, but as preterm infants of younger
gestational ages and lower birth weights are surviving, ROP has reemerged
(3). In countries that have resources to successfully regulate and monitor
oxygenation of infants, the risk of ROP has shifted from affecting larger and
older premature infants to those who are extremely premature, including
infants born with extremely low birth weight (ELBW < 1,000 g) or at
extremely low gestational age (ELGAN < 28 weeks GA). However, in
emerging countries that do not have resources to regulate oxygen delivery,
ROP occurs in infants of older gestational age and larger birth weights (4,5).
As smaller and less developmentally mature premature infants have
survived, it has also become increasingly recognized that stresses besides
high oxygen at birth are involved in the pathophysiology of ROP (6,7).
Oxygen regulation, prenatal care, and nutrition remain important challenges
to overcome in all settings, but other risks have been identified and include
fluctuations in oxygenation (8–11), including intermittent hypoxemic
episodes (12), nutritional concerns, oxidative stress leading to dysregulated
signaling pathways that affect angiogenesis (13) (see also Chapter 41), and
potentially light-induced effects (14) including to the fetus and after birth that
involve and trigger effects in opsin-specific retinal ganglion cells (15,16).
Clinical trials that tested increased (17) or decreased oxygen saturation
targets (18–21) have not led to universally accepted guidelines regarding
oxygen saturation (see also Chapters 40 and 52), because lower saturation
targets have led to increased mortality in some units (18,22). Biphasic oxygen
saturation targets that change the oxygen saturation target based on the
postgestational age, which is the sum of gestational and chronologic ages,
have been proposed and studied (23–27).
Therefore, today, there are differences in ROP throughout the world and
are related to oxygen regulation, nutrition, and resources to provide
optimized prenatal and perinatal care. In countries that have recently
developed methods to save premature infants but have limited resources to
provide optimal prenatal and perinatal care, ROP is seen in larger and older
infants (4) (see Chapter 50).
Phenotype and Treatment of ROP
Variability in Training
As a result of different resources worldwide, ROP is seen in larger and more
developmentally mature infants in countries with limited resources compared
to countries with adequate resources and trained ophthalmologists to
accurately screen, diagnose, and classify ROP by indirect ophthalmoscopy or
using a contact camera (see Chapters 20, 50, and 51). Indirect delivery of
laser also requires skill and may not be delivered at a sufficient spot density
or location in the retina. Follow-up of infants to determine additional
treatment or surgical intervention requires skill.
Variability in Prematurity and Safety of Anti-VEGF

If anti-VEGF is given, care must be taken on the preparation of the drug and
the accuracy of the dose or volume delivered. If anti-VEGF is delivered into
the vitreous of a larger infant, it is likely more dilute as it enters a greater
blood volume than that in a growth-restricted or extremely preterm infant.
Infants born at older gestational ages are also more developmentally mature,
so the effects of anti-VEGF on the development of systemic organs could
differ from that in younger extremely premature infants (28).
Genotypic Variability
Some infants carry mutations of genes, such as in genes that regulate the
Norrie disease (NDP)/frizzled-4 (FZD4) pathway (29–32). Those infants
manifest severe ROP despite being of older gestational age or larger birth
weight than otherwise would be considered at risk (30). Other infants may
survive extreme prematurity by not having these genotypes but lack
protective mechanisms to reduce risk of ROP (33), and the predispositions
might be picked up only on genetic testing that may not be present
universally. For example, in a candidate gene study from 1,000 extremely
low–birthweight infants, intronic variants in the gene encoding brain-derived
neurotrophic factor (BDNF) were associated with severe (treatment-
warranted) ROP, and previous studies showed low BDNF in the bloodstream
in association with more severe ROP (33,34). Therefore, the genotype–
phenotype relationship complicates the picture.
Diagnostic Variability
The decision to treat ROP is based on clinical trials that used different
classifications of severe or treatment-warranted ROP. Two clinical trials
defined “treatment-warranted” ROP differently from one another (35)
(ClinicalTrials.gov), and both definitions differed from type 1 ROP tested in
another clinical study (36) or threshold ROP that was the definition used at
the time of the CRYO-ROP study (see Chapter 52). These concerns reduce
the ability to relate studies that use different treatment modalities or are from
different regions of the world to one another. Of course, clinical trials are
needed, but because of differences in infant genotype and phenotype, ability
to diagnose treatment-warranted ROP, and effect of treatment, the diagnosis
of what constitutes treatment-warranted ROP makes comparing clinical trials
from different regions of the world challenging.
Oxygen and Angiogenesis

Retinal vascular development begins by vasculogenesis, which is de novo
development of blood vessels from precursors, such as angioblasts or
astrocytes depending on species, and ensues through a process of
angiogenesis, which involves budding from existing blood vessels to extend
vascularization (37). This process has been shown in several species and is
believed to occur in human (see also Chapter 5). Angiogenesis is physiologic
when it occurs in the developing retina or can be pathologic, such as when it
occurs in stage 3 ROP with intravitreal neovascularization (IVNV).
Physiologic and pathologic angiogenesis is often prompted by hypoxia-
induced transcription factors that are stabilized in low oxygen and transcribe
a number of angiogenic factors, most notably vascular endothelial growth
factor (VEGF) (see Chapter 43), but also other growth factors. VEGF is
important for physiologic retinal vascular development (PRVD) (38) but,
when overactive signaling occurs through its angiogenic receptor, leads to
disoriented divisions of endothelial cells (39) and aberrant angiogenesis as
IVNV (40,41).
VEGF/Oxygen in ROP
VEGF was first described as vasopermeability factor (42). Numerous
investigators have studied VEGF related to adult diseases. When VEGF was
recognized as an important factor in adult pathologic retinopathies, like
diabetic retinopathy and retinal vein occlusion, and in neovascular age-related
macular degeneration, preclinical studies used animal models of oxygen-
induced retinopathy to test the effect of anti-VEGF agents (43–50). These
models share some features of human ROP. Therefore, it was logical that
VEGF would also be involved in pathologic IVNV in ROP. However, VEGF
is important in physiologic angiogenesis in retinal vascular development (38).
Therefore, there was a concern that inhibiting VEGF in the premature infant
eye, in which retinal blood vessel development was ongoing, might not only
reduce pathologic IVNV but also interfere with normal retinal vascular
development. This set of events could set up a scenario in which persistent
avascular retina would lead to recurrent IVNV and also reduce visual field.
Therefore, our lab was interested in extending PRVD in an effort to reduce
the stimulus for recurrent IVNV in treatment-warranted ROP. We focused on
the question why blood vessels grew into the vitreous instead of into the
retina proposed to be hypoxic in the premature infant eye once the infant was
removed from high supplemental oxygen.
When ROP was first described in the 1940s and 1950s, premature infants
were about 2 months premature and exposed to constant high oxygen but
may have had more peripheral retinal vascular development than extremely
premature infants, who are often 4 or more months premature when they
develop ROP today. Therefore, the original two-phase hypothesis, posed by
Ashton in the 1950s, has changed. Ashton posited that high oxygen damaged
newly formed retinal capillaries (termed, “vaso-obliteration” in phase I),
thereby creating broad areas of avascular retina. When the infant was
removed from high supplemental oxygen and placed into ambient air, the
avascular retina became hypoxic, which stimulated angiogenesis that led to
blood vessels growing into the vitreous rather than into the retina (termed,
“vasoproliferation” in phase II). Today, phase I not only includes the oxygen
damage to newly formed vasculature, or “compromised physiologic
vascularity,” but also includes a delay in PRVD, proposed to occur as a result
of fluctuations in oxygenation that affect VEGF expression (see also
Chapters 40 and 43) and potentially through oxidative damage or signaling
(51) (see also Chapter 41). Phase II involves vasoproliferation into the
vitreous as IVNV and plus disease, which is characterized by increased
tortuosity and dilation of retina arterioles and veins (see Chapter 52) (Figure
38-1). VEGF is an important angiogenic factor in ROP, but other factors may
also play a role, including the angiopoietins and erythropoietins. In addition,
other risk factors besides high oxygen at birth have been identified, including
poor nutrition and growth (see Chapter 39) and activation of oxidative
signaling mechanisms (see Chapter 41).
Figure 38-1 The rat OIR model represents features of
human ROP. A:Superior view of early ROP demonstrating
avascular retina and ridge (yellow arrow). B:Temporal
view of vascular ROP demonstrating neovascular tufts
showing stage 3 ROP (white arrow) and thickened
demarcation line (yellow arrow). C:Postnatal day 14 rat
retinal flatmount with delayed PRVD (white arrow) and
compromised physiologic vascularity with capillary
crossings (yellow box). D:Postnatal day 20 rat retinal
flatmount with intravitreal neovascularization (white
arrow) and peripheral avascular retina. OIR, oxygen-
induced retinopathy; P14, postnatal day 14; P20, postnatal
day 20; PRVD, physiologic retinal vascular development;
ROP, retinopathy of prematurity.
CLINICAL EFFECTS AND CONCERNS

OF ANTI-VEGF IN ROP
In clinical retinovascular disease, the current methods to inhibit aberrant
VEGF signaling in the eye are the use of intravitreal neutralizing full length
(bevacizumab) or fragments (ranibizumab) of antibodies or fusion proteins
(aflibercept) to bind the VEGF protein, so less bioactive VEGF is available to
activate its receptors to trigger signaling cascades that lead to biologic events,
including angiogenesis (35,52–54).
VEGF Binding Receptors

VEGF binds a number of receptors and coreceptors that are activated through
tyrosine kinases to initiate signaling cascades and lead to biologic events
(Figure 38-2). VEGF is mostly secreted but has forms that associate with the
extracellular matrix as part of basement membranes of cells within the retina
(Müller cells, astrocytes, RPE, photoreceptors) (55). Once VEGF is bound to
an anti-VEGF drug, access to receptors on endothelial cells is reduced as well
as on receptors of other retinal cells, including neurons and glia. As a result,
angiogenesis is affected, but VEGF’s beneficial effects on neural and glial
retina are also reduced. VEGF has been found to be neuroprotective,
including in the retina (56). This raises the concern that anti-VEGF agents
used in ROP may be detrimental to the developing retina. Anti-VEGF agents
are also able to access the blood circulation and may affect the developing
organs (28) (see also Chapter 53).
Figure 38-2 Graphical depiction of VEGF signaling and
interventions in clinical and experimental ROP. A: Under
physiologic conditions, VEGF/VEGFR2 signaling
activates STAT3 to cause a biologic response in the
experimental ROP model. Our working hypothesis is that
overactivation of the VEGF/VEGFR2 signaling pathway
leads to IVNV and delayed PRVD in ROP. B: Clinical
intervention using anti-VEGF agents limits the
bioavailability of VEGF, which reduces activation of
VEGFR2 signaling and subsequent IVNV; however, these
agents may also cause side effects since VEGF is
important for normal vascular development and
neuroprotection. C: Experimental regulation of
VEGF/VEGFR2 signaling reduces pathologic features of
ROP (see Table 38-1 for details). IVNV, intravitreal
neovascularization; PRVD, physiologic retinal vascular
development; shRNA, short hairpin RNA; STAT3, signal
transducer and activator of transcription 3; VEGF,
vascular endothelial growth factor; VEGFR2, VEGF
receptor 2. (Image created with BioRender.)
A difficulty related to the use of anti-VEGF is how to determine an effective

and safe dose to the individual infant eye. It is unknown what the
concentration of VEGF is during normal retinal development in the
premature infant vitreous or how much VEGF is in the vitreous at any
pathophysiologic step of severe ROP in the individual infant eye. ROP
severity can differ even with the same diagnosis of “treatment-warranted
ROP.” Therefore, it is unknown how much VEGF needs to be neutralized.
Also, doses are based on adult doses that are diluted, and this process may
lead to errors in concentration. The infant eye is about one-quarter the size of
the adult eye, so volumes delivered are reduced compared to adult volumes.
This means that even a small error in volume or concentration could
potentially have a larger effect in the infant compared to the adult eye.
Various syringes currently used to inject small volumes of agents into the eye
can add variability to the anti-VEGF dose received into the vitreous. In
addition, there is risk from long needles that are safe in adults but potentially
dangerous in the small infant eyes. Safer methods have been recommended
(57), but there remains a need for better methods to accurately deliver small
reproducible doses in preterm infant eyes (see also Chapter 53).
Safety of Anti-VEGF
The anti-VEGF agent, bevacizumab, has been used at doses of 1 to 10 mg/kg
as an intravenous drug for cancers. Known adverse events include impaired
wound healing, thromboembolism, proteinuria, and gastrointestinal
perforation (58). The concentration of bevacizumab in the eye is much lower
(52), but the drug accesses the systemic circulation and is potentially less
diluted in the small preterm infant blood volume compared to that of the adult
(59). Lower doses are being evaluated with reported efficacy for treating
severe ROP (36). Measured levels of VEGF in the bloodstream are reduced
in premature infants for at least 2 months after a single intravitreal injection
of bevacizumab or aflibercept (60), but VEGF levels in the blood are less
affected with ranibizumab (61). The effect of intravitreal anti-VEGF is also
noted in adult eyes (54) with certain diseases without systemic effects noted.
Nonetheless, the duration and effect of anti-VEGF in the developing infant
raise concerns and are being studied (see also Chapter 53).
VEGF has been reported as essential for the development of the kidney,
lung, brain, and other organs (28), and inhibition of VEGF has been
associated with damage to organs. Experimental studies reported pulmonary
hypertension in a rat model following an injection of an anti-VEGF agent
(62). Also, renal VEGF was shown to be regulated by changes between high
and low oxygen in the rat (63). Hypoxia can increase the expression of VEGF
and be pathologic to retinal vessels, but potentially protective in various
oxygen stresses. Some clinical studies report impairment of neurocognitive
development, but there is controversy, and many studies have had limitations
(64,65). However, a recent report from the Neonatal Research Network in the
United States again raises concerns (66). Therefore, the effect on premature
infants treated with anti-VEGF agents raises concerns about the development
of systemic organs (see Chapter 53).
In order to study the molecular mechanisms involved in the
pathophysiology of ROP and potential treatments on development
systemically, experiments are performed using models of oxygen-induced
retinopathy (OIR) most representative of human ROP.
STUDYING THE PATHOPHYSIOLOGY

OF ROP
Studying the pathophysiology and molecular mechanisms involved in ROP is
done through interventions in animal models of OIR that are reproducible
with much less variability than what occurs in preterm infants. To incorporate
both high oxygen and fluctuations in oxygenation, we adapted a rat model of
OIR (11), which This model employs oxygen cycling that causes pup arterial
oxygen levels to be similar in extremes to the transcutaneous oxygen levels
of preterm infants with severe ROP (8–11) and reduces postnatal growth
(11,67). Both growth restriction and the oxygen profile created in the model
reflect conditions in premature infants who develop treatment-warranted
ROP. The rat OIR model reproduces the initial compromised vascularity
from oxygen stress and delayed PRVD in phase I followed by
vasoproliferation as IVNV in phase II; both are appearances similar to human
severe ROP (7) (Figure 38-1). Retinal features can be quantified and
analyzed and include areas of peripheral avascular retina as a measure of
delayed PRVD, area of capillary crossings for compromise of physiologic
vascularity in phase I (68), and area of IVNV and tortuosity for plus disease
in phase II (Figure 38-1).
VEGF and its signaling have been studied in relation to diseases. Many
investigators have paved the way and studied ROP (1,8,9,11,69–71),
retinopathy, and VEGF signaling related to human disease (45,72–74), but
fewer have studied mechanisms using a representative model of human ROP,
such as the beagle (75–77) or the rat OIR model (78,79). In part, this was
because there was not a way to measure molecular or signaling mechanisms
in specific cells very easily. Mouse models of OIR have been used by others
to demonstrate the effect of anti-VEGF antibodies on angiogenesis and high
oxygen-induced effects on the retinal vasculature (69,80). Mice can be
genetically manipulated, but the model has less similarity to human stresses
leading to ROP than the rat OIR model (see Chapter 40). The VEGF
signaling pathway is challenging to study since a single-allele knockout of
VEGF or a receptor is lethal in mice (49,81,82). Our laboratory has used a
representative model of human ROP (11) and developed methods to
understand the specific effects of broad inhibition of VEGF on certain cells in
the retina (41,83–85). We focused not only on IVNV but also on PRVD and
on effects in neural and glial cells of the retina. To do this, we developed a
method to specifically knock down, but not totally inhibit, genes in specific
cells of the rat retina.
Studies on Broad Anti-VEGF Effects in Retinal

Vasculature With Intravitreal Inhibitors
From early experimental studies, we surprisingly found that inhibiting VEGF
either with a neutralizing intravitreal antibody to the ligand, VEGF164
(henceforth referred to as anti-VEGF), or with an intravitreal receptor
tyrosine kinase inhibitor reducing signaling through the angiogenic receptor,
VEGF receptor 2 (VEGFR2), reduced IVNV, as anticipated, but did not
reduce PRVD in the rat model of ROP (86,87). This was surprising since
neutralizing VEGF, an angiogenic factor, would be predicted to inhibit both
physiologic and pathologic angiogenesis. However, compared to control rat
IgG, an effective dose of neutralizing anti-rat VEGF antibody led to recurrent
IVNV about 7 days later in association with the activation of angiogenic
pathways in the retina (88).
Studying the VEGF signaling pathway is difficult. A single-allele
knockout of VEGF or a receptor is lethal in mice. To study angiogenic
signaling of VEGF through its main angiogenic receptor, VEGFR2, we used
an embryonic stem cell model in which VEGFR1 was knocked out in
endothelial cells using a cell-specific platelet endothelial cell adhesion
molecule (PECAM) promoter that also drove the expression of a green
fluorescent protein tag to identify endothelial cells. This caused most of the
VEGF produced to bind and trigger signaling through VEGFR2 and not
VEGFR1. In development, VEGFR1 can act as a decoy receptor and is
believed to regulate the amount of VEGF available to trigger signaling
through VEGFR2. Overactive VEGFR2 signaling led to a pattern of
endothelial cell growth that was disordered due to disoriented cleavage planes
of dividing endothelial cells. The pattern was reminiscent of stage 3 ROP and
was rescued by a transgene that replaced VEGFR1 to reduce VEGF/VEGFR2
signaling (39). This finding suggested that VEGF signaling through VEGFR2
interfered with normal angiogenesis necessary to extend physiologic
vascularization and also allowed for disordered growth as IVNV. Therefore,
regulation of VEGFR2 signaling was predicted to order growth and thereby
inhibit IVNV as well as extend PRVD (Figure 38-2).
Optimal Anti-VEGF Dose Inhibits IVNV but Thins

Neural Retina
It is not possible to specifically inhibit VEGFR2 signaling in endothelial cells
in the human infant retina to date, so our initial studies focused on creating an
“optimal anti-VEGF” dose in the rat model to address the problem of
variability of drug concentrations in the vitreous in human infant studies. We
identified the expression of VEGF and VEGFR2, but not VEGFR1, as being
elevated in the rat OIR model compared to room air control rat pups of the
same developmental ages at day 14 (phase I) when peripheral avascular retina
was present and day 18 (phase II) when IVNV occurred (89). Using in situ
hybridization, we found several cell types, including cells in the inner nuclear
layer, Müller cells, RPE, and photoreceptors that express VEGF mRNA
splice variants (85). Since Müller cells span the retinal layers where inner and
deep plexuses of retinal vessels exist, we sought to downregulate, but not
abolish, the expression of VEGF in these cells. We developed a lentiviral
gene therapy delivery method to introduce short hairpin RNAs to knockdown
VEGFA or the splice variant, VEGF164 mRNA, and specifically targeted
Müller cell expression using a cell-specific promoter CD44 with a green
fluorescent protein tag. We cloned effective shRNAs to VEGFA or
VEGF164 into plasmids embedded within a microRNA 30 context and
created lentiviral vectors. Subretinal injections of lentiviral vectors with
VEGFA shRNA, VEGF164 shRNA, or luciferase shRNA control were
delivered into 8-day-old rat pups in the rat OIR model. The lentiviral gene
therapy approach reduced retinal VEGF protein in the VEGFA shRNA–
treated pups significantly compared to luciferase shRNA–treated pups and
brought it to the level of room air–raised rat pups of the same developmental
ages. By reducing the expression of VEGF from Müller cells, we effectively
reduced the amount of VEGF secreted into the vitreous and defined this
reduction in retinal VEGF as creating an “optimal anti-VEGF dose.”
Using this method, we found reduced IVNV, stable peripheral avascular
retina, and no reduction in the physiologic retinal vascularity as measured by
pixels of fluorescence compared to control lentiviral vectors that delivered
shRNAs to luciferase. Furthermore, there was no recurrent IVNV in contrast
to an intravitreal anti-VEGF antibody (83) (Table 38-1).
Table 38-1 Summary of outcomes when regulating

VEGF signaling in the rat OIR model
Anti-VEGF and VEGFR2 inhibitors were delivered by intravitreal injection at P12. Lentiviral vectors
(MC-VEGFAshRNA, MC-VEGF164shRNA, EC-VEGFR2shRNA, and EC-STAT3shRNA) were
delivered by subretinal injection at P8. All comparisons are relative to control.
aAnti-VEGF agents caused recurrent IVNV and reduced PRVD at P25.
—, no change; ↓, reduced; ↑, increased; EC, endothelial cell; ERG, electroretinograms; IVNV,
intravitreal neovascularization; MC, Müller cell; N.D., not determined; PRVD, physiologic retinal
vascular development; shRNA, short hairpin RNA; VEGF, vascular endothelial growth factor;
VEGFR2, VEGF receptor 2.
To evaluate whether a neutralizing antibody to VEGF affected the avascular

retina and caused a compensatory increase in angiogenic signaling, we
compared the anti-VEGF intravitreal antibody to its intravitreal IgG control
with the “optimal anti-VEGF dose” created by a subretinal lentiviral vector
targeting Müller cell–expressed VEGFA compared to its subretinal lentiviral
vector targeting luciferase. We measured the peripheral avascular retina as a
marker of PRVD and the pixels of fluorescence within vascularized retina as
a measure of compromised physiologic vascularity. We found no difference
in the peripheral avascular retinal areas but found a significant reduction in
pixels of fluorescence in anti-VEGF antibody-treated eyes compared to the
IgG control. In contrast, there was no decrease in pixels of fluorescence in the
VEGFA shRNA–treated eyes compared to its control luciferase shRNA (90).
These data supported the idea that the optimal anti-VEGF dose was less
likely to lead to recurrent IVNV and that damaged vascularized retina that led
to capillary non-perfusion was involved in recurrent IVNV. However, it also
brings out the concern that too high a dose of anti-VEGF or anti-VEGF
treatment before severe ROP develops may harm the existing vasculature and
potentially lead to later IVNV (Table 38-1).
Even with the “optimal anti-VEGF dose,” thinning of the outer nuclear
occurred with the Müller cell-specific shRNAs to VEGFA compared to
control. Surprisingly, ERG a- and b-wave amplitudes were no worse and
sometimes better than the control lentivirus (Table 38-1). Compared to
control lentivirus or the VEGF164 shRNA lentivirus, there was increased
mRNA expression of neuroprotective factors in the retina, specifically
erythropoietin, BDNF, nerve growth factor, glial cell–derived neurotrophic
factor (GDNF), and neurotrophin 3 in the VEGFA shRNA–treated eyes.
Knockdown of VEGFA in cultured Müller cells confirmed increased
expression of BDNF, EPO, and NT-3 mRNA. From these findings, it is
surmised that knockdown of the full length of VEGFA reduced
neuroprotection from VEGFA that was important to the developing and
oxygen-stressed retina and that other neuroprotective factors from Müller and
retinal cells were expressed. Furthermore, even when VEGF164 was knocked
down, the peripheral avascular retina persisted without extension of PRVD
(83), and there was reduced outer nuclear and photoreceptor thickness
compared to luciferase shRNA control. However, there was no difference in
the thickness of the ONL in VEGF164 shRNA–treated compared to
luciferase control, whereas outer nuclear layer (ONL) thickness was
significantly reduced in the VEGFA shRNA compared to control (Table 38-
1).
VEGF has been reported as neuroprotective in the retina (56). Even with
the ideal anti-VEGF dose, the method is not specific to the endothelial cells
or to the angiogenic receptors of VEGF. These findings suggest that a single
dose of anti-VEGF could potentially be harmful to existing, newly developed
capillaries and lead to recurrent disease and that even the “optimal
physiologic dose” may affect the neural retina. Although knockdown of
VEGF164 did not increase neuroprotective factors or thin the outer retina, it
also did not lead to extension of PRVD.
We, therefore, determined whether regulating VEGFR2 signaling
specifically in retinal endothelial cells would safely reduce IVNV and extend
PRVD. Although this is not something possible in the human infant, it was
important to assess in order to enhance understanding of the pathophysiology
and safety of anti-VEGF agents being tested in human infants with ROP.
Toward Targeted VEGFR2 Inhibition in Endothelial

Cells
As we previously reported, overactivation of VEGF/VEGFR2 signaling by
knocking out VEGF receptor 1 (VEGFR1) in an embryonic stem cell model
led to disordered angiogenesis (39) similar in appearance to IVNV seen in
stage 3 ROP (Figure 38-1). Addition of the VEGFR1 plasmid “trapped”
excessive VEGF and allowed ordered angiogenesis of dividing endothelial
cells (39). We wished to test if knockdown of VEGFR2 specifically in retinal
endothelial cells would order physiologic angiogenesis and reduce pathologic
angiogenesis in the rat model of ROP. We used the lentiviral gene therapy
approach to target VEGFR2 specifically in retinal endothelial cells with a
Cdh5 promoter. shRNAs were created to knock down VEGFR2 and its
activation, and a downstream signaling protein of VEGFR2, STAT3, that we
had found was involved in IVNV (91). The control was Cdh5-driven shRNA
to luciferase (41) (Figure 38-2).
Knockdown of VEGFR2 specifically in endothelial cells inhibited IVNV
and extended PRVD retinal compared to control luciferase. Knockdown of
EPOR only extended PRVD, whereas knockdown of STAT3 only inhibited
IVNV. No shRNA caused thinning of the retina or reduced ERGs (41) (Table
38-1).
From these experiments, it appears that specific retinal endothelial cell—
VEGFR2 signaling—is important to regulate in ROP in order to inhibit
aberrant angiogenesis and extend normal retinal vascularization. We cannot
safely perform subretinal injections on premature infant eyes and are testing
alternate approaches to target retinal endothelial cells in pathologic
angiogenesis. It remains unknown if the extremely premature infant retina is
mature enough to support extended vascularization of the entire peripheral
retina. Future studies are needed.
SUMMARY
ROP has evolved from the time RLF was first described by Terry in the
United States in 1942 (2). However, throughout the world, ROP varies based
on the developmental age and size of the infants potentially because of
differences in oxygen regulation and monitoring, prenatal and perinatal
resources to deliver care, ability to diagnose ROP, agreement of what
constitutes “treatment-warranted ROP,” and potentially genetic or epigenetic
factors. Now, infants are less developmentally mature and of smaller size in
the United States and developed nations than in emerging nations. This may
have impact on treatments recommended to reduce the bioactivity of VEGF,
which is important to the developing infant and retina. Anti-VEGF agents
may affect neural development in the retina and CNS and when delivered into
the eye of an extremely premature infant be less diluted in the small blood
volume and have greater deleterious impact in the less developmentally
mature than in a larger and more developmentally mature infant. Anti-VEGF
agents delivered into the vitreous affect not only the VEGFR2 signaling
pathway in endothelial cells of the retina but also VEGFR2 and other VEGF
receptors in cells of the neural retina and glia. Experimentally, anti-VEGF at
too high a dose may lead to thinner retinas in oxygen-stressed eyes. Taken
together, these studies support the continued work toward better treatments
for ROP for neuroprotection and vascular health.
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39
Weight Gain and Retinopathy of
Prematurity
Ann Hellström, Lois E. H. Smith, and Anna-Lena Hård
INTRODUCTION
For developing children, a normal increase in weight is an indicator of health.
Extrauterine growth restriction is common among very preterm infants, and
subnormal weight gain, both before and after birth, has been identified as an
important risk factor for severe retinopathy of prematurity (ROP) (1). This
knowledge can be used to predict severe ROP and potentially to prevent ROP
through improvements in neonatal care that improve growth.
Low gestational age (GA) is the most important risk factor for ROP. Low
weight at birth has also been described as an important risk factor, but low
birth weight (BW) may simply reflect low GA if the BW is appropriate for
gestational age (AGA).
In this chapter, we will address
Poor prenatal weight gain as a risk factor for ROP

Poor postnatal weight gain as a risk factor for retinopathy in animal
studies
Poor postnatal weight gain as a risk factor for ROP
Factors contributing to poor weight gain in the very preterm infant
Possible future interventions aiming to improve growth and reduce ROP
POOR PRENATAL WEIGHT GAIN AS A

RISK FACTOR FOR ROP
When analyzing weight, standard deviation scores (SDSs) are often used as
SDS adjusts for sex and age. A low weight SDS indicates that the weight is
low in relationship to norms for age and sex, whereas a high weight SDS
indicates a weight that is high in relationship to age and sex norms. In
general, being born small for gestational age (SGA) (with a low BW SDS)
appears to positively correlate with the development of ROP in preterm
infants of older compared to younger preterm infants.
In a large cohort study of 2,521 infants from the United States, Canada,
and Sweden, a lower BW SDS was associated with ROP of any severity in
infants born at 25 to 30 weeks GA, especially in infants at the higher end of
the GA range (Figure 39-1) (2). Another large study of 2,941 infants born
<32 weeks GA from North America and Sweden also reported that a low BW
SDS was a less important risk factor for severe ROP needing treatment in
infants born <26 weeks GA than in infants born at older GA (Figure 39-2)
(3).
FIGURE 39-1 Difference in birth weight SD score
according to GA group between infants with and without
retinopathy of prematurity. Data are shown as mean and
95% CI. Analyses are adjusted for exact GA at birth, sex,
and center. (From Klevebro S, Lundgren P, Hammar U, et
al. Cohort study of growth patterns by gestational age in
preterm infants developing morbidity. BMJ Open
2016;6(11):e012872.
http://creativecommons.org/licenses/by/4.0/.)
FIGURE 39-2 Birth weight (BW) SDS odds ratio for

infants for ROP requiring treatment in relation to
immaturity. The BWSDS was calculated using a Swedish
or a Canadian growth chart reference for different GA
cutoffs. (From Lundgren P, Kistner A, Andersson EM, et
al. Low birth weight is a risk factor for severe retinopathy
of prematurity depending on gestational age. PLoS One
2014;9(10):e109460. Copyright © 2014 Lundgren et al.
https://creativecommons.org/licenses/by/4.0/.)
Most studies on the influence of a low BW (birth weight standard deviation

score [BWSDS]) on ROP risk do not compare BWSDS directly, but instead
investigate the effect on ROP risk of being born SGA. However, SGA and
low BWSDS are not the same. Definitions of SGA vary with respect to
growth standards used as well as definition of the range of BW included in
the definition of SGA. SGA is commonly defined as having a BW that is
either more than two SDs below the mean BW for GA corresponding
approximately to the second percentile [2.3 SD] or sometimes defined as
below the 10th percentile for GA, thus classifying heavier infants as being
SGA. Currently used growth standards are either based on weights recorded
at birth in large populations that include preterm births (4,5) or on weight
estimations based on ultrasound measurements during normal pregnancies
(6). Fetal curves based on ultrasound may be more appropriate than weights
recorded at birth, which include BWs from abnormal pregnancies resulting in
preterm delivery. When a neonatal (7) or a fetal (8) growth standard was used
on the same population of infants born at or <34 weeks gestation, the results
differed greatly. When the neonatal standard was used, 11.6% had a BW
below the 10th percentile and were classified as SGA, whereas 23.3% using
the fetal growth standard were classified as SGA, demonstrating the
substantial discrepancy found depending on the standard used (9) and the
difficulty comparing studies with varying standards.
The impact of a low BW SDS on risk of severe ROP appears to differ
with degree of immaturity. SGA at birth was not associated with increased
risk of severe ROP in extremely preterm infants born <27 weeks GA in two
studies (10,11). In accordance, Dhaliwal et al. (12) found no association
between ROP and SGA at birth in infants born ≤25 weeks GA. However,
more mature infants born at 26 to 31 weeks GA appeared to be more likely to
develop any stage of ROP and specifically more likely to develop stage 3 to 5
ROP if they were born SGA compared to AGA (12). Several larger studies
report a significantly increased risk of severe ROP (3,12–18) and some also
for any stage ROP (9,12,13) in infants born SGA (9,13). Darlow et al found a
“dose–response” relationship between lower BW for GA and risk of severe
ROP indicating that the more growth restricted the infant, the greater the risk
for severe ROP (3,16). Suggested causes for the lower impact of SGA at birth
in infants born at low GA include that there are small numbers of immature
growth-restricted infants due to decreased survival, and that there is a high
risk of ROP in infants of extremely low GA regardless of being SGA or AGA
at birth (3).
A recent study of 6,708 infants born <32 weeks GA from 717 U.S.
hospitals defined SGA as BW below the 10th percentile on an intrauterine
growth curve based on U.S. data (19). The risk increase for stage 3 to 5 ROP
associated with SGA at birth in surviving infants was significant only for
those born at or <25 weeks GA. The greatest excess risk for mortality and
stage 3 to 5 ROP was found among low or extremely low GA infants
confirming a higher mortality rate in these infants (20) (Figure 39-3).
FIGURE 39-3 Unadjusted rates and adjusted risk
differences for death and retinopathy of prematurity
among survivors and morbidity–mortality composite
outcomes stratified by gestational age week. The adjusted
risk difference values above zero quantify the excess
proportion of risk for the study outcome independently
associated with small for gestational age (SGA) birth
among SGA babies. Risk difference values are adjusted
for infant sex, race/ethnicity, insurance type, caesarean
birth, multiple gestation pregnancy, maternal initiation of
prenatal care during the first trimester, preeclampsia or
eclampsia, gestational diabetes, gestational hypertension,
treatment with antenatal corticosteroids, and rupture of
amniotic membranes ≥18 hours prior to delivery. A robust
variance estimator for cluster-correlated data was used in
all models to account for potential within-hospital
outcome correlation. ROP, retinopathy of prematurity; wk,
week. (Reprinted from Jensen EA, Foglia EE, Dysart KC,
et al. Adverse effects of small for gestational age differ by
gestational week among very preterm infants. Arch Dis
Child Fetal Neonatal Ed 2019;104:F192–F198. With
permission from BMJ Publishing Group Ltd.)
Thus, it appears that prenatal growth restriction is a risk factor for ROP at
least in some older GA groups. Differences in impact of SGA at birth on
ROP depends on different degrees of immaturity (GA) at birth may be due to
the use of different definitions of SGA and growth curves used in studies as
well as higher mortality in the most infants born at the lowest GA.
In 1953, Lubchenco et al. reported that retrolental fibroplasia, the end
stage of ROP, occurred in infants with low BW but also in infants with
greater weight loss during the neonatal period followed by a relatively fast
weight gain. This observation sheds light on the importance of postnatal
growth (21).
POOR POSTNATAL WEIGHT GAIN AS

A RISK FACTOR FOR RETINOPATHY
IN ANIMAL STUDIES
Animal studies have shown that poor postnatal weight gain is associated with
increased risk for oxygen-induced retinopathy (OIR). At birth, mice and rats
born with normal weight have incomplete retinal vascularization, similar to
premature human infants. Retinal vascularization continues to develop during
the first few postnatal weeks in rodents. To study the effect of poor postnatal
growth on ROP, OIR rodent models are used with the addition of extra pups
to one dam to reduce nutrition to individual pups, which cause poor weight
gain.
More than 50 years ago, the Swedish researcher Bo Hellström found that
starved mice with poor postnatal weight gain had poor retinal vascular
development. He studied the influence of nutrition on the development of
oxygen-induced retinal changes in mice, but first studied the effect of
starvation on retinal vascularization in pups raised in room air (22).
In starved animals in room air, the outgrowth of vessels of the nerve fiber
layer was slightly delayed. Vessels extended to the ora serrata in starved
animals at 15 days of age, whereas in normally fed animals, this occurred at
10 days of age. In addition, there were other developmental delays noted in
retinal development including in the separation of the nuclear layers, the
relative increase in the inner plexiform layer, the outgrowth of rods and
cones, the development of secondary vitreous, and the involution of hyaloid
vessels. In a later study of rat pups raised in room air, reduced nutrition
(accomplished by rearing large litters of pups) delayed vascularization of the
retina and impaired total retinal growth. The ratio of vascularized to total
retinal area was 54% in larger litters (18 pups/mother) compared to 67% in
smaller litters (10 pups/mother) (23). Thus, starvation per se appears to have
a negative impact on both retinal vascularization and development.
Bo Hellström (22) also found that exposure to 98% to 100% oxygen
almost completely inhibited retinal vessel growth regardless of nutritional
state of the pups. When mouse pups were transferred to room air, a
vasoproliferative stage occurred, which was almost completely suppressed if
starvation continued. Only 2 of 14 animals with continued starvation and
poor growth during the room air course developed pathologic
neovascularization compared with 12 of 12 animals that were starved only
during oxygen exposure. These findings support the notion that poor weight
gain in the first few weeks of life followed by later catch-up growth, as
commonly seen in preterm babies, is associated with increased risk of retinal
neovascularization.
Variable hyperoxia/hypoxia also appears to negatively influence retinal
neovascularization. In rat pups exposed to cyclic hyperoxia and hypoxia
followed by room air exposure, retinal neovascularization occurred in 53% of
rats in large litters (n = 25) versus 15% in smaller litters (n = 10). Total
retinal area as well as the ratio of vascularized to total retina was smaller in
rats raised in large litters. In addition, the median number of clock hours of
neovascularization was 3.5 in large litters and 2.0 in small litters. Smaller
ratios of vascularized to total retinal areas were associated with a greater
incidence and severity of abnormal neovascularization. This finding agrees
with observations that the size of the peripheral avascular zone determines
neovascularization in rats exposed to variable oxygen (21,24,25).
Acidosis in the neonatal period results in acidosis-induced retinopathy
(AIR), which has similar pathologic features as OIR models. AIR induces
more retinal neovascularization in undernourished than in normally fed rat
pups (26).
Poor postnatal weight gain in a mouse OIR model is also associated with
a delayed onset and prolonged course of neovascularization in association
with increased retinal vascular endothelial growth factor (VEGF) expression.
In addition, electroretinography responses were attenuated with significantly
reduced amplitudes corresponding to bipolar cell and inner retinal function
compared with pups having medium or extensive weight gain (27).
POOR POSTNATAL WEIGHT GAIN AS

A RISK FACTOR FOR RETINOPATHY
OF PREMATURITY IN CLINICAL
STUDIES
At very preterm birth, the supply of nutrients and growth factors provided by
the maternal/fetal interaction is suddenly lost, and general growth, as well as
vascular growth, slows down. In addition, it is likely that the relative
hyperoxia of the extrauterine environment together with supplemental
oxygen causes obliteration and regression of already formed vessels as seen
in animal studies (28). Poor postnatal growth in very preterm infants is a
major problem and results in reduced weight gain relative to age and sex
(29).
In an early paper, investigators reported that poor postnatal weight gain
was associated with increased ROP severity in four surviving quintuplets
with identical GAs and similar BWs (30). Other studies have found an
association between poor postnatal weight gain during the first 6 weeks of
life and severity of ROP (15,31,32). In infants with GA < 30 weeks, each 60
g below the expected weight at 6 weeks was associated with a 20% increase
in the risk of stage 3 ROP (31). In one study, poor weight gain during the first
6 weeks of life in infants born with BW ≤ 1,500 g and GA ≤ 32 weeks was
reported to be an important and independent risk factor for severe ROP (32).
Weight gain at 6 weeks that was <51.2% of BW was associated with a
threefold increase in risk of severe ROP. Very low BW infants are expected
to have regained weight, approximately 50% of their BW, after 6 weeks of
life (33). In the Extremely Preterm Infants Study in Sweden (EXPRESS), low
weight SDS at 36 weeks’ postmenstrual age (PMA) was highly associated (P
< 0.001) with ROP stage 3 or worse (10), suggesting that poor postnatal
weight gain up to 36 weeks’ PMA is a strong risk factor for ROP, whereas
appropriate postnatal weight gain through 36 weeks’ PMA is associated with
lower risk.
Insulin-like growth factor I (IGF-I) promotes anabolism and is essential
for growth during the third trimester and for normal vascularization of the
human retina (34). In a study of 84 infants born between 24 and 32 weeks
GA, low serum IGF-I values correlated both with the presence of ROP and
increased ROP severity (35). The mean ± SEM serum IGF-1 level during 30
to 33 weeks PMA was lowest (IGF-1 of 25 μg/L) in patients with severe
ROP, intermediate (29 μg/L) with moderate ROP, and highest (33 μg/L) with
no ROP. In addition, the duration of low IGF-I levels correlated with
increased severity of ROP. The age at which serum levels of IGF-I reached
33 μg/L was 52 days for severe ROP, 44 days for moderate ROP, and 23 days
for no ROP. Other prematurity-related morbidities, such as
bronchopulmonary dysplasia, intraventricular hemorrhage (IVH), and
necrotizing enterocolitis (NEC), also correlated with low IGF-I values.
Postnatal Weight Gain and Prediction of Severe ROP

The relationship between postnatal weight gain and ROP can be used very
early in the neonatal course to predict the risk for severe ROP. The value of
this prediction is that care can be concentrated on those infants more likely to
develop severe ROP, and fewer examinations performed on infants at low
risk. In Figure 39-4, postnatal weight development is presented as SDSs in
129 infants with no, mild, and proliferative ROP, respectively, from two
populations (36,37).
FIGURE 39-4 Postnatal weight development in 129
infants from two study populations. (From Löfqvist C,
Andersson E, Sigurdsson J, et al. Longitudinal postnatal
weight and insulin-like growth factor I measurements in
the prediction of retinopathy of prematurity. Arch
Ophthalmol 2006;124:1711–1718; Löfqvist C, Hansen-
Pupp I, Andersson E, et al. Validation of a new
retinopathy of prematurity screening method monitoring
longitudinal postnatal weight and insulin-like growth
factor I. Arch Ophthalmol 2009;127:622–627.) (From
Hellström A, Ley D, Hansen-Pupp I, et al. New insights
into the development of retinopathy of prematurity—
importance of early weight gain. Acta Paediatr
2010;99:502–508. Copyright © 2009 The
Author(s)/Journal Compilation © 2009 Foundation Acta
Pædiatrica. Reprinted by permission of John Wiley &
Sons, Inc.)
With current screening criteria based on GA and sometimes BW, only 5% to

10% of infants who undergo repeated painful and stressful eye examinations
develop severe ROP requiring treatment. Therefore, more efficient ways to
detect infants at risk are needed.
The finding that risk of severe ROP is increased in infants with low
serum levels of IGF-I, which is essential for growth during the third
trimester, suggested that early identification of infants at risk for severe ROP
might be possible using the variables IGF-I and weight gain. Hence, the
algorithm Weight IGF-I Neonatal ROP (WINROP) was constructed to detect
deviations of any child from the expected weight gain of those infants who
developed no ROP. Initially in the same way, deviations from the IGF-I level
were also used in WINROP using online statistical surveillance (36)
(www.winrop.com). Each week after birth, weight and IGF-I levels are
entered into the algorithm, and if the accumulated deviation reaches a certain
level, which is set to include all infants at risk of severe ROP, an alarm is
shown that indicates that the child is at high risk for proliferative ROP. The
algorithm is based on initial data from 79 Swedish infants born at GA < 32
weeks, 38 with no or mild ROP, and 13 with proliferative ROP. WINROP
was first validated in 50 infants born at GA < 31 weeks in Lund, Sweden. In
this cohort, WINROP correctly identified 9/9 infants who developed
proliferative ROP on average 9 weeks (1.1 to 21.6 weeks) later (100%
sensitivity) with a specificity of 54% (37).
Serial analyses of serum IGF-I are costly and not universally available.
Furthermore, weight gain also reflects IGF-1 levels. Therefore, studies were
performed to determine if predictions of severe ROP could be accurately
made without using IGF-1 concentrations but using weight gain only. In a
retrospective study, weekly weights of 353 infants were entered into
WINROP. Infants with hydrocephalus, which causes nonphysiologic weight
gain due to accumulation of fluid in the head, were excluded. Using the level
of accumulated deviation in weight gain that triggers an alarm before 33
weeks’ PMA to predict stage 3 ROP, the sensitivity of WINROP was 100%
and the specificity 84.5%. As in the initial WINROP study, the alarms came
well before the development of stage 3, severe ROP. For 35 infants who
developed proliferative ROP, the chronologic median age when there was an
indication of high risk for proliferative ROP was 3.1 weeks (range 2 to 10
weeks), and the median time from the alarm to diagnosis of stage 3 ROP was
7.7 weeks (range 1 to 16 weeks) (Figure 39-5). It was estimated that by using
WINROP, 76% of infants fulfilling screening criteria in this population
would not need any eye examinations (40).
FIGURE 39-5 Time after birth for alarm, first observed

proliferative ROP (diagnosis), and treatment for children
with proliferative ROP (n = 35).
(Reprinted with permission from Hellström A, Hård A-L, Engström E, et al. Early weight gain predicts
retinopathy in preterm infants: new, simple, efficient approach to screening. Pediatrics
2009;123:e638–e645. Copyright © 2009 by the American Academy of Pediatrics.)
WINROP has now been validated in many different populations in Europe,

America, and Asia with varying sensitivity in predicting stage 3 or more or
treatment-warranting ROP. The sensitivity and specificity in different
populations of infants are presented in Table 39-1 (39–59). Differences in
nutrition polices and oxygen control may influence the impact of different
risk factors and affect WINROP results. In addition, the predictive power of
WINROP is dependent on weekly weight inclusion and exclusion of infants
with GA < 23 weeks or ≥32 weeks as well as those with hydrocephalus who
have unphysiologic weight gain and high risk of needing ROP treatment.
Varying adherence to these prerequisites may contribute to the variability
between studies.
Table 39-1 The sensitivity and specificity of

WINROP (using weight only) in predicting severe
ROP in different populations of infants
WINROP, Weight IGF-I Neonatal ROP; ROP, retinopathy of prematurity.
At our clinic in Gothenburg, Sweden, more mature infants (GA ≥ 30 weeks)

without alarms are examined first at PMA 35 to 37 weeks and examined no
further if no ROP is found at that time. However, after an increase in oxygen
saturation targets from 88%–92% to 91%–95% starting in 2014–2015,
treatment-warranting ROP without preceding WINROP alarms became more
common. An analysis of infants who were now subjected to the higher
oxygen saturation targets revealed that poor postnatal weight gain was no
longer a significant risk factor for ROP needing treatment. There was an
increase in the proportion of infants treated for ROP from 12.6% in the earlier
time period with lower oxygen saturation targets to 17.6% in the later period
that did not reach significance (60). In the time period with lower oxygen
saturation targets, other models based on postnatal weight gain to predict
severe ROP have been developed and validated (61,62). None of these
models have been retested in the era of higher oxygen saturation targets. One
clinical prediction model including GA, BW, and rate of weight gain was
developed in a cohort of 451 infants born at BW < 1,000 g in the Premature
Infants in Need of Transfusion ROP (PINT-ROP) study (61). A nomogram
was developed for the actual GA, and a line drawn between values for BW
and daily weight gain calculated from weekly weight gain. This line crosses a
“probability line” in which the estimated risk is found at the intersection. The
model was run weekly and developed to identify all infants (n = 33) with
severe ROP (defined as stage 3 or treatment), and it correctly identified 66 of
a total of 67 with severe ROP in this cohort. The median time between the
alarm and diagnosis of severe ROP was 10.8 weeks (range 1.9 to 17.6). The
sensitivity for predicting severe ROP was 99% and the specificity 36%
(Figure 39-6).
FIGURE 39-6 Sample nomogram to determine risk of
ROP in infants with various GAs at birth. A straight line is
drawn between the values for BW and daily weight gain
rate. The intersection of this line with the gray auxiliary
axis is then connected to the value for GA. The
intersection of this second line with the probability line
provides the predicted probability of severe ROP. At a
predefined risk, in this case 0.085, eye examinations are
considered indicated.* Because of the small number of
infants with higher GAs in the cohort, the point for GA 28
weeks falls out of its expected sequence. (Reprinted with
permission from Binenbaum G, Ying GS, Quinn GE, et al.
A clinical prediction model to stratify retinopathy of
prematurity risk using postnatal weight gain. Pediatrics
2011;127(3):e607–e614. Copyright © 2011 by the
American Academy of Pediatrics.)
Again in the time period with lower oxygen saturation targets, further
evaluation and modifications were performed in the Children’s Hospital in
Philadelphia Retinopathy Prematurity model (CHOP-ROP) (63) and in the
Postnatal Growth and Retinopathy of Prematurity study (G-ROP), which
includes 7,483 infants from 29 hospitals in the United States and Canada
(64). The CHOP-ROP model was validated in the G-ROP cohort regarding
ability to predict type 1 ROP (sensitivity 98.5%, specificity 36.4%) and
potential to reduce the number of infants requiring eye examinations. If the
cut point for a risk that was warranting eye examination was lowered to
capture all type 1 ROP cases, the specificity was reduced to 7.8% (65).
Based on results of the G-ROP study, modified criteria for inclusion in
ROP screening were proposed. Infants meeting any of six criteria would
undergo eye examinations: a GA younger than 28 weeks; a BW of <1,051 g;
a weight gain of <120 g, 180 g, or 170 g during ages 10 to 19, 20 to 29, or 30
to 39 days, respectively; or hydrocephalus. These criteria have yet to be
validated in other cohorts and in infants subjected to higher oxygen saturation
targets (66). In the Colorado ROP model (CO-ROP), infants with GA at birth
of ≤30 weeks and BW of ≤1,500 g and a net weight gain of ≤650 g between
birth and 4 weeks of age are screened for ROP (67). Validation in cohorts
other than the original, on which the model was developed, has yielded high
but not 100% sensitivities and highly variable specificities in detecting ROP
needing treatment (68–70).
Another prediction method (ROP Score) developed in a single cohort is
based on BW, GA, proportional weight gain, use of oxygen in mechanical
ventilation, and need for blood transfusions from birth to 6 weeks of life (62).
Five percent of the infants developed severe ROP and were identified with
96% sensitivity and 56% specificity.
In a comparison between WINROP, ROP Score, and CHOP-ROP on an
Italian cohort of infants with GA ≤ 30 weeks and/or BW ≤ 1,500 g, ROP-
Score and CHOP-ROP identified all infants with type 1 ROP while WINROP
missed three cases (55). It is thus clear that there is a strong relationship
between poor early postnatal weight gain and later development of ROP. It is
possible to use this information to predict severe ROP at an early age, often
weeks to months before treatment is required. However, sensitivity has rarely
been 100% for predicting type 1 ROP. Thus, weight gain–based models
cannot be used to exclude infants from screening since a missed case is likely
to result in lifelong blindness. Instead, weight gain–based prediction can be
used in clinical practice as an adjunct to standard care to reduce traditional
screening in infants with no alarm and attend most to infants with an alarm.
This is important since current ROP screening requires repeated diagnostic
eye examinations mainly by specially trained ophthalmologists. There are too
few trained ophthalmologists able to perform these examinations adequately
throughout the world. The examinations and eye drops have potential adverse
effects on fragile preterm infants and are costly to the health care system, and
90% to 95% of infants examined do not need treatment. In addition, early
identification of infants with poor weight gain and increased risk of severe
ROP may lead to interventions to promote appropriate postnatal weight gain
and possibly prevent disease.
FACTORS CONTRIBUTING TO POOR

WEIGHT GAIN IN THE VERY
PRETERM BABY
Infants born very preterm (low GA) usually become growth restricted soon
after birth compared to GA-matched normal fetuses in utero, and they remain
growth restricted during the first 1 to 2 years of life (38,39,71,72). Infants
with major morbidities such as chronic lung disease, severe IVH, NEC, or
late-onset sepsis grow even more slowly than those without these
comorbidities (33). The etiology of the growth restriction is multifactorial,
and associated morbidities may be causes as well as results of insufficient
growth.
Increased Metabolic Rate

The metabolic rate in utero is less than the postnatal metabolic rate. In
mammals, small size is associated with a high metabolic rate, which is
thought to compensate for heat loss through a larger relative surface area.
Preterm infants adapting/adapted metabolically to the extrauterine
environment have increased calorie requirements exceeding 60 kcal/kg/d (73)
compared to in the in utero environment (24 to 38 kcal/kg/d) (74).
Inappropriate Nutrition
Nutrient supply from the mother is suddenly withdrawn at birth, and in very
preterm infants, the immaturity of the gastrointestinal tract prevents sufficient
enteral feeding. Therefore, immature infants at risk of ROP generally need
parenteral nutrition during the first weeks of life. However, solutions for
parenteral nutrition have been developed for adults, not for preterm babies.
Gut maturation normally starts during the third trimester when the fetus
swallows approximately 200- to 250-mL amniotic fluid/kg fetal weight each
day. Both amniotic fluid and breast milk contain nutrients and bioactive
factors that promote gut maturation, protect against infection, and facilitate
growth (75,76). The mother’s own milk is the recommended food for all
infants and especially those born preterm (77), and in current clinical
practice, small amounts of breast milk, enteral (so called trophic) feeds are
given early after birth to stimulate gut function. Providing maternal breast
milk after very preterm delivery is often challenging. Mothers are encouraged
to express milk for their infants but milk donated from other mothers or
preterm infant formula are often used as a complementary food or used
exclusively.
Maternal milk is generally consumed raw. Donor milk needs to be
pasteurized for destruction of pathogens, usually with Holder pasteurization,
which implies heating to 62.5°C for 30 minutes. Donor milk is mainly
produced after a period of lactation after term delivery and contains less
protein, fat, and energy than early milk from mothers who deliver preterm
(78). Some breast milk factors involved in infection defense, such as
secretory immunoglobulin-A, lactoferrin, and lysozyme, are reduced by
pasteurization. In addition, growth promoting factors such as bile salt
stimulated lipase, IGF-I, insulin, and transforming growth factor β are
reduced or inactivated by pasteurization, which also destroys living breast
milk cells such as leucocytes and stem cells with unknown consequences for
preterm infants (79). Neither maternal nor donor milk can fulfill the
nutritional needs of very preterm infants and both are fortified to fulfill
current nutritional recommendations.
Recent studies have revealed that donor milk and maternal milk are not
equivalent. Studies have shown a dose-dependent beneficial effect on early
growth with the use of fortified maternal milk compared to fortified
pasteurized donor milk (80–83) as well as better cognitive scores at 2 years of
age (82). Slower weight gain during the first postnatal month and lower
cognitive scores at 1 and 2 years of age were found in infants with BWs
<1,000 g if supplemented with more than 50% donor milk compared to
infants fed exclusively maternal milk (82). In another study, a dose–response
relationship between maternal milk and weight gain was found in infants with
GA 32 weeks or less or BW 1,800 g. The mean weight gain velocity before
36 weeks PMA or discharge from the neonatal intensive care unit decreased
by 0.17 g/kg/d and the mean head circumference velocity by 0.01 cm/week
for every 10% increase in donor milk intake compared to mother’s own milk
(83). Dose-dependent relationships between mother’s milk feeding during the
neonatal period and reduced frequency of ROP have been reported
(81,84,85).
Lack of Growth Factors and Essential Nutrients

(80–83)
IGF-I serves an important anabolic role during the third trimester, and fetal
serum concentrations of IGF-I are related to fetal size from 33 gestational
weeks. IGF-1 is increasingly produced by the fetus during the third trimester
and concentrations increase 2- to 3-fold from week 33 to term (86). After
preterm birth, IGF-1 levels fall dramatically. A strong association between
low postnatal serum IGF-I concentrations and ROP and other prematurity-
related morbidities has been found (35).
Lack of Essential Polyunsaturated Fatty Acids

Omega 3 and 6 long chain polyunsaturated fatty acids (LCPUFAs) are
insufficiently synthesized by the fetus and mother and are obtained through
diet or, in the case of the fetus, from the mother’s diet. During the third
trimester, there is a massive transfer of omega-3 and omega-6 LCPUFAs
from the mother to the fetus. Therefore, some of these essential fatty acids are
lacking in infants born preterm (87).
In retinal rod outer segments, the omega-3 LCPUFA, docosahexaenoic
acid (DHA), is found in higher concentrations than in any other part of the
body suggesting an essential role of DHA in retinal function. The omega-6
LCPUFA, arachidonic acid, is essential for infant growth and development
(88). Low serum concentrations of arachidonic acid have been strongly
associated with later ROP development (89). The association of arachidonic
acid and DHA levels in preterm infant with growth and health is under
investigation (www.clinicaltrials.gov #NCT03201588).
Oxygen Supplementation
The influence of oxygen treatment on general growth in preterm infants is
largely unknown. In rat pups, exposure to alternating hyperoxia and hypoxia,
which is common in preterm babies, leads to severe OIR in association with
low body weight and low systemic IGF-I levels (90). Very preterm (low GA)
infants are especially vulnerable to oxidative stress secondary to both
hypoxia and hyperoxia and also secondary to inflammation, parenteral
nutrition, and high levels of free iron in combination with inadequate
antioxidant protection (91–93).
Optimal oxygenation of very preterm (low GA) infants is still unknown.
Combined data from five studies with similar designs comparing higher (91%
to 95%) and lower (85% to 89%) oxygen saturation targets were analyzed in
the NeOProM study (Neonatal Oxygen Prospective Meta-analysis)
Collaboration. A lower oxygen saturation target was associated with
decreased frequency of ROP needing treatment. However, there was no
difference in weight at discharge between the two oxygen target groups (94).
Oxygen supplementation is a very strong risk factor for ROP, and the value
of postnatal weight gain in the prediction of severe ROP is influenced by
oxygen supplementation strategies (60).
POSSIBLE FUTURE INTERVENTIONS

AIMING TO IMPROVE POSTNATAL
GROWTH AND REDUCE ROP
Nutrition
An association between low intake of energy, carbohydrates, and fat but not
protein during the first weeks of life and later severe ROP was reported in
two studies of extremely preterm infants with GA < 28 weeks (95) and 27
weeks (96). This might indicate a possible preventive effect of increased
provision of these nutrients, but it may also reflect feeding intolerance
preventing more aggressive nutrition.
It is evident that maternal milk during the first weeks is superior to both
donor milk and preterm formula in promoting growth and reducing
morbidities, including ROP. Large regional European differences in
provision of maternal milk exist with only 36% of very preterm infants
receiving any breast milk at discharge in one unit compared to 80% in
another. In Sweden, the rates of exclusive maternal milk feeding at discharge
of extremely preterm infants have declined from 55% to 16% from 2004 to
2013 (97). Therefore, efforts for improvements seem warranted. The World
Health Organization and the United Nations Children’s Fund have launched a
special version of a Baby-Friendly Hospital Initiative for preterm and sick
infants to implement practices that promote breast-feeding. It defines 10 steps
to successful breast-feeding (98). Allocating resources to follow these
guidelines could be an efficient method to increase growth and reduce ROP
and other morbidities.
Timing of Postnatal Growth

Control of growth is most important early after preterm birth. This is shown
with WINROP, which identifies infants at risk for severe ROP on average 3
weeks after birth if the postnatal weight gain is poor. ROP is a two-phased
disease. The first phase of poor retinal vessel growth resulting in greater areas
of avascular retina is seen simultaneously with delayed general growth. Phase
I occurs from birth until approximately 30 weeks’ PMA. Thereafter, catch-up
of general growth occurs, and during this period, some infants, mainly those
with the poorest initial weight gain, develop phase 2 ROP with pathologic
retinal neovascularization that threatens their vision.
With increasing knowledge about the neonatal physiology of the very
preterm infant and identification of factors necessary for normal growth, care
can be improved and ROP hopefully prevented, with fewer infants needing
destructive laser therapy or antiangiogenic drugs with potentially serious side
effects. It is likely that general health will also benefit from these
improvements addressing early postnatal growth of the very preterm infant.
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40
Oxygen as a Pathogenic Factor in
Retinopathy of Prematurity (ROP)
Dolly A. Padovani-Claudio, Megan E. Capozzi, Colin A. Bretz, and
John S. Penn
INTRODUCTION TO RETINOPATHY
OF PREMATURITY
Although therapeutic oxygen in preterm infants is vital for tissue, organ, and
cognitive development, it can have detrimental consequences even when
strictly managed. One such consequence is retinopathy of prematurity (ROP).
ROP results from dysregulated tissue oxygenation in the retinas of preterm
infants and is a major cause of irreversible vision loss in children worldwide.
Development of the retinal vasculature begins at approximately 15 to 16
weeks of gestation in humans and proceeds radially in a circular wave from
the optic nerve head to the retinal periphery leading to the formation of the
primary superficial retinal vascular plexus. Angiogenesis and vascular
remodeling contribute to the radial expansion of the superficial plexus and to
the growth of the secondary deep retinal vascular plexus (1). These events are
driven largely by tissue growth and the subsequent increased oxygen demand
within the relatively hypoxic uterine environment (arterial blood partial
pressures of oxygen [PaO2] in the range of 20 to 30 mm Hg) (2). Tissue
hypoxia promotes the production of angiogenic growth factors such as
vascular endothelial growth factor (VEGF), an endothelial cell mitogen
tightly regulated by tissue oxygen tension (3–7). Peripheral retinal
vascularization increases with gestational age, and, under normal
developmental conditions, vascularization of the retina is completed by 38 to
40 weeks (8). In full-term infants, the blood-borne oxygen delivered by the
newly grown vasculature in combination with the relatively hyperoxic
postnatal environment (PaO2 between 55 and 85 mm Hg or higher) into
which an infant is born prevents persistent vascular proliferation by inhibiting
the production of VEGF and other growth factors (2,9,10).
In premature infants, the retinal vasculature is incompletely developed,
and the peripheral retina is avascular at the time of birth. Exposure to the
hyperoxic postnatal environment leads to suppression of VEGF production
and delayed physiologic retinal vascularization, which poses increased risk of
insufficient peripheral retina vascularization. Under these circumstances,
retinal oxygen levels are temporarily maintained primarily by the neighboring
choroidal circulation, which is unique in that its autoregulatory response to
altered oxygen tension results in limited vasoconstriction (11,12). Oxygen
diffuses from the choroidal circulation into the retinal tissue, bathing the
retina and worsening the inhibitory effect of hyperoxia on the growth and
maintenance of sensitive retinal capillaries (13,14). In combination, these
events define the first phase of ROP: stunted physiologic retinal vascular
development. Clinically, the radial location of the junction of the vascularized
and the peripherally avascular retina determines the zone of ROP (Table 40-
1).
TABLE 40-1 Definitions and illustrations of

retinopathy of prematurity pathophysiologic and
clinical progression highlighting the presumed role
of oxygen in ROP development
N, nasal ora; T, temporal ora; ON, optic nerve; F, fovea; Z1, zone 1; Z2, zone 2; Z3, zone 3; NV,
neovascularization; RD, retinal detachment; O2, oxygen; VEGF, vascular endothelial growth factor.
Eventually, the oxygen demands imposed by the growing retina and the rapid
development of retinal neurons are unfulfilled, leading to hypoxia in the
avascular peripheral retina. Retinal hypoxia induces the second phase of
ROP: vasoproliferation. During the vasoproliferative phase of ROP, the
hypoxic retina dramatically up-regulates VEGF production, leading to
increased vascular growth. In some infants, this vascular growth is abnormal,
with vascular beds that are poorly perfused, and fragile, worsening tissue
hypoxia. Moreover, dysfunctional autoregulatory mechanisms that may
further restrict retinal blood flow can potentiate this retinal hypoxia (15). The
consequence of these alterations is the persistent growth of abnormal vessels.
Fragile vessels can leak or rupture and bleed, therefore posing a threat to
retinal and visual development. In addition, vascular growth, intended to be
oriented toward the retinal periphery, can be misdirected to the vitreous
cavity. Ultimately, if this intravitreal neovascularization is left untreated, it
may cause fibrovascular contraction of the vitreous leading to tractional
retinal detachment and blindness (16,17). As this pathologic retinal vascular
growth is highly hypoxia-driven, weaning premature infants from oxygen
therapy during the vasoproliferative phase, although not required for ROP
development, is thought to predispose to ROP or contribute to ROP
worsening (18). Clinically, the severity of retinal abnormalities resulting from
the vasoproliferative response determines the stage of ROP (Table 40-1) (see
Chapter 52).
THE EFFECT OF OXYGEN

MANAGEMENT ON RETINOPATHY OF
PREMATURITY: HISTORICAL
PERSPECTIVE
Early studies suggested that the cardiopulmonary complications associated
with prematurity could be addressed by consistent exposure to fractional
inspired oxygen (FiO2) levels of 0.7 (19). Thus, exposure of preterm infants
to intensive oxygen treatment became the standard of care in the early 1940s.
This treatment paradigm improved general patient outcomes for many
neonates; however, the change was accompanied by a rise in the occurrence
of retinal vascular abnormalities leading to Theodore Terry’s landmark
description of retrolental fibroplasia (RLF) in 1942 (20).
Terry first identified severe ROP (initially termed RLF) as a condition
associated with premature birth, albeit sporadically seen in full-term infants
(20). Subsequently, in 1951, Campbell suggested that supplying uncontrolled
oxygen to newborns was responsible for the ROP epidemic in premature
infants emphasizing the importance of limiting the use of prophylactic
oxygen therapy on premature infants and reserving it for the treatment of
cyanosis (21,22). Subsequent studies in the 1950s describe an association
between high levels of oxygen, attenuation of retinal vasculature, and the
most severe forms of RLF (23,24). Later, this period was coined the “first
epidemic” of ROP. Soon after, Patz demonstrated in a small prospective,
randomized clinical trial that limiting therapeutic oxygen levels resulted in a
significant reduction in ROP incidence (14). He stressed the importance of
measuring actual incubator oxygen levels rather than relying on oxygen flow
rates to determine oxygen exposure in these infants. Oxygen administration
protocols were revolutionized by these findings coincident with a drop in the
incidence of ROP-associated blindness (25,26). Unfortunately, systemic
complications, including death, increased due to low oxygen administration
trends in neonatal care (27).
The rise and fall in ROP incidence in developed and developing countries
since its discovery in the 1940s has been attributed to several factors, of
which oxygen delivery is only one. Although gestational age and birth weight
consistently emerge as the strongest factors associated with ROP incidence,
enhanced screening and advances in ophthalmic management of ROP (both
surgically and medically) are thought to contribute to the variability in rates
of ROP-related blindness. Despite the careful monitoring of oxygen delivery
to neonates in the 1960s to 1980s coincident with cutting-edge advances in
neonatal care, the incidence of ROP again soared in what was coined the
“second epidemic” (28,29). This epidemic was thought to be due to increased
survival rates of very low–birth weight premature infants (<1,000 g) rather
than their clinical management. This new wave of ROP suggested that the
developmental stage of premature infants may impact responses of the
immature retina to oxygen (29–31). The conundrum regarding optimal
oxygen supplementation resurfaced, and, although limiting oxygen
supplementation led to marked decreases in ROP incidence, it again resulted
in unacceptable morbidity and mortality in these more vulnerable infants.
With time and technologic and medical advances in developing countries,
a new rise in ROP has emerged during this century. This “third epidemic” is
not limited to extremely low–birth weight infants as seen in the “second
epidemic” but is characterized by an unusually higher rate of ROP incidence
in older (and heavier) premature infants than is currently seen in developed
countries. This “third epidemic” is thought to be partly due to enhanced
access to neonatal intensive care facilities with less sophisticated technologic
advances, inadequately regulated oxygen supplementation protocols, and
inconsistent access to ophthalmic care, reminiscent of the “first epidemic”
(32). Table 40-2 summarizes the relative role of uncontrolled oxygen
supplementation and extreme prematurity in the various ROP epidemics.
TABLE 40-2 Relative contributions of oxygen

control versus extreme prematurity in the various
ROP epidemics
Adapted from Gilbert C. Retinopathy of prematurity: a global perspective of the epidemics, population
of babies at risk and implications for control. Early Hum Dev 2008;84:77–82. Ref. (33).
ANIMAL STUDIES OF OXYGEN IN

Due to the variability in causal mechanisms that trigger premature delivery of
preterm infants and the ethical and safety limitations of performing clinical
trials in this patient population, animal models of ROP were developed soon
after ROP was identified. Multiple species and iterations of these models
were tested with the goal of best approximating human ROP
pathophysiology.
Early experimental models exposed newborn, healthy mammals to
constant high oxygen levels equivalent to those used in neonatal intensive
care units (NICUs) at the time. The healthy newborn animals had robust
pulmonary function leading to very high retinal tissue oxygen tensions from
exposure to the experimental levels of inspired oxygen and were not
representative of premature infants who developed ROP. This physiologic
discrepancy limited the usefulness of the models in investigations of ROP
pathogenesis, particularly in the current day. As our understanding of the
ROP progressed, models were developed with lower and fluctuating oxygen
levels chosen to recreate blood oxygen profiles that better represented those
experienced by preterm infants in whom ROP develops. The term oxygen-
induced retinopathy (OIR) is, therefore, routinely used to describe animal
models that develop features of human ROP (34).
Effects of Oxygen on Physiologic Retinal Vascular

Development and Pathogenic Vasoproliferation in
Animal Models
One of the earliest models of OIR was the kitten model used by Norman
Ashton in the 1950s (35). Vascular development in a newborn kitten is
similar to that of a preterm infant at 28 weeks of gestation comparable to the
at-risk population at the time that ROP was discovered (36). Ashton used the
kitten model to investigate how variables such as postnatal age at the time of
oxygen administration, oxygen concentration, and length of oxygen exposure
affect the severity of OIR (37). He found that oxygen concentrations between
0.7 and 0.8 FiO2 produced the most severe retinopathy and that FiO2 levels of
at least 0.35 were necessary to induce vascular loss and later intravitreal
neovascularization in even the most susceptible kittens (24). His work
demonstrated the destruction of retinal capillaries by high oxygen
concentrations within a span of hours in younger kittens, whereas more
mature blood vessels in older kittens were less susceptible to high oxygen
concentrations (24). Thus, the kitten was instrumental in our early
understanding of the role of oxygen stress on developing retinal vasculature
and reinforced the notion that the retina’s developmental stage can affect its
susceptibility to oxygen.
Decades later, the mouse has become the most commonly used animal in
studies of retinal angiogenesis largely due to the ease of genetic manipulation
of the species. However, in 1954 long before the advent of transgenic
technologies, Hellstrom and Gyllensten reported that 1 to 3 weeks exposure
to 1.0 FiO2 resulted in vitreous and anterior chamber hemorrhages in
newborn mice. However, no preretinal vascular proliferation was observed
unless the pups were exposed to room air following the hyperoxic insult
(38,39). The relative hypoxia, as in the case of prematurely delivered infants,
was thought to be the trigger of the vasoproliferative phase. Despite its early
adoption by ROP research groups, the mouse model of OIR did not produce
reliable results for nearly half a century.
It was only after Smith and colleagues described modifications in the
model leading to reproducible results, that the mouse OIR model became
widely used to study intravitreal neovascularization and ROP. Smith’s group
exposed mice to 0.75 FiO2 at postnatal day 7 for 5 days to induce
vasoattenuation, which manifests as an avascular central retina due to the
destruction of central capillaries. After 5 days in 75% oxygen, mice are
removed to room air and develop vasoproliferation and partial
revascularization of the central retina (40). Unfortunately, this pattern is
opposite that seen in the vast majority of human ROP in developed countries,
where the avascular retina and preretinal neovascularization are found at the
retinal periphery.
There remained a critical need for animal models that both reflected the
blood oxygen concentrations experienced by preterm infants in modern-day
NICUs and recreated a human-like pattern and distribution of vessel growth
delay, with a peripheral avascular region as seen in humans. The most widely
used model that accomplishes this is the variable oxygen rat model first
developed by Penn and colleagues in 1993. Retinal vascularization in rats
usually completes around postnatal day 15, unlike humans, in which
vascularization completes at birth. Compromised lung function and
variability in oxygen delivery from clinical activities in the NICU result in
fluctuations in PaO2 in preterm infants (41–43). Therefore, Penn and
colleagues developed a model to recapitulate oxygenation fluctuations in
their rat model. Penn’s group exposed rat pups to oscillating levels of oxygen
followed by a postexposure period in room air (44). In the model, rat pups
experienced oxygen concentrations that cycled between 0.8 and 0.4 FiO2
during 12-hour intervals. The variable oxygen model induced intravitreal
neovascularization in two-thirds of oxygen-exposed pups, whereas constant
exposure to 0.8 FiO2 rarely (range from 0% to 13%) resulted in intravitreal
neovascularization.
Penn and coworkers subsequently improved the clinical relevance of the
model by testing a range of alternating hyperoxic and hypoxic cycles to
reproduce PaO2 values and fluctuations in the rats, which mimicked those
experienced by premature infants in whom ROP develops (45). Their
findings suggested that PaO2 variability was the most critical factor in
determining OIR outcome. The results also suggested that an FiO2 difference
(delta FiO2) of at least 0.2 between the hyperoxic and hypoxic cycles and an
FiO2 below 0.125 in the hypoxic cycle were needed to reliably induce
intravitreal neovascularization in rats. They further found that alternating
daily cycles of 0.5 and 0.1 FiO2 for 14 days followed by 6 days in room air
resulted in the most robust and consistent peripheral retinal avascularity and
subsequent intravitreal neovascularization (46). The “50/10 model” (referring
to percentages of ambient oxygen) is still widely used to investigate both
retinal vascular pathophysiology and response to candidate drug therapies for
ROP and other neovascular retinal conditions (47,48).
It is important to note the intravitreal neovascularization that develops in
OIR animal models regresses without the retinal traction or detachments that
characterize the most severe stages of human ROP. Thus, the most salient
benefit of the rat model is its production of relevant, fluctuating PaO2 levels
resulting in a human-like pattern of retinal vascular pathology with delayed
physiologic retinal vascular development and subsequent intravitreal
neovascularization. In the rat, as in the human, the resulting intravitreal
neovascularization develops peripherally at the boundary between vascular
and avascular retinas. A comparison of the rat and mouse OIR models and of
hypoxia-induced retinal vascular patterns among the mouse OIR, rat OIR,
and human ROP is shown in Figure 40-1.
FIGURE 40-1 Comparison of oxygen exposure and return
to room air parameters in the oxygen-induced retinopathy
(OIR) mouse (A) and rat (B) models and of preretinal
neovascularization patterns between human ROP (G, H)
and mouse (C, D magnified insert) and rat (E, F magnified
insert) OIR. The pattern of pathology in the rat retina (F)
more accurately mimics the radial distribution of
peripheral avascularity and preretinal neovascular tuft
development seen in human ROP (G, H).
Oxygen Fluctuations and OIR Severity

Hypothesizing that supplemental oxygen may be beneficial for ROP
recovery, Phelps and Rosenbaum exposed 3-day-old kittens to 0.8 FiO2 for
65 hours and then allowed them to recover in one of the three oxygen
environments: room air (0.21 FiO2), a variable hyperoxic/hypoxic
environment (~0.43 to 0.8 FiO2), or a tapered withdrawal of oxygen to room
air. The result of gradual withdrawal from hyperoxia was not significantly
different from that of direct removal to room air (49). Interestingly, kittens
that were exposed to variable oxygen during the postexposure recovery
period had significantly less severe retinopathy than those exposed to
constant room air. This early study prompted later studies testing
supplemental oxygen in different OIR recovery models. Several investigators
used the rat OIR model for this purpose (48,50). In one study, following
exposure to the 50/10 OIR model, rat pups were returned to room air or a
supplemental ambient oxygen environment (0.28 FiO2) for 4 days (50).
Whereas VEGF levels were reduced with supplemental oxygen treatment, no
difference in neovascularization severity was observed. This conclusion
opposed early findings from the kitten model and was a more accurate
predictor of the outcome of supplemental oxygen clinical trials (see below).
Fluctuating blood arterial oxygen levels in the rat model of OIR were also
critical to our understanding of the molecular mechanisms involved in ROP
pathogenesis. The extended bouts of hyperoxemia and hypoxemia imposed
on the newborn rats, which recapitulated those in sick preterm infants,
dramatically altered the levels of angiogenic and antiangiogenic factors in the
developing eye. Werdich and collaborators demonstrated the effect of
inspired oxygen fluctuations on retinal VEGF production at various time
points in rat OIR (51). They showed that episodic hyperoxemia arrested or
delayed physiologic retinal vascular development in association with reduced
retinal VEGF and that subsequent preretinal neovascularization was
associated with increased retinal VEGF when the animals were returned to
room air. Furthermore, both the level of retinal VEGF after return of rats to
room air and the severity of neovascularization were positively correlated
with the degree of FiO2 fluctuation during exposure.
Oxygen and Oxidative Stress in OIR: A Quest for

Potential Antioxidant Therapies
In addition to expanding our understanding of the mechanisms and factors
that contribute to ROP, OIR models have been used to develop and test
therapeutic interventions. The notion that inspired oxygen attenuated retinal
capillaries through endothelial cell destruction by reactive forms of molecular
oxygen led researchers to investigate the potential therapeutic efficacy of
antioxidants in OIR. In the mid-1970s, Phelps and Rosenbaum administered
alpha-tocopherol (vitamin E) to kittens exposed to 2 to 3 days of constant
approximately 0.8 FiO2 and found effective reduction in vessel attenuation
(52). They hypothesized that vessel closure during hyperoxemia was due to
damage from lipid-soluble oxygen free radicals, and their results suggested
that vitamin E might be efficacious in the treatment of ROP. Investigations of
antioxidant therapies for OIR expanded to other OIR models in other species.
To determine the effect of oxygen exposure on retinal levels of
antioxidants, Penn and coworkers exposed newborn albino rats to 0.6 FiO2
for 14 days and found reductions in retinal alpha-tocopherol and ascorbic
acid (vitamin C), but no change in glutathione reductase, glutathione S-
transferase, or glutathione peroxidase compared to room air controls (53).
They then manipulated the diet of nursing mothers through exogenous
administration of vitamin C or E to raise the nursing pups’ retinal ascorbic
acid and alpha-tocopherol levels, respectively. Vitamin C supplementation of
mothers did not significantly alter the retinal ascorbic acid levels in the pups
regardless of their exposure to oxygen. Interestingly, vitamin C
supplementation increased retinal alpha-tocopherol levels in room air–reared
pups but not in oxygen-exposed pups, and no changes in vasoattenuation
were observed. Conversely, vitamin E dietary intake in mothers induced a
three- to fourfold increase in retinal alpha-tocopherol in both room air– and
oxygen-exposed pups. However, both the low–vitamin E and high–vitamin E
groups exhibited reduced avascular retinal area and increased retinal vessel
density when compared to pups from regular chow-fed mothers (53). The
research group concluded that altering retinal levels of one antioxidant
molecule may affect levels of others, making it challenging to tease out the
effect of each oxidative stress–altering molecule on retinal vascular
development. Consistent with this, clinical trials evaluating the efficacy of
vitamin E supplementation in premature infants yielded inconsistent results
(54).
Using the mouse OIR model, Spierer and coworkers overexpressed the
naturally occurring antioxidant enzyme copper–zinc superoxide dismutase
(CuZnSOD) in mice. Overexpression of the CuZnSOD antioxidant
significantly reduced central vasoconstriction, vessel tortuosity, and
intravitreal neovascularization compared to wild-type mice (55). In addition,
prompted by the knowledge that nitric oxide (NO)-derived peroxynitrite is a
known mediator of oxidative injury in endothelial cells and is elevated in
infants with bronchopulmonary dysplasia, Brooks and colleagues used
transgenic mice to determine the potential pathogenic role of NO produced
by endothelial NO synthase (eNOS) (56,57). They demonstrated decreased
oxidative stress in eNOS-deficient mice or after pharmacologic inhibition of
the enzyme in the mouse OIR model, showing that eNOS specifically
affected immature vessels. Furthermore, the findings stimulated further study
of eNOS activation in other animal models including rat OIR. Using the
50/10 OIR model, Hartnett and colleagues demonstrated that, although eNOS
accumulation coincided with the peak of vascular tortuosity, a feature similar
to human plus disease in ROP, it was not caused by elevated VEGF, or
responsible for the tortuosity, as blocking VEGF signaling could reduce
tortuosity without changing the levels or activation state of eNOS (58).
CLINICALLY TARGETING OXYGEN

AND ITS BY-PRODUCTS
Current Treatment Strategies
Based on data derived from OIR models, clinical trials have addressed phase
2 of ROP by interfering with VEGF up-regulation in order to curb
vasoproliferation. The two most accepted modalities to achieve this are by
retina ablation with laser or cryotherapy and by intravitreal delivery of drugs
targeting VEGF signaling (59,60). This section will focus specifically on the
oxygen stress–related functions of these two interventions.
Cryo- or laser treatment are postulated to reduce oxygen-demanding
retinal cells in the peripheral avascular retina and, therefore, decrease the
generation of VEGF in response to tissue hypoxia while sparing the available
oxygen to the posterior pole (17,59). Under normal circumstances, oxygen
and nutrients diffuse from the choriocapillaris into the retina. As
demonstrated by Linsenmeier (Figure 40-2), most of this choriocapillaris-
derived oxygen is consumed by the photoreceptors, which have a high
density of mitochondria and high oxygen consumption (61,62).
FIGURE 40-2 Oxygen profile data (dotted trace) with
outer retinal model fit (solid trace) superimposed. A
simplified schematic of the retina is shown to illustrate the
anatomic correspondence of each model layer.
(Republished with permission of Association for Research
in Vision & Ophthmology, from Padnick-Silver L,
Linsenmeier RA. Effect of acute hyperglycemia on oxygen
and oxidative metabolism in the intact cat retina. Invest
Ophthalmol Vis Sci 2003;44(2):745–750; permission
conveyed through Copyright Clearance Center, Inc. Ref.
(62).)
Hypothetically, tissue ablation allows these peripheral retinal areas to

function as photoreceptor free windows through which choroidal oxygen is
free to diffuse into the inner retina, leading to increased oxygen tension and
decreased VEGF production (60). Several studies have evaluated oxygen
tension in the inner retina following laser treatment in animal models of
neovascularization or in diabetic patients. Improved tissue oxygenation
following retinal laser ablation was first shown in rhesus monkeys (63,64)
and was subsequently confirmed in several studies (65–67). In addition,
decreased levels of VEGF have been documented following laser ablation in
humans. It is reasonable to hypothesize that laser-induced destruction of the
glial cells most responsible for VEGF production also contributes to VEGF
decrease.
Aiello and colleagues analyzed vitreous samples from diabetic patients to
determine the effect of laser treatment on VEGF release. Patients receiving
laser photocoagulation therapy had reduced vitreous VEGF protein levels
compared to patients with untreated proliferative diabetic retinopathy (68).
Similarly, in postmortem human retinal tissue, VEGF immunoreactivity was
decreased following successful laser photocoagulation therapy (69,70).
Finally, reduction in serum VEGF levels has been reported following
panretinal photocoagulation in patients with proliferative diabetic retinopathy
(71) and ROP (72). Despite its benefits, panretinal laser photocoagulation
may require general anesthesia, is time-consuming, and is associated with
side effects and potential complications, including peripheral visual field
constriction, poor night vision, inflammation, and potential development of
cataracts and myopia. Therefore, investigators have continued to seek new
and improved therapies for premature infants at risk of vision loss from ROP.
Because of the strong association between VEGF expression and the
development and progression of ROP, anti-VEGF agents have been tested as
therapies in ROP. In an attempt to reduce the intravitreal neovascularization
of ROP, the Bevacizumab Eliminates the Angiogenic Threat for Retinopathy
of Prematurity (BEAT-ROP) trials employed a VEGF monoclonal antibody,
bevacizumab (see Chapter 52). This strategy was designed to curb the effects
of up-regulated retinal VEGF in the retinas of premature infants. Anti-VEGF
agents, including ranibizumab and aflibercept, are relatively easy to
administer, reduce vascular activity in many eyes, and may potentially reduce
side effects of laser treatment. However, a Cochrane review published in
2018 evaluated data from six clinical trials that compared laser and anti-
VEGF therapies and found insufficient evidence to support anti-VEGF agents
to reduce ROP recurrence or retinal detachment (73). In addition, there are
safety concerns related to the use of anti-VEGF therapies as they access the
systemic circulation and may adversely affect other organs that require VEGF
for development (see chapter on Anti-VEGF Agents Chapter 53). VEGF-
targeted therapies are being tested in large-scale clinical trials and published
results are awaited. Except for zone 1 ROP or aggressive posterior ROP,
near-confluent laser ablation of the peripheral avascular retina remains the
standard of care for ROP therapy (74,75).
Oxidative Stress–Related Therapies

The retina is particularly susceptible to oxygen-mediated damage because
retinal cells have a high rate of oxygen consumption. In addition,
polyunsaturated fatty acids, preferred substrates for harmful lipid
peroxidation reactions, are highly prevalent in the retina (76). As the light-
sensing organ, the retina is repeatedly exposed to photon energy, which can
initiate oxygen free radical formation (48,77). Free radicals derived from
molecular oxygen can lead to DNA damage and to protein degradation
through polypeptide cleavage. In addition, reactive oxygen radicals can
directly induce retinal VEGF production by stabilizing hypoxia-driven
transcription factors (78) (see also Chapter 41). Oxygen consumption in the
retina is high under normal development and, theoretically, more so with
therapeutic oxygen treatment in vulnerable premature infants in whom the
partial pressure of oxygen in the retina is likely to be episodically elevated. In
premature infants, repeated fluctuations in oxygenation increase the
generation of reactive oxygen species and the expression of VEGF, which
can predispose to the development of ROP (79,80).
Antioxidants in the retina can combat peroxidation by enzymatic
detoxification of reactive oxygen molecules or termination of radical chain
propagation. One example of these antioxidants is vitamin E, which
scavenges reactive oxygen species that leads to peroxidation. It remains
unclear whether, at premature birth, the retinal antioxidant system is
sufficiently developed. In the case of vitamin E, premature infants are born
with only 10% of the adult retinal vitamin E levels (81). However, the tissue
levels of other antioxidants (glutathione peroxidase, glutathione S-transferase,
ascorbic acid) are substantially greater in premature infant retinas than in
mature retinas (82).
As discussed above, antioxidant therapy in the kitten and rat models
proved efficacious in reducing vasoattenuation in OIR. This prompted human
studies to assess whether antioxidant supplementation could effectively
prevent progression to threshold ROP. Vitamin E was the first biologic
antioxidant agent introduced as a potential ROP therapy in 1949 (83). More
extensive clinical trials were conducted in the 1980s to assess the effect of
prophylactic and therapeutic supplementation of vitamin E (54,84–89).
Unfortunately, despite an abundance of data on its use in the treatment of
ROP, there remains no conclusive case for its efficacy. Brion and colleagues
performed a meta-analysis of studies in which vitamin E was administered to
premature infants (90). Based on this compilation of data, they found that
vitamin E supplementation did not significantly affect the risk of developing
all grades of ROP but only significantly reduced progression to severe grades
of ROP and ROP-associated blindness in a small cohort of infants. However,
the dose necessary to afford ROP protection in this study was associated with
an increased risk of sepsis suggesting a potential harmful effect of
supplementation (90).
A major caveat to interpretation of this Cochrane meta-analysis is that
many of the contributing studies were performed in the 1980s, when standard
of care and technology in the NICU were much different than what is
encountered today. Therefore, a multicenter, randomized trial with a large
patient population would still be required to determine the potential benefit of
vitamin E supplementation under treatment conditions that premature infants
encounter currently. Important variables in the study design would include
the degree of prematurity and birth weight; vitamin E dosage; route, duration,
and schedule of administration; and the achieved plasma concentration of
alpha-tocopherol.
Small clinical studies have been performed to assess the potential
therapeutic utility of other antioxidants, such as ascorbic acid (vitamin C) and
superoxide dismutase. A group in New Zealand supplemented ascorbic acid
to infants in the NICU that were either <1,500 g or <32 weeks of gestation at
the time of birth (91). Only infants admitted within 48 hours of birth were
included in the trial. Selected infants were supplemented for 28 days. The
primary outcomes of this trial were the oxygen supplementation required at
28 days and 36 weeks of gestation, the total amount of oxygen
supplementation needed, and the incidence of ROP. The treatment regimen
with vitamin C did not yield significant benefit or harmful effects. However,
the study may have lacked statistical power because only 119 infants were
enrolled (91).
Recombinant human Cu/Zn superoxide dismutase (rhSOD) has been
tested in the clinic for several oxidative stress–related diseases. The North
American rhSOD study group administered the antioxidant supplement
intratracheally in an attempt to prevent bronchopulmonary dysplasia (92).
Three hundred and two patients were enrolled to receive either intratracheal
rhSOD or placebo every 48 hours for a month, beginning at birth. Initial
analysis of the population indicated a pulmonary benefit of intratracheal
rhSOD in premature infants at 1-year adjusted age. This positive result
prompted Parad and colleagues to perform a retrospective analysis of this
multicenter trial to determine if rhSOD also decreased the incidence of ROP
(93). No significant difference in the incidence of ROP was found within the
cohort. However, subgroup analysis identified a 22% reduction in ROP for
infants born <26 weeks (n = 72) and a 53% reduction in ROP for infants born
<25 weeks (n = 24). While post hoc analysis suggested a benefit for rhSOD
in the highest-risk newborns, a larger study is needed to confirm the efficacy
of rhSOD in the NICU (93).
Finally, ambient light in the NICU has also been suggested to be a
potential mechanism for the generation of reactive oxygen radicals in the
preterm infant. The Light Reduction in Retinopathy of Prematurity (Light-
ROP) trial enrolled 409 patients that were randomized to two groups: one in
which the infants were exposed to standard ambient light and the other in
which light-reducing goggles were worn (94). The goggles, which reduced
visible light by 97% and ultraviolet light by 100%, were placed on the infants
in the experimental group within 24 hours of birth. The goggles remained on
the infants for the longer of two options: 31 weeks postconception or 4 weeks
after entering the trial. The primary outcome of the study was the presence of
ROP, and the secondary outcome was the stage of disease. This study
determined that the reduction of ambient light did not decrease the incidence
of any stage of ROP, while no adverse effects resulted from wearing the
goggles (94). However, the retina is more metabolically active in dark, and
this may be part of the reason light reduction was not found to reduce ROP
severity.
As can be concluded from these examples, although potentially
promising as strategies to reduce ROP and its related blindness, the data
derived from trials of antioxidant therapy for ROP prevention have been
inconsistent or inconclusive. Several factors contribute to this, including
evolving NICU clinical management protocols, evolving characteristics
(gestational age and weight and overall health) of surviving premature
infants, and underpowered studies, among others. Therefore, antioxidant
therapy has not been adopted as the standard of care for premature babies at
risk of ROP. In addition, the studies done on antioxidants do not address the
potential role of oxidative compounds as signaling molecules involved in the
pathogenesis of ROP (see Chapter 41). However, with the advent of advances
in genomics, proteomics, and metabolomics, it may be possible to identify
biomarkers that could aid in the interpretation of results of clinical trials, help
predict susceptibility, or guide personalized medical interventions for ROP
(95).
Oxygen Monitoring and Management

Understanding the deleterious effects that oxygen can have on retinal
vascular development and the challenges in treating advanced ROP stages
has raised questions regarding the length of time preterm infants should
remain on oxygen therapy. Early on, clinicians were reluctant to provide
oxygen supplementation for an extended length of time because they feared
oxygen toxicity. Therefore, neonatologists transitioned infants from oxygen
to room air as soon as lung function allowed. However, kitten studies by
Phelps suggested that oxygen supplementation following hyperoxic exposure
might be beneficial (49,96). Askie and Henderson-Smart retrospectively
analyzed a study by Engle, designed to compare gradual versus abrupt
oxygen discontinuation across various gestational age groups and birth
weights in humans (97,98). Growth, neurodevelopment, and the incidences of
ROP and lung disease were assessed after gradually weaning infants from
oxygen supplementation. No significant benefit was found between gradual
and abrupt discontinuation of supplemental oxygen. Since then, the interest
has instead turned to determining the safest concentration range for oxygen
treatment of susceptible infants.
Several trials have specifically focused on oxygen supplementation levels
in the different phases of ROP and their role in ROP pathogenesis. Seeking to
design a more stringent oxygen-monitoring paradigm that would reduce ROP
progression, Flynn used noninvasive transcutaneous electrodes to
continuously measure transcutaneous partial pressure of oxygen (TcPO2) in
preterm infants (99). The amount of time at TcPO2 > 80 mm Hg was
correlated with more severe ROP. Subsequently, using continuous
transcutaneous measurements, Cunningham demonstrated that TcPO2 levels
fluctuate on a minute-to-minute basis in the first 2 weeks of life in preterm
infants who develop ROP (43). In fact, TcPO2 variability was shown to be a
significant predictor of ROP severity. Furthermore, a retrospective study of
arterial blood oxygen levels in catheterized infants by York and colleagues
further demonstrated the importance of oxygen fluctuation, as opposed to
average PaO2 over the course of therapy, in the development of ROP (42).
Using a cohort of 231 infants, these investigators found that very low birth
weight infants who experienced fluctuating PaO2 due to either ambient
oxygen variability or poor lung function were at a higher risk for progressing
to threshold ROP. These studies further emphasized the important role of
oxygen in ROP pathogenesis and suggested that the average level and degree
of fluctuation in circulating oxygen as measured by pulse oximetry, TcPO2 or
PaO2, should be closely monitored in order to limit ROP progression.
The range of ambient oxygen saturation immediately after birth and
during recovery became a major focus of clinical trials (see also Chapter 52).
The Supplemental Therapeutic Oxygen for Prethreshold ROP (STOP-ROP)
trial was conducted from February 1994 to March 1999 to assess how
supplementing oxygen to reach a higher than conventional target SpO2 would
affect the progression to threshold ROP and the need for avascular retinal
ablation in infants with prethreshold ROP (100). Six hundred and forty-nine
infants with moderately severe ROP were enrolled, excluding those with
oxygen saturation of >94% at the time of enrollment. Infants were
randomized to oxygen administration at conventional levels (89% to 94%
saturation) versus supplemental levels (96% to 99% saturation). The infants
remained on oxygen therapy for at least 2 weeks, until both eyes reached
study end points (ranging from reaching threshold criteria for ablation,
reaching full retinal vascularization, or regressing to zone 3 ROP for >2
weeks). The use of supplemental oxygen to reach higher SpO2 target did not
change the rates of threshold ROP or need for retinal ablation. Supplemental
oxygen only reduced the progression to threshold ROP in a narrow subgroup
of infants without plus disease (101). In this small cohort, only 32% of
infants on supplemental oxygen progressed to threshold ROP, while 46%
progressed with conventional oxygen saturation. The STOP-ROP trial did not
lead to changes in standard of care practices for ROP.
The High Oxygen Percentage in ROP (HOPE-ROP) study followed the
STOP-ROP trial. The goal of HOPE-ROP was to analyze infants that were
excluded from STOP-ROP due to arterial oxygen saturations of >94% at the
time of prethreshold diagnosis. One hundred thirty-six HOPE-ROP infants
were compared to 229 STOP-ROP patients. After controlling for gestational
age, race, zone 1 disease, and plus disease at prethreshold diagnosis, the
analysis determined that HOPE-ROP infants were significantly less likely to
progress to threshold ROP, suggesting that arterial oxygen saturation values
at prethreshold diagnosis might provide a prognostic indicator of ROP
severity and risk of progression (102).
Although healthy preterm infants maintain an oxygen saturation >95%
most of the time, Tin and colleagues have shown that attempts to maintain
arterial oxygenation at this level may do more harm than good. As shown in
Table 40-3 and Figure 40-3, infants maintained at 70% to 90% oxygen
saturation had a 6.2% chance of developing severe retinopathy, whereas
those maintained at 88% to 98% oxygen saturation had a 27.7% chance
(103). Additional studies were performed in infants who were maintained at
low (80% to 90%) and high (94% to 98%) oxygen saturations to determine
long-term (10-year) outcomes. Cognitive, adaptive, and behavioral
performances were assessed (104,105). Unfavorable visual outcomes occurred
in 2.5% of the eyes in the low saturation group and in 12.7% of the eyes in
the high saturation group. Measures of cognitive and behavioral performance
were similar between the two groups.
TABLE 40-3 The effect of O2 saturation range on

survival, cerebral palsy, and the development of
threshold ROP
Figure adapted from Tin W, Milligan DW, Pennefather P, Hey E. Pulse oximetry, severe retinopathy,
and outcome at one year in babies of less than 28 weeks gestation. Arch Dis Child Fetal Neonatal Ed
2001;84:F106–F110. PMID 11207226. Ref. (103).
FIGURE 40-3 The effect of O2 saturation range on

survival, cerebral palsy, and the development of threshold
ROP. (Reproduced from Tin W, Milligan DW,
Pennefather P, et al. Pulse oximetry, severe retinopathy,
and outcome at one year in babies of less than 28 weeks
gestation. Arch Dis Child Fetal Neonatal Ed
2001;84:F106–F110. With permission from BMJ
Publishing Group Ltd. Ref. (103).)
In 2003, an Australian multicenter study known as the Benefits of Oxygen

Saturation Targeting (BOOST) was completed (26). The primary aim of this
study was to determine if maintaining oxygen saturation at a higher than
standard level (95% to 98% vs. the standard 91% to 94%) would improve
long-term growth and development among oxygen-dependent preterm
infants. The study found no clear long-term growth or developmental benefits
to maintaining higher than standard levels of oxygen saturation. In fact,
higher than standard levels have a cost disadvantage, owing to increased
length of time on oxygen and increased cost associated with home
oxygenation.
The most recent attempt to close the evidence gap is the Neonatal
Oxygenation Prospective Meta-analysis (NeOProM) Collaboration, a large-
scale, prospective meta-analysis of five trials including SUPPORT-USA,
BOOST II-Australia, BOOST-New Zealand, BOOST II-UK, and COT-
Canada. The goal of this collaboration was to determine the optimal oxygen
saturation in the first few weeks of a preterm infant’s life that would increase
survival and decrease morbidity (106). The group hypothesized that oxygen
saturations between 85% and 89% (vs. 91% and 95%) would reduce
morbidity and disability and promote a better ROP outcome. The primary
outcome was a composite outcome of death or major disability at a corrected
age of 1.5 to 2 years of age. Secondary outcomes included several measures
of neonatal morbidity, including ROP. For the almost 5,000 randomized
infants with mean gestational age of 26 weeks and mean birth weight of 832
g, there was no significant difference in primary composite outcome between
the lower (85% to 89%) and higher (91% to 95%) SpO2 target range groups.
The lower target group showed a 4% decrease in risk for advancing to need
ROP treatment, but a 2.3% and 2.8% increase in risk of necrotizing
enterocolitis and death, respectively.
Our capacity to manage an infant’s blood oxygen is limited. Although
there is evidence to support important roles for both the average
concentration of therapeutic oxygen and the degree to which oxygen
fluctuates about that average over time, questions remain about their relative
pathogenic contributions. In fact, the two are likely to be directly related. The
technologic constraints of oxygen-sensing equipment, the infant’s clinical
course, and NICU management practices dictate that saturation values will
fluctuate within a finite percent deviation from the intended level, meaning
higher target saturations will yield larger variation ranges. This has led to the
suggestion that the benefit of maintaining lower oxygen saturation levels is
derived from a reduction in fluctuations due to these parameters (42). By
today’s standard of care, when an infant moves outside the range of
normoxemia, tissue oxygenation is manually regulated by frequent
adjustment of FiO2—a method that is ineffective for managing the constantly
fluctuating blood oxygen levels of a compromised preterm infant (107).
Automated control systems are becoming more prevalent in the medical arena
(100,108,109) and could offer valuable assistance to NICU staff by providing
immediate minor adjustments in FiO2 and reducing the frequency of full
saturation and desaturation events believed to be pathogenic in modern-day
ROP (107,110).
Despite numerous trials and years of research, there is still no consensus
on a specific oxygen target for NICUs, and more research is needed to
determine the role that oxygen saturation plays in ROP pathology. Ideally,
maximum and minimum thresholds would be determined for patient groups
based on gestational age and body weight, which are known factors
predisposing to ROP. However, care must be taken to assess the effect of
these oxygen limits on other organ systems that are still developing and
therefore dependent on appropriate oxygenation. What is optimal for one
organ system may not be optimal for another, and oxygen saturation must be
finely tuned to balance a variety of critical needs. Regrettably, since Terry’s
landmark observation, fundamental questions remain regarding the
pathogenesis of ROP, the precise influences of oxygen therapy on disease
onset and progression, and the best strategies to prevent and to treat ROP. As
technology advances and NICU resources continue to improve allowing
survival of more vulnerable premature infants, continuous research will be
needed to solve this conundrum, and it is likely that, as answers are found,
more questions will be uncovered.
CONCLUSIONS
In 1945, the esteemed surgeon and scientist, Julius H. Comroe, Jr., stated “the
clinician must bear in mind that oxygen is a drug and must be used in
accordance with well recognized pharmacologic principles; i.e., since it has
certain toxic effects and is not completely harmless (as widely believed in
clinical circles) it should be given only in the lowest dosage or concentration
required by the particular patient.” This admonition is particularly pertinent
to the care of premature infants at risk for ROP. From the time of the initial
identification and characterization of ROP, investigators have strived to
define the precise role of oxygen in its pathogenesis. Much of this work was
conducted using animal models of OIR and tightly controlled experimental
conditions. Subsequently, large-scale clinical trials have sought to further
refine our understanding of the role of oxygen in disease progression. Great
effort has been expended to understand the pathogenic role of oxygen and
oxidative stress at the molecular, cellular, and tissue levels and to define
optimal oxygen supplementation protocols to establish appropriate standards
of care for preterm infants. Despite this, we have yet to arrive at an optimally
efficacious course of action. Additional studies directed at better
understanding the role of oxygen in retinal vascular development and disease
progression remain crucial for better management and treatment of ROP.
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41
Oxidative Mechanisms and Signaling
Pathways Related to Pathogenesis of
Haibo Wang, and M. Elizabeth Hartnett
INTRODUCTION
Retinopathy of prematurity (ROP) is complex and is affected by heritable and
environmental factors (1). Some preterm infants may be predisposed or
vulnerable to environmental risks based on genotype or genetic expression. In
addition, maternal or environmental factors may modulate the regulation of
gene expression through processes, such as gene–gene interactions or
epigenetic modifications.
The goals of this chapter are to address the current understanding of ROP
with focus on the role of factors, for example, oxygen concentration and
oxidative stress, and how these trigger or regulate signaling pathways to
cause pathologic features of severe ROP, involving first damaged or
incompletely vascularized retina and second aberrant intravitreal
neovascularization. We also will discuss briefly the temporal effects of retinal
neural development and the evidence regarding the interrelationship of this
with retinal vascular development in ROP.
BACKGROUND
The Infant
The in Utero and Perinatal Setting of the Infant with
ROP
In the human fetus, early retinal vessels first appear around the optic nerve at
about 14 weeks’ gestational age (2) and, by term birth, have extended to the
ora serrata in at least two major vascular plexuses, the inner plexus at the
nerve fiber layer and the deeper vascular plexus at the inner nuclear layer (see
Chapter 5). The preterm infant, therefore, has incompletely vascularized
retina at birth. However, after birth, some preterm infants have a further delay
in physiologic retinal vascular development. There are several hypotheses for
this occurrence. One is based on lack of maternally supplied growth factors
and nutrients during normal birth in the third trimester, which the preterm
infant is unable to make and that are essential for retinal vascular
development and also overall growth (2,3). Also, the nascent vessels of the
preterm infant are vulnerable to oxidative compounds and high oxygen.
Preterm birth increases generation of oxidative compounds (3), and the
preterm infant is unable to adequately quench oxidative compounds because
of insufficient antioxidant enzyme systems. In addition, in utero, the oxygen
concentration to the fetus is believed to be <40 mm Hg. At birth, the relative
change in oxygen concentration, particularly if high oxygen is used, can
potentially injure newly formed retinal capillaries, leading to areas of
avascular retina, which stimulate later development of intravitreal
neovascularization (Figure 41-1). There is also a developing hypothesis in
which some infants can call upon potential mechanisms that protect against
ROP despite extreme prematurity and oxygen/oxidative insults. These
include possible ischemic preconditioning that may occur because of repeated
fluctuations in oxygenation (4). Further studies are indicated to test this
hypothesis.
FIGURE 41-1 Factors contribute to the development of
ROP. Low serum insulin-like growth factor-1 (IGF-1) and
its binding protein 3 IGFBP3 derived from preterm birth
lead to impaired retinal vascular development. At birth,
particularly in the units that have not implemented oxygen
monitoring, relatively high oxygen concentration increases
ROS generation, which can damage newly formed
endothelial cells and leads to capillary constriction.
Postnatal oxygen stresses including fluctuations in oxygen
delay ongoing physiologic retinal vascular development,
leading to the phase of reduced peripheral retinal vascular
development. Reduced retinal vascularization and
increased oxygen demand by photoreceptor differentiation
cause retinal hypoxia, which leads to uncontrolled
disordered angiogenesis and results in intravitreal
neovascularization, the phase of vasoproliferation.
Postnatal Stresses That Increase Risk of ROP

Postnatally, stresses associated with increased risk of ROP include
fluctuations in oxygenation (5), poor postnatal weight gain (6), perinatal
morbidities including sepsis and necrotizing enterocolitis, and other
complications of prematurity (7). Several of these can be linked to increased
generation of oxidative compounds that can injure blood vessels and/or
activate pathways of apoptosis or angiogenesis, and these processes
contribute to the pathologic features of persistent avascular retina and
intravitreal neovascularization in ROP (5).
Postnatal Oxygen
When ROP was first described in the 1940s prior to the implementation of
oxygen regulation, very high oxygen likely caused damage to newly formed
capillaries and led to ROP (8,9). To study the causes of ROP described in the
1940s, several models of oxygen-induced retinopathy (OIR models) were
developed in which animals that vascularize their retinas postnatally were
exposed to oxygen at different concentrations (10–13). The results of this
early research led to improvements in oxygen monitoring and the avoidance
of excessive, unregulated oxygen at birth (14). However, as infants of
younger gestational ages and smaller birth weights survived, ROP reemerged.
It is recognized now that, besides high oxygen at birth, postnatal fluctuations
in oxygen levels are also associated with severe ROP. It is not clear if it is
fluctuations in inspired oxygen alone that are associated with ROP risk or
those caused secondarily from bradycardia, apnea of prematurity, ventilation–
perfusion deficits related to lung disease, and other conditions of prematurity.
However, healthy newborn animals exposed to repeated fluctuations in
inspired oxygen consistently develop features of acute severe ROP
supporting the role of oxygen fluctuations in causing severe ROP features
(13). There are also conflicting reports about the role of supplemental oxygen
in the development of severe ROP. The Supplemental Therapeutic Oxygen
for Prethreshold Retinopathy of Prematurity study found no adverse effect
from supplemental oxygen and perhaps a beneficial effect in a subgroup (15).
Others have advocated later supplemental oxygen to reduce the risk of severe
ROP (16,17). However, several studies have also found correlations between
high oxygen saturations and ROP (18,19). Two multicenter clinical trials
(SUPPORT, BOOSTII) (20) found that low oxygen saturation targets were
associated with less severe ROP, but there was increased mortality, although
another multicenter study did not find a relationship (21). Currently, neonatal
units tend to develop guidelines regarding oxygen saturation targets based on
their units and outcomes.
The initial phase of avascular retina from compromised physiologic
retinal vascularity and delayed physiologic retinal vascular development is
postulated to become hypoxic once the infant is removed from supplemental
oxygen. Currently, there is no available technique to accurately measure
retinal tissue oxygenation in the human infant in vivo, but experimental
evidence provides support that retinal hypoxia precedes the development of
disordered angiogenesis (22,23) that grows outside the retinal plane and into
the vitreous as intravitreal neovascularization, rather than into the retina as
postnatal retinal vascularization. Most studies strive to understand the
mechanisms leading to this intravitreal neovascularization in ROP, but also
important is finding ways to promote retinal vascularization of the avascular
areas, protect vascularized retina from damage, and promote neuroprotective
mechanisms for normal retinal development.
Animal models of OIR are used to study how oxygen levels activate
signaling pathways that cause features of severe acute ROP; however they
only mimic the vascularly active phases of ROP, that is, plus disease and
intravitreal neovascularization (stage 3 ROP), and do not generally develop
fibrovascular features, that is, stages 4 and 5 ROP.
Oxidative Stress in Preterm Infant

Oxidative stress is postulated to play an important role in ROP. The preterm
infant is predisposed to increased oxidative stress because of an imbalance
between generation of reactive oxygen species (ROS) and antioxidant
capacity. A number of factors contribute to increased ROS generation in the
retina of the preterm infant, including hypoxia, hyperoxia, and fluctuations in
oxygen concentration; and the high metabolic rate and rapid rate of oxygen
consumption of the developing photoreceptors (3). The major sources of ROS
generation in living cells are the mitochondria and enzymes, including
nicotinamide adenine dinucleotide phosphate oxidase (NADPH) oxidase,
lipoxygenase, nitric oxide synthetase (NOS), and others. The retina is
believed to be particularly vulnerable to ROS because of abundant
polyunsaturated fatty acids in the retina, which can form oxidative
compounds, and also because of inadequate antioxidant defenses. Therefore,
if excessive ROS are not quenched, tissue damage can result. However, low
levels of ROS have physiologic functions to regulate cell growth, survival,
differentiation, and metabolism, which all benefit the developing preterm
infant. Therefore, a balance in ROS generation and quenching is needed for
physiologic growth and to prevent pathologic processes in the preterm infant.
MODELS OF ROP (OXYGEN-INDUCED

RETINOPATHY MODELS)
It is virtually impossible to study mechanisms of ROP in human preterm
infants’ eyes, because it is unsafe to obtain retinal samples, and biochemical
and signaling pathways affected in the retina may not be affected in other
tissues that are easier to sample. A great deal learned about ROP first came
from clinical observations that drove research. Subsequently, models were
developed to identify genes regulated (24) or proteins activated that caused
features of severe ROP. Animals that vascularize their retinas after birth (cat
(25), mouse (10,26), rat (13,27–32), and beagle (11)) were subjected to
oxygen stresses and developed features of severe ROP (Table 41-1). These
models have limitations. The animals used are not premature; therefore, the
effect of oxygen concentration may differ from what occurs in preterm
infants. The retina is complex with numerous cells, and interactions between
retinal cells can be difficult to study in whole retinas from animals. The retina
is also developing and differentiating, and these processes create oxygen
demands within different retinal regions at various times. These caveats are
considered when choosing animal models to address hypotheses and when
designing experiments and interpreting data.
TABLE 41-1 Animal models of retinopathy of

prematurity (oxygen-induced retinopathy)
Retinas can be removed from eyes, flattened, and stained to visualize the
vasculature in retinal flat mounts. Areas of normal retinal vascularization,
intravitreal neovascularization, and avascular retina can be measured as
compared to total retinal area. There are two broad categories of models:
those in which high oxygen damages newly formed capillaries and variable
oxygen models in which oxygen fluctuations delay ongoing vascular
development and also compromise newly formed capillaries. Each is useful
to address different experimental questions (Figure 41-2).
FIGURE 41-2 The features of retinal vessels visualized

by retinal flat mount with isolectin staining in models of
OIR. In mouse OIR model, mouse pups are exposed to
75% O2 from postnatal day 7 to day 12. This model
produces vaso-obliteration of newly formed capillaries in
the posterior retina at day 12 with decreased retinal
vascular endothelial growth factor (VEGF), IGF-1, and
erythropoietin (EPO) and intravitreal neovascularization at
the junction of vascular and avascular retina at day 17 with
increased retinal IGF-1, VEGF, ROS, and EPO signaling.
In rat OIR model, newborn rat pups are exposed to 50%
O2 followed by 10% O2 cycled every 24 hours until
postnatal day 14. This model produces features of ROP
including delayed physiologic retinal vascular
development and compromised physiologic vascularity
with decreased retinal EPO and IGF-1 and intravitreal
neovascularization at the junction of vascular and
avascular retina with increased VEGF, IGF-1, and ROS
signaling.
Hyperoxia-Induced Vaso-Obliteration Followed by

Relative Hypoxia
Initial OIR models tested the role of high oxygen at birth on the development
of ROP (Table 41-1). The mouse OIR model developed by Smith and
D’Amore is the most recognized and used (10). Mice are exposed to constant
oxygen (75%) at day 7 of life. High oxygen can change the fate of endothelial
precursors (33) and cause newly developed capillaries to constrict in part,
from apoptotic death of endothelial cells, which leads to avascular retina. In
these animal models, this event is termed vaso-obliteration and occurs in the
central, previously vascularized retina (10). Following return to room air,
endothelial budding into the vitreous is driven by relative hypoxia in the
central avascular retinal regions (25). The mouse OIR model is particularly
valuable to study genetic mechanisms and the effects of high oxygen stresses
and relative retinal hypoxia on angiogenesis, potentially as what occurred in
ROP of the 1940s. It is also useful to measure vascular repair although there
is little evidence that this process occurs in the human preterm infant. Today,
high oxygen at birth is avoided in neonatal units that have implemented
technology to monitor and regulate oxygen. High oxygen remains a concern,
for example, in units in which resources or technologic advances are
insufficient to implement oxygen monitoring and regulation (34).
Variable Oxygen
Variable oxygen is common in neonatal units that regulate oxygen. There are
a number of models of variable oxygen stresses (35–37) (Table 41-1). The
most characterized is the rat OIR model developed by John Penn, in which
newborn rat pups are exposed to fluctuations in inspired oxygen, first 50%
oxygen for 24 hours, followed by 10% oxygen for 24 hours (13). The 24-hour
cycles are repeated until day 14, at which time animals are returned to room
air. At postnatal day 14, there is peripheral avascular retina and by day 18,
aberrant intravitreal neovascularization at the junction of vascular and
avascular retina. The rat OIR model may simulate the pattern of oxygen
exposure experienced by preterm infants in the United States today. The
oxygen extremes cause arterial oxygen levels similar to transcutaneous
oxygen levels measured in human preterm infants (5). The appearance of the
retina is similar to zone II, stage 3 ROP characterized by a central
vascularized retina with some compromised physiologic vascularity and a
peripheral avascular area. Hypoxic retina is present in the avascular
peripheral retina as well as in avascular regions around retinal vessels (22).
Uteroplacental insufficiency (UPI) has been considered a risk factor of ROP.
To determine the potential effects of maternal UPI on ROP, a modification of
the rat OIR model was made in which dams underwent bilateral uterine artery
ligation and pups were born and placed into oxygen fluctuations (38). In the
modified rat OIR model, oxygen concentrations were maintained at 50% O2
for the first 48 hours instead of for the first 24 hours as in the standard rat
OIR model, and subsequently cycled between 10% and 50% O2 until day 15
(4).
Signaling Pathways Activated by Postnatal Stresses in

Animal Models
Numerous signaling pathways have been reported to be involved in retinal
development and in OIR. Here, we focus on those involved in oxidative
signaling, particularly pathways that can be translated into clinics for treating
ROP.
Oxidative Signaling Pathways

Using OIR models, it has been found that retinal ROS are generated in
association with avascular retinal areas and pathologic disordered
angiogenesis leading to intravitreal neovascularization (stage 3 ROP) (22).
Many signaling pathways are triggered by oxidative compounds and lead to
apoptosis, angiogenesis, and inflammation (39,40). Intravitreal lipid
hydroperoxides (LHP), end products of ROS, induced a cascade of
angiogenic growth factors and cytokines in the rabbit eye and led to the
development of intravitreal neovascularization (41). Using the rat OIR model,
a trend toward increased LHP was found in temporal association with the
development of intravitreal neovascularization (42). However, neither
intravitreal neovascularization nor avascular retina was reduced by the broad
antioxidant, N-acetylcysteine (NAC), given at a dose that inhibited LHP
production in the retina (42). However, in a mouse model of OIR, NAC given
during hyperoxia reduced both vaso-obliteration and later intravitreal
Treatment with vitamin C or E (43) or liposomal superoxide dismutase
(44) reduced peripheral avascular retina and vascular leakage and increased
capillary density compared to sham-injected controls but did not reduce
pathologic intravitreal neovascularization in the rat OIR model.
Several studies have looked at the effect of the NADPH oxidases on both
avascular retina and intravitreal neovascularization. In both the mouse and rat
OIR models, quenching ROS by apocynin, an antioxidant, reduced avascular
retina by promoting revascularization and inhibiting apoptosis (42,45). Using
the mouse model of OIR, activated NADPH oxidase induced VEGF
expression and intravitreal neovascularization (43) in part, by inhibiting the
anti-inflammatory effect of the transcription factor, peroxisome proliferator–
activated receptor gamma (46). In the rat OIR model rescued with
supplemental oxygen instead of room air, activated NADPH oxidase
mediated intravitreal neovascularization (22), in part, through Janus
kinase/signal transducer and activator of transcription (JAK/STAT) signaling
(47).
Besides NADPH oxidase, other enzymes catalyze ROS generation in
endothelial cells including eNOS and cyclooxygenase. eNOS catalyzes the
release of nitric oxide (NO). NO is an important vasodilator and has both
protective and proangiogenic properties in the eye that preserve endothelial
cell barrier integrity by reducing apoptosis. However, NO can react with ROS
and cause retinal microvascular degeneration by processing nitro-oxidative
stress and generating damaging peroxynitrite (48). Enos−/− mice subjected to
the OIR model developed less avascular retina and intravitreal
neovascularization, and overexpression of eNOS increased intravitreal
neovascularization, providing evidence of the importance of eNOS and nitro-
oxidative stress in ROP (48,49). However, there was no change in retinal
VEGF levels in either Enos−/− mice or eNOS-GFP mice with eNOS
overexpression, suggesting that eNOS-regulated nitro-oxidative stress did not
mediate VEGF expression in this model (48). In the rat OIR model, activated
eNOS was found in association with increased arteriolar tortuosity. However,
eNOS was not found to be activated by VEGF. Reactive nitrogen species,
such as NO2*, can isomerize arachidonic acid to trans–arachidonic acid,
which contributes to hyperoxia-induced vaso-obliteration by up-regulating
the antiangiogenic agent, thrombospondin-1, in the mouse OIR model.
Retinal polyunsaturated fatty acids, prevalent particularly in photoreceptor
outer segments, are susceptible to trans–arachidonic acid formation when
arachidonic acid interacts with reactive nitrogen species (50). Therefore,
evidence suggests that ROS signaling can cause pathologic features
independent of and in association with VEGF signaling.
Bringing It Back to the Infant: Clinical Studies of

Antioxidants on ROP
The signaling of oxidative compounds is evolving and complex, but given the
preterm infant’s predisposition to increased ROS and the evidence from
animal models of oxygen stresses, it appears that oxidative stress does
contribute to features of severe ROP. Several clinical studies were performed
to test the effect of vitamin E supplementation on ROP in infants, but many
of the studies were inconclusive or stopped because of complications,
including neonatal sepsis and necrotizing enterocolitis (51). Clinical trials
tested the effects of antioxidants in preventing ROP. Recombinant human
Cu/Zn superoxide dismutase (52) or vitamin E (53) reduces the risk of ROP
in extremely low gestational age or extremely low birth weight newborn
infants; however, this beneficial effect is not supported by studies with other
antioxidants, such as NAC, lutein, and zeaxanthin. A clinical trial testing
low-dose NAC in premature infants born under 1,000 g birth weight for the
first 6 days after birth showed no significant difference in survival, body
weight gain, respiratory disease, or ROP (54). Randomized and double-blind
clinical studies tested the effect of lutein administered to preterm infants born
≤32 weeks’ gestational age found no effect on the incidence of ROP at any
stage compared to controls (55,56). Future studies are, therefore, needed to
confirm the benefits of antioxidants to reduce ROP.
Vascular Endothelial Growth Factor and Other

Angiogenic Factors
A number of angiogenic factors are involved in models of OIR and include
VEGFA, erythropoietin (EPO), insulin-like growth factor 1 (IGF-1),
angiopoietins, stromal-derived factor, and hypoxia-inducible factor-1α (HIF-
1α). Of these, VEGFA is perhaps the best studied in the pathogenesis of
ROP. VEGFA was identified originally as a vasopermeability factor and later
as an angiogenic factor (57). The human VEGFA gene produces at least six
proangiogenic splice variants through alternative mRNA splicing (58). Due
to differences in the affinity for heparin binding, the splice variants were
found to have different biologic functions (59). VEGF165 is a predominant
splice variant and creates a chemoattractant gradient for migrating endothelial
cells during retinal vascular development (60), but also causes inflammatory
leukostasis and endothelial apoptosis, the signaling events that are involved
in avascular retina and pathologic angiogenesis in OIR (61). VEGF promotes
angiogenesis by signaling through its receptors (VEGFRs). There are three
known VEGFRs and several coreceptors, including VEGFR1 (Flt-1),
VEGFR2 (KDR/Flk-1), VEGFR3 (Flt4), and the neuropilins (NRP1 and
NRP2). VEGFR1 and VEGFR2 both have very high affinity for VEGF
binding, and VEGFR3 only binds VEGFC and VEGFD. All the VEGFRs are
members of receptor tyrosine kinases. VEGFR2 has strong tyrosine kinase
activity and is the receptor believed to be most involved with angiogenic
processes (62). VEGFRs are located on retinal vascular endothelial cells,
choroidal capillaries, photoreceptors, and Müller cells, but in the early
developing retina of mouse, VEGFR1 and VEGFR2 are also localized in
retinal neurons of the inner nuclear and ganglion layers. NRPs act as
coreceptors for specific VEGFRs. NRP1 is a coreceptor for VEGFR2 and
enhances the binding and biologic activity of VEGF165.
An increase in VEGFA protein was found in the ocular fluid of human
infants with ROP (63), and VEGF mRNA was detected in the avascular
retina of a human infant with stage 3 ROP (60). Inhibition of VEGF
bioactivity with a VEGF antibody reduced severe ROP (64). However,
VEGF is also essential in retinal vascular development and as an endothelial
and neuronal survival factor (65). In experimental models, VEGF signaling
regulates the number and length of filopodia in endothelial tip cells (66). The
filopodia are considered the guidance-seeking protrusions from neurons and
may serve a similar role in endothelial tip cells at the migrating front during
retinal vascular development. VEGF signaling also interacts with the delta-
like ligand 4/Notch1 signaling pathway, regulating the endothelial tip-to-stalk
cell ratio at the junction of the vascular and avascular retina. This ratio is
important in ordered developmental angiogenesis, and proliferation of stalk
cells causes intravitreal neovascularization (67). An axon guidance signaling
Slit2/Roundabout 4 (Robo4) pathway has been found important in stabilizing
retinal vessels by inhibiting VEGF-induced pathologic angiogenesis (68).
Evidence exists that the receptors VEGFR1 and VEGFR2 are required for
normal neural retinal development (69).
VEGF is up-regulated by a number of stimuli that affect preterm infants,
including hypoxia, ROS, and inflammatory cytokines. Hypoxia-induced
VEGF expression is regulated by mechanisms including transcriptional
activation, alternative translational initiation, and mRNA stabilization. The
transcriptional activation involves stabilization of the subunit HIF-α. In
normoxia, HIF-α is hydroxylated by prolyl hydroxylase enzymes (70).
However, hypoxic conditions lower the activity of prolyl hydroxylases,
preventing HIF-α from hydroxylation. In the mouse OIR model, inhibition of
prolyl hydroxylases promoted stabilization of HIF during hyperoxia, which
then reduced vaso-obliteration and subsequent endothelial budding into the
vitreous and reduced vessel tortuosity in the model (71).
In the mouse OIR model, high constant inspired oxygen at 75% reduced
retinal VEGF expression in association with vaso-obliteration (Figure 41-2).
Prior to this phase, if VEGF (72,73), IGF-1 (74), IGF-1–binding protein 3
(IGFBP3) (75), EPO (76), omega-3 fatty acids (77), or certain peptides (78)
were given to mice, vaso-obliteration was reduced. When mouse pups were
then placed into room air following high oxygen, relative tissue hypoxia
occurred in the vaso-obliterated retina, followed by overexpression of VEGF
(79) and EPO (80) and endothelial budding above the internal limiting
membrane.
In the rat OIR model, repeated fluctuations in oxygen cycled between
50% and 10% increased retinal VEGF164 (rat homolog to human VEGF165)
expression at both mRNA and protein levels beginning at day 8 of life
compared to that in the pups raised in room air or to a single hypoxic episode
(10% O2) (81). Retinal VEGF was increased in association with persistent
avascular retina, suggesting that increased VEGF was not able to support
ongoing physiologic retinal vascularization. When rat pups were returned to
room air from oxygen cycling, retinal VEGF was increased compared to that
in the same-aged pup raised in room air, in association with intravitreal
Retinal hypoxia has been measured in the rat OIR model following
repeated fluctuations in oxygen and in the mouse OIR model when mice are
brought out of hyperoxia into room air (22,23). Retinal hypoxia can cause
VEGF-induced aberrant intravitreal neovascularization (46,47,73,79,83). In
the beagle model, a low dose of a “VEGF trap” reduced intravitreal
neovascularization without adversely affecting ongoing retinal vascular
development; however, a high dose caused persistent avascular retina (83). In
the mouse OIR, inhibition of VEGF reduced endothelial budding into the
vitreous during the vasoproliferative phase and did not appear to affect the
avascular retina; however, the area of avascular retina was not quantified
(84). In the rat OIR model, inhibition of retinal VEGF by a neutralizing
antibody against VEGF164 significantly reduced clock hours of intravitreal
neovascularization by 50% and did not interfere with retinal vascular
development (46). At first, the finding that an antiangiogenic compound
would not interfere with ongoing vascular development at certain doses
seemed counterintuitive.
However, further study showed that VEGF neutralization that reduced
signaling through VEGFR2 affected the orientation of dividing vascular cells.
The orientation of the cleavage planes of dividing endothelial cells to the long
axes of vessels predicts the future location of daughter endothelial cells that
extend vessels (Figure 41-3). If cells divided and the cleavage planes were
parallel to the long axis of the vessels, then the resulting vessels were dilated.
If the cleavage planes of the dividing endothelial cells were perpendicular to
the long axis of the vessels, vessels were elongated. When cleavage planes
were disordered relative to the long axis of the vessels, the result was
disordering of the vasculature. A single allele knockout of a splice variant or
receptor in the VEGF signaling pathway is lethal. Therefore, to study the
effect of excessive VEGF/VEGFR2 signaling on endothelial cell divisions,
investigators used an embryonic stem cell model that had an Flt-1 knockout
(knockout to VEGFR1). VEGFR1 acts as a decoy receptor in development,
limiting the amount of VEGF available to signal through the angiogenic
receptor, VEGFR2. In the Flt-1 knockout model, VEGF could not bind
VEGFR1 and triggered signaling through VEGFR2. In the flt1−/− model,
cleavage angels were not clustered in one direction but instead were
disoriented. This led to a pattern of growth similar to the appearance as seen
in human ROP with plus disease. Ordered angiogenesis was restored in the
flt-1−/− when a CD31 transgene containing flt-1 was introduced to the model,
thus restoring the ability to bind excess VEGF and limit angiogenic signaling
(85). In an analogous study in the rat OIR model, the angles between
cleavage planes of dividing vascular endothelial cells and the long axes of the
vessels (i.e., cleavage angles) were found to be less likely to predict
lengthening of the retinal vessels (Figure 41-3). Vessels were also more
tortuous in the rat OIR model than those in room air. A neutralizing antibody
to VEGF, given at a dose found to reduce intravitreal neovascularization,
restored cleavage angles to predict lengthening of vessels and reduced
tortuosity (47). This study suggests a way in which excessive
VEGF/VEGFR2 signaling may lead to plus disease (83).
FIGURE 41-3 Diagram of the orientation of the cleavage
planes of dividing endothelial cells and vascular
development. Vessel elongation results when the cleavage
planes of the dividing endothelial cells are perpendicular
to the long axis of the vessels, and when the cleavage
planes of dividing endothelial cells are parallel to the long
axis of the vessels, the vessels are elongated.
Pups treated with an intravitreal neutralizing antibody to rat VEGF at a dose

effective to inhibit intravitreal neovascularization had recurrent intravitreal
neovascularization, reduced serum VEGF, and less postnatal weight gain than
did pups treated with an intravitreal injection of IgG (86). These findings
raised safety concerns about intravitreal agents to inhibit VEGF to treat ROP.
Scientists sought a way to identify an optimal anti-VEGF dose to inhibit
intravitreal neovascularization and preserve or promote physiologic retinal
vascular development safely. One way to inhibit intravitreal
neovascularization safely is to target the cells that overproduce VEGF. In the
rat OIR model, targeted inhibition of Müller cell–derived VEGFA by short
hairpin RNA mediated–knockdown reduced intravitreal neovascularization
without interfering with physiologic retinal vascular development. In
addition, targeted knockdown of up-regulated VEGF in Müller cells
preserved retinal physiologic vascularity in vascularized retina in the primary
and deep plexi, whereas anti-VEGF antibody reduced physiologic vascularity
in the inner and deep plexi in association with recurrent intravitreal
neovascularization (87). This finding suggests a mechanism whereby high
anti-VEGF dose in human infants leads to recurrent intravitreal
neovascularization.
In the rat OIR model, phosphorylated VEGFR2 was increased in the
retina of OIR pups when maximal intravitreal neovascularization occurred.
However, VEGFR2 is expressed not only on retinal vascular endothelial cells
but also on other retinal cells, making broad inhibition of retinal VEGFR2
potentially unsafe. Therefore, regulating VEGFR2 in endothelial cells may
safely inhibit intravitreal neovascularization. Studies found that knockdown
of endothelial VEGFR2 using lentiviral-derived shRNA driven by cdh5
promoter inhibited intravitreal neovascularization and extended physiologic
retinal vascular development without adverse effects on retinal function (88).
This finding together with earlier studies measuring cleavage angles in the
flt1−/− model suggests that excessive VEGF/VEGFR2 signaling disorients
dividing endothelial cells and allows vessels to become intravitreal
neovascularization while interfering with extension of vessels into the
peripheral retina. Regulation of VEGF signaling through VEGFR2 in
endothelial cells inhibits intravitreal neovascularization and permits extension
of physiologic retinal vascular development.
Erythropoietin
Erythropoietin (EPO) is believed to be an oxygen-regulated hormone,
produced by the kidney in response to ischemia and hypoxia. EPO promotes
erythrocyte formation in the bone marrow. Recombinant EPO is used to treat
anemia in patients with chronic kidney failure secondary to diabetes mellitus
or preterm infants with anemia of prematurity. In these groups of patients,
some studies have found an associated increased risk of severe ROP with
EPO use (89). In the mouse OIR model, retinal EPO expression was
suppressed during hyperoxia-induced vaso-obliteration and elevated during
intravitreal neovascularization (76). Exogenous EPO administered during
hyperoxia prevented both hyperoxia-induced vessel loss and later hypoxia-
induced intravitreal neovascularization and retinal neuron apoptosis;
however, exogenous erythropoietin treatment during hypoxia-induced
proliferation did not protect against retinal vessel loss and worsened
pathologic intravitreal neovascularization (90).
In the rat OIR model, down-regulation of retinal EPO contributed to
avascular retina prior to the development of intravitreal neovascularization.
The mechanism for the reduced expression appeared to be activation of the
JAK/STAT pathway (91). STAT3 was activated in Müller cells by increased
retinal VEGF following repeated oxygen fluctuations. Activated STAT3
delayed retinal vascularization by down-regulating EPO mRNA expression in
Müller cells. Exogenous EPO given early at days of life 2, 4, and 6 increased
peripheral retinal vascularization without causing increased intravitreal
neovascularization (91). Others reported that EPO recruited proangiogenic
bone marrow–derived progenitor cells and activated NF-κB signaling by the
activated EPO receptor on retinal vessels and neurons (76). In a combined rat
model of maternal UPI and OIR, increased circulating EPO was associated
with extension of physiologic retinal vascular development and reduced
pathologic intravitreal neovascularization in rat pups (4). However, EPO has
been associated with worse OIR features experimentally depending on timing
of administration.
Bringing It Back to the Infant: Clinical Studies
Anti-VEGF
The current approved treatment for severe ROP is laser (preferred to
cryotherapy) delivered in a nearly confluent pattern to the peripheral
avascular retina (see Chapter 52). Even though these treatments reduce the
risk of untoward structural outcomes and poor vision, peripheral vision lost is
still inevitable in some cases. Several clinical studies tested the effect of a
complete VEGFA-specific neutralizing antibody on severe ROP. Initial
results appeared promising with reduction in intravitreal neovascularization
(stage 3 ROP), plus disease, and avascular retina (64). However, recurrent
intravitreal neovascularization (92), persistent avascular retina and adverse
outcomes including reduced serum free VEGF and IGF-1 (93) are being
reported. There is also concern about fluorescein angiographic evidence of
areas of nonperfused retina within the already vascularized central retina in
infants treated with bevacizumab (94). Greater study of the inhibitory effects
of different VEGF doses or agents on pathologic and normal vascularization
is needed. Several clinical trials have tested de-escalating doses of
bevacizumab and have compared ranibizumab to laser and are covered in
greater detail (95) (see Chapters 38 and 53).
Insulin-Like Growth Factor

IGF-1 is a hormone essential for fetal development during all the stages of
pregnancy and is signaled via IGF-1 receptor (IGF-1R). Plasma
concentrations of IGF-1 increase with gestational age, rising during the third
trimester of pregnancy and peaking at full-term birth (96). In utero, IGF-1 for
the fetus is partly supplied by the maternal and placental circulation. When
infants are born preterm, they are unable to produce sufficient IGF-1 and,
therefore, have reduced levels due to loss of maternal and placental supplies
(97). Investigators developed an algorithm to predict the risk of ROP based
on serum levels of IGF-1 and IGFBP3 and postnatal weight gain (98). They
found evidence that low serum levels of IGF-1 might predict the risk of
severe ROP in some populations. This model has subsequently been revised
to include only postnatal weight gain as a risk of ROP. A phase 2 clinical trial
to test the effect of IGF-1 on reducing ROP did not find beneficial effects of
ROP from treatment with human recombinant IGF-1 and IGF-1 binding
protein-3 (99). However, ongoing studies to assess its role in preventing
intraventricular hemorrhage are being performed (see Chapter 41).
Erythropoietin in ROP and Cognitive Development

With greater survival rates of extremely low birth weight infants, it has
become recognized that these infants are at high risk of not only ROP but
also later neurodevelopmental impairment. EPO use is gaining interest as a
neuroprotective agent. A previous study reported that recombinant EPO
given to preterm infants was associated with better cognition in childhood
(84). From the laboratory studies, a protective effect of early EPO use was
found in both the mouse and rat OIR models, suggesting that early treatment
may be beneficial. Ongoing clinical trials are being performed to test the role
of EPO in preterm infants on cognitive development and on ROP (100).
Neural and Vascular Interaction in ROP Pathogenesis

Although abnormal retinal vasculature is the most recognized feature of ROP,
dysfunction of the neural retina has been reported in OIR models and in
human preterm infants. In the rat OIR model, abnormalities of rod structure
and function antedate retinal vascular abnormalities and persist after vascular
abnormalities have mostly resolved. Deficits in rod sensitivity are
documented in infants and children with a history of ROP (101). In a rat
model of high (80%) oxygen followed by room air exposure, there was
thinning of the ganglion cell and inner nuclear and plexiform layers and
abnormalities in amacrine, bipolar, and Müller cells during hypoxia and
particularly noted in the peripheral retina (102). A midperipheral “transition
zone” existed wherein neurons and glia were affected differently depending
on their local vascular supply. Furthermore, evidence of changes in
metabolism within the retina has been shown by magnetic resonance imaging
in the rat OIR model (103).
As the hyaloidal circulation regresses and metabolically active
photoreceptors and other neuronal cells have differentiated and demand
oxygen (101), the incompletely vascularized retina becomes hypoxic (23,25).
Metabolic demand from photoreceptors begins posteriorly and proceeds
peripherally. Infants born at young gestational ages may manifest aggressive
posterior ROP at young postgestational ages for this reason. Following
apparently successful laser treatment, new stage 3 ROP can develop around
37 weeks’ postgestational age when oxygen demand increases in the
peripheral retina as photoreceptor differentiation extends peripherally and as
the retina expands in area stretching regions of untreated hypoxic retina
between laser spots. Further, laser can cause regression of flat
neovascularization that occurs posteriorly creating new “skip” lesions of
hypoxic retina. These events may explain the often recurrent stage 3 ROP
seen in infants following laser for aggressive posterior ROP.
Besides oxygen-related events, there is also evidence that several of the
neural guidance molecules play a role in the orientation and growth of blood
vessels within the retina or vitreous. Most studies have been done on mice.
Semaphorin 3A, which can compete for VEGFR2 (104), has been shown in
association with vaso-obliteration and later intravitreal neovascular budding
in the mouse OIR (105). Semaphorin 3E binding its receptor, plexin D1, was
found to be important in restoring normal endothelial orientation and retinal
vascularization during hypoxia-induced VEGF expression (106).
ACKNOWLEDGMENT
We thank James Gilman, CRA, FOPS, for his help in creating the figures and
table.
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42
Signaling Pathways in Wnt and
Effects on Disease
Eric Nudleman, Geoffrey Weiner, and Kimberly A. Drenser
OVERVIEW OF WNT SIGNALING

The Wnt/β-catenin signaling pathway is an evolutionarily highly conserved
pathway that regulates embryogenesis and homeostasis in adult tissues
(reviewed in Ref. (1)). Wnt was first discovered in mice, and the critical role
it plays in organismal development was first appreciated in Drosophila
melanogaster mutants (2–5). Since its discovery more than 35 years ago, the
Wnt/β-catenin pathway has been demonstrated to play a role in many
biologic processes, including specifying body axis in mammals (6), stem cell
function (7–9), and mammalian central nervous system (CNS) development
(10,11). In the eye, Wnt signaling has been linked to familial exudative
vitreoretinopathy (FEVR) and Norrie disease, and emerging evidence
suggests that Wnt dysregulation may play a role in more prevalent diseases
like diabetic retinopathy, age-related macular degeneration, and retinopathy
of prematurity (ROP).
Molecular Pathways in Wnt Signaling

Wnt signaling in mammals has been studied extensively (reviewed in Ref.
(12)). Broadly, the Wnt pathway consists of an extracellular ligand capable of
binding to a multiprotein receptor complex on a target cell, an event that
leads to intracellular changes that promote the stabilization of the
transcription factor, β-catenin. The Wnt ligand is a hydrophobic, lipid-
modified (13,14) protein secreted into the extracellular space. Signal
transduction is regulated in the cytoplasm by the interaction of many distinct
Wnt pathway–associated proteins, and these inputs converge on stabilization
of β-catenin. Stabilized β-catenin translocates to the nucleus and activates the
transcription of Wnt pathway–associated genes with the corresponding
proteins affecting multiple downstream cellular changes (15).
When no Wnt ligand is present, β-catenin is constitutively degraded by a
process involving N-terminal phosphorylation by glycogen synthase kinase-3
(GSK-3) and casein kinase 1 (CK1), ubiquitination, followed by proteasomal
degradation (16,17). Wnt ligands bind to a receptor complex composed of a
frizzled (FZD) and lipoprotein receptor–related protein (LRP) at the plasma
membrane (18,19). This binding results in the phosphorylation of LRP (20),
which allows the membrane- associated receptor to sequester the intracellular
protein complex containing Axin, adenomatous polyposis coli (APC), GSK-
3, and CK1 (reviewed in Ref. (21)). The Axin/APC/GSK-3/CK1 complex
constitutively degrades β-catenin in the cytoplasm; thus, sequestering this
complex in response to WNT/FZD/LRP interaction allows β-catenin to
accumulate in the cytoplasm and translocate to the nucleus (22–24). In the
nucleus, β-catenin binds to T-cell factors/lymphoid-enhancing factors
(TCFs/LEFs) that, as a complex, activates the expression of a number of
target genes (25–27).
The general architecture of the Wnt/β-catenin signaling pathway is
conserved across tissues and organisms, but the specific Wnt ligand and the
LRP/FZD complex are varied and confer tissue specificity (reviewed in Ref.
(28)). There are 19 known WNT ligands, 10 FZD receptors, and 2 LRPs. In
the CNS, Wnt signaling is known to be region specific with different WNT
ligands regulating vascular development and blood–brain barrier/blood–
retina barrier functions in different brain or retinal regions (29). Furthermore,
the expression of WNT genes is highly dynamic during development (30–32).
WNT SIGNALING IN THE

DEVELOPMENT OF RETINAL
VASCULATURE
Wnt signaling is known to regulate the development of the mammalian CNS.
There are Wnt signaling proteins specific to the retina, one of the most well-
studied being Norrin, encoded by the NDP gene. Norrin, a cysteine-knot
protein, is a Wnt ligand in the retina despite not being a classical Wnt protein
(33–36). It is a member of the mucin-like subgroup of 10-membered
cysteine-knot proteins. The cysteine-knot motif is highly conserved in many
growth factors, including transforming growth factor-β, human chorionic
gonadotropin, nerve growth factor, and platelet-derived growth factor. Norrin
has two primary domains, a signal peptide that directs localization of the
molecule and a cysteine knot that provides the structural conformation
required for receptor binding and activation of signal transduction. Structural
determination supports the importance of the cysteine residues and their
disulfide bonds in the structural conformation and functional capacity of
Norrin (33,37,38). During development, Norrin is secreted by Müller glia and
regulates development of the retinal vasculature (39–42) and the blood–retina
barrier (BRB) (33). Wnt receptor complexes are generally composed of
LRPs, FZDs, and other coactivators. The LRP expressed in the retina is LRP5
(43,44), and the FZD receptor is FZD4 (39). Additionally, TSPAN12 is a
Norrin-specific coreceptor for the FZD4/LRP5 complex (45). Studies in
knockout mice have demonstrated that Norrin, LRP5, and TSPAN12 are
essential to normal retinal vascular development, as well as BRB function. In
animal models, mutations in any of these genes result in an avascular
peripheral retina, BRB dysfunction, as well as a reduction in the development
of the deep vascular plexi. Rather than forming a superficial vascular plexus
within the nerve fiber layer, and two additional plexi surrounding the inner
nuclear layer, mutations in the Wnt pathway result in superficial vessels that
form blunted endings and fail to form the deep plexi (39,45–47). Thus, the
retina expresses its own unique complement of Wnt pathway genes that
regulate vascular development and function.
NORRIE DISEASE AND FEVR ARE

WNT SPECTRUM DISORDERS
History and Overview
Although classically considered to be two distinct diseases, recent discoveries
in the genetics of FEVR and Norrie disease (ND) have led to the
understanding that both of these inherited retinopathies are due to defects in
the Wnt signaling pathway and thus are more appropriately classified as Wnt
spectrum disorders. Their asynchronous description and spectrum of clinical
features have obscured the true relationship between FEVR and ND. ND is a
rare X-linked recessive disorder with clinical manifestations extensively
characterized (48–52), whereas FEVR has multiple inheritance patterns,
although predominantly autosomal dominant, first described by Criswick and
Schepens (53). Because FEVR was reported after the description of ROP and
the ocular findings resembled those in ROP but were observed in full-term
babies, it has been referred to by some as ROP in full-term infants.
In ND, ocular manifestations are often severe and are the presenting
finding at the time of diagnosis. Additionally, nearly all patients develop
hearing loss by the third decade of life, and one-third to two-thirds have
cognitive impairments (38,54–56). Boys with Norrie disease (ND) can have
limited vision with no light perception possible from birth or shortly
thereafter. Findings in infancy may include leukocoria due to dense
retrolental plaque, iris atrophy, vitreous and subretinal hemorrhage, and
dysplastic retinae with folds and detachments (Figure 42-1). Classically,
findings in FEVR are less severe. FEVR is an abnormality of the retinal
vasculature resulting in a disruption in the development of the peripheral
retinal vessels, leaving varying degrees of avascular retina. The disease is
often asymmetric; however, it is always bilateral (Figure 42-1). Recent
evidence has demonstrated that FEVR is not only an abnormality or retinal
vascular development; it is a lifelong active retinal vascular disease with
variable periods of quiescence. At the juncture of avascular and vascularized
retina, there often is a line of vascular buds that are distinct from the typical
neovascularization of ROP and the telangiectatic vessels of Coats disease.
FEVR has great variability in presentation, course, and underlying genetics.
Benson (57) reported the natural history of FEVR, noting that people who
present with FEVR prior to age 3 years have a worse prognosis and that there
can be very long (a decade of more) quiet periods.
FIGURE 42-1 Presentations of FEVR are highly variable.
Shown are images from three patients with mutations in
NDP. A–C: Bilateral leukocoria, with higher
magnification view of the right eye (B) and left eye (C). A
dense retrolental plaque is present with dragging of the
ciliary processes centrally. D–E: Asymmetric disease is
common. In this patient, the right eye (D) demonstrates an
avascular periphery with hemorrhage, a retinal fold
involving the inferotemporal arcade, and extensive
subretinal exudate. The left eye (E) has a total retinal
detachment with subretinal hemorrhage and exudate. F–G:
Fluorescein angiography is often critical to determine the
extent of avascularity. In this patient, the right eye appears
unaffected in the color image (F). The left eye (G) has
retinal fold involving the macula. Fluorescein angiography
demonstrates extensive temporal avascularity in the right
eye (H) with a bud of neovascularization and mild
temporal dragging of the retinal vessels. Extensive
peripheral avascularity is also evident in the left eye (I),
with leakage from the vessels within the fold.
Genetics
The Wnt pathway in the retina consists of Norrin, FZD4, LRP5, and
TSPAN12. Mutations in all of those genes have been described to cause one
of the ND/FEVR spectrum disorders. ND is associated with mutations in the
NDP gene, which is located on the short arm of chromosome X at position
11.3 and thus is inherited in an X-linked recessive pattern (58–60). Note that
mutations in NDP also cause EVR2 or X-linked EVR, reflecting
nomenclature that predates the cloning of NDP. FEVR can be associated with
mutations in FZD4, LRP5, or TSPAN12, and the specific mutation determines
the inheritance pattern (58,61,62). Mutations in FZD4, located at 11q14.2,
cause EVR1 and are autosomal dominant (63). Mutations in LRP5, located at
11q13.2, cause EVR4 and can be autosomal dominant or recessive (64).
Mutations in TSPAN12, located at 7q31.31, cause EVR5 and are also
autosomal dominant (62,65). Overall, the most common inheritance pattern
for FEVR is autosomal dominant. (Also see Chapter 66.)
There are 107 single-nucleotide polymorphisms (SNPs) reported in the
NDP gene, 28 of which are nonsense and 64 are missense mutations and
almost always cause ND (59). In vitro studies have confirmed that the most
common pathogenic mutations in NDP result in dysfunctional Norrin protein
and impaired Wnt signaling (38,39,66). Although the cysteine-knot motif
appears to be important in conferring biologic activity to Norrin both
biologically and computationally, untranslated regions within NDP are also
sometimes mutated and have regulatory functions that control the expression
and stability of Norrin (38,67,68). To date, 37 different SNPs in FZD4 have
been described, 12 of which are nonsense and 24 are missense mutations. A
small number of mutations have been tested in vitro, with all mutants
showing at least 30% reduction in activity compared to wild type (39,66,69).
There are 90 known mutations in LRP5, 27 of which are nonsense and 58 are
missense. Many mutations do not result in FEVR but do result in dysfunction
in other body systems, reflecting the widespread expression of LRP5 in
various tissues and developmental stages. When studied in vitro, LRP5
mutants show the most variability in the degree of Wnt pathway inhibition,
likely due to the fact that LRP5 is the target of endogenous Wnt inhibitors
and thus the effect of any given mutation is highly base pair specific
(66,70–72). TSPAN12 is the most recently described FEVR-associated gene,
and only eight mutations are reported, with four of them being nonsense
mutations and two of them missense mutations (65,73).
Although it might be expected that the clinical phenotype of mutations in
the numerous genes of the Wnt pathway would correlate with the known
effects of those mutations, this is not generally observed. This lack of strong
genotype–phenotype correlation could be due to genetic interactions with
other loci in the genome. Or it may suggest that the Wnt pathway subserves
additional functions in the retina that have yet to be understood. Some of
those other functions are just beginning to be investigated. For example,
recent evidence in animal models suggests that Norrin serves a
neuroprotective function (41,42,74).
Clinical Diagnosis
Diagnosis is based on birth history, family history, and clinical examination
and is supported by genetic mutations in the NDP, FZD4, TSPAN12, or LRP5
genes. Classic ND typically presents with leukocoria and poor vision in both
eyes (49,54). Extraocular manifestations (hearing loss and cognitive
impairments) are often identified subsequently (38,54–56). The eyes are of
average size (corneal diameter 9 to 10 mm) initially but typically become
smaller with time as the traction tethering the posterior lens capsule and
posterior retina limits eye growth (Figure 42-1). Incomplete maturity of the
iris vessels is common with elongation of the ciliary processes due to traction
(Figsures 42-1 and 42-2). A fold of retina (resembling a stalk in persistent
fetal vasculature syndrome [PFVS]) may be present and attach anteriorly to
the peripheral retina, ciliary processes, or posterior aspect of the lens with
variable size footplates. The base of the fold generally involves the optic
nerve, often with exudation (Figure 42-2). Unbranched retinal vessels can be
seen coursing through the fold tissue (Figure 42-2). The periphery is
avascular and can have underlying areas of pigment changes.
FIGURE 42-2 Retinal fold in FEVR in a patient with a
mutation in FZD4. A: External view demonstrating
leukocoria in the left eye with a slightly smaller cornea. In
the right eye, a small opacity is visible inferotemporally.
B: Fundus photo of the right eye, demonstrating a tight
(falciform) retinal fold with retinal dragging
inferotemporally. C–E: Fluorescein angiogram of the right
eye. The retinal vessels are seen pulled into the fold (C)
forming a sheet of retina-to-retina apposition. The
remaining retina is attached with some evidence of
vasculature but extensive peripheral retina. D: Insertion of
the fold into the inferotemporal periphery and ciliary
processes. E: Peripheral retinal avascularity. F,G: Wide-
field fluorescein angiography of the patient’s
asymptomatic mother helps to confirm the diagnosis,
demonstrating typical avascular periphery in the left eye
(G).
The FEVR classification system has five stages (Table 42-1) (75). Classic
FEVR features avascular peripheral retina and findings of vascular buds at
the junction of the avascular and vascularized retina. The vessels are often
dragged in the posterior pole, and retinal folds are common (Figure 42-2).
Consistent with animal models, OCT-A has recently demonstrated a
reduction in the vascular density, particularly in the deep vascular plexi, and
blunted vessels in the superficial plexus in humans with FEVR (76–78).
These vessels often exhibit dysfunction of the blood-retinal barrier, and large
amounts of subretinal exudate may be present. In eyes with retinal
detachment, both traction from fibrosis and subretinal exudate can be seen.
Histologically, inflammatory elements are found that reflect the chronic
process (79). A positive family history is very helpful in making a diagnosis
but not essential. The lack of a diagnosis of FEVR in family members does
not rule out a diagnosis of FEVR, either because of de novo mutations or
because of undiagnosed asymptomatic FEVR (80). Identical mutations
causing FEVR can have variable expression and incomplete penetrance.
Therefore, if the diagnosis of FEVR is suspected, a thorough peripheral
retinal examination including wide-field fluorescein angiography is
recommended for all family members (Figure 42-2).
TABLE 42-1 Differential diagnosis
Presentations of FEVR are both variable and asymmetric. It is not unusual for
one eye to present with advanced disease while the fellow eye has an
asymptomatic avascular periphery. Therefore, examination of the fellow eye
is essential to confirming the diagnosis. In a patient referred with presumed
PFVS based on microphthalmia and white pupil in one eye, it is prudent to
perform an EUA with a wide angle FA to assure the fellow eye is normal
(81). If FEVR is suspected in patients presenting with retinal dysgenesis
and/or disorganized retinal detachments, genetic testing should also be
offered, which is now done most cost-effectively by whole genome
sequencing (WGS). A mutation in one of the known genes is helpful to
confirm the diagnosis. However, WGS fails to identify a mutation in one of
the known genes in approximately one-half to one-third of cases (73,82) and
does not definitively rule out FEVR. Rather, the results may be helpful in
identifying novel new variants. Patients with ND/FEVR may benefit from
genetic counseling to better understand the implications for themselves and
other family members.
Clinical Course
Warburg’s seminal study of ND described only rare cases with vision beyond
infancy (83). Out of 24 patients in her series, all had no light perception
except for one boy who could count fingers until age 12 when light
perception was lost and another boy who could perceive light. In Warburg’s
review of the literature, she identified an additional 106 patients with ND. Of
these, only six patients were noted to have light perception or pupillary
reactivity after 3 months of age. Untreated, this process will continue to
contract resulting in a retrolental plaque with pseudogliosis and eventual
phthisis.
However, the FEVR patient population is characterized by a spectrum of
disease presentation (75). FEVR eyes that present in the first year of life have
a worse prognosis, but many children present later in life due to asymmetric
disease identified during school vision screening or strabismus. With
appropriate management (discussed in detail in Chapter 66), improved vision
can be achieved. FEVR likely is underdiagnosed, with earlier milder stages of
FEVR often asymptomatic, and advanced disease commonly confused for
other retinal disorders (Table 42-2). Therefore, a thorough family history,
wide-field angiography, and genetic testing have become invaluable tools in
distinguishing between disease entities.
TABLE 42-2 Clinical classification of familial

exudative vitreoretinopathy
RD, retinal detachment.
Treatment
Surgical approaches to ND/FEVR will be addressed in Chapter 66. Here we
consider the evidence for ablative therapy, anti-VEGF agents, and emerging
therapies. FEVR often has a progressive long-term course, with periods of
waxing and waning, and requires lifelong screening, particularly with new
symptoms such as reduced vision or floaters. In animal models of FEVR,
elevated expression of vascular endothelial growth factor (VEGF) have been
measured (84,85). It is likely that avascular retina and capillary dropout
contribute to this finding. Because the stimulus for neovascularization in the
avascular retina is believed to be similar to that in ROP, treating the avascular
peripheral retina with laser ablation is recommended if hemorrhage, or
leakage on fluorescein angiography, is present. In children younger than 3
years with FEVR of stage 2 or greater (Table 42-2), treating any avascular
retina is recommended. Laser ablation is performed using a pattern similar to
the retinal ablation for ROP, adding near-confluent spots, one-half spot size
from one another (see also Chapter 58). It is not recommended to monitor
these eyes until they develop exudates. Rather, treatment is recommended to
prevent exudation before it occurs, because ablative therapy becomes less
effective once subretinal fluid and exudation develop. Drainage of subretinal
fluid followed by laser or cryotherapy often is disappointing, because the
fluid is thick and drainage incomplete. Aggressive treatment of avascular
retina with laser ablation results in improved long-term control. Small studies
of anti-VEGF therapies have reported improved outcomes, but larger trials
are needed (86–89). In the future, new pharmacologic therapy may be the
best way to approach ND/FEVR to offset the effects of growth factors and
abnormal Wnt signaling. As the genetics of ND/FEVR are further explored,
the disease may become a candidate for modulation with specific molecules,
such as adding Norrin to conditions that have mutatons in NDP.
EMERGING CONNECTIONS BETWEEN

WNT SIGNALING AND OTHER
RETINAL DISORDERS
Wnt signaling has been implicated in ROP pathogenesis (90). Mutations in
NDP and FZD4 are associated with ROP and IUGR (91–93). The picture of
Wnt signaling in ROP is significantly muddled by diagnostic uncertainty. It
appears that mutations in Wnt pathway genes are associated with preterm
labor, which would put them at risk for ROP. However, the high-risk Wnt
pathway variants may also put them at risk for FEVR. At this time, it is
difficult to distinguish ROP due to preterm labor from FEVR due to a
pathologic Wnt variant in a premature child. In the oxygen-induced
retinopathy (OIR) mouse model of ROP, exogenous Murine protein helps
normalize vessels (94) even though other Wnt ligands are overexpressed in
abnormal vessels, suggesting complex and widespread dysfunction of Wnt
signaling in this disorder (47). The interaction of genetic Wnt variation on
premature labor, ROP, and FEVR is an important issue to clarify in future
research.
Osteoporosis Pseudoglioma Syndrome
Osteoporosis pseudoglioma syndrome (OPPG) is a FEVR variant that is due
to mutations in LRP5 (95). It is described here, because it has a distinct
clinical presentation but can be considered part of the FEVR spectrum. In
addition to the classic and variable manifestations of FEVR, OPPG patients
present with child-onset osteoporosis, skeletal thinning, and fractures. This is
due to the fact that Wnt signaling plays an important role in bone homeostasis
(96). The mutations in LRP5 causing OPPG have been to be both compound
heterozygote and autosomal recessive (97,98). At this time, it is not clear
whether specific LRP5 mutations are sufficient to distinguish OPPG from
nonosteoporotic forms of FEVR or whether other genes contribute to
producing OPPG or FEVR in different patients.
Coats Disease
Coats disease is a congenital retinopathy that presents with unilateral
telangiectasia, microaneurysm, and lipid exudates (see also Chapter 64). It
typically presents in males in the first to second decade of life. Most cases are
sporadic, and although the genetic driver is not definitively established,
somatic mutations in NDP have been reported in retinal tissue from a Coats
patient (99), and in one case, a FZD4 polymorphism was present (100).
However, the majority of patients with Coats disease do not have identifiable
Wnt pathway mutations or polymorphisms known to be pathologic. It is
possible that somatic mutations in Wnt-associated genes are drivers for the
disease, but at this point, the link between Coats disease and Wnt is not
definitively established.
Coloboma
Coloboma is the result of variably incomplete closure of the optic fissure or
choroidal fissure during development. It is associated with a number of
systemic congenital malformations. In animal models of Wnt pathway
dysfunction, coloboma is also sometimes present (101,102). In at least one
case, a mutation in FZD5 has been found to cause autosomal dominant
coloboma (103). Given the locus heterogeneity of mutations associated with
this condition, it is likely that mutations in the many pathways that contribute
to patterning during development are capable of causing coloboma, including
Wnt and others (see also Chapter 22).
Persistent Fetal Vasculature Syndrome

Familial exudative vitreoretinopathy (FEVR) patients can present with a
persistent fetal vasculature. However, PFVS is typically an isolated finding
with an independent underlying cause. Many cases are sporadic with no
known causative mutation. There are several cases reported of PFVS
associated with mutations in NDP; these are likely cases of patients who fall
somewhere on the Wnt spectrum described above. Inherited PFVS has been
reported, more common in Middle Eastern families, where the causative
mutations have been traced to a noncoding region of the genome that affects
the expression of the gene ATOH7 (104). The fact that fetal vasculature is
persistent in ND/FEVR and in Wnt pathway knockouts in animals raises the
question of whether ATOH7 is a Wnt-associated gene. At this time, this
question is unresolved and no mutations in ATOH7 have been associated with
FEVR. (See also Chapter 65.)
Microcornea, Posterior Megalolenticonus, PFVS, and

Coloboma Syndrome
Microcornea, posterior megalolenticonus, PFVS, and coloboma (MPPC) is an
extremely rare condition that presents with bilateral microcornea, posterior
megalolenticonus, PFVS, and coloboma (105). The clinical presentation ties
together several of the elements of ND/FEVR, PFVS, and coloboma
described above. No causative mutations have been definitively established,
although a recent report identifies a likely pathologic variant in ZNF408
associated with MPPC (106). Interestingly, a mutation in ZNF408 was also
reported in a pedigree with autosomal dominant FEVR, and in an animal
model knockout, development of the retinal vasculature was impaired (107).
Whether ZNF408 is a bona fide Wnt pathway gene remains to be established.
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43
Hypoxia-Inducible Factors and
Prevention of ROP
Jonathan E. Sears, and George Hoppe
INTRODUCTION
Survival after premature birth requires oxygen supplementation that is
simultaneously toxic to some premature developing tissues (see also Chapter
40). The fetal circulation is designed to provide oxygenation to the brain
while decreasing negative outcomes of oxygen toxicity. This is achieved by
creating a large oxygen gradient across the placenta. Maternal oxygen
concentration preplacenta is 90 to 100 mm Hg, whereas the fetal umbilical
vein has an oxygen concentration of 32 to 35 mm Hg (for reviews see (1,2)).
The normal oxygen saturation of a fetus in utero at the inferior vena cava
streaming to the foramen ovale to enter the left atrium is 80% to 85%. This
creates a carotid and coronary artery concentration of 25 to 28 mm Hg (58%
to 65% SaO2), which contrasts sharply with accepted target saturations in
premature infants after birth of 91% to 95% that yield a mean PaO2 close to
80 mm Hg (nl adult arterial saturations = 75 to 100 mm Hg). Furthermore,
oxygen saturations >93% can yield PaO2 higher than 100 mm Hg (3). An
additional factor contributing to the effect of oxygen is fetal hemoglobin,
which shifts the oxygen disassociation curve to the left, resulting in greater
binding of oxygen to fetal hemoglobin than it would to adult hemoglobin
under high oxygen tension. The destructive effects of oxygen create
retinopathy of prematurity (ROP), a vasoproliferative disease that blinds
170,000 infants annually worldwide (see Chapter 52). Recent randomized
clinical trials have tested “static” or continuous low oxygen targets to high
oxygen targets, and determined lower oxygen targets are associated with
reduced ROP but increased mortality (4). Although infant targets are often
outside of these ranges, it may be the high continuous set-point that affects
gene expression, not the range or the low set-point, as discussed below.
Balancing the needs of oxygen for survival with oxygen-induced defects
in multiple organ systems such as the brain, eye, lung, and kidney led to the
thinking that hypoxic preconditioning might be effective in preventing ROP
because it induced stabilization of the oxygen-sensitive transcription factors
and hypoxia-inducible factors (HIFs), which are heterodimeric transcription
factors that regulate the expression of cytoprotective, erythropoietic, and
angiogenic genes (5). Normally, HIFs are degraded in hyperoxia but
stabilized in hypoxia. Experiments that demonstrated the central role vascular
endothelial growth factor (VEGF) played in the development of oxygen-
induced retinopathy (OIR) provided a platform for prevention of ROP. VEGF
supplementation during the hyperoxic phase 1 of ROP prevented vascular
obliteration and retinovascular growth attenuation and thereby removed the
stimulus for ROP by permitting physiologic vascular growth even in
hyperoxia (6,7). VEGF expression was later found to be regulated by HIF (8).
These discoveries led to the hypothesis that stimulating angiogenesis during
hyperoxia by activating HIF (hypoxiamimesis) would reduce avascular retina
that becomes the stimulus for aberrant angiogenesis during hypoxia in OIR
(9).
EVIDENCE THAT OXYGEN IS A

PRIMARY DETERMINANT OF ROP
Although ROP was first reported by Terry in 1942, he initially speculated
that precocious exposure to light stimulated the fetal vasculature to overgrow
the lens (10). Kinsey and Zacharias next postulated a relationship between
disease severity and vitamin E, iron, and oxygen (11). The association of
oxygen toxicity was developed by Campbell and Ryan who saw a direct link
between oxygen and ROP, further confirmed by Crosse who noted a dose
response between oxygen exposure and ROP severity (12–14) (see also
Chapters 40 and 52). This was confirmed by Patz and Kinsey who conducted
a randomized multicenter trial to prove the causal effect of hyperoxia on ROP
seen at that time (15,16).
Follow-up studies on oxygen saturations in humans have noted that high
supplemental oxygen saturation (STOP-ROP) did not decrease ROP but
reduced plus disease in eyes with Stage 3, prethreshold ROP, suggesting that
hyperoxia had an effect on neovascularization and vasodilatation once they
occurred (17). Conversely, in multiple randomized clinical trials, decreasing
oxygen saturation in early life drastically reduced ROP but increased
mortality (4,18). One problem with these studies was that lower oxygen
saturations were maintained until 36 weeks corrected gestational age (CGA)
and hence did not test biphasic oxygen set-points. The proposed benefit of
hypoxic preconditioning is to induce normal retinovascular growth until 32 to
34 weeks CGA when saturations are increased in order to flip phase 1
hyperoxia and phase 2 ischemia (19–21). A nonrandomized retrospective
study compared biphasic oxygen supplementation (85% to 89% saturation
until 34 weeks followed by >95% at 34 weeks and older) to standard care,
defined as static 91% to 95% SaO2, and reported reduced overall ROP and
type 1 ROP with a biphasic approach without an increase in overall mortality
(22). These studies support the hypothesis that hypoxic preconditioning
reduces ROP. It is from this concept that the idea of using hypoxiamimesis,
that is, stabilization of an oxygen-sensitive transcription factor central to
vascular development, was developed. Further studies will provide guidance.
HYPOXIA-INDUCIBLE FACTOR
HIFs are heterodimeric transcription factors comprised of HIFα and HIFβ.
HIFβ is an aryl hydrocarbon nuclear translocator that regulates the expression
of hundreds of genes relevant to cytoprotection, erythropoiesis, angiogenesis,
and metabolism (23). HIFs were originally discovered in 1992 by Semenza as
a factor that bound the 3″ hypoxia-inducible enhancer to the erythropoietin
(EPO) gene (5). The hypoxia response element (HRE) is a 5 nucleotide
sequence (5′-G/ACTCG-3′) found in all DNA binding sites for HIFs (24).
HIFs include two abundant isoforms, HIF-1 and HIF-2, composed of α
subunits that have 48% homology that dimerize with the identical β subunit
(aryl nuclear translocator receptor, ARNT). A third isoform HIF-3 is less well
described and thought to function as a soluble receptor. HIFβ is constitutively
expressed. It is an 87 kD member of the basic helix-loop-helix family of
proteins that contains a per-ARNT-sim (PAS) binding site. Although RNA
levels of HIF are fairly stable, the gene product is regulated
posttranslationally. The stability of HIF can be regulated by multiple
pathways including hydroxylation and ubiquination, phosphorylation,
sumoylation, and inhibition by intermediary metabolites such as succinate
and 3-OH-pyruvate (25,26). Nevertheless, HIF is predominantly regulated
through posttranslational hydroxylation within the oxygen-dependent
degradation domain (ODDD) at Pro 405 and Pro 564 of HIF (Figure 43-1A)
(27). This hydroxylation occurs in normoxia and hyperoxia and serves to
make HIFα a substrate for E3-ubiquitin ligases by permitting ubiquitination
at lysine residues on HIFα at Lys 532, 538, and 547 (Figure 43-1B) (28,29).
Hydroxylation of these proline residues within the ODDD is at the N-
terminal domain compared to hydroxylation of the asparaginyl residues at the
C-terminal domain. When the C-terminal domain is hydroxylated, HIFα is
allowed to bind factor inhibiting HIF (FIH) and interfere with chaperone
p300 from binding the HIF heterodimer, thereby preventing transcriptional
activity (30). In terms of different functions of HIF-1 versus HIF-2, we and
others have found evidence experimentally that Müller cell HIF-2 controls
pathologic angiogenesis, whereas stimulation of HIF-1 controls the normal
physiologic development of retinal blood vessels (31).
FIGURE 43-1 A: Structure of the HIFα gene
demonstrates key proline residues within the oxygen-
dependent degradation domain are trans-4-hydroxylated to
make HIα a substrate of the VHL protein, an E3 ubiquitin
ligase. The N-terminal activation domain (NTAD) has an
asparaginyl residue that is also hydroxylated, but this
makes HIFα a binding partner to HIF, which blocks
binding of HIF to the hypoxia response element. B:
Overall schema of the oxygen sensor within all cells.
Prolyl hydroxylase enzymes, part of the dioxygenase
family, are Fe metalloenzymes sensitive to local oxygen
concentrations. In hypoxia, HIF PHD is inhibited, whereas
hyperoxia induces rapid hydroxylation and subsequent
catabolism of HIFα. C: Replacing α-ketoglutarate with a
competitive inhibitor such as a carbonyl glycine,
benzolamides, or hydrazones. The latter competes for Fe,
whereas the two former small molecules are bound by the
catalytic site of HIF PHD. (C, Cleveland Clinic.)
HIF PROLYLHYDROXYLASES
The oxygen sensor within all cells is HIF prolylhydroxylase, a member of the
dioxygenase enzyme family that requires the cofactors Fe, oxygen, and 2-
oxoglutarate. The fact that these enzymes regulate HIFs by using oxygen and
2-oxoglutarate cofactors makes them central to metabolic plasticity. Oxygen
naturally is the final electron acceptor in respiration, and 2-oxoglutarate is a
tricarboxylic acid (TCA) cycle intermediate that stands at the crossroads of
oxidative phosphorylation and glutamate metabolism. The
prolylhydroxylases are nonheme iron–containing enzymes that possess an
octahedral ferrous iron active site that oxidizes Fe II to Fe IV with carbon–
oxygen bond formation with the α keto group of 2-oxoglutarate. Following
this reaction, the carbon 1-2 bond is cleaved to trans-4-proline that
hydroxylates HIFα in the ODDD with the reaction products being succinate
and carbon dioxide (CO2) (32). The oxygens necessary for hydroxylation of
the ODDD and the formation of succinate are derived from molecular
dioxygen (Figure 43-2A). The affinity of prolyl hydroxylases for oxygen is
relatively low (Km 250 μM) (33–35), which allows prolylhydroxylases to be
sensitive to local oxygen concentrations. It is key that prolylhydroxylase uses
oxygen and α-keg, a central substrate of the TCA cycle and an acceptor of
ammonia to make glutamate, a key player of neuronal transmission,
demonstrating the interplay between oxygen tension and metabolism.
FIGURE 43-2 A:Overall reaction accomplished by HIF
prolyl hydroxylases. Molecular dioxygen is split in the
oxidation of Fe II to Fe IV adding one oxygen to
oxoglutarate and second to proline on the target protein
while decarboxylating α-ketoglutarate to succinate. B: HIF
prolyl hydroxylase inhibition works in the mouse OIR
model and (C) the rat 50/10 model. (B reprinted from
Sears JE, Hoppe G, Ebrahem Q, et al. Prolyl hydroxylase
inhibition during hyperoxia prevents oxygen-induced
retinopathy. Proc Natl Acad Sci U S A
2008;105(50):198980903. Copyright © 2008 National
Academy of Sciences, U.S.A. C republished with
permission of Association for Research in Vision &
Ophthmology, from Trichonas G, Lee TJ, Hoppe G, et al.
Prolyl hydroxylase inhibition during hyperoxia prevents
oxygen-induced retinopathy in the rat 50/10 model. Invest
Opthalmol Vis Sci 2013;54(7):4919–4926; permission
conveyed through Copyright Clearance Center, Inc.)
There are three isoforms of HIF PHD, PHD 1, 2, and 3. These enzymes are
also known as ELGN-2, ELGN-1, and ELGN-3 from their original cloning
from Caenorhabditis elegans. PHD 1 is constitutively found in the nucleus
and hydroxylates both proline residues within the ODDD. The most abundant
and potent isoform is PHD 2, which occurs in the cytoplasm and
hydroxylates both proline residues. PHD 3 hydroxylates only Pro562 and is
found in both the cytoplasm and nucleus. Conditional knockout of all three
enzymes, PHD 1, 2, and 3, is embryonically lethal, whereas inducible
knockout of PHD2 leads to a hypervascular state (36). Each PHD enzyme is
only about 50% homologous in the C-terminal half of the protein. Although
there has been preliminary evidence that suggests a preference of PHD2 for
HIF-1α in vitro, this finding has been difficult to recapitulate in vivo. It is
known that all the PHDs are selective for the C- and not the N-terminal
activation domain. This is important because the biologic effects of PHDs or
of FIH can alter the extent of inhibition of hypoxia-induced transcription of
factors (37).
HIF-REGULATED GENES AND

GENERAL PATHWAY ANALYSIS
HIFs are linked to metabolic plasticity, cytoprotection, erythropoiesis, and
angiogenesis based on the gene products they regulate and the phenotypes
produced when active. Therefore, it has been proposed that the therapeutic
indications for HIFs are anemia, myocardial ischemia, wound healing,
inflammatory bowel disease, and, as will be outlined below, ROP. Of the
hundreds of genes that HIFs regulate, and thousands that are secondarily
affected, the most prominent are EPO, VEGF and VEGFR2, as well as a
cadre of glycolytic genes, including pyruvate dehydrogenase kinase (Table
43-1). The cytoprotection that HIF confers is likely related to protection
against energy-deficient states and protection against hypoxia and ischemia.
However, activation of HIFs during hyperoxia is a strategy that can allow
tissue protection during oxygen toxicity and shift the antiangiogenic state to a
proangiogenic state during hyperoxia.
TABLE 43-1 HIF-dependent genes
From Hong SS, Lee H, Kim KW. HIF-1alpha: a valid therapeutic target for tumor therapy. Cancer Res
Treat 2004;36:343. Copyright © 2004 Korean Cancer Association.
HIF STABILIZATION IN
EXPERIMENTAL ROP BY SMALL
MOLECULES
HIF stabilization is proangiogenic, whereas HIF inhibition is antiangiogenic.
Early in studies of HIF stabilization, we postulated that ROP is a reflection of
systemic oxygen toxicity, which also leads to bronchopulmonary dysplasia
and nephron loss (38). Therefore, we sought to develop a therapy that could
be applied systemically in order to address the global toxicity of oxygen. Our
early studies involved the use of dimethyl oxaloylglycine, a carbonyl amide
that mimics the 2-oxoglutarate cofactor to HIF PHD but inhibits the enzyme
by occupying the binding site of this cofactor to HIF PHD, which in turn
inhibits the trans-4-prolyl hydroxylation of the ODDD and the subsequent
activation or stabilization of HIFα (Figure 43-1C). Administering 200 μg/g
pup weight of DMOG completely abrogated oxygen-induced vaso-
obliteration and attenuation of retinovascular growth in both the mouse and
rat experimental OIR models (9,39). These findings were striking because
stabilization of HIF was effective in two species (Figure 43-2B and C) and
satisfied the different oxygen stresses that are believed to be involved in the
pathophysiology of ROP. The success of this strategy in two different models
also supported the thinking that the overall high set-point of oxygen
saturations is important in ROP pathophysiology, as described by previous
investigators.
SYNERGY BETWEEN THE LIVER AND

THE RETINA
Initial experimentation in OIR models failed to show retinal HIF protein at
anticipated doses and times following an intraperitoneal injection of DMOG
(Figure 43-3A). However, HIF-1 was up-regulated in the liver and confirmed
as the target of DMOG in a luciferase assay using the in vivo reporter
transgene, luciferase ODDD comprised of the ODD fused to luciferase
(Figure 43-3B) (40). We next conditionally ablated hepatic HIF-1 and found
that hepatic HIF-1, with DMOG as a drug, was necessary and sufficient to
transduce protection against oxygen-induced vascular damage in phase 1 in
OIR models (Figure 43-3C) (41). We found further evidence in support of
these findings using RNA-seq by comparing DMOG to roxadustat (42).
Transcriptional analysis was nearly identical in the liver when either drug
was used but vastly different in the retina where DMOG had little
transcriptional effect, but roxadustat increased retinal HIF-1. Finally, using
liquid chromatography-mass spectrometry, we found evidence that DMOG
was broken down in the liver and never entered the blood. Roxadustat, on the
other hand, not only entered the blood but also entered the retina. Therefore,
roxadustat could override hepatic HIF-1α knockout and worked
synergistically to provide near total protection to the retina. In summary,
western blot, luc-ODD reporter gene analysis, RNA-seq, conditional
knockout experiments, and LC–MS provided evidence that the liver protected
a distal capillary bed and supported the hypothesis that low and intermittent
dosage of HIF stabilizers might be safe and effective in fragile premature
infants during hyperoxia.
FIGURE 43-3 A: Western blot shows no dose or time
response of retinal HIF-α after DMOG except in the liver.
(From Sears HE, et al. Prolyl hydroxylase inhibition
during hyperoxia prevents oxygen-induced retinopathy.
Proc Natl Acad Sci U S A 2008;105(50):198980903.)
B:Luciferase–oxygen-dependent degradation domain (luc-
ODD) can act as an in vivo reporter gene to demonstrate
where small molecules that block HIF PHD action are
functional. Note that both subcutaneous and
intraperitoneal injections have the same result.
Hydralazine, a hydrazone, targets the kidneys. Roxadustat
targets both the liver and the retina with some activity in
the kidney. Ref.9 C:HIF-1α ablation in the liver (HIF-
1α2lox/2lox; albumin-cre) abolishes protection by DMOG.
(From Hoppe G, Lee TJ, Yoon S, et al. Inducing a visceral
organ to protect a peripheral capillary bed: stabilizing
hepatic HIF-1α prevents oxygen-induced retinopathy. Am
J Pathol 2014;184(6):1890–1899.). Ref.41
SAFETY AND EFFICACY OF HIF PHD

INHIBITION IN PRECLINICAL
MODELS
There have been multiple trials in humans using PHD inhibitors to correct
anemia of chronic kidney disease in adults. In phase 3 trials, there have been
no safety issues. Nevertheless, we examined liver function tests, renal
function tests, and changes in hematocrit in animals in the mouse and rat OIR
models (see Chapters 40 and 52) and animals in diabetic models that were
chronically administered DMOG weekly. Diabetic animals had no
statistically significant elevation of hematocrit, and the rat and mouse OIR
models had normal serum chemistries whether using DMOG or roxadustat
(41). In addition, electroretinograms of treated animals had normalization of
B-waves even in hyperoxia compared to controls. Light microscopy of the
brain, liver, lung, kidney, and retina revealed no abnormal vascular
proliferation or scar formation. The weights of animals compared to controls
were not different. Finally, when comparing retinovascular development
between treated and control animals, there is no vascular overgrowth but
rather vascular stabilization in hyperoxia in the DMOG-treated animals.
Finally, there are concerns regarding the timing of HIF stabilization. ROP
has been a fairly predictable vasoproliferative disease with onset usually at
about 31 weeks postgestational age (43). And the use of a dilated fundus
exam can accurately depict the different stages of ROP. Therefore, it may be
reasonable to dose HIF stabilization through intravenous administration once
every 5 days, similar to how adult clinical trials are dosed, but begin at birth
and end at 30 weeks CGA. Our studies would predict this regimen would
drive normal, sequential, retinovascular development.
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SECTION VI
Tumors
44
Retinoblastoma
Jesse L. Berry, and Joan Marie O'Brien
INTRODUCTION
Retinoblastoma (RB) is a disease that has advanced our understanding of the
management of pediatric solid tumors and our fundamental understanding of
the etiology of cancer. With steady advances in knowledge of the natural
history, epidemiology, pathogenesis, cellular biology, and therapeutics for
RB, in Western societies more than 90% of children are treated at an early
stage while the tumor is still confined to the globe with excellent prospects
for life (1). Morbidity today has also been dramatically reduced, and a large
majority of children affected by RB can look forward to good vision in at
least one eye and excellent cosmesis (2,3). The study of RB has also led to
concomitant advances in our understanding of the genetic basis for malignant
transformation as the RB1 gene was the first described tumor-suppressor
gene. Although our understanding of the genetics and molecular biology of
RB has undoubtedly had an impact on screening for the disease, surprisingly
to date, this knowledge has had little direct influence on directed therapy or
clinical prognosis, which has advanced empirically.
HISTORICAL CONTEXT
The foundations for modern understanding of RB can be traced back to the
early 19th century, when the Scottish surgeon James Wardrop first
recognized RB as a distinct pathologic entity and published detailed
descriptions of this condition (4). Through his meticulous work, Wardrop
identified the retinal origin of the tumor, characterized the clinical course of
advanced disease, and described optic nerve invasion. He also proposed that
enucleation could be curative. His observations and clinical predications have
proved to be remarkably accurate, and they remain of clinical relevance
today. It was not until the development of the ophthalmoscope by Helmholtz
in 1851 that Wardrop’s ideas regarding treatment could be tested. The
ophthalmoscope allowed diagnosis of intraocular disease, and enucleation
was accomplished before the tumor had spread outside the globe (5). The
work of von Graefe and others improved surgical techniques for enucleation
(6–8). By isolating the rectus muscles, von Graefe achieved proptosis of the
globe and obtained a longer section of optic nerve, thereby reducing the risk
of unresected tumor remaining at the surgical margin of the optic nerve. This
technique improved survival from 5% in 1869 to 57% by the turn of the
century (9,10).
Subsequent decades witnessed advances in understanding the pathology
of RB. In 1864, Virchow (11) proposed the term retinal glioma for this
tumor, reflecting his now rejected hypothesis that the tumor arose from cells
of glial origin. Later, Flexner (12) and Wintersteiner (13) proposed a
neuroepithelial origin for RB and interpreted the rosettes that bear their
names as attempts at photoreceptor differentiation. Verhoeff and others
stressed the embryonic nature of RB and suggested that the tumor was
derived from undifferentiated embryonic retinal cells or retinoblasts. He
proposed the term retinoblastoma to reflect this cell of origin (14). In recent
decades, more differentiated tumors displaying benign characteristics have
been recognized and called retinocytoma or, more recently, retinoma (15,16).
While studies point toward a cone precursor cell of origin (17,18), the
histogenesis of RB continues to be a subject of considerable research interest
and debate.
Hilgartner (19), followed by Schoenberg (20), first advocated the use of
radiotherapy to treat RB. Radiotherapy provided a successful alternative to
enucleation and had particular application in bilaterally affected patients.
While effective for tumor control, the resultant bony midface hypoplasia
(21–23) and, worse, an increase in secondary neoplasms (24) limited its
success. Beginning in the 1990s, systemic chemotherapy used in conjunction
with local consolidative therapy (laser, cryotherapy) improved globe salvage
rates minimizing the need for enucleation and for external beam radiotherapy
(25,26).
In parallel with progress in clinical understanding, significant progress
has been made in understanding the genetics of RB. From RB pedigree
studies in 1971, Knudson concluded that RB could be caused by a germinal
or a somatic mutation and proposed his “two-hit hypothesis.” In this
hypothesis, Knudson proposed that two “hits” or mutations were required to
produce RB. Knudson also collated information regarding clinical
characteristics of patients, including patterns of disease, laterality of disease,
age of onset, and family history, which would pave the way for precise
localization and eventual cloning of the RB tumor susceptibility gene
(27–30).
The identification and cloning of the RB tumor suppressor gene (RB1) in
1987 produced a quantum leap in our understanding of the etiology of cancer
(28–30). Prior to the identification of RB1, it was widely believed that cancer
was caused by activation of oncogenes. Even in the 1980s, studies involving
the fusion of normal cells with cancer cells in culture suggested the existence
of elements within normal cells that could suppress the malignant phenotype.
These elements remained enigmatic until the work of Dryja, Murphree,
Gallie, Friend, and Cavanee that led to the cloning of RB1 in 1987, which
provided the first unequivocal proof of the existence of tumor suppressor
genes for carcinogenesis (28–32). Since the identification of RB1, many other
tumor suppressor genes have been identified. The protein products of these
genes are important in regulating cellular growth and differentiation. When
the function of the retinoblastoma protein (pRB) is reduced through mutation
or deletion in the germ line or as a somatic mutation in the retina, RB may
develop. Abnormalities in RB1 have also been detected in many other
malignancies aside from RB, such as breast, lung, and prostate cancer (33).
These observations suggested that RB1 dysfunction or pathway deregulation
plays a central role in the development of many cancers aside from just RB.
Why germ-line loss of one copy of the RB1 gene so uniquely predisposes to
tumor formation in the retina in young children is not fully understood;
however, cone precursor cells in the developing retina display a particular
tropism for RB1 and a heightened sensitivity to loss of a functional protein
product (17,18).
Basic science understanding of the RB1 gene and its protein product pRB
has so far preceded our ability to apply this knowledge to the human disease
in the form of precision therapies. It is likely that advances in our
understanding of this disease at the molecular level will yield dramatic
improvements in the lives of RB patients and their families.
EPIDEMIOLOGY AND PREVALENCE
RB is the most common primary eye cancer of childhood. The disease affects
approximately 1 in 20,000 live births leading to approximately 9,000 new
cases of RB diagnosed annually worldwide (34–36). An increasing incidence
of RB and other pediatric cancers has been reported in the developed world;
the basis for this increase is not currently understood (37). Patients in
developing regions present with RB at a much later stage in the course of
their disease and have a considerably worsened outcome, suggesting a need
for strategies to improve diagnosis and treatment of this disease in the
developing world (37).
The vast majority of RBs arise from mutations in both copies of the RB1
gene. The first mutation may either involve the germ line or be confined to
the tumor itself. This distinction produces two fundamentally different forms
of the disease: heritable RB and nonheritable RB. Heritable and nonheritable
RBs have quite distinct epidemiologic and clinical features. In brief,
approximately 40% of patients have heritable or germ-line RB; only 10% of
these patients have a positive family history and the others are from a new
sporadic germ-line mutation. These patients are at nearly 45% risk for the
development of RB because the trait is transmitted in an autosomal dominant
fashion with 90% penetrance. Patients with heritable RB have an 80% to 90%
risk of developing multiple tumors and bilateral eye disease in infancy, and
these patients have a lifetime predisposition to develop other secondary
nonocular cancers (38,39). Approximately 5% of germ-line RB patients will
develop a second primary intracranial malignancy in childhood, termed
trilateral RB (36). These tumors represent primitive neuroectodermal
malignancies for which treatment is less effective than for primary intraocular
RB, although there have been recent improvements in treatment modalities
and survival (40–44). Thirty percent of heritable RB patients will develop a
nonocular neoplasm by age 40 years (45).
The remaining 60% of RB patients have the nonheritable or somatic form
of the disease. In these patients, both RB1 mutations occurred within a single
retinal cell, resulting in unilateral, unifocal eye disease. The ocular disease
remains unilateral in these patients, and the cancer syndrome is not
transmissible to future offspring of the patient. There is no predisposition to
develop later nonocular malignancies in these somatic RB patients. A third
group of patients who have mosaic mutations in some portions of their
cellular makeup has also been recognized.
The mean age at diagnosis for RB patients is 13 months for bilateral RB
and 24 months for unilateral cases (46). Patients diagnosed as a result of
family screening are considerably younger, and many cases are detected near
birth. More than two-thirds of RB cases develop in children younger than 2
years, and 95% occur in children younger than 5 years (46). The onset of new
tumors after age 7 years is unusual, although rare cases presenting in
adolescence or adulthood have been reported that may represent malignant
conversion of a benign retinoma (47–53). Patients diagnosed with RB must
be very closely monitored for the development of new retinal tumors, which
are likely to occur in early childhood in those patients with heritable disease.
In those with somatic disease, close monitoring is also required as tumors
may recur and require further treatment.
New germ-line mutations in the human RB gene are known to arise
preferentially on paternally derived chromosomes. Some studies have
suggested an increased parental age in sporadic hereditary RB (54,55).
Approximately 80% of heritable RB cases have a paternally derived mutant
allele, although no differences in paternal age between paternally and
maternally derived mutations have been observed (56). We do not yet
understand what environmental factors, if any, influence the development of
initial mutations in the RB1 gene. Some have postulated that human
papillomavirus (HPV) may play a role in sporadic RB. Environmental
factors, however, likely have a role in the predisposition to second cancers
observed in adults with the heritable form of RB, because the distribution of
second cancers in these patients is influenced by previous exposure to
radiotherapy.
WORLDWIDE IMPACT
The exact number of cases of RB worldwide is unknown. It has been
estimated at approximately 9,000 cases annually (34). Countries with the
highest number of annual births, China and India, are thought to harbor the
largest number of new cases. Limited access to cancer registries in
developing countries makes accurate assessments difficult.
PATHOPHYSIOLOGY
Thorough understanding of pathologic features, including growth patterns
and mode of tumor extension, is critical for diagnosis, staging, and
management decisions in RB patients. RB represents a malignant tumor of
neuroectodermal origin arising from the nucleated layers of the retina.
Malignant cells usually are poorly differentiated, rounded cells of variable
size and shape with a high nuclear-to-cytoplasmic ratio and numerous
mitoses. RB may display variable cellular differentiation, although most
cases encountered are relatively undifferentiated (57,58). RB is composed of
a solid well-vascularized tumor mass with regions of viable tumor cells
surrounding vessels and interspersed zones of necrosis and calcification, a
hallmark of RB. Additionally, apoptotic cells, cells of the mononuclear
phagocyte series, lymphocytes, and reactive glial cells have been observed in
these tumors (59,60).
Photoreceptor differentiation is suggested by tumor cell rosette and
fleurette formation; the morphologic features of these structures have been
meticulously demonstrated by a number of histopathologic and ultrastructural
studies (Figure 44-1) (12,13,61,62). The Flexner-Wintersteiner rosette is not
specific for RB, but it does represent a characteristic pathologic feature.
These rosettes have also been observed in pineoblastoma and
medulloepithelioma (63). The Homer-Wright rosette is less frequently
observed in RB and is less specific; this rosette is found in a variety of other
neuroblastic tumors (64). Glial differentiation may be observed in RB (65).
Although true glial differentiation is difficult to distinguish from reactive
gliosis, studies in tissue culture have demonstrated the potential of RB cells
to differentiate along a neuronal or glial cell lineage (59,60,65).
FIGURE 44-1 A: Hematoxylin and eosin–stained section
from an RB specimen. A region of viable tumor cells
demonstrating rosette formation. A prominent Flexner-
Wintersteiner rosette is indicated (arrow). A tumor
vascular complex is present at the top right-hand corner
(×25). B: Toluidine blue–stained section from an RB
specimen showing fleurette formation, an expression of
photoreceptor differentiation (×100).
Cell death is usually a prominent characteristic in RB specimens, and

extensive areas of cell death are interspersed with zones of viable tumor cells
surrounding blood vessels. A number of investigations suggest that tumor
cells displaced from feeder vessels by more than 100 μm undergo ischemic
necrosis. This observation has been attributed to the metabolic demands of
this highly active tumor outstripping the supply of nutrients (66). Although
necrotic portions of RB tumors do not characteristically provoke an
inflammatory response, massive necrosis in large tumors may occasionally
become clinically relevant by producing marked inflammation that involves
surrounding tissues and clinically simulates orbital cellulitis (67).
Apoptosis plays a critical role in growth regulation of many tumor types.
This form of cell death has been reported in RB primary tumors and in cell
lines and has been recognized as the primary mechanism of RB cell death in
response to radiation, cytotoxic chemotherapy, and other therapeutic
approaches (68,69). Cell death in RB is frequently accompanied by
dystrophic calcification, a hallmark seen on imaging and clinical examination
as well as pathologic analysis. In one clinical series, more than 80% of RB
tumors showed calcification, and the degree of calcification appeared to be
related to tumor size, with small tumors tending to be less calcified.
Extraocular RB tumor does not typically demonstrate calcification (70–72).
Tumor vasculature is another critical histopathologic feature of RB. The
development of an intrinsic tumor blood supply, termed tumor angiogenesis,
is critical to the growth of primary tumors and also influences metastatic
extension by providing access to the systemic vasculature.
One significant mechanism promoting angiogenesis in tumors is the
production of vascular endothelial growth factor (VEGF). VEGF mRNA and
protein have been demonstrated in RB cells, and the expression of VEGF is
increased by hypoxia (73–76). Vascular endothelia at intraocular sites distant
from the tumor may respond to high intraocular levels of VEGF by
increasing expression of VEGF receptors. An increase in VEGF receptors
could have relevance for neoplasm-related ocular angiogenesis, such as iris
neovascularization, which is observed in RB patients with large intraocular
tumors (75). Leukocytes, particularly cells of the mononuclear phagocyte
series, have been observed in RB primary specimens in association with
tumor vasculature. The role of these cells in immune surveillance, tumor
development, and angiogenesis remains to be determined (59,60).
Left untreated, RB grows relentlessly, filling the ocular cavity and
eventually spreading extraocularly. Four primary patterns of intraocular RB
growth have been described (77,78). The endophytic growth pattern is
characterized by growth of tumor toward the vitreous space; in this case, the
tumor mass may be directly visualized with the ophthalmoscope (Figure 44-
2). In the exophytic growth pattern, tumor grows from the outer retina toward
the choroid, elevating the retina and presenting as serous retinal detachment
(Figure 44-2). In practice, most RB patients display a combination of these
two growth patterns; this is termed a mixed growth pattern. A fourth pattern,
known as diffuse RB, is clinically significant. Although this form is rare,
accounting for only 1% to 2% of all RB cases, the tumor insidiously
infiltrates the retina, producing plaque-like thickening. The clinical signs in
this form are subtle, with consequent delays in diagnosis (79).
FIGURE 44-2 A: Gross pathology specimen
demonstrating the exophytic pattern of RB tumor growth.
The chalky white tumor mass at the posterior pole had
prominent calcification, arose from the retina, and grew in
a scleral direction. Such tumors have a tendency to spread
in the subretinal space. B: Clinical ophthalmoscopic
appearance of an exophytic RB. The tumor demonstrates a
pale yellow color and is covered by the overlying retina,
which often has a serous detachment. The retinal vessels
overlying the tumor tend to be dilated and tortuous.
C:Multiple endophytic tumors in a patient with heritable
RB. These tumors appear as pale chalky masses due to
prominent calcification.
RB cells are poorly cohesive; they readily disseminate throughout the globe,
a clinical phenomenon called seeding. One mechanism underlying loss of
cellular cohesion in RB may include deranged expression of intercellular
adhesion molecules such as N-cadherin (80,81). RB cells are shed into the
vitreous as small clumps of tumor cells called vitreous seeds (from
endophytic growth) or into the subretinal space (from exophytic lesions),
called subretinal seeds. (See clinical presentation below.) RB tumor cells
frequently disseminate away from the main tumor mass, involving ocular
structures such as distant retina, lens capsule, or anterior chamber. This
dissemination can result in secondary ocular complications such as glaucoma
or pseudouveitis (57). Retinal seeding can occasionally mimic multicentric
RB, leading to the inaccurate diagnosis of germinal mutation. In large tumors
with extensive vitreous and subretinal seeding, the distinction between
unifocal and multifocal disease may be impossible to determine clinically.
RISK OF METASTASIS
RB may spread extraocularly and extend outside the eye by a number of
routes. The most frequently observed route of extension is through invasion
of the central nervous system (CNS) through the optic nerve, followed by
direct extraocular spread and involvement of the orbit. Tumor cells may
invade the optic nerve directly or may invade the leptomeninges, gaining
access to the orbital apex, optic chiasm, base of the brain, and subarachnoid
space (57). Extension of RB cells into the optic nerve is considered a major
histopathologic risk factor for the development of metastases, particularly
when the tumor extends beyond the lamina cribrosa. If the optic nerve is
involved posterior to the line of surgical transection at the time of
enucleation, patients demonstrate a dramatically worsened clinical course
(82–85). RB tumor cells may also invade the retinal pigment epithelium,
Bruch membrane, and choroid (86,87). Exposure to the rich choroidal
vasculature provides tumor cells access to emissary vascular channels and
ultimately to the systemic circulation. The sclera may also be directly invaded
by tumor cells. It has been suggested that the exophytic growth pattern is
associated with increased risk for choroidal or orbital involvement and
metastases, although this hypothesis remains clinically unproven (86). It has
also been suggested that massive invasion of the choroid is an independent
risk factor for disease dissemination (88–95). Tumor cells that spread
hematogenously produce widespread metastases to the lungs, brain, bone,
and other viscera. Lymphatic dissemination of RB may occasionally occur in
cases that extend anteriorly and allow tumor cells access to the bulbar
conjunctiva and to the eyelid lymphatics (57).
The complex molecular interactions among tumor cells, parenchymal
tissues, and extracellular matrices are currently not fully understood. Recent
research suggests that a growth and invasion advantage may be conferred on
subgroups of tumor cells that become independent of normal mechanisms for
cell death or apoptosis and avoid tissue constraints, allowing spread to distant
sites. These clones of more aggressive cells arise through ongoing mutagenic
and epigenetic phenomena within the tumor. Some molecular mechanisms
identified in RB and other tumors that could facilitate these changes include
up-regulation of telomerase activity, N-myc oncogene amplification, p53
inactivation, changes in cell adhesion molecules (e.g., cadherins), altered
expression of integrin subunits by tumor cells, and increased expression of
angiogenic factors (81,96–98). Greater understanding of these critical
molecular processes is likely to yield better and more specific therapies for
this disease in the future.
OVERVIEW OF RETINOBLASTOMA
GENETICS
Retinoblastoma Gene and Protein Product
RB is one of very few cancers in which a single genetic mutation predicts
disease development at very high penetrance. The presence of an underlying
genetic mutation in RB was predicted by Knudson in 1971, and this
prediction led to identification of the RB susceptibility gene locus, with
subsequent cloning of this tumor suppressor gene in 1987. Identification of
the RB1 gene provided definitive evidence that loss of tumor suppressor gene
function is important in the etiology of all human cancers. Although it
appears that other genes may be involved in the subsequent tumorigenesis of
RB (99), in order for RB to arise, a mutation on both alleles of the RB1 gene
is required in the great majority of cases (16). Rarely, RB is initiated by
primary MYCN amplification in the absence of a RB1 mutation. This occurs
in approximately 2% of unilateral cases (100).
The RB1 gene encompasses 180,000 base pairs on chromosome 13q14
and contains 27 exons. The gene encodes a 4.7-kDa message that results in a
105-kDa nuclear phosphoprotein. The RB protein, known as pRB, is a
transcriptional modulator expressed throughout development in all cells of
adult humans (101–104). The protein has two conserved regions where
important functional domains reside and which also represent sites for viral
oncogene binding (e.g., SV40 T-antigen, adenovirus E1A, and HPV E7).
Oncogenic viruses are capable of inactivating pRB through binding, thereby
inducing malignant transformation. Many of the clinically relevant mutations
that have been observed in RB1 affect these conserved regions of the protein
(38,105–107). Mutations have also been observed in the promoter region of
the gene; these mutations affect recognition sequences for transcription
factors (e.g., ATF and SP-1) (108).
Mutations affecting the RB1 locus appear to be very heterogeneous. Few
preferential sites for mutation, or “hotspots,” have been identified (38,109).
Only 3% to 5% of constitutional mutations in RB1 are cytogenetically
recognizable; these include interstitial deletions or translocations involving
13q. The remainder of mutations are submicroscopic, including point
mutations, microdeletions, and duplications. In more than 90% of RB1
mutations, deoxyribonucleic acid (DNA) alterations result in a truncated
mRNA product (38,101). The second mutation that occurs in the retinal cell
of origin of the tumor is a result of chromosomal abnormalities, such as
nondisjunction, loss of heterozygosity due to mitotic recombination, or large
deletion (101). This second event may be more influenced by environmental
factors, including ionizing radiation. Therapeutic radiation exposure
influences the distribution of nonocular tumors.
The RB protein has a critical and complex role in the regulation of
cellular growth and differentiation. pRB binds to more than 30 separate
cellular proteins, including transcription factors, nuclear matrix proteins,
growth regulators, protein phosphatases, and protein kinases (110). One of
the best understood functions of pRB involves regulation of the cell cycle
through inhibition of the transcription factor E2F (111–113). In this role, the
active hypophosphorylated form of pRB acts at a restriction point in late G1
of the cell cycle to inhibit the E2F family of transcription factors. These
factors in turn regulate expression of a large set of genes, including, among
others, c-myc, b-myb, cdc2, E2F-1, and the dihydrofolate reductase gene
associated with cell division in S phase (112,113). The phosphorylation and
dephosphorylation of pRB itself are controlled by a complex series of cell
cycle–dependent and cell cycle–independent enzymes, including cyclin–
cyclin-dependent kinase (cyclin–CDK) complexes and cyclin-dependent
kinase inhibitors (CDKIs) (114,115) (Figure 44-3).
FIGURE 44-3 Schematic representation of pRB function

in the regulation of the cell cycle. Active
hypophosphorylated pRB blocks cell cycle progression by
repressing E2F target genes through binding promoter-
bound E2F and by recruiting chromatin remodeling factors
(CRFs) including histone deacetylase. Phosphorylation of
pRB is controlled by cyclin–cyclin-dependent kinase
(cyclin–CDK) complexes resulting in inactivation of pRB,
allowing transcriptional activation of E2F target genes.
Cyclin-dependent kinase inhibitors (CDKIs) positively
regulate pRB by inhibiting the negative regulators, the
cyclin–CDK complexes.
In addition to its role in cell cycle regulation, pRB has important effects on
cellular differentiation. These effects include cell cycle arrest, suppression of
apoptosis, and transcriptional activation of tissue-specific genes (103). These
effects may be particularly important in tissues such as muscle, bone, fat, and
nerve (116–120). Despite the ubiquitous expression of pRB in all dividing
cells, germ-line mutations leading to loss of functional RB protein in humans
seem to uniquely cause RB and, later, tumors of mesenchymal origin.
These mesenchymal tumors include osteosarcoma, leiomyosarcoma, and
malignant fibrous histiocytoma (121). In contrast, somatic alterations in pRB,
which occur mainly through functional inactivation of upstream regulators,
are involved in the progression of many common cancers. These include
carcinoma of the breast, bladder, and prostate; small cell lung cancer;
leukemia; and glioblastoma (33). For example, mutation of p16Ink4a (a CDKI)
and amplification of cyclin D, both upstream regulators of pRB, are
frequently observed in these malignancies (122,123). The vulnerability of
RB1-deficient cells in the retina and certain connective tissues to tumor
development in the presence of RB1 germ-line mutation has never been
adequately explained. It is possible that this vulnerability arises from a
critical requirement for pRB in terminal differentiation of these tissues.
Although mutation in both RB1 alleles initiates RB, other secondary
chromosomal abnormalities and mutations have also been consistently
observed. These other consistent genetic and genomic abnormalities include
trisomy 1p, loss of heterozygosity for chromosome 17, and isochromosome
formation at 6p (124–127). These abnormalities could represent secondary
changes due to genetic instability and could play a role in mediating
malignant progression, allowing clones of malignant cells to acquire selective
growth advantage. Interestingly, reconstitution of RB cells, RB-null
carcinoma cells, or sarcoma cells with functional pRB only partially
modulates the malignant phenotype (128). This suggests that other consistent
genomic alterations such as the highly recurrent chromosomal aberrations
include gain of 1q, 2p, and 6p and loss of 13q and 16q. A better
understanding of the target genes contained in these regions may inform
understanding of RB tumorigenesis and phenotype (99,129).
Clinical Genetics
Heritable Retinoblastoma
Although only 10% of RB patients have a family history of this disease, new
germ-line mutations account for an additional 30%, meaning that 40% of RB
patients have a heritable form of RB. Patients with heritable disease have a
mutation (or first “hit,” according to Knudson’s hypothesis) involving RB1 in
every cell of their bodies. This is termed a germ-line mutation because the
mutation either was inherited from a parent or, more commonly, developed at
the one-cell stage of embryonic development in the affected individual (or as
a mosaic in which the mutation developed during embryogenesis but after the
one-cell stage; the % of mosaicism in a patient is determined by the timing of
this mutation). Individuals with germ-line mutations in RB1 are capable of
passing on the mutation to their offspring. Mutation in the other copy of RB1
(the second “hit” described by Knudson) occurs in an undifferentiated
embryonic retinal cell that gives rise to the malignant tumor. In this heritable
form of RB, patients usually demonstrate bilateral, multifocal retinal tumors,
which are pathognomonic for heritable disease. Although the RB gene at the
cellular level is recessive (i.e., both RB alleles must be abnormal in order for
the tumor phenotype to be expressed) at the level of the individual, the
mutation behaves in an autosomal dominant manner, with 80% to 90%
penetrance (130) (Table 44-1).
TABLE 44-1 Clinical and genetic forms of RB

aSome of these cases may represent parental mosaicism.
bFifteen percent of total germ-line mutations present as unilateral diseases.
RB, retinoblastoma.
Nonheritable Retinoblastoma
The remaining 60% of RB cases represent nonheritable or somatic disease. In
this nonheritable form, mutations in both RB1 alleles (i.e., both the first and
the second “hits”) occur within a single retinal cell of origin for the tumor.
Such events are rare; therefore, these patients demonstrate a single unilateral,
unifocal tumor. Unilateral disease presents, on average, at a later age than
does bilateral disease given the additional time needed to acquire the first and
then second mutation in these cases. This observation was the basis for much
of Knudson’s hypothesis. These children with unilateral involvement also
present at a later age because they retain vision in the unaffected eye.
It should be recognized that approximately 15% of patients who present
with unilateral RB have the heritable form of this disease, but by chance they
have developed a tumor in only one eye (131). For this reason, all children
with RB should undergo genetic testing for evaluation of somatic mutations,
and this can be critically impactful in unilateral cases as it dictates risk of
subsequent ocular and nonocular tumors and risk to future offspring. The
relationship between the clinical and the genetic forms of RB is summarized
in Table 44-1.
Second Tumor Predisposition

As a consequence of germ-line mutation in RB1, patients with heritable RB
should be regarded as having a genetic cancer predisposition syndrome. The
clinical consequences of this syndrome are that these patients develop not
only multiple, bilateral retinal tumors, but they also demonstrate a severalfold
increased risk for developing second malignancies in later life, including
osteogenic sarcoma, fibrosarcoma, and melanoma (38,39). These second
nonocular tumors may occur anywhere throughout the body, although their
distribution appears to be increased within the radiation field following
radiotherapy, particularly if radiation was delivered before 12 months of age
(24,132). By age 40 years, 30% of heritable RB patients will have developed
a second nonocular cancer (45). Additionally, approximately 5% of
hereditary RB patients will develop midline intracranial primitive
neuroectodermal tumors (PNETs, formerly called trilateral RB), which
frequently occur in the pineal region (44). These tumors usually occur in
conjunction with bilateral disease, although PNETs have also been described
in patients with unilateral retinal tumors and in patients with RB1 mutations
(133).
Genetic Mosaicism
It has been recognized that at least 10% of RB families demonstrate germ-
line or somatic mosaicism (134). Mosaicism refers to the presence of two or
more cell lineages differing in genotype in an individual. If a mutation
develops at the RB1 locus during later stages of embryonic development, the
lineage of cells derived from this original mutation-bearing cell will carry an
identical RB1 mutation. In contrast, cells derived from unaffected lineages
will carry normal copies of RB1. If the mutant RB1 is carried in the gametes
but not in the eye, the affected individual could pass the mutation to offspring
without personally demonstrating clinical evidence of RB. The practical
significance of mosaicism in one of a proband’s parents is that siblings of this
proband would be at higher risk for developing the disease than previously
believed (134). Therefore, siblings should receive early and regular screening
for development of this disease or genetic testing to better define the risk
(135). Parents who are mosaic for mutant RB1 may also be at personal risk to
develop malignancies in cell lineages that carry the mutant gene. These
individuals should receive prompt referral to an oncologist if systemic
complaints suggestive of malignancy develop. It should also be noted that
while the initial familial mutation may present in a mosaic fashion, future
generations will only develop RB if the mosaic RB mutation involves the
gametes of the affected individual.
Low-Penetrance Retinoblastoma
Forms of RB have been described in families where a proportion of RB1
mutation carriers do not develop eye tumors. This clinical circumstance is
termed low-penetrance RB. In other families, RB1 mutation carriers either
develop unilateral disease or demonstrate a benign variant of RB called
retinoma. This clinical circumstance is described as low-expressivity RB.
Low-penetrance and low-expressivity forms of RB have been associated with
RB1 mutations that either partially inactivate the protein or reduce protein
expression (136). Recognition of these clinical situations is important
because they allow more accurate genetic counseling for affected families
(130,137).
Current Status of Genetic Counseling and DNA

Testing
It is important that all ophthalmologists who treat patients with RB inform
patients and families of the potential heritable nature of this disease. Families
also should be made aware that germ-line mutation in RB1 confers a lifetime
second tumor predisposition. Referral to a pediatric oncologist for lifelong
second tumor surveillance is indicated.
Genetic counseling is often performed by the ophthalmologist in
conjunction with a clinical genetics service. A careful and complete family
history should be taken at the time of the initial referral. Examination of
relatives for retinoma or regressed RB can be very helpful in some cases, as
this suggests carrier status for the affected relative. Regardless of age at
presentation or laterality of disease, it is recommended that all patients have
serum testing for a RB1 mutation; if tumor is available, that may also be
tested (138–141).

When there is a family history of RB (10% of cases), generally these patients
are detected very early as part of RB screening programs. In these siblings or
offspring of RB patients, the diagnosis usually is made at a very early stage in
infancy when the tumors are small, offering the best opportunity for
treatment aimed at preserving the eye and whenever possible the vision.
In unaffected families, RB presents either incidentally or following
referral by a family physician or pediatrician. Often, the first sign of the
disease is a white pupillary light reflex, called leukocoria, or strabismus,
usually noticed initially by a patient’s parent or relative. Leukocoria was first
described as a presentation for RB by Hayes in 1767 and unfortunately
remains the most common presenting sign of this disease today (in ~60% of
cases) (Figure 44-4) (142,143). Strabismus is the second most frequent
presentation of RB (in ~20% of cases), and this observation reinforces the
need to perform a dilated retinal examination in all new patients who present
with strabismus. Unfortunately, both of these presenting signs are generally
found when the tumor is quite advanced and has affected vision. Other less
frequent presentations for RB include intraocular inflammation, iris
neovascularization, hyphema, pseudohypopyon, preseptal/orbital cellulitis,
and glaucoma (Figure 44-4) (144). With increased awareness and the advent
of early screening programs, some patients may be found at a very early stage
with no signs apart from the presence of a small retinal lesion. However,
given the rarity of this condition, and the low proportion of familial cases,
this has been minimally successful even in developed countries.
FIGURE 44-4 A: Heritable RB patient presenting with
bilateral leukocoria, the most commonly reported
presenting sign of this condition. B: RB cells have a
propensity to detach from the primary tumor mass and
spread throughout the eye, as illustrated by a patient with
endophytic RB who developed marked tumor cells in the
vitreous, resembling a vitritis. C: Occasionally, tumor
necrosis may produce an inflammatory response
associated with a red eye and conjunctival injection as
seen in this patient. The patient also has a tumor
hypopyon. D: Massive tumor necrosis in a phthisical eye
of a patient with RB has resulted in marked inflammation
involving the orbit, simulating an orbital cellulitis with lid
swelling and pain on eye movement.
The clinical presentations of RB that simulate inflammation are associated

with the greatest diagnostic difficulty. True inflammation may arise in an eye
with RB due to extensive tumor cell necrosis. This situation occurs with very
large tumors, and the accompanying inflammation may simulate preseptal or
orbital cellulitis. If RB is suspected, identifying the apparent cellulitis as a
masquerade syndrome is often straightforward. Children with true orbital
cellulitis exhibit fever, leukocytosis, and sinusitis. These findings are
generally absent in patients with RB. The presence of normal sinuses and
evidence for an intraocular mass are easily documented with magnetic
resonance imaging (MRI) (Figure 44-4). In the setting of massive tumor
necrosis, it has also been reported that the lens may dislocate posteriorly and
there may be loss of view from vitreous hemorrhage, hyphema, or early
phthisis (145). Children with RB may present with hyphema, for example,
and prior to any decision to “wash out” the hyphema, ultrasound is indicated
to exclude RB. In any case with loss of view, the clinician should have a high
level of suspicion for RB particularly if calcification is seen on intraocular
imaging (B-scan, CT, or MRI; of note, CT should not be the imaging
modality of choice if RB is suspected) (146).
Another less straightforward clinical presentation for RB involves friable
tumor cells that detach from the main tumor mass and subsequently spread
throughout the eye. These tumor cells may simulate the white blood cells of a
hypopyon and may suggest uveitis as an initial diagnosis (Figure 44-4).
Secondary glaucoma, due to mass effect from the tumor producing angle
closure or due to secondary iris neovascularization, occasionally may
produce a diagnostic challenge.
Rarely in the developed world, patients may present with evidence of
extraocular spread, metastatic disease, or CNS symptoms due to a PNET.
Proptosis, although rare as a presenting sign of RB in industrialized societies,
unfortunately is a common presentation in less-developed countries.
Proptosis was the most common presentation of RB (65% of cases) in one
study from India (147). This presentation carries a poor prognosis because it
is frequently associated with advanced disease and extraocular spread into the
orbit.
RB occasionally may occur in association with other congenital
abnormalities, including cleft palate, cardiovascular defects, infantile cortical
hyperostosis, dentinogenesis imperfecta, incontinentia pigmenti, or familial
congenital cataracts. Patients also may demonstrate features of 13q deletion
syndrome, associated with deletions in the distal part of chromosome 13q.
These patients demonstrate mental retardation, broad nasal bridge,
hypertelorism, microphthalmia, micrognathia, cleft palate, or foot and toe
abnormalities (148,149).
Clinically, on indirect funduscopy, an endophytic tumor appears as a
white- or cream-colored mass that penetrates the internal limiting membrane
of the retina and demonstrates prominent white chalky areas that represent
intrinsic calcification.
The vessels associated with endophytic RB usually are small, irregular
intrinsic tumor vessels that stand in marked contrast to the dilated,
aneurysmal blood vessels observed in Coats disease (150). Advanced
endophytic RB may be associated with seeding of tumor cells into the
vitreous space. These clumps of cell can present as dust-like debris,
snowball-like spheres (151), or massive clouds (152,153) within the vitreous
cavity (Figure 44-5). Sometimes there are intervening clear spaces, and
sometimes the seeding is so extensive that it obscures the view of the
underlying ocular structures (154). Rarely, vitreous seeding from RB may be
so extensive that the anterior chamber is involved. This presentation with a
tumor hypopyon can resemble endogenous endophthalmitis (Figure 44-4),
and care should be taken to perform a detailed B-scan ultrasound in a young
child in this setting, particularly in the absence of positive blood cultures or
other infectious findings (155).
FIGURE 44-5 The clinical applications of handheld

optical coherence tomography (OCT) imaging (Bioptigen,
Morrisville, NC) for retinoblastoma continue to evolve and
include characterization of small tumors, tumor
recurrences, evaluation of seeding, and retinal anatomy.
OCT was performed at staging examination of the left eye
in a 13-month-old child diagnosed with bilateral
retinoblastoma (Group C right eye, Group D left eye) who
demonstrated normal foveal architecture, preretinal
dusting of small hyperreflective seeds, and a hollow
reflective cystic structure floating above the retina, with
shadowing posteriorly, which correlated clinically with a
large translucent spherical seed in the vitreous cavity
(asterisk). (Reprinted from Berry JL, Anulao K, Kim JW.
Optical coherence tomography imaging of a large
spherical seed in retinoblastoma. Ophthalmology
2017;124(8):1208. Copyright © 2017 by the American
Academy of Ophthalmology. With permission.)
In its exophytic form, RB tumors usually are yellow to white in coloration
and are observed deep to the overlying retina, which is often serously
detached from fluid and the tumor beneath the retina. The retinal vessels
overlying the tumor may be dilated and tortuous. In contrast to serous
detachments from other causes, an underlying mass with characteristic
features usually can be demonstrated by ultrasound or MRI. Occasionally,
intraretinal and subretinal exudation may mimic Coats disease, making
diagnosis difficult (Figure 44-2) (156,157).
Seeding in either the vitreous or subretinal space has the capacity to
implant and to grow at other retinal sites, producing multiple tumor foci.
Most advanced tumors have both exophytic and endophytic features. Very
small RBs may appear as pale grayish masses confined to the retina. Tumors
so small as to be clinically invisible have now been noted from optical
coherence tomography (OCT) evaluation of young children (Figure 44-6)
(158,159).
FIGURE 44-6 A: Color fundus photograph of the left eye
demonstrates three retinoblastoma tumors. B: Color
fundus photograph of the left eye demonstrates three
retinoblastoma tumors labeled Rb1–Rb3, marked by lines
through the body of the tumors correlating with the OCT
slice. The smallest one, barely visible on funduscopy, lies
just superior to the optic nerve. C: Spectral-domain OCT
of the three tumors shows homogenous dome-shaped
masses with overlying inner retinal draping. Tumor No. 3
is located in the outer retina involving the ONL and
possibly the OPL. The INL and IPL drape over the tumor.
There is also an outer retinal abnormality in all tumors
affecting the external limiting membrane (ELM), ellipsoid
zone (EZ), and interdigitation zone (IZ). There is
shadowing on OCT from the retinal vessels overlying the
tumor, which are also seen clinically. (Reproduced with
permission from Berry JL, Cobrinik D, Kim JW. Detection
and intraretinal localization of an ‘invisible’
retinoblastoma using optical coherence tomography. Ocul
Oncol Pathol 2016;2(3):148–52.)
The diffuse infiltrating form of RB is the most difficult form to accurately

diagnose. Diffuse RB often presents in older children as a resistant uveitis
with mild conjunctival hyperemia and pseudohypopyon. Ophthalmoscopy
may reveal a diffuse gray-white opacification and retinal thickening with
vitreous seeds. Frequently, this form of RB is situated in a more anterior
location, underscoring the importance of dilated fundus examination with
scleral indentation. In the absence of a well-defined mass with a lower
incidence of intrinsic calcification, ultrasound and MRI may be less
informative in establishing the diagnosis of diffuse RB (160–163).
The clinical diagnosis of RB always requires that this malignant tumor be
distinguished from other simulating disease processes by an ocular
oncologist. In a study of 500 patients referred to the Wills Eye Hospital for
suspected RB, only 58% of referred patients were found to have RB on
clinical evaluation. The remaining 212 patients received diagnoses of 23
different forms of pseudoretinoblastoma (157). The most common simulating
lesions were persistent hyperplastic primary vitreous (PHPV) (28%), Coats
disease (16%), and presumed ocular toxocariasis (16%), followed by
retinopathy of prematurity and retinal hamartoma. Other simulating lesions
included hereditary conditions, such as familial exudative vitreoretinopathy,
congenital retinoschisis, Norrie disease, and incontinentia pigmenti. Similar
findings and referral accuracy were found in a study from Sloan Kettering
Cancer Center (164). Developmental conditions that are mistaken for RB
included retinal or optic nerve coloboma, retinal dysplasia, congenital retinal
folds, and myelinated nerve fibers. Inflammatory or infectious conditions,
including congenital toxoplasmosis or toxocariasis or cytomegalovirus or
herpesvirus retinitis, can also simulate RB. Tumors such as
medulloepithelioma, choroidal hemangioma, glioneuroma, capillary
hemangioma, and ocular infiltrates from leukemia occasionally can present
initially as RB. It should always be recalled in the setting of media opacities,
such as hyphema, cataract, or vitreous hemorrhage, that the diagnosis of RB
is not excluded until the retina has been thoroughly examined or imaged to
exclude a concomitant tumor. RB coexisting with other conditions, such as
PHPV or cataract, has been reported (165). In practice, most clinical cases of
RB can be diagnosed through careful clinical evaluation by an experienced
examiner and the judicious application of supplemental investigations. Table
44-2 summarizes the salient diagnostic features of the more common lesions
that simulate RB.
TABLE 44-2 Diagnostic features of most common

lesions simulating RB
aRB can produce a Coats-like reaction with a large amount of retinal exudation. In these eyes, US can
usually detect the retinal tumor beneath the exudate.
bCorrespond to the refractile cholesterol particles seen clinically. These particles are much less
reflective than the calcium particles seen with RB. (From Shields JA, Shields CL, Parsons HM.
Differential diagnosis of retinoblastoma. Retina 1991;11:232–243.)
cCalcification is not specific to RB, although it is very characteristic. The calcification in astrocytomas
appears clinically as refractile yellow areas versus the chalky white appearance in RB.
RB, retinoblastoma; PHPV (PFV), persistent hyperplastic primary vitreous (persistent fetal
vasculature); CT, computed tomography; Dx, diagnosis; FA, fluorescein angiography; HA, hyaloid
artery; HI, high intensity; Hx, history; ILM, internal limiting membrane; LBW, low birth weight; MI,
medium intensity; MRI, magnetic resonance imaging; RD, retinal detachment; RPE, retinal pigment
epithelium; US, ultrasound; AD, autosomal dominant.
STAGING EXAMINATION
Whenever RB is suspected, a detailed medical history should be taken at
initial examination, including a prenatal and birth history. Historical features
suggesting a pseudoretinoblastoma should be specifically investigated (Table
44-2). A family history should be obtained, and a detailed family tree should
be diagrammed in the medical record. A history of RB or other forms of
cancer, as well as a history of ocular malformations, blindness, or enucleation
in other family members, should be explored. Enucleation or blindness
occurring within a family is often better remembered than the precise
diagnosis of RB. The family tree should document the ages and health status
of all siblings and near relatives.
In every child with suspected RB, a complete physical examination is
required. Specific features associated with simulating lesions and/or RB
syndromes or systemic metastases should be evaluated. For example, the
child should be examined for evidence of tuberous sclerosis (e.g., ash leaf
spots) or dysmorphic features suggesting a 13q deletion syndrome. Evidence
for underlying systemic disease or metastases, such as cachexia, wasting, or
developmental delay, should be investigated. Clinical features of PNETs
associated with trilateral RB include abnormal eye movements, vomiting,
seizure, headache, and changes in cognition. We routinely refer all patients
with suspected RB to a knowledgeable pediatric oncologist for a complete
history and physical examination.
The ophthalmologic evaluation of a child with suspected RB should be
detailed, complete, and performed under general anesthesia. While there is a
priority placed on visual acuity and visual function, the primary goal is
always cancer control. The visual potential of each eye is an important
determinant for future therapy, and patients with unilateral disease and eyes
that have been functionally destroyed by the tumor should be considered for
enucleation as a curative procedure. External examination of the patient
should evaluate for dysmorphic features, lymphadenopathy, or proptosis.
Pupils and eye movements should be evaluated for evidence of a concomitant
intracranial disease process. Careful anterior segment examination is
necessary to evaluate the size of the corneas and the clinical appearance of
anterior segment structures. For example, a unilateral small corneal diameter
may suggest PHPV particularly in the setting of anteriorly rotated ciliary
body processes, rather than RB. An enlarged diameter may be seen in the
setting of buphthalmos from an eye with advanced disease and high
intraocular pressure. Signs of intraocular inflammation should be sought on
slit lamp evaluation (handheld is often used during examination under
anesthesia [EUA]). Although RB may rarely produce a pseudouveitis or
tumor hypopyon, signs of intraocular inflammation are more often associated
with granulomatous uveitis, such as toxocariasis, or with other pediatric
uveitis syndromes. Intraocular pressures for each eye should be recorded, and
the cause of any pressure elevation including angle closure or
neovascularization of the iris should be investigated.
Bilateral dilated fundus examination with scleral depression is performed
under general anesthesia. Scleral indentation must be performed for 360
degrees of the globe in order to inspect the entire surface of the retina to
detect anteriorly located tumors. In one series, RB in the periphery of the
fundus was detected using scleral indentation in 65% of cases (166). Color
fundus photos with a wide-angle contact camera and retinal drawings should
be performed to document the size and location of all retinal tumors. This
documentation is essential to plan treatment and to monitor subsequent
treatment response (167).
DIAGNOSTIC STUDIES
Supplementary investigations are frequently used to aid in the diagnosis and
staging of patients referred with RB. Ultrasonography is a useful tool,
particularly for confirming the initial diagnosis and for evaluating features of
intraocular disease. In larger tumors, A-scan echography typically
demonstrates high-intensity echoes within the tumor that correspond to areas
of tumor calcification. The tumor surface nearest the sclera may be anechoic.
B-scan often demonstrates an intraocular mass with scattered highly
reflective echoes within the tumor and with attenuation of normal soft tissue
signals posterior to the tumor. In patients with advanced disease, exudative
retinal detachment, vitreous seeding, and subretinal seeding also may be
demonstrated by ultrasound (Figure 44-7) (168). Fluorescein angiography
(FA) can also be performed. This is useful for staging in two ways; the first is
that if there is a concern for Coats disease, it can help to characterize the
lesion vasculature. More often in the setting of RB, FA can be used to
determine subclinical neovascularization of the iris, which in the setting of
increased intraocular pressure affects staging (150,169).
FIGURE 44-7 A: Ultrasound scan of RB showing a
retinal mass located at the posterior pole. Both (A) and (B)
scans demonstrate that the mass has areas of high internal
reflectivity due to intrinsic calcification. Intrinsic tumor
calcium may produce a shadowing of the soft tissues
behind the tumor. B: CT scanning performed with contrast
demonstrates bilateral RB tumors involving the posterior
poles with dense calcium deposits. C: T1-weighted MRI
scan with gadolinium enhancement demonstrating an
enhancing PNET in a patient with heritable RB.
MRI is used to evaluate for extraocular extension of disease via the optic
nerve, orbit, subarachnoid space, and meninges, as well as to exclude all
PNETs. Gadolinium-enhanced MRI is particularly valuable for evaluating
optic nerve extension or CNS disease. RB is hyperintense on T1-weighted
images and hypointense on T2-weighted images (Figure 44-7) (170–172).
Most experts prefer MRI over CT to avoid any radiation exposure in this
high-risk population.
Bone marrow aspiration and lumbar puncture may be obtained to
evaluate for systemic spread, although it is not routinely performed in most
centers. Other investigations, such as bone scans, are performed based on the
overall clinical assessment of the patient. In most cases, complete metastatic
evaluation is reserved for patients who present with features of highly
advanced disease.
GROUP CLASSIFICATION
At the time of initial clinical evaluation, the patient should be staged, and the
extent of intraocular disease should be classified as a means of assisting the
ocular oncologists in assessing the likelihood of salvaging the eye. The first
such system was proposed by Reese and Ellsworth in the 1960s (173). This
presurgical grouping system proved highly useful for decades throughout the
primary radiotherapy era. However, once this treatment modality was
supplanted by systemic intravenous chemotherapy, a new system was needed
to fill this clinical void. In 2005, Murphree proposed the International
Intraocular Retinoblastoma Classification (IIRC) system for retinoblastoma
(169), which groups eyes from A to E by the extent of disease with Group A
eyes having small tumors far from critical structures and Group E eyes being
anatomically and functionally destroyed by the cancer. The IIRC was
developed with international cooperation among oncologists and
ophthalmologists; however, it has not been strictly followed, and other
classifications with small differences are used across centers (174). The
controversy over differences in classification system has limited the ability of
the RB community to truly compare outcomes across centers (particularly for
advanced Group D and E eyes) and has led to calls for uniformity in
classification. Multicenter clinical trials to assess the management of RB,
such as those performed through the Children’s Oncology Group, also require
a uniform staging system and used the IIRC clinical classification with a
minor variation, which is the size of subretinal fluid allowable in Group B
eyes (3 vs. 5 mm).
In 2017, the eighth edition of the AJCC (175) was published, which
introduced a comprehensive reclassification of the RB grouping system
including the addition of the H “heritable trait” for genetic status of the
patient. This was based on retrospective evaluation between 2001 and 2011
of 1,728 eyes diagnosed from multiple RB centers. Kaplan-Meier survival
analyses of the proportion of eyes salvaged (without the need for external
beam radiation) were evaluated based on clinical features. The salvage of
eyes based on the TNM grouping was the most predictive of all clinical
classification systems and is growing in acceptance among ocular oncologists
and RB centers worldwide.
Families with a new diagnosis of RB require considerable psychological
and emotional support during this period of initial diagnosis. The attending
ophthalmologist, pediatrician, pediatric oncologist, and other health
professionals play an important role in this and in responding to the needs of
the patient, the parents, and other family members with regard to emotional
support and information. The attending ophthalmologist needs to be available
to spend considerable time with the family of a child with newly diagnosed
RB. Showing the parents images of the tumor will frequently assist in the
process of enabling them to accept the recommended treatment plan. Support,
information, and visual documentation of the disease are particularly helpful
when enucleation is being considered. Parents need to be given time to deal
with a new diagnosis of RB. Often, parents are surprised by the initial
diagnosis, and little information from the first consultation is assimilated.
Discussion of the child’s case at a multidisciplinary pediatric tumor board
with consensus on appropriate treatment is frequently reassuring to families.
Many families obtain benefit by referral to a multidisciplinary RB follow-up
clinic, where it is apparent that older children with this diagnosis are leading
full and healthy lives after successful treatment. An inordinate delay in
proceeding to treatment should be discouraged, however, because a delay of
weeks can make a difference in staging and subsequently in salvage. There
should be very minimal delay in definitive therapy for an eye that is unsafe
for the patient due to high pressure from advanced intraocular disease or
signs of optic nerve involvement on MRI imaging (132,176).
MANAGEMENT
A treatment plan should be individualized for each patient and will be
determined by many factors, including the age of the patient and laterality of
disease; the size and location of the ocular tumor(s); the presence of optic
nerve, orbital, or metastatic disease; and, finally, the visual potential of the
affected eye (see also Chapter 45).
In recent years, significant changes have occurred in initial approaches to
the therapy of RB. As the therapy for this disease is complex and involves
expertise across a variety of specialties, management usually is performed
through a multidisciplinary approach at centers of expertise in pediatric and
ocular cancer. Treatment modalities, including surgery, local therapies (laser
ablation, diode hyperthermia, or cryotherapy), intravenous and intra-arterial
chemotherapy (IAC), and brachytherapy or external beam radiotherapy, may
be used. Also important are psychological and social support for the child and
family, visual rehabilitation, achievement of best cosmesis involving experts
in prosthetics, and finally the scheduling of ongoing examinations for the
patient, genetic counseling for the family, and screening for other at-risk
pediatric family members. Given this complexity and the diverse clinical
presentations, the therapeutic approach is tailored to the extent of disease,
needs of the family, and expectation of ability to save the eye and to save any
functional vision.
The majority of children with unilateral RB present with leukocoria. This
presentation usually represents advanced local disease with >60%
involvement of the globe. Enucleation should certainly be considered in these
children because it spares them the toxicity of systemic chemotherapy or
radiation and provides an acceptable cosmetic outcome. It is also a curative
procedure for the vast majority of these patients (177–181). Other clinical
indications for enucleation include advanced neovascular glaucoma
producing pain or RB in an eye that is devoid of visual potential. It should be
noted that while enucleation may not be chosen as primary therapy (e.g., the
therapy initiated at diagnosis), it is often required for persistent or recurrent
tumors not responsive to salvage therapy; this is termed secondary
enucleation.
Whether primary or secondary, the surgical approach to enucleation in
children with RB should facilitate obtaining a long section of optic nerve and
should avoid inadvertent globe penetration. Any tumor at the surgical margin
is an undesirable outcome that is associated with a significantly increased risk
for the development of metastatic disease, and this situation mandates
significant further therapy (86,178,179). If RB is confined to the globe, an
orbital prosthesis is routinely placed at the time of enucleation; therefore, it is
important to preserve the conjunctiva, Tenon capsule, and extraocular
muscles for integration of the implant. It also is important to completely
cover the prosthesis with Tenon capsule and conjunctiva; we use a
meticulous three-level closure to minimize exposure and extrusion and to
provide the best long-term cosmesis. Both porous and nonporous implants
have been used successfully in the management of the anophthalmic socket in
children with RB, and each has a separate side effect profile (177,180,182).
Unilateral cases, particularly if detected at an early stage with reasonable
visual potential, are usually successfully managed by modalities that preserve
the globe. These approaches include systemic or IAC with use of local
therapies including brachytherapy. For small tumors detected early, serial
application of laser ablation, diode hyperthermia, and cryotherapy may
occasionally be sufficient. However, the presence of vitreous seeding
indicates more advanced disease and consolidative therapy alone is rarely
indicated. It is worth noting that for larger localized tumors located more than
2 mm away from the macula and optic nerve or in patients with focal relapse
after local therapy, good responses to brachytherapy may be obtained,
including successful control of mild-to-moderate vitreous seeding close to the
main retinal lesion. Tumors considered for brachytherapy should ideally be
smaller than 15 mm in basal diameter and <8 mm in thickness (183).
Complications associated with plaque placement include radiation
retinopathy and papillopathy and early cataract formation (183). Patients who
have had previous chemotherapy have an increased risk for development of
neovascular complications following brachytherapy, and dose reduction may
be considered (184–187).
Although external beam radiation therapy (EBRT) very effectively treats
RB, it is associated with significant complications, including secondary
tumors inside and outside the radiation field and midfacial bony hypoplasia,
especially if radiation is administered at age <1 year (24). Less serious
complications include growth hormone deficiency; radiation retinopathy,
vasculopathy, and optic neuropathy; neovascular glaucoma; and dry eye
(188–190). A common problem in management following radiation is the
high incidence of cataract (~10%), especially when an anterior port for
therapy is used. Although cataracts can be successfully removed surgically,
an eye with RB cannot undergo intraocular surgery when any concern exists
regarding disease activity, because tumor cells could be disseminated.
Delaying cataract extraction results in visual deprivation during a critical
period and has produced amblyopia in these RB patients. The presence of
cataract can also make small intraocular tumor recurrences that would be
amenable to local therapy difficult to detect (191,192).
Radiation therapy in RB patients has been associated with increased
frequency of second primary malignancies within the radiation field
(24,39,45,132,193–195). The original studies describing this problem
reviewed patient outcomes following treatment with older radiotherapy
techniques, such as orthovoltage sources and single-port delivery systems.
Newer radiation techniques, including lens-sparing techniques, have
considerably reduced the dose received by surrounding tissues during the
treatment of RB (132). RB patients who receive EBRT after age 1 year may
show a lower rate of second tumor development, suggesting that EBRT may
still have a role if therapy is delayed (24).
Intravenous chemoreduction using three-drug systemic chemotherapy
combined with local therapy (photoablation, cryotherapy, or thermotherapy)
has been a mainstay of therapy for bilateral disease since the 1990s; it has
replaced enucleation and EBRT in many patients with heritable RB. The goal
of chemoreduction is to reduce the volume of retinal tumors in order to allow
ablation of residual disease with local therapy. This approach frequently
preserves the eye often with visual potential; however, the likelihood of this
depends on the stage of the eye (25,196–201). With more advanced Group D
disease successfully avoiding EBRT for salvage only in half of the cases
(197), although with newer modalities this is somewhat improved (202),
concomitant local therapy is needed to definitively treat the tumor. Thus, a
treatment plan consists of a number of cycles of chemotherapy during which
patients also receive local therapy. A number of studies suggest that local
therapy acts synergistically with chemotherapy, possibly by increasing the
uptake of the chemotherapeutic agents into the eye. The agents currently used
for systemic chemotherapy include carboplatin, vincristine, etoposide (VP-
16), and sometimes cyclosporin A. The dosage range varies across centers,
although standard regimens have been developed by protocols sponsored by
the Children’s Oncology Group of the National Institutes of Health. In
general, higher-dose regimens are used for patients with larger tumors
associated with vitreous and subretinal seeding. Children with smaller tumors
may respond well to lower-dose or fewer cycle protocols (201).
All chemotherapeutic agents are associated with potential toxicity.
Carboplatin may produce otologic toxicity and high-frequency hearing loss
that is unacceptable in children who already may have reduced vision.
Pretreatment audiologic testing and ongoing surveillance for the development
of hearing loss are imperative (203). Long-term mutagenesis and reduced
fertility have also been reported. Vincristine (VP-16) may have major
neurologic toxicities, including numbness and tingling in the extremities and
loss of deep tendon reflexes, which may be reversible with dose reduction or
cessation of therapy (204). Severe constipation may also be observed with
this agent; to prevent paralytic ileus, it is not prescribed for children <3
months of age per the Children’s Hospital Los Angeles protocol (196). The
dose-limiting toxicity for VP-16 is leukopenia that occurs during treatment.
Leukopenia may be treated by bone marrow–stimulating medications. The
risk also exists for development of nonlymphocytic leukemia associated with
a translocation at chromosome 11q23. This disease appears a short time
interval (usually between 1 and 3 years) after cessation of VP-16
chemotherapy. Although rare, it appears that the standard doses used for
chemoreduction in RB may be associated with a low risk for the development
of this devastating complication (205). Additionally, nausea, vomiting,
diarrhea, and stomatitis are observed in 15% of patients receiving VP-16
intravenously (204).
Systemic chemotherapy may be associated with other problems. Multiple
anesthetics are needed, both for application of local treatment and for
increased surveillance required to detect new tumors following completion of
this treatment. Chemotherapy remains less effective against very large tumors
and vitreous and subretinal seeding. In advanced cases, enucleation may be
required for the worse eye in bilateral disease. Finally, for the heritable form
of RB, we have not defined the potential mutagenicity of these
chemotherapeutic agents will have upon the incidence of second cancers in
later life.
Patients with persistent or relapsed intraocular disease that cannot be
controlled by standard consolidative therapy have a number of treatment
options still available to them. Such patients may be candidates for increased
dose chemotherapy with the addition of subconjunctival carboplatin, although
given the associated periocular fibrosis, this therapeutic approach is less
frequently utilized (206). Some centers employ cyclosporin A that may be
added to the chemotherapeutic regimen; this agent has been shown to be
effective in managing advanced intraocular disease, including vitreous and
subretinal seeding (207). Cyclosporin A competitively inhibits the P-
glycoprotein (P-gp) pump. Up-regulation of this pump reduces the
concentration of some chemotherapeutic agents in cancer cells by pumping
agents out of the cell into the extracellular space. The expression of P-gp
appears to be up-regulated on tumor cells by prior exposure to
chemotherapeutic agents and may be a source of resistance to chemotherapy.
Unfortunately, cyclosporin A is associated with increased toxicity, including
renal dysfunction, tremor, hypertension, hyperlipidemia, hirsutism, and gum
hyperplasia (204). EBRT remains an effective option in the treatment of RB,
and ERBT may have a role in salvaging an only eye that has not responded to
other therapies when the only alternative is enucleation. The majority of eyes
that are initially treated with intravenous chemotherapy and consolidation,
without appropriate disease response, are currently treated with IAC.
Starting in the 2000s, with the goal of increasing rates of salvage for
advanced disease, and decreasing systemic toxicity, there has been a move
toward a more localized approach to chemotherapy by administering the
drug(s) directly into the ophthalmic vasculature and bypassing the systemic
circulation (208–223). IAC was initially developed by Kaneko in Japan
decades ago (224). He devised a technique in which a balloon catheter was
introduced into the femoral artery and guided just past the ostia of the
ophthalmic artery via the carotid under fluoroscopy. After inflation of the
balloon, melphalan was injected into the intraocular vasculature. Abramson et
al. modified the technique by inserting the catheter directly into the lumen of
the ophthalmic artery. Initially, it was described in the setting of advanced
unilateral disease but has since been subsequently modified for bilateral RB.
It can be administered as a primary and salvage technique with various agents
such as carboplatin, topotecan, and methotrexate injected in a similar fashion,
although melphalan is the most commonly used agent. Intraoperative ERG
has been used to monitor the ocular toxicity of these agents on the retina
(225). In many cases, three injections of single-agent IAC with adjuvant local
therapy can cure an eye of advanced RB with higher success rates per group
than the standard six cycles of intravenous chemotherapy and with reasonable
visual outcomes. As with any therapy, there are associated complications
including vascular events, intraocular hemorrhage, and stroke (226–228).
While IAC is more effective at ocular salvage than intravenous
chemotherapy, it does not provide appropriate systemic coverage for high-
risk pathologic tumor invasion. It is hypothesized that due to this, the risk of
metastatic disease is slightly (1% to 3%) higher with IAC over intravenous
chemotherapy, but the exact incidence is hard to define (cite). Recently,
deaths from pathologic spread of RB have been reported in the scientific
literature (208) and popular press.
INTRAVITREAL CHEMOTHERAPY
Intravitreal chemotherapy (IVC) was introduced initially in the 1962 to treat
vitreous seeding, a common cause of tumor relapse, but was abandoned due
to concerns of extraocular spread (229). In 2012, Munier introduced a safety-
enhanced procedure for intravitreal injection of chemotherapy (230). As an
initial safety mechanism, a limbal paracentesis with removal of aqueous
humor is performed to lower the intraocular pressure and prevent reflux of
active seeds to the scleral injection site. The chemotherapy injection can then
be performed via the pars plana in a quadrant of the eye confirmed free of
tumor by ultrasound biomicroscopy. A 32-gauge needle is placed into the
midvitreous cavity and visualized behind the lens during injection.
Cryotherapy is then applied at the injection site as the needle is
withdrawn. The eye is shaken to distribute the chemotherapy, and the surface
is bathed in sterile water, which is cytotoxic. The described toxicities of this
treatment modality are retinal pigment epithelium mottling, hypotony,
anterior segment inflammation, and cataract formation (153).
Rarely, severe complications such as maculopathy, optic neuropathy,
phthisis, and acute hemorrhagic retinopathy (231) can occur (Figure 44-8).
The efficacy for treating vitreous seeds with intravitreal injections has been
reported to be 100% at many centers and has enabled many children to be
successfully treated without EBRT (152,202,232–234).
FIGURE 44-8 A–E: Fundus photographs of the left eye

of patient 1 (A) and the right eye of patient 2 (E) revealing
a partially treated retinoblastoma with associated vitreous
seeding located in the macula and nasal to the optic disc,
respectively. B and F: Fundus photographs of patient 1
(B) and patient 2 (F) revealing diffuse retinal edema with
associated intra- and preretinal hemorrhages. C and G:
Midphase fluorescein angiography of the left eye of
patient 1 (C) and the right eye of patient 2 (G) revealing
areas of blockage corresponding to the areas of retinal
hemorrhage and intraocular tumor, with vascular
sheathing. D and H:Fundus photographs at the follow-up
of patient 1 (D) and patient 2 (H) revealing diffuse
chorioretinal atrophy in the posterior pole with a
demarcation between the normal and the atrophic retina.
(Reproduced with permission from Aziz HA, Kim JW,
Munier FL, et al. Acute hemorrhagic retinopathy
following intravitreal melphalan injection for
retinoblastoma: a report of two cases and technical
modifications to enhance the prevention of retinal toxicity.
Ocul Oncol Pathol 2017;3(1):34–40.)
METASTATIC DISEASEMetastatic RB has traditionally been associated

with a poor prognosis. However, encouraging results from the Children’s
Oncology Group (235) and others (236–238) have been reported with the use
of high-dose systemic and intrathecal chemotherapy and total body
irradiation with bone marrow rescue, resulting in disease-free survival up to
80 months after diagnosis
Other studies have demonstrated effective treatment of orbital disease
with a combination of surgery and high-dose chemotherapy (239,240).
Despite attempts at aggressive treatments for PNET with systemic and
intrathecal chemotherapy and craniospinal irradiation, outcomes for this
condition remain disappointing (41–43).
Beyond the life- and sight-threatening implications, the diagnosis of RB
has many far-reaching and long-term consequences for the lives of the
affected individual and family members. This is particularly true for some
patients carrying a genetic cancer predisposition syndrome, which can be
passed on to future children. Other challenges for RB patients and families
include concerns around cosmesis and body image, visual rehabilitation,
special educational needs, and success at mastery of knowledge and tasks that
will have an impact on the patient’s psychosocial adaptation and integration.
Psychological effects on other members of the family may also play a role in
how successfully the entire family copes with the patient’s disease process.
Because RB involves accepting a very serious diagnosis, an often long,
difficult treatment process, and many years of follow-up examinations, the
developmental, social, and psychological health of the patient and family
should be considered throughout their experience with this disease.
FOLLOW-UP AND MONITORING OF

CHILDREN WITH RETINOBLASTOMA
In the heritable form of RB, significant risk for the development of further
retinal tumors exists throughout childhood, and children should be closely
and serially monitored during this period. Close follow-up is particularly
necessary with chemoreduction and associated serial local treatment. The
frequency of examination depends upon the age of the patient, stage of
treatment, and estimated risk for further tumors in that individual. Children
with sporadic unilateral RB should also be examined under anesthesia
serially if genetic testing has not ruled out a germ-line mutation, as 15% of
unilateral cases represent heritable disease. Any patient with a germ-line RB1
mutation remains at risk to develop disease in the other eye, develop
secondary nonocular tumors later in life, and risk of passing the mutation to
future offspring. Some centers screen heritable patients for development of
PNETs with MRI every 6 months until the age of 3.
Heritable RB is a lifelong illness, potentially familial, which confers an
increased risk for the development of other malignancies in the patient and
their future children. For this reason, we follow all children with RB in a
multidisciplinary clinic composed of experts with different perspectives and
unique areas of expertise. As a group, these physicians monitor, evaluate, and
intervene in a timely fashion to minimize adverse consequences of RB as a
diagnosis. After age 7 years, we see RB patients annually and schedule
siblings on the same day to reduce the number of required visits for the
family.
In the ocular oncology clinic, the patient’s vision and refraction are
checked. A complete ophthalmologic examination is undertaken to exclude
reactivations in the primary tumor(s) and to look for evidence of treatment
complications, including radiation- or chemotherapy-related retinopathy or
vasculopathy. Whenever possible, visual rehabilitation is undertaken. An
ocularist is available to optimize the fitting of the prosthesis over the orbital
implant to maximize cosmesis. A pediatric oncologist and a school specialist
perform a complete medical history and physical examination to evaluate
general health and to exclude problems related to previous therapy. Detailed
review of systems and physical examination are undertaken to exclude the
development of second malignancies. We have not found that routine bone
scans used to detect osteogenic sarcoma yield any earlier diagnoses, although
some centers perform routine MRI of the long bones for screening purposes.
If applicable, patients are also monitored for endocrine deficiencies as
well as for problems with midfacial hypoplasia that can develop in the setting
of radiation therapy. Educational and nursing specialists evaluate the children
to identify behavioral or learning difficulties. The multidisciplinary
environment also fosters strong support between RB families. Newly
diagnosed families are often relieved to see healthy older children who carry
the RB diagnosis. Information on support services, visual devices, braille
books, and other practical considerations are provided by clinic personnel as
well as by discussions between families. Long-term friendships between RB
families emerge in this setting.
Retinoblastoma Screening
When the RB mutation is detected in a family, precise individualized genetic
counseling is provided. However, in some cases, the RB gene mutation is not
identified, usually because of lack of access to genetic testing due to
insurance restrictions (241). In this situation, even if the proband has
unilateral RB, offspring and siblings born into a family with RB are
examined first at birth and then again within the first weeks of life in order to
detect intraretinal tumors while they are small and amenable to local
therapies. OCT can be critical during this time period (158,242). Periodic
dilated ophthalmoscopy with a depressed examination to the ora 360 degrees
is continued throughout infancy and childhood as siblings with initially
normal examinations occasionally can develop tumors months after the initial
normal examination (135). We now recognize that germ-line mosaicism for
RB1 mutations occurs in at least 10% of RB families (134). This finding,
along with recognition of low-penetrance kindreds, suggests that a significant
minority of patients exists who have inherited a mutant RB1 gene from an
asymptomatic carrier. Without definitive genetic testing available for some
families, at the present time, we regard these RB patients as potentially
heritable cases. If genetic testing is not available to inform the true risk of
disease, serial screening examinations under anesthesia are justified in order
to detect this significant minority of patients with heritable disease before
symptoms arise.
PROGNOSIS
With steady advances in the diagnosis and treatment of RB, children with this
disease now have an excellent prognosis for life with decreased ocular and
visual morbidity. In developed countries, the mortality rate associated with
RB has fallen to <5%. Although the risk for metastatic disease in children
with bilateral RB appears to be higher than for individuals with unilateral RB,
the survival from metastatic disease is similar for both groups (243).
Survivors with heritable RB have a worse long-term prognosis than patients
with nonheritable RB because germ-line RB1 mutation confers a
predisposition for the development of second nonocular cancers (45). A 50%
overall mortality has been observed 25 years after the diagnosis of bilateral
RB (244). This statistic emphasizes the need to counsel patients with
heritable RB that careful follow-up for second tumor development is required
and that annual second tumor screening should be a family priority.
The outlook for vision in RB has improved with the development of
treatments that preserve vision and avoid enucleation. In one recent series of
74 children, the enucleation rate was 62%, and 58% of patients had a visual
acuity in the better eye between 20/20 and 20/40. Only 9% of patients
demonstrated visual acuity of 20/400 or worse. The final visual acuity in this
series was influenced by multiple factors, most notably tumor location. The
final visual outcome could not be predicted by the disease stage at initial
presentation (2) although macular tumor location and extensive retinal
detachment remain risk factors for poorer visual outcomes (3).
Management of metastatic RB continues to be a difficult clinical
problem. One factor that improves the prognosis for survival in patients with
metastatic disease is early detection. A number of studies have examined risk
factors for the development of metastatic disease. Early recognition of these
risk factors and prompt treatment may help to reduce mortality associated
with disease dissemination (86). Major clinical risk factors for the
development of metastatic RB include advanced intraocular disease at
diagnosis, delays in diagnosis, or history of prior intraocular surgery
(86,244–246). Accepted histopathologic risk factors include tumor cells
extending to the surgical margin of the optic nerve and microscopic
extraocular involvement by tumor cells (84,240). While multimodal therapies
have improved the overall survival for extraocular disease (235,238), the
importance of primary enucleation for highly advanced eyes (247) with
adjuvant therapy for high-risk pathologic features (248) cannot be
understated.
The three priorities in the management of patients with RB in descending
order are salvage of life, salvage of eye(s), and salvage of vision. In all
unilateral cases with a normal contralateral globe, patients should be screened
for refractive errors, placed in shatterproof lenses, and follow monocular
precautions.
It may be difficult to predict final visual acuity in cases of advanced
bilateral disease. Patients with significant focal scarring in the macula and
fovea of their remaining eye have been known to retain better than expected
vision. As described elsewhere, cataract extraction may need to be deferred
until all intraocular tumors are stable and completely regressed. A pediatric
ophthalmologist can serve a critical role on the RB team by aiding in
decisions regarding patching, maximizing refraction, and visual rehabilitation
for these children.

As outlined throughout this chapter, the treatment of RB is multidisciplinary
in nature and requires highly subspecialized care (see the above section on
Follow-Up and Monitoring of Children with Retinoblastoma). In addition to
ocular and pediatric oncologists, other tertiary fields of medicine play a
central role in managing this disease including ocular pathology, radiation
oncology, anesthesiology, genetics, and interventional radiology. Ocularists,
psychologists, and pediatric specialists are additional members of the heath
care team that can significantly affect the rehabilitation of these children.
ETHICAL CONSIDERATION
Access to specialized care remains a significant obstacle to the care of
children with RB in the developing world. As newer modalities such as IAC
become available worldwide, it is critical that experts acknowledge the
lifesaving role that early detection and primary enucleation still play in areas
where more expensive modalities may not be available. World outreach
programs are underway to reduce mortality from RB in the developing world
(249).
SUMMARY AND FUTURE

TREATMENTS
Despite significant advances in our understanding and management of RB,
challenges remain ahead. A better understanding is needed of the molecular
events surrounding RB cell death, angiogenesis, and metastasis. While
genetic testing for RB has become the standard in most specialist centers,
insurance coverage is still a real-life barrier.
An immediate and ongoing priority is the discovery of new therapeutic
options with better efficacy and lower toxicity, as well as a clearer definition
of which treatments should be applied for different stages of disease. A
number of different approaches are being studied at various centers. Porcine
and primate animal models for IAC are being developed to further assess this
modality. The recent safety-enhanced procedure for intravitreal injection of
chemotherapy has changed the management of vitreous seeds dramatically
and led to improved outcomes for globe salvage (230). Other alternatives to
local therapies are being investigated including a transscleral implant, which
is nearing phase 1/2 clinical trials and was recently successfully used for the
first time in Canada on a relapsed eye (personal communication with the
author). While the increasing trend toward local therapy and minimizing
systemic toxicity is a benefit for this disease, critical steps are needed in order
to personalize the care of RB patients. Use of tumor biomarkers has
revolutionized the diagnosis, prognosis, and approach to breast, lung, and
other cancers. This has been limited in RB due to contraindication to tumor
biopsy in this disease. However, recent research on use of the aqueous humor
as a surrogate to tumor biopsy has opened the door for evaluation of
biomarkers and correlation with clinical therapeutic response (250,251). This
will no doubt provide critical insight into tumorigenesis for this rare cancer as
well as possibly targets for personalized, directed therapy. Finally, more
effective clinical screening remains a clinical need. Education of primary care
physicians and the community, particularly parents of young children, is a
priority in order to detect this disease at an earlier stage.
ACKNOWLEDGMENT
This work was supported by the National Eye Institute, Bethesda, Maryland
(grant #1RO1EY023557-01) and the Department of Ophthalmology at the
Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA.
Funds also come from the Vision Research Core Grant (P30 EY001583),
F.M. Kirby Foundation, Research to Prevent Blindness, The UPenn Hospital
Board of Women Visitors, The Paul and Evanina Bell Mackall Foundation
Trust, and, the NEI intramural program under eyeGENE® - protocol 06-EI-
0236 funded in part under Contract No. HHS-N-263-2017-00001-C. The
sponsor or funding organization had no role in the design or conduct of this
research. Dr. Berry has grant support from National Cancer Institute of the
National Institute of Health Award Number K08CA232344, TheWright
Foundation, the Knights Templar Eye Foundation, American Cancer Society
#IRG-16- 181-57, Hyundai Hope on Wheels, Childhood Eye Cancer Trust.
Dr. Berry has indirect support from The Larry and Celia Moh Foundation,
The Institute for Families, Inc., Children’s Hospital Los Angeles, an
unrestricted departmental grant from Research to Prevent Blindness.
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45
Treatment of Retinoblastoma
Lauren A. Dalvin, David Ancona-Lezama, and
Carol L. Shields
INTRODUCTION
Retinoblastoma management has evolved over the past several decades, but
the ultimate goals of care remain unchanged with preservation of life and the
globe, if possible, and maximization of visual acuity (1). The primary goal of
retinoblastoma therapy is cure of the malignancy with prevention of
metastatic disease. Secondary goals are to preserve the eye, prevent related
pineoblastoma, and minimize secondary systemic cancers (1). There is a
tertiary goal in management that focuses on protection of useful visual acuity.
With earlier detection and remarkable treatment advances, globe salvage with
preservation of visual acuity is possible in more cases than ever before.
Treatment of retinoblastoma can be complex and depends on several
factors such as patient age, germline genetic status, anticipated visual acuity,
globe classification, and ultimate risks for metastasis. Treatment is
individually tailored to each patient (Figure 45-1). The various treatment
modalities include systemic intravenous chemotherapy (IVC), intra-arterial
chemotherapy (IAC), intravitreal chemotherapy (IvitC), intracameral
chemotherapy (ICC), focal therapies (cryotherapy, laser photocoagulation,
transpupillary thermotherapy [TTT]), external beam radiotherapy (EBRT),
plaque radiotherapy, and enucleation. In this chapter, we discuss current
retinoblastoma treatment modalities, including applications, outcomes, and
side effects.
FIGURE 45-1 Management of retinoblastoma: decision
tree by patient presentation. Decision tree is a rough
guideline only. Real life management can be much more
complex. If tumor control cannot be achieved, optic nerve
becomes involved or cannot be visualized for 3
consecutive months, or the eye becomes blind and painful,
consider enucleation with or without adjuvant systemic
chemotherapy. IAC, intra-arterial chemotherapy; IvitC,
intravitreal chemotherapy; EBRT, external beam radiation
therapy.
RETINOBLASTOMA CLASSIFICATION
Retinoblastoma is classified prior to any treatment administration, according
to the features found in each globe independently (2). The classification is
listed in Table 45-1.
TABLE 45-1 International classification of

retinoblastoma (ICRB)
INTRAVENOUS CHEMOTHERAPY
Systemic IVC for retinoblastoma was introduced in the early 1990s and
remains an essential component of treatment in many cases. For many
patients, IVC is a preferred first treatment choice, particularly in patients with
bilateral disease, family history of retinoblastoma, known germline mutation,
presenting patient weight <6 kg, or cases suspicious for optic nerve or
choroidal invasion (Figure 45-2) (3). IVC can treat local disease in the eye as
well as prevent distant metastasis and second cancers. In patients with
germline retinoblastoma, IVC is especially important because it can protect
against the development of pineoblastoma and secondary cancers (4–6).
FIGURE 45-2 Group B retinoblastoma managed with

systemic intravenous chemotherapy (IVC). (A) Before and
(B) 2 years after six cycles of IVC using vincristine,
etoposide, and carboplatin with transpupillary
thermotherapy (TTT) consolidation.
IVC typically consists of two, three, or four chemotherapy agents delivered

each month for a total of six to nine cycles (3). The most frequently used
regimen consists of three drugs, including vincristine, etoposide, and
carboplatin (VEC) (7). Drugs are dosed by patient weight for patients with
presenting age <36 months or weight <10 kg, with dosing per square meter
for patients exceeding such limits (Table 45-2) (1). IVC is sometimes referred
to as chemoreduction due to the chemotherapy impact on the tumor with
reduction in size, and focal consolidation with cryotherapy or TTT can
improve tumor control. Focal treatment is generally performed on the same
day, immediately preceding IVC, to improve chemotherapy penetration into
the tumor (8). In most centers, focal consolidation is key to IVC success.
TABLE 45-2 Management of retinoblastoma:
dosing schedule for systemic, intra-arterial, and
intravitreal chemotherapy
Various regression patterns can be observed following IVC, including

complete disappearance of the tumor with no residual scar (type 0), a
completely calcified mass (type I), a completely noncalcified mass (type II), a
partially calcified mass (type III), or a flat atrophic scar (type IV) (9). Thinner
tumors usually exhibit type 0, type II, or type IV regression, while thicker
tumors tend to have regression type I or type III (9). Efficacy of IVC for
tumor control and globe salvage varies by International Classification of
Retinoblastoma (ICRB) group. In a study of 249 consecutive eyes, IVC
achieved globe salvage in 100% of group A eyes, 93% of group B, 90% of
group C, 47% of group D, and 25% of group E eyes (7). Alternative or
adjuvant treatments further discussed in this chapter can improve globe
salvage for advanced group D and E eyes.
Systemic toxicity from IVC is mild. As with most systemic
chemotherapy, transient alopecia, cytopenia, and fever can occur (10,11).
While transfusion of blood products is occasionally required, support with
hematopoietic growth factor is not required with standard VEC dosing (10).
Chemotherapy can often be delivered through peripheral intravenous access
without the need for a central line. Thus, broad-spectrum antibiotic coverage
is required for febrile neutropenia but not fever in the setting of normal blood
counts (10). Patients should, however, receive routine prophylaxis for
Pneumocystis jirovecii pneumonia (10). Chemotherapy-related nausea,
vomiting, and constipation can be medically managed. Ophthalmic toxicities
from IVC have not been observed.
Long-term renal toxicity is rare when medications are appropriately
dosed. Likewise, hearing loss is uncommon, found in 0 of 164 and 2 of 175
children treated with systemic carboplatin (12,13). When used in the
recommended doses, infertility following IVC generally does not occur, but
if additional therapy with melphalan is needed, then infertility in males can
occur, especially when delivered in excess of 140 mg/m2 cumulatively (14).
While there have been reports of secondary acute myelogenous leukemia
(AML) following IVC for retinoblastoma, these cases have been associated
with higher than recommended doses of chemotherapy, concomitant EBRT,
or other predisposing systemic conditions (6,15). Thus, the risk of secondary
AML following standard dose VEC for retinoblastoma in the absence of
adjuvant EBRT is low (16).
INTRA-ARTERIAL CHEMOTHERAPY
The use of IAC for retinoblastoma was introduced in 2004 in Japan and later
in the United States as a targeted therapy for retinoblastoma. This modality
has earned a particularly important role in retinoblastoma management,
especially for unilateral tumors (17–20). IAC is employed as primary therapy
for nongermline, unilateral, group B, C, D, or E retinoblastoma or as
secondary therapy for unilateral or bilateral advanced disease with recurrence
or failure to regress following IVC or plaque radiotherapy (Figures 45-3 and
45-4) (1,3,21,22). Some centers use IAC as primary therapy for bilateral
disease as tandem therapy. IAC is effective for treatment of subretinal seeds,
even those that are extensive and involving two or more quadrants. IAC can
also be effective for treatment of vitreous seeds, especially when seeding is
limited and in close proximity to the retina (3). If IAC is used as primary
therapy in patients with known or suspected germline mutation, careful
consideration must be given to the need for subsequent systemic IVC for
prevention of metastasis, pineoblastoma, and second cancers (4–6). Likewise,
caution is advised, especially for group D or E eyes, especially if there is
suspicion for high-risk features, including tumor invasion of the optic nerve,
uvea, or anterior chamber. (See section on high-risk retinoblastoma below.)
High-risk retinoblastoma warrants enucleation and additional systemic
chemotherapy to prevent metastatic disease (23). Due to anatomic challenges
with arterial catheterization in small-caliber vessels, use of IAC is typically
limited to patients older than 3 or 4 months (3). In younger patients, IVC can
be used as bridge therapy until weight reaches 6 kg, allowing for safe
delivery of IAC (24).
FIGURE 45-3 Group D retinoblastoma managed with

intra-arterial chemotherapy (IAC) without complications.
(A) Before and (B) 1 year after three cycles of IAC using
melphalan 5 mg and topotecan 1 mg. (C) Before and (D) 1
year after three cycles of IAC using melphalan 5 mg.
FIGURE 45-4 Group E retinoblastoma managed with
intra-arterial chemotherapy (IAC) without complications.
(A) Inadequate tumor regression after systemic
chemotherapy confirmed by (B) B-scan ultrasonography.
(C) Tumor regression 2 years after IAC using melphalan 5
mg and topotecan 1 mg confirmed by (D) B-scan
ultrasonography.
IAC is a complex procedure ideally performed by an experienced

neurosurgeon or interventional neuroradiologist (25). A microcatheter is
inserted into the femoral artery and fluoroscopically guided to allow
supraselective delivery of potent chemotherapy via the ophthalmic artery
(25). Chemotherapy consists of one or two drugs, typically delivered once per
month for a total of three sessions (3). Additional sessions can be delivered
depending on tumor response, with careful attention to dose-limiting local
and systemic side effects. For moderate disease, treatment usually consists of
single-drug therapy with melphalan dosed at 5 mg (3). More severe disease
can be treated either by escalating the dose or by adding other agents. A
commonly used combination is two-drug therapy consisting of 5 mg or 7.5
mg melphalan plus 1 mg topotecan (Table 45-2) (3). While carboplatin was
used in the early IAC era, high rate of ophthalmic toxicity has limited its
utility in some centers (26,27).
Compared with IVC, IAC results in 10 times the chemotherapy dose
delivered directly to the eye, which accelerates tumor regression (23). A 4-
year experience study found that IAC achieved globe salvage in 82% of eyes
when used as primary therapy and 58% of eyes when used as secondary
therapy (19). Subsequent studies found that efficacy of IAC for tumor control
and globe salvage varied by ICRB group. In a 5-year experience study of 70
consecutive eyes, IAC achieved globe salvage in 100% of group B, 100% of
group C, 94% of group D, and 36% of group E eyes, representing an
improvement in globe salvage compared to IVC for group D (94% vs. 47%)
and group E (36% vs. 25%) eyes (7,28). Comparing location of tumor
burden, IAC achieved regression in 94% of cases for solid tumor, 95% for
subretinal seeds, and 87% for vitreous seeds (28). A retrospective
comparative analysis of IAC versus IVC for unilateral retinoblastoma in 91
cases revealed significantly improved globe salvage with IAC (vs. IVC) for
group D (91% vs. 48%, P = 0.004), but no difference for groups B, C, or E
(29). In that analysis, tumor control was significantly better with IAC for
solid tumor (92% vs. 62%, P = 0.002), subretinal seeds (86% vs. 31%, P =
0.006), and vitreous seeds (74% vs. 25%, P = 0.006) (29). Adjuvant IvitC,
discussed later in this chapter, can improve globe salvage for advanced group
D and E eyes manifesting extensive vitreous seeds.
Despite localized delivery of chemotherapy, systemic toxicity following
IAC can occur, including neutropenia in up to 12% of patients, which is
usually transient and does not require transfusion (30). Hypoxia, hypotension,
and bradycardia can occur in 24%, and patients must be monitored for
contrast-related nephrotoxicity (23,31). Systemic vascular events, such as
femoral artery occlusion and blue toe syndrome, can be managed and
reversed with anticoagulation (19,32). Serious neurologic complications,
including carotid artery dissection, stroke, and death, are rare but can occur
(23). Patient selection is critical, as undetected optic nerve or choroidal
invasion can lead to metastasis when patients are managed with IAC alone
without systemic chemotherapy (23).
Local periocular side effects and serious ophthalmic vascular events can
be observed with IAC due to chemotherapy drug toxicity and IAC technique.
Side effects, which are often self-limited, can include periorbital edema,
cutaneous hyperemia, madarosis, blepharoptosis, scalp hair loss, and
extraocular motility abnormalities (26,33,34). Serious ophthalmic vascular
events include choroidal ischemia, branch or central retinal artery occlusion,
ophthalmic artery spasm or occlusion (Figure 45-5), vitreous hemorrhage,
and others (26–28,35–37). A recent study of 243 infusions in 76 eyes
performed over 9 years demonstrated that event rates have decreased over
time, correlating to increased experience of the treating neurosurgeon or
interventional neuroradiologist (27). Vascular events did not correlate to
patient age, number of IAC infusions, cumulative dosage of chemotherapy, or
use of melphalan alone versus melphalan plus topotecan, highlighting the
importance of physician experience in performing this procedure (27).
Vascular events do not correlate with decreased globe salvage but can limit
visual acuity (27). Risk for vascular events is similar when IAC is used as
primary or secondary therapy (38).
intra-arterial chemotherapy (IAC) with ophthalmic artery
occlusion. (A) Intact retinal and choroidal circulation with
active tumor confirmed by (B) fluorescein angiography.
(C) Retinal and choroidal atrophy 7 years after treatment
with IAC using melphalan 5 mg and carboplatin 30 mg,
with poor retinal and choroidal perfusion confirmed by (D)
fluorescein angiography.
INTRAVITREAL CHEMOTHERAPY
The initial use of IvitC in the United States was by our team in 2006, based
on early observations from Japanese investigators (39). This therapy was later
widely adopted in the United States and Europe in 2012 (40–42). Despite
significant improvements in survival, tumor control, and globe salvage in the
IVC and IAC eras, group D and E eyes often necessitated enucleation or
external beam radiation for vitreous seed recurrence (3,43,44). In 2003,
Kaneko and Suzuki introduced IvitC to the modern armamentarium of
available treatments for retinoblastoma (1,39,45). IvitC is now commonly
used in combination with IAC to achieve tumor control in eyes that otherwise
would have been enucleated, especially in advanced group D and E tumors
(Figures 45-6 and 45-7). IvitC is currently indicated for the presence of
persistent (refractory) or recurrent vitreous seeds from retinoblastoma
following other treatments. Most ocular oncologists use IvitC as secondary
therapy given limited efficacy on the primary tumor. Relative
contraindications for IvitC include presence of tumor or vitreous seeds at the
planned site of needle entry, tumor invasion of the pars plana, and anterior
chamber seeding. Careful clinical examination with or without ultrasound
biomicroscopy can be employed to rule out pars plana involvement prior to
IvitC.

intra-arterial chemotherapy (IAC) and intravitreal
chemotherapy (IvitC). (A) Extensive vitreous seeding with
(B) regression 2 months after IvitC using 30 μg melphalan
for 6 cycles.
FIGURE 45-7 Group E retinoblastoma with anterior
chamber seeding managed with intra-arterial
chemotherapy (IAC) and intravitreal chemotherapy
(IvitC). (A,B) Extensive vitreous and anterior chamber
seeding with (C,D) regression of seeds after 6 cycles of
IvitC.
Melphalan was confirmed to be the most efficient drug in suppressing

colonies of retinoblastoma compared to 12 other chemotherapy regimens, and
topotecan was subsequently found to have similarly potent in vivo activity
(40,41,46,47). Early reports from Kaneko and Suzuki using low doses (8 μg)
of intravitreal melphalan in the treatment of retinoblastoma showed positive
results with 51% globe salvage. Higher doses (50 μg) were also explored, but
tumor control had a high cost of toxic side effects. Based on animal models,
the extrapolated human dose of melphalan at which a balance was met
between efficacy and toxicity was 20 to 30 μg (40,48). We recommend
intermediate range doses of 20 to 30 μg every 2 to 4 weeks to balance tumor
control with toxic effects (Table 45-2). The standard dose is 20 μg, but we
employ 30 μg in cases that show dense vitreous seeding. Topotecan is used at
the same dosage, either alone or in combination with intravitreal melphalan
(Table 45-2). Some centers base dose on patient age (49).
After confirming the absence of contraindications, the injection site is
chosen, preferably at least two clock hours away from vitreous seeding to
avoid contact with active tumor and potential for extraocular extension (41).
When the intended injection volume surpasses 0.1 mL, especially when
injecting more than one drug, an anterior chamber paracentesis is performed
prior to the intravitreal injection. The volume withdrawn from the aqueous
humor is generally 0.1 mL and accommodates the planned volume of
intravitreal drug to be injected in order to both prevent optic nerve damage
from elevated intraocular pressure and prevent vitreous reflux and seed
dissemination secondary to ocular hypertension (41). Some centers use
digital massage to lower eye pressure prior to injection (50). The distance
from the limbus for injection is chosen based on patient age (51,52). The
injection is administered 1.5 mm from the limbus for children age 0 to 6
months old, 2.0 mm for children 6 to 12 months old, 2.5 mm for children 1 to
2 years old, 3.0 mm for children 2 to 3 years old, and 3.5 mm for children >3
years old (51). A 32-gauge needle is mounted on a syringe with the mixed
chemotherapeutic agent. A separate needle and syringe is used for each drug
if multiple drugs are to be given. If melphalan is used, care must be taken to
use the drug promptly after mixing, because melphalan has a short half-life of
approximately 60 minutes. The needle is pointed toward the center of the
vitreous cavity, and the bolus is injected into the mid vitreous as a continuous
jet. Following injection, the needle is withdrawn while simultaneous triple
freeze–thaw cryotherapy is delivered to the entry site. The needle is not
completely withdrawn until an ice ball has formed around the needle track.
The eye is gently jiggled back and forth continuously for 30 seconds to
achieve a homogenous distribution of the chemotherapeutic agent throughout
the entire vitreous cavity, and copious irrigation of the ocular surface is
performed with sterile saline to prevent toxic keratopathy. Indirect
ophthalmoscopy is performed to identify possible immediate complications.
Topical steroid antibiotic ointment is applied without patch. Parents are
instructed to avoid eye drops, swimming, rubbing, or other manipulation of
the eye for 7 days following the procedure.
The treatment response and complications are assessed every 2 to 4
weeks prior to the next injection. The treatment success is established if seeds
show signs of regression, including complete disappearance (regression type
0), calcification (regression type I), amorphous morphology (regression type
II), or combination of the latter two (regression type III) (49). Francis et al.
reported different types of clinical response to IvitC based on vitreous seed
classification as dust, spheres, or clouds (50). Dust responded after a mean of
3 weeks and three injections of intravitreal therapy, whereas spheres
responded at a mean of 7 weeks and five injections and clouds at a mean of 8
months and eight injections (50). Thus, clouds can require a longer course of
treatment.
IvitC is most beneficial for advanced group D and E eyes. In particular,
in group E eyes, globe salvage improved from 25% with IAC alone to 73%
with combined therapy (IAC + IvitC) (43). Since then, further studies have
reported impressive rates of tumor control and globe salvage with
combination therapy, ranging from 87% to 100% (3,41–43,53–55).
Despite the proven efficacy of IvitC in treating vitreous retinoblastoma
seeds, ocular toxic side effects can occur. Based on animal studies by
Shimoda et al., the estimated toxic threshold in humans lies between 23 and
35 μg (48,55). Ghassemi and Shields reported that high-dose IvitC could
result in serious ocular adverse events, including cataract, vitreous and
subretinal hemorrhage, ocular hypotony, and phthisis bulbi eventually
leading to enucleation (40). Around the same time, Munier et al. found retinal
toxicity in the form of salt-and-pepper retinopathy near the injection site in
nearly half of eyes treated with therapeutic doses of IvitC (56). Toxicity was
further studied by Francis et al., where a 5.8 μV decrease in
electroretinogram recordings, corresponding to an estimated 5% decrease in
retinal function, was observed per each single injection of melphalan 30 μg
(55). Despite these toxic effects, Munier et al. reported a success rate of 83%
in tumor control and globe salvage. Shields et al. reported minor
complications in a review of 55 intravitreal melphalan injections for recurrent
vitreous seeding in 16 retinoblastoma patients that included focal retinal
pigment epithelial mottling near the injection site and posterior lens
opacification (41). Shields et al. later studied 192 consecutive injections and
found therapeutic success in all eyes (n = 40) with side effects of focal retinal
pigment epithelial mottling at the injection site (n = 12), focal paraxial lens
opacity (n = 11), focal vitreous hemorrhage (n = 5), hypotony (n = 3), retinal
hemorrhage (n = 2), optic disc edema (n = 1), and hemorrhagic retinal
necrosis (n = 1) (53). Other ocular IvitC-related adverse events include
anterior segment toxicities of angle recession and depigmentation, iris and
scleral thinning, and cataract, which appear to occur more commonly in the
same meridian as the injection, where the drug concentration is highest (57).
Prior to the implementation of antireflux safety measures, extraocular
extension was reported in 0.4% of cases, but recent studies report a
considerable decrease in risk, which ranges from 0% to 0.08% using current
injection techniques (41,49,58).
Precision Intravitreal Chemotherapy

The use of precision intravitreal chemotherapy (p-IvitC) was introduced by
our team to treat localized vitreous seeding (Figure 45-8). In contrast to the
standard technique aimed to treat diffuse vitreous seeds, p-IvitC involves
injection of the chemotherapeutic agent(s) in close proximity to a single or
localized group of vitreous seeds under indirect ophthalmoscopic guidance,
rather than directing the needle toward the center of the globe and dispersing
the drug(s) throughout the vitreous cavity (59). In this technique, the eye is
not jiggled in order to avoid dispersion of the injected drug(s) via advective
mass transport. Instead, the eye is kept still, and the head is positioned with
the vitreous seed(s) located inferiorly, using gravitational forces to help settle
the injected agent(s) toward the seed(s) and reducing drug exposure to the
macula and other structures critical for vision. This modality seems to
optimize drug functionality, translating into a reduction in range of 4 to 5
injections to a mean of 2.6 injections. Yu et al. reported eight eyes of eight
retinoblastoma patients treated with p-IvitC in which focal retinal pigment
epithelial mottling was observed in only one case (13%) and remained
outside the fovea (59). No recurrence was observed in any case after a mean
follow-up of 10 months.
FIGURE 45-8 Localized vitreous seeding from
retinoblastoma managed with precision intravitreal
chemotherapy (p-IvitC). A: Persistent vitreous seed
superotemporal to the fovea with (arrow) B: resolution
(arrow) after three cycles of p-IvitC. C: Persistent vitreous
seed near the foveola (arrow) with D: resolution after
three cycles of p-IvitC.
Intracameral Chemotherapy
The use of ICC was recently introduced by Munier et al. to provide sufficient
drug bioavailability for treatment of anterior chamber tumor retinoblastoma
seeding (60). Previously, aqueous seeding remained an indication for
enucleation or anterior chamber plaque radiotherapy due to pharmacokinetic
properties that impeded tumoricidal doses of chemotherapy from reaching the
anterior chamber via conventional routes of administration. In Munier’s
technique, patients were given oral acetazolamide 5 mg/kg preoperatively to
suppress aqueous humor secretion in order to avoid drug dilution (60).
Aqueous humor was then aspirated from the anterior and posterior chambers
through a transcorneal approach with a 34-gauge long needle, facilitated by
contralateral limbus indentation. Without removing the needle, a syringe
exchange was performed to replace a comparable volume of aqueous with
melphalan at a concentration of 15 to 20 μg/0.05 mL (Table 45-2), minus any
volume planned to be injected into the vitreous. The dose was
compartmentalized, distributing 1/3 of the dose to the anterior chamber, and
the remaining 2/3 to the posterior chamber via a transiridal approach.
Following injection, cryotherapy was applied to the entry site at the time of
needle removal. In a case report, the authors reported tumor regression
following seven injections of ICC with melphalan into the anterior chamber
(total dose 15.8 μg) with recurrence 3.5 months later, ultimately controlled by
an additional six injections of ICC with melphalan targeting the posterior
chamber (total dose 35 μg) (60). Intracameral topotecan (7.5 μg/0.015 mL)
injected into the anterior chamber for retinoblastoma seeds has also been
explored after failure of intravitreal melphalan (20 μg/0.05 mL) and yielded
tumor regression after a single injection without toxic effects at 9-month
follow-up (61). ICC has also been investigated in combination with plaque
brachytherapy with complete tumor control in one case after 3-year follow-up
(62).
Complications at 5-year follow-up of intracameral delivery of
chemotherapy include iris heterochromia and progressive cataract formation
in the treated eye with stable corneal endothelial cell density (60). Others
retrospectively analyzed the outcomes of patients treated with ICC combined
with other therapies and achieved globe preservation in 71% of patients with
intact anterior hyaloid membrane and 25% in patients with disrupted anterior
hyaloid membrane secondary to previous vitrectomy with silicone oil or
intracapsular lensectomy (62). The authors attributed outcomes to two
possible mechanisms: drug dilution and/or tumoral cross-contamination
between anterior and posterior segments (62). Salvaged eyes remained event-
free after 17-month follow-up.
FOCAL THERAPIES
Focal therapies are often used for tumor consolidation in conjunction with
IVC or IAC (63). IVC alone achieves suboptimal tumor control in many
cases, with up to 28% of cases requiring additional adjuvant therapy to
achieve tumor control (64). Therefore, targeted local treatments, including
cryotherapy, laser photocoagulation, and TTT, remain important in disease
management. The choice of adjuvant treatment is individualized to the tumor
size and location. Less commonly, focal therapies can be used alone as
primary therapy when patients present with isolated, small, unilateral tumors.
In our center, we have had success with tumor consolidation after IVC with
the single exception of foveal tumors in which we do not treat the tumor
directly. For eyes treated with IAC, we do not need to provide consolidation.
Cryotherapy
Cryotherapy is a reliable treatment that is still regularly used in management
of intraocular retinoblastoma. Indications include treatment of small tumors
and foci of subretinal seeds <3 mm in diameter (63). Cryotherapy to the
peripheral ora serrata is often performed on the same day as systemic
chemotherapy, immediately preceding chemotherapy delivery, to facilitate
improved drug penetration to the tumor(s).
Treatment is performed under indirect ophthalmoscopic visualization,
placing the cryotherapy probe on the conjunctiva for peripheral lesions or
directly on the sclera through a conjunctival cut down incision for more
posteriorly located lesions. A triple freeze–thaw technique is employed,
visualizing the tumor becoming entirely encased in an ice ball and then
waiting for a complete thaw prior to applying the second and third freeze–
thaw cycles. Typically, three sessions are required, administered on a
monthly basis, with the end point being a completely flat tumor scar. In a
study of 67 retinoblastomas treated with cryotherapy, tumor control was
achieved in 53 (79%) tumors, with complete control of all tumors ≤2.5 mm in
largest basal diameter and ≤1.0 mm in thickness confined to the neurosensory
retina without adjacent vitreous seeding (65). Presently, cryotherapy is rarely
used alone and is more frequently used in combination with chemotherapy.
Laser Photocoagulation
Laser photocoagulation is not often used for treatment of retinoblastoma
anymore, as it has been replaced by chemotherapy. For those eyes receiving
IVC, laser is certainly not used, as laser coagulates the vascular supply of the
tumor, preventing adequate chemotherapy delivery. In the past, laser was
performed with xenon arc, argon, or yttrium aluminum garnet lasers.
Indications included small tumors <3 mm in diameter and 2 mm in thickness
with no vitreous seeding and that were located posterior to the equator where
cryotherapy application would be technically difficult (66,67).
Laser was applied with approximately 200 μm spot size and sufficient
power to achieve a white burn. A double row of laser spots encircling the
tumor was applied to occlude tumor vasculature, with median of two laser
sessions required to achieve the desired flat chorioretinal scar. Direct tumor
treatment with laser must be avoided, as rupture of the overlying internal
limiting membrane could potentially cause iatrogenic vitreous seeding
(49,68). In one study of 30 tumors in 20 eyes, complete tumor regression was
seen in 21 eyes (70%) following treatment with laser photocoagulation (66).
Transpupillary Thermotherapy
TTT using diode laser has now largely supplanted laser photocoagulation for
treatment of retinoblastoma. As with cryotherapy, TTT can be used in
combination with chemotherapy or as primary treatment for small tumors <3
mm in diameter (63).
TTT is administered under indirect ophthalmoscopic visualization using
an 810-nm diode laser on continuous mode. Spot size when using indirect
ophthalmoscopy and a 20 diopter lens is 1.2 mm, with multiple spots often
required to cover the entire tumor. Laser power can be increased from 300 to
800 mW and is titrated to induce a gray-white discoloration of the treated
tumor after several seconds, with higher power required for thicker, more
calcified lesions. The entire tumor surface is treated, and time of application
depends on visualized response to the laser, with larger tumors requiring
longer application times to achieve a satisfactory gray-white color change.
Other signs, including microhemorrhages and retinal edema surrounding the
lesion, can signal when treatment is complete (63). Eyes with darker retinal
pigment epithelium pigmentation and darker choroid tend to respond to lower
power and shorter duration treatments. Multiple sessions of TTT, ranging
from 2 to 6, are usually required at 4- to 6-week intervals to achieve the end
point of a flat scar or completely calcified tumor (69). Persistence of
noncalcified fish flesh is a risk factor for tumor recurrence (70,71).
TTT can be effective as a primary therapy when used appropriately. In a
series of 91 small tumors in 22 eyes, 84 tumors (92%) were cured with TTT
alone (72).
Indocyanine green (ICG) can be used to enhance the effects of TTT in
cases of suboptimal response, tumor recurrence, or in a lightly pigmented
fundus. ICG is infused at a dose of 0.3 to 0.5 mg/kg approximately 1 minute
prior to TTT application. The technique for TTT application is otherwise as
above. Prior studies have demonstrated tumor control with this technique in
30 of 30 treated eyes (73), and ICG-enhanced TTT can be combined with
both systemic chemotherapy and IAC (Figure 45-9) (74).
FIGURE 45-9 Group B retinoblastoma managed with

systemic intravenous chemotherapy (IVC) and
indocyanine green–enhanced (ICG-enhanced)
transpupillary thermotherapy (TTT). (A) Tumor touching
the optic disc with (B) regression after IVC and
consolidation with ICG-enhanced TTT.
Complications
Anterior segment damage can occur following laser or TTT application,
including iris atrophy, anterior or posterior synechiae, and focal cataract,
especially if the pupil is not well dilated during the procedure (69,75). Sight-
threatening complications are rare with appropriate use and include retinal
vein occlusion, vitreous hemorrhage, retinal neovascularization, vitreoretinal
traction, and retinal detachment (71,76,77). Exudative or rhegmatogenous
retinal detachment can also occur following extensive cryotherapy (78,79).
All focal therapies result in chorioretinal scarring and can lead to
reduction in visual field or visual acuity if lesions are treated in the macula.
TTT in particular, which is more easily applied to posteriorly located lesions
than cryotherapy, can be performed in a fovea-sparing fashion, and caution
should be applied when choosing to use these therapies for macular tumors
(69,71). Consideration should be given to alternative chemotherapy-based
treatment regimens for tumors involving the fovea, especially if both eyes are
affected.
External Beam Radiotherapy

Prior to the introduction of systemic chemotherapy, EBRT was used as a
globe salvage therapy. The standard dose for EBRT was 40 to 45 Gy,
delivered in 20 fractions over 4 weeks (80). Techniques evolved throughout
the years to provide more targeted radiation to the tumor with lower doses to
the cutaneous tissue, lens, and brain (80). In the 1970s, tumor control and
globe salvage rates with EBRT were reported at 100% for Reese-Ellsworth
groups I and II, 86% for group III, and 75% for groups IV and V (81).
Abramson et al. later reported tumor control in 49% of 63 treated eyes, with
recurrence in 41% and new tumor in 13%; globe salvage at 1 year was 81%
and at 10 years was 53% (80,82). Today in the United States, EBRT is
largely of historical significance due to the many associated side effects and
improved outcomes following introduction of effective chemotherapy for
retinoblastoma. EBRT does, however, maintain a role in the setting of
extraocular tumor extension, orbital recurrence, and involvement of the
surgical margin of the optic nerve following enucleation (80). Reports of
EBRT plus systemic chemotherapy for treatment of orbital retinoblastoma
describe local tumor control in 71% of patients (83).
Radiation side effects associated with EBRT include tear deficiency and
dry eye with or without filamentary keratopathy, cataract, radiation
retinopathy, optic neuropathy, and orbital growth retardation causing facial
deformity (80,84–86). The most serious side effects of EBRT, however, are
subsequent development of second primary tumors in the field of radiation
with especially elevated risk in patients with germline retinoblastoma. The
most common second primary tumor is osteosarcoma, but other malignancies
of the bone, soft tissue, and skin include fibrosarcoma, rhabdomyosarcoma,
chondrosarcoma, Ewing sarcoma, basal cell carcinoma, squamous cell
carcinoma, cutaneous melanoma, transitional cell carcinoma, breast
carcinoma, lung carcinoma, leukemia, lymphoma, and pineoblastoma (80).
The risk of second primary malignancies in patients with germline
retinoblastoma treated with EBRT is reported to be as high as 53% by age 50,
making patients more likely to die from a second cancer than from the
retinoblastoma itself (87–89). Other studies demonstrated that the risk could
be most elevated for patients treated with EBRT prior to age 12 months (87).
In a review of 813 retinoblastoma survivors treated with radiotherapy, 265
(33%) patients developed at least one second primary tumor, with 46 (6%)
patients developing more than one second cancer (90). The most common
second primary cancers included bone tumors, soft tissue sarcoma,
melanoma, and epithelial tumors (bladder, breast, colorectal, kidney, lung,
nasal cavity, prostate, retroperitoneum, thyroid, tongue, uterus). Due to these
side effects, we recommend avoiding treatment with EBRT if other effective
treatment methods are available.
PLAQUE RADIOTHERAPY
Plaque radiotherapy for retinoblastoma was first described in 1929 and was
initially used as globe salvage therapy for recurrent tumor following EBRT
(91–93). Other indications for plaque radiotherapy prior to the introduction of
IAC and IvitC included primary therapy for small- to medium-sized
retinoblastoma and secondary therapy for localized tumor recurrence
following failure of cryotherapy, laser photocoagulation, or TTT (94–97). In
the present era, plaque radiotherapy is typically used as secondary treatment
for small- to medium-sized (≤16 mm in largest basal diameter and >3 to ≤9
mm in thickness), chemoresistant tumors with or without localized vitreous
or subretinal seeding following recurrence after systemic chemotherapy or
IAC (98). Plaque radiotherapy can also be used to manage diffuse anterior
segment retinoblastoma with or without systemic chemotherapy in the
absence of choroidal or retinal tumors (Figure 45-10) (99).
FIGURE 45-10 Anterior chamber retinoblastoma treated

with plaque radiotherapy. (A) Retinoblastoma in anterior
chamber treated with (B) radioactive plaque for the entire
anterior segment applied after (C) 360-degree conjunctival
peritomy with (D) nylon sutures covered with (E) a
Gunderson flap. (F) Complete regression after treatment.
Accurate tumor sizing and localization is important for planning purposes

prior to undertaking treatment with plaque radiotherapy. Tumor largest basal
diameter, thickness, and distances to the foveola and optic nerve must be
measured using indirect ophthalmoscopy and ultrasonography, including
areas of localized vitreous or subretinal seeding. A 2-mm safety margin is
typically added to the largest basal diameter. Tumors within 2 mm of the
optic nerve require a notched plaque with a deep notch for 3 or more clock
hours of tumor around the nerve. Iodine-125 is currently the most commonly
used isotope for plaque radiotherapy for retinoblastoma in the Unites States
(96). The tumor apex should be treated with a dose of 35 to 40 Gy to provide
adequate tumor control while minimizing radiation side effects (100). If
possible, secondary plaque radiotherapy is delivered 1 to 2 months after
systemic chemotherapy is completed in order to minimize side effects (100).
However, tumor measurements should be performed no longer than 3 weeks
prior to treatment to avoid treatment failure from uncalculated tumor growth.
Plaque application is performed under general anesthesia, beginning with
a conjunctival peritomy and tenotomy in the quadrant(s) where the plaque
will be placed. The rectus muscles are isolated with silk sutures to rotate the
globe for exposure during tumor localization and plaque placement. Tumor
localization is performed using transillumination and scleral indentation with
indirect ophthalmoscopy. Even in the presence of a good shadow on
transillumination, ophthalmoscopy should be used to confirm tumor
localization, as thin tumors, irregular margins, and tumor seeding do not cast
a reliable shadow.
Occasionally, a rectus or oblique muscle needs to be temporarily
disinserted for accurate plaque placement. The plaque is left in place for a
calculated number of hours based on tumor thickness, tumor type, and
activity of the radioactive seeds.
Compared to EBRT, plaque radiotherapy is convenient, given that it only
takes 2 to 4 days to deliver the complete dose, with the minor disadvantage of
requiring two surgeries to place and remove the plaque (96,100). Plaque
radiotherapy avoids many of the serious side effects associated with EBRT,
particularly ipsilateral orbit and facial hypoplasia, and most seriously, second
cancers (80,94). Our initial experience using this technique for the treatment
of retinoblastoma dates back to 1976 when cobalt-60 plaques were used for
smaller tumors (95). After observing continued success using this technique,
episcleral plaque radiotherapy was further employed both as a primary and a
secondary treatment, such as following failure of EBRT, photocoagulation,
and/or cryotherapy, in 51 eyes of 50 patients with unilateral sporadic
retinoblastoma and resulted in good functional outcomes in both groups with
better tumor control in the primary treatment group (87%) compared to the
secondary group (61%) (95). In a subsequent analysis of a larger cohort of
103 patients, similar treatment success was observed between eyes treated
with primary (84%) versus secondary (88%) plaque radiotherapy (94).
Success of plaque brachytherapy as secondary treatment for retinoblastoma
following failure of other treatments has been reported to be 92% after
cryotherapy, 92% after laser photocoagulation, 92% after IVC, 75% after
EBRT, and 66% after a combination of IVC and EBRT (96,98). In an
analysis of 208 tumors in 147 eyes of 141 patients with longer follow-up,
tumor control was maintained in 79% of cases at 5 years (96). More recently,
success of plaque radiotherapy as secondary treatment following IAC was
reported to be 79% even in the presence of localized vitreous seeding (101).
Tumor recurrence following plaque radiotherapy usually occurs within 1 year
of treatment and tends to be lower with primary compared to secondary
treatment (12% vs. 8%) (102). Predictors of recurrence have been identified,
including presence of subretinal or vitreous seeds and older age at treatment
(96).
Despite excellent tumor control following plaque brachytherapy with
good visual outcomes in over 62% of cases, side effects can occur and
include cataract (20% to 43%), radiation maculopathy (25%), radiation
papillopathy (26%), and vitreous hemorrhage (54%) (Figure 45-11)
(94,96,98,100,101). A comparison between primary plaque radiotherapy and
secondary plaque radiotherapy following IVC showed a higher incidence of
radiation retinopathy (27% vs. 40%) and cataract formation (33% vs. 43%)
with the latter (96). Treatment of radiation complications has been reported
(103–105). Intravitreal antivascular endothelial growth factor (anti-VEGF)
medications can be used to treat macular edema following plaque treatment.
However, prior to confirmed tumor regression, intravitreal injections of
nonchemotherapeutic agents should be avoided to prevent extraocular tumor
extension. A more conservative approach to prevent or treat macular edema
in such cases is the use of sub-Tenon triamcinolone. Sector panretinal laser
photocoagulation can be used in combination with prophylactic sub-Tenon
triamcinolone to prevent macular edema or proliferative radiation retinopathy
following plaque radiotherapy (106).
FIGURE 45-11 Retinoblastoma managed with plaque

radiotherapy with vitreous hemorrhage. (A) Recurrent
tumor with (B) regression and vitreous hemorrhage after
plaque radiotherapy.
ENUCLEATION
Enucleation was previously the standard of care for advanced unilateral or
bilateral retinoblastoma. However, the introduction of systemic
chemotherapy and IAC have allowed for globe salvage in many cases that
would have been previously treated with primary enucleation. Nevertheless,
enucleation remains an important treatment modality for advanced unilateral
group E disease (Figure 45-12), anterior chamber involvement, neovascular
glaucoma, pars plana tumor seeding, long-standing retinal detachment, poor
tumor visualization (e.g., due to vitreous hemorrhage), optic nerve invasion,
orbital invasion, after failure of other treatment modalities, and if there is no
expectation of useful vision (107–111).
FIGURE 45-12 Group E retinoblastoma managed with
enucleation. A:Retinoblastoma with total retinal
detachment. B:Left prosthesis with good cosmesis 1 year
after enucleation.
At the time of surgery, the correct eye should be confirmed by marking the
site in front of the parent(s) or guardian(s). Under general anesthesia in the
operative suite, the correct eye should again be confirmed using indirect
ophthalmoscopy, review of consent forms, and agreement during surgical
time out by the entire surgical team. In cases of a small palpebral fissure,
particularly in the Asian population, a lateral canthotomy can be performed to
facilitate globe removal. Following conjunctival peritomy, each of the four
rectus muscles is isolated and secured with double-armed 5-0 Vicryl sutures
before disinsertion from the globe, and the medial rectus stump is left long
for globe manipulation. The two oblique muscles are then disinserted. Using
long, straight enucleation scissors placed in the orbit nasally and pointing to
the orbital apex, the optic nerve is cut, attempting to obtain a long segment.
The globe is removed and an implant is placed. The conjunctival tissue is
closed. Fresh tissue is harvested from the globe for genetic assessment on a
separate surgical field, and the surgeon re-gloves prior to returning to the
patient to avoid contamination of the orbit with malignant cells.
Complications following enucleation include chemosis, conjunctival cyst,
pyogenic granuloma, blepharoptosis, lagophthalmos, superior sulcus defect,
enophthalmos, symblepharon, implant exposure, and infection (112). Implant
exposure requires return to the operating room for wound repair (112).
Infection can be managed with topical and/or systemic antibiotics but can
require implant removal in severe cases. Giant papillary conjunctivitis can
occur secondary to prosthesis wear and can be managed with a combination
of antibiotic–steroid ointment and refitting of the prosthesis in severe or
recurrent cases. Following enucleation, the globe should be evaluated
histopathologically to confirm the diagnosis of retinoblastoma and to assess
for invasive (high-risk) features.
High-Risk Retinoblastoma
On histopathologic evaluation, the enucleated globe should be evaluated for
high-risk features, including postlaminar optic nerve invasion, massive
choroidal invasion (>3 mm diameter), and any degree of optic nerve and
choroidal invasion (113,114). In an analysis of 519 enucleated eyes with
retinoblastoma, high-risk features were found in 17% of group D and 24% of
group E eyes (115). There were no enucleated eyes in that study classified as
groups A, B, or C.
Patients with high-risk features require adjuvant systemic chemotherapy
to prevent systemic metastasis. A VEC regimen, with doses comparable to
those used in IVC, can be used in these cases. Kaliki et al., in a study of 55
patients with high-risk retinoblastoma treated with six cycles of adjuvant
VEC, found no case of metastatic disease over mean follow-up of 66 months
(116).
ORBITAL DISEASE
Orbital involvement of retinoblastoma can be due to delayed detection of
intraocular retinoblastoma or secondarily due to recurrence following
enucleation (117). Orbital disease must be managed with combination
therapy, including surgical excision, systemic chemotherapy, and
radiotherapy to effectively eradicate orbital disease and prevent recurrence or
metastasis (117). The initial therapy consists of high-dose systemic
chemotherapy with a VEC regimen for three to six cycles with serial imaging
to monitor for complete resolution of the orbital component. Following
complete resolution of the orbital component or after completion of three to
six cycles of VEC, enucleation surgery is performed and followed by
radiotherapy in fractionated doses, totaling 50 Gy. The systemic
chemotherapy is continued to complete a total of 12 cycles.
Orbital retinoblastoma imparts poor prognosis, with a 10 to 27 times
higher risk of systemic metastasis and mortality rates ranging from 25% to
100% (118–122). However, combination therapy has improved treatment
success. In a series of 16 patients presenting with orbital retinoblastoma and
no intracranial extension or systemic metastasis, combination therapy
achieved complete tumor resolution without recurrence or metastatic disease
after 36-month follow-up in 14 patients (87.5%) (123,124). Efforts toward
earlier detection, especially in less developed countries, are critical to
improving vision and life prognosis for retinoblastoma patients.
METASTATIC DISEASE
Metastatic disease is rare in developed countries, affecting <5% of patients
(125). However, metastases remain a significant cause of mortality in
retinoblastoma patients in the developing world likely due to delayed disease
detection (126,127). Limited evidence is available for the treatment of
metastatic retinoblastoma. Given that most tumors are sensitive to
chemotherapy, successful treatment involves intensive high-dose
chemotherapy with autologous stem cell rescue. Radiotherapy can be added
for residual bulky disease.
Reports have been limited to small numbers of patients, but eradication of
cancer can be achieved in 60% to 70% with better prognosis in patients
without central nervous system involvement (128–133). Unfortunately, such
intensive treatment regimens are not often available in developing countries
where metastatic disease is most prevalent. Education of patients and
physicians with more timely disease detection remain important goals in the
developing world.
GENETIC TESTING
We recommend genetic testing in all cases of retinoblastoma both for the
patient and subsequently for the parents and siblings if germline disease is
confirmed. Patients with germline mutation should be managed with IVC,
often in combination with other treatment modalities, to prevent
pineoblastoma, and such patients require careful monitoring with MRI of the
brain and orbits twice per year until the child is 5 years old (4–6). Patients
and parents should be counseled about the risk for retinoblastoma in future
offspring, and in vitro fertilization with preimplantation diagnosis can be
discussed as part of family planning. In the case of pregnancy with a possible
germline retinoblastoma mutation, amniocentesis can be used to aid in
diagnosis with in utero ultrasonographic monitoring throughout the
pregnancy (134).
CONCLUSION
Management of retinoblastoma is complex. Each case is unique, and
treatment regimens must be carefully customized for varying disease
presentations and family goals of care. Close cooperation between the
treating ocular oncologist and pediatric oncologist is critical to patient
success, with both parties prioritizing patient safety and preservation of life.
ACKNOWLEDGMENTS
Support provided in part by an unrestricted grant from Research to Prevent
Blindness, Inc., New York, New York (LAD); the Heed Ophthalmic
Foundation, San Francisco, CA (LAD); and the Eye Tumor Research
Foundation, Philadelphia, PA (CLS). The funders had no role in the design
and conduct of the study; in the collection, analysis and interpretation of the
data; and in the preparation, review, or approval of the manuscript. Carol L.
Shields, MD has had full access to all the data in the study and takes
responsibility for the integrity of the data and the accuracy of the data
analysis.
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46
Tumors in Infants and Children
Victor M. Villegas, and Timothy G. Murray
CIRCUMSCRIBED CHOROIDAL
HEMANGIOMA
Prevalence
Circumscribed choroidal hemangioma is the most common benign vascular
tumor of the choroid. These tumors may be classified into two types, diffuse
and circumscribed, based on the degree of choroidal involvement (1–4). The
diffuse process occurs in Sturge-Weber syndrome (SWS) (discussed later in
this chapter), whereas the circumscribed form is usually not associated with
cutaneous lesions or other systemic findings (2). In 1976, Witschel and Font
(5) reported the clinicopathologic characteristics of 71 choroidal
hemangiomas. In their study, 37% represented the diffuse type and 63% were
circumscribed.
The exact incidence of circumscribed choroidal hemangioma is unknown.
However, choroidal hemangioma is diagnosed in approximately 1 case per 40
cases of posterior uveal melanoma (3). Because these lesions are often
asymptomatic and may remain undiagnosed, their prevalence may be much
higher than expected.

No known environmental factors contribute to the development of choroidal
hemangioma. No genetic associations have been identified to date.
Worldwide Impact
Because these tumors are mostly unilateral and may not affect visual acuity,
they are likely to have a limited role on worldwide childhood blindness.
Pathology and Pathophysiology

Choroidal hemangioma contains microscopically congested vessels. The
histologic appearance of these tumors demonstrates complete absence of
cellular proliferation in the blood vessel walls, suggesting that these lesions
are nonproliferative in origin (5). Histopathologically, choroidal hemangioma
has been described as a circumscribed tumor with sharply demarcated
margins, leading to compression of surrounding melanocytes and choroidal
lamellae (5).
Circumscribed choroidal hemangioma is believed to be a congenital
hamartomatous tumor. The exact cause and pathophysiology are unknown,
although some have suggested that the underlying process involves
arteriovenous (AV) shunts that are present in embryogenesis and
subsequently disappear during normal development (6).

Patients with circumscribed choroidal hemangioma often present with ocular
symptoms during adulthood, in contrast to patients who have the diffuse type.
Both types of choroidal hemangioma are likely to be present at birth, with the
circumscribed variant frequently remaining undetected until adulthood. The
circumscribed form should be included in the differential diagnosis of a
pediatric intraocular tumor. These tumors typically present with blurry vision,
metamorphopsia due to serous subretinal and intraretinal fluid, and/or
hyperopic shift associated with subretinal mass or fluid effect in subfoveal
tumors (7,8).
Choroidal hemangiomas are typically nonpigmented elevated masses
with a characteristic orange-red color (Figure 46-1). Small tumors may blend
with the surrounding choroidal pattern and may be difficult to identify during
routine ophthalmoscopic examination. Irregular pigmentation may be present
at the tumor surface or on its edges, especially in cases with chronic
subretinal or intraretinal fluid. These tumors are almost always unilateral and
solitary.
FIGURE 46-1 The orange-red elevated mass with apical
fibrosis demonstrates the characteristic appearance of
circumscribed choroidal hemangiomas.
Most circumscribed choroidal hemangiomas are located in the postequatorial

fundus (5,9). The posterior margin of the majority of these circumscribed
lesions is located within two disc diameters of the optic disc or the fovea (9).
Most of these lesions are less than 19 mm in diameter and have a mean
elevation of 3 mm (9). Most choroidal hemangiomas do not grow
significantly over time. However, tumor growth has been reported (10).
Subretinal fluid is commonly associated with this tumor and is
responsible for the most common presenting symptoms (7). Circumscribed
choroidal hemangiomas have also been associated with choroidal
neovascularization (CNV) (11). Spontaneous reabsorption into a flat
chorioretinal scar has been reported.
Visual acuity in patients with circumscribed choroidal hemangioma can
be completely normal or may be decreased from a number of mechanisms.
Vision loss and visual field defects associated with circumscribed choroidal
hemangiomas may result from exudative retinal detachment, hyperopic shift,
macular edema, retinal degeneration, photoreceptor loss, chorioretinal
adhesions, cystoid degeneration, and amblyopia.
Diagnostic Studies
The diagnosis of choroidal hemangioma is usually made based on
ophthalmoscopic findings; however, several ancillary studies can be used to
confirm the diagnosis. Ultrasonography classically shows high internal
reflectivity on standardized A-scan and a homogenous hyperechoic mass with
vascularity on B-scan images. Fluorescein angiography (FA) demonstrates
early filling within large vascular channels constituting the tumor. Late
frames show diffuse hyperfluorescence due to leakage of dye from the tumor
surface (12). Indocyanine green angiography (ICG) shows accumulation of
dye early with subsequent washout later in the study (13). ICG pattern may
be particularly helpful in challenging cases, such as choroidal hemangiomas
with significant retinal pigment epithelium (RPE) changes or preretinal
fibrosis. Optical coherence tomography (OCT) shows a smooth, gently
sloping choroidal mass without compression of the choriocapillaris (14).
Subretinal fluid, intraretinal fluid, or retinoschisis may be present on OCT.
Diffuse vascular anomalies can be seen in optical coherence tomography
angiography (OCTA) (15).
Despite the characteristic clinical features of choroidal hemangioma, this
tumor can be challenging to distinguish from other choroidal tumors
including amelanotic melanoma, choroidal osteoma, choroidal metastasis, or
retinoblastoma. In addition, these lesions can mimic posterior scleritis and
other inflammatory conditions.
Management
Treatment of circumscribed choroidal hemangiomas is generally limited to
those that are vision threatening, and treatment planning depends upon tumor
size and location (13). Hemangiomas that do not affect vision and are not
associated with subretinal fluid are generally observed. Most lesions are
characteristically nonprogressive in nature.
Laser photocoagulation can be an important treatment and is indicated
when the tumor causes loss of vision secondary to serous retinal detachment
(9,16). Historically, focal lesions were treated with laser to cause resorption
of subretinal fluid and readhesion of the neurosensory retina and RPE.
Subretinal fluid has been reported to resolve in 62% to 100% of cases
following laser therapy (7).
Radiation therapy has also been used frequently in the treatment of
choroidal hemangioma, including external beam radiation, plaque
radiotherapy, or proton beam irradiation. Numerous reports have indicated
that these treatments offer excellent anatomical outcomes (17–19). Previous
studies (7) have recommended early low-dose plaque radiotherapy or external
beam radiation in cases with recurrent subretinal fluid after laser therapy and
subfoveal lesions. Metamorphopsia may improve significantly following
brachytherapy.
Photodynamic therapy (PDT) has become a commonly used therapy for
subfoveal circumscribed choroidal hemangiomas. Multiple studies have
reported on the efficacy of PDT for subretinal fluid in subfoveal lesions
(16,20–22). Most patients show positive anatomical and visual outcomes
after only one cycle of PDT (16,21). However, choroidal ischemia may limit
visual acuity following PDT therapy.
Despite the benign nature of these tumors, approximately 50% of diagnosed
patients have long-term visual acuity of 20/200 or worse (7). Final visual
acuity is dependent upon numerous factors including initial visual acuity,
failure of previous laser treatment, multiple quadrants of subretinal fluid,
chronic submacular fluid, chronic cystoid macular edema, or chronic retinal
pigment epithelial changes (7).
Children with circumscribed choroidal hemangiomas may also develop
amblyopia in the affected eye. Amblyopia may be secondary to hyperopic
shift, astigmatism, or subretinal fluid. Because early intervention can be
effective in achieving resolution of subretinal fluid and restoration of visual
function, prompt referral to an ophthalmologist with experience treating
circumscribed choroidal hemangioma is especially important in children
suspected of having this lesion.

In select cases where radiation therapy is indicated, a consultation with a
radiation oncologist is warranted. They can review the risks and benefits of
radiation exposure associated with a plaque versus external techniques in a
pediatric patient.
Not applicable.
Future Treatments
Intravitreal bevacizumab has been used in the primary treatment of
circumscribed choroidal hemangioma with positive outcomes (23). Most
recently, Arevalo and associates have reported faster resolution of subretinal
fluid when PDT with verteporfin is used in combination with intravitreal
bevacizumab (24). Other authors have had similar results with combination
PDT and intravitreal bevacizumab. Further studies are needed to evaluate the
role of intravitreal bevacizumab in the treatment of circumscribed choroidal
hemangioma.
MEDULLOEPITHELIOMA
Prevalence
Medulloepithelioma is a rare embryonal tumor that arises from the medullary
epithelium on the inner layer of the developing optic cup (25). The exact
prevalence is still unclear. Verhoeff (26) first described this tumor
histopathologically as a teratoneuroma in 1904. Medulloepithelioma is
classified into teratoid and nonteratoid forms. The nonteratoid form,
otherwise known as a dictyoma, represents only medullary epithelial cells. In
contrast, teratoid forms of this tumor contain heterotopic tissues including
cartilage, skeletal muscle, and brain-like tissue (27). Either form can be
benign or malignant (25,28,29).
Intraocular medulloepithelioma often has a malignant histologic
appearance. Extraocular extension with distant metastasis to the lungs,
mediastinum, and lymph nodes has been documented (28,30). Broughton and
Zimmerman (28) reported 56 cases in which 66% were malignant and 30%
were teratoid. Other authors have reported up to 90% malignancy (31).
Despite a malignant histologic appearance, distant metastases are rarely seen
in the absence of local extraocular extension (28,29,31).

No reported environmental or genetic factors have been associated with
medulloepithelioma.
Worldwide Impact
The worldwide epidemiology of this malignancy is unclear due to the rarity
of this malignancy. As with other intraocular tumors, delay in diagnosis,
particularly in less developed countries, can result in metastasis and death.
Pathophysiology
Differentiation of the medullary epithelium that lines the optic cup normally
produces the retinal photoreceptors, glial and neuronal tissues, nonpigmented
ciliary epithelium, pigmented epithelium of the iris, muscles of the iris, and
components of the vitreous.
Histopathologic studies demonstrate that medulloepithelioma contains
elements that resemble the medullary epithelium, optic cup and/or vessel,
pigmented and nonpigmented ciliary epithelium, vitreous, or neuroglia (28).
Medulloepitheliomas often contain heterotopic tissues such as cartilage,
rhabdomyoblasts, skeletal muscle, and brain-like tissues (27). This feature
results from the capacity of the primitive medullary epithelium to
differentiate into a variety of neural and mesenchymal tissue types.
Medulloepithelioma characteristically presents before 10 years of age as a
white fimbriated ciliary body mass; however, it may also arise in adults (29).
The most common location at presentation is the ciliary body. However,
primary optic nerve, retinal, and orbital involvement has been reported
(32–35).
Medulloepithelioma frequently presents with decreased vision and pain.
Other common signs include leukocoria and a nonpigmented iris or ciliary
body (Figure 46-2) tumoral mass (28). One of the earliest signs may be a
lenticular notch in the quadrant of the tumor (36). This coloboma occurs
because the embryonal tumor prevents normal zonular development in the
associated lens quadrant.
FIGURE 46-2 Medulloepithelioma typically presents as a

white fimbriated mass in the anterior segment.
Medulloepithelioma has the capacity to progress and become locally invasive

and destructive to surrounding intraocular tissues. Loss of vision occurs
secondary to cataract, subluxed lens, lenticular astigmatism, neoplastic
cyclitic membrane formation, glaucoma, and/or amblyopia (31). Other
findings associated with this tumor include uveitis and retinal detachment
(28). Iris neovascularization has also been reported in association with
primary and recurrent medulloepithelioma (31,37). Neovascular glaucoma
discovered in a child with a normal fundus examination should raise
suspicion for occult medulloepithelioma.
The presence of cysts within a tumor is highly suggestive of
medulloepithelioma (31). Foci of cartilage, appearing as whitish opacities
within the tumor, are also a characteristic finding in the teratoid form of
medulloepithelioma (31).
Diagnostic Studies
The diagnosis of medulloepithelioma is primarily a clinical one, and the role
of additional studies to confirm the diagnosis is not well established. A-scan
ultrasonography has been reported to show irregular high internal reflectivity
with areas of moderate reflectivity, and B-scan can demonstrate characteristic
cysts within the tumor (38). Dense hyperechoic areas in medulloepithelioma
may be similar in appearance to calcifications in retinoblastoma (39).
Intrinsic tumor vascularity has been demonstrated with FA with associated
leakage (37,39).
Magnetic resonance imaging (MRI) of medulloepithelioma is
hyperintense on T1-weighted images and hypointense on T2-weighted
images similarly to malignant melanoma (40). Ultrasound biomicroscopy has
recently emerged as a helpful diagnostic tool for evaluating tumors involving
the anterior uveal tract; this approach may be particularly useful in
demonstrating the multicystic appearance of medulloepithelioma (39).
Anterior segment OCT may also help evaluate growth over time.
Given the wide variety of presentations and possible locations for
medulloepithelioma, the differential diagnosis is broad. Diagnoses that
should be considered include congenital and inflammatory conditions, such
as persistent hyperplastic primary vitreous (persistent fetal vasculature
syndrome), pars planitis, or vascular malformations, and numerous tumors,
including retinoblastoma, melanoma, melanocytoma, iridociliary cyst,
rhabdomyosarcoma, neuroblastoma, or teratoma.
Management
Definitive treatment of medulloepithelioma continues to be enucleation.
Medulloepithelioma frequently recurs following local resection (29,37).
Some cases may be observed until definitive change is documented. Cataract
surgery should be avoided if medulloepithelioma is suspected. Patients with
extrascleral extension may require exenteration.
The roles of cryotherapy, radiation, and chemotherapy are not well
established for the treatment of medulloepithelioma. In cases involving risk
for recurrence of the tumor, these methods may be used as adjunctive
therapy.
The visual outcome in pediatric patients diagnosed with medulloepithelioma
is poor mostly due to amblyopia, cataract, and glaucoma. Despite the
malignant nature of medulloepithelioma, patients with complete tumor
resection have excellent survival rates. In the largest series of 56 patients with
medulloepithelioma, four known tumor-associated deaths were reported. The
most important prognostic feature was extraocular extension (29).
As with other intraocular tumors, early diagnosis is important in
determining the final clinical outcome. Unfortunately, diagnosis is often
delayed in patients with medulloepithelioma; most of these patients
experience a delay in surgical treatment for more than 1 year after the initial
onset of symptoms (29).

The diagnosis and management of these rare malignancies require the
evaluation by ocular pathology, radiation oncology, and/or a pediatric
oncology. These patients are best treated in a tertiary center with
multidisciplinary expertise in pediatric solid tumors.
Developing countries often lack the multidisciplinary expertise to manage
these rare tumors. Delay in diagnosis and limited access to specialty health
care can lead to orbital invasion and increased morbidity and mortality. If
progressive medulloepithelioma is suspected, enucleation is the gold
standard.
Future Treatments
Recently, vincristine (1.5 mg/m2), carboplatin (560 mg/m2), and etoposide
(150 mg/m2) have been used with tumor burden control in an adult patient
(41). Cyclophosphamide has also been used (42) in combination with
vincristine, carboplatin, and etoposide with positive outcomes.
Radiation has also been used in primary treatment of ocular
medulloepithelioma (29,42,43). Most reports have prescribed a dose between
40 and 50 Gy with encouraging outcomes. Brachytherapy is preferred over
external beam radiation to minimize the morbidity associated with therapy.
CHOROIDAL OSTEOMA
Prevalence
Choroidal osteoma is a benign ossifying choroidal tumor that was first
described by Gass et al. (44) in 1978. Choroidal osteoma may present at any
age, is more common in females, and typically is located adjacent to the optic
nerve or involving the macula.
The prevalence of choroidal osteoma is unknown, and it has been
suggested that it may be more common than initially suspected (45).
However, since many lesions are asymptomatic, most choroidal osteomas
may be undiagnosed.

Choroidal osteoma has no known association with environmental factors. No
genetic associations have been identified.
Worldwide Impact
Worldwide impact is unknown as many of these cases are believed to be
asymptomatic. Cases with asymmetric visual acuity may not come to medical
attention in many developing parts of the world.
Pathophysiology
Choroidal osteoma is composed of bony trabeculae with osteoblasts,
osteocytes, and osteoclasts (44,46). Large, endothelial-lined, cavernous
spaces are covered with small capillary blood vessels. The choriocapillaris
can be partially or completely dysplastic in the involved areas. Overlying
RPE can undergo degeneration.
The pathogenesis of choroidal osteoma is not known. Previous reports
have suggested that focal choroidal inflammation may lead to calcification of
the choroid (47–49). It may also represent an osseous choristoma: normal
bone tissue in an ectopic location (48).
Intraocular calcification can be associated with trauma, long-standing
retinal detachment, inflammation, abnormalities in calcium and phosphorus
levels, and phthisis bulbi. Serum calcium, phosphorus, and alkaline
phosphatase levels are, however, found to be within the normal range in
patients with choroidal osteoma.
Females are more frequently affected than males, but endocrine
abnormalities have not been documented in these patients. A hereditary
component in the pathogenesis has been suggested by some familial cases
(50). Noble (50) hypothesized that a congenital, possibly inherited, defect in
the choroid may remain undetectable until osseous calcification is initiated
through the influence of additional factors.

Signs and symptoms of choroidal osteoma vary and depend mostly on the
location of the tumor. Although choroidal osteomas may be diagnosed at any
age, a congenital etiology has been suggested. Choroidal osteoma may
frequently remain asymptomatic and undiagnosed, especially in the
community setting (44). Foveal involvement by the tumor typically results in
decreased visual acuity. Subretinal neovascularization, exudative retinal
detachment, visual field defects, and metamorphopsia may be present.
Choroidal osteoma is characteristically an elevated, yellow-orange mass
(Figure 46-3). The tumor surface may appear mottled or spotted due to
clumping of orange or brown pigment on their surface (51). Most choroidal
osteomas are solitary lesions, although multifocal bilateral lesions may also
be present. Small distinctive tufts of vessels are often found on the tumor
surface, are easily visualized on FA, and demonstrate no leakage (52).
FIGURE 46-3 Choroidal osteoma is characteristically a
well-demarcated yellowish lesion at the posterior pole.
Choroidal osteoma commonly presents in the posterior pole. Approximately

75% of cases are unilateral, with tumor dimensions of up to 22 mm in
diameter and up to 2.5 mm in height (46). Subretinal fluid can be present
overlying a choroidal osteoma and should raise the suspicion of CNV with
subsequent accumulation of subretinal fluid and/or development of
hemorrhage (44,52). Subretinal fluid may also be present due to RPE
dysfunction (52).
Slow growth of choroidal osteoma occurs in approximately 50% of
patients. Growth is demonstrated either in an overall increase in size or in the
formation of pseudopod projections extending out from the tumor’s central
mass (53). Rapid growth in choroidal osteoma has been reported (54).
Spontaneous involution of the tumor has also been documented (55).
Diagnostic Studies
Choroidal osteoma can be highly suspected clinically. However, confirmation
of calcium with other imaging modalities is paramount for diagnosis.
Ultrasonography is particularly useful in differentiating choroidal
osteoma from other lesions. A-scan ultrasonography demonstrates a high-
intensity echo spike from the inner surface of the tumor, with decreased
amplitudes of orbital soft tissue echoes posterior to the tumor. B-scan shows
a homogenous hyperechoic choroidal mass with acoustic shadowing posterior
to the tumor (46,52).
A retrospective case series that evaluated fundus autofluorescence in
patients with choroidal osteoma found hyperautofluorescence in areas of
decalcification and granular hypoautofluorescence in areas of RPE mottling
(56). FA demonstrates early diffuse, irregular hyperfluorescent vascular tufts
with associated late staining. In the late FA frames, the spider-like vascular
tufts may stand out in negative relief as hypofluorescent lines against a bright
background (46,52). Choroidal neovascular membranes exhibit an early lacy
pattern of hyperfluorescence with early leakage. Recently, OCT and OCTA
have been used to detect CNV in a patient with choroidal osteoma (57).
On computed tomography (CT) scan, choroidal osteoma demonstrates the
same density as bone. On MRI, these tumors are hyperintense compared to
the vitreous on T1-weighted imaging and hypointense on T2-weighted
imaging (58).
The differential diagnosis of choroidal osteoma includes other intraocular
tumors, such as sclerochoroidal calcification, amelanotic choroidal nevi,
choroidal melanoma, choroidal metastasis, and circumscribed choroidal
hemangioma. Choroidal osteoma may also appear similar to osseous
metaplasia that can occur in other choroidal tumors. Choroidal osteoma
should be carefully distinguished from posterior scleritis or subretinal
hemorrhage. Idiopathic sclerochoroidal calcifications should be included in
the differential diagnosis for choroidal osteoma; however, the former are
typically multiple, often bilateral, and classically found along the
superotemporal vascular arcades (51).
Linear nevus sebaceous syndrome (59) presents with osseous choroidal
choristoma, which should be carefully distinguished from choroidal
hemangioma. This syndrome, however, also presents with characteristic
systemic findings: midline facial linear nevus of Jadassohn, seizures,
cognitive developmental deficits, and multiple eye findings including
lipodermoids; colobomas of the lids, iris, and choroid; and choroidal
calcification (59).
Management
Most cases of choroidal osteoma are asymptomatic and may be observed.
Periodic dilated fundus examinations and self-monitoring with the use of an
Amsler grid can facilitate timely diagnosis in patients who develop choroidal
neovascular membranes or subretinal fluid.
Argon laser photoablation has been associated with regression of
choroidal osteoma (60) and been efficacious for treatment of a subfoveal
neovascularization threatening the fovea (45,61). Laser treatment may cause
retinal anastomosis with both arteriolar and venular vessels within the tumor
in select cases (45). Successful treatment of focal RPE leakage with laser has
been reported (45). The use of PDT for subretinal neovascular membranes in
choroidal osteoma has also been described (62).
Because the etiology of choroidal osteoma is unknown at this time, no
preventive measures have been suggested.
Visual loss to 20/200 or worse has been reported to occur in approximately
60% of patients with choroidal osteoma after 20 years of follow-up (52). The
most common cause of severe vision loss is CNV. The long-term CNV
incidence is over 50%. Previous studies have reported a low success rate
(25%) after laser ablation of neovascular membranes (52). Hemorrhage
associated with choroidal osteoma typically resolves but may cause a
subretinal, vision-limiting, disciform scar. Prognosis is poor once subfoveal
hemorrhage is present.
Severe visual loss can occur in bilateral cases. Low-vision rehabilitation
with appropriate aids such as magnifiers and specialty lenses may be
indicated.
Role of Other Physicians and Health Care Providers

In some instances, a low-vision expert may be beneficial to address severe
bilateral visual loss—often in advanced long-standing disease with CNV.
Not applicable.
Future Treatments
Intravitreal bevacizumab monotherapy has been used in the treatment of
secondary subretinal neovascular membranes associated with choroidal
osteoma with success (57). Other vascular endothelial growth factor (VEGF)
antagonists have also been used successfully (63,64). PDT combined with
intravitreal bevacizumab has also been reported (65). Intravitreal
bevacizumab has been used with positive results in patients with exudative
retinal detachment without neovascular membranes in patients with choroidal
osteoma (66). Additional studies that evaluate the role of VEGF antagonists
for the treatment of secondary neovascular membranes and/or exudative
detachments are needed to better understand the optimal treatment regimen.
CONGENITAL HYPERTROPHY OF THE
RETINAL PIGMENT EPITHELIUM
Prevalence
Congenital hypertrophy of the retinal pigment epithelium (CHRPE) was first
described by Buettner (67) in 1975. CHRPE are hyperpigmented, isolated
lesions that are flat, circumscribed, and well demarcated and represent
congenital hypertrophy of the pigment epithelium with no primary
involvement of the overlying retina (Figure 46-4). Hypopigmented lacunae
are typically present. Because most CHRPE lesions are asymptomatic and
likely undiagnosed, the exact prevalence of this disease is unknown.
FIGURE 46-4 Peripheral CHRPE epithelium,

demonstrating characteristic lacunae.
Multifocal lesions appear in two forms. One form of multifocal disease is
unilateral and consists of group lesions typically in only one sector of the
fundus. These are sometimes referred to as “bear tracks,” or congenital
grouped pigmentation of the retina (68). The second multifocal form of
CHRPE is bilateral. The multifocal lesions in the bilateral form assume a
random distribution and are not grouped together in a single sector of the
fundus (68,69) (Figure 46-5). Bilateral multifocal CHRPE is associated with
familial adenomatous polyposis (FAP) (70–78).
FIGURE 46-5 Multifocal pigmented lesions in a patient

with familial adenomatous polyposis.
FAP is an autosomal dominant disease characterized by more numerous
polyps of the colon and rectum that predisposes patients to colon cancer
before the age of 40. Gardner syndrome involves the development of
extracolonic manifestations of FAP and consists of intestinal polyps, CHRPE,
skeletal hamartomas, and other soft tissue tumors (74). Both intracolonic and
extracolonic presentations are now recognized as phenotypic variations of the
same disease. CHRPE is the most common extraintestinal manifestation
associated with FAP (67). FAP accounts for approximately 1% of all colon
cancers, affecting 1 in 7,500 to 10,000 individuals (67).

The gene responsible for FAP, adenomatous polyposis coli (APC) gene, was
cloned in 1991 and mapped to chromosome 5 (79–81). APC is a tumor
suppressor gene, with a demonstrated involvement in the carcinogenesis of
colon cancer (80). This oncogene plays a role in tumor formation in the
gastrointestinal tract, soft tissue, and bone. One study suggests that APC gene
alterations may lead to defects in RPE melanogenesis and to focal RPE
lesions (82).
Mutation analysis of APC is now an option for many families affected by
FAP. A direct correlation has been demonstrated between the locus of the
APC mutation and the retinal phenotypic disease expression. Careful
delineation of the phenotype of the CHRPE lesions allows focused
investigation of the APC mutation within certain coding regions. CHRPE
lesions are only demonstrated when the mutation is located between codons
464 and 1,387 of the APC gene (83).
No environmental factors have been reported in association with CHRPE.
Worldwide Impact
None, as these lesions rarely affect vision and malignant transformation is
rare.
Pathophysiology
Idiopathic solitary CHRPE histopathologically exhibits a monolayer of
hypertrophied RPE cells with large pigment granules and photoreceptor
degeneration overlying the RPE (67).
In contrast, the histopathology described in Gardner syndrome
demonstrates a more pervasive melanogenesis of the RPE (68). Several
different configurations are described: a monolayer of hypertrophied cells, a
mound of pigmented RPE cells between the RPE basement membrane and
Bruch membrane, or a multilayered mound of hyperplastic RPE in a nodular
or mushroom-shaped configuration (82,84). All of these conformations show
cellular hyperplasia of the RPE. Although the fundus lesions in Gardner
syndrome are referred to as CHRPE, this distinctive feature of cellular
hyperplasia corresponds to a hamartomatous malformation of the RPE, not
hypertrophy alone.
Electron microscopy has shown absence of autofluorescent lipofuscin
granules in CHRPE lesions, suggesting that RPE cells in CHRPE lack the
catabolic functions of normal RPE cells (85).

Congenital hypertrophy of the RPE can present as a solitary lesion or as a
collection of multiple lesions. Although some investigators classify these two
disease forms differently, both forms are included in this discussion due to
their similarities. Several of their clinical and histopathologic differences
have previously been described in this section.
CHRPE lesions are congenital and have been observed in newborns (71).
Solitary CHRPE lesions are demonstrated in the normal population as a
benign incidental finding. A patient presenting with CHRPE typically has
normal vision and normal anterior segment. Fundus examination reveals a
flat, circumscribed, nonprogressive lesion. Size, shape, and degree of
pigmentation of the lesions are quite variable (86). CHRPE lesions can be
oval or round, with areas of pigmentation and depigmentation, and are often
surrounded by a characteristic halo of depigmentation. CHRPE can present
anywhere in the fundus; approximately 70% of these lesions are located in
the temporal quadrant (87).
While CHRPE lesions are typically flat and nonprogressive, a nodular
lesion arising from CHRPE has been reported (88). Nodular growth may
cause exudative retinal detachment and chronic cystoid macular edema (88).
This presentation may represent secondary reactive RPE proliferation or an
acquired adenoma from CHRPE (88). Adenocarcinoma has also been
reported to arise from CHRPE in rare occasions, suggesting that CHRPE
lesions should be observed for neoplastic development, although this disease
course is unusual (89).
The Association of Congenital Hypertrophy of the

Retinal Pigment Epithelium With Gardner Syndrome
and Familial Adenomatous Polyposis
FAP, Gardner syndrome, and CHRPE are all closely related. The ocular
lesions in FAP have been reported to be bilateral in 86% of cases (90). Ocular
lesions are observed in the presence or absence of other systemic
manifestations of Gardner syndrome. The presence of multiple fundus lesions
(more than four) or bilateral lesions has been reported to be a highly specific
and sensitive phenotypic marker for Gardner syndrome (90). CHRPE lesions
have been reported to be present in about two-thirds of families with FAP
(91).
Families with FAP can differ in the presenting number and type of RPE
lesions, although studies have demonstrated that affected individuals within a
single family have similar pigmented lesions (91). In families with known
FAP, the presence of retinal lesions revealed by fundus examination is highly
predictive for the development of intestinal polyps; however, a negative
examination cannot exclude risk for polyp formation (91).
Diagnostic Studies
CHRPE epithelium lesions are very distinctive and are generally diagnosed
clinically. Studies, however, can support the diagnosis. FA demonstrates
blockage of choroidal fluorescence in the hyperpigmented areas of the lesions
in all phases of the study (69). Hypopigmented areas can exhibit
hyperfluorescence, both early and late (window defect). CHRPE lesions do
not leak on FA, as the overlying retinal vasculature and choriocapillaris are
normal (69).
OCT of CHRPE is characterized by retinal thinning, photoreceptor loss,
disorganized retinal anatomy, increased RPE thickness, and decreased
reflectivity of the choroid (69,92,93). Choroidal cavitation has been reported
on OCT (94). Fundus autofluorescence is characterized by
hypoautofluorescence, although the lacunae may show mild
hyperautofluorescence. Fluorescein angiography typically demonstrates
blocking, although the lacunae may show window defects. A corresponding
visual field defect is typically associated (69). Intraretinal extension of the
RPE is a feature not seen in CHPRE lesions that may be unique to some of
the pigmented lesions of FAP and has been detected on OCT (70).
Visual field testing may show scotomas that correspond to the location of
these lesions, presumably representing progressive degeneration of the
overlying photoreceptors (67). Because CHRPE lesions are flat, ultrasound
studies may be unremarkable.
The differential diagnosis of solitary CHRPE lesion should include choroidal
nevus, choroidal melanoma, RPE adenoma, RPE adenocarcinoma, and
combined hamartoma of the RPE. Multiple bilateral CHRPE lesions indicate
greater suspicion of an underlying polyposis syndrome. Chorioretinal scars
from toxoplasmosis, secondary hyperplasia of the RPE, sickle cell disease
(sunburst lesions), sector retinitis pigmentosa, and pigmented retinopathies
should also be considered in the differential diagnosis of CHRPE lesions.
Management
Treatment is not indicated for either solitary or multifocal CHRPE lesions.
These lesions should be observed routinely, and if the patient’s medical
history is suggestive, FAP or Gardner syndrome should be investigated. As
rare malignant transformation has been described, an annual dilated
examination is recommended.
Vision Rehabilitation and Prognosis

The prognoses for vision and for life are very good with solitary CHRPE
lesions. In patients with FAP or Gardner syndrome, the prognosis for life is
altered by the risk of malignant transformation of the colonic polyps.
Ophthalmologic examinations revealing multiple bilateral CHRPE lesions
may identify children and families at risk for developing polyposis and colon
cancer. Rare malignant transformation of a solitary CHRPE to
adenocarcinoma has been described.

Patients with FAP or Gardner syndrome should be referred to a
gastrointestinal specialist with expertise in managing colonic polyps. In some
instances, surgical intervention and referral to an oncologist may be
indicated.
Not applicable.
Future Treatments
Most cases of CHRPE do not require therapy. However, atypical cases may
develop exudation and may respond to VEGF antagonist therapy (95,96).
Because of the rarity of the exudative complications associated with CHRPE,
prospective studies may be limited, and treatment should be performed on an
individualized basis after a complete explanation of risks and potential
benefits.
COMBINED HAMARTOMA OF THE

RETINA AND RETINAL PIGMENT
EPITHELIUM
Prevalence
Combined hamartoma of the retina and of the RPE was first described by
Gass (97) in 1973. Gass organized these lesions into categories based on
location within the fundus: on the disc, next to the disc, in the macula, and in
the periphery.
This ocular hamartomatous malformation involves the sensory retina, the
RPE, the retinal vasculature, and the overlying vitreous. Combined
hamartoma of the retina and RPE is a rare lesion, and its exact prevalence is
unknown.

No known environmental factors or genetic associations have been identified
for combined hamartoma of the retina and RPE.
Worldwide Impact
Unknown, but likely very small as most cases are unilateral.
Pathophysiology
A hamartoma is a benign proliferation of cells that normally are found in the
affected area. Combined hamartoma of the retina and RPE appears
histopathologically as dysplastic retina and overlying retinal vascular
tortuosity (14). Hyperplastic RPE cells migrate into the retina (97). Gliosis
and epiretinal membrane formation commonly are present at the retinal
surface leading to retinal distortion and folding (98). The vitreoretinal
interface is also altered, and tractional changes may be present (99).
These ocular hamartomatous lesions are characterized by benign growth
of glial, vascular, or pigmented tissue from the RPE (100). Some lesions
contain prominent vascular tissue, while others are composed predominantly
of glial tissue, which can lead to formation of preretinal or epiretinal
membranes (97–102).
The pathogenesis of combined hamartoma of the RPE and retina is
uncertain. Although the majority of patients with combined hamartoma do
not present with other systemic diseases, these lesions have been reported in
association with neurofibromatosis (NF), tuberous sclerosis (TS),
incontinentia pigmenti, bilateral colobomas of the optic disc, optic nerve head
drusen, juvenile retinoschisis, juvenile nasopharyngeal angiofibroma, Gorlin
syndrome, and sickle cell anemia (103–116). These associations may indicate
a developmental etiology, as suggested by Gass (97).

Combined hamartoma of the retina and RPE presents most frequently as
painless loss of vision. Other presenting symptoms include metamorphopsia,
floaters, strabismus, leukocoria, and occasionally ocular pain (98). Combined
hamartoma may also be discovered as an incidental finding on routine
ophthalmologic examination. Changes in vision associated with these lesions
occur due to a variety of factors: involvement of the optic nerve or fovea,
alterations at the vitreoretinal surface with epiretinal membrane formation
and subsequent macular traction, or subretinal and intraretinal exudation
(97–102).
On ophthalmologic examination, combined hamartoma of the retina and
RPE appears pigmented and elevated, often with distortion of the retina and
tortuous overlying vessels (Figure 46-6). Because the pigmentation in these
lesions is variable, the diagnosis may be clinically challenging. The location
of the lesion, either juxtapapillary or peripheral, may also have an effect on
its appearance (101). A juxtapapillary lesion characteristically appears as a
solitary elevated mass adjacent to or immediately overlying the optic disc. In
these lesions, contraction of overlying glial tissues often results in striae and
distortion of the retina. Peripheral lesions can resemble an elevated ridge with
accompanying traction of the vessels toward the lesion (105).
FIGURE 46-6 Combined hamartomas of the retina and
RPE are pigmented elevated masses that are frequently
associated with overlying glial tissues that produce retinal
distortion and striae.
Combined hamartoma of the retina and RPE is typically unilateral, although

bilateral cases have been described (100). When bilateral, type 1 or type 2 NF
should be suspected (106–110). Ocular combined hamartoma is a congenital
lesion. The diagnosis can be made at any age, including infancy and most
frequently in childhood (97–102). Combined hamartoma of the retina and
RPE is characteristically nonprogressive, although cases involving growth
have been reported (113,114). Complications associated with these lesions
may include retinoschisis, retinal holes, vitreous hemorrhage, CNV, retinal
hemorrhages, and exudative retinal detachment (117–119).
Diagnostic Studies
Historically, FA has been a useful tool in evaluating combined hamartoma of
the retina and the RPE. Early FA phases demonstrate large tortuous vessels
within the tumor, abnormal retinal capillaries, and late leakage from
anomalous tumor vessels.
Ultrasound studies may show a hyperechoic mass with or without
calcification (120). Ultrasound may also aid in detecting tractional
components.
OCT shows an elevated hyperreflective retinal thickening, retinal
disorganization, hyporeflective shadowing of the underlying tissues, retinal
striae, retinal edema, epiretinal membrane formation, and tractional
components (121,122). OCTA may also aid the diagnosis by highlighting the
anomalous vascular complexes (123).
Combined hamartoma can be a challenging diagnosis due to variable
pigmentation and mass effect (124). Epiretinal membrane formation typically
leads to dysplastic vascular alterations that can be seen in OCT and OCTA
(122,123). Pigmentation and elevation may be similar to choroidal
melanoma; however, dysplastic retinal changes are typically not associated
with melanoma. Early retinoblastoma may also show retinal thickening with
anomalous vasculature; however, pigmentation is not a prominent feature in
early retinoblastoma. Calcification may be present in some combined
hamartomas of the retina and RPE (120).
Peripapillary combined hamartoma has a presentation similar to morning
glory disc anomaly, although the latter condition does not exhibit the
characteristic elevation of combined hamartoma. Advanced cases may show
prominent vascular anomalies that may be similar to those in cicatricial
retinopathy of prematurity.
Management
No well-established treatment plan has been developed for combined
hamartoma of the retina and RPE. Therapy for this lesion has included
treatment for amblyopia, which has demonstrated utility in selected patients
(98). Vitreous surgery with epiretinal membrane removal is another treatment
with reported success (125–129). Other patients with combined hamartoma
have been treated with pars plana vitrectomy and membrane peeling without
any subsequent improvement in their visual acuity (128). Visual prognosis is
guarded.
Patients with combined hamartoma of the retina and RPE demonstrate a wide
spectrum of visual acuity. In one study, approximately one-quarter of the
patients lost at least two lines of visual acuity; approximately one-third of
patients exhibited vision of 20/200 or worse (102). A patient affected with the
characteristically unilateral form of this disease typically experiences normal
vision in the unaffected eye. No report has been made of malignant
transformation of combined hamartoma of the retina and RPE lesions. The
prognosis for life in these patients is normal.
In cases with extreme unilateral visual loss, amblyopia management may
be considered.

Consideration of NF or TS is important, and it can be helpful to work with
geneticists.
Not applicable.
Future Treatments
Currently, children in the amblyopic age range may benefit from early
treatment prior to amblyopia therapy especially with newer small-gauge
vitreoretinal platforms. Vitreomacular and vitreopapillary traction should be
addressed surgically in order to maximize the potential visual outcomes
(126,129). Postsurgical treatment of macular edema with intravitreal
triamcinolone acetonide has also been performed (130). Recently, macular
edema has been treated successfully with VEGF antagonists in atypical cases
(96,131). Recalcitrant vascular activity may also be treated with laser
photocoagulation or PDT (132).
Prospective studies on combined hamartoma of the retina and RPE are
limited due to the rarity of the disease and treatment is typically
individualized depending on the extent of the disease and potential visual
acuity prognosis.
PHAKOMATOSES
The phakomatoses are a group of syndromes characterized by multiple
associated lesions in multiple organ systems. Characteristically, patients with
phakomatoses demonstrate hamartomatous malformations, which are
abnormal proliferations of tissues that are normally found in the affected
organ system. All these syndromes demonstrate ocular manifestations upon
fundus examination. Phakomatoses discussed individually in this chapter
include von Hippel-Lindau (VHL) syndrome, TS, NF, Wyburn-Mason
syndrome (WMS), and SWS.
Von Hippel-Lindau Syndrome
Prevalence
VHL syndrome is an autosomal dominant condition characterized by
hemangioblastomas of the retina, cerebellum, brain stem, and spine,
accompanied by adenomas, angiomas, and cysts of the kidney, liver, and
pancreas. Renal cell carcinoma occurs in approximately 25% of VHL
patients, and pheochromocytoma occurs in approximately 10% (133).
Incidence of VHL is estimated at 1 in 36,000 births (133).
Pathophysiology
Histopathologically, the retinal lesions of VHL are hemangioblastomas
identical to the lesions also found in the central nervous system (CNS) (134).
These vascular masses are composed of retinal capillaries exhibiting normal
endothelium, basement membrane, and pericytes. Capillaries within the
lesion may demonstrate abnormal fenestrations (134,135). Plump vacuolated
interstitial cells with foamy cytoplasm, which are likely of glial origin,
separate the capillary channels (134–136). Growth of these retinal lesions can
extend inward toward the vitreous (endophytic type) or outward toward the
choroid (exophytic type) (137–140).

The pathogenesis of VHL disease is associated with a mutation in the VHL
tumor suppressor gene (141). This syndrome follows an autosomal dominant
inheritance pattern with variable penetrance. The VHL gene was mapped to
chromosome 3p25 in 1988 and was cloned in 1993 (142,143). Previous
studies have suggested that tumor formation in VHL disease follows the
“two-hit” model initially hypothesized by Knudson (144,145) for
retinoblastoma. According to this model, germline transmission of one
mutation (first hit) is followed by a genetic alteration of the second allele in
specific somatic tissues (second hit) (146). Loss of both normal functioning
alleles of the VHL gene results in subsequent loss of functional VHL protein.
Normal VHL protein appears to down-regulate production of VEGF
(147,148). Both the absence of functioning VHL gene product and the
expected up-regulation of VEGF have been demonstrated in retinal capillary
hemangioblastoma (148). Mutations in the VHL gene are highly variable and
include large deletions, small deletions or insertions, and nonsense or
missense point mutations (141). Genetic diagnosis by direct mutation
analysis may be possible in up to 75% of families with VHL (149–151).
DNA testing is a valuable tool in patients with retinal findings suggestive of
VHL, particularly in families having no known history of the disease (152).
No specific environmental factors have been associated with VHL.
Clinical Features
Retinal hemangioblastoma is observed in approximately two-thirds of
patients with VHL disease (153,154). Of reported VHL patients with retinal
tumors, 25% will demonstrate associated cerebellar hemangioblastoma.
Many other organs are affected by cysts, including the pancreas, kidneys,
liver, adrenal glands, and epididymis. VHL is also characterized by renal cell
carcinoma (22%), and pheochromocytoma is a less common but serious
association (154). Although patients with VHL demonstrate brain and CNS
tumor involvement, they exhibit normal cognitive capacity.
A patient with VHL disease may present with decreased visual acuity or
may be asymptomatic with retinal tumors discovered as an incidental finding
upon routine ophthalmologic examination. Alternatively, a patient may
become symptomatic when retinal lesions cause a visual field defect
associated with subretinal fluid or retinal detachment; these lesions may also
cause metamorphopsia, macular edema, traction, full-thickness macular hole,
or epiretinal membrane formation (155). Retinal lesions in VHL may become
visible upon ophthalmologic examination during childhood. Because patients
are frequently asymptomatic, they may be diagnosed at any time throughout
adulthood.
In VHL disease, retinal capillary hemangioblastoma appears as globular
red-orange masses in the fundus. A characteristic feature of these retinal
tumors is a pair of dilated, tortuous feeding and draining vessels traveling
between the lesion and the optic nerve (Figure 46-7). Hemangioblastomas
may form anywhere in the fundus: rarely at the posterior pole (1%), more
commonly at the optic disc (8%), and most frequently in the temporal
FIGURE 46-7 Retinal capillary hemangioblastoma in
VHL syndrome has a striking appearance. This example
demonstrates a pair of tortuous feeder vessels.
Patients with VHL disease demonstrate retinal masses that are solitary or
multiple, with the mean number of these lesions in genetic carriers of the
disease reported as 1.85 (range of 0 to 15) (153). Lesions may present either
unilaterally or bilaterally. Solitary unilateral lesions are less likely to be
associated with VHL disease, whereas multiple bilateral lesions are
invariably indicative of VHL syndrome. The size of these lesions varies,
ranging from small vascular tufts to large masses. Retinal tumors in VHL
demonstrate limited ability to proliferate; however, leakage of thin vessels
with subsequent fluid buildup may result in the appearance of tumor growth.
Lipid exudate in the macula is a common cause of decreased visual acuity in
these patients.
Complications of retinal hemangioblastoma in VHL include exudation of
fluid into the subretinal space and subsequent retinal detachment. Preretinal
membranes may form causing traction on the retina. Patients who develop
retinal detachments are at significant risk for secondary glaucoma, cataract,
or vitreous hemorrhage. Disc and retinal neovascularization may also occur
(156).
The differential diagnosis of retinal capillary hemangioblastoma associated
with VHL disease varies with the location of the lesions. Peripheral lesions
with dilated feeding and draining vessels are highly characteristic and are
frequently diagnosed based on ophthalmoscopic appearance. In the presence
of massive subretinal exudation with retinal detachment, however, the lesion
may resemble other entities. In children, the differential diagnosis should
primarily include Coats disease, familial exudative vitreoretinopathy, and
retinoblastoma. Additional lesions considered in the differential may include
racemose hemangioma, retinal cavernous hemangioma, sickle cell
retinopathy, retinal astrocytoma, and nematode endophthalmitis (153).
The differential diagnosis for juxtapapillary capillary hemangioblastoma,
particularly the exophytic form, involves entities distinct from those
considered in the diagnosis of peripheral lesions. Exophytic juxtapapillary
hemangioblastoma may obscure the disc margin, making this lesion
challenging to distinguish from other causes of disc edema. The swollen disc
appearance in juxtapapillary capillary hemangioblastoma is more frequently
unilateral, although bilateral cases have been reported. Papilledema
subsequent to increased intracranial pressure may be a diagnostic
consideration in patients presenting with intracranial VHL lesions. CNV and
choroidal hemangioblastoma may also enter the differential diagnosis in VHL
(156,157).
Diagnostic Studies
The diagnosis of peripheral capillary hemangioblastoma is frequently made
prior to performing any ancillary studies because of the characteristic
appearance of the lesion on ophthalmoscopy. Standard fundus photography is
helpful to document lesion growth, stability, or response to treatment. FA
shows rapid filling of the tumor by the feeding artery in early phases.
Midphase photographs show intense staining of the tumor, and late frames
show leakage of dye from the tumor into the vitreous (156). FA may be
particularly useful in distinguishing juxtapapillary capillary
hemangioblastoma from other causes of disc edema.
OCT and OCTA may also aid in the diagnosis by allowing visualization
of the retinal tumoral mass, highlighting the feeder and draining vessels, and
showing retinal edema in the juxtatumoral retina (158,159). OCT helps to
identify exophytic and endophytic growth. However, to localize small
tumors, FA may be preferable since some may be undetected by
ophthalmoscopy.
A detailed family history should be taken in all patients diagnosed with
retinal hemangioblastoma, with suspicion of VHL disease. In addition,
patients presenting with retinal hemangioblastoma should be screened for
other manifestations of VHL, with an MRI of the head and spine, as well as a
CT scan of the abdomen.
Management
Management of retinal capillary hemangioblastoma is oriented toward
reducing the destructive exudation associated with these lesions. Various
methods for ablating retinal capillary hemangioblastoma have been
previously reported; mainstays of treatment, however, have included laser
photocoagulation and cryotherapy (160–168). Small asymptomatic lesions
may remain stable for many years, and observation on regular follow-up is an
option in these patients (162). For symptomatic patients, laser
photocoagulation is effective in treating smaller and more posterior lesions
not associated with significant retinal detachment. Cryotherapy is efficacious
in larger and more anterior lesions. In a large series, laser photocoagulation
effectively controlled 18 of 18 (100%) extrapapillary hemangioblastomas
measuring 1.5 mm or smaller in diameter and 8 of 17 (47%) larger lesions
(162). Seven of eight juxtapapillary lesions were controlled with laser.
Extrapapillary lesions larger than 1.5 mm were successfully controlled with
cryotherapy in 28 of 39 (72%) cases (162). More than one session of
cryotherapy or laser treatment may be required for control of the exudative
process. For lesions larger than 3.5 to 4 mm, treatment with plaque
radiotherapy (Figure 46-8) has been demonstrated to be more effective than
cryotherapy (162,169). In cases involving large exudative detachments,
rhegmatogenous detachments, or tractional detachments of the macula,
operative intervention with scleral buckling or vitrectomy may be beneficial
(170–175).
FIGURE 46-8 Plaque brachytherapy for large solitary

retinal capillary hemangioblastomas.
Treatment of retinal capillary hemangioblastoma with transpupillary
thermotherapy and PDT has been recently reported (176–180). The role for
these modalities in the treatment of capillary hemangioblastoma remains
unclear. PDT may demonstrate theoretical advantages over other treatments
(179,180).
Recent studies have also reported rapid and significant improvement in
visual acuity in patients with VHL hemangioblastoma following systemic
treatment with the VEGF receptor inhibitor, SU5416 (181,182). Although
lesion size remained unchanged by treatment in these studies, associated
cystoid macular edema was significantly decreased (181,182). In patients
with VHL-associated or solitary non-VHL angiomas, long-term follow-up is
important to promptly detect and, if necessary, to treat any further
development of symptoms, growth of lesions, or new retinal tumors. Some
studies have suggested limited benefit of intraocular injection of anti-VEGF
compounds in treating intraocular hemangioblastomas.
Untreated, retinal hemangioblastoma typically has a poor prognosis. Severe
vision loss has been associated with presentation of retinal lesions at an early
age (153). In one study, the prognosis for vision in patients presenting with
retinal hemangioblastoma was reported as better in those individuals whose
diagnosis did not include VHL disease than in those who were diagnosed
with the VHL syndrome (183). Another study found equal visual outcome in
both these groups (184).

All patients with VHL should be under the care of a pediatrician or internist
with expertise in this disorder. There is a significant risk of nonocular
malignancies, particularly renal cell carcinoma. The most common causes of
mortality in patients with VHL are cerebellar hemangioblastoma and renal
cell carcinoma (154). Given the genetic associations of this disease, referral
and consultation with a geneticist are indicated.
Not applicable.
Future Treatments
Discovery of the VHL protein targeting hypoxia-inducible factors and the
protein’s role in angiogenesis may facilitate the development of new
treatments. Development of drugs specifically active on hypoxia-inducible
factors and VEGF may play a role in the future in treating VHL disease
(185). Recent clinical studies evaluating the potential benefits of ranibizumab
only showed efficacy with intravitreal VEGF antagonists when tumors were
small (186). Recent studies have shown that intravitreal VEGF antagonists
may prove beneficial either alone or in combination with other modalities
(187,188). Intravitreal corticosteroids have also been used with success for
secondary macular edema (189). In pediatric patients, laser photocoagulation
and PDT may be preferable in select cases to decrease the burden of
intravitreal injections (190). Advanced cases may be managed with low-dose
external beam radiotherapy and plaque brachytherapy (191). Selective intra-
arterial bevacizumab has also been used with positive outcomes (192). Future
prospective studies are needed to better understand how intravitreal VEGF
antagonists and corticosteroids may be used with optimal outcomes.
Tuberous Sclerosis
Prevalence
TS is a rare, hereditary phakomatosis first described by Bourneville (193) in
1880. The disorder has been classically characterized by a triad of presenting
symptoms: seizures, cognitive developmental deficits, and adenoma
sebaceum of the skin (194). Clinical expression of TS, however, is highly
variable. The National Tuberous Sclerosis Association has established a more
complex set of diagnostic criteria based on the presence of one or more of the
following features: adenoma sebaceum, ungual fibroma, cortical tubers,
subependymal nodules, retinal astrocytic hamartoma, cardiac rhabdomyoma,
hypopigmented (ash-leaf) skin patches, and infantile spasms, among other
features (195). The incidence of TS has been estimated at 1 in 15,000 live
births. Retinal astrocytic hamartoma develops in approximately 50% of
patients with TS (196,197).

TS is an autosomal dominant syndrome, demonstrating incomplete
penetrance and variable expression. It is estimated that up to 80% of cases
represent new mutations. Two genetic loci, 9q34 and 16p13.3, have been
identified in association with TS. The TSC2 gene was identified in 1993; its
protein product is called tuberin (198,199). Tuberin is a tumor suppressor,
high levels of which are found in the brain, kidney, heart, skin, and vessels.
The protein product of TSC1, hamartin, is also thought to function as a tumor
suppressor. Hamartin and tuberin are believed to act in synergy to regulate
cell growth and differentiation (198,199).
No specific environmental associations are known in TS.
Pathophysiology
Histologically, retinal astrocytic hamartoma in TS is composed of spindle-
shaped fibrous astrocytes containing small oval nuclei arising from the nerve
fiber layer of the retina (200,201). Larger lesions may have areas of calcific
degeneration and may contain cystoid spaces filled with serous exudate or
blood. A rare histopathologic variant, giant cell astrocytoma, has also been
reported (202).

Although the most common ocular manifestation of TS is the retinal
astrocytic hamartoma, this lesion is not pathognomonic for the syndrome.
Retinal astrocytoma is a benign lesion that can also occur in patients with NF
and rarely in the normal population. These retinal lesions are most commonly
located near the macula and, more rarely, may involve the optic disc.
However, they typically do not significantly affect vision. When associated
with TS (or NF), retinal astrocytoma is more likely to be found as multifocal
lesions than when not associated with an underlying systemic condition
(203).
Three basic types of retinal astrocytoma have been described based on
appearance (204). The first type, often found in young children, is flat with a
smooth, translucent appearance. Frequently observed in older patients, the
second type is nodular and elevated with a calcified appearance. Although
age associations with the type of lesion are observed, either of these two
types may be demonstrated in patients of any age. Translucent retinal
astrocytoma is more challenging to detect clinically, and in some
presentations, only the obscuration of the retinal vessels is observed. The
nodular lesions are frequently more easily visualized and have a characteristic
yellow-to-white coloration and a cluster-like appearance. The third type of
these astrocytic lesions exhibits characteristics of both of the other two forms.
These mixed-type lesions (Figure 46-9) may result from the translucent
hamartoma form evolving over time into the mulberry-cluster, calcified form,
although this process has not been completely elucidated.
FIGURE 46-9 Astrocytic hamartomas in TS vary greatly

in appearance.
Retinal astrocytic hamartoma is frequently congenital and nonprogressive.
These lesions have been reported in infancy, diagnosed within the first few
weeks of life (205). In some cases, these retinal tumors have been reported to
progress (206), and some have been observed to calcify over time (206). New
lesions may arise from the retina that appeared normal on presentation in
some patients with TS, suggesting that not all retinal astrocytomas are
congenital (204). Spontaneous regression of these retinal lesions in the setting
of TS has been reported (207).
Complications of retinal astrocytic tumors reported in rare cases include
vitreous seeding, vitreous hemorrhage, macular edema, and vitreoretinal
traction at the tumor surface (208–210). A rare invasive giant cell
astrocytoma of the retina was reported in one patient presenting in infancy
with TS. The tumor’s steadily aggressive growth was associated with
subsequent development of neovascular glaucoma and spontaneous
perforation of the sclera. Because the eye became blind and painful,
enucleation was performed when the patient was 12 years of age (210).
Other ocular manifestations described in association with TS include
fundus depigmentation, iris depigmentation, subconjunctival nodules, eyelid
angiofibroma, hamartoma of the iris and ciliary epithelium, and ocular
coloboma (211,212).
Nonophthalmologic features of TS include facial angiofibroma, ungual
fibroma, benign lesions of the CNS (cortical tubers and cerebral
astrocytoma), and hamartomatous tumors of virtually any organ in the body,
including the kidneys, heart, liver, and lungs (213). Patients may demonstrate
skin lesions such as hypopigmented macules (“ash-leaf spots”), which are
characteristic of TS and are sometimes present at birth. Also characteristic of
TS are shagreen patches, which are thickened skin lesions commonly found
on the back. Facial angiofibroma, referred to as adenoma sebaceum, is a
condition associated with TS that presents in childhood and must be carefully
distinguished from acne. Tuberous cortical lesions and nodular lesions in the
basal ganglia and periventricular area are also presentations in TS. Seizures
are frequently observed in patients with TS, and cognitive deficits or
developmental delay is present in approximately half of TS patients.
Malignant astrocytoma may present in TS, although this lesion is observed
infrequently in this syndrome.
Diagnostic Studies
In patients with TS, retinal astrocytoma demonstrates early hypofluorescence
on FA with late staining. A-scan ultrasound indicates a mass of medium
reflectivity, while B-scan may demonstrate focal calcifications similar to
those observed in retinoblastoma. The ultrasound pattern may closely
correspond to that associated with choroidal osteoma and some cases of
retinoblastoma (200).
OCT is particularly helpful because tumors are hyperreflective and
located in the retinal nerve fiber layer (214,215). Small lesions show a
smooth dome-shaped mass, while larger lesions may show cavitation,
posterior shadowing, and a surface “moth-eaten” appearance. OCT may also
help distinguish it from presumed solitary circumscribed retinal astrocytic
proliferation and in some cases of early retinoblastoma (215). OCTA has also
shown intrinsic inner retinal vascular ectasia (215,216).
Diagnostic studies are frequently selected to distinguish TS from NF.
Neuroimaging is helpful in demonstrating the subependymal nodules of TS,
as well as in distinguishing the sphenoid dysplasia, meningioma, or vestibular
schwannoma of NF.
Retinoblastoma is an important consideration in the differential diagnosis of
retinal astrocytic hamartoma in TS, due to the frequent characteristic
presence of focal areas of calcification in both types of lesions. Other entities
within the differential diagnosis include presumed solitary circumscribed
retinal astrocytic proliferation, amelanotic choroidal melanoma, Coats
disease, myelinated nerve fiber layer, and choroiditis. In patients with TS,
systemic findings of the disease, such as the various skin lesions and
intracranial findings, provide specific clues to aid in diagnosis. Retinal nerve
fiber layer involvement in OCT without outer retinal involvement is highly
characteristics of astrocytic hamartomas.
Management
Except in rare cases, retinal astrocytic hamartoma of TS requires no
treatment. Current treatments for this syndrome are supportive rather than
curative. One complication of TS requiring long-term care is seizure,
necessitating neurologic consultation and management. Antiepileptic drugs,
dermatology treatments, and occupational therapy for developmental deficits
are commonly employed in treating TS patients.
In patients with ocular lesions associated with TS, the prognosis for vision is
good, because visual acuity is frequently unaffected unless retinal tumors
involve the fovea. The most common causes of morbidity in patients with TS
are neurologic complications of intractable seizures and hydrocephalus.
Renal hamartoma in TS patients may also be associated with significant
complications. Cardiac or pulmonary complications may arise in the presence
of lesions in the heart or lungs.

These patients are often managed in concert with pediatric neurologists given
the significant risk of seizure disorder. A consultation with a geneticist is
recommended.
Not applicable.
Future Treatments
Intravitreal VEGF antagonists and corticosteroids have been used
successfully in cases of exudative retinopathy and macular edema in patients
with TS (217). Vitreoretinal surgery combined with VEGF antagonists has
also been reported to be beneficial in a patient with exudative retinal
detachment (218). Recent studies have shown systemic everolimus and
sirolimus may be efficacious at inducing regression of astrocytic hamartomas
in select cases (219,220).
Neurofibromatosis
Prevalence
NF is a heritable phakomatosis characterized by lesions composed of
melanocytes or neuroglial cells (221). Initial signs of the disease may be
present at birth or develop throughout childhood and adolescence as the
disease progresses. NF-1 and NF-2 are two distinct forms of NF, each
demonstrating different clinical and genetic features. The primary retinal
lesions in NF-1 are astrocytic hamartomas, which can be identical to those
found in TS. NF-1, the much more common form, affects approximately 1 in
4,000, while NF-2 affects approximately 1 in 50,000.

NF-1 and NF-2 are both autosomal dominant diseases with high penetrance.
Nearly 50% of cases of NF-1 are sporadic, and the other half of cases is
transmitted genetically. The gene for NF-1 is located on chromosome 17; the
gene for NF-2 is on chromosome 22 (222).
Neurofibromin has been identified as the protein product of the NF-1
gene; this protein is believed to play a role in the regulation of cellular
proliferation and tumor suppression. Mutation of the NF-1 gene leads to
subsequent formation of the many different tumors of this syndrome.
Pathogenesis of the tumors is believed to involve the Ras pathways (222). No
known environmental factors are associated with NF.
Pathophysiology
NF is a disorder of the neuroectodermal cell line. NF tumors originate from
neural crest cells, such as sensory neurons, Schwann cells, and melanocytes.
Clinical Features
NF-1 is a progressive disease with numerous and variable manifestations
affecting multiple organ systems including the eye, skin, and CNS. Retinal
tumors associated with NF-1 include retinal astrocytic hamartoma (most
common), retinal capillary hemangioblastoma, vasoproliferative tumors, and
combined hamartoma of the RPE and retina. These lesions may all cause
vision loss in these patients.
Similar to the retinal astrocytic lesions in TS, retinal astrocytic
hamartomas in NF-1 are benign and are often located near the optic disc. One
series reported 42 cases of astrocytic retinal tumors. Of these patients, 14%
had NF, and their tumors were more frequently located adjacent to or on the
disc (222). These lesions demonstrate the white, mulberry-cluster appearance
of astrocytic hamartoma also observed in TS.
Combined hamartoma of the retina and RPE in NF has been reported
(101,223). Retinal capillary hemangioblastoma has also been described in
association with NF (224). Optic nerve glioma is another characteristic tumor
in NF-1, which can cause significant vision loss or proptosis (225). These
lesions may be unilateral or bilateral and may also affect the optic chiasm.
Optic nerve glioma typically presents symptomatically in young children.
Nonocular complications of this tumor include pituitary dysfunction and
hydrocephalus. Optic nerve sheath meningioma may also be associated with
NF-1, although this lesion is less common in childhood.
NF-1 is characterized by uveal masses of the iris and choroid. On dark
irides, the iris lesions, termed “Lisch nodules,” may appear hypopigmented,
whereas on light irides, the masses demonstrate darker pigmentation. The iris
lesions increase in number with age and are observed nearly universally in
adults with NF-1 (226). Choroidal lesions in NF-1 are less common than
those in the iris but are still demonstrated in approximately one-third of these
patients as flat masses, ranging in coloration from white to yellow to darkly
pigmented. Patients with NF-1 are also believed to have an increased risk for
development of uveal melanoma.
Numerous nonocular findings are demonstrated in NF-1. Flat,
hyperpigmented macules (café au lait spots) are the most common cutaneous
presentation in this disease (221). The number and size of these cutaneous
macules vary, and they become larger and more numerous with increasing
age. Other distinctive findings in NF-1 include nodular cutaneous and
subcutaneous neurofibroma, plexiform neurofibroma, and bony lesions (221).
Plexiform neurofibroma in NF-1 may involve the eyelid, giving it an S-
shaped appearance. These eyelid lesions may cause ptosis and may be
challenging to resect.
Pheochromocytoma, other soft tissue tumors, and benign and malignant
CNS tumors have also been reported in association with NF-1. Patients with
CNS abnormalities may demonstrate hydrocephalus, seizures, cognitive
deficits, and developmental delay. Other findings of NF-1 include visceral
tumors such as gastrointestinal neurofibromas, enlarged corneal nerves,
choroidal ovoid bodies, and orbital neurofibromas.
NF-2 is less prevalent than NF-1 and is characterized by acoustic
neuroma, neurofibroma, meningioma, glioma, and/or schwannoma. The most
common eye finding in NF-2 is either cortical or posterior subcapsular
cataract (227). NF-2 patients often have secondary keratopathy due to facial
palsies. As in NF-1, retinal hamartoma and Lisch nodules may be present,
although these lesions are much less common in NF-2 (226). Complications
of both NF-1 and NF-2 arise primarily due to progression of the
hamartomatous tumors in the eye, skin, and CNS.
Association of Neurofibromatosis 1 and Glaucoma

NF-1 is associated with glaucoma, which is most commonly found ipsilateral
to the eyelid involved by plexiform neurofibroma. In infancy, glaucoma in
patients with NF-1 is associated with buphthalmos. NF-1 patients with
glaucoma may demonstrate abnormalities of the trabecular meshwork,
whereas others may exhibit angle closure due to progression of lesions
infiltrating the angle. Neovascular glaucoma has also been reported in
pediatric patients (228).
Diagnostic Studies
Patients demonstrating any findings suggestive of NF-1 should have a
complete ophthalmologic examination to evaluate optic nerve function, iris,
disc appearance, choroid, and intraocular pressure. On FA, retinal
astrocytoma, the most common retinal lesion in NF-1, is hypofluorescent on
early phases with late staining. B-scan ultrasonography may reveal focal
calcifications in these lesions. CT scanning and MRI are helpful in evaluating
extension of optic nerve or optic chiasm glioma, although routine screening
with MRI in the absence of nerve abnormalities on ophthalmologic
examination may not be indicated in all cases (229).
OCT has been used to differentiate lesions and monitor progression in
patients with intraocular and orbital tumors associated with NF-1 and NF-2
(230–233). Decrease in retinal nerve fiber layer may characteristically be
seen in patients with optic nerve gliomas.
When NF-1 or NF-2 is suspected, the differential diagnosis of a retinal mass
resembling an astrocytic hamartoma in a child or teenager should also include
retinoblastoma, Coats disease, toxoplasmosis, toxocariasis, and choroidal
melanoma. Presentation with other associated manifestations of NF,
including cutaneous lesions, is also diagnostically helpful.
Management
Retinal astrocytomas associated with NF are typically observed. However, in
one series of patients with NF-1, these retinal lesions progressed requiring
surgical intervention including retinal detachment repair, photocoagulation,
or cryopexy (110).
Optic nerve glioma continues to be a therapeutic challenge due to
variable progression. Optic nerve tumors in NF-1 frequently do not progress
in the years immediately following diagnosis (234). Unless these lesions
exhibit aggressive growth, they are often observed. Surgical excision of optic
nerve glioma can be globe preserving, although resection frequently does not
preserve vision in the affected eye. Radiation and chemotherapy have been
employed as treatments in optic nerve glioma with varying degrees of success
(101,235,236). Optic nerve glioma has also been reported to exhibit
spontaneous regression (237).
A medical regimen is typically the initial treatment for NF-1–associated
glaucoma, although this condition may prove intractable to medical therapy.
Surgical intervention is frequently required including goniotomy,
trabeculotomy, trabeculectomy, and installation of an aqueous shunt.
Treatment of cutaneous manifestations of NF includes possible resection
of the neurofibroma tumors. However, these lesions often recur and are only
excised in cases in which the patient is experiencing pain or significant
impairment. Pigmentation defects in NF are not treated. CNS manifestations
of this disease may require treatment with anticonvulsants, and in the setting
of hydrocephalus, neurosurgical treatment may be necessary. Visceral tumors
may also require surgical resection.
Visual acuity in NF-1 patients with optic nerve abnormalities is often
reduced, particularly in patients demonstrating concurrent amblyopia in the
affected eye. Patients may have increased morbidity and mortality when they
exhibit more severe presentations of NF, such as CNS tumors, severe
seizures, or other aggressive malignancies.

NF is best treated in a multidisciplinary setting with neurologists providing
primary care. In cases of optic nerve glioma associated with visual loss,
medical and radiation oncology should be involved. If a surgical approach is
elected, an experienced orbital surgeon working in concert with neurologic
surgery should be considered.
Not applicable.
Future Treatments
Knowledge of the role in tumorigenesis of the Ras protein and VEGF has led
to treatment of some patients with VEGF antagonists (238,239). Most
recently, intravitreal bevacizumab has been used to treat retinal
vasoproliferative tumors in NF-1 (240). Intravitreal VEGF antagonists and
corticosteroids have also used successfully in cases of exudative retinopathy
and macular edema in patients with astrocytic hamartomas (217).
Vitreoretinal surgery combined with VEGF antagonists has also been
beneficial in select cases (218). Recent studies have shown systemic
everolimus and sirolimus may be efficacious at inducing regression of some
astrocytic hamartomas (219,220). Patients with optic nerve gliomas and NF-1
may also benefit from systemic VEGF antagonists (241).
Wyburn-Mason Syndrome
Prevalence
Initially described and named after its discoverer in 1943, WMS is a rare,
nonheritable disorder characterized by AV malformations of the eye and CNS
(242). Congenital AV malformations in WMS primarily involve the retina,
optic disc, and midbrain; the retinal lesion is known as racemose
hemangioma. AV malformations may also occur elsewhere in the body in
WMS, including the skin, nasopharynx, orbit, lung, and spine. Although the
exact incidence is unreported, WMS is considered a rare condition
worldwide.

WMS is nonheritable, and no known genetic associations or environmental
factors have been found.
Pathophysiology
Although it has been determined that WMS is a congenital, nonheritable
disorder, the pathogenesis has not otherwise been delineated (243,244).
Vessel walls demonstrate fibromuscular medial coats of variable thickness
and acellular fibrohyaline adventitial coverings. Dilated vascular channels
may occupy the entire thickness of the retina. Cystoid changes may be
observed, as well as loss of ganglion cell bodies and axons (245,246). Further
details of the histopathology of racemose hemangioma in WMS remain to be
discovered.
Clinical Features
Most patients with racemose hemangioma in WMS experience reduced visual
acuity. The extent of the vascular malformation varies widely, and the lesions
have been divided into three groups with clinicopathologic characteristics
ranging from least to most severe (244). Group I is comprised of patients
demonstrating interposition of an abnormal capillary plexus between a major
communicating artery and vein. These patients are typically asymptomatic,
and retinal lesions in this group are rarely associated with cerebrovascular
malformations. Group II patients demonstrate direct AV communications
without the interposition of capillary elements. Microvasculature adjacent to
AV lesions may be altered, and beading and multiple fusiform dilations of the
large vessel walls may be observed. Group III patients demonstrate many
anastomosing channels of large caliber. These channels are so intertwined
and convoluted that separation into their arterial and venous components may
be difficult. Perivascular sheathing, exudation, and pigmentary degeneration
may also be observed. Fundus changes in group III patients are similar to
those originally described by Wyburn-Mason (242,243), and visual acuity in
this group is frequently poor. Group III patients also demonstrate a high
incidence of CNS lesions (244). One study suggests that group I and II retinal
vascular lesions are typically isolated. If patients in these first two groups are
asymptomatic, systemic workup is not indicated (247). More severe cases
may be associated with retinal vascular occlusion, retinal ischemia, and focal
exudation, which can lead to progressive vision loss (248–250). Racemose
hemangioma manifests unilaterally and is typically nonprogressive although
the pattern of vascular tortuosity may alter over time.
CNS AV malformations in WMS are more frequently observed ipsilateral
to the retinal lesions. In one series of 80 cases of retinal AV malformations,
30% of these patients also demonstrated CNS malformations (251). Retinal
lesions in WMS may extend from an intracranial AV malformation, traveling
along the optic nerve to the retinal vasculature.
Visual acuity in WMS patients with retinal AV malformations depends
upon the size and extent of the defective vasculature and ranges from normal
to severely reduced. Patients with diffuse, large, markedly dilated, and
tortuous vessels are frequently severely impaired and are consequently often
diagnosed earlier than asymptomatic patients with milder forms of WMS.
Reported visual field defects in patients with this syndrome indicate scotoma
associated with retinal AV malformations (252). Complications of the retinal
AV malformations reported in WMS include intraocular hemorrhage,
secondary neovascular glaucoma, macular hole, central retinal vein
obstruction, macroaneurysm, retinal hemorrhage, and vitreous hemorrhage
(248–253). Extensive peripheral retinal ischemia, neovascularization, and
choroidal infarction have also been reported (254). Retinal detachment and
exudation are not typical in racemose hemangioma.
Neurologic symptoms in WMS depend on the location and size of the
CNS lesions. Patients demonstrating these lesions may develop headaches,
cranial nerve palsies, visual field abnormalities, seizures, weakness, mental
status changes, and papilledema. CNS lesions in WMS are commonly
hemorrhagic.
Diagnostic Studies
In most cases, the diagnosis of racemose hemangioma in WMS may be
determined upon examination by indirect ophthalmoscopy. FA demonstrates
rapid filling of the vascular malformation and provides dramatic
documentation of the lesion. In advanced cases, differentiation between
artery and vein may not be possible. MRI of the brain is indicated to delineate
CNS manifestations in patients with more severe forms of WMS. OCT may
show diffuse or focal retinal thickening, vascular ectasia, and vitreoretinal
interface anomalies (255,256). B-scan shows hyperechoic fundus thickening
with vascularity.
Other retinal vascular abnormalities, such as retinal arterial and venous
collaterals, retinal telangiectasis, retinal capillary hemangioblastoma, retinal
cavernous hemangioma, and retinal vasoproliferative tumors, should be
included in the differential diagnosis of racemose hemangioma in WMS.
Management
No treatment is indicated for the primary lesions in WMS. The utility of laser
photocoagulation or cryotherapy for these lesions has yet to be clearly
defined.
Visual prognosis for patients with ocular AV malformations in WMS varies
widely depending upon the extent of retinal or optic nerve involvement. In
patients demonstrating less severe forms of this disease, the prognosis for
vision and quality of life is typically good. For patients with more severe
manifestations of WMS, associated CNS vascular malformations can lead to
cerebral hemorrhage with potentially devastating consequences.
WS is best treated in a multidisciplinary setting with neurologists providing
primary care.
Not applicable.
Future Treatments
Intravitreal VEGF antagonists have been used with visual and anatomic
improvements in a patient with WMS and central retinal vein occlusion
(257). These may play a role in patients with WMS that develop serous
retinal detachment and cystoid macular edema (258). The medical literature
continues to be scarce regarding potential targets for ocular therapy in
patients with WMS.
Sturge-Weber Syndrome
Prevalence
SWS, also referred to as encephalotrigeminal angiomatosis, is a
nonhereditary phakomatosis manifesting a range of symptoms from partial to
complete expression of the disease (259). In its complete form, SWS is
characterized by ipsilateral angiomatous malformations involving the face,
brain, and eye (260). Patients frequently demonstrate seizures and intracranial
calcifications (196,206). Characteristic ocular findings of SWS include
diffuse choroidal hemangioma and glaucoma (5,260). The exact incidence
and prevalence of SWS are not known.

SWS is associated with no known environmental or genetic factors.
Pathophysiology
In contrast to circumscribed choroidal hemangioma, the diffuse type
observed in SWS demonstrates a gradual transition at the margin of the lesion
with progressively less engorgement of the vessels as observed on light
microscopy (5). In one large histopathologic series, diffuse choroidal
hemangiomas were classified as mixed cavernous and capillary-type tumors
exhibiting both large and small blood vessels (5).
The pathogenesis of SWS is poorly understood. SWS is believed to be
associated with a defect in neural crest cell migration and differentiation
(261). These precursor cells give rise to ocular tissues, meninges, and the
dermis. Overproduction of angiogenic factors may play a role in the
pathogenesis of this disease.

Patients with SWS are usually diagnosed at birth or in early infancy when the
presence of a facial hemangioma (port-wine stain or nevus flammeus)
prompts evaluation. Choroidal thickening associated with diffuse choroidal
hemangioma may produce prominent hyperopia with associated
anisometropic amblyopia. Retinal striae may also be present (Figure 46-10).
Glaucoma may result from iris neovascularization or from episcleral vascular
changes associated with SWS. On fundus examination of patients with
choroidal hemangioma, a diffuse red or orange thickening of the choroid is
observed posteriorly. In some cases, the fundus of the affected eye
demonstrates a dramatically deeper red coloration than that of the opposite
eye, and the term “tomato-catsup fundus” has been used to describe this
appearance (262). Diffuse choroidal hemangioma in SWS is usually thickest
in the macular region and then blends imperceptibly into the normal choroid
anteriorly.
FIGURE 46-10 Diffuse choroidal hemangioma with
secondary subretinal fluid and retinal striae.
Dilated and tortuous retinal vessels are commonly observed in eyes with
diffuse choroidal hemangioma. Patients with SWS may also demonstrate
exudative retinal detachment with cystoid degeneration of the macula. These
exudative detachments are associated with subretinal fluid that shifts with
movement of the patient’s head. Total retinal detachment with secondary
cataract and leukocoria may be observed (263). Development of CNV
associated with choroidal hemangioma has also been described (11).
Glaucoma is the most common and most serious ocular manifestation in
SWS, presenting in approximately 70% of SWS patients (260). Intraocular
pressure may be elevated at birth as a form of congenital glaucoma, or
glaucoma may become symptomatic later during childhood. Intraocular
pressure in patients with SWS may be elevated secondary to increased
episcleral venous pressure or a developmental defect in the angle. Additional
ocular manifestations in patients with SWS include vascular malformations
of the eyelids, episclera, conjunctiva, retina, and choroid. Patients with SWS
may demonstrate retinal vascular tortuosity, iris heterochromia, and
strabismus (260). SWS with bilateral optic neuropathy has been reported
(264).
Cutaneous and CNS lesions of SWS are also congenital. Cutaneous
lesions, or nevus flammeus, present ipsilaterally to brain vascular
malformations. Characteristically, SWS patients demonstrate a sharply
demarcated port-wine–colored lesion that may involve the scalp, forehead,
eyelids, and lower face. These cutaneous lesions may undergo thickening
over time, with hypertrophy of underlying bone and soft tissues. CNS
angiomatosis in SWS may lead to calcium deposition in the brain. Decreased
cerebral volume with venous abnormalities and enlargement of the choroid
plexus may also be observed. Clinically, these lesions may present in SWS
patients as seizures, cognitive developmental deficits, hemiplegia, and other
focal neurologic deficits.
Diagnostic Studies
SWS is frequently diagnosed clinically based upon the presence of a port-
wine stain and choroidal thickening due to diffuse choroidal hemangioma, as
well as upon other characteristic ocular and CNS symptoms. FA shows
diffuse early choroidal enhancement (263). In cases where chronic exudative
detachments are present, patchy blocking of the choroidal pattern may be
seen in areas of corresponding hyperpigmentation. B-scan ultrasonography
demonstrates marked thickening of the choroid, often with overlying retinal
detachment, while A-scan ultrasound demonstrates high internal reflectivity.
On FA, widespread early filling of the tumor with late leakage is observed
(263). OCT invariably shows increased choroidal thickness (265). Subretinal
fluid, deep retinal alterations, and intraretinal fluid may also be detected on
OCT (265). Although not commonly utilized in SWS, CT and MRI scans can
be helpful because CT scan can demonstrate abnormal thickening of the
choroid with enhancement of the globe while MRI exhibits a distinctive high
signal on T1-weighted images (266).
The differential diagnosis of choroidal hemangioma in SWS includes
circumscribed choroidal hemangioma unassociated with SWS, amelanotic
choroidal nevus or melanoma, choroidal osteoma, retinal pigment epithelial
detachment, scleritis, and retinal capillary hemangioblastoma.
Management and Visual Rehabilitation

In SWS, children with diffuse choroidal hemangioma may demonstrate
associated hyperopic shift, glaucoma, and exudative retinal detachment.
Refraction, corrective lenses, and amblyopia therapy are indicated for
hyperopia and anisometropic amblyopia. Port-wine lesions may be treated
with laser to decrease vascularity for improved cosmesis. When glaucoma is
treated with filtering surgery, subretinal exudation may worsen in the early
postoperative period. Visual acuity in patients with SWS-associated
glaucoma has been reported to be 20/40 or better in two-thirds of patients
(260).
Several modalities have been described in the treatment of exudative
retinal detachment associated with diffuse choroidal hemangioma. Because
hemangioma and associated retinal detachment are more extensive in SWS,
treatment for diffuse choroidal hemangioma in this syndrome has not been as
successful as in the circumscribed variety. Xenon and argon laser
photocoagulations have been employed in treating these lesions although
achieving resolution of subretinal fluid may be challenging (267). Radiation
treatment in diffuse choroidal hemangioma has been used with some success
(18,267–269). In a series of five patients with SWS, treatment of diffuse
choroidal hemangioma with lens-sparing external beam radiotherapy was
examined. A total dose ranging from 1,200 to 4,000 cGy in fraction sizes
from 150 to 200 cGy was used (268). Radiation treatment resulted in
complete resolution of retinal detachment in all five cases. With a mean
follow-up of 40 months, two patients had significant improvement in visual
acuity, and the other three had no change. No recurrence of subretinal fluid or
cataract formation was reported during the follow-up period in any of the five
patients (268).

It is important to consult a pediatric radiation oncologist for patients with
diffuse choroidal hemangiomas. If external beam radiation is administered, a
tailored radiation plan should be developed to limit exposure to adjacent and
developing orbital structures (i.e., orbital bones).
Not applicable.
Future Treatments
Intravitreal VEGF antagonists have been used with visual and anatomic
improvements in a patient with SWS and exudative retinal detachment (270).
PDT has also been employed successfully to treat focal subretinal exudation
in patients with diffuse choroidal disease (271). Some authors have reported
successful therapy with oral propranolol 60 mg twice a day (272). Some
diffuse choroidal hemangiomas may not respond to oral beta-blocker therapy
(273). Most recently, low-dose brachytherapy has been employed with visual
and anatomic improvements (274).
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SECTION VII
Uveitis in Infants and Children
47
Uveitis and Endophthalmitis Affecting
Christopher D. Conrady, Stephen D. Anesi, C. Stephen Foster, and
Albert T. Vitale
INTRODUCTION
Pediatric uveitis encompasses a wide array of infectious and noninfectious
entities that may be isolated to the eye or the manifestation of an underlying
systemic condition. This subject requires special attention due to important
diagnostic and therapeutic challenges specific to this patient population. To
start, a comprehensive history and review of systems may not be possible in a
preverbal child and a complete ocular examination, including measurement of
intraocular pressure and indirect ophthalmoscopy, may require general
anesthesia in an uncooperative youngster. The differential diagnosis can vary
considerably with age, an important consideration given the
overrepresentation of infectious etiologies among younger children and the
presence of pediatric-specific neoplastic masquerade syndromes such as
retinoblastoma (RB) and acute leukemia. Certain endogenous noninfectious
entities such as juvenile idiopathic arthritis (JIA), Kawasaki disease,
tubulointerstitial nephritis and uveitis (TINU) syndrome, chronic infantile
neurologic cutaneous and articular/neonatal-onset multisystem inflammatory
disease (CINCA/NOMID) syndrome, and familial juvenile systemic
granulomatosis (Blau syndrome) are diseases found only in children and
young adults. The clinical presentation of other entities such as early-onset
sarcoidosis (EOS) differs from that seen in the adult while the choice and
interpretation of laboratory data must be modified to reflect differences in the
normative values of certain tests in children such as a physiologically
elevated angiotensin-converting enzyme (ACE).
Ocular inflammatory disease in children is typically chronic, recurrent,
and difficult to manage with the frequent development of ocular structural
complications. Presentation with established ocular pathology is itself not
only a risk factor for the development of further complications but also
graphic testimony to diagnostic delay and screening failure in this population.
To further complicate matters, ocular structural complications, such as the
development of a cataract in the pediatric population, require more prompt
attention than do those found in adults due to the risk of amblyopia in
children <10 years of age. These factors influence judgments with respect to
the aggressiveness of medical therapy and the timing of surgical intervention.
Therapeutic dilemmas in the medical management of noninfectious
pediatric uveitis include concerns regarding the promotion of cataract,
glaucoma, and growth retardation with the chronic use of corticosteroids
versus therapeutic timidity surrounding the early introduction of steroid-
sparing immunomodulatory therapy (IMT) for fears of potential toxicity
associated with these agents. Corticosteroid monotherapy and the late or
subtherapeutic introduction of IMT conspire to place children at greater risk
for visual loss from ocular structural complication both iatrogenically and
from uncontrolled inflammation. Children represent greater surgical risks due
to their well-known exuberant inflammatory response to surgical trauma as
well as the inherent complications associated with specific disease entities
such as JIA.
While the incidence and prevalence of childhood uveitis is significantly
less than that in adults, the proportion of children at risk for blindness and
severe visual loss and the impact of this visual debility over the life span of
the patient may be greater in children. Tolerance of low-grade intraocular
inflammation, prolonged use of corticosteroids, and reluctance to employ
steroid-sparing IMT when clinically indicated results in ongoing and
irreversible damage to ocular structures critical for visual function with
devastating effects for these young patients and their parents. Retina
specialists should be keenly aware of these issues in their approach to the
management of pediatric uveitides as accurate diagnosis, the timely and
appropriate use of antimicrobial agents in cases of infectious uveitis, and
early implementation of steroid-sparing IMT for chronic noninfectious
entities are critical for the maintenance and preservation of vision in this most
vulnerable population.
EPIDEMIOLOGY
Children account for anywhere from 2.2% to 21.65% of patients in various
uveitis clinics (1–5). A review of recent U.S. insurance claims found rates of
29 uveitis cases per 100,000 children (6). This is in contrast to much higher
prevalence rates in adults of 58 to 115.3 cases of uveitis per 100,000 adults in
the United States (7–9). Despite a lower prevalence and fewer children in
uveitis clinics compared to adults, the rate of vision loss and ocular
complications is higher in the pediatric population (10,11).
Large retrospective studies have identified a slight female predominance
and bilateral involvement in pediatric uveitis. Chronic or recurrent disease
represents up to 87% of cases, and there is a delay between diagnosis and
referral to a uveitis specialist that can range from <6 months to 14 years (5).
Anterior uveitis is the most frequent anatomical location, constituting nearly
half of all cases of pediatric uveitis (1,3–5,12–16). Intermediate uveitis
accounts for approximately another quarter of all disease, while panuveitis
and posterior uveitis are much less common, representing a quarter of disease
combined (3–5,13–17). JIA-associated uveitis is the most common type of
pediatric uveitis, and is easily the most frequent cause of anterior uveitis with
rates as high as 47% of all anterior uveitides (3–5,12–17). Pars planitis is the
most common form of intermediate uveitis, while toxoplasmic
retinochoroiditis is the most frequent etiology of posterior uveitis (17).
Systemic disease can be identified in 42% of cases with JIA being the most
common underlying etiology (5). Despite recent advances in molecular and
immunologic diagnostic tools, upward of 60% of pediatric patients’ disease is
considered idiopathic or undifferentiated (4,5,17).
There is significant geographic variation with respect to the anatomic
distribution, the prevalence of infectious etiologies, and so, the diagnosis of
uveitis among children. For example, toxoplasmosis is seen in only 2% to
10% of all cases of pediatric uveitis cases in Western industrialized countries,
but accounts for much higher rates in the Middle East and India with rates
approaching 35% to 40% of posterior uveitides in some reports from India
(13,17–20). Likewise, Behçet disease, while uncommon in the United States,
is more frequently encountered among patients along the ancient “Silk Road”
with Korean, Japanese, Chinese, Turkish, and Middle Eastern ancestry
(17,21–24). Finally, there has been a notable increase in panuveitis with
concurrent decrease in posterior uveitis over the last several decades. This is
likely due to the decrease in toxoplasmosis and toxocariasis in children
within developed countries (5). Table 47-1 illustrates the anatomic
distribution, association with systemic disease, infectious etiologies, and
geographic differences of pediatric uveitis (50).
TABLE 47-1 Epidemiology of pediatric uveitis

throughout the world
The number of patients included in each study is shown by in the sample size column, while the
affected intraocular locations and specific diagnoses are expressed in percentages
(1–5,10,12–17,25–49).
The differential diagnosis for pediatric uveitis can be divided into three broad
categories: infectious, noninfectious (autoimmune, autoinflammatory, or
idiopathic in origin), and masquerade syndromes. The most common
infectious causes of anterior uveitis are herpes viruses (HSV and VZV), and
toxoplasmosis is the most common cause of posterior infectious uveitis (51).
In one of the only large retrospective cohorts of its kind to evaluate cases of
infectious uveitides in children, an infectious etiology was identified in 17%
with T. gondii representing 60% of cases and viral infections another 30%
(51). The most common noninfectious cause of anterior uveitis in children is
JIA, which represents at least a third of all pediatric uveitis cases at tertiary
referral centers (25,52). There are also specific forms of uveitis much more
prevalent in the pediatric population such as toxocariasis, TINU syndrome,
Kawasaki disease, postviral, and diffuse unilateral subacute neuroretinitis
(DUSN). Intermediate uveitis may also represent a slightly larger portion of
cases in children than adults. This is supported by estimates of burden of
disease as high as one-fifth of all cases of pediatric uveitis being diagnosed
with intermediate uveitis and approximately 85% of all cases of pars planitis
occurring in children <14 years of age in one study (53). Masquerade
syndromes in children are also not uncommon and include primary
intraocular neoplasms such as RB and ocular involvement of systemic
leukemia (Figure 47-1, Table 47-2).
FIGURE 47-1 Masqueraders: Retinoblastoma. A: A child

with retinoblastoma and an associated total retinal
detachment with significant hyperreflective deposits
consistent with calcium on B-scan (B). Images courtesy of
Hakan Demirci, MD.
TABLE 47-2 Types of uveitis in children
CINCA/NOMID, chronic infantile neurologic cutaneous and articular/neonatal-onset multisystem

inflammatory disease syndrome.
The same anatomical categories of uveitis based on anatomic classification

(anterior, intermediate, posterior, panuveitis); grading of anterior chamber
cell, flare, and vitreous haze; and descriptors of the onset (sudden or
insidious), duration (limited or persistent), and clinical course of uveitis
(acute, chronic, recurrent) as defined by the Standardization of Uveitis
Nomenclature Working Group (SUN) for adults apply to ocular inflammatory
disease in children (54). A helpful approach to narrow the broad differential
of anterior uveitides is to subdivide the uveitis into nongranulomatous or
granulomatous disease (Table 47-3). This may serve to differentiate common
anterior nongranulomatous diseases such as JIA, herpetic diseases, and
human leukocyte antigen (HLA)-B27–associated iridocyclitis from
granulomatous uveitides such as sarcoidosis, inflammatory bowel disease,
syphilis, and Lyme disease (LD). Both noninfectious and infectious etiologies
are important considerations in the differential diagnosis of intermediate
uveitis (Table 47-3). The clinical hallmarks include anterior vitreal
inflammation, vitritis, peripheral retinal vasculitis, pars plana exudation, and
the not infrequent concurrence of moderate anterior cellular inflammation in
contrast to adults with intermediate uveitis in which there is usually only
minimal anterior chamber cellular reaction. Common structural complications
include optic nerve swelling and cystoid macular edema (CME). Posterior
uveitides can be further divided into those with or without retinal vasculitis
based on an extension of international uveitis study group classification
system (55). Retinal vasculitis may be observed in association with
toxoplasmosis, necrotizing herpetic vasculitis, sarcoidosis, syphilis, LD, and
systemic vasculitic syndromes. In contrast, toxocariasis, tuberculosis (TB),
Vogt-Koyanagi-Harada (VKH) syndrome, and presumed ocular
histoplasmosis syndrome (POHS) lack a prominent vasculitic component.
Several of these aforementioned diseases can also affect all anatomic regions
of the eye to cause a panuveitis (Table 47-3).
TABLE 47-3 Differential diagnosis of pediatric

uveitis based on most common anatomical location
affected
HIV, human immunodeficiency virus; VZV, varicella–zoster virus; HSV, herpes simplex virus.
As a complement to the SUN classification, it may be useful to think of

pediatric differential diagnoses in terms of the age of presentation (Table 47-
4). It is far more likely for an infant with intraocular inflammation to have a
congenital infectious syndrome such as rubella or a malignant masquerade
such as RB, whereas a child of 2 to 10 years of age would be more apt to
have an acquired infection such as toxocariasis, LD, or catscratch disease
(CSD) or an underlying systemic condition, such as JIA. Adolescents may
present with toxoplasmic retinochoroiditis, pars planitis, or even multifocal
chorioretinal disease, which is distinctly uncommon in younger age groups.
Finally, it must be borne in mind that many of these noninfectious and
infectious disease entities may present at any childhood age, including VKH
and sympathetic ophthalmia.
TABLE 47-4 Differential diagnosis of intermediate

and posterior pediatric uveitides based on age at
presentation
HIV, human immunodeficiency virus; VZV, varicella–zoster virus; HSV, herpes simplex virus.
DIAGNOSTIC APPROACH
A comprehensive medical history and review of systems, together with a
complete ocular examination, employing general anesthesia, as necessary,
form the cornerstones to the diagnostic approach in pediatric uveitis.
Laboratory tests and ancillary investigations are guided by the above and by
the putative differential diagnosis (Table 47-5). Laboratory tests are used in
most cases to confirm the clinical impression or to exclude diagnostic
possibilities, especially infectious entities rather than a “shotgun approach”
where false-negative and false-positive results can obfuscate the underlying
diagnosis. As such, disease-specific testing should be performed when there
is an elevated pretest probability for the disease based off of the history and
ocular examination or the risk of missing the diagnosis could lead to
permanent loss of vision or the eye altogether.
TABLE 47-5 Guidelines for further evaluation

depending on anatomic location
ACE, angiotensin converting enzyme; ANA, antinuclear antibody titers; CBC, complete blood count;
FTA-Abs, fluorescent treponemal antibody absorption test; LD, lyme disease antibody titers and
Western blot; PPD, purified protein derivative skin testing; RPR, rapid plasma reagin; UA, urinalysis.
COMPLICATIONS OF PEDIATRIC
UVEITIS
The risk for the development of ocular complications among the pediatric
population is greater than in adults with uveitis due to the chronicity of
inflammation, the frequent lack of symptoms and longer delays between
diagnosis and initiation of appropriate treatment. Ocular structural
complications include band keratopathy, secondary glaucoma, posterior
synechiae, cataract formation, vitreous hemorrhage, rubeosis, papillitis,
papilledema, CME, epiretinal/neovascular/cyclitic membranes, retinal
detachment (RD), hypotony, and phthisis. To complicate matters, ocular
complications associated with pediatric uveitis such as visually significant
media opacities, especially in very young children, can lead to the
development of amblyopia. This necessitates aggressive medical and surgical
treatment to prevent profound and permanent vision loss. The presence of
ocular complications on baseline examination is an important risk factor for
the development of subsequent complications (56,57).
The most common complications of pediatric uveitis are posterior
synechiae, cataract formation, CME, secondary glaucoma, band keratopathy,
and hypotony (4,5,15,16,58,59). Moreover, ocular hypertension affects a
significant minority of children with noninfectious uveitis and requires
vigilant monitoring particularly when strong risk factors, including local
corticosteroid use (in a dose- and route of administration–dependent
relationship) and contralateral IOP elevation, are present due to the risk of
developing a glaucomatous optic neuropathy (60). Complication rates have
been shown in studies from tertiary referral centers on pediatric uveitis to be
as high as 64% of children will develop cataracts, 58% posterior synechiae,
46% band keratopathy, and/or 25% glaucoma. (4,5,15,16). Furthermore, most
children will develop at least one complication from uveitis (4,5,15,16,25).
Children with JIA-associated iridocyclitis are particularly vulnerable as
compared to other forms of uveitis with the risk of developing any ocular
complication of 0.33 per eye-year and at least one ocular complication noted
in 67% of patients at time of diagnosis (4,5,56,59,61,62). Maculopathy
(including CME) and epiretinal and neovascular membranes are most
commonly found in association with intermediate uveitis, while papillitis and
optic nerve edema occur most frequently in posterior uveitis and panuveitis
(4,5).
Flare is also an important marker of anterior segment inflammation and
blood–aqueous breakdown. Its presence is thought to represent a dynamic
sign of disease activity. There are emerging imaging modalities currently
available that are being tested for their ability to objectively and accurately
quantitate the presence of flare. The most commonly employed method is
laser flare photometry (LFP). Measurements with this device have correlated
well with complications noted at baseline and over time and overall outcomes
in children with chronic anterior uveitis (63). High baseline and first flare
values also correlate well with risk of vision loss to <20/50 and vision-
threatening ocular complications independent of the presence of anterior
chamber cells (64). Anterior segment ocular coherence tomography (OCT)
has recently been shown to correlate well with LFP and clinical examination
in measuring anterior chamber flare (65). Consequently, the measurement of
aqueous flare is likely to become an important prognostic tool in uveitis. In
contrast, the presence of anterior chamber cell is indicative of active disease
and is more responsive to treatment, making it less helpful in defining long-
term prognosis.
VISUAL OUTCOMES
Not surprisingly, vision loss has been reported to be more common in
pediatric uveitis than in adults given the high rates of complications and
chronicity of disease observed in many childhood entities (66). In the 1960s,
30% of children with chronic uveitis became blind (26). More recent analyses
have estimated that between a quarter and a third of children with uveitis will
have severe vision loss (66). The most frequent causes of vision loss to
20/200 or worse include band keratopathy, cataract, vitreous opacities,
glaucoma, CME, and choroidal neovascular membranes and/or scars
(4,12,14). Interestingly, patients with posterior uveitis and panuveitis were
referred sooner to tertiary centers (11 and 7 months, respectively) than those
with intermediate (19 months) or anterior uveitis (33 months), likely due to
more vision-threatening disease (5). A more recent study suggests that cases
of posterior infectious uveitis have the worst visual prognosis, requiring
timely and accurate diagnosis, including the use of aqueous humor analysis
and prompt, directed therapy (11).
In cases of JIA-associated uveitis, the most extensively studied entity, a
worse prognosis has been historically associated with increasing severity of
disease. Ocular complications at initial evaluation, longer duration of uveitis,
younger age of presentation, male sex (56,67), and delayed referrals to a
uveitis specialist are all known risk factors of more severe disease (5,68).
Blindness from disease complications ranges from 0% to 17.5% and is much
more common in African Americans than non-Hispanic Caucasians (68–70).
However, visual outcomes have improved with early treatment with IMT
with 92% of patients having 20/40 or better vision 5 years after presentation
(17,52).
In our view, the currently guarded visual prognosis for pediatric uveitis is
potentially modifiable with early diagnosis and referral to a uveitis specialist
with appropriately aggressive anti-inflammatory and/or antimicrobial
therapy. Our experience and that of others suggests that the judicious
implementation of steroid-sparing IMT is essential in the treatment of chronic
noninfectious uveitis in children as this approach is effective both in
achieving inflammatory control and in reducing structural and steroid-
associated complications and so, improving visual outcomes (71,72).
Inflammatory quiescence for a minimum of 3 months prior to intraocular
surgery is recommended to improve outcomes and reduce the risk of
perioperative complications (61,73). Finally, the novel application of imaging
modalities such as OCT and laser-flare photometry to more quantitatively
assess inflammation and a reassessment of the current screening guidelines
may serve to more accurately identify those children at greatest risk of vision
loss and the development of ocular complications, thereby reducing the long-
term burden, socioeconomic, and psychosocial impact of these ocular
diseases (74).
SPECIFIC UVEITIC DISORDERS IN

CHILDREN
Noninfectious Uveitides
Juvenile Idiopathic Arthritis–Associated (JIA) Uveitis

JIA, a multisystemic autoimmune disease, is the most common form of
arthritis of childhood affecting approximately 70,000 children in the United
States and is the most common identifiable systemic association with
iridocyclitis, with an average cumulative incidence of approximately 8.3%,
representing 2,500 to 5,000 cases and up to 38% of those seen in a tertiary
care setting (75,76). Its pathogenesis appears to be multifactorial, as in most
autoimmune diseases, with an identified multigene genetic predisposition
possibly being triggered by microbial contact, with distinct mechanisms of
onset possibly differing between disease subtypes. The peak age of onset of
JIA is between ages 6 months and 4 years. The International League of
Associations for Rheumatology (ILAR) classifies JIA into the following
categories with the attendant associated risk of uveitis: systemic (rare);
oligoarticular, persistent, or extended (10% to 30%); rheumatoid factor (RF)–
negative polyarthritis (5% to 10%); RF-positive polyarthritis (rare, scleritis);
psoriatic (10% to 20%); and enthesitis related (10% to 15%, HLA-B27,
acute, recurrent, uniocular) (77,78). New possible classification criteria are
currently being studied by the Pediatric Rheumatology International Trials
Organization (PRINTO) (79).
As many as 12% of JIA patients may have uveitis, most frequently
anterior uveitis (83%), but with intermediate (9%), posterior (1%), and
panuveitis (7%) presenting as well (80). Disease is typically silent with the
patient being completely asymptomatic and unaware of active intraocular
inflammation. This is an extraordinarily curious aspect of the disorder, since
the vast majority of other individuals who develop uveitis experience pain,
ocular redness, light sensitivity, and/or diminished vision. Arthritis normally
precedes intraocular inflammation, but the converse may be true. As many as
10% may have chronic intraocular inflammation for prolonged periods of
time prior to entering into the health care system and resulting in delay in
diagnosis, treatment, and poorer outcome. In one study, between 3% and 7%
of patients may have uveitis onset occur up to 7 years prior to arthritis (81).
Furthermore, the systemic and ocular inflammatory activity may not be
synchronous, and their management should be considered independently.
Hence, recommendations by the American Academy of Pediatrics (AAP) for
screening children with JIA for uveitis were introduced in 2006 to ensure
proper monitoring of children with higher risk of developing ocular
inflammation, with the highest-risk group of ANA positive oligo- or
polyarthritis patients of disease onset earlier than age 7 and disease length of
4 years or less requiring ocular examinations every 3 months, and all other
groups every 6 to 12 months (Table 47-6) (82).
TABLE 47-6 International League of Associations

for Rheumatology (ILAR) classification of JIA and
associated risk of uveitis
a. Psoriasis or a history of psoriasis in the patient or first-degree relative.

b. Arthritis in an HLA-B27 positive male beginning after the sixth birthday.
c. Ankylosing spondylitis, enthesitis-related arthritis, sacroiliitis with inflammatory bowel disease,
reactive arthritis, or acute anterior uveitis, or a history of one of these disorders in a first-degree
relative.
d. The presence of IgM rheumatoid factor on at least two occasions at least 3 months apart.
e. The presence of systemic JIA in the patient.
* Petty RE, Southwood TR, Manners P, et al. International League of Associations for Rheumatology
classification of juvenile idiopathic arthritis: second revision, Edmonton, 2001. J Rheumatol
2004;31:390–392, Ref. (77).
† Angeles-Han ST, Yeh S, Vogler LB. Updates on the risk markers and outcomes of severe juvenile
idiopathic arthritis-associated uveitis. Int J Clin Rheumatol 2013; 8(1).
‡ Paroli MP, Speranza S, Marino M, et al. Prognosis of juvenile rheumatoid arthritis-associated uveitis.
Eur J Ophthalmol 2003;13:616–621.
HLA-B27, human leukocyte antigen B-27; JIA, juvenile idiopathic arthritis; RF, rheumatoid factor.
In addition to JIA subtype, factors increasing risk of development of uveitis

include ANA positivity, younger age at onset of arthritis (2 to 5 years),
female sex, positivity for multiple class I and II HLA genes including HLA-
DR5 (and possibly HLA-DR11, HLA-DRB1*1104, and HLA-DRB1*1301
though controversial), and moderate or high systemic disease activity.
Calandra et al., however, reported female sex not to be a risk factor in their
large cohort of 278 JIA uveitis patients (83). ANA, while a risk factor for
ocular involvement, does not correlate with disease activity. Predictors of
poor visual prognosis were associated with increased severity of ocular
disease at presentation and include active inflammation, presence of posterior
synechiae or hypotony, abnormal IOP, uveitis onset before or concurrent with
diagnosis of arthritis, shorter interval from onset of arthritis to uveitis,
prolonged referral to a uveitis specialist, prior intraocular surgery, and male
gender (54,61,62,72,83–95). Data from the large multicenter Systemic
Immunosuppressive Therapy for Eye Diseases (SITE) study showed
increased risk of vision loss to 20/50 or worse with use of oral corticosteroid,
and reduced risk of uveitis and vision loss with immunosuppressive therapy,
with the latter findings corroborated in other reports (72,85,96,97). HLA-
DRB1*0405 and HLA-DR1 have also been implied as protective against
uveitis (87,98).
Numerous biomarkers, in addition to serum ANA, have been implicated
as possible observable risk factors for onset of uveitis and as markers of
disease activity including elevated serum erythrocyte sedimentation rate
(ESR), Th1/Th2 lymphocyte ratio, and S100A8/A9 and S100A12 levels, the
latter of which was only significantly elevated in JIA with active uveitis and
inactive arthritis (86,99,100). Although not statistically significant, elevated
serum antibodies to the dense fine speckled 70 kDa antigen, also known as
“lens epithelium–derived growth factor” (LEDGF), were seen overall in
uveitis patients, and in a higher percentage of JIA patients with uveitis than
without uveitis (101). Decreased aqueous levels of interleukin-29 (IL-
29)/interferon-λ1 (IFNλ1) and increased aqueous levels of latency-associated
peptide, osteoprotegerin, and transthyretin have been seen in JIA patients
compared to non-JIA chronic anterior uveitis or control patients (102,103).
Other aqueous markers seen to be increased in JIA include transforming
growth factor beta-1, -2, and -3 (TGFβ-1, -2, -3); TNFα; serum amyloid A
(SAA); interleukin-8 (IL-8); and matrix metalloproteinase-2, -3, and -9
(MMP-2, -3, -9). Genetic analysis of various single nucleotide
polymorphisms (SNPs) suggests that the AA genotype of TRAF1-C5 may be
associated with increased risk of developing uveitis. Most recently, tear
analysis has shown a similar cytokine profile to aqueous humor in children
with anterior uveitis, with significant differences seen between patients with
JIA- and non–JIA-associated anterior uveitis (86,87,99–107).
While joint activity in JIA is driven by synovial influx of various cells
and cytokines activating a proinflammatory T-lymphocyte population, the
pathophysiology of JIA uveitis is still poorly understood. Histopathologic and
immunohistochemical studies performed directly on biopsied iris tissue of
JIA patients and indirectly via analysis of binding patterns of serum
autoantibodies of JIA patients to ocular cryosections of porcine eyes have
shown prominent plasma cell and macrophage infiltrate in the JIA specimens
compared to controls, and a higher rate of autoantibody binding, especially to
iris and ciliary body, in JIA patients with uveitis (94%) compared to JIA
patients without uveitis or idiopathic anterior uveitis patients (75% each) and
healthy controls (29%), with this increase correlating only with the presence
of ocular complications (108). Together these findings, while not contributing
to feasible screening methods, may help to more specifically direct future
therapy in this disease.
Ocular structural complications associated with visual loss include band
keratopathy, posterior synechiae, hypotony, glaucoma, macular edema, RD,
optic nerve edema, vitreous haze, and epiretinal membrane with an incidence
of any complication occurring at 33% per eye per year (61). Ocular
complications at presentation are common, being observed in up to 45% of
JIA children in a large population-based study and in 67% of 75 patients with
JIA followed over a 20-year period at the Wilmer Eye Institute (57,59). In the
latter study, complications at presentation were associated with active
inflammation, ANA positivity, and a shorter duration between the diagnosis
of arthritis and uveitis. Poor vision at presentation due to cataract, central
band-shaped keratopathy, or glaucoma to a level of 20/50 and 20/200 was
seen in 36% and 24% of patients respectively. The presence of active anterior
segment inflammation and a history of previous ocular surgery before
presentation were significantly associated with both levels of visual loss,
while the additional finding of posterior synechiae at presentation was
associated with 20/200 (legal blindness) (59). In other studies, additional risk
factors for the development of ocular complications have included uveitis
diagnosis before arthritis and African descent; however, it is unknown if the
latter is more attributable to biologic variance or other factors (109,110).
Ocular hypotony has been associated with a particularly poor prognosis and
is associated with longer total duration of uveitis, bilateral disease, and signs
of more severe disease such as posterior synechiae, presence of panuveitis,
anterior chamber cells or flare ≥3+, and use of oral corticosteroid (111,112).
The reported outcomes of JIA uveitis in the literature are variable. Kump
et al. (62), Chylack et al. (113), and Wolf et al. (114) reported poor outcomes
in their JIA populations over 30 years ago; however, Sherry et al. noted that
the prevalence of uveitis had been steadily decreasing as a consequence of
better JIA management by rheumatologists (115). Chalom et al. emphasized
that the outcome of their pediatric population of JIA who developed uveitis
was excellent 95% of the time in a 1997 publication (116). Kotaniemi’s
group reported ocular complications in 24% of their patients, but an overall
good outcome visually in 97% in a publication in 2001 and 100% in the latter
of 2 groups reported in another publication in 2014 (117,118). In stark
contrast, however; Cabral et al. emphasized that the occurrence of ocular
complications associated with JIA-associated uveitis was common (32%) and
vision impairment was not rare (15%) (119). While Tappeiner et al. reported
significant decreases in uveitis prevalence and secondary complications as
well as an increase in achievement of uveitis inactivity when comparing
cross-sectional data from a national German database over a decade, which
they attributed to more frequent use of IMT, the decrease in uveitis
prevalence was not large (13.0% to 11.6%) and prevalence of secondary
complications was still relatively common (33.6% to 23.9%) (120).
Regrettably, even today, despite adherence by ophthalmologists and
rheumatologists alike to the AAP recommendations for biomicroscopic
screening of children with JIA, the proportion of severe disease presentation
appears to be unchanged with as many as 25% developing significant visual
impairment and, in some studies, as many as 12% becoming blind in at least
one eye (Table 47-7) (122,123). This fact is underscored by Ozdal et al., who
reported that final visual acuity in their JIA population followed into
adulthood was impaired in 40%, quite poor in 20%, and completely lost in
10% of eyes (124). All 18 adult patients, with a mean duration of JIA of 24.9
years and uveitis for 20.5 years, had ocular complications at the time of first
evaluation, with visual acuity <20/150 in 30% of the patients and three eyes
in phthisis. The authors emphasized that chronic uveitis among patients with
JIA may persist into adulthood with attendant structural complications that
compromise visual function and adversely affect the long-term visual
outcomes resulting in an overall poor prognosis. Moreover, they argue that
the currently guarded outcomes may be improved only with earlier disease
detection and increased vigor of therapy by physicians skilled in the use of
immunomodulatory agents, and suggest that the more favorable ocular
prognosis reported in some publications in children with JIA-associated
uveitis suffer from case selection bias and/or inadequate longitudinal follow-
up.
TABLE 47-7 American Academy of Pediatric

recommendations of frequency of ophthalmic
examinations in patients with juvenile idiopathic
arthritis
Recommendations for follow-up continue through childhood and adolescence.

ANA, antinuclear antibodies; NA, not applicable.
Adapted from Cassidy J, Kivlin J, Lindsley C, et al. Ophthalmologic examinations in children with
juvenile rheumatoid arthritis. Pediatrics 2006;117:1843–1845 and Cassidy L, et al. Ophthalmic
complications of childhood medulloblastoma. Med Pediatr Oncol 2000;34(1):43–47, Ref. (121).
We support this opinion as far too many children with JIA-associated uveitis
lose vision to the point of being permanently visually handicapped, as a
consequence of delayed diagnosis, inadequate treatment including reliance on
or reluctance to taper off corticosteroid therapy, and prolonged duration of
disease. We reported on 77 adult JIA uveitis patients (135 eyes, mean age
29.72 years) at their last visit exhibiting the presence of at least one
complication in 72% of eyes and ongoing inflammation in 52% of patients,
with patients presenting for tertiary care older than age 16 having more
complications and worse vision (125). De Boer et al. showed a positive linear
correlation between macular thickening and duration of JIA uveitis follow-
up, with 13 of 61 patients developing macular edema after mean follow-up of
11.6 years, and eventual macular thinning after longer follow-up (126).
Additional risks of developing complications may exist as older children are
lost to follow-up during transition to adult care, with a preadult dropout rate
reported as high as 22% versus 3% for younger children and 10% for adults
(127). A study from the Bascom Palmer Eye Institute reported on 34 JIA
anterior uveitis patients among a series of 112 noninfectious pediatric uveitis
patients followed for up to 10 years, finding JIA patients with the highest risk
ratio for band keratopathy, cataract, and posterior synechiae, with the
cumulative proportion of patients with complications continuing to rise
through time, with substantial numbers of eyes and patients with visual acuity
of 20/200 or less in the better eye (4). As previously mentioned, the disparity
in the literature with respect to visual outcome in patients with JIA-associated
iridocyclitis, with more favorable outcomes reported among population-based
studies and surveys of JIA-associated iridocyclitis referred to pediatric
hospitals as opposed to poorer outcomes in our experience and those reported
from other tertiary care ophthalmic centers, may be more the reflection of
sampling bias and lack of long-term follow-up, obfuscating the harsh reality
of significant visual debility often associated with this disease
(4,27,62,75,124,125,128).
Our therapeutic approach to JIA uveitis is not unlike our approach to
other forms of noninfectious uveitis and includes the elimination of all active
inflammation with limited tolerance for steroids and the early introduction of
steroid-sparing IMT (129). The use of IMT in recalcitrant disease has been
associated with decreased risk of uveitis onset and frequency, vision loss,
ocular complications, and improved outcomes after surgery (72,97,130,131).
We recently reported a series of 30 JIA uveitis patients from 1990 to 2011
who achieved successful corticosteroid-free (no use of topical or systemic
corticosteroid) remission using IMT for 2 years followed by medication
withdrawal and maintenance of remission for at least 1 year in which 56.7%
remained in long-term remission thereafter, with a total population median
time of 7 years, with longer times significantly associated with patients
receiving IMT at an earlier age and earlier in the course of their disease
(132). Conversely, Acharya et al. reported a relapse rate of 68% in 19
patients (including 82% of 11 on tumor necrosis factor [TNF] inhibitors)
from 1988 to 2011 after achieving corticosteroid-sparing control of
inflammation (allowed use being prednisone ≤10 mg, prednisolone acetate
dose <3 drops/day) followed by withdrawal of IMT with a shorter median
time to relapse of 288 days, however; 69% of these patients were on topical
corticosteroids, with 17% on oral prednisone, and discontinuation of IMT in
some was due to reasons other than achieving steroid-sparing control of
inflammation (i.e., adverse reaction, poor efficacy, cost, pregnancy) (133).
Differences in outcomes between these reports likely reflect the notable
differences in length of use of IMT as well as allowed or acceptable amounts
of corticosteroid during steroid-sparing therapy.
Initial treatment of JIA-associated uveitis is commenced with topical
corticosteroids with regional corticosteroid injections (generally requiring
anesthesia) being added to the program should the uveitis not respond. We
use a so-called stepladder algorithmic paradigm in our care of patients with
JIA-associated uveitis, with brief (not >3 months) administration of systemic
corticosteroids representing the first step. Chronic oral nonsteroidal anti-
inflammatory drug (NSAID) therapy, commonly with naproxen or tolmetin,
represents step 2, and may be employed in instances in which uveitis of mild
to moderate severity recurs with every attempt at tapering topical steroids.
Some children with JIA may already be on NSAIDs for their arthritis. The
usual caveats pertain, of course, with respect to NSAID use, and therapy
should be coordinated with the patient’s pediatrician and/or pediatric
rheumatologist. Some patients with JIA-associated uveitis have recurrences
of the uveitis despite the chronic use of NSAIDs, or more severe disease that
would not be expected to respond to lower level anti-inflammatory therapy
(134). These patients must be advanced to the third step on the stepladder:
IMT.
A variety of immunomodulatory medications have been used in this
context (Table 47-8); however, the antimetabolite methotrexate has, by far,
the longest safety and efficacy record and is the agent of first choice in the
treatment of pediatric uveitis in general, and in JIA-associated iridocyclitis
specifically among treating physicians across multiple specialties (135–139).
We typically commence therapy at 0.15-mg/kg body weight once weekly by
mouth with 1 mg folic acid daily; weekly dosing reduces the risk of
hepatotoxicity (140) We advance the dose of methotrexate every 6 to 8
weeks, as needed and tolerated, to achieve the goal of complete freedom of
all recurrences of all inflammation at all times off all corticosteroids. Uveitis
associated with JIA often requires much higher doses of methotrexate to
induce durable remission than does arthritis. Fortuitously, children tolerate
such doses quite well, with no significant differences in side effects or ocular
complications seen between patients started on lower versus higher doses
(141). The bioavailability of the medication is variable after oral
administration; therefore, we switch patients to subcutaneous administration
at doses above 17.5 mg. Potential side effects include hepatotoxicity, bone
marrow suppression, interstitial pneumonitis, mucositis, nausea, fatigue, and
alopecia. For this reason, it is mandatory that patients be regularly monitored
by an expert in the prescribing and monitoring of IMT. We monitor patients
every 6 weeks, both by personal observation and query and by hematologic
assay of liver enzymes, renal function, and complete blood count (CBC) with
differential. Suppression of the bone marrow, in our experience, is
extraordinarily rare. Rising liver enzymes is an indication of hepatotoxicity,
and we typically back off on the dose if the patient has elevated enzymes on
two occasions separated by 2 weeks. Interstitial pneumonitis and chemical
hepatitis are reversible with cessation of drug if detected early in the course
of development.
TABLE 47-8 Commonly employed IMT for

noninfectious pediatric uveitis
ANC, absolute neutrophil count; BM, bone marrow; CD, cluster of differentiation; GI, gastrointestinal;
IL, interleukin; IMT, immunomodulatory therapy; kg, kilogram; m, meter; mg, milligram; mm,
millimeter; TB, tuberculosis; TNF, tumor necrosis factor.
We currently treat our JIA-associated uveitis patients on IMT for a minimum

of 2 years, after the uveitis has been in remission, off all steroids.
Inflammatory control has been observed in 60% to 82% of patients with a
steroid-sparing effect in >50% of patients (69,142–144). A recent study
confirmed our clinical impressions and practice in that inflammatory relapse
was observed in 69% of patients within 1 year of withdrawal of medication.
Both a longer period of inactivity prior to withdrawal and longer treatment
periods with MTX drug for up to 3 years are associated with a reduction of
the chance of relapse after drug discontinuation (142). This means, of course,
that 20% to 40% of patients do not succeed with methotrexate as
monotherapy. For them, alternative agents are explored, until the agent or
combination of agents is found that accomplishes the aforementioned goals
for them. Consensus recommendations specifically for childhood anterior
uveitis created by an expert panel of rheumatologists and ophthalmologists of
the Childhood Arthritis and Rheumatology Research Alliance (CARRA)
outline a similar approach intended for use in reducing interpractice
variability for future comparative analyses rather than as standard treatment
guidelines, but emphasize the now more widely accepted progression through
a “stepladder” approach to steroid-sparing treatment as needed to control
uveitis activity (138). Likewise, the Single Hub and Access point for
pediatric Rheumatology in Europe (SHARE) initiative aimed to optimize
diagnostic and therapeutic measures in children and young adults with
rheumatologic diseases, and presented their own guidelines specifically for
use in JIA uveitis (139). This trial-and-error exercise can be extremely
frustrating for patient, family, and doctor alike. However, to give up on the
quest for disease remission is, in our view, an enormous mistake. One must
stay in the hunt, never quitting on the patient and prevailing over the problem
in the end.
In those patients not responding to methotrexate, alternative
antimetabolites such as azathioprine (3 mg/kg/d) or mycophenolate mofetil
(MMF) (600 mg/m2/twice daily) may be substituted. Chang et al. reported on
52 children using MMF to control chronic uveitis showing at least short-term
efficacy in 73%, including 84% of 25 JIA patients (145). Cyclosporine (up to
5 mg/kg/d) may be used in combination with antimetabolites, but is not seen
to be very effective as monotherapy (146). Leflunomide is not often used in
uveitis therapy due to history of poor efficacy (147). Intravenous
immunoglobulin (2 g/kg/cycle, once monthly) has been successfully
employed in some patients in our experience. We have also used cytotoxic
agents such as chlorambucil (0.1 mg/kg/d) and intravenous
cyclophosphamide (1 g/m2 once monthly) in the treatment of recalcitrant
childhood uveitis, however, they are employed infrequently due to very real
concerns regarding their serious long-term side effects.
Biologic response modifiers, particularly the TNF inhibitors infliximab
and adalimumab, are the agents to which children with methotrexate-resistant
JIA-associated iridocyclitis are typically advanced (135,148). A systematic
review of anti-TNF agents in immune-mediated uveitis suggests that use of
these medications has increased given improved treatment outcomes and
excellent tolerance in the pediatric population, including 316 patients with
JIA uveitis (149). Improved postoperative outcomes in surgical patients using
anti-TNF agents have also been shown; however, disease relapse is often
seen after discontinuation (150). Typically reported adverse events include
infections, infusion or injection site reactions, and aminotransferase
elevation, and vary by study and medication as discussed below (151).
Etanercept, while effective in treating the articular manifestations of JIA,
has mixed results with respect to treatment of iridocyclitis, is questionably
associated with de novo inflammation, and has been mostly shown to be
ineffective both in preventing relapses and in the treatment of JIA-associated
iridocyclitis (137,152–155). Moreover, etanercept was the only one of three
TNF inhibitors (including infliximab and adalimumab) in a 10-year
longitudinal study to be discontinued due to new-onset uveitis (156).
Therefore, it is currently recommended that patients with new-onset or
incompletely controlled uveitis should not be started on etanercept, and those
who are on etanercept be switched to either infliximab or adalimumab
(128,139).
Infliximab has demonstrated consistent anti-inflammatory activity at
higher doses and shorter infusion intervals than that used for the treatment of
rheumatoid arthritis, allowing a reduction in both topical and systemic
corticosteroids and permitting systemic IMT taper (157–163). Our current
preferred regimen starts at 5 mg/kg monthly after induction, increased to 10
mg/kg or even to 20 mg/kg if needed and tolerated, with treatment intervals
extended as allowed with disease remission until 8 weeks, and then an
eventual trial of dose de-escalation. As this monoclonal antibody is a mouse–
human chimera, it is more immunogenic, and so, it has been recommended
that patients be maintained on low-dose antimetabolite therapy for the
prevention of the development of human antichimeric antibodies that are
associated with younger age of therapy initiation and much higher rates of
infusion reactions. Higher infliximab doses have actually been associated
with lower incidence in antibody formation, thought to be responsible for one
study finding uveitis as an indication for treatment being protective against
their formation (159,164). Unfortunately, infliximab does not consistently
produce disease remission with its efficacy waning over time (90), and, in
several comparative studies, has not proven as effective as adalimumab in
inducing steroid-sparing remission during continuation of therapy
(157,158,165).
Adalimumab is an effective and safe treatment option given after
methotrexate failure, as maintenance therapy following induction with
infliximab, or as initial monotherapy (160,166–171). Adalimumab is fully
humanized, and so, less immunogenic, and may be administered
subcutaneously rather than intravenously, ideally now in a formulation
without citrate buffers that have been associated with pain during injection.
We employ a standard dose of 20 or 40 mg given subcutaneously every 2
weeks, or weekly as needed, based on weight, disease severity, or medication
tolerance. Our review of 17 pediatric uveitis patients using adalimumab on
this regimen, with or without concomitant medications, including 14 JIA
uveitis patients, showed that 77% were able to achieve steroid-free
quiescence of disease at 1 year follow-up (166). Antiadalimumab antibodies
may form during therapy and have been associated with more severe uveitis,
active uveitis or loss of response to medication, and lack of concomitant
immunosuppressive therapy (172,173).
Adalimumab, in particular, has received considerable attention in recent
years with the awaited publication of two double-blind, randomized, placebo-
controlled trials studying the efficacy and safety of adalimumab in JIA-
associated uveitis patients failing methotrexate therapy: the SYCAMORE and
ADJUVITE trials (167,174). SYCAMORE studied patients age 2 years and
older with active uveitis comparing adalimumab to placebo in a time to
treatment failure protocol over 18 months in the UK. Treatment failure was
met by 18 of 30 (60%) patients on placebo versus 16 of 60 (27%) on
adalimumab, allowing the study to stop before enrollment goals had been met
due to adequate fulfillment of predetermined criteria for efficacy. Patients on
adalimumab also had a significantly longer mean duration of inactive disease
and no flares of arthritis, and there were no differences between health-
related quality of life scores; however, adverse and serious adverse events
were more common in adalimumab patients than placebo by a modest
margin, respectively (174). ADJUVITE looked at chronic active pediatric
uveitis, including JIA and idiopathic anterior uveitis, in children age 4 and
older comparing adalimumab to placebo over 2 months using LFP, and then
continuing both arms in an open-label adalimumab treatment trial for the
remainder of a year. Although no significant differences were seen in these
two groups in SUN-defined uveitis scores through month 2, LFP was seen to
improve to primary endpoint of 30% or better in 9 of 16 adalimumab patients
compared to 3 of 15 placebo. The authors reason this may show early
efficacy in treatment not discernable on slit lamp evaluation, even before
SUN criteria are met, however they caution the interpretation of these results
due to the small sample size (167). These trials were instrumental in
corroborating the expanded indication for the use of adalimumab as the first
anti-TNF medication for use in JIA-associated noninfectious pediatric uveitis
by the United States Federal Drug Administration (FDA) in 2018 (175).
The formation of anti-DNA antibodies may rarely produce a drug-
induced lupus syndrome with anti-TNF agents, but there are more significant
concerns surrounding long-term use of the agent including an increased risk
of infection including TB and histoplasmosis, hepatotoxicity, demyelination,
retrobulbar optic neuritis, heart failure, and a potentially increased risk of
lymphoma in children and young adults (176–181). This has led the FDA to
issue a “black box” warning for these medications with respect to malignancy
risk in this setting. Interestingly, Okihiro et al. reported on a cohort of 357
Canadian children with rheumatic diseases and long-term follow-up treated
with anti-TNF therapy over 16 years showing a low incidence of
malignancies (6/357 or 1.68%) with a 5-year malignancy-free probability in
JIA patients treated with biologic therapy of 97%; however, the few
malignancies seen were rare and unusual in the pediatric population (182).
Caution must be exercised in patients with intermediate uveitis or pars
planitis given the associated risk for the development of demyelinating
disease. TNF inhibitors are contraindicated in patients with symptoms or
magnetic resonance imaging (MRI) findings consistent with multiple
sclerosis (MS).
Several other biologic medications have been trialed in pediatric, and
specifically JIA-associated, uveitis. The TNF-inhibitor golimumab was
effective in limited case reports with or without concomitant methotrexate in
uveitis refractory to other biologic therapy (183–185). Retrospective data on
use of abatacept, an anti-CD28 fusion protein, on JIA uveitis have shown
some efficacy as well in recalcitrant disease, albeit less so than other more
commonly used biologic therapy and with eventual flare in several of these
patients shortly after achievement of remission (186–188). Two reports show
rituximab, with the standard cycle of 1,000 mg given twice at 2-week
intervals every 6 months, has been effective in treating JIA uveitis refractory
to IMT and anti-TNF agents, and we have unpublished experience of using a
more aggressive regimen with some success as well (189,190). Other groups
have reported on slightly larger cohorts employing tocilizumab in refractory
JIA uveitis demonstrating improvement in both inflammation and CME, as
well as complete resolution of the first documented JIA uveitis–associated
vasoproliferative mass and associated subretinal fluid, with better efficacy
being noted with 8 mg/kg monthly intravenous versus subcutaneous
administration (191–194).
The complications of JIA-associated iridocyclitis may require surgical
management. Cataract and glaucoma, in particular, occur in many patients
with this disorder, and the glaucoma that develops is frequently resistant to
adequate control medically. Many patients already have glaucoma at the time
of first referral. They generally also have cataract. Therefore, the decision-
making becomes complex. Should the cataract come out? Should the patient
have a lens implant at the time of cataract removal? Should a glaucoma
procedure be performed simultaneously? If the patient is judged to be a poor
risk for the presence of an intraocular foreign body such as a lens implant,
should contact lens wear or aphakic spectacles be planned? Should
trabeculectomy or valve tube shunt be employed for the glaucoma surgery
versus more traditional surgeries in childhood glaucoma such as goniotomy
or trabeculotomy? Should the surgery be performed soon or deferred until
better control of the uveitis can be accomplished? This latter question is
especially challenging in the child who still has the potential for avoidance of
amblyopia. However, of course, operating on the inflamed eye, particularly
performing cataract surgery, is notorious for poor outcomes. Therefore, the
urgency of prevailing over the active uveitis is even more critical.
Historically, the experience of most uveitis specialists was that children
with JIA-associated iridocyclitis are poor candidates for the incorporation of
a lens implant (195–197). The formation of inflammatory membranes behind
or around the implant, with membrane contraction and progressive hypotony,
has resulted in many such eyes being destroyed by phthisis. Some have
suggested that the best approach to this problem is cataract removal and total
vitrectomy with subsequent aphakic soft contact lens fitting; however, this
makes trabeculectomy for coexistent glaucoma a poor choice since the risk of
endophthalmitis in a contact lens wearer with a filtering bleb is unacceptable
(198–200). We favor the use of a valve tube shunt, generally placed through
the pars plana (201,202). Some studies suggest an increased risk of
postoperative IOP elevation, possibly requiring surgical intervention, with
primary lens placement at time of cataract surgery, despite good visual
outcome, with one study suggesting that secondary lens placement later may
decrease this risk (203,204). Lastly, the increasingly popular minimally
invasive glaucoma surgery (MIGS) devices now available are contraindicated
in uveitis and have not been well studied in children, and thus are not
recommended.
We think that in well-selected cases, with adequate control of
perioperative inflammation and proper follow-up, an intraocular lens (IOL)
can be well tolerated. Advances in surgical techniques and devices in cataract
surgery have popularized primary placement of an IOL in the pediatric
population. A multicenter analysis by Nemet et al. found no significant
difference in postoperative course or complications when comparing non–
JIA-associated uveitis patients and JIA (205). Lam et al. reported favorable
visual outcomes after cataract surgery with primary IOL implantation in
children with JIA-associated uveitis (206). We described improved
postoperative acuity in 92% of pediatric patients and found that placement of
an IOL did not significantly raise the level of postoperative inflammation
(207). Kulik et al. recently reported that 65% of eyes had BCVA of 20/40 or
better after cataract surgery with IOL placement with median follow-up time
of 10.9 years; however, 15% developed macular edema and 6% eventually
developed phthisis (208). Prudence absolutely dictates that one be very
thoughtful on this matter, counseling parents carefully, preparing them for the
possibility for the need for additional surgery for IOL explanation in the
event that their child demonstrates that the presence of the IOL is clearly
producing adverse consequences that threaten vision over the long term.
Implantable or injectional steroids have also been studied in the pediatric
population, including JIA uveitis, and tend to be used more often in cases of
intermediate or posterior inflammation. Similar to adult studies, the 0.7-mg
injectable dexamethasone implant use has shown efficacy in a variety of
pediatric uveitis types, allowing reduction or discontinuation of topical
corticosteroid therapy and IMT while improving vision and CME, and was
almost always associated with side effects or complications that were
transient and treatable medically, with only one patient requiring surgical
intervention (for high IOP) (209–211). The implantable 0.59-mg fluocinolone
acetonide implant is not often used in this population, but was shown to be
effective in 6 of 6 eyes in 4 patients with pediatric posterior uveitis, including
2 eyes in a single bilaterally phakic JIA patient; this patient eventually
underwent successful bilateral cataract surgery and valve implantation for
worsening cataract and steroid-induced ocular hypertension (212).
Seronegative Spondyloarthropathies
The juvenile seronegative spondyloarthropathies (JSpA) are a series of
related rheumatic diseases in children under age 16 characterized by
enthesitis (inflammation of tendons, particularly at their points of insertion
into bone) and involvement of peripheral large joints and with a
predisposition to sacroiliitis, uveitis, and the HLA-B27 gene. By definition
(“seronegative”), the patients are RF negative. Classification has varied over
the last few decades with the development of various criteria, with
considerable overlap found between these disorders and some forms of JIA,
given their similarities, and includes ankylosing spondylitis (AS), reactive
arthritis, psoriatic arthritis, and inflammatory bowel disease. These are very
similar to the adult variants; however, differences exist with respect to
symptoms at disease onset and severity through course of disease. Juvenile
psoriatic arthritis, by ILAR criteria, is classified as a subtype of JIA; however
it is, perhaps more so related to a form of JSpA. MRI has increasingly been
thought to be more sensitive than radiography in detecting early
inflammation, including prior to axial involvement, and may show early
characteristic findings that may differ by disease subtype (213). For the
ophthalmologist, the symptomatic presentation of acute anterior uveitis
(AAU) seems to be more often found in patients with the juvenile
spondyloarthropathies than JIA and may help to distinguish these patients in
order to better predict disease course.
Ankylosing spondylitis
Pediatric AS is a well-characterized syndrome. As in adults, HLA-B27–
positive males predominate in the pediatric population. Conversely, unlike
the adult form, in which initial onset is typically arthralgia with morning,
lower back stiffness and axial involvement, enthesitis with exquisite
tenderness in the areas of tendon inflammation, and peripheral arthritis are
more prominent early in the children with AS and usually involve knees,
shoulders, hips, or even ankles. As with JIA, the temporomandibular joint
(TMJ) may be involved and may be the single peripheral joint involved. AS
in children typically begins after age 10. Anterior uveitis associated with AS
as with all of the seronegative spondyloarthropathies is generally acute,
unilateral, and recurrent but may become bilateral and alternating. As many
as 15% of children with juvenile AS may develop chronic iridocyclitis (214).
HLA-B27 positivity is not required for the diagnosis; 5% to 10% of patients
with AS are HLA-B27 negative. Unlike JIA uveitis, the uveitis associated
with JSpA is often symptomatic and presents with a red eye, ocular
discomfort, and photophobia and may become severe quite rapidly or in a
delayed manner 1 to 2 weeks after initial symptoms despite initiation of
topical therapy. Therapy should include aggressive topical corticosteroids
often administered every 30 to 60 minutes while awake along with
cycloplegia; however; in the presence of severe inflammation, indicated by
the presence of fibrin in the anterior chamber on slit lamp evaluation or when
topical therapy does not result in a significant reduction in the intensity of
anterior chamber reaction, a short burst of systemic corticosteroid (1.0
mg/kg/d) with a rapid taper, such as a dosage reduction every 5 to 7 days, and
aggressive cycloplegia is appropriate. More recalcitrant cases may require
regional corticosteroid injections, and this generally requires brief general
anesthesia in children.
HLA-B27–associated uveitis tends to be recurrent, and our experience
suggests that oral NSAIDs, especially celecoxib and diflunisal, are
particularly useful in long-term management of patients with
nongranulomatous, acute idiopathic, or HLA-B27–associated recurrent
anterior uveitis, as well as with concurrent active systemic manifestations,
allowing a reduction in the amount of corticosteroid required to achieve
inflammatory quiescence and often enabling patients to maintain quiescence
once steroids have been discontinued (215). One should be guided by the
child’s rheumatologist as to the choice and dosing of the NSAID.
Antimetabolites may be used if the NSAID is not able to control disease.
TNF-α has been shown to be involved in the pathophysiology of both adult
and juvenile AS; thus, TNF-inhibitors have been widely studied in the
treatment of systemic disease and have been effective in one double-blind
study versus placebo; however, patients are not often able to discontinue
therapy without relapse (216–218).
Reactive arthritis
Reactive arthritis (previously “Reiter syndrome”) was originally defined as
the triad of arthritis, conjunctivitis, and noninfectious urethritis, but uveitis
may substitute for or be present along with conjunctivitis. It may rarely occur
in children, and may present after infections, such as Shigella, Salmonella,
Yersinia, Clostridium difficile, or streptococcal pharyngitis, the last of which
is differentiated in part from acute rheumatic fever by duration of arthritis and
response to therapy (219). Whereas 75% of adults who develop reactive
arthritis are HLA-B27 positive, 90% of children with it are HLA-B27
positive, and 2% of pediatric patients with reactive arthritis develop uveitis.
The intraocular inflammation is identical to that seen with AS or HLA-B27–
associated uveitis without spondyloarthropathy and is generally anterior,
nongranulomatous, and recurrent with the initial attack being typically
unilateral and acute. Hypopyon uveitis may also occur. Therapy is similar to
that described above for AS-associated intraocular inflammation.
Psoriasis
Children may develop uveitis in association with psoriasis, with or without
the development of arthritis. Stoll et al. first reported that disease onset is
bimodal, which was later corroborated by the CARRA cohort that showed an
average early onset near age 3 and late onset at approximately age 11.
However, these groups differed on whether uveitis onset was more common
in the early group or similar between the two (220,221). Girls are slightly
more likely than boys to develop psoriasis (3:2) with a mean age of onset of
approximately 9 years. The uveitis associated with psoriasis without arthritis
is slightly different from that of uveitis associated with psoriatic arthritis or
HLA-B27–associated uveitis (222). The mean age of onset is older, and it
tends to be bilateral and of longer duration and requires oral treatment with
NSAIDs. Retinal vasculitis, macular edema, and papillitis are more
frequently seen.
The skin lesions of psoriasis typically precede the development of either
joint or ocular inflammation; however, we have made the diagnosis of
psoriasis in numerous instances while examining the skin of patients during
their initial evaluation of uveitis, which was confirmed by subsequent
dermatologic referral. Small hidden patches of itchy, scaly dermatitis may
come and go, in the axilla, on the umbilicus, on the genitalia, on the back, or
on the scalp. Dactylitis and nail pitting may also be present. The prevalence
of uveitis in patients with psoriasis is unknown and may occur in up to 20%
in pediatric psoriatic arthritis (223,224). As with AS and reactive arthritis,
psoriasis-associated uveitis presents with acute, anterior, recurrent,
nongranulomatous, unilateral inflammation, but with time can develop into
bilateral involvement. The uveitis is more severe and more recalcitrant in
patients who are HLA-B27 positive. Patients who develop uveitis associated
with psoriatic arthritis are also more commonly ANA positive (80%) with
oligoarticular arthritis presenting as long as 10 years before the onset of the
dermatologic manifestations of psoriasis; hence, the diagnosis of seronegative
oligoarticular JIA has often been (erroneously) made earlier in these children.
Therapy is with aggressive topical corticosteroids and cycloplegics.
Chronic oral NSAID therapy may also be helpful in reducing the likelihood
of recurrent inflammation. Treatment-resistant cases may respond to low-
dose once-weekly methotrexate or daily cyclosporine. A recent small case
series of 6 patients suggested that younger patients (age 6 or less) may suffer
more severe and chronic disease requiring TNF-α inhibition to control
disease (225).
Inflammatory bowel disease (IBD) (enteropathic) arthritis

Enteropathic arthritis–associated uveitis may occur in 10% of patients with
Crohn disease (CD), ulcerative colitis (UC), or inflammatory bowel disease
unclassified. These disorders are associated with peripheral arthritis (20% in
CD and 10% in UC), sacroiliitis (10% in either CD or UC), and the HLA-
B27 haplotype (46% in IBD) (226). Uveitis occurs more often in patients
with CD than UC (223). Active disease has been found in as many as 23.1%
of asymptomatic patients during screening, again more frequently in CD than
UC (227). Uveitis more often presents years after IBD but may precede
intestinal symptoms, and age does not appear to affect risk of onset
(228,229). Episcleritis may be seen with anterior uveitis. In most cases, the
clinical presentation is similar to the other seronegative spondyloarthropathy-
associated uveitides as is its treatment. Although much less frequent,
posterior uveitis may also occur and is characterized by granulomatous
panuveitis with choroidal infiltrates, occlusive retinal vasculitis, serous RD,
retrobulbar neuritis, and papillitis (230–232). Surgical excision of the
inflamed bowel may be beneficial in management of the extraintestinal
symptoms, including peripheral arthritis and uveitis. IMT (especially with 6-
mercaptopurine) and anticytokine therapy (especially with anti-TNF agents)
may be critical agents in the eventual successful control of the patient’s
intestinal and extraintestinal disease.
Tubulointerstitial Nephritis and Uveitis

Uveitis may occur in patients with tubulointerstitial nephritis, an association
initially described by Dobrin et al. It is classically thought to be a relatively
rare form of uveitis; however, several groups worldwide have reported a few
hundred cases of this condition, suggesting that it may be more common than
initially reported, possibly attributable to a paucity of symptoms at
presentation or poor screening for renal and/or uveal inflammation (233). In
fact, a prospective multicenter study in Finland showed that 8 of 16 children
presented with asymptomatic uveitis (234). Most reports show disease
predominantly occurs in adolescent females in the early to mid-teenage years
but may occur frequently in women up to their early 30s. Uveitis is typically
simultaneous and bilateral 75% of the time and anterior in 80% of cases.
Although there is no racial or geographic predilection, we and others have
identified strong HLA associations with the development of this syndrome
including HLA-DR14, HLA-DQB1*0503, and HLA-DRB1*0102 (235–238).
Both HLA-DQA1*0104 and HLA-DRB1*14 associations were found
independently among cohorts in China and Finland (235,237,239). Although
the exact pathogenesis is unknown, Tan et al. showed that antibodies against
modified C-reactive protein (mCRP), shown to be associated with intensity of
inflammation in lupus nephritis, are also distinctly elevated in TINU patients,
and that mCRP was co-localized with IgG in both tubular epithelial and uveal
cells in TINU as compared to controls, suggesting that mCRP may be a target
autoantigen in TINU (240).
Tubulointerstitial nephritis typically develops acutely beginning with
fever and malaise, followed by flank tenderness and pyuria. Proteinuria,
hematuria, rising blood urea nitrogen (BUN) and creatinine, and, on biopsy,
edema and inflammatory cells in the renal interstitium are the hallmarks of
this usually self-limited disorder. The nephritis generally resolves without
permanent renal deficit, either spontaneously or following systemic
corticosteroid therapy, but may progress resulting in severe compromise in
renal function requiring dialysis or transplantation, underscoring the
importance of comanagement with a pediatric nephrologist. Uveitis may
present before the development of systemic symptoms; however, it more
commonly presents as acute bilateral anterior iritis following the onset of
nephritis with redness, variable pain, blurred vision, and photophobia. In
some cases, the course may be recurrent or chronic and evolve into an
intermediate, posterior, or panuveitis. Reddy et al. reported 6 of 6 biopsy-
confirmed TINU children had panuveitis and suggested this uveitis anatomic
subtype may be an underappreciated presentation of this disease (241). In
these cases, uveitis may be nongranulomatous or granulomatous and may
include the presence of frank retinal vasculitis, with intraretinal hemorrhages
and exudates. Younger patients (<20 years) were more likely to be chronic
with complications reported in 21% including macular edema, macular
pucker, and chorioretinal scarring. Systemic corticosteroid therapy–
associated cataract and glaucoma, along with iatrogenic Cushing syndrome,
were also described in these patients (241).
Mandeville et al. proposed diagnostic criteria for TINU, categorizing
cases as “definite” if interstitial nephritis had been diagnosed by biopsy, as
“probable” if the nephritis had been diagnosed clinically, and as “possible” if
the nephritis had been diagnosed clinically and the uveitis is “atypical” (242).
A clinical diagnosis of TINU may be made by fulfilling the following
criteria: abnormal serum creatinine or decreased creatinine clearance;
abnormal findings on urinalysis (UA) (elevated β2-microglobulin,
proteinuria, eosinophiluria, pyuria or hematuria, white cell casts,
normoglycemic glycosuria); and associated systemic findings (fever,
anorexia, weight loss, arthralgias, myalgias, abnormal liver function,
eosinophilia, and elevated ESR) (243). In a prospective study by Hettinga et
al., elevated β2-microglobulin combined with increased serum creatinine had
a positive predictive value of 100% (n = 8) for detecting definite or probable
TINU, although only two of these were biopsy proven (244). The importance
of renal biopsy, when feasible, cannot be overstated as illustrated by a case
reported by Pepple et al. in which a 9-year-old boy with bilateral anterior
uveitis with elevated creatinine and significantly elevated β2-microglobulin
underwent biopsy that showed results consistent with sarcoidosis rather than
TINU (245). Serial renal biopsies have shown chronic active inflammation up
to 2 years after initial presentation and biopsy, suggesting these patients may
require long-term anti-inflammatory therapy with IMT (246).
In most cases, topical corticosteroids are adequate in controlling the
uveitis, but some patients require systemic corticosteroid therapy. In cases of
steroid-resistant, recurrent, or chronic inflammation, immunomodulatory
agents can provide lasting control and prevent relapse of uveitis attacks as
well as nephropathy. Immunomodulatory agents and biologic therapy
(methotrexate, azathioprine, cyclosporine, MMF, and adalimumab) have been
used in the care of patients with TINU who were inadequately responsive to
steroids or who were developing steroid- or IMT-associated toxicity
(236,247).
Sarcoidosis
Sarcoidosis is a chronic multisystem noncaseating granulomatous disease of
unknown cause that occurs rarely in children. There are two presentations in
children: EOS, <4 years of age, and late onset, between 8 and 15 years of age
(248,249). EOS patients are primarily Caucasian, who present most
commonly with the triad of arthritis, skin lesions, and uveitis. Pulmonary
involvement is less common (35%), making it more difficult to establish the
diagnosis, especially since articular involvement may mimic JIA (250,251).
In contrast, older children and adolescents usually present with multisystem
disease similar to that seen in adults with sarcoidosis. There is generally no
familial history of sarcoidosis, and bilateral hilar lymphadenopathy or
pulmonary involvement may be present on chest radiographs in 90% of
cases. Eye and skin lesions occur in 30% of late-onset juvenile sarcoidosis.
Since other diseases, including mycobacterial or fungal infection and
berylliosis, as well as foreign body reaction, can also produce noncaseating
granulomas, the histologic diagnosis of sarcoidosis is made by exclusion.
Vasculitis is a relatively unique complication associated with juvenile
sarcoidosis.
Blau syndrome, also known as familial juvenile systemic granulomatosis
and/or Jabs syndrome, is an autosomal dominant granulomatous disease of
childhood with clinical features almost identical to EOS. Patients with Blau
syndrome have mutations in the NOD2 gene (CARD15) on chromosome 16.
Renal and hepatic granulomas have been described in this disease (252–255).
Granulomas of the skin and synovial biopsy from Blau syndrome are
identical to those seen in sarcoidosis (255).
Sarcoidosis is rare in children, but a patient as young as 3 months old has
been reported (256). The annual incidence has been estimated at two to three
cases per million with a higher prevalence in females; however, the majority
of reported pediatric cases are similar between genders (257,258). In the
southeastern United States, sarcoidosis has a higher incidence among African
Americans (259,260). In children age 4 years and younger with sarcoidosis,
7% to 28% were African Americans, whereas among children aged 8 to 15
years, the percentage of African Americans increases to 72% to 81%
(248,261,262). Within the United States, approximately 80% of childhood
cases have been reported in the southeastern and south central states,
suggesting they are an endemic area for childhood sarcoidosis
(259,261,263–265). The disorder is common in Scandinavian countries but
rare in India, Southeast Asia, New Zealand, and China (264).
As mentioned above, EOS is characterized by the triad of skin eruptions,
arthritis, and uveitis without bilateral hilar lymphadenopathy, which may
mimic JIA. Intermittent fever and synovial swelling may further contribute to
this masquerade. Early differentiation of sarcoidosis from JIA is important in
planning treatment strategies and in counseling patients and families with
skin changes serving to help differentiate these diseases at their onset. The
rash seen in JIA is transient and composed of pink macules, whereas that
observed in sarcoidosis presents with variable erythema, papules, plaques,
and ichthyosiform lesions (266). The cutaneous manifestations may be the
earliest sign and occurs before joint or eye involvement with papules,
plaques, nodules, erythema nodosum, and hypo- or hyperpigmented areas.
Lupus pernio is frequent in adults but rare in children (267,268). A skin
biopsy should be performed if lesions are present given their accessibility in
an effort to confirm the diagnosis. Sarcoid arthritis in children is
characterized by painless, boggy synovial and tendon sheath effusions with
mild limitation of motion, whereas in JIA, there is pain, limitation of
movement, and destructive changes found on radiographs. Parotid gland
enlargement is a frequent finding in children with sarcoidosis, especially
EOS, and represents an additional easily accessible biopsy site.
Late-onset juvenile sarcoidosis presents similarly to that seen in adults—
a multisystem disease with lymphadenopathy, hepatosplenomegaly, parotid
fullness, and pulmonary involvement as well as generalized constitutional
signs and symptoms (fever, anorexia, and malaise) (267,268).
A wide spectrum of systemic vasculitis in pediatric sarcoidosis has been
reported, including leukocytoclastic vasculitis, vasculitis of small- to
medium-sized vessels, and large-vessel vasculitis. EOS patients should be
carefully followed for development of vasculitis.
Neurologic dysfunction secondary to sarcoidosis is rare in children.
Granulomas are most common in the basal area of the meninges and brain,
causing seventh nerve palsy and hydrocephalus (269). Growth deficiency has
been reported in association with brain MRI abnormalities, and hypothalamic
infiltration can manifest as diabetes insipidus (270,271).
Hypercalcemia may be present due to increased 1-α hydroxylase activity
and interferon (IFN)-γ production, both leading to elevated levels of active
1,25-vitamin D and increased gut absorption of calcium (272,273). Renal
involvement may be related to the presence of granuloma in the renal
parenchyma or to hypercalciuria with nephrocalcinosis (270,274–279).
Granulomatous tubulointerstitial nephritis may present with uveitis and
appear as TINU, with biopsy being required to differentiate it from
sarcoidosis (245). Sarcoid liver granuloma may be present in up to 90% of
patients, and liver enzymes are often mildly elevated in such instances. A
needle biopsy may identify the lesions (280).
The ocular manifestations of sarcoidosis in children are similar to those
seen in adults and were the second most common initial presenting findings
in a study in Louisiana (263). Anterior uveitis is the most common ocular
manifestation in both the younger (81%) and older pediatric groups (21% to
48%) (263). The inflammation may be chronic and granulomatous with
minimal pain and photophobia. Multiple areas of interstitial keratitis (IK),
corneal limbal nodules, mutton fat keratic precipitates, and iris nodules may
be observed (Figure 47-2) (274,275). An acute, nongranulomatous type of
uveitis can also occur, typically characterized by pain, redness, and
photophobia. Posterior segment involvement is the most vision threatening,
manifesting with vitritis, pars planitis, papillitis, chorioretinal granulomas,
macular edema, branch retinal vein occlusion, retinal periphlebitis, subretinal
neovascular membranes, and optic nerve granulomas. Lacrimal gland
involvement, commonly found in adult patients with sarcoidosis, rarely
occurs in children (281).
FIGURE 47-2 Iris nodules in pediatric sarcoidosis.
ACE, produced by macrophages and epithelioid cells, reflects the total body
granuloma load. ACE levels are commonly higher in children than in adults,
and ACE is not specific for sarcoidosis, because it is elevated in several other
systemic diseases that affect the lungs or the liver. When ACE levels are
compared to age-matched controls, 80% of children with pediatric
sarcoidosis have elevated ACE (282). Gundlach et al. reported that elevated
levels of soluble interleukin-2 receptor (sIL2R), also nonspecific and known
to be released activated T lymphocytes, may be a useful screening tool in
newly diagnosed adult uveitis. sIL2R has higher sensitivity (98%) compared
to ACE (22%) or chest radiography (50%) in known sarcoidosis; there is
comparable high specificity among all three tests; however, no pediatric
patients were included (283). Hilar adenopathy may be seen with or without
parenchymal involvement. Gallium scanning and thin-cut spiral CT scanning
are more sensitive imaging methods to detect early lung involvement.
Contrast-enhanced MRI with fast imaging sequencing has been found to be
comparable to CT in evaluating pulmonary and cardiac lesions without the
same risk of radiation to children as CT and has been useful in establishing
the diagnosis of sarcoidosis in children who present with fever of unknown
origin by revealing multifocal nodular lesions in the bone marrow of the
lower extremities (284,285). Fine needle aspiration biopsy cytology has been
suggested as an adjunct in the diagnosis of children with suspected
sarcoidosis with the demonstration of epithelioid histiocytes and
multinucleated foreign-body–type giant cells without accompanying necrosis
or acute inflammation (280). Ideally, biopsy of easily accessible sites, such as
skin, lymph nodes, and conjunctiva, especially when conjunctival nodules are
present, is highly recommended in this diagnostic effort (286). As in adults,
sarcoidosis in children may act as a “great imitator,” appearing similar to
other entities such as JIA uveitis or even early-onset birdshot
retinochoroidopathy, but where hilar node biopsy may confirm the diagnosis
(287). Likewise, sarcoid may be suspected in cases where another etiology is
the masquerade, as in a case of a 6-year-old girl with nodular anterior uveitis
with corneal nodules responding to “anti-inflammation treatment,” which,
after aqueous analysis, turned out to be diffuse infiltrating anterior RB (288).
The course of sarcoidosis is unpredictable with the prognosis of
childhood sarcoidosis not significantly different from that in adults. The
disease is characterized by either progressive chronicity or periods of activity
interspersed with remission, sometimes permanent and spontaneous, or
initiated by steroid therapy. Overall, most affected patients recover with
minimal or no residual manifestations, whereas a small portion have
permanent loss of some pulmonary function or permanent visual impairment.
Some patients die of cardiac or central nervous system (CNS) damage.
The therapy of choice for sarcoidosis in children with multisystem
involvement is oral corticosteroids, which produce rapid symptomatic
improvement but may not affect the long-term prognosis. Corticosteroid
treatment should be instituted in the presence of respiratory symptoms or in
the presence of severe impairment of pulmonary function tests and are the
primary treatment for ocular manifestations of this disease. Methotrexate is a
safe and effective steroid-sparing strategy in children with severe chronic
uveitis who are dependent on but not well controlled by or develop
unacceptable side effects from systemic corticosteroid monotherapy
(289,290).
Other immunomodulatory medications used successfully in the care of
both adults and children with sarcoidosis include azathioprine, cyclosporine,
cyclophosphamide, and the TNF inhibitors infliximab and adalimumab
(145,166,291–295). Some debate exists as to the efficacy of anti-interleukin-
1β therapy with anakinra for Blau syndrome, and studies for EOS or late-
onset pediatric sarcoid uveitis have not been reported to date (296,297).
Familial Juvenile Systemic Granulomatosis (Blau

Syndrome, Jabs Syndrome)
Blau syndrome, or familial juvenile systemic granulomatosis (also known as
Jabs syndrome), is a rare, autosomal dominant disorder characterized by
early-onset granulomatous arthritis, uveitis, and skin rashes. Camptodactyly
(flexion deformity at the proximal interphalangeal joint) may occur. It has
variable expressivity and is known to appear in children younger than 4 years
of age. Ocular manifestations also include subconjunctival nodules, cataract,
glaucoma, and RD (252,298). The susceptibility locus has been mapped to
chromosome 16p12–q21, and a large number of CARD15/NOD2 mutations
have been reported (299,300).
EOS and Blau syndrome may represent the sporadic and familial forms
of the same disease and are often described together in the literature, since
both entities share genetic mutations in NOD2 (nucleotide-binding
oligomerization domain 2), also referred to as capsule recruitment domain
family member 15 (CARD15), in 50% to 90% of cases (301). A recent
review by Caso et al. discusses possible mechanisms by which changes in
NOD2 function might lead to a phenotypic presentation of granulomatous
disease and catalogs several reports of specific mutations (many shared)
found in both Blau and EOS over a decade (302). The term “pediatric
granulomatous arthritis” has been used to describe both disorders, although
arthritis is not a universal finding (303). Clinical resemblance between Blau
syndrome and EOS suggests genetic homogeneity. A study by Rybicki et al.
found no linkage between sarcoidosis and the Blau syndrome locus; however,
this might be explained as the group only studied cases of late-onset
sarcoidosis (304). Blau syndrome does not have pulmonary involvement,
which is also uncommon in EOS.
Arthritis is the most common manifestation of the disease, classically
described as boggy tenosynovitis, usually manifested in the first decade of
life as nonerosive cystic swelling of the wrists, fingers, ankles, and elbows
with preserved range of motion. The skin lesions are intermittent, red, and
papular rashes (252,298). An international prospective study of 50 patients
showed ocular involvement in over 80% with median age onset of 5 years
(305). Uveitis is classified as typically bilateral granulomatous panuveitis,
phenotypically similar to sarcoidosis, but anterior uveitis and intermediate
uveitis, as well as multifocal choroiditis and disk changes (pallor and
peripapillary nodules), have been described in children. Secondary
complications of cataract, glaucoma, and macular edema are also seen.
Vision loss may progress despite aggressive therapy (305,306). Common
laboratory abnormalities associated with the syndrome include
hypercalcemia, hypercalciuria, elevated ACE level, elevated ESR, elevated
immunoglobulins, leukopenia, eosinophilia, hematuria, proteinuria, pyuria,
and abnormal liver function tests (260,282). RF is often positive, a finding
not typical of patients with JIA. IL-1β levels are variably elevated, including
in some patients felt to be have more severe disease. Histopathology shows
ill-defined, superficial, dermal granulomatous inflammation. The granulomas
contain dense lymphocytic coronas, epithelioid cells, occasional Langerhans-
type multinucleated giant cells, and a few lymphoid cells, without evidence
of necrosis. Perivascular lymphomononuclear infiltration around dermal
vasculature is also typical. Immunofluorescence studies have shown high
expression of cytokines such as IL-6, TGF-β, IFN-γ, and IL-17 (307).
The initial presentation of joint manifestations in the absence of ocular or
skin findings is frequently mistaken for JIA. However, Blau syndrome has
milder systemic symptoms and characteristic joint abnormalities not seen in
JIA. The previous section on sarcoidosis also describes differences in
presenting skin lesions of EOS and JIA, which one can apply to Blau
syndrome during workup. The diagnosis is supported by the demonstration of
noncaseating granulomas within skin, synovial or conjunctival biopsies, and
the presence of CARD15/NOD2 mutations with familial history of disease
(253).
Therapy for Blau syndrome is similar to that of EOS with progression
from topical and systemic corticosteroid for acute inflammation, to steroid-
sparing therapy for chronic disease. Agents used with reported success
include NSAIDs and IMT with various agents including methotrexate,
cyclosporine, MMF, adalimumab, infliximab, as well as tocilizumab
(305,308). Some reports describing the use of anakinra, an anti–IL-1β
antibody, exist, with debate over the efficacy and effect on systemic
inflammatory cytokine levels, though arguments against cytokine-lowering
ability were only performed in vitro (296,297). Canakinumab, another IL-1β
inhibitor, was used to treat refractory panuveitis with macular edema in a 4-
year-old Blau boy who had failed several other antimetabolite and biologic
therapies, resolving inflammation and also reducing systemic markers of
innate immunity (309). Corticosteroid implants may play a role in controlling
ocular inflammation; however, one must also be cognizant of untreated
systemic disease with purely local anti-inflammatory modalities.
Cryopyrin-Associated Periodic Syndromes and Other

Periodic Fever Syndromes
Cryopyrin-associated periodic syndromes (CAPS): These syndromes include
a spectrum of three rare overlapping diseases, previously felt to be distinct,
but linked by a common causative autosomal dominant gain-of-function
genetic mutation with variable penetrance (or at times presenting via somatic
mosaicism) in the NLRP3 (CIAS1) gene on chromosome 1q44 (310). These
include, in increasing severity of presentation, familial cold autoinflammatory
syndrome (FCAS), Muckle-Wells syndrome (MWS), and CINCA/NOMID.
These conditions are also considered to be among a larger family of unrelated
disorders, the periodic fever syndromes, due to their tendency to cause
episodic noninfectious fever along with other distinguishing disease
manifestations, which include familial Mediterranean fever (FMF); periodic
fever, aphthous stomatitis, pharyngitis, adenitis (PFAPA) syndrome; and
TNF receptor–associated periodic syndrome (TRAPS). NLRP3 encodes the
protein cryopyrin, and mutations in this key regulatory protein of the
inflammasome complex lead to overproduction of IL-1β in various
leukocytes and chondrocytes, eventually causing the various phenotypes seen
with CAPS.
All three forms of CAPS typically develop early in the first few years of
life but may have features that present later in adulthood and frequently lead
to misdiagnosis once or several times throughout the course of disease. The
mean age of ocular symptoms is 4.5 years of age (311). All 3 forms of CAPS
are very rare, occurring in about 1 per million. FCAS is the mildest form of
CAPS, causing a recurrent nonpruritic urticarial rash, arthralgia, and fever
triggered by exposure to cold temperatures. MWS may have some of the
same features, but also typically includes sensorineural hearing loss,
secondary amyloidosis with nephropathy, and conjunctivitis.
CINCA/NOMID is the most severe and may occur in the neonatal period,
eventually causing CNS changes such as headaches, intellectual disability,
aseptic meningoencephalitis and papilledema, as well as a characteristic body
habitus, arthropathy, urticaria, and fevers.
Ocular manifestations of CAPS range from anterior segment disease such
as conjunctivitis and interstitial keratitis (FCAS and MWS) to more severe
intraocular entities such as uveitis and disk edema (MWS and
CINCA/NOMID). A recent expert consensus categorized recurrent eye
inflammation, described as nonallergic, noninfectious conjunctivitis with or
without other inflammatory ocular manifestations, as one of six criteria for
diagnosis of CAPS along with elevated systemic inflammatory markers, such
as CRP, ESR, or SAA (312). Available case reports detail nongranulomatous
uveitis involvement in about 50% of patients. We have reported adult cases
with juvenile onsets of disease that have either anterior or panuveitis with
papillitis and optic atrophy (311,313,314). Given the variable presentation of
these diseases, they may at times be difficult to discern from JIA early on;
however, one may less often see posterior synechiae, significant cataract, or
glaucoma, and find inclusion of CNS disease in CAPS (311). Successful
treatment of systemic and ocular inflammation has been achieved with use of
IL-1β blocking agents such as anakinra and canakinumab, whereas
inflammation may not respond to more typical conventional anti-
inflammatory agents (313–317).
Familial Mediterranean fever

FMF is an autosomal recessive or possibly dominant with limited penetrance
disease that occurs mostly in Mediterranean populations and is caused by
mutations in the MEFV gene, which encodes pyrin, a protein that helps to
regulate production of IL-1β. It is believed to be the most common of the
periodic fever syndromes. Symptoms are generally characterized by 1 to 3
days of episodic fever, arthropathy, abdominal pain, and pleurisy. A
neutrophil-dominant leukocytosis occurs with elevation of acute phase
reactant proteins as well as secondary amyloidosis. FMF has been shown to
be associated with various autoimmune diseases such as Behçet disease, MS,
JIA, and systemic lupus erythematosus (SLE). Ophthalmic manifestations of
FMF are rare with limited reports documenting only sparse cases in these
patients of concurrent anterior uveitis, intermediate uveitis, panuveitis, as
well as acute posterior multifocal placoid posterior epitheliopathy
(APMPPE); however, it is not at all clear whether these are coincidental or
truly associated with FMF (318–320).
Periodic fever, aphthous stomatitis, pharyngitis, and cervical

adenitis (PFAPA) syndrome
PFAPA is considered the most common nonhereditary autoinflammatory
disease in childhood. As the name suggests, findings include episodic fever
with the distinct morphologic features listed: aphthous stomatitis, pharyngitis,
and cervical adenitis. The pathogenesis is poorly understood, but it is
hypothesized by some that it is driven by IL-1-dependent innate immune
mechanisms driving T-lymphocyte recruitment (321,322). Onset of disease is
usually earlier than age 5 with spontaneous resolution in most cases by
puberty; however, 20% of cases may recur into adulthood (323).
Symptomatic episodes last a week or less and recur every 2 to 8 weeks but
less often in adults. Other symptoms including arthralgia, myalgia, headache,
fatigue, rash, and conjunctivitis are found more often in adults with little to
no long-term sequelae from disease. Corticosteroids are the mainstay of
treatment and lead to rapid improvement of many symptoms as soon as 24
hours after initial administration. Other agents have also been employed with
variable success including NSAIDs, colchicine, cimetidine, and anti-IL-1β
therapy with anakinra and canakinumab (322,324).
We described 2 children as the first documented cases of PFAPA-
associated uveitis. Both had intermediate uveitis with classic presentation of
vitreous and pars plana exudates as well as secondary complications by
CME, vitreous hemorrhages, peripheral retinal neovascularization, ocular
hypertension, and tractional RD (325). Both patients were diagnosed with
PFAPA in their first 3 years of life with chronic intraocular inflammation
refractory to conventional corticosteroid therapy. One was eventually
transitioned to steroid-sparing IMT using methotrexate and the other to
adalimumab and infliximab. Each achieved a steroid-free remission.
Tumor necrosis factor receptor–associated periodic syndrome
Formerly known as familial Hibernian fever or familial periodic fever,
TRAPS is a rare autosomal dominant disease with variable penetrance caused
by a defect in the TNFR1 gene encoding the 55 kDa receptor for TNF. The
disease normally starts in infancy or presents later in the first decade of life.
Symptoms include several days of myalgia, rash, abdominal pain, arthralgia,
and ocular inflammation manifested as conjunctivitis, periorbital edema, and
less commonly, uveitis. The first case of uveitis in TRAPS was reported in a
7-year-old boy with bilateral panuveitis, who was described to have yellow
creamy choroidal lesions, vitreous cells and haze, and widespread capillaritis.
The uveitis finally responded well to adalimumab (326). Other cases of
systemic TRAPS have been treated with anti-TNF agents, as well as anti-IL-
1β agents and tocilizumab.
Intermediate Uveitis
Intermediate uveitis including the idiopathic variant, pars planitis, is
estimated to represent about one-fifth of all cases of pediatric uveitis, with no
definite predilection for race or gender. The prevalence peaks between
childhood and the fourth decade (327). Seventy to ninety percent of cases are
bilateral. About a third of patients with unilateral involvement at the time of
presentation develop bilateral involvement. No inheritance pattern has been
defined, but isolated reports of familial cases support the idea that this disease
may have genetic predispositions (328–337).
Patients with intermediate uveitis generally present with floaters and
blurred vision and typically have a “quiet” and white eye. In some children,
including the preverbal age group, intermediate uveitis is found accidentally
on routine screening or may be found because of leukocoria, amblyopia, or
strabismus. Pain, redness, tearing, and photophobia are signs and symptoms
of anterior segment inflammation, which are seen more commonly in
children with intermediate uveitis than in adults. Moderate to severe cells and
flare, keratic precipitates, posterior subcapsular cataract, band keratopathy,
and even posterior synechiae may be found. Children presenting with
intermediate uveitis have a worse visual acuity both at initial diagnosis and at
follow-up than do adults (338).
Vitreous cells are the most important finding in intermediate uveitis and
are the sine qua non for making the diagnosis. Cellular exudates in the
vitreous and on the pars plana are seen on depressed peripheral examination
of retina in patients with active pars planitis, whereas old cellular debris,
crenated residua of prior vitreal exudates, and a collagen band are generally
present in the patient whose pars planitis is inactive. The terms “snowballs”
and “snowbank” are frequently used in describing examination findings in
this disease; however, we prefer to avoid these terms in practice as they can
be misleading, oftentimes being mistakenly used interchangeably to describe
either active or old inactive inflammatory changes in vitreous and along pars
plana, respectively. Extent of inflammation may vary from a few clock hours,
mostly inferiorly, to 360 degrees of the retinal periphery. Collagen banding
may be acellular or contain fibrous astrocytes, and reflects inflammation
involving the peripheral retina and the vitreous base (339). Although vitreous
and pars plana exudates are not required to make the diagnosis of
intermediate uveitis, they are prominent clinical features of pars planitis and
are found in 74.1% and 66.7%, respectively, in one case series and serve to
delineate pars planitis as a subset of intermediate uveitis (340). The presence
of pars plana exudate is associated with worse visual outcome (341,342).
Other signs of disease include diffuse or peripheral retinal vasculitis with
sheathing of both venules and arterioles (Figure 47-3). Vasculitis may lead to
occlusion, peripheral retinal nonperfusion, and neovascularization of
peripheral retina and optic nerve (343–345).
FIGURE 47-3 Pars planitis. Mid-phase wide field
fluorescein angiogram of the right eye of a 16-year-old
female with active, severe pars planitis demonstrating
periphlebitis in a fern-like pattern with leakage of the
macular and peripheral capillaries, optic nerve head
leakage, and peripheral, inferotemporal nonperfusion.
Autoimmune endotheliopathy is a rare finding in the anterior segment

associated with intermediate uveitis. Peripheral corneal edema evolves to
include keratic precipitates lying in a typically linear fashion inferiorly
marking a distinct border between edematous and normal cornea. This
transplant rejection line–like appearance suggests an autoimmune etiology of
intermediate uveitis, further supported by flow cytometry of peripheral blood
mononuclear cells (PBMCs) in pars planitis that reveals elevated levels of
Th1- and Th17-lymphocytes (346–349).
Children are generally at higher risk for complications from pars planitis,
perhaps explained by a more asymptomatic course than with adults, later
referral, and a greater degree of irreversible damage on initial presentation.
Complications include cataract, macular edema, secondary glaucoma, band
keratopathy, RD, optic neuropathy, neovascularization of the peripheral
retina or optic nerve, vitreous hemorrhage, vasoproliferative tumors, and
formation of inflammatory membranes (350,351). Amblyopia may develop
as a result of prolonged inflammation and the development of these
complications.
Cataract develops in about 50% of patients with intermediate uveitis and
is associated with both uncontrolled inflammation and use of corticosteroids,
especially posterior subcapsular cataract. Cataract is less common in patients
treated with IMT early in the course of their disease (341,352,353).
CME and maculopathy are the most common causes of severe visual
impairment and occur in 12% to 50% of patients (4,5,15). Left untreated,
CME leads to permanent loss of vision through evolution of fixed cysts in the
macula, macular pucker from epiretinal membrane, macular atrophy, and
macular hole formation. Visual acuity may be predicted by assessing the
integrity of the photoreceptor outer segment ellipsoid zone (EZ) as visualized
on OCT (354).
Secondary glaucoma occurs predominantly as a result of topical
corticosteroid therapy. In most studies, ocular hypertension has not been
reported in association with intermediate uveitis (4,355). Paroli et al. found
glaucoma to be more common in adults with pars planitis than in children
(350). In our series, however, 24% of children with pars planitis developed
elevation in intraocular pressure (5).
RD is a complication seen with considerable frequency in patients with
intermediate uveitis. Exudative RDs occur in 5% to 17% of patients with
intermediate uveitis (343,353,356,357). Such high prevalence is uncommonly
seen in other uveitic entities except for the VKH syndrome. In addition, up to
19% of patients with pars planitis may present with bullous retinoschisis,
mostly bilateral and located in the inferior retinal periphery (340,358). Both
exudative RDs and retinoschisis may be related to a Coats-like vascular
response secondary to chronic inflammation (359). Vitreoretinal traction is
reported in 3% to 22% of patients with intermediate uveitis, leading to retinal
tears and combined rhegmatogenous–tractional RD (360,361).
Optic nerve involvement in intermediate uveitis is observed much more
frequently in children than in adults (328,335,350). Disk edema is present in
3% to 50% of eyes (328,356,357,362). Long-standing edema may lead to
optic nerve atrophy, and extensive retinal ischemia may result in optic disk
neovascularization. Optic neuritis in pars planitis patients occurs in
association with MS in up to 38.5% of cases (363,364). Even though MS is
exceptionally rare in children, one might consider a systemic workup if a
patient is a female, has bilateral intermediate uveitis, is HLA-DR15 positive,
and exhibits extraocular symptoms or neurologic signs suggestive of this
disease (365,366). Due to the rarity of the disease in children, we do not
routinely order neuroimaging unless signs and symptoms are suggestive of
neurologic involvement or if anti-TNF therapy is being considered as these
agents are contraindicated in the presence of demyelinating disease.
Peripheral retinal neovascularization is commonly reported in patients
with intermediate uveitis and may become vision threatening with the
development of recurrent vitreous hemorrhages. In many instances, vitreous
hemorrhages will clear, but nonclearing hemorrhages require surgical
intervention (365,366). Children with pars planitis are more likely than adults
to experience vitreous hemorrhage. Peripheral retinal neovascularization may
lead to formation of vascular cyclitic membranes. Although epiretinal
membranes in pars planitis are rarely considered visually significant, their
presence is grossly underestimated (250,367). Secondary vasoproliferative
tumors have been described in pars planitis as well. These lesions typically
develop inferiorly sometimes years after the initial diagnosis and although
benign, may cause significant vision decline by promoting CME, vitreous
hemorrhage, and epiretinal membrane (368).
The diagnosis of intermediate uveitis is based on clinical findings.
Absence of chorioretinal involvement, variable degrees of anterior segment
inflammation in children, and presence of vitreous exudates and pars plana
exudates, which are pathognomonic for pars planitis, papillitis, and
associated retinal phlebitis suggest the diagnosis of intermediate uveitis.
Intermediate uveitis may be associated with various noninfectious entities
such as sarcoidosis, MS, and primary Sjögren syndrome or infectious
diseases such as LD, Whipple disease, peripheral toxocariasis, toxoplasmosis,
syphilis, TB, and human T-cell lymphotropic virus type 1 (HTLV-1). Pars
planitis refers to the subset of intermediate uveitis in which these findings are
present in the absence of an associated infection or systemic disease.
Laboratory and ancillary investigations are guided by a careful history,
review of systems, and examination. Exposure of a child to ticks, skin
changes, or arthritis may suggest the possibility of LD. Contact with cats,
constitutional symptoms, and adenopathy may suggest Bartonella infection.
Fevers, night sweats, and fatigue may be associated with TB or sarcoidosis,
and chest x-rays may be diagnostic. A gallium scan or chest CT is done in
patients with equivocal or negative results and may reveal subclinical
sarcoidosis. Elevated ACE and lysozyme may suggest sarcoidosis; however,
in children, ACE levels are physiologically elevated, and systemic steroids
may suppress ACE levels yielding a false-normal value. Furthermore, these
indices are not specific for sarcoidosis with elevated ACE and lysozyme
levels found in patients with other granulomatous disorders such as TB and
leprosy.
Severe anterior uveitis treated previously with topical steroids may create
confusion due to spillover of inflammatory cells into anterior vitreous and
present a picture of intermediate uveitis. In such cases, if the history is not
typical for AAU, one may choose to observe the course of the disease.
Inflammatory cellular aggregates in the vitreous are found rarely in
iridocyclitis and never in pure iritis.
Although intermediate uveitis has often been considered one of the most
benign forms of uveitis, our experience suggests that early and aggressive
treatment is more effective in preserving good visual function than waiting
until acuity has decreased. Variable reported rates of inducible remission and
relapse thereafter suggest that many patients will need long- term
management, and that remission may be more easily achieved when
treatment is initiated concomitant with initial diagnosis (327,369). We use a
modified stepladder approach for the treatment of intermediate uveitis as
initially proposed by Kaplan after treatable infectious and noninfectious
entities that may simulate or present a clinical picture of intermediate uveitis
have been ruled out (370). Step one may be topical corticosteroids or regional
corticosteroid injections, followed by step two with oral NSAIDs, if
inflammation recurs, and topical NSAIDs in the presence of CME. Step three
involves systemic corticosteroid therapy, such as prednisone 1 mg/kg/d orally
and taper thereafter guided by the clinical response, limiting to no more than
3 months of therapy.
In cases of recalcitrant inflammation, step four consists of systemic IMT
versus therapeutic pars plana vitrectomy (PPV) with endolaser and/or
cryopexy peripheral retinal ablation. It is not clear, however, whether
vitrectomy should precede systemic IMT or whether all patients should
receive preoperative IMT. Nineteen of twenty patients in our study treated
with IMT were able to successfully completely discontinue corticosteroid
therapy. Shin et al. showed that use of IMT prior to PPV decreased
postoperative uveitis flares and need for local and systemic medication (371).
Immunomodulatory drugs used in our practice include methotrexate and
MMF as a first choice. Azathioprine is the second choice, and cyclosporine
may be used to supplement therapy. Biologic therapy, especially with TNF-
blocking agents such as infliximab and adalimumab, may also be used in
those with refractory inflammation (71). As intermediate uveitis, especially
pars planitis, may be associated with an increased risk of MS, one must
exercise caution with vigilant monitoring of pediatric patients with
intermediate uveitis on long-term anti-TNF treatment. This is especially true
in those with the HLA-DR15 allele, and many uveitis specialists defer to
other therapy with review of systems or MRI findings suggestive of possible
demyelinating disease. In addition, there is a still a questionable association
of TNF blocking agents with the development of lymphoma in some
children. Cyclophosphamide and chlorambucil are the drugs of last resort due
to their potential for significant toxicity.
Therapeutic surgical care of noninfectious intermediate uveitis can
include intraocular corticosteroids and anti-VEGF agents, cryoablation, and
PPV. Dexamethasone implants have been used with efficacy and decrease the
need for IMT but increase the risk of cataract and glaucoma (209,210).
Peripheral cryoablative therapy to areas of active inflammation compares
favorably with conventional application of topical, regional, and systemic
corticosteroids (372). PPV with possible adjunct endolaser or cryotherapy,
although not universally accepted, is now at least more widely considered in
the management of noninfectious, posterior intraocular inflammation
associated with intermediate and pediatric uveitis. Several studies in the last
decade have looked at PPV and found attenuation of disease activity and
inflammatory recurrences, a reduction in CME, a diminished requirement for
anti-inflammatory medications, and an improvement in vision in carefully
selected patients (327,371,373–376). Our cohort showed 96% of patients
achieved inflammatory control with or without concomitant medical therapy
including five of six patients with persistent retinal vasculitis, a group known
to be particularly resistant to medical therapy. Although there was only a
modest reduction in the need for systemic IMT postoperatively, it should be
noted that no eyes had adequate inflammatory control prior to surgery. Intra-
and postoperative complications may include retinal tears or detachment,
acceleration of cataract formation, and uveitis flare. Cataract formation may
be a particular deterrent as eventual removal negates the advantageous
accommodative capacity of the crystalline lens in children. Careful
consideration of the relative risks and benefits and judicious case selection
for children undergoing PPV for this indication is paramount with those
having poorly controlled inflammation despite extended therapy with IMT or
biologic therapies being the most likely to experience a therapeutic benefit
from surgery.
Adamantiades-Behçet Disease
Adamantiades-Behçet disease (ABD) is a chronic, recurrent–remitting, and
progressive, idiopathic multisystem inflammatory vascular disorder, which is
characterized by the presence of recurrent oral aphthous and genital ulcers,
skin lesions, and intraocular inflammatory disease together with other
systemic findings. The diagnosis is a clinical one based on specific criteria,
which vary depending on the classification system employed (377–379).
Onset is usually after puberty, predominantly in young adults between ages
20 and 40, but there are observations of putative patients with onset before
puberty (380,381). Since Mundy and Miller reported the first pediatric case in
1978, additional reports from different parts of the world have followed, most
of which include small numbers of patients. Even fewer studies compare the
expression of childhood- to adult-onset ABD (380,382–387). Most of these
reports defined pediatric ABD based on age of presentation before 16 years
(387).
There are no definitive diagnostic laboratory findings for the disease. The
diagnosis of ABD relies on the identification of specified clinical criteria as
described by several classification systems; the most commonly employed
are those enumerated by the International Study Group for Behcet’s Disease.
Juvenile-onset disease is characterized by an increase in familial cases, a
lower incidence of severe complications, and a delay of the clinical course; it
is less severe than adult-onset disease, especially because the frequency of
systemic involvement is higher in adults (387–389). A clinical study of ABD
in childhood compared to adult-onset disease showed that all patients had
recurrent oral ulcers, similar ocular and skin lesions, positive pathergy test,
and similar amounts of arthritis, vascular involvement, and recurrent
headaches (390). Genital ulcers were significantly more common in adults, as
were nonspecific GI symptoms and CNS involvement other than headaches.
ABD is classically felt to be a polygenic autoinflammatory disease with
strong association with the HLA-B51 allele along with other newly
discovered risk loci (391). More recently, a separate entity has been
described, which closely mimics ABD, and is caused by an autosomal
dominant mutation in the TNFAIP3 gene encoding the TNFα-induced protein
3 (TNF/AIP3), also called A20. A20 is a down-regulator of the NF-κB
pathway. Mutations lead to haploinsufficiency of A20 (HA20), which in turn
leads to increased production of inflammatory cytokines such as IL-1β, IL-6,
and TNF and facilitation of differentiation and activation of various
proinflammatory lymphocytes including Th17 cells (392,393).
Children with this disorder often meet major criteria for ABD; however,
significant differences exist between this condition and more classical ABD,
including infrequent HLA-B51 association, early age onset (median age 5.5
years), ubiquitous geographic distribution, preponderance of females,
recurrent fever, more significant abdominal findings, and poor response to
colchicine (394). It is unclear as to the extent to which this entity has been
included in previous studies of ABD and requires further investigation to
distinguish the two seemingly separate disorders. At least a similar pathway
may exist between them that may be important in the development of ocular
inflammation, since decreased A20 has been shown in peripheral blood
monocytes and dendritic cells in ABD patients with active versus inactive
uveitis (393).
Age distribution of disease onset in children has been reviewed in several
studies (384–390,395–398). Most patients were Caucasian of Turkish,
Iranian, or European descent with mean age at onset of 8.4 years (0 to 16
years), and mean age at diagnosis of 13 years. A transient neonatal form of
(ABD) disease has been described in infants born to mothers with the
syndrome with three criteria for the diagnosis of (ABD) disease: stomatitis,
genital ulcers, and bullous skin lesions. There is no consensus regarding the
age at which juvenile disease should be differentiated from that in adults, nor
whether juvenile ABD should be defined by fulfillment of the classical versus
updated or new classification criteria (377,378). More recent studies confirm
the following clinical features for children believed to have ABD as
described below (384,398,399).
Buccal aphthosis (painful ulcers found on lip, cheek, tongue, palate,
tonsil, gingiva, and pharynx mucosa) is present in almost all (96% to 100%)
children and may be the presenting symptom in 60% with mean age of onset
of 4 years. Lesions are discrete, round or oval white ulcerations 3 to 15 mm
in diameter with red a rim. Attacks vary from 3 per year to being present
almost constantly. Genital ulcers occur in 70% of cases and are similar in
appearance, usually appearing later in children who are older at disease onset
and affecting the vulva, scrotum, and penis as well as the perianal region
(384,395).
Skin lesions are present in 90% of pediatric ABD patients and include
papules, pustules, acneiform lesions, pyoderma gangrenosum–type lesions,
palpable purpura, hypopigmented areas, and purulent bullae; pustular
eruptions are the commonest skin lesion (400). A pathergy test is positive in
80% of the patients. Arthritis is usually pauciarticular, involving knee, ankle,
hip, metatarsophalangeal joints, the shoulder, and the sternoclavicular joint.
Gastrointestinal (GI) involvement varies widely, depending on region or the
report, and includes abdominal pain, vomiting, flatulence, diarrhea, and
constipation. Radiologic examinations demonstrate thickened mucosal folds,
pseudopolyps, deformity of bowel loops, ulcerations, and fistulae.
Ulcerations are localized or diffuse, with the majority (76%) occurring in the
ileocecal region, and rarely in esophagus (401–404). The vasculitic process
affects both arterial and venous systems. Venous thrombosis has been
observed in 15% of children with ABD, most often in the lower extremities
but can occur in the cerebral circulation, inferior vena cava, and central retina
artery or vein, whereas arterial complications, including aneurysms and
thrombosis, may be fatal. Pulmonary artery involvement can lead to life-
threatening hemoptysis from pulmonary hemorrhage. Pulmonary signs
include generalized lymphadenopathy, parenchymal infarction related to
venous thrombosis, chest pain, and hemoptysis (397). Renal involvement
results in proteinuria and hematuria. Urethritis and orchitis, or epididymitis,
may occur. Myocarditis and arrhythmia may also occur in children with
ABD.
Neurologic signs are present in 15% of pediatric ABD patients and are
the most serious manifestations of disease. They include headache,
meningitis, benign intracranial hypertension, brainstem involvement,
neuropsychiatric symptoms, and meningoencephalitis, hemiparesis or
paraparesis with spastic quadriparesis, and seizures. MRI and CT findings
include ventricular dilatation and hyperintensity signals in the pons,
brainstem, putamen, and upper medulla (390). A cerebral venous and dural
sinus thrombosis may occur (405,406). Cerebral spinal fluid analysis may
show elevated protein and hypercellularity, with lymphocytosis and negative
culture. Organic psychiatric disturbances were reported with severe
neurologic symptoms. Depression, loss of memory, and personality changes
may occur.
Eye lesions are present in 60% of ABD children and include
conjunctivitis, scleritis, uveitis, optic disk edema, retinal vasculitis, and optic
atrophy (407,408). An international collaborative study of 86 cases showed
that uveitis was strictly anterior in 8% of the cases, strictly posterior in 9%,
and panuveitic in 28%, usually bilateral. Eighty-nine percent of patients had
severe uveitis, retinal vasculitis, or both, and uveitis was significantly more
frequent in boys than girls (407). Panuveitis was the most common type of
uveitis in cases with childhood-onset Behçet disease in a study on children
from Turkey. Cataract was the most common anterior segment complication,
and optic atrophy was the most common posterior segment complication
(395). Cataract, maculopathy, and optic nerve atrophy were found to be the
most common complications and seen in over 40% of all eyes each (408).
The choice of medical therapy is based on the severity of the disease. In
general, treatment should be more aggressive when the following are present:
complete ABD, vascular involvement, retinal and bilateral involvement, CNS
involvement, male sex, and a geographic origin in the Mediterranean basin or
Far East. A wide variety of agents have been employed in the treatment of
severe uveitis associated with ocular ABD including corticosteroids,
azathioprine, chlorambucil, cyclophosphamide, cyclosporine, methotrexate,
MMF, α-interferon, and TNF inhibitors such as infliximab, the latter being
the first-line drug of choice by many uveitis experts for this indication (148).
Other treatment modalities have included colchicine, plasmapheresis,
penicillin, thalidomide, and other various biologic medications including
anakinra and rituximab (409–411).
Kawasaki Disease
Kawasaki disease (KD), also known as mucocutaneous lymph node
syndrome, is a primary vasculitis often seen in childhood. It is mediated by
immunoglobulin A (IgA), affects mostly small- and medium-sized vessels,
and can lead to a fibrinoid necrosis in vessel walls (412). The disease
damages the intima layer to perivascular area of vessels, leading to aneurysm
formation in several different stages in a childhood polyarteritis. This is a
systemic vasculitis, but preferentially manifests in coronary arteries (413).
KD is the leading cause of acquired heart disease in children. Although
approximately 80% of patients are <5 years of age, older children and
teenagers may exhibit this disease. KD is more common in boys than girls,
and the majority of cases are diagnosed in the winter and early spring.
Although it is more prevalent among children of Asian and Pacific Island
descent, KD affects people of all racial and ethnic groups.
The cause of KD is unknown. Modified criteria for the diagnosis of KD
include the major criterion of fever persisting for at least 5 days, plus four of
the following five features: changes in peripheral extremities or perineal area,
polymorphous exanthema, bilateral conjunctival injection, changes of lips
and oral cavity or injection of oral and pharyngeal mucosa, and cervical
lymphadenopathy. In the presence of coronary artery involvement detected
on echocardiography and fever, fewer than four of the remaining five criteria
are sufficient (414). Common symptoms leading parents to bring their
children to medical attention include high fever that lasts for 5 or more days;
rash, often worse in the groin area; red bloodshot eyes without drainage or
crusting; bright red, swollen, cracked lips; “strawberry” tongue, which
appears with shiny bright red spots after the top coating sloughs off; swollen
hands and feet and redness of the palms and soles of the feet; and enlarged
lymph nodes in the neck (Figure 47-4).
FIGURE 47-4 Kawasaki disease. A 5-year-old child with
fever for 5 days and diagnosed with Kawasaki disease
with characteristic findings of (A) desquamative rash of
the fingers and feet, (B) rash often worse in the groin area,
and (C) swollen, cracked lips.
The most severe complication of the disease is coronary vasculitis leading to

coronary artery changes, which may develop in 15% to 20% of patients if left
untreated (415). These changes include aneurysms, coronary artery ectasias,
and stenoses, and carry a 2% mortality rate. The administration of
intravenous immunoglobulin G over the first 10 days of the disease leads to a
reduction of coronary artery impairment of 3% to 8%, and a decrease in
mortality to 0.2% (415,416). Patients also receive high-dose aspirin therapy.
Aneurysms of other arteries may occur. Gastrointestinal complications in KD
are similar to those observed in Henoch-Schönlein purpura and include
intestinal obstruction, colonic edema, intestinal ischemia, intestinal pseudo-
obstruction, and acute abdomen (417–421). KD can also manifest as
necrotizing vasculitis, progressing to peripheral gangrene (422–424).
The neurologic complications include meningoencephalitis, subdural
effusion, cerebral hypoperfusion, cerebral ischemia and infarct, and
cerebellar infarction presenting with seizures, chorea, hemiplegia, mental
confusion, lethargy, and coma (425–431). Other reported neurologic
complications include ataxia, facial palsy, and sensorineural auditory loss
(430,432–438). Behavioral changes may develop with attention and learning
deficits; emotional disorders, such as emotional lability, fear of night, and
night terrors; and internalization problems including anxious, depressive, or
aggressive behavior (439,440).
Ophthalmologic findings associated with KD include iridocyclitis,
punctate keratitis, ptosis, conjunctival hemorrhage, optic neuritis, amaurosis
fugax, and central retinal artery occlusion (441–446). Ophthalmologic
complications occur during the acute and subacute phases. They are usually
transient and clear within 2 to 3 months of the acute phase (442). Bilateral
bulbar conjunctival erythema is present in more than 90% of affected
children. Chemosis, follicles, and papillae are characteristically absent. Burns
et al. assessed 41 patients with KD during the acute phase and found 27
patients with anterior uveitis (25 bilaterally), 5 patients with punctate
keratitis, and 3 patients with keratouveitis (447). Anterior uveitis is usually
mild and bilateral and is sometimes associated with keratic precipitates,
which develop in as many as 83% usually in about a week after fever onset
and commonly resolve without any of the usual structural complications of
uveitis (447). Optic disk edema, when present, may last longer than the other
ocular features and has been seen as long as 6 months after presentation
despite therapy (448). Retinal microvascular changes were studied by Chen et
al. in two large groups of KD patients 2 years after their acute illness, and in
one group, significant retinal venule enlargement, a feature also associated
with coronary artery disease in adults, was seen in comparison to controls
using computer-assisted analysis of standardized fundus photography (449).
Uveitis and conjunctival inflammation are treated with topical
corticosteroids. Punctate keratitis requires supportive care, such as artificial
tears and ointments. Posterior segment disease may require more aggressive
anti-inflammatory therapy with regional or even systemic corticosteroids.
Vogt-Koyanagi-Harada Disease
VKH disease is a multisystem disorder involving the ocular, auditory,
nervous, and integumentary systems. Ocular manifestations include most
commonly bilateral granulomatous panuveitis with exudative RDs. VKH
disease is an uncommon cause of uveitis in children. Children with VKH
represented 16% of all children with uveitis in a tertiary referral center in
Saudi Arabia and 8.2% in India (28,450). In the Western Hemisphere, the
disease is rare. Treatment is usually commenced with systemic
corticosteroids with the majority of pediatric patients requiring
immunomodulatory agents. Methotrexate, azathioprine, cyclosporine,
cyclophosphamide, chlorambucil, MMF, and biologic agents such as
infliximab and rituximab may be successfully used to treat pediatric VKH
disease (451,452).
Systemic Lupus Erythematosus

SLE is an autoimmune disease that is characterized by periodic episodes of
inflammation of and damage to the joints, tendons, other connective tissues,
and organs, including the heart, lungs, blood vessels, eye, brain, kidneys, and
skin. Production of numerous autoantibodies and deposition of pathogenic
immune complexes occur. The disease is known to have periods of flare-ups
and periods of remission. Children with SLE can have a high degree of renal
involvement, which is more frequent than in adults. Overall, 60% to 80% of
children with SLE have urinary or renal function abnormalities early in the
disease course. In 90% of patients, renal disease occurs within 2 years from
disease onset (453). Clinically significant renal involvement ranges from
asymptomatic urinary findings to nephrotic syndrome and renal failure.
Incidence and prevalence rates among children younger than age 15 years
have been reported to be 0.5 to 0.6 and 4 to 250 per 100,000 persons,
respectively. SLE occurs more often in Native Americans, Asian Americans,
Latin Americans, and blacks. African American children may represent up to
60% of patients younger than age 20 years. SLE in children occurs most
often at age 15 years and older and is rare in children younger than 5 years. A
female-to-male ratio of approximately 4:1 occurs before puberty, and a ratio
of 8:1 occurs after puberty. At one time, lupus was thought to be more severe
in children than in adults, but currently, there is no evidence to support this
assertion (454).
The diagnosis of SLE is made based on clinical and laboratory findings
with the identification of 4 of 11 criteria enumerated by the American
College of Rheumatology or by the newer criteria put forth by the Systemic
Lupus International Collaborating Clinics in 2012 in which 4 of 17 criteria,
including at least one clinical and one immunologic, must be met or the
patient must have biopsy-proven lupus nephritis in the presence of ANA or
anti–double-stranded DNA antibodies (Table 47-9) (456,457). Initial
symptoms may include hematologic abnormalities, cutaneous symptoms,
musculoskeletal abnormalities, renal symptoms, and fever. Children in whom
SLE is suspected should undergo a serologic evaluation, including ANA,
ESR, anti-dsDNA, anti-Sm, anti-RNP, anti-Ro (SSA), and anti-La (SSB), as
well as measurement of complement levels. Sometimes ocular manifestations
may precede systemic signs. A high level of suspicion should be present
when evaluating a child with cotton-wool spots or intraretinal hemorrhages.
TABLE 47-9 Classification criteria for systemic

lupus erythematosus
aTan EM, Cohen AS, Fries JF, et al. The 1982 revised criteria for the classification of systemic lupus
erythematosus. Arthritis Rheum 1982;25(11):1271–1277, Ref. (455).
bPetri M, Orbai A-M, Alarcon GS, et al. Derivation and validation of Systemic Lupus International
Collaborating Clinics classification criteria for systemic lupus erythematosus. Arthritis Rheum
2012;64(8):2677–2686.
C, complement factor; EKG, electrocardiography; dsDNA, double-stranded DNA; Ig, immunoglobulin;
C3, complement 3; C4, complement 4; CH50, total complement activity (50% hemolytic component);
pHTN, portal hypertension; SLE, systemic lupus; Sm, Smith; TTP, thrombocytopenic purpura.
Ocular manifestations in children most often include keratoconjunctivitis

sicca and also include episcleritis, scleritis, keratopathy, uveitis, retinal and
choroidal microangiopathy, papillitis, and neuro-ophthalmic disease. Lupus
retinopathy is a particular poor prognostic sign for survival in adults and
almost always portends active disease (458,459). Neuro-ophthalmic
involvement is apparent when strabismus develops from cranial nerve
dysfunction. Uveitis is rare in children with SLE. One recent cohort of 40
children reported no cases of uveitis; however, dry eye, retinal vascular
changes, and cataract were seen (460). A large retrospective study of
Brazilian childhood SLE patients over 30 years showed that only 7 of 852
patients 16 years or younger had uveitis with mean age of onset of 10 years
and typically presenting within 6 months of systemic diagnosis (461).
Symptoms usually included redness and blurring. Posterior, anterior, and
panuveitis were seen, as were retinal and choroidal exudates and
hemorrhages. Complications in this series included cataract and retinal
vasculitis and ischemia with subsequent neovascularization. One child died
from SLE complications shortly after uveitis diagnosis. In this cohort, uveitis
in childhood SLE was associated with increased SLE disease activity index
2000 (SLEDIA-2K), fever, lymphadenopathy, arthritis, and use of
intravenous methylprednisolone (461).
The goal of therapeutic intervention in pediatric systemic and ocular
lupus is to control disease manifestations and allow the child to have a good
quality of life without major disease exacerbations while preventing serious
organ damage that adversely affects function or life span. NSAIDs;
antimalarial drugs; corticosteroids; immunosuppressive medications such as
cyclophosphamide, MMF, azathioprine, methotrexate, and cyclosporine;
anticoagulants; and monoclonal antibodies all have been used to treat the
underlying systemic disease and the ocular manifestations of lupus. Biologic
therapies may be effective treatments for lupus, including rituximab, other B
cell– and T cell–directed therapy, anticomplement and anticytokine therapies,
and peptide manipulation to promote tolerance. Stem cell transplantation and
high-dose immunoablative therapies are being studied, as well. The 5-year
survival rate for children with SLE is more than 90% (462–467). Most deaths
in children with SLE are the result of infection, nephritis, renal failure,
neurologic disease, or pulmonary hemorrhage. Plasmapheresis has been used
along with aggressive IMT in treating adult patients with active retinal
vasculitis and in two pediatric cases of lupus retinal vasculitis, combination
rituximab and cyclophosphamide was required (468,469). Visual prognosis
for children with SLE depends on the prompt diagnosis and treatment of
underlying systemic disease.
Juvenile Dermatomyositis
Juvenile dermatomyositis (JDM) is a rare autoimmune disease mainly
affecting striated muscle and skin and is also associated with systemic
vasculitis and calcinosis involving the heart, lungs, and gastrointestinal tract.
Symptoms generally include bilateral symmetric proximal muscle weakness
along with dermatologic manifestations such as Gottron papules seen on
hands and knuckles as well as the classic violaceous periorbital heliotropic
rash. Other systemic features may include fever, arthralgia, asthenia,
anorexia, and general malaise. Serum creatine kinase may be significantly
elevated. Overlap features with other diseases may be present, such as
systemic sclerosis, SLE, and mixed connective tissue disease. The incidence
is 3.2 per 1 million children yearly in the United States and is more common
in females. The typical presenting age is between 5 and 10 years. A genetic
susceptibility has been suspected given association with various HLA class I
and class II alleles, such as HLA-DQA1*0501. Unlike the adult form, JDM is
not associated with malignancy (470,471).
Ocular manifestations of JDM include the above mentioned periorbital
heliotropic rash in 45% to 100% (pathognomonic for JDM), steroid-induced
cataracts in 17%, and retinal findings, possibly incidental, in a small number
of patients (472–474). Eleven cases of retinal pathology have been described
in the literature, with findings including diffuse retinal exudates centered over
the macula and optic nerve, intraretinal hemorrhage, macular edema, and a
Purtscher-like retinopathy. We described the first JDM-associated case of
paracentral acute middle maculopathy (PAMM) in a 13-year-old girl,
presenting with inner retinal whitening, macular edema, and Purtscher
flecken. Fluorescein angiography demonstrated bilateral diffuse retinal
perivenous leakage and punctate choroidopathy. OCT showed diffuse
intraretinal and subretinal fluid initially, and later in the course of disease, a
hyperreflective band along the inner nuclear layer and outer plexiform layer
junction became evident. These acute findings resolved completely while the
patient received aggressive IMT from her rheumatologist and left behind
atrophic retinal changes consistent with PAMM (473). Hence, patients with
JDM and vision changes should have a thorough dilated evaluation for
possible retinal involvement.
Polyarteritis Nodosa
Polyarteritis nodosa (PAN) is a necrotizing medium-sized–vessel vasculitis
associated with aneurysmal nodules along the walls of medium-sized
muscular arteries. In adults in Europe and the United States, PAN has an
estimated annual incidence of 2.0 to 9.0/million (475). While rare in
childhood, it is the most common form of systemic vasculitis after Henoch-
Schönlein purpura and KD (476,477). Peak age of onset in childhood is 7 to
11 years, often with a male preponderance. Ocular manifestations include
scleritis, peripheral ulcerative keratitis, and retinal vasculitis, albeit extremely
rare in children. A recent review by Iudici et al. of systemic characteristics in
childhood versus adult PAN showed no ocular findings in their 21 pediatric
patients (478). In addition, patients with deficiency in the protein adenosine
deaminase 2 (DADA2) have been shown to have symptoms, which closely
mimic those of PAN (479).
Granulomatosis with Polyangiitis

Granulomatosis with polyangiitis (Wegener) in children is also rare. The
estimated incidence in childhood is <1 per 2 million per year (480). During
the 10-year period from 1979 to 1988, granulomatosis with polyangiitis
(Wegener) was listed as the cause of death on 1,784 death certificates; 22 of
these deaths were of children younger than 15 years (481). The US Renal
Data Systems Annual Data Report from 2002 to 2006 stated that
granulomatosis with polyangiitis (Wegener) was the underlying cause of end-
stage renal disease (ESRD) in 54 (0.9%) children (0 to 19 years) (482). The
North American Pediatric Renal Transplant Cooperative Study 2007 Annual
Report listed the causes of renal failure in their pediatric dialysis database
from 1992 to 2006 (483). Of the more than 9,500 children transplanted in the
registry, granulomatosis with polyangiitis (Wegener) was listed as the
primary disease in 52 cases (0.5%). Permanent morbidity from disease or its
treatment occurs in 87% of patients. The 1st-year mortality for children in the
United States with granulomatosis with polyangiitis (Wegener) and ESRD
was 3.7% (482). Most patients with granulomatosis with polyangiitis
(Wegener) are Caucasian (484,485). Of the pediatric patients with
granulomatosis with polyangiitis (Wegener) who developed ESRD in the
United States, 82% were white (481). A female preponderance is found in
patients with childhood-onset granulomatosis with polyangiitis (Wegener)
(484). The mean age of patients varies from 6 to 15.4 years old in different
studies (484).
Most pediatric patients present with ear, nose, and throat pathology that
includes sinusitis (61%), nasal disease (48%), otitis media (39%), hearing
loss (26%), and ear pain (22%) (486). Ultimately, 91% of pediatric patients
develop upper respiratory problems, with sinus involvement in 83% of
pediatric patients. Granulomatosis with polyangiitis (Wegener) may involve
the larynx, subglottic space, trachea, and mainstem bronchi. Subglottic
stenosis ultimately developed in 48% of pediatric patients. Pulmonary
involvement can be asymptomatic, insidious in onset, or severe and
fulminant. In the pediatric series, lung disease occurred in 22% of patients at
presentation and eventually developed in 74% of patients (486). Pulmonary
infiltrates (61%), pulmonary nodules (43%), and hemoptysis (26%) were the
most common abnormalities. Radiographic abnormalities may occur in the
absence of symptoms. Patients with granulomatosis with polyangiitis
(Wegener) may develop life-threatening diffuse alveolar hemorrhage,
accompanied by progressive glomerulonephritis (GN). GN was present in
only 9% of patients at presentation but ultimately developed in 61% (14 of
23) of patients at some point during the course of the disease. GN manifests
as hematuria, proteinuria, and renal insufficiency. Hypertension and gross
hematuria are typically uncommon. Systemic symptoms in children,
including arthralgia/arthritis (30%), fever (22%), weight loss (13%), and rash
(9%), were also present (486). At some point during the disease,
musculoskeletal symptoms occurred in 78% of patients, fever in 43% of
patients, weight loss in 26% of patients, and rash in 52% of patients (486).
Eye disease occurred in 13% of pediatric patients at the outset of disease
and 48% of patients overall (486). In contrast to studies in adult patients
where orbital inflammation and scleritis are most frequently seen, a series of
pediatric patients with granulomatosis with polyangiitis (Wegener) found that
dacryoadenitis, proptosis, eye pain, and episcleritis were the most common
abnormalities (486). Another more recent report of the PRINTO vasculitis
database showed that 35% of children presented with eye findings such as
conjunctivitis and keratitis (487).
The histologic diagnosis is established through biopsy of affected tissues.
Imaging of the chest and sinuses may reveal nodular, diffuse, or cavitary
lesions on x-ray and/or CT, while laboratory studies may show proteinuria or
hematuria, elevated sedimentation rate, C-reactive protein, and the presence
of antineutrophil cytoplasmic antibodies (ANCAs). ANCAs are antibodies
directed against cytoplasmic azurophilic granules of neutrophils and
monocytes, which are specific markers for a group of related systemic
vasculitides, including granulomatosis with polyangiitis (Wegener), PAN,
microscopic PAN, Churg-Strauss syndrome, and pauci-
immunoglomerulonephritis. Two principal types have been described based
on the immunofluorescence staining pattern on ethanol-fixed neutrophils and
the main target antigen. The cytoplasmic pattern, c-ANCA, is both sensitive
and specific for granulomatosis with polyangiitis (Wegener) and is present in
up to 95% of patients with proteinase 3 (PR3), the most common target
antigen. The perinuclear pattern, or p-ANCA, is associated with PAN,
microscopic PAN, relapsing polychondritis, and renal vasculitis, with
myeloperoxidase (MPO) being the most common antigenic target. In contrast
to granulomatosis with polyangiitis (Wegener), the diagnostic sensitivity of
c-ANCA and p-ANCA for PAN is only 5% and 15%, respectively; in patients
with microscopic PAN, p-ANCA (MPO) positivity is more common (50% to
80%), with a smaller percentage (40%) having the c-ANCA (PR3) marker
(488).
It must be emphasized that this diagnosis, as with PAN, mandates the
implementation of systemic corticosteroids and IMT, specifically with
cyclophosphamide or rituximab, at the outset as such therapy not only
reduces ocular morbidity but significantly improves overall mortality as
compared to treatment with corticosteroids alone. Pediatric deaths have been
reported in 9% to 12% of patients (484).
Sympathetic Ophthalmia
Sympathetic ophthalmia (SO) is a bilateral autoimmune granulomatous
panuveitis, which presents after penetrating insult to the globe, be it
accidental or surgical, as well as nonpenetrating procedures such as proton
beam irradiation. Inflammation ensues, typically 2 weeks to 3 months after
the insult but may occur decades after and is from exposure of uveal proteins
to the systemic immune system that leads to excitation and up-regulation of a
pro-inflammatory T-lymphocyte population. These T lymphocytes can occur
in both the excitatory and sympathizing, that is, nontraumatized, fellow eye.
The incidence was seen to be as low as 0.24% (6/2511) in a southern Indian
cohort of children presenting with open globes over 10 years (489). HLA
associations have been made, including HLA-DRβ1*04, an allele also found
in increased frequency in VKH, to which this disease is often compared
because of similar phenotypic and immunopathologic features.
Clinical examination may show anterior segment findings such as
anterior uveitis and corneal endothelial attenuation with bullous keratopathy.
Posterior segment choroidal thickening, best visualized on enhanced depth
OCT imaging, with multiple white-yellow lesions, known as Dalen-Fuchs
nodules, is seen in about half of patients with disease. Papillitis with optic
nerve hyperemia is also characteristically observed. Complications include
glaucoma, cataract, optic atrophy, and RD. Fluorescein angiography presents
similar to VKH with early multifocal hyperfluorescent patchy leakage and
expand and coalesce late consistent with an exudative RD. Aggressive
topical, regional, or systemic corticosteroid therapy is the mainstay of
treatment for acute disease with early introduction of steroid-sparing IMT in
an effort to prevent and reduce the severity of recurrent inflammatory disease.
Various agents ranging from antimetabolites to biologic therapy have been
successfully employed, including adalimumab and infliximab in treatment-
resistant cases (490,491). Also, the 0.7-mg intravitreal dexamethasone
implant has been shown to have efficacy (209).
Masquerade Syndromes
There are other ocular conditions, both malignant and nonmalignant, that are
not primarily inflammatory in nature but may “masquerade” as uveitis (see
Chapters 44, 45 and 49). Malignancy is an infrequent, but important,
masquerader of pediatric uveitis. Childhood neoplasias that may have
intraocular manifestations include acute leukemias, posttransplant
lymphoproliferative disorders, RB, uveal melanoma, intraocular–CNS
lymphomas, and metastatic cancer. As these malignant entities are
exceedingly rare in the pediatric population, clinicians may fail to even
consider them in cases of steroid-resistant uveitis, and diagnosis and
treatment of the conditions may be delayed. The acute leukemias (acute
myelogenous leukemia and acute lymphoblastic leukemias) may masquerade
as a pediatric uveitis, and intraocular involvement can be the initial
manifestation of systemic disease (492). Intraocular involvement may include
cancerous cells resembling normal lymphocytes within the anterior chamber
and/or the vitreous masquerading as inflammation and even a
pseudohypopyon (Figure 47-5). This mass of leukemic cells is creamy white
in color and may shift depending on the patient’s position. In contrast, a
nonshift or delayed shift is characteristic of a true inflammatory hypopyon
due to fibrin deposition within the white cell mass (493). Cytology obtained
from an anterior chamber paracentesis is consistent with the malignant blood
dyscrasia. A secondary glaucoma from leukemic infiltration of the trabecular
meshwork is rare but can also occur. Diffuse invasion of the iris can produce
heterochromia, and more focal lesions can create the appearance of iris
nodules. Perivascular sheathing and/or avascular leukemic exudates
mimicking retinal vasculitis are frequent findings of posterior segment
involvement (494,495). The choroid is the most common anatomical site
affected with a subretinal mass, serous RD, or diffuse choroidal thickening
most apparent on ultrasound and these findings are highly suggestive in a
patient with a history of leukemia (494,496). These clinical signs are in
contrast to those of leukemic retinopathy in which Roth spots, cotton-wool
spots, and extensive preretinal and retinal hemorrhages are the heralding
features of ongoing thrombocytopenia and anemia (496,497). Orbital
infiltration can also occur in both acute and chronic forms of leukemia and
can range from an insignificant mass to a large, space-occupying lesion. The
presentation of orbital involvement is variable but may include proptosis,
ecchymosis, chemosis, diplopia, visual disturbances, and/or extraocular
motility deficits. Orbital involvement is much more common in acute
leukemias in children than adults.
FIGURE 47-5 Pseudohypopyon in a patient with acute
lymphocytic leukemia.
The eye is an immunologically privileged site as is the brain due to the

blood–ocular barrier, which is composed of tight junctions that prevent
noxious molecules from entering extravascular space of the retina and
promote retention of important compounds within ocular tissue. Due to this
barrier, adequate intravenous therapy may not effectively eradicate
intraocular neoplastic disease. As such, local radiation and/or intraocular
chemotherapy for primary intraocular lymphoma may be required to treat the
sequestered ocular component of the disease. In cases where the intraocular
component is found initially, an exhaustive search for extraocular disease is
required. Importantly, the development of ocular involvement of leukemia in
a patient believed to be in remission should raise serious concerns for
systemic relapse, especially with CNS involvement (498).
RB is the most frequent primary malignant intraocular tumor in children
and is found in 1 in 20,000 infants. The malignancy is usually identified
before the age of 6 and is familial in 40% of cases and sporadic in the other
60%. In two-thirds of cases, the tumor is confined to only one eye (499).
Trilateral RB is an entity in which an intracranial tumor is found in
association with heritable RB necessitating systemic chemotherapy and
unfortunately carries a poorer prognosis (500). The most common ocular
findings in RB are leukocoria and strabismus. Less commonly, a
pseudouveitis due either to an inflammatory response to tumor necrosis or to
seeding of the tumor in the anterior chamber resembling inflammatory cells
may be the initial finding in a child with RB. Findings at presentation may
include a red, painful eye with a pseudohypopyon and/or hyphema that may
lead to diagnostic confusion with RB being misdiagnosed as an anterior
uveitis (501). It is likewise critical to consider RB in the differential diagnosis
of any child who is thought to have endogenous endophthalmitis but who
lacks systemic signs of infection. In such cases, a careful examination for
subtle structural stigmata of concurrent or prior intraocular inflammation, that
is posterior synechiae, is essential as their presence would be exceedingly
unusual in RB. (502). Finally, diagnostic anterior chamber paracentesis is
discouraged due to concern of extraocular spread of the tumor but may be
performed by an experienced ocular oncologist in the most difficult cases
such as the diffuse infiltrating anterior RB variant (503).
Juvenile xanthogranuloma (JXG) occurs in infants and young children
with the predominant manifestation being widespread cutaneous orange-red
macules or papules involving the upper body. Ocular involvement is
relatively frequent with iris and ciliary body tumors. The most common
ocular manifestation resulting in “uveitis masquerade” is the development of
an iris nodule or nodules with cells in the anterior chamber. Glaucoma in the
affected eye is also typical, and heterochromia and spontaneous hyphema
may develop. More diffuse intraocular involvement of JXG is exceedingly
rare, and most intraocular lesions will not improve spontaneously and require
treatment due to the risks of glaucoma, recurrent hyphema, and amblyopia.
The use of topical and periocular steroids induce tumor regression without
risk of recurrence (504), and intraocular pressure spikes can be managed with
topical antihypertensive medications. The off-label use of intravitreal
bevacizumab to induce tumor regression has shown some promise in steroid-
refractory cases (505). For eyes in which the tumor is unresponsive to
medical therapy, low-dose radiation (300 to 400 cGy) and/or excisional
biopsy may be considered (506,507); however, surgical resection should be
reserved for only the most resistant cases. The visual prognosis varies in
relation to the development of ocular complications with the poorest
prognoses found in those patients with secondary glaucoma and hyphemas
with corneal bloodstaining.
Extraocular involvement of JXG includes the testes, spleen, lung,
pericardium, bone, and gastrointestinal tract. Treatment for extraocular
disease is usually deferred, as the course is most frequently benign and self-
limited. The vast majority of children present with the disease by age 2, and
approximately 85% of cases with ocular involvement develop within the first
year of life (508,509). The diagnosis of JXG should be considered in any
child with “uveitis” and iris nodules, secondary glaucoma, and/or hyphema in
the setting of suspicious skin lesions. In addition to JXG, important
differential diagnostic considerations of an iris mass include leukemia,
lymphangioma, hemangioma, melanoma, iris leiomyoma, RB, and
sarcoidosis. A biopsy of the skin lesions is diagnostic with identification of
Touton giant cells and foamy histiocytes (510). If skin lesions are absent,
biopsy of an iris nodule can be considered to confirm the diagnosis; however,
both giant cells and foamy histiocytes are fewer in number than those found
in skin specimens (511). Finally, the high frequency of typical skin lesions in
JXG underscores the importance extending one’s examination skills beyond
the eye in sharpening diagnostic acumen.
Nonmalignant entities that can masquerade as uveitis include RDs and
various retinal degenerations, intraocular foreign bodies, ocular ischemic
syndrome, and pigment dispersion syndrome. There are also systemic
infections found predominantly in the immune naive infant or
immunocompromised child that can manifest as endogenous bacterial or
fungal chorioretinitis or endophthalmitis. In most cases, an extraocular focus
can be identified as the primary source of disseminated Candida, Staph
species, Strep species, Pseudomonas aeruginosa, Haemophilus, Klebsiella
pneumonia, and less commonly Neisseria meningitidis (Figure 47-6)
(512,513). Postoperative endophthalmitis is an important consideration given
the guarded visual outcomes with the best treatment and meticulous
preoperative preventative strategies (512).
FIGURE 47-6 Candida endophthalmitis. A young patient
receiving chemotherapy developed Candida albicans
endophthalmitis while neutropenic as evident by the
whitish, chorioretinal lesion with overlying vitritis.
INFECTIOUS UVEITIDES
Toxoplasmosis
Toxoplasmosis is caused by the obligate intracellular protozoan, Toxoplasma
gondii that may be transmitted by the injection of undercooked meats,
contaminated water, fruit, unpasteurized milk, and/or vegetables, or
inadvertent contact with cat feces or litter or soil containing oocysts. The
organism can also be transmitted transplacentally if primary maternal
infection occurs during pregnancy or through inadvertent direct inoculation
from skin puncture, blood transfusion, or organ transplant. In most cases,
infection with the organism is asymptomatic; however, the parasite is
responsible for as much as 70% of pediatric posterior and panuveitis cases
and is the leading cause of posterior uveitis in all age groups (514).
Congenital toxoplasmosis occurs in approximately 0.23 to 0.50 cases per
10,000 live births in the United States; however, the true incidence may
actually be higher due to lower sensitivity rates of newborn screenings and
fetal losses not included in these data (515). The risk of acquisition is greatest
the later in pregnancy the mother develops acute disease with rates
approaching 60% during the third trimester if left untreated (516). Early
treatment of the mother during pregnancy reduces this risk substantially
(517). On the other hand, the severity of disease is inversely proportional to
the gestational age at time of acquisition. Earlier infections can result in
spontaneous abortions or severe congenital disease. Congenital infections
may cause a wide range of findings including peripheral retinal scars, low
birth weight, jaundice, and the classic triad of convulsions, cerebral
calcifications, and chorioretinitis (518). The virulence of the toxoplasma
strain can vary tremendously based on geographic location resulting in a
spectrum of disease severity worldwide (519). Of the three major clonal
lineages initially described, type II is the most common in North America and
Europe but also the least virulent (520). Atypical genotypes such as those
circulating in Southern Brazil have been shown to cause high rates of ocular
involvement (520). In most cases, congenital infection results in a subclinical
infection with ocular and CNS sequelae appearing much later. In long-term
studies of infants with congenital toxoplasmosis, 85% will develop one or
more episodes of chorioretinitis within 4 years resulting in significant visual
impairment and even blindness (521,522). Historically, it was believed that
the vast majority of toxoplasmic inflammatory foci were the reactivation of
congenitally acquired infection (523,524). More recent observations have
challenged this view such that it is currently held that the majority of ocular
toxoplasmosis is acquired postnatally and that ocular disease can present
following this infection without concurrent systemic signs or symptoms
(525,526). These observations not only underscore a paradigm shift in the
pathogenesis of ocular toxoplasmosis but also suggest a change in the focus
of primary preventative strategies that target children and adults, as well as
pregnant women who may be at risk for developing ocular disease as a result
of postnatal infection.
The diagnosis of toxoplasmic chorioretinitis or congenital toxoplasmosis
in most cases is made clinically by the identification of characteristic lesions
on dilated fundus examination, and in the case of congenital disease, other
systemic findings (Figure 47-7). Serologic evidence is used to support the
diagnosis especially when the ocular findings are atypical. Caution should be
exercised when interpreting IgG antibody levels in any neonate as IgG may
cross the placenta, but IgM cannot. This can result in positive passive titers
for over a year and false-positive tests. Elevated IgM and IgA levels within
the first year of life are indicative of prenatal infection and usually decline to
undetectable levels by the child’s first birthday. The presence of IgG in older
children could be the result of either congenital or postnatal acquisition but is
at least indicative of prior exposure. PCR analysis of aqueous or vitreous
specimens can also help confirm the diagnosis, and in congenital disease, be
used to identify the pathogen within the CSF (527–529). The measurement of
intraocular and serologic antibody titers can be used to establish the
Goldmann-Witmer coefficient with a ratio greater than three being
considered diagnostic of intraocular toxoplasmosis (530). Lastly, tissue
culture techniques have also been used successfully to detect tachyzoites in
vitro (531).
FIGURE 47-7 Congenital toxoplasmosis. A large,
pigmented chorioretinal scar within the macula in a patient
with congenital toxoplasmosis with a satellite lesion
superiorly.
Treatment of ocular toxoplasmosis is almost always indicated for newborns,

children with an active-appearing lesion, immunosuppressed patients, or
pregnant women with acquired disease. Early and prolonged treatment during
the first year of life has been shown to greatly reduce recurrence rates in
newborns from undertreated or untreated rates of almost 85% to 4%–13%
following specific antibiotic therapy (519,521,522,532). This reduction in
recurrence occurs even with prolonged intermittent prophylaxis for
Toxoplasma chorioretinitis in adults as shown in a recent double-masked
randomized placebo-controlled trial (533). These data support the notion that
all infants with congenital toxoplasmosis be treated during the first year of
life irrespective of retinochoroidal lesion activity.
The indications for treatment in older children are similar to those in
adults, albeit with a distinctly lower threshold, and include large or multiple
active lesions; those within the macula, abutting the optic nerve or a large
blood vessel; lesions associated with moderate of severe vitritis or
hemorrhage; and those producing moderate decrease in visual acuity (534). In
the immunocompetent host, toxoplasmic retinochoroiditis is a self-limited
process lasting approximately 6 weeks; however, recurrence is quite common
(nearly 80% of cases) (535). The suggested regimen for infants and children
is usually a combination of pyrimethamine, sulfadiazine, and folinic acid to
prevent leukopenia and thrombocytopenia. Clindamycin, azithromycin, and
trimethoprim–sulfamethoxazole (Bactrim) can serve as reasonable
alternatives for sulfadiazine. Atovaquone (Mepron) is yet another option, and
unlike the other antimicrobial agents, has activity against both tachyzoites
and cystic forms of T. gondii. Systemic prednisone (1 mg/kg/d) may be added
24 to 48 hours following the commencement of antimicrobial medication, so-
called triple therapy, in an effort to treat accompanying intraocular
inflammation, especially when a lesion is in the macula and vision
threatening. In a recent double-masked, randomized control trial, quadruple
therapy (the addition of another antibiotic to triple therapy) was employed
and led to faster resolution of chorioretinitis than the triple therapy (536).
Periocular steroids are contraindicated due to localized immune suppression
and the possibility of unabated replication of the organism within the eye.
Topical steroids and cycloplegic agents are used to treat anterior chamber
inflammation. Vitreoretinal surgical techniques may be required to obtain
diagnostic vitreous or retinochoroidal specimens in atypical cases with a
broad differential diagnosis and/or therapeutically to address posterior
segment structural complications, such as epiretinal membrane formation
and/or tractional and/or rhegmatogenous RD related to prolonged intraocular
inflammation.
Overall, the visual prognosis of ocular toxoplasmosis is favorable among
immunocompetent patients with extramacular lesions. Although the disease is
frequently bilateral, it is rarely simultaneously active at least in otherwise
healthy individuals. In contrast, eyes with macular involvement and
immunocompromised patients are at significant risk for visual loss. In a
review of 154 patients with ocular toxoplasmosis, 24% would become blind
in one or both eyes, and bilateral blindness was more common in patients
with congenital ocular disease (535). A more recent prospective study aligns
more closely with anecdotal evidence in which children with congenital
toxoplasmosis who were treated for 1 year and followed for at least another
16 years had a good visual prognosis and a low rate of late ocular
complications (537). In some cases, ocular lesions developed as late as 12
years after birth emphasizing the need for long-term follow-up (537). A more
recent and larger prospective study identified unilateral lesions in 69% of
cases with no vision loss in 80.6% (538). Similarly, in a small study from a
community in Brazil with a recent toxoplasmosis outbreak, ocular
involvement in congenital toxoplasmosis seemed to correlate with an
increased frequency of learning disabilities, but ocular outcomes were
favorable (539). Although visual outcomes are usually good in toxoplasmosic
chorioretinitis, long-term surveillance is necessary due to the risk of late
reactivation of the disease.
Toxocariasis
Toxocariasis is a worldwide zoonotic disease caused by the roundworm
larvae of Toxocara canis or Toxocara cati. The parasitic worms complete
their life cycles within the small intestines of dogs or cats. Transmission to
humans occurs through contact with infected fomites, fecal–oral routes, or
ingestion of contaminated foods, mainly undercooked meats (540). A history
of pica and/or contact with dogs, specifically puppies, can usually be elicited
in children with toxocariasis. Once the eggs have been ingested, the
roundworm larvae hatch and travel through the bloodstream to the brain,
eyes, heart, liver, lungs, and/or muscles. Most people remain asymptomatic
during this time; however, two forms of toxocariasis can occur: visceral larva
migrans (VLM) or ocular larva migrans (OLM). In VML, a self-limited
condition of fever, pulmonary symptoms (cough, bronchospasms, and
asthma), hepatosplenomegaly, and pronounced eosinophilia occurs in young
children (usually 15 to 30 months old) (541). Pronounced eosinophilia is so
common in this disease that some authors have suggested that patients of any
age with a chronic eosinophilia of unknown origin should be evaluated for
toxocariasis (542–544). In those children that develop VLM, OLM is
unlikely to appear as evident by a large, retrospective study that identified
that only 5% of children with VML developed OLM at any point in time
(540). Furthermore, OLM usually presents in older children (7.5 to 8.6 years
of age), and eosinophilia is not a typical feature (545). However, the disease
has been reported in young adults and those up to the age of 77 (546).
Ocular toxocariasis is most commonly a disease of children presenting
with painless, unilateral vision loss, strabismus, and/or leukocoria. Bilateral
ocular involvement is extremely rare but has been reported (547). The
anterior chamber is usually quiet, and the posterior segment findings can vary
depending on the primary anatomical site of involvement. The four forms of
the disease are as follows: peripheral granuloma, posterior pole granuloma
(Figure 47-8), chronic endophthalmitis, and atypical presentations. The
posterior pole granuloma is the most common presentation and found in 44%
of cases, whereas the localized forms (peripheral and posterior pole
granulomas) are believed to arise from cicatricial changes following
resolution of the acute inflammatory phase of the disease (545). A unilateral
pars planitis with extensive inflammatory exudates is an uncommon variant
of the peripheral granuloma (548). Atypical presentations include
neuroretinitis, mobile subretinal nematodes, scleritis, cataracts, anterior
segment involvement, and a diffuse chorioretinitis (546,549).
FIGURE 47-8 Macular granuloma in the left eye of a
child with toxocara chorioretinitis.
The diagnosis of OML is essentially clinical, based on characteristic lesion

morphology, supportive laboratory data, and imaging studies in the
appropriate context. Identification of a retinal granuloma before or after
inflammation is important in narrowing the differential diagnosis. A positive
serum ELISA titer for T. canis is both sensitive and specific for prior
exposure, and the titers significantly decrease over time (550). As such, any
positive titer should be considered abnormal in the proper clinical context.
Although not specific to toxocariasis, elevated IgE levels are also common
(551). Imaging studies, particularly ultrasound examination and computed
tomography, are extremely useful noninvasive modalities in the setting of
media opacification in order to detect the presence of calcification, typically
found in RB, and to define the posterior segment anatomy. Although
exquisitely sensitive for detection intraocular calcification, CT cannot
absolutely differentiate OML or other simulating entities from RB, and there
is significant concern with exposure of children with RB to ionizing radiation
(552–554). Moreover, intraocular calcification, while more typical of RB, is
not uniformly present, and may be seen in eyes with OML with significant
ocular disruption or phthisis (555). In these complex cases where imaging
results are difficult to interpret, more invasive testing may be considered with
the consultation of other specialists, including ocular oncologists (556,557).
Normal aqueous levels of lactate dehydrogenase and phosphoglucose
isomerase and the presence of eosinophils or anti–T. canis antibodies are
more consistent with OLM (556). As RB is the most common malignant
intraocular neoplasm of childhood, it is critically important to distinguish it,
particularly the sporadic, unilateral variant, from OT. The lack of
inflammatory stigmata (anterior segment pathology, cataracts, and cyclitic,
epiretinal, or transvitreal membranes), a younger mean age of presentation
(22 to 23 months for RB vs. 7.5 to 8.9 years for OML), and growth of the
lesion would be more suggestive of RB (545,558). On the other hand, a
migrating lesion would be much more suggestive of OLM (551). As
previously mentioned elsewhere, diagnostic taps of the anterior chamber or
vitreous should be performed with extreme caution by only by an
experienced ocular oncologist when RB is suspected.
There is no uniformly satisfactory treatment for ocular toxocariasis,
which raises the importance of strategies to reduce transmission to humans.
Medical therapy with periocular and systemic corticosteroids is aimed at
reducing the inflammatory response in an effort to prevent the structural
complications of the disease. The use of antihelminthic agents such
thiabendazole, mebendazole, or albendazole is not firmly established for
ocular disease; however, successful treatment of 9 children with OLM with
albendazole has been reported (559). Vitreoretinal surgical techniques can be
used to manage tractional and rhegmatogenous detachments, and laser
photocoagulation of live larvae, if identified, may be considered (560,561).
Some authors have argued for near-complete removal of the granuloma with
subretinal surgical techniques (560). The visual prognosis depends on the
severity and location of the intraocular inflammatory foci; the development of
tractional membranes involving the macula, optic nerve, and ciliary body;
and the presence or absence of CME. In the absence of foveal involvement,
vision at presentation is usually best when the granulomata are located in the
posterior pole as these eyes are less likely than those with peripheral
granulomas to have macular traction. Eyes presenting with endophthalmitis
have the worst vision at presentation; however, the long-term outcomes are
similar among the different clinical presentations (548,562).
Diffuse Unilateral Subacute Neuroretinitis

DUSN is an uncommon but important intraocular nematode infection to
consider in the differential diagnosis of pediatric posterior uveitis and other
nematodes (Figure 47-9A). Early recognition and treatment may preserve
vision (563). The nematode presents in otherwise healthy children of mean
age 14 years (range 11 to 65 years) and is caused by one of two solitary
nematodes that migrate into the subretinal space (564). The smaller worm,
presumably Ancylostoma caninum (the dog roundworm) or Toxocara canis,
is approximately 400 to 1,000 μm in length and is endemic to the
southeastern United States, Caribbean, and Brazil (565). The larger nematode
(1,500 to 2,000 μm in length) is thought to be Baylisascaris procyonis (the
raccoon roundworm) and is found in the northern Midwest and Canada (564).
Isolated cases of DUSN have also been described in Germany, China, and
South Korea (566,567).
FIGURE 47-9 Early- and late-stage DUSN. A:

Ophthalmomyiasis can appear similar to DUSN but is
defined by extensive worm tracts as seen in the
photograph and the worm in the inset. (Image courtesy of
Danielle Wiscombe.) B: DUSN with associated inset
showing magnification of the pathogenic worm. (Image
courtesy of Robin Ray, MD.) C: Late-stage, untreated
DUSN with diffuse pigment changes.
The clinical course of DUSN is characterized by recurrent episodes of focal,

multifocal, and diffuse inflammation of the retina, retinal pigment epithelium
(RPE), and optic nerve with the insidious onset of unilateral visual loss (568).
The earliest stages of the disease are defined by multiple 1,200 to 1,500 μm
grayish-white retinal lesions with moderate to severe vitritis and optic nerve
swelling. These posterior equatorial lesions may be associated with overlying
serous RDs. It is during this phase of the disease that the subretinal nematode
is most easily visualized and may be amenable to laser photocoagulation
(Figure 47-9B) (569). In later stages of the disease, retinal arteriolar
narrowing, optic nerve atrophy, diffuse RPE degeneration, and an abnormal
electroretinogram (ERG) are common (Figure 47-9C). Although very
unusual, cases of DUSN can be bilateral and include the CNS (neural larvae
migrans) (570,571).
The diagnosis is made in the appropriate clinical context and is most
strongly supported by the observation of a subretinal worm. Unfortunately
from both a diagnostic and therapeutic perspective, the worm cannot be
located in most cases. Laboratory tests are unremarkable and changes in the
ERG are not diagnostic as they may occur even early in the disease.
Interestingly, OCT angiography may help visualize the worm when it cannot
be readily identified clinically due to a presumable movement artifact of the
worm (572). Treatment in the early stages of the disease with direct laser
photocoagulation of the worm is usually curative and leads to improvement
in visual acuity (573). Fortunately, applying laser burns directly onto the
nematode does not usually induce a significant worsening of intraocular
inflammation (573).
Medical therapy with corticosteroids may achieve transient improvement
in inflammatory control; however, recurrence of inflammation and
progression of visual loss is typical. The use of intravitreal triamcinolone in
the presence of significant intraocular inflammation has been suggested to
improve visualization of the nematode and permit laser photocoagulation
(574). Initial experience with antihelminthic therapy was disappointing;
however, successful treatment with oral thiabendazole (22 mg/kg twice daily
for 2 to 4 days with a maximum dose of 3 g) has been reported in patients
with moderate to severe inflammation. Treatment with albendazole (200 mg
twice daily for 30 days) may be a better-tolerated alternative. Immobilization
of the subretinal nematode has been observed following systemic
antihelminthic therapy, and so it has been recommended that patients with
DUSN in whom the worm cannot be initially identified receive a course of
such therapy in order to maximize the chances of identifying and treating the
offending organism (575). Uncommonly, a second nematode, presumably
from reinfection, may occasionally be observed in patients who have
undergone successful previous photocoagulation of a subretinal worm. These
patients may also benefit from a course of systemic antihelminthic therapy,
particularly if inflammation does not abate promptly following laser
photocoagulation. Finally, close follow-up is important as there is a small
chance that repeated application of laser to the worm may be required (576).
Herpes Simplex Viruses

Herpes simplex virus (HSV) belongs to a family of encapsulated and
enveloped, double-stranded, epitheliotropic DNA viruses. Clinically relevant
strains include human herpesvirus (HHV)-6, -7, and -8; cytomegalovirus
(CMV); varicella–zoster virus (VZV); and Epstein-Barr virus (EBV)
(577,578). HSV, CMV, VZV, and EBV are important causes of pediatric
uveitis and will be discussed in further detail (51). There are two distinct
antigenic variants: HSV-1, which commonly causes orofacial lesions and
encephalitis, and HSV-2, which is sexually transmitted and can cause
recurrent genital ulcers (579). There has been significant crossover in
presentation making the viruses more difficult to distinguish clinically as
shown by HSV-1 accounting for nearly half of new genital herpes cases
diagnosed each year in the developed world (580). Both viruses are capable
of establishing latency in sensory ganglia and reactivating to cause distant
vesicular eruptions within the innervated dermatome or other neurologic
complications (581,582). While the distinction between the two viruses has
been traditionally important from a public health perspective, they share
similarities in pathogenesis, clinical course, and treatment.
Perinatal exposure to HSV has become increasingly common with a
retrospective study suggesting rates as high as 1 in 3,200 births (583).
Transmission from mother to child can occur in utero (5%), during the
peripartum period (85%), or postnatally (10%) (583). Transplacental
infection is believed to be due to viremia in the mother during pregnancy
with highest rates of transmission during the first 20 weeks of gestation.
Consistent with newly diagnosed genital herpes cases, a majority of perinatal
herpes infections are due to HSV-1, and there is an increased risk of
transmission with premature rupture of membranes, vaginal delivery, positive
HSV-1 or -2 PCR results from the cervix or genitalia, invasive
instrumentation, the use of a fetal scalp electrode, and primary genital
infections (583,584). Postnatal transmission is thought to occur by direct
contact with individuals who have an active oral–labial lesion or other
cutaneous vesicle or are asymptomatic viral shedders.
The ocular and systemic manifestations of perinatal HSV infections can
vary tremendously from one infant to another. Systemic findings from in
utero exposure include higher rates of spontaneous abortion and prematurity,
growth and psychomotor retardation, microcephaly, hydranencephaly,
intracranial calcifications, encephalitis, seizures, pneumonitis, hepatomegaly,
aplasia cutis, and recurrent vesicles at birth (585,586). Ocular findings occur
in approximately 13% of infants and include keratitis, vitritis, chorioretinitis,
chorioretinal scars, and developmental malformations (microphthalmia, optic
atrophy, retrocorneal masses, lenticular opacities, and retinal dysplasia)
(Figure 47-10A) (585,587–589). Ocular involvement is the sole finding in
approximately a third of cases (590). The timing of in utero exposure has
been hypothesized to affect the severity and presentation of the ocular
complication(s) much like other congenital infections. Exposure during the
first trimester is associated with teratogenic complications, second trimester
infections with inactive chorioretinal scarring, and third trimester acquisition
being indistinguishable from active, neonatally acquired disease discussed
below (590).
FIGURE 47-10 Herpetic-related ocular disease. A:
Diffuse neonatal acute retinal necrosis. (Image courtesy of
Andrea Zinn, MD, PhD.) B: Acute retinal necrosis with
confluent peripheral retinitis, retinal vasculitis, and
intraretinal hemorrhage. C: The classic presentation of
CMV retinitis in the posterior pole with white retinitis and
intraretinal hemorrhages located predominantly along the
vascular arcades and cotton-wool spots. D: Peripheral
frosted branch angiitis associated with CMV retinitis.
E:Epstein-Barr virus–related peripheral retinitis and
vasculitis.
Infants with natal or postnatal herpes infections can be classified into one of
three groups: those involving the skin, eye, and/or mouth; those involving the
CNS; or disseminated disease. Forty-five percent of cases are isolated to
mucocutaneous sites, 30% to the CNS, and the remaining 25% to
disseminated variants (585). CNS involvement may include any of the
following: lethargy, body temperature instability, irritability, seizures, and a
bulging fontanelle with untreated mortality rates as high a 50% (591).
Disseminated disease usually presents with a clinical presentation similar to
that of sepsis with ongoing respiratory and hepatic failure, lethargy, and
disseminated intravascular coagulation and can affect almost any organ
system. Untreated disseminated disease has a mortality rate as high as 85%
with only 50% of those who survive having normal neurologic development
(591). The use of high-dose intravenous acyclovir (60 mg/kg/d) has reduced
mortality rates to 29% in disseminated disease and 4% with CNS infections.
High-dose intravenous acyclovir also substantially improved neurologic
outcomes (592). Ocular manifestations of neonatally acquired HSV include
corneal ulcers and even perforation, anterior uveitis, cataracts, vitritis,
chorioretinitis, optic atrophy, and rarely, persistent fetal vasculature and acute
retinal necrosis (ARN) (589,593,594).
The diagnosis of intrauterine and perinatal herpetic infection is made in
the appropriate clinical context and by isolation of the virus from vesicular
fluid, nasal or conjunctival secretions, whole blood, or cerebral spinal fluid. It
is important to differentiate quiescent or active ocular HSV from other
nonherpetic infectious diseases such as Toxoplasma, rubella, and CMV
infections as the infection and its associated findings may or may not be
present at the time of birth. Diagnosing reactivated ARN in a child may be
especially difficult in cases where there is nonspecific peripherally
chorioretinal scarring with concurrent inflammation. For these reasons, a past
medical history of a neonatal ocular or systemic herpes infection is a key
component to make the correct diagnosis (595). In unclear cases, a diagnostic
paracentesis of the anterior chamber or vitreous biopsy should be performed.
The drug of choice for disseminated (including HSV retinitis) or CNS-
involving HSV infections in neonates is intravenous acyclovir at a dose of 30
to 60 mg/kg/d for 10 days to 4 weeks depending on the age of the child.
Nephrotoxicity is the most frequently encountered side effect of systemic
acyclovir and is more common with doses above 15 mg/kg/d (596). There is
emerging evidence of HSV strains resistant to acyclovir, so an incomplete or
poor response to the drug should warrant further questions including
reevaluating the diagnosis and the possibility of resistance (597). In cases of
ARN, oral valacyclovir at 20 mg/kg may obviate the need for multiple rounds
of intravenous medications (598,599). Vidarabine, which is an equally
effective alternative to acyclovir, may also be administered as a single
intravenous infusion, 15 to 30 mg/kg/d over a 12-hour period, with potential
side effects including hepatic toxicity and bone marrow suppression (600). In
cases in which ARN worsens despite intravenous acyclovir, intravenous or
intravitreal foscarnet or cidofovir are effective alternatives (601).
Varicella–Zoster Virus
VZV is another herpes virus family member that causes two distinct diseases:
a primary cutaneous infection that is usually relatively benign but extremely
contagious during childhood (chickenpox) or a reactivation of the virus
within a sensory dermatome resulting in an eruption of painful, cutaneous
vesicles in older adults (herpes zoster, shingles). Vaccines are currently
available to induce immunity and boost waning vaccine-induced immunity to
the virus (602,603). Ocular-related complications with primary infections or
reactivation of the virus in children include orbital myositis, ARN, keratitis,
orbital apex syndrome, and dacryoadenitis (604).
Congenital varicella is rare, and if acquired within the first 20 weeks of
gestation, can cause congenital varicella syndrome (CVS) in approximately
2% of infants (605,606). The syndrome is defined by one or more of the
following; low birth weight, hypertrophic cicatrix of the skin, hypoplastic
limbs and digits, cortical atrophy, ventriculomegaly, microcephaly, and/or
psychomotor retardation. These children are at risk for developing
intracerebral VZV reactivation later in life (607). If maternal disease should
develop 5 days prior to delivery, mortality may be as high as 50% due to
fulminant systemic disease in an infant with a rudimentary immune system
and the absence of maternally supplied antibodies (608). Ocular findings in
CVS include congenital cataracts, unilateral or bilateral microphthalmia,
pendular nystagmus, chorioretinal scarring, atrophy and hypoplasia of the
optic nerve, Horner syndrome, and bulbar palsies (609,610). Chorioretinal
scarring consists of a central area of pigment clumping surrounded by a halo
of hypopigmentation with normal-appearing retina between scars and is
usually found in the macula or peripheral retina of one eye. The scars may
resemble those seen with congenital rubella, syphilis, and inactive
toxoplasmosis, especially in cases with more posterior involvement. Active
chorioretinitis has never been reported in cases of CVS as the chorioretinitis
has likely resolved by time of delivery.
Congenital disease, primary infections, and reactivation of VZV in
children can result in necrotizing retinitis in which disease severity varies
depending on both host and viral factors (611). ARN syndrome was first
described in 1971 as part of a spectrum of necrotizing herpetic retinopathies
(612,613) (Figure 47-10B). In immunocompetent children, the development
of peripheral retinitis, vasculitis, iridocyclitis, and vitritis defines ARN. A
variant of ARN, progressive outer retinal necrosis (PORN), is seen in the
immunocompromised and is unique in that it may affect the macula initially
with a paucity of intraocular inflammation (614). Although the spectrum of
necrotizing herpetic retinopathies is more common in adults despite
acquisition of the virus during childhood, there are a few case reports in
children with the youngest patient at time of diagnosis being 4 years old
(615–618).
The diagnosis of CVS is essentially clinical with the recognition of the
characteristic defects in the neonate and a history of maternal varicella. In
cases of inactive chorioretinal scars and with an unclear history, other
TORCH (toxoplasmosis, others, rubella, CMV, and HSV [“others” include
syphilis, VZV, EBV, HIV, and lymphocytic choriomeningitis virus
(LCMV)]) infections should be considered. Serologic testing of the infant and
mother may then be pursued based on the clinical history and examination to
identify the offending pathogen. In cases of VZV, the diagnosis is also
clinical with the appearance of the typical vesicular lesions that respect the
midline in the setting of prior exposure. The treatment for CVS is directed
toward reducing the risk of acquisition and by the administration of exposure
prophylaxis in nonimmune mothers and neonates with varicella–zoster
immune globulin (619,620). In infants that acquire the virus after birth,
acyclovir dosed at 30 to 100 m/kg/d should be administered for 5 to 8 days
(621).
Cytomegalovirus
CMV is another extremely common herpes virus that establishes latency
within monocytes with characteristic large intracellular inclusion bodies
(622). It is the most common congenital viral infection affecting one out of
every 200 live births in the United States as reported by the Centers for
Disease Control and Prevention (CDC). Transmission may occur prenatally
during maternal viremia secondary to primary infection or reinfection or due
to reactivation of a latent infection while exposure to genital secretions
during delivery or ingestion of breast milk may produce viral infection during
the natal and postnatal periods (623). A majority of congenital CMV
infections are asymptomatic or subclinical, but intrauterine growth
retardation, thrombocytopenic purpura, jaundice, hepatosplenomegaly,
pneumonia, microcephaly, seizures, and sensorineural hearing loss can occur
(624). The most common intraocular manifestation of congenital CMV is
chorioretinitis with subsequent scarring similar to that seen in toxoplasmosis
(625). As in the adult, CMV retinitis may present as one of three common
variants in children: classical or fulminant retinitis, a peripheral granular
retinitis, and a perivasculitis. In the classic form, there are large areas of
retinal hemorrhage against a background of whitened, edematous, and
necrotic retina, typically appearing in the posterior pole, from the disk to the
vascular arcades in the distribution of the nerve fiber layer (Figure 47-10C).
The granular variant is more often peripheral and characterized by less retinal
edema and hemorrhage with active retinitis appearing at the borders of the
lesion having a “brush fire” appearance. In the perivascular form often
described as a variant of frosted branch angiitis, a retinal perivasculitis
initially described in immunocompetent children, there is predominance of
retinal periphlebitis seen in association with peripheral retinitis (Figure 47-
10D). It is important to recognize these variations in presentation of CMV
retinitis, as the diagnosis is essentially clinical and made in the context of
immunosuppression. Strabismus, cortical visual impairment, nystagmus, and
optic nerve atrophy/coloboma/hypoplasia are other ocular complications of
the disease (626). In this study, symptomatic patients experienced more
ophthalmologic sequelae and significantly worse visual outcomes than did
asymptomatic CMV and control patients. Risk factors of severe visual
impairment were symptomatic status, optic nerve atrophy, chorioretinitis,
cortical visual impairment, and sensorineural hearing loss (626). Ocular
findings are also much more commonly associated with cases of systemic
disease but have also been described in isolation (627,628).
The diagnosis of congenital disease is suggested by clinical appearance of
chorioretinal scarring in combination with inclusion bodies or positive real-
time (RT)-PCR results of urine, saliva, blood, and/or subretinal fluid (629).
RT-PCR and other emerging PCR methods have supplanted more traditional
virus isolation techniques from tissue samples that are both more expensive
and labor intensive (630). Complement-fixation tests for CMV-specific IgM
may be helpful after loss of maternal antibodies that occurs approximately 6
to 12 months after birth. Treatment of clinically significant CMV requires
intravenous ganciclovir and/or foscarnet; however, once therapy is
discontinued, viral shedding in urine and saliva reconvenes (631). In cases
where systemic therapy is contraindicated, intravitreal ganciclovir can be
used for local control (632).
Epstein-Barr Virus
EBV is a ubiquitous double-stranded DNA herpesvirus that is transmitted
through bodily fluids, mainly the saliva, and commonly causes a flu-like
illness in childhood (633). If acquired during adolescence or adulthood, EBV
is most commonly associated with infectious mononucleosis (IM), which
causes a systemic illness with fever, lymphadenopathy, splenomegaly, and
extreme fatigue (633). The virus establishes latency in B lymphocytes where
it may cause cellular transformation and has been implicated in the
pathogenesis in various types of cancer (i.e., Burkitt and Hodgkin
lymphomas, and nasopharyngeal and gastric carcinomas) (634). Ocular
manifestations of EBV may arise from congenital infections, or more
commonly, during primary infection in the context of IM (635). Congenital
cataract has been reported in association with congenital EBV infection
(636). The most common ocular manifestation of IM is a self-limited
follicular conjunctivitis during the early stages of the disease (637).
Episcleritis or stromal or epithelial keratitis may also occur. Less commonly,
a severe, bilateral granulomatous iridocyclitis may appear (638,639). Less
frequent complications of IM include dacryoadenitis, Parinaud
oculoglandular syndrome, conjunctival masses, and cranial nerve palsies
(637,640).
Various posterior segment abnormalities may arise secondary to EBV
including optic disk edema or optic neuritis, vitritis, retinal hemorrhages,
retinitis, choroiditis, and macular edema (Figure 47-10E) (638,641–649).
Case reports of children with primary EBV infections have also described
“punctate outer retinitis” and progressive subretinal fibrosis and uveitis with
secondary choroidal neovascularization (642,643). Most, if not all, of these
cases were associated with clinical IM; however, there are isolated reports,
mainly in adults, of anterior or posterior uveitis without clinical IM. Wong et
al. described three patients ranging from 15 to 30 years of age with serologic
evidence of chronic EBV infection that developed bilateral uveitis, ranging
from steroid-responsive granulomatous anterior uveitis to severe panuveitis,
cataract, optic disk swelling, macular edema, and chorioretinal scarring (644).
Usui and Sakai reported three otherwise healthy young adults with elevated
EBV viral capsid antibody titers in the aqueous humor who presented with a
viral prodrome with the subsequent development of acute, bilateral, fibrinous,
anterior uveitis followed by a chronic, granulomatous inflammatory reaction
marked by disk hyperemia, minimal vitritis, and the development of an early
“sunset glow” fundus. The ocular changes were responsive to systemic
steroids resulting in good visual outcomes in all three cases (639). Finally,
EBV has been implicated but not substantiated in the pathogenesis of certain
uveitic syndromes, including multifocal choroiditis and panuveitis (MCP)
(645,646).
The diagnosis is made clinically and based on serologic analysis of
antibodies against a variety of EBV-specific capsid antigens. During IM, the
VCA IgM antibody, followed by the VCA IgG antibody, rises during viral
incubation 4 to 5 weeks following exposure to the virus to a maximum IgG
titer approximately 120 days after symptom onset. The early antigen (EA)
rises with the onset of clinical disease usually within 5 to 10 weeks of
exposure and rises to a maximum approximately 2 months after exposure.
The antibody titers fall to undetectable levels 6 to 12 months after resolution
of the infection. The EBV nuclear antigen antibody appears slowly within 2
months and persists for life, whereas the VCA IgM titer falls to undetectable
levels after resolution of the infection. The diagnosis of chronic EBV
infection is best supported by abnormally elevated anti-VCA IgM and anti-
EA antibody levels (647,648). Most ocular disease is self-limited and does
not require treatment. However, the presence of iridocyclitis may necessitate
the use of topical corticosteroids and cycloplegia, and systemic steroids may
be required to treat posterior segment inflammation. Little is known
regarding the efficacy of antiviral therapy for ocular involvement, and their
use is controversial even in severe systemic infections among
immunocompetent patients (650).
Rubella
The rubella virus is an enveloped, single-stranded RNA virus of the
Togaviridae family that is highly contagious and spread through aerosolized
droplets to cause rubella (German measles). In most cases, the virus causes a
mild illness of a low-grade fever, sore throat, conjunctivitis, and
maculopapular rash that starts on the face and spreads to involve the patient’s
entire body. The disease is exceedingly rare in the United States due to
widespread vaccination programs (651). In adults, the demonstration of
rubella virus in the aqueous humor of patients with Fuchs uveitis syndrome
and the concurrent decline in its incidence concurrent with widespread
vaccination against the virus in the United States has implicated it in the
pathogenesis of this syndrome (652–654). In contradistinction, congenitally
acquired rubella is associated with more significant morbidity including the
classic triad of sensorineural deafness, eye abnormalities, and congenital
heart defects. The risk of congenital birth defects is greatest if the virus is
acquired during the first trimester. Ocular findings include unilateral or
bilateral pigmentary retinopathy (“salt-and-pepper”) in 25% to 50%, cataracts
in 15%, and glaucoma in 10% (655,656). The salt-and-pepper fundus can
range from finely stippled, black, bone spicule-like irregularities to gross
pigment changes with coarse mottling. These posterior segment findings may
be stationary or progressive. Even with extreme pigmentary abnormalities
and loss of the foveal light reflex, vision and ERG usually remain unaffected.
Congenital cataracts, the development of glaucoma, and microphthalmia are
the most common causes of vision loss (657).
Rubella is part of the TORCH group of pathogens, and thus, an important
cause of congenital disease. A detailed clinical history including pregnancy
history, serologic evidence from both the mother and child, and retinal
examination findings are important in differentiating the various TORCH
pathogens and in the diagnosis of the congenital rubella syndrome. A
fourfold increase in antirubella IgG titers between paired sera 1 to 2 weeks
apart or the emergence of IgM titers is diagnostic for an ongoing or recent
rubella infection (655). Cord blood may be used to confirm the diagnosis, as
the fetus is capable of mounting an immune response to the virus. Treatment
of rubella is guided by the specific ocular findings and symptoms including
both surgical and medical treatment of glaucoma and cataract surgery.
Lymphocytic Choriomeningitis Virus

LCMV is an underrecognized fetal teratogen with potential devastating
sequelae, including intrauterine death, hydrocephalus, and chorioretinitis. It is
listed among “others” in the TORCH group of congenital infections. LCMV
is an enveloped, negative-sense RNA virus of the Arenaviridae family with
rodents being the primary hosts and reservoirs for the virus, especially
common house mouse and hamsters. It is transmitted to humans by
aerosolized rodent saliva, urine, or feces or through contaminated food and
fomites (658). Contraction of the virus most commonly manifests as an
asymptomatic infection or a mild febrile illness. Symptomatic disease
typically has 2 phases: a flu-like illness within 8 to 13 days of exposure,
followed by the development of a meningoencephalitis (659). Fortunately,
fatalities are rare (<1%), and a full recovery is the norm (659). Congenital
LCMV arises through transplacental infection of the fetus during maternal
viremia producing sequelae, the seriousness of which are inversely related to
gestational age: first trimester infections result in spontaneous abortion and
infections during the second and third trimesters produce permanent birth
defects as part of the TORCH group of congenital infections (660). Women
with infected infants have been noted to reside in substandard housing or
domiciles where they may be at greater risk for exposure to rodents during
pregnancy (661). Interestingly, individuals with an occupational exposure to
mice are also apparently at greater risk for acquiring LCMV as evidenced by
the 2012 outbreak in the United States (662).
Congenital LCMV was first reported in 1955 in England (663). Since that
time, there have been 75 total cases described in the literature, 47 of those
occurring in the United States (664). Unlike acquired cases that cause little to
no long-term effects, congenital LCMV has a mortality rate as high as 30%
and survivors often have profound neurologic deficits (665,666). Literature
reviews of these cases have identified macrocephaly, microcephaly,
hydrocephalus, and intracranial calcifications as the most common systemic
findings of congenital LCMV (666–668). Other neurologic manifestations
such as mental retardation, seizures, and visual impairment are much more
common than hearing loss and/or hepatosplenomegaly (666). In contrast to
the other TORCH infections, LCMV is rarely accompanied by the ocular
and/or systemic stigmata of other congenital infections such as the
hepatosplenomegaly of CMV; cataracts, cardiac disease, and deafness with
rubella; or bony and/or hepatic abnormalities of syphilis.
The most common ocular finding of congenital LCMV are chorioretinal
scars that predominantly involve the peripheral retina but also can involve the
macula. These consist of alternating patches of hyperpigmentation and
atrophy most closely resembling those seen in congenital toxoplasmosis but
distinct from the salt-and-pepper retinopathy of rubella and syphilis (669).
The areas of chorioretinal scarring and dysgenesis of the corpus callosum
may also resemble that of the X-linked dominant Aicardi syndrome
(664,670). An important distinction between toxoplasmosis and LCMV is the
location of intracerebral calcifications. In toxoplasmosis, the calcifications
are diffuse, whereas in LCMV they tend to be isolated to periventricular
locations (669). Other less common ocular findings include bilateral optic
nerve atrophy, nystagmus, and eso- or exotropia from profound vision loss.
Unilateral microphthalmia and cataract formation have also been described.
Isolated ocular findings in congenital LCMV are rare, but chorioretinal
scarring as the only manifestation of disease has been reported in two
children (663). Any child with unexplained hydrocephalus, micro- or
macrocephaly, intracranial calcifications, chorioretinal scars, and nonimmune
hydrops should be evaluated for congenital LCMV (671). Although the
aforementioned clinical findings may help differentiate many of the TORCH
infections, serologic testing is essential in establishing a specific diagnosis.
The diagnosis of congenital LCMV is made by confirmatory serologic
testing of the mother and infant using immunofluorescent antibody (IFA)
tests and Western blots or ELISA-based techniques to detect IgM and IgG
antibodies (671,672). Virus can also be detected by PCR-based assays (672).
The differential diagnosis of congenital LCMV includes the pathogens
compromising the TORCH mnemonic. The use of ribavirin has shown anti-
LCMV activity in vitro, but there is no evidence to support its routine use
clinically for congenital or acquired infections, so treatment is largely
supportive (659). Recombinant LCMV vector-based vaccines are currently in
development but have shown mixed results in mouse models (673–675).
Measles Virus
The measles virus is a highly contagious, enveloped, single-stranded RNA
virus that is transmitted either directly or via aerosolization of
nasopharyngeal secretions to mucus membranes of the conjunctiva or
respiratory tract of susceptible individuals. Humans and monkeys are the only
natural hosts. Acquired infection results in a self-limited prodromal phase
exemplified by high fever, fatigue, cough, coryza, and conjunctivitis, which
is followed by a maculopapular rash that develops first on the face and
quickly spreads centrifugally to the rest of the body (676). The early signs
and symptoms of cough, conjunctivitis, and coryza, define the classic triad of
measles. Clustered, blue-white lesions surrounded by red areolae develop on
the buccal mucosa (Koplik spots) or on the conjunctiva (Hirschberg spots)
within a few days and just prior to the onset of the rash. Patients are
contagious 4 days prior to the rash and remain contagious 4 days after the
rash appears. Measles can also be transmitted transplacentally from the
infected mother. However, the disease is usually one of childhood and a
vaccine against the disease had nearly eradicated the disease from the United
States until recently, when some parents have refused to vaccinate their
children (677,678). This has led to the reemergence of the disease and a
localized loss of herd immunity (677).
The most common ocular manifestations of measles are mild, papillary,
nonpurulent conjunctivitis and epithelial keratitis. In the United States, both
the keratitis and conjunctivitis resolve without sequelae; however, in other
parts of the world, “postmeasles blindness” can occur with corneal
complications leading to severe visual impairment. Measles retinopathy may
also occur and is more common in acquired cases of the disease. Retinopathy
manifests 6 to 12 days after the appearance of the exanthema in the presence
or absence of encephalitis. Ophthalmic findings include attenuated arterioles,
scattered retinal hemorrhages, diffuse retinal edema, a macular star, blurred
disk margins, and clear media (679). Following resolution of systemic and
ocular symptoms, optic disk pallor, persistence of arteriole attenuation with
or without perivascular sheathing, and a pigmentary retinopathy with either a
“bone spicule” or “salt-and-pepper” appearance develop in the posterior pole.
A retinopathy of pigmented paravenous retinochoroidal atrophy with visual
field and ERG abnormalities developing years later has also been described
as long-term sequela (680).
In patients with acute retinopathy from measles, fluorescein angiography
is likely to show diffuse leakage from retinal edema. In the resolving phases
of the disease, generalized choroidal hyperfluorescence is found due to retinal
pigment epithelial disruption (680). Widespread retinal dysfunction results in
an extinguished ERG during the acute phase. Similarly, visual field changes
may include severe, generalized constriction, ring scotomas, or small
peripheral, residual islands of vision. Useful vision and ERG activity may
return following resolution of acquired measles retinopathy; however, the
visual prognosis is guarded (679,680). In one study, 43.7% of students in
blind school institutions were severely visually impaired as a consequence of
measles infection (681).
The diagnosis of measles and attendant ocular sequelae is made clinically
by observation of the sequence of symptoms. The virus may be recovered
from the nasopharynx, conjunctiva, lymphoid tissues, respiratory mucus
membranes, urine, and blood for a few days prior to the appearance of the
rash and for several days thereafter (682).
Leukopenia is common during the prodromal phase of the disease. A
variety of tests are available for serologic confirmation of measles infection,
including complement fixation, ELISA, immunofluorescent and
hemagglutination inhibition assays. The differential diagnosis of measles
retinopathy includes VKH syndrome, toxoplasmic retinochoroiditis, central
serous chorioretinopathy, retinitis pigmentosa, Leber stellate idiopathic
neuroretinitis, and other viral retinopathies. These diseases can be readily
distinguished with a detailed patient history, differences in clinical findings,
and serologic testing.
The treatment of measles is supportive as the disease is generally self-
limited. In certain high-risk populations such as the immunocompromised,
children under 1, and pregnant women, gamma globulin administered within
5 days of exposure may prevent the disease, and its use was supported by a
recent Cochrane review (683,684). Likewise, the ocular manifestations of
measles are treated symptomatically with topical antiviral or antibiotics to
prevent secondary infections in patients with keratitis or conjunctivitis.
Consideration should be given to the use of systemic corticosteroids for cases
of acute measles retinopathy. Finally some have suggested that in children
with severe cases of measles, intramuscular vitamin A should be provided to
prevent vision loss; however, there are insufficient data to support this
recommendation (685).
Late Complications of Measles: Subacute Sclerosing

Panencephalitis
Subacute sclerosing panencephalitis (SSPE) is a rare, late complication of
acquired measles infection during childhood with devastating neurologic
manifestations including encephalomyelitis and SSPE. Previously, it was
thought to be relatively uncommon; however, recent evidence has shown that
it occurs in 1 out every 609 children in the United States infected with
measles before their first birthday (686). Worldwide, SSPE is more common
due to lower vaccination rates with some areas reporting an annual incidence
>100/million/year (687). While still rare in the United States, SSPE usually
arises in unvaccinated children 6 to 8 years after the primary infection and is
characterized by the development of visual impairment, behavioral issues,
and memory loss. The symptoms progress to myoclonus, spastic
quadriparesis, dementia, and death within 1 to 3 years.
Ocular findings are found in 10% to 50% of patients and may precede
neurologic changes by several weeks to years (688,689). The most common
ocular finding is a focal macular retinitis found in 36% of patients with
minimal vitritis followed by mottling and scarring of the RPE as the retinitis
resolves (Figure 47-11) (690–694). The retinitis may progress peripherally
within days. Many other ocular findings have been reported and include optic
nerve edema, optic neuritis, optic atrophy, small intraretinal hemorrhages,
macular edema, macular pigment epithelial disturbances, whitish retinal
infiltrates, serous macular detachments, drusen, preretinal membranes, gliotic
scars, macular holes, cortical blindness, hemianopsia, horizontal nystagmus,
extraocular muscle paresis, and ptosis (693–700). Fluorescein angiography
typically reveals optic nerve staining, precapillary arteriole and postcapillary
venous occlusion involving the vessels surrounding the macula, focal defects
in the RPE in the absence of choroidal exudates, and cystic areas of
hyperfluorescence and RPE window defects (693,698).
FIGURE 47-11 Subacute sclerosing panencephalitis–
associated macular retinitis. A: Fundus photograph
showing macular scarring and (B) OCT through the scar
shows extensive full-thickness loss of the retina and cystic
spaces. (Images courtesy of Nuriye Ilknur Tutkun, MD.)
The diagnosis of SSPE is made based on clinical manifestations, a history of
previous measles infection, characteristic electroencephalographic
discharges, and elevated antimeasles IgG antibody titers in the serum and
CSF of the afflicted. MRI can be normal early in the disease, but diffuse
white matter changes develop as the disease progresses (701). In cases where
the diagnosis remains unclear, a brain biopsy may be necessary.
Histopathologic findings consistent with panencephalitis with the
demonstration of Cowdry type A inclusion bodies in the brain or retina may
help confirm the diagnosis (700,702,703). The differential diagnosis of the
posterior segment findings associated with SSPE include those seen with
necrotizing viral retinitis due to HSV, VZV, and CMV as well as those with
MS. In contrast to SSPE, MS is not a panencephalitis, with MRI imaging
revealing focal periventricular white matter lesions in the latter. Furthermore,
CME and retinal vasculitis are prominent features of MS but have been rarely
reported in patients with SSPE (689,704). Significant RPE changes within the
macula can mimic toxoplasmosis scars (705). The most effective treatment
for SSPE is currently unknown. The use of intraventricular interferon-α with
or without oral Isoprinosine (inosiplex) shows the most promise if started
early in the disease process; however, lifelong therapy is required (690).
Unfortunately, there is variability in response to treatment, relapse is
common, and medication-associated toxicity is not uncommon (690,706).
Emerging Pathogens: West Nile, Dengue, Zika,

Chikungunya, and Ebola Viruses
There are several emerging pathogens that are worth briefly mentioning
although there may not be specific case reports of ocular involvement in
children. The diagnosis of these viruses can be complicated due to cross-
reactivity among them with IgM-related tests and the potential for
coinfections with other viruses. The differential diagnosis of each virus
includes the other viruses in this section. Similar methods for detection are
employed for each of the viruses and include immunoassays to detect IgM
and IgG, plaque-reduction neutralization tests, viral cultures, reverse
transcriptase PCR (RT-PCR), and immunohistochemistry. Pan-organism
PCR may help in cases where multiple pathogens could be responsible for a
particular clinical presentation (707). In most cases, multiple tests are
performed concurrently to confirm the diagnosis and repeat testing may be
required to rule out the disease, as a single negative test may not be
sufficient.
West Nile virus (WNV), dengue fever (DF), and Zika virus are all
positive-sense, single-stranded flaviviruses transmitted by mosquitoes that
can cause similar symptoms. WNV is a neuroinvasive virus maintained in an
enzootic cycle that was first isolated in Uganda in 1937 and is now endemic
to North and South America, Europe, Asia, Australia, and Africa. The virus
gained significant notoriety in 1999 when an extremely neurotropic outbreak
was first reported in New York City and quickly spread throughout the
United States and Canada in which 51% of the reported 663 cases manifested
as either meningitis or encephalitis with a mortality rate of 4.5% (708).
Fortunately, the vast majority of WNV infections are subclinical (80%) or
self-limited, febrile flu-like illnesses (709). Risks factors associated with
more severe neurologic disease include older age at presentation and
diabetes.
The most common ocular manifestation in WNV is a self-limited,
asymptomatic, multifocal choroiditis (Figure 47-12). Active chorioretinal
lesions (200 to 1,000 μm in size) appear whitish to yellow in color; are flat
and deep, and evolve with varying degrees of pigmentation and atrophy.
They typically occur in linear clusters within the midperipheral retina
following the course of retinal nerve fibers and are more frequently seen in
patients with concurrent severe neurologic involvement (710). Other less
common ocular findings include anterior uveitis, intraretinal hemorrhages,
vitritis, disk edema, optic atrophy, and, less commonly, retinal vascular
sheathing and occlusion, and VI nerve palsies (709). Intrauterine exposure to
the virus has been reported to result in chorioretinal scarring (711).
Fluorescein angiography (FA) reveals central hypofluorescence and late
staining of active lesions and early hyperfluorescence with late staining of
inactive lesions. Many inactive lesions exhibit a characteristic target-like
appearance angiographically with central hypofluorescence from blockage of
fluorescein dye by pigment and peripheral hyperfluorescence due to atrophy
(Figure 47-12). ICG angiography reveals hypofluorescent spots that are more
numerous than those seen on FA or funduscopy (712). The presence of this
unique pattern of chorioretinal lesions in patients with a febrile illness and
systemic symptoms suggestive of WNV can help establish the diagnosis
while serology is still pending. Conversely, a complete ocular evaluation,
including dilated funduscopy and FA, may be very helpful in suggesting the
diagnosis of WNV infection in patients presenting with meningoencephalitis.
FIGURE 47-12 West Nile virus–associated

chorioretinitis. Late-phase fluorescein angiogram of the
right eye in a patient with WNV-associated chorioretinitis
demonstrating typical targetoid appearance of subacute
lesions with central hypofluorescence and peripheral
hyperfluorescence. Note linear clustering of lesions within
the inferotemporal arcade.
Commercially available immunoassays to detect WNV-specific IgM titers are

the most commonly serologic tests employed; however, a negative result does
not rule out the diagnosis, and repeat testing is required such as early in the
disease course when antibodies may not be detected or when clinical
suspicion remains high. The differential diagnosis includes syphilis, MCP,
histoplasmosis, sarcoidosis, and TB. All of these infectious diseases may be
distinguished based on history, systemic signs and symptoms (or lack
thereof), serology, and the pattern of chorioretinitis. Unfortunately, there is
no proven therapy for systemic or ocular infections with WNV, but case
reports have suggested that interferon-α therapy has some efficacy in
extremely ill patients with encephalitis (713,714). Treatment of anterior
uveitis with topical corticosteroids is certainly indicated, but the efficacy of
systemic and periocular corticosteroids for the chorioretinal lesions of WNV
infection remain uncertain. Strategies directed at prevention of WNV
infections are the mainstay of infection control.
Infection with dengue virus is most often asymptomatic; however, it may
cause high fevers and severe arthralgias in patients residing near the equator
where it is known as “breakbone fever”. Uncommonly, more severe
manifestations including hemorrhagic fever and shock with mortalities rates
as high as 5% have been reported (715). All four DF serotypes are capable of
producing the full spectrum of disease. The most common ocular symptoms
of DF are blurring of vision and central scotomas. While ocular
manifestations are relatively rare with DF, young adults seem more prone to
develop posterior pole findings such as vasculitis, foveolitis, dengue-related
maculopathy, acute macular neuroretinopathy, and optic neuropathy that can
either be self-limited or result in permanent vision loss (716,717). Current
treatment is supportive, and unfortunately, recent widespread vaccine trials
have shown a worsening of disease in DF-naive patients once the pathogen is
naturally acquired by a mosquito bite (718). In cases where DF-related ocular
pathology persists, topical and systemic steroids may be used to treat
inflammatory complications. There are case reports showing the efficacy of
IVIg as a salvage therapy in patients with worsening vision despite
corticosteroid therapy (716). Detection of DF is similar to that of WNF, but
cross-reactivity of the flaviviruses can cloud the interpretation of IgM-based
assays. Coinfections with Zika and chikungunya virus are not uncommon and
only complicate the diagnosis.
Zika virus is found throughout Central and South America, central
Africa, and Southeast Asia. As with WNV and DF, most patients infected
with Zika are asymptomatic or develop a mild flu-like syndrome with
conjunctivitis. More clinically significant manifestation includes bilateral
hypertensive, anterior, nongranulomatous uveitis and posterior uveitis with
mid-peripheral yellow-white chorioretinal lesions (719–721). Congenital
transmission is much more severe as it can result in profound microcephaly.
More severe ocular manifestations also occurs in these infants and include
strabismus, iris colobomas, macular pigment mottling, optic nerve
hypoplasia, chorioretinal atrophy, hemorrhagic retinopathy, and torpedo
maculopathy (Figure 47-13) (722–725). Unfortunately, the ocular findings
that are not self-limited and/or responsive to topical or systemic steroids can
cause significant and irreversible vision loss. The differential diagnosis of
Zika virus includes the TORCH infections, isolated congenital
malformations, DF, chikungunya virus, and WNV. Laboratory tests have
been designed to detect Zika virus in both serum and urine. Current treatment
is supportive for systemic disease, and the CDC currently recommends
pregnant women to avoid areas with risk of Zika transmission. Topical
steroids and ocular antihypertensives have been effectively used to treat
ocular inflammation and elevated intraocular pressure. Of note, Zika virus
has been found in the tear film of patients 30 days after onset of symptoms,
so special care should be taken when performing ocular examinations as this
fluid may serve as a latent infectious reservoir (726).
FIGURE 47-13 Zika-associated viral retinitis. Macular
chorioretinal atrophic lesion in associating with congenital
Zika virus infection. (Images courtesy of Camila Ventura,
MD, PhD.)
Chikungunya virus is a vector-borne, positive-sense, single-stranded RNA

virus of the family Togaviridae that, in most cases, causes a self-limited high-
grade fever, photophobia, and polyarthralgia in countries near the equator.
These symptoms are common to DF and Zika virus as well and occur 3 to 7
days after exposure. The virus can cause a myriad of ocular findings ranging
from conjunctivitis to panuveitis, but a bilateral, pigmented granulomatous or
nongranulomatous anterior uveitis with an elevated intraocular pressure is
most commonly reported (727–729). Testing for the virus is similar to that
for other viruses and unfortunately requires analysis by the CDC due to
biosafety concerns. The differential diagnosis includes the other emerging
pathogens discussed in this section as well as HSV and VZV (Figure 47-14).
There is no proven treatment for the systemic disease, but anterior uveitis
with elevated intraocular pressure should be treated with topical steroids and
ocular antihypertensive medications.
FIGURE 47-14 Chikungunya-associated viral retinitis. A:

Peripheral retinal whitening and hemorrhages seen in
chikungunya-associated viral retinitis. B: Late-stage
diffuse atrophy of pediatric chikungunya virus infection.
(Images courtesy of Andrea Zinn, MD, PhD.)
Ebola virus is a potentially lethal, hemorrhagic, single-stranded RNA virus of

the family Filoviridae transmitted by direct contact with infected blood or
bodily fluids, sexual transmission, or consumption of contaminated meat in
sub-Saharan Africa (730,731). Mortality rates average 65% but depend
significantly on the virus species and proximity to care (730). The most
recent outbreaks have brought to light a “post-Ebola virus syndrome” that
develops in approximately 15% of survivors that is characterized by extreme
fatigue, arthralgias, and visual and hearing loss (732). Chronic anterior
uveitis, posterior uveitis, and panuveitis may lead to significant ocular
structural complications (hypotony, macular edema, epiretinal membranes,
and cataract) and visual impairment in these patients (733). The uveitis is
usually steroid responsive, but approximately a third of patients with uveitis
will have visual acuities worse than 20/400 in the affected eye (733). The
demonstration of persistence of viable virus in the eye months after it had
been cleared from the blood poses unique challenges in the surgical
rehabilitation of patients with cataract and post-Ebola virus syndrome (734).
Laboratory testing for Ebola should be performed only by specially trained
staff due to high risk of transmission and the need for established quarantine
protocols. The differential diagnosis includes sarcoidosis, syphilis, TB,
Lyme, and endophthalmitis depending on the specific presentation. Current
treatment is supportive, and infection control is paramount to reduce the risk
of periodic outbreaks. While uncommon, awareness of the presentation and
ocular complications of these emerging pathogens is critically important in
helping arrive at an accurate diagnosis and in limiting vision loss particularly
at a time when international travel is common.
Syphilis
Syphilis is an uncommon cause of uveitis in the pediatric age group with
ocular involvement occurring in both the acquired and congenital forms of
the disease as a result of infection with the spirochete, Treponema pallidum.
Acquired infection is most commonly due to sexual contact with infected
individuals. The progression of untreated, acquired syphilis has been well
described, the signs and symptoms of both ocular and systemic disease
varying with the stage of the disease. Classically, the primary infection is
characterized by a single chancre teeming with spirochetes that erupts
approximately 3 weeks after exposure and spontaneously resolves within 12
weeks. Six to eight weeks following the appearance of the chancre, the
disseminated portion of the disease, secondary syphilis, appears as a diffuse
maculopapular rash involving both the palms of the hands and soles of the
feet and generalized lymphadenopathy. Multiple organ systems including the
eye can be involved during this stage with uveitis occurring in approximately
10% (735). Latent syphilis then develops and is defined by positive serologic
markers of infection in asymptomatic patients; however, congenital
transmission is still possible during this time (736). One-third of untreated
patients with latent syphilis will progress to tertiary syphilis with
manifestations of gummas, noncancerous growths, obliterative endarteritis,
and cardiac and neurologic abnormalities years to decades later. Intraocular
inflammation may occur in any stage of the disease and should regarded as
neurosyphilis.
Congenital syphilis may arise either due to transplacental infection after
the 10th week of pregnancy or, less commonly, through direct contact with a
genital chancre during birth. Acquired disease in a child should raise
questions of sexual abuse or precocity. Furthermore, any child found to have
acquired HIV due to sexual abuse should be evaluated for syphilis due to
high rates of coinfection (736). Pregnant women with untreated or
inadequately treated syphilis may transmit the spirochete to their fetus during
any stage of disease and time throughout pregnancy. In fact, untreated
women have about a 70% chance of transmitting the infection to their fetus
during the first 4 years of disease (737). In general, the longer the interval
between maternal infection and pregnancy, the more benign the outcome in
the infant (Kassowitz law) (738). If the infection is transmitted late in
pregnancy, the infant may appear completely normal at birth, and if left
untreated, develop manifestations of the disease over time (739). For this
reason, the CDC recommends that all pregnant women be tested for syphilis
(non–treponemal-based testing) during their first prenatal visit, and if at risk
of contracting the disease, again during the third trimester (740). If
nontreponemal testing is positive, confirmatory treponemal serology should
be performed with complete medical evaluation. Although this strategy will
fail to detect primary maternal syphilis acquired late in pregnancy, a careful
prenatal examination of the mother for signs of infection and prompt
treatment may mitigate this risk. Another important consideration is the
development of the “prozone” effect when nontreponemal antibody titers are
so high that the test is erroneously interpreted as negative if not properly
diluted (741). Maternal surveillance is paramount to reducing the risk of
congenital syphilis in the neonate.
The prevalence of syphilis drastically declined with widespread
availability of penicillin following World War II. In 2000 and 2001, the CDC
reported the lowest rates of syphilis since reporting began in 1941. Despite
continued efforts to reduce disease burden further, syphilis rates have instead
risen since 2001, and in 2015–2016 alone, the rate among women rose
35.7%. This mirrors the rise in congenital syphilis cases that have increased
86.9% from 2012 to 2016. In total, of 628 cases of congenital syphilis and 41
stillbirths were reported in 2016, due in large part to an absence of testing
among pregnant until late in pregnancy, if at all (742,743). Syphilis should be
always be in the differential of any patient with unexplained intraocular
inflammation, including young children, as a delay in diagnosis and treatment
can lead to significant ocular and systemic morbidity, especially considering
that the infection is curable with appropriate antibiotic therapy and the
disease is on the rise in the United States.
The clinical manifestations of congenital syphilis have been arbitrarily
divided into early and late onset (after age 2) (738). The most commonly
reported systemic findings in early congenital syphilis include desquamative
skin rash, low birth weight, failure to thrive, severe anemia, generalized
lymphadenopathy, distended abdomen, hepatosplenomegaly, radiographic
evidence of characteristic and symmetric long bone changes of the lower
extremities, and pneumonia (738). Other less common manifestations include
rhinitis, bloodstained coryza or snuffles, and CNS involvement such as
meningitis, hydrocephalus, cognitive delay, and seizures. Late manifestations
of congenital syphilis develop as the result of scarring from early systemic
disease and include fissures and mucous patches (rhagades), Hutchinson
incisors, mulberry molars, perforation of the hard palate, abnormal facies,
bossing of the frontal and parietal bones, saber shins, progressive
sensorineural deafness, and neurosyphilis (737). Cardiovascular
complications are an uncommon complication of congenital syphilis.
Ocular manifestations of congenital syphilis are uncommon but
increasing in frequency according to retrospective data analysis and in line
with the rising caseload of the disease (744,745). Ocular inflammation related
to congenital syphilis may be present at birth or not present for years.
Spirochetes have been isolated from aqueous humor, choroid, retina, and
inflamed corneal stroma of neonates with congenital syphilis (746,747).
Ocular findings of congenital syphilis include uveitis, interstitial keratitis,
iridoschisis, congenital cataracts, optic neuritis, and glaucoma (748).
Nonulcerative interstitial keratitis (IK) is the most common inflammatory
finding of untreated, late congenital syphilis (20% to 50% of cases) and is
usually accompanied by anterior uveitis (749). IK is more common in girls
and is typically bilateral in congenital cases while unilateral in acquired
disease. The most common uveitis is a multifocal chorioretinitis, and less
commonly, retinal vasculitis that results in the development of a salt-and-
pepper fundus (750). The salt-and-pepper appearance is due to alternating
RPE hyperplasia and atrophy usually most prominent in the peripheral fundus
but may include the entire retina.
In acquired syphilis, intraocular inflammation may occur as an isolated
finding in any anatomic region of the eye (anterior, intermediate, posterior or
panuveitis), but most commonly presents with posterior segment findings,
including vitritis, papillitis, neuroretinitis, chorioretinitis, necrotizing retinitis,
and retinal vasculitis. Two distinctive posterior segment presentations
deserve special mention: acute syphilitic posterior placoid chorioretinitis
(ASPPC) and panuveitis with superficial retinal precipitates as their
biomicroscopic and imaging findings are highly suggestive of the diagnosis
of ocular syphilis (751–753) (Figure 47-15). In ASPPC, uniform inner/outer
choroidal inflammation manifests as discrete, nonelevated, pale yellow,
oval/circular lesions in the posterior pole that, on fluorescein angiography
(FA), become progressively hyperfluorescent, while indocyanine green
angiography (ICGA) shows confluent hypofluorescence. OCT of these
lesions demonstrates focal thickening and nodularity of the RPE with
disruption of the overlying IS-OS junction, which resolves with treatment
(Figure 47-15) (752,754,755). These findings warrant a lumbar puncture and
treatment with intravenous penicillin G.
FIGURE 47-15 Syphilitic posterior uveitis. A: Acute

syphilitic posterior placoid chorioretinitis (ASPPC) and
(B) panuveitis with superficial retinal precipitates.
Direct visualization of spirochetes from infected tissue confirms the

diagnosis. In the absence of this information, serologic tests are required.
There are two types of serologic tests available: nontreponemal (rapid plasma
regain [RPR] or Venereal Disease Research Laboratories [VDRL]) and
treponemal (fluorescent treponemal antigen absorption test [FTA-ABS])
antigen tests. The sensitivity and specificity of these tests can vary depending
on the stage of the disease and history of treatment; thus a single type of
serologic test is not sufficient for diagnosis. Classically, the recommendation
has been to screen patients with nontreponemal tests and then positive results
are confirmed by treponemal-based tests. There is good evidence to support
using both treponemal and nontreponemal tests to diagnose ocular syphilis. In
a small retrospective study, 15% of patients had a negative RPR despite a
positive FTA-ABS and no prior treatment (756). In our own clinical
experience, we have seen similar findings with ocular syphilis in which all
patients tested positive for FTA-ABS tests but only 68% had a reactive
VDRL (749).
Due to time limitations in high-volume labs and the expertise needed to
perform the tests, clinical laboratories have begun to use an algorithm termed
reverse sequence screening. The sample is first tested with an immunoassay
(enzyme immunoassay or multiplex flow immunoassay). If positive, the
sample is then evaluated for RPR status, and this level is then followed to
monitor treatment response. Discordant results are resolved with a T.
pallidum particle agglutination test. This algorithm has shown increased
sensitivity and specificity for syphilis, but improvements in the technique are
still ongoing (757,758). The principal advantage to screening with reverse
sequence testing is that syphilis IgG antibody detection is much more highly
sensitive than nontreponemal testing during primary, latent, and late syphilis
(759). While high sensitivity is a desirable criterion for screening test, with it
comes the expected disadvantage of increased false-positive tests, the
proportion of which increases among all positives as disease prevalence
decreases (760). In every case of syphilis-associated uveitis, CSF evaluation
with a lumbar puncture should be performed to aid in diagnosis, staging of
disease, and treatment plan as one cannot predict from clinical findings which
patients will also have neurosyphilis (761). A recent analysis in fact found
that 72% of patients with ocular manifestations of syphilis had abnormal CSF
results (762). PCR analysis of CSF fluid can be used to make the diagnosis of
neurosyphilis as well (763). Furthermore, any patient diagnosed with syphilis
should be tested for HIV and vice versa as demographics, clinical
presentation, duration of treatment for adequate cure, and severity of disease
can vary tremendously among individuals with HIV coinfection
(761,764–766).
In congenital syphilis, the CDC has segregated congenital syphilis into
four possible categories to guide diagnosis and treatment: proven or highly
probable, possible, less likely, and unlikely. In proven or highly probably
cases, there must be physical examination findings, a positive dark-field
microscopy or PCR of lesions or body fluid, or serum nontreponemal
quantitative serologic titer that is fourfold higher than the mother’s to make
the diagnosis. Possible cases are those in which the mother was inadequately
treated per CDC guidelines or received treatment <4 weeks before delivery.
For either proven or possible cases, the CDC advises 10 days of intravenous
penicillin G and CSF evaluation. In cases that are “less likely” or “unlikely,”
the mother has been treated appropriately and the neonate’s serology is
normal. In less likely cases, either the infant can be closely monitored or a
single dose of benzathine penicillin G can be administered. “Unlikely” cases
require periodic serologic evaluation but no further intervention provided
follow-up can be assured and serology remains unremarkable (740).
Penicillin G is the mainstay of treatment for any syphilis infection. The
treatment of infants with congenital or acquired syphilis according to CDC
guidelines is as follows: 50,000 units/kg/dose intravenous aqueous (IV)
crystalline penicillin G every 12 hours for the first 7 days and every 8 hours
for an additional 3 days or a single 50,000 units/kg/dose of intramuscular
procaine penicillin G for 10 days for congenital syphilis. In infants older than
1 month, 50,000-units/kg/dose IV aqueous crystalline penicillin G every 4 to
6 hours for 10 days is recommended (740). Acquired ocular syphilis should
be considered a CNS infection and treated with 18 to 24 million units per day
divided into 3 to 4 million units IV every 4 hours or as a continuous infusion
for 10 to 14 days per CDC recommendations for the treatment of
neurosyphilis. If compliance can be ensured, 2.4 million units of
intramuscular procaine penicillin G once daily with 500 mg of probenecid
four times daily for 10 to 14 days can be considered as an alternative (740).
In cases of penicillin allergy in infants with congenital syphilis, the CDC
advises oral or intravenous desensitization due to lack of evidence for
efficacy of other therapies in children (740,767,768). A recent case report has
advocated for the use of intravitreal ceftazidime while desensitization is
pursued to avoid further vision loss (769). IK can be exceptionally difficult to
treat. It can be refractory to penicillin once established, but maternal
treatment or treatment of the infant before 3 months of age may prevent the
development altogether (738). The development of a Jarisch-Herxheimer
reaction in which malaise, headache, tachypnea, tachycardia, vasodilation,
leukocytosis, chills, and fever can occur 2 to 12 hours after initiation of
systemic treatment but usually resolves within a day. Therapy should
continue despite this reaction, and prednisone may be used to help alleviate
the symptoms until they have resolved. In a recent meta-analysis, oral
prednisone did not affect visual outcomes for ocular syphilis (770). If therapy
can be administered in a timely fashion for ocular syphilis, patients with or
without coinfection with HIV have favorable outcomes (771,772).
Lyme Disease
LD is a multisystem disorder caused by the spirochete Borrelia burgdorferi
sensu lato (773). The disease was first recognized in 1975 and is now
considered the most common vector-borne disease of Europe and the United
States (774,775). According to the CDC, in 2016 there were 26,203 proven
cases of LD and another 10,226 that were considered probable cases. Boys
aged 5 to 9 years are the most commonly affected (776). The clinical course
of systemic LD has been divided three stages analogous to that of syphilis.
Early disease is characterized by the development of constitutional
symptoms, an expanding targetoid-appearing macular rash known as
erythema chronicum migrans within 2 weeks at the site of the tick bite in
60% to 80% of individuals (stage 1). If left untreated, the spirochete
disseminates (stage 2) to cause arthralgias (monoarticular or oligoarthritis
with preference for the knee); facial nerve palsies, meningitis, and
atrioventricular heart block may ensue (777). The disease can then progress
to cause persistent arthritis, polyneuropathies, encephalopathy, and frank
psychosis in late LD (stage 3) (777,778).
Ocular findings associated with LD vary with the stage of the disease
(779). The most common ocular finding in stage 1 of the disease is a
follicular conjunctivitis, and keratitis predominates the persistent stages
(780). Intermediate uveitis is the most common intraocular manifestation
during stages 2 and 3 of the disease in adults (781). A granulomatous anterior
chamber reaction, severe vitritis, papillitis, neuroretinitis, choroiditis, and/or
panuveitis may also occur (782–784). In children, case reports have described
an isolated anterior uveitis; however, anterior chamber reactions are much
more common in association with cases of keratitis or intermediate uveitis
(782–789). Intermediate uveitis is rare in pediatric LD (790). LD can also
rarely cause ocular motor abnormalities, paralytic strabismus, optic
neuropathies, posterior uveitis with fibrosing scars of the macula, and an
acute transverse myelitis in children (791–794). In a cohort of 84 children
with LD-associated arthritis from Europe, only 4% of patients developed
ocular involvement (1 each of keratitis, anterior uveitis, and intermediate
uveitis) (785). The authors concluded that regular screening examinations
were unnecessary due to the very low rate of affected children (785). This is
in contradistinction to the most important differential diagnosis in LD-
associated uveitis, JIA-associated uveitis, in which screening guidelines for
the detection of intraocular inflammation have been recommended for all new
diagnoses of JIA.
The diagnosis is clinical with identification of residence in or travel to
endemic areas, exposure to blacklegged ticks, and objective physical
examination findings (erythema chronicum migrans). Laboratory data are
used to support the presumed diagnosis as the tests available lack
standardization of positive values and cross-reactivity among the various
spirochetes causes high rates of false-negative and false-positive results. Due
to this, the CDC currently recommends a two-step test to diagnose LD. The
first step is an enzyme-linked immunosorbent assay or indirect
immunofluorescence assay. If the first step is positive, a Western immunoblot
should be performed as the second step. The results are considered positive
only if both steps are positive. PCR-based assays have been successfully used
to amplify both genomic and plasmid B. burgdorferi DNA from a variety of
tissues, including ocular fluids, and the highest yields are obtained from skin
(782,795). LD-associated intraocular inflammation should be considered a
manifestation of CNS infection and careful neurologic evaluation including a
lumbar puncture is required in these cases. Antibiotic dosing should be dosed
for CNS involvement and guided by an infectious disease expert.
If the tick is removed within 36 hours of implanting, the transmission of
LD is significantly reduced (796). If LD should develop, prompt
administration of antibiotics, preferably doxycycline, is usually curative
(797). In children under age 8 in whom doxycycline is contraindicated,
amoxicillin, cefuroxime, and azithromycin are other options. In more severe
infections, intravenous ceftriaxone should be considered. In cases where there
is intraocular involvement, topical corticosteroids and mydriatics should be
used to quiet an anterior chamber reaction while under appropriate antibiotic
coverage. Systemic corticosteroids have been used in the management of LD;
unfortunately, their use is controversial due to higher rates of treatment
failures. As with syphilis, a Jarisch-Herxheimer reaction, a hypersensitivity to
spirochete antigens released during the first 24 hours of treatment that cause
constitutional symptoms and increased intraocular inflammation, may
complicate antibiotic treatment. In these cases, local and/or systemic
corticosteroids may be required, but resolution without long-term visual
sequelae occurs in the majority of cases with observation alone.
Tuberculosis
TB kills over a million people per year worldwide (798). According to the
CDC, in 2017 there were 9,093 cases reported in the United States, a decline
from 2016 and the lowest count on record (799). Foreign-born children and
adults are much more likely to have TB than U.S. born peers (799). In the
United States in 2016, children (<15 years old) represented approximately
4% of all TB cases with those aged 10 to 14 years having rates of
extrapulmonary involvement approaching 35% according to the CDC (799).
Fortunately, TB-related ocular and systemic diseases are relatively
uncommon in the United States. Although TB is considered one of the “great
mimickers” of various other uveitis entities and may cause intraocular
inflammation in any anatomical subtype of uveitis, ocular involvement is
uncommon even in parts of the world where TB is endemic (Figure 47-16)
(800). Nevertheless, TB should always be considered in the differential of
new onset of uveitis and excluded in children being considered for treatment
with IMT, particularly with TNF inhibitors. Ocular TB has been identified in
children as young as 1 year of age and in children with disseminated disease
(801,802). In areas of the world where TB is much more common, clinical
observations have noted worse intraocular inflammation from TB in children
compared to adults and thus children require more aggressive anti-
inflammatory treatment (803).
FIGURE 47-16 TB-associated macular star and choroidal
granuloma. A large macular star associated with active
tuberculosis. Inset shows a choroidal granuloma within the
macula in this same patient.
While a pediatric infectious disease specialist will guide most TB treatment,

it is imperative for the ophthalmologist to be familiar with current treatment
guidelines, importance of medication compliance, and ocular toxicity of anti-
TB drugs to guide appropriate monitoring. Visual field assessment,
measurement of retinal nerve fiber layers, and red–green color testing should
all be utilized examinations to monitor for optic nerve toxicity although there
is no literary consensus on their frequency. Nine months of four-drug therapy
should be advocated for in cases of presumed ocular TB to reduce the chance
of recurrence (804).
Catscratch Disease
CSD is a feline-associated zoonotic disease caused by a genus of gram-
negative bacilli known as Bartonella. There are currently 21 recognized
species with eight known to cause disease in humans. The most well known
is Bartonella henselae, a pathogen distributed worldwide and responsible for
CSD and the principal cause of neuroretinitis (805). CSD, as its name
implies, is transmitted most frequently from immature cats to humans
through a bite or scratch; however, there are confirmed cases where no
animal encounter can be elicited (806). The syndrome is defined by regional
lymphadenopathy, low-grade fever, and an erythematous papule or pustule
that develops within 3 to 10 days of exposure at the inoculation site. The
systemic signs and symptoms such as lymphadenopathy and fever develop 1
to 2 weeks after the local pustule has appeared (807). CSD occurs in the
immunocompetent of all age groups, but the highest age-specific incidence is
found among children (808). In fact, B. henselae is one of the most common
causes of chronic lymphadenopathy in children (808).
Ocular manifestations of CSD occur in approximately 5% to 7% of
patients, and the most common is Parinaud oculoglandular syndrome in
which there is fever, a unilateral granulomatous conjunctivitis, and ipsilateral
preauricular lymphadenopathy (809,810). Facial nerve palsies and Horner
syndrome have also been recorded (811,812). Intraocular manifestations
include unilateral or bilateral neuroretinitis, optic neuritis, optic neuropathies,
inflammatory and vascular lesions of the optic nerve, vascular occlusions,
uveitis (anterior, posterior, or panuveitis), multifocal chorioretinitis, and
serous detachments of the macula (Figure 47-17) (813–823).
FIGURE 47-17 Neuroretinitis in a patient with catscratch
disease. Optic disk swelling, moderate vascular
engorgement, and partial macular star right eye.
The diagnosis of CSD is clinical with supportive serologic testing. Indirect

fluorescent antibody (IFA) tests developed by the CDC are both sensitive and
specific except in patients with HIV (824,825). Enzyme immunoassays (EIA)
and Western blot procedures have shown similar to slightly improved
sensitivities and specificities (826,827). Unfortunately, there is a potential for
cross-reactivity among the various Bartonella species that cause disease in
humans. The recent development of PCR-based techniques is not yet widely
available but will likely supplant other techniques in the near future for ocular
specimens due to the high degree of sensitivity and specificity and the
paucity of sample required for testing (707,828).
In the vast majority of cases, CSD is a self-limited disease with an
excellent systemic and visual prognosis (821,829). While most clinicians
observe mild to moderate inflammation in the immunocompetent patients,
more significant inflammation or disease found in the immunocompromised
should warrant treatment with systemic antibiotics. Treatment should ideally
be comanaged with a pediatric infectious disease expert. There is no
consensus on antibiotic choice, but erythromycin, doxycycline, erythromycin,
rifampin, trimethoprim–sulfamethoxazole, ciprofloxacin, and gentamicin
have all been employed successfully (821,830,831). Systemic doxycycline is
preferred to erythromycin due to its superior intraocular and CNS
penetration; however, the drug is contraindicated in children <9 years of age
due to the risk of permanent tooth discoloration. In more severe infections,
intravenous erythromycin or doxycycline should be considered in
combination with rifampin. The treatment duration can vary from 2 weeks up
to 4 months in the immunocompromised patient. Longer treatments can be
administered to prevent disease recurrence in these vulnerable populations
(832). The use of oral steroids significantly improved visual outcomes in one
large retrospective study; however, their use has not been systematically
evaluated in this disease (821).
Postinfectious and Vaccination-Associated Uveitis

An infectious agent can be identified in over a third of patients with ocular
inflammation (833), and postinfectious inflammation is much less common.
The most common culprit is streptococcus species in which a bilateral,
nongranulomatous anterior uveitis can occur in the setting of significantly
elevated antistreptococcal lysin O titers 1 to 8 weeks after initial infection
(834). However, in a third of patients, there is some form of posterior
involvement such as vitritis, focal retinitis, optic nerve swelling, multifocal
choroiditis, or panuveitis, and it may be the only manifestation of the disease
(834–836). The uveitis most frequently affects younger patients and responds
to systemic corticosteroids but has a propensity to recur (836). Patients with
poststreptococcal uveitis syndrome may present first to the ophthalmologist
where early diagnosis and treatment is essential in preventing possible
sequelae of this infection including rheumatic fever and glomerulonephritis.
The pathogenesis is currently unclear but is believed to be similar to other
poststreptococcal autoimmune conditions such as rheumatic fever and
Sydenham chorea in which molecular mimicry is the underlying
pathophysiologic mechanism (837). The differential diagnosis of
simultaneous bilateral nongranulomatous anterior uveitis in children includes
postinfectious or drug induced, TINU syndrome, Kawasaki disease, and
inflammatory bowel disease, and uncommonly, HLA-B27–associated
anterior uveitis (838).
There are several immunizations associated with a postvaccination-
associated uveitis. The Bacille Calmette-Guerin (BCG) vaccine for TB can
cause a bilateral granulomatous or nongranulomatous anterior uveitis that
readily responds to topical corticosteroids and cycloplegics and is the vaccine
most strongly linked to uveitis (839). Less commonly, the vaccine, and even
purified protein derivative (PPD), can lead to chorioretinitis, panuveitis, or
optic neuritis (839). The measles, mumps, and rubella (MMR) vaccine may
cause an anterior uveitis or panuveitis usually weeks after immunization. The
inflammation can last much longer than that seen with BCG and may require
both topical and oral corticosteroids (839). There are also case reports of
acute posterior multifocal placoid pigment epitheliopathy (APMPPE)
following influenza and hepatitis B vaccine administration (839). The
varicella vaccine has also been linked to an anterior uveitis and/or ARN
presumably due to the attenuated virus used in the vaccine. Treatment
requires antivirals and local steroids (839). Anecdotally, the diphtheria–
pertussis–tetanus (DPT) and smallpox vaccines have also been shown to be
associated with uveitis, but case reports remain rare (839). Taken together,
postvaccination-associated uveitis is still a relatively uncommon event
despite the widespread use of these vaccines and should not deter children
from receiving these lifesaving therapies unless a significant contraindication
exists.
Conclusions
Uveitis in children is relatively common in ophthalmology clinics and has a
devastating impact on their lives, those of their families, and society as a
whole. Despite significant advances the diagnosis, management, and the
availability of new therapeutic agents, legal blindness still occurs in around
8% to 10% of cases (15,840). With the concerted efforts of uveitis specialists,
pediatric ophthalmologists and pediatric and adult retina specialists, pediatric
rheumatologists, and pediatricians, this risk of lifelong blindness is
potentially modifiable through earlier diagnoses, referrals to subspecialists,
and aggressive measures to suppress intraocular inflammation and place it in
remission. The broader armamentarium of anti-inflammatory drugs with the
introduction of biologic agents has expanded our therapeutic options;
however, postsurveillance marketing and randomized controlled trials are
necessary to minimize risk of harm and define efficacy of these newer
immunomodulatory agents in this vulnerable population. The novel
applications of imaging modalities such as OCT and laser-flare photometry to
more quantitatively assess inflammation and a reassessment of the current
screening guidelines may serve to more accurately identify those children at
greatest risk of vision loss and the development of ocular complications.
Moving forward, the use of real time, quantitative, PCR-based assays will
help to classify more infectious causes of uveitis especially for cases of
unsuspected or unusual pathogens (707). Moreover, the advent of unbiased
metagenomic deep sequencing may identify infectious organisms (fungi,
parasites, DNA and RNA viruses) in eyes with otherwise idiopathic or
undifferentiated uveitis requiring very small volumes of intraocular
specimens, possibly changing our paradigms of the etiopathogenesis for
many uveitic entities (841).
Lastly, we propose the following changes or areas of emphasis to help
identify children at risk of developing uveitis and the complications related to
ocular inflammation that would benefit from early referral to specialists in the
field:
1. The widespread establishment of vision screening programs in

elementary schools.
2. Informing schools of local pediatric uveitis specialists.
3. Identification of children at risk for developing uveitis and early referral
to subspecialists such as pediatric rheumatologists and uveitis experts.
4. Emphasizing the need for early referrals to uveitis specialists in cases of
persistent inflammation among ophthalmology and rheumatology
communities.
5. Educating the community and primary care providers of the need for
lifelong follow-up for patients with pediatric uveitis.
6. Coordination and frequent communication between subspecialists caring
for children with uveitis.
7. Educating optometrists and ophthalmologists about the detrimental
effects of long-term systemic or local steroid use to manage uveitis and
that long-term, low levels of inflammation should not be tolerated.
8. Develop therapeutic guidelines for cases of persistent inflammation with
IMT use no <2 years to increase steroid-free remission rates.
9. In cases of suspected infectious uveitis, the use of PCR-based techniques
should be more readily employed when available to improve diagnostic
yields and pathogen identification, direct specific antimicrobial
treatment, and avoid potentially devastating corticosteroid monotherapy.
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48
Surgical Approaches to Uveitis
Stavros N. Moysidis, Edward H. Wood, Baruch D. Kuppermann,
and Lisa J. Faia
INTRODUCTION
Pediatric uveitis is a subspecialty of vitreoretinal diseases within the field of
ophthalmology, which may be due to infectious and noninfectious etiologies.
The goal of treatment is to achieve control of ocular inflammation, whether
by local therapies, systemic corticosteroids, or other immunomodulatory
drugs. The workup and treatment vary based on the patient and etiology of
the uveitis and are based on the best clinical judgment of the physician.
Occasionally, surgical intervention is necessary in a child with uveitis.
Diagnostic surgery may be needed in the pursuit of the underlying diagnosis.
Therapeutic surgery may be needed to treat the sequelae of chronic
inflammation, including vitreous hemorrhage, iridocyclitic membranes,
epiretinal membranes, retinal detachments, and hypotony. The
implementation of small-gauge instrumentation has allowed for an expansion
of the indications for surgical intervention in pediatric uveitis. Prior to the
advent of 23-gauge vitrectomy, there was a higher incidence of postoperative
complications in these patients, including uncontrolled inflammation,
hypotony, and phthisis. Advances in small-gauge instrumentation have
contributed to safer surgical intervention in uveitic eyes. In addition to
surgical advances, more aggressive control of chronic and recurrent
inflammation by ophthalmologists using systemic immunosuppressants has
led to improved outcomes (1,2).
Corticosteroids and systemic immunosuppression play critical roles in the
treatment of pediatric uveitis. Tight control of intraocular inflammation can
prevent irreversible complications of uveitis. Aggressive treatment with high-
frequency topical corticosteroid dosing is crucial at the onset of
inflammation. Periocular, intravitreal, or systemic corticosteroids may also be
needed to achieve control of the inflammation. Steroids should be tapered
when possible, and patients should be monitored for adverse effects related to
chronic steroid use. Patients who are on steroids long-term may require
immunosuppressant medications to allow for tapering of the steroid, as long-
term steroid use is not recommended in the pediatric population. The treating
ophthalmologist may need to coordinate care with a rheumatologist or
primary care physician for immunosuppressant therapy and the monitoring of
toxicity with physical examination and laboratory testing (see also Chapter
47).
The decision to pursue therapeutic surgery in a child with uveitis should
be carefully considered (Table 48-1). Surgery in a pediatric eye with chronic
or recurring inflammation is a daunting task and is accompanied by unknown
intraoperative risks and an uncertain postoperative course (3,4). In order to
try to eliminate as many postoperative complications as possible, these eyes,
if possible, should have at least 3 months of quiescence of their disease
before surgical intervention. Surgery alone can be daunting without also
having to deal with the postoperative complications, which can result in the
loss of the eye. Surgery in these eyes can be technically challenging due to
corneal opacification, posterior synechiae, delicate iris vasculature,
hemorrhage diathesis, iridocyclitic membranes, and miotic pupils (5). The
combination of the pediatric heightened immune response and a uveitic eye
may result in a recalcitrant postoperative inflammatory response and may
lead to hypotony or phthisis. The decision to operate must be carefully
weighed.
TABLE 48-1 Surgical considerations in pediatric

uveitis
On the other hand, surgery should not be delayed to the point that the patient
suffers irreversible damage and, sometimes, a surgeon must operate on an
active eye. A special concern in young children is also the risk of amblyopia
and strabismus, and while the visual cortex and neuro-ophthalmic pathways
are largely developed by the age of 4, the Pediatric Eye Disease Investigator
Group trials have shown visual benefit to amblyopia treatment even in the 7-
to 17-year-old age group (6). The decision to operate should be carefully
weighed by the clinician considering surgery for a nonclearing vitreous
hemorrhage in a child with uveitis. It is critical to fully address the risks and
benefits of surgery with the patient and his or her guardian; children over the
age of 14 are capable of providing assent, but the responsibility of consent is
in the hands of the parents or guardian. Establishing control of intraocular
inflammation is crucial for a successful surgical outcome. Treatment options
for achieving this control of inflammation during and after surgery are
discussed in the sections that follow. The physician managing the child’s
uveitis should communicate closely with the pediatric ophthalmologist to
manage any possible amblyopia and, if the patient is on systemic
immunosuppression, with the pediatric rheumatologist to stay ahead of the
ocular inflammation. The age of the child, the presence or absence of
amblyopia (whether from anisometropia, strabismus, or deprivation), and the
presence or absence of irreversible retinal or optic nerve pathology are
important factors affecting the surgical outcomes of children with uveitis.
This chapter first discusses surgical procedures in the pursuit of an
etiology of uveitis. The chapter then discusses an array of therapeutic surgical
procedures used to treat the sequelae of uveitis.
DIAGNOSTIC SURGERY
Diagnostic surgery may be necessary to establish the etiology of uveitis.
Diagnosis begins with a careful history, ocular examination, and focused
laboratory testing with an understanding of the natural history of the patient’s
disease and potential complications. A diagnostic surgical procedure may be
reasonable if (a) the history-taking, examination, and noninvasive workup
have failed to reveal the underlying diagnosis; (b) the patient is not
responding to medical therapy and the diagnosis is being reconsidered; (c) the
patient develops significant complications during treatment and additional
diagnostic information is needed to guide therapy; or (d) malignancy is
suspected.
Anterior Chamber Paracentesis
Indications
Anterior chamber paracentesis is safe and easily performed; it provides a
small volume of aqueous humor for laboratory analysis. Inflammation leads
to breakdown of the blood–ocular barrier and can lead to higher than normal
antibody titers and immunoglobulin concentrations in the aqueous humor. To
compare antibody concentrations in aqueous and serum, both samples must
be obtained on the same day. The Goldmann-Witmer (GW) coefficient can be
used to determine whether there is a localized ocular antibody production; it
is measured by enzyme-linked immunosorbent assay (ELISA) and defined as
GW = A/S, where A is the specific antibody concentration in the aqueous
divided by the total IgG in the aqueous and S is the specific antibody
concentration in the serum divided by the total IgG in the serum (Figure 48-
1) (7).
FIGURE 48-1 Goldmann-Witmer coefficient.
Procedure
In a child, anterior chamber paracentesis may need to be performed under
anesthesia, but it varies based on the age and the individual patient’s ability
to remain still for the procedure. The anterior chamber is entered at the
limbus with a 30-gauge needle on a 1-cc syringe, and about 0.2 mL of
aqueous humor is removed. The aqueous can be sent for cell studies,
antibody measurement, bacterial or fungal culture, or viral antigen detection
with polymerase chain reaction. A vitreous sample may be higher yield in an
infectious workup.
Vitrectomy
Indications
Diagnostic vitrectomy may be considered when less invasive methods have
failed to establish a diagnosis. This is especially important for patients who
are not responding to medical treatment for the presumed diagnoses, patients
who are developing complications from current therapies, or in the case of a
suspected malignancy or other masquerader. The goal of the diagnostic
vitrectomy is to determine the etiology of inflammation in order to guide
treatment. A vitreous specimen may be obtained by direct aspiration,
typically performed using a 25- or 27-gauge needle through the pars plana or
by surgical vitrectomy. A diagnostic vitrectomy may be preferable to direct
aspiration, as it is more controlled; allows for a larger sample; and carries a
lower theoretical risk of retinal tear or detachment than direct aspiration and
traction on the more adherent vitreous base.
Procedure
A diagnostic vitreous sample is obtained via a three-port vitrectomy. A
longer infusion (4 to 6 mm) may be required in eyes with uveitis, as patients
may have scleral thickening, choroidal edema, or retinal separation that may
result in inadvertent suprachoroidal infusion placement (7). Direct
visualization of the infusion cannula within the vitreous cavity is critical prior
to turning on the infusion.
The initial specimen in a diagnostic vitrectomy should be obtained before
turning on the infusion, so that the sample is undiluted. A sample of vitreous,
typically 0.5 to 1 mL, depending on the child’s age, can be obtained with the
vitrector on low suction and with the infusion turned off. The specimen can
be aspirated directly into a syringe from the vitrector. Next, the infusion line
is opened and 1 mL of vitreous and fluid is aspirated into a separate syringe
from the vitrector. A therapeutic vitrectomy, again, depending on the child’s
age, may then be performed, if necessary. Although it may be beneficial for
the adult patient to have a posterior vitreous detachment induced at the time
of surgery, this is not the case in a child, because the strong adherence of the
vitreous to the retina may cause a retinal tear or detachment.
Both the undiluted vitreous sample and the diluted vitreous aspirate
should be immediately sent for microbiologic and cytologic analysis.
Millipore filtration can be performed on the dilute sample to concentrate any
microorganisms and cellular elements (2). Immunohistochemistry can be
performed and cellular specimens can be sorted for monoclonality and cell
subtyping. Polymerase chain reaction and cDNA probes may be used to
detect DNA from many infectious agents, such as the herpetic viruses and
Toxoplasma gondii. If flow cytometry is indicated, Roswell Park Memorial
Institute (RPMI) solution (Thermo Fisher Scientific, Grand Island, NE)
should be obtained from the flow cytometry lab on the day of surgery, and
the concentrated sample of vitreous cells should be immediately stored in the
RPMI solution and sent to the flow lab. Otherwise, samples should be sent
fresh and not in any fixatives.
Retinal or Chorioretinal Biopsy
Indications
In some conditions, the infectious or inflammatory process is localized
primarily to the retina or choroid, and aqueous or vitreous testing may be
negative. Chorioretinal biopsy may be considered in patients with sight-
threatening disease or inflammation refractory to medical treatment.
Procedure
The retinal or chorioretinal biopsy site should be selected to maximize the
diagnostic value and minimize potential complications. Biopsy sites located
in the superior retina allow for better and longer gas tamponade, although gas
is not necessarily needed during these cases. An optimal chorioretinal biopsy
is performed at the border of normal retinal and active retinal disease to
afford the pathologist more clinical information.
A vitrectomy is performed, and the retinal or chorioretinal biopsy site is
identified. In rare cases where there is simultaneously a retinal detachment,
retinal scissors or the vitrector can be used to perform the biopsy. In most
cases, the retina is attached; in those cases, a retinotomy can be made, and
saline can be injected subretinally to create a bleb. If the retinotomy is large
enough, retinal forceps can be used to grasp the tissue and remove it. If
visualization is not sufficient, a partial retinectomy can be performed to gain
access to the biopsy site (7). The intraocular pressure (IOP) may be elevated
to minimize bleeding. Endodiathermy may also be used for a deeper
chorioretinal biopsy to ensure hemostasis. If a bimanual technique is
required, a chandelier can be used for lighting, but it is not required in most
cases (8). The specimen should be removed with care, so that the specimen is
not lost in the trocar or the sclerotomy. The physician should keep in mind
the gauge of the instrumentation; a larger specimen may be more difficult to
remove through a 25-gauge trocar (0.55 mm) than a 23-gauge trocar (0.72
mm). Alternatively, the trocar can be removed and pulled out over the forceps
as the forceps is being removed from the eye, so as to allow the specimen to
exit through a larger space, directly through the sclerotomy. In rare cases, a
larger scleral incision can be made in order to remove the specimen with as
little damage as possible and closed with nylon sutures. A fluid–air exchange
is then performed, and all retinal breaks are treated with endolaser. Some
physicians prefer to perform endolaser prior to creating the retinotomy. Many
physicians will then exchange air with gas for tamponade, but for smaller
retinotomies, leaving the eye filled with air may also be appropriate.
In cases where a full-thickness choroidal biopsy is desired, this can also
be performed externally, with or without vitrectomy, in cases where a
specimen is located peripherally, preferably anterior to the equator. If the
physician elects for vitrectomy prior to the biopsy, the vitrectomy can be
performed first, and the biopsy site is identified. Scleral depression can be
performed with direct visualization under wide-field viewing system, or with
indirect ophthalmoscopy, and the biopsy site can be marked with indentation
with the scleral depressor and a marking pen. A scleral flap is performed with
a crescent blade or another knife of the surgeon’s preference, and a 6 mm × 6
mm flap is typically sufficient to achieve the biopsy. The hinge for the scleral
flap is often left at the posterior-most location, and the scleral cut-down
should be as thick as possible without penetrating the globe. After a flap of
sufficient thickness, the inner wall will appear gray or brown in color,
indicating to the surgeon that the choroid is less than a few hundred microns
away. Diathermy is performed on the inner bed of the sclera to minimize
bleeding. A smaller rectangle of sclera and choroid is then incised with a
needle or blade and removed with retinal scissors; typically, a 4 mm × 3 mm
or 3 mm × 3 mm biopsy is taken. The specimen is then grasped with forceps
and removed, and the scleral flap is sutured closed. If a vitrectomy was
performed, the eye may be filled with gas.
The retinal or chorioretinal biopsy specimen should be carefully
subdivided into three sections. The first section is sent for light and electron
microscopy, the second section is sent for viral or fungal culture or
polymerase chain reaction analysis, and the third section is sent for
immunohistochemical studies to determine the type of predominant immune
response (2).
THERAPEUTIC SURGERY
Despite appropriate medical therapy to control ocular inflammation,
therapeutic surgery may be needed to treat the sequelae of chronic
inflammation. The physician should be prepared to encounter surgical
challenges unique to a chronically or recurrently inflamed eye. Even a
minimally traumatic and efficient surgery may elicit severe postoperative
inflammation in a child with uveitis. Careful control of inflammation
preoperatively, intraoperatively, and postoperatively allows the greatest
chance of a successful surgery.
Band Keratopathy
Indications
Calcific band keratopathy is a degeneration of the superficial cornea with the
accumulation of calcium hydroxyapatite primarily in Bowman membrane but
also in the epithelial basement membrane and the basal epithelium. There can
be multiple etiologies of band keratopathy; however, in this chapter we focus
on the causes secondary to chronic uveitis in children, which include juvenile
idiopathic arthritis (JIA), sarcoidosis, or chronic intermediate uveitis (9). The
calcific deposits can lead to a rough corneal surface that elevates the
epithelium and leads to pain, foreign body sensation, recurrent corneal
erosions, and decreased vision. Indications for surgery include rehabilitation
of vision, improvement of ocular surface and pain, or improving the view to
allow for adequate clinical examination.
Band keratopathy can be removed by disodium
ethylenediaminetetraacetic acid (EDTA) chelation and superficial, lamellar
keratectomy. A second method for removing band keratopathy is by excimer
laser phototherapeutic keratectomy (PTK); however, this typically results in a
highly irregular base due to the nonuniformity of the calcium density in the
band across the cornea (10). PTK effectively treats ocular surface symptoms,
but improvement in visual acuity can be variable. Stewart and Morrell (11)
reported that 55% of patients who underwent PTK for visual rehabilitation of
band keratopathy had improvement or no change in visual acuity, and 45% of
patients had a decrease in visual acuity; meanwhile, 83% of patients who
underwent PTK for ocular surface improvement noted an improvement in
symptoms. Chelation with EDTA is the preferred treatment option for band
keratopathy, and multiple treatments are likely to be needed in chronically
inflamed eyes, due to recurrences.
Procedure
Although chelation can be performed with local anesthesia, general
anesthesia is preferred for most children. Foster has previously described his
technique for EDTA chelation and superficial keratectomy (1). The corneal
epithelium is scraped off with a no. 15 Bard-Parker blade, taking care not to
penetrate through Bowman’s. A well can then be made by cutting off the
back end of a 3-mL plastic syringe, which can be positioned over the affected
area with the cut side up. A few milliliters solution of 0.35% EDTA is placed
in the watertight well and held in position for approximately 5 minutes. This
allows for chelation of calcium in the affected area while simultaneously
protecting the limbal stem cells. The well is then removed, and the cornea is
irrigated with multiple washes of balanced salt solution. The no. 15 blade is
then used to scrape off the loosened flakes of calcium. The procedure is
repeated as many times as necessary to remove the calcium from the central
cornea. Care must be taken to avoid scarring of the Bowman’s within the
visual axis. Cycloplegic and antibiotic drops are given, a continuous-wear
bandage soft contact lens is placed, and the eye is patched. Topical antibiotic
and steroid are used postoperatively. Bandage contact lens, cycloplegia, and
oral analgesics help with pain control. The epithelium generally has covered
the corneal surface within 1 week following surgery, and the bandage soft
contact lens can be removed.
Glaucoma
Mechanisms of Glaucoma in the Setting of Uveitis

Glaucoma may occur in children with uveitis via a number of complex
mechanisms. The five major causes of secondary glaucoma in patients with
uveitis are (a) inflammatory cells and debris in the trabecular meshwork
leading to impaired aqueous outflow, (b) secondary angle closure from
posterior synechiae leading to pupillary block, (c) secondary angle closure by
peripheral anterior synechiae, (d) active trabeculitis, or (e) steroid-induced
IOP rise. Alternatively, inflammation may lead to the production of
prostaglandins and result in decreased IOP; indeed, this observation led to the
invention of latanoprost. Active inflammation may also suppress ciliary body
function and, in the chronic state, can lead to hypotony. Topical steroid
administration may relieve inflammation or trabeculitis and reduce IOP.
Paradoxically, topical corticosteroids may increase IOP from either
reactivated ciliary body function or a steroid-induced IOP rise. Thus,
evaluating and managing IOP in a child with uveitis is a complex task.
Angle-Closure Glaucoma
Indications
Children with uveitis may develop posterior synechiae, which can lead to
pupillary block, iris bombé, and secondary angle closure. The initial
treatment of choice for acute angle closure is laser peripheral iridotomy;
however, there is a high incidence of posttreatment iridotomy scarring and
closure in children with active inflammation. Thus, laser iridotomy may serve
as an appropriate initial choice but likely will not offer a definitive, long-term
treatment. Furthermore, many children will not tolerate a laser iridotomy in
clinic. A larger, controlled, surgical iridectomy under general anesthesia is
the preferred procedure in a child with uveitis and secondary angle closure.
Procedure
Surgical iridectomy consists of five steps: (a) incision, (b) exteriorization of
the iris, (c) excision of the iris, (d) repositioning of the iris, and (e) closure of
the incision. The incision may be made in the sclera under a conjunctival flap
or through clear cornea. It is technically more challenging to perform the
procedure through a clear corneal incision (CCI), but this minimizes
conjunctival scarring in anticipation of possible future glaucoma surgery.
A partial-thickness incision is made perpendicularly into clear cornea.
Sutures are then preplaced on either side of the incision. The corneal incision
is then extended perpendicularly until the anterior chamber is entered. Gentle
pressure can be applied to the posterior lip of the wound to prolapse and
exteriorize the iris. If the iris does not prolapse, then nontoothed forceps can
be used to grasp and externalize it. The iris is then excised. Often, the iris will
internalize into the anterior chamber after the iridectomy, but if it remains
prolapsed, viscoelastic or an iris spatula can be used to reposition it. The
cornea is then closed with 10-0 nylon suture. Corticosteroids may be given
subconjunctivally intraoperatively or topically postoperatively.
Uveitic Glaucoma
Children with chronic or recurrent uveitis who also have uveitic glaucoma are
challenging to treat (12). When medical therapy or glaucoma laser procedures
are inadequate to attain the IOP goal, surgical intervention may be necessary.
Failure rates for trabeculectomy are higher in eyes with uveitis than in
noninflamed eyes, although some studies report successful 1- and 2-year
results of trabeculectomy with or without antimetabolites in patients with
uveitis (13–16). Glaucoma drainage implants appear in some studies to trend
toward better long-term success rates than trabeculectomy in patients with
uveitis; however, a large clinical trial in children would be required to better
answer this question. Either technique can be used successfully according to
the physician’s preference (14,17–20). Choroidal effusions, hypotony, and
cystoid macular edema comprise the most common postoperative
complications for either surgery.
After incisional surgery, an extensive inflammatory response should be
anticipated in the pediatric eye with uveitis. The recurrent or chronic nature
of inflammation can result in fibrous tissue growth and subsequent closure of
the filtering bleb in a patient who underwent trabeculectomy. Adjunctive
antimetabolites have been shown to be effective in preventing this from
occurring in the short-term, but late or persistent inflammation may later
close the filtering bleb. One- and two-year success rates after trabeculectomy
without antimetabolite therapy range from 67% to 92% (13–16). However, 5-
year success rates decrease dramatically, with Towler et al. (16) reporting a
drop from 80% success at 2 years to 30% at 5 years.
The 1- and 2-year success rates after glaucoma drainage devices range
from 92% to 94% (14,17–20). A 91.7% success rate at both 1 and 2 years
with IOPs of 5 to 21 mm Hg was reported in 24 eyes treated with Baerveldt
glaucoma drainage implant (17). A 94% success rate at 1 year with IOP ≤ 21
mm Hg was reported in 21 eyes treated with Ahmed valve implantation (18),
whereas a 79% success rate was obtained at both 1 and 2 years with IOP of 6
to 21 mm Hg with Molteno device implantation (14). A 95% success at 27
months and a 90% success at 52 months with IOP ≤ 22 mm Hg with Molteno
device implantation were reported in 27 eyes with secondary glaucoma
caused by JIA (19). In a prospective case series of 40 eyes, the probability of
the Molteno implant to control IOP ≤ 21 mm Hg was calculated to be 87% at
5 years and 77% at 10 years after surgery (20).
In the child with uveitis, surgery is best undertaken after the
inflammation is maximally controlled with corticosteroid or
immunosuppressant medications for as long as possible preoperatively,
especially in patients undergoing nonurgent glaucoma surgery. In general, it
is preferable for the eye to be quiet for at least 3 months prior to incisional
surgery, although this is not always possible in more urgent cases. A
glaucoma drainage device may be implanted in the standard fashion between
Tenon fascia and sclera and secured in place. The silicone tube is tied off if
the Baerveldt implant is used. The tube is placed through either the limbus or
the pars plana in vitrectomized eyes. The tube and its insertion site are
covered with a patch of preserved donor sclera or cornea. Topical
corticosteroids are used postoperatively. Maximum control of inflammation
preoperatively, intraoperatively, and postoperatively allows the greatest
chance of a successful surgery.
In the pediatric aphakic uveitic patient, postoperative amblyopia therapy
is vital with proper spectacles or an aphakic contact lens. If a glaucoma
drainage implant is placed in a vitrectomized, aphakic, pediatric patient, a
pars plana tube placement will better allow contact lens fitting following
surgery (1).
Cataract
Cataract is one of the most common complications of uveitis, with a
prevalence of up to 50%, and can be attributed to chronic inflammation and
corticosteroid use (21,22). Cataract surgery in pediatric uveitic eyes is
challenging, with unknown risks faced intraoperatively and an uncertain
postoperative course (3,4). Posterior synechiae, delicate iris vessels, pupillary
membranes, miotic pupils, and corneal opacification can increase the
complexity of the case (5).
Indications
Indications for cataract surgery in a child with a uveitic cataract include
visual rehabilitation, prevention or treatment of amblyopia, and improving
the ophthalmologist’s view of the posterior segment.
Cataract extraction with intraocular lens (IOL) implantation has become
increasingly successful over the past 20 years as a result of improved
microsurgical techniques and viscoelastics, as well as more rigorous control
of recurrent inflammation and chronic low-grade inflammation (1,2).
Currently, cataract extraction with endocapsular posterior chamber IOL is an
accepted practice in the management of many adults with uveitic cataract,
provided the inflammation has been quiescent for a sustained period before
surgery (23). However, the decision whether to implant an IOL in a pediatric
patient with uveitis remains controversial.
In the setting of aphakia with inadequate capsular support, there are
several options to correct the resultant large hyperopic refractive error
including (a) aphakic spectacles (best if bilaterally aphakic), (b) contact lens
use, or (c) IOL implantation in the form of (i) anterior chamber IOL (ACIOL)
or (ii) fixation of IOL either with sutures to the iris/sclera or sutureless within
the sclera. Although the sutureless scleral fixated lens technique has been
successfully used in the pediatric population (24), it has only been used a
handful of times in the pediatric uveitic eye, and longer-term studies are
needed. This technique has been successfully used in the adult population
with uveitis (25) with good long-term outcomes. In pediatric eyes with 70
years or more of life ahead, this technique minimizes the associated risks of
corneal decompensation or anterior uveitis reactivation from ACIOLs or
suture erosion from sutured IOLs and maximizes visual pathway
development and visual potential (24).
Procedure
The surgical technique for cataract extraction in a child with uveitis should
include removal of the cataract, including the posterior capsule, all
retrolenticular and cyclitic membranes, and most of the vitreous. It is usually
not possible to fully separate the posterior hyaloid from the retina in a child.
Pars plana lensectomy with pars plana vitrectomy is the preferred approach
(26). For the pars plana approach, the lens and posterior and anterior capsules
are cut and aspirated. This is followed by a near-total vitrectomy. Removal of
the lens capsule and anterior hyaloid must be performed to prevent posterior
capsular opacification (PCO) and to remove the scaffold on which secondary
membranes can form and grow (21). This reduces the likelihood of
subsequent posterior synechiae and cyclitic membrane formation and
decreases the need for postoperative neodymium-doped yttrium aluminum
garnet (Nd:YAG) laser capsulotomy, which may not be feasible in many
children (26–29). A surgical peripheral iridectomy should be considered as
prophylaxis against secondary angle closure if recurrent or chronic
inflammation is anticipated.
Cataract extraction may also be accomplished by a combined limbal
approach with phacoemulsification and pars plana vitrectomy. During the
limbal approach, synechiolysis and pupilloplasty may be performed, or iris
hooks may be used to ensure adequate exposure. Then a capsulorrhexis,
hydrodissection, and phacoemulsification are performed in standard fashion,
and the incision is closed with a suture. This is followed by a pars plana
approach for complete posterior capsule excision, followed by a meticulous
vitrectomy. It is usually not possible, nor desirable, to surgically induce a
posterior vitreous detachment in a child.
The decision for IOL implantation in a pediatric patient with uveitis
remains controversial. The decision must carefully consider the individual
patient and the risk of recurrent inflammation. If chronic or recurrent
inflammation is anticipated, there is a high risk of developing perilenticular
membranes and iris capture. The IOL itself may provide a stimulus for
intraocular inflammation or a scaffold for inflammatory membranes, which
can lead to ciliary body dysfunction, ocular hypotony, and chronic macular
edema. Treatment of these membranes with Nd:YAG capsulotomy may incite
further inflammation and reformation of membranes. This also can be
difficult to perform in the pediatric population. Surgical posterior
capsulectomy, retrolental membranectomy, and near-total vitrectomy
represent more definitive treatments, but even with this approach, significant
perilenticular membranes may persist or recur. In addition, chronic
inflammation refractory to therapy or cyclitic membranes can result in
hypotony, which may necessitate the removal of the IOL (23). Younger age
and recurrent inflammation are risk factors for subsequent IOL explantation
(23). IOL implantation is contraindicated in children with JIA-associated
uveitic cataracts (1,2,21,30). Some will use a staged approach in which the
cataract is removed and the surgeon waits until full quiescence is reached
again. An IOL is then placed several months later.
The choice of IOL may affect postoperative inflammation and formation
of perilenticular membranes. Acrylic lenses are thought to be the least
inflammatory (31,32). In a randomized, double-masked trial of 36 adult
uveitic eyes that underwent cataract extraction with IOL placement with four
types of IOLs (silicone, polymethylmethacrylate [PMMA], heparin-coated
PMMA, or acrylic lenses), acrylic IOLs had the lowest rates of postoperative
inflammation, PCO, and cystoid macular edema and the best postoperative
final visual acuity (31). Another study also showed a lower PCO rate in
acrylic IOLs, when comparing silicone, PMMA, heparin-coated PMMA, and
acrylic IOLs in adults with uveitis (32). Secondary IOL implantation after
cataract removal in children with JIA-associated uveitis resulted in lower
incidence of secondary glaucoma, compared to primary IOL implantation in a
study of 40 eyes of 40 patients (33). A study assessing primary IOL
implantation in 34 eyes of 24 children with chronic JIA-associated uveitis
showed 65% of patients achieved final visual acuity of 20/40 or better,
secondary glaucoma in 24% of eyes, PCO and secondary membrane
formation in 44%, macular edema in 15%, and phthisis in 6%; the median
age at diagnosis was 5.3 years and median age of cataract extraction with IOL
placement was 9.7 years of age (34). In another study of 80 eyes in children
with uveitis, females were more likely to experience recurrent uveitis
postoperatively, and a heparin-surface-modified IOL appeared to reduce the
incidence of postoperative inflammation (35). The use of heparin in the
infusion solution for pediatric uveitic cataract surgery is controversial
(36,37).
If structural issues do not allow for lens placement in the usual position, a
three-piece lens can be fixated with scleral tunnels (Video 48-1 ). Walsh
et al. (24) describe the procedure in the pediatric population. The technique
begins by making a conjunctival peritomy superiorly and inferiorly at the
limbus. Diathermy is typically not used to avoid the risk of scleromalacia. A
standard three-port vitrectomy setup is initiated, with the trocars placed 2 to 3
mm posterior to the limbus depending on the patient’s age. Measuring 1.5 to
2.5 mm from the limbus at 12:00 and 6:00, a 2- to 3-mm long intrascleral
tunnel is made with 25-g trocars (achieved by maintaining a 30- to 40-degree
bevel) both superiorly and inferiorly. A toric marker is used to ensure a
consistent 180-degree separation of the tunnels, and mild conjunctival
displacement prior to tunnel creation helps bury the haptic following
externalization. Biasing the distal end of the tunnel slightly closer (0.25 to 0.5
mm) to the limbus may decrease lens tilt and decentration. For the left eye,
these tunnels end up extending nasally from the 12:00 sclerotomy and
temporally from the 6:00 sclerotomy and vice versa for the right eye. In the
pediatric setting, if not already done so, a vitrectomy is completed. When
implanting a three-piece IOL, a superotemporal paracentesis is created with a
no. 75 supersharp blade, and viscoelastic is injected into the anterior
chamber. One then makes a standard CCI with a 2.8-mm keratome. It is best
to choose the IOL power in conjunction with the pediatric ophthalmologist
who will be managing the patient’s refractive error for years to come. A
three-piece lens is then inserted using a monarch injector, either leaving the
trailing, superior haptic outside the eye through the CCI or suspended on iris.
Intraocular forceps are inserted through the inferior 6:00 cannula comprising
the intrascleral tunnel, and under direct visualization, one engages the tip of
the leading haptic in the mid-vitreous cavity with forceps. It is important to
attempt to grasp the tip of the haptic to prevent haptic kinking and/or
breakage. The haptic tip is then brought to the internal opening of the
cannula, and one then slowly slides the cannula up the shaft of the forceps.
Finally, the haptic is externalized through the sclerotomy. Once externalized,
the haptic is left to rest on the surface. For the trailing haptic, the same
forceps are used through the superior 6:00 cannula into the anterior vitreous
with the technique of externalization performed, as above. If there is
difficulty in grabbing the trailing haptic, one may enter the anterior chamber
via the paracentesis with another intraocular forceps and “hand” the haptic to
the forceps engaged in the cannula. Once both haptics are externalized, the
tips may be flanged with gentle low-temp cautery and/or gently reengaged
into the tunnel proper. The conjunctiva is reapproximated to overly the
haptics, and one may choose to further fixate the haptics to the sclera with 9-
0 nylon suture if the child is very young with potential for significant eye
rubbing. A repeat limited vitrectomy is performed to remove any debris
and/or viscoelastic, the main wound is sutured with 10-0 nylon, and the
trocars are removed. We typically suture all sclerotomies in a pediatric case
with 7-0 absorbable polyglactin (Vicryl) sutures and close the conjunctiva
with 6-0 plain gut suture. The use of atropine is avoided, but acetylcholine
chloride intraocular solution (Miochol-E, Novartis, East Hanover, NJ) may
be used at the end of the case.
Management of Inflammation
Vigilant control of inflammation preoperatively, perioperatively, and
postoperatively is critical for children with uveitis undergoing cataract
extraction. Some surgeons will prophylactically increase a patient’s
immunosuppression and/or add oral steroids before surgery in order to
maintain better control after cataract surgery. Cataract surgery has an
increased chance for a successful outcome if it is performed after the
inflammation has been quiescent for as long as possible, ideally more than 3
months. Surgery in the presence of active inflammation carries a much higher
potential for complications. The optimal time to operate is after the active
inflammatory phase of the uveitis has burned out (38). In some patients with
chronic inflammation, it is not possible to achieve complete quiescence.
Although attempts should be made to maximally quiet the eye, surgery
should not be delayed if the delay would lead to irreversible loss of vision or
damage to the eye.
Systemic immunosuppressive therapy may be needed for adequate
preoperative control of inflammation in children with JIA-associated uveitis.
In 2003, a favorable outcome was reported in six eyes of five children
younger than 13 years with JIA-associated uveitis, all of whom underwent
standard phacoemulsification cataract extraction without a posterior
capsulectomy and with posterior chamber IOL implantation (7).
Postoperative visual acuity was 20/40 or better in all six eyes. Median follow-
up was 44 months. Preoperatively, four children (five eyes) were on low-dose
methotrexate immunosuppressive therapy for a median length of 1.25 years
before surgery. Preoperatively, two children (three eyes) were on additional
immunosuppressants or systemic corticosteroids. All eyes received frequent
topical corticosteroid therapy for a median of 2 weeks preoperatively and 8.5
weeks postoperatively. Five eyes required Nd:YAG capsulotomy. One eye
needed a subsequent surgical posterior capsulectomy and pars plana
vitrectomy. The authors concluded that children with JIA-associated uveitis
might have favorable surgical outcomes after cataract surgery with posterior
chamber IOL implantation provided they had adequate long-term
preoperative and postoperative control of intraocular inflammation with
systemic immunosuppressive therapy and intensive perioperative topical
corticosteroid therapy (39). As previously mentioned, a second study
assessing primary IOL implantation in 34 eyes of 24 children with chronic
JIA-associated uveitis showed 65% of patients achieved final visual acuity of
20/40 or better, secondary glaucoma in 24% of eyes, PCO and secondary
membrane formation in 44%, macular edema in 15%, and phthisis in 6% of
eyes (34).
Perioperative control of inflammation is vital, as the pediatric uveitic eye
often has an intense postoperative inflammatory reaction. The goal of
perioperative medications is to control and reduce the anticipated severe
postoperative inflammation. Although specific medications and dosing
schedules vary, the concepts are similar. In general, surgeons will
prophylactically add oral steroids or increase immunosuppression 1 to 3 days
preoperatively, even if there is no evidence of active inflammatory disease. If
the child is not on systemic immunosuppression and the uveitis has been
quiescent for an extended period of time, aggressive topical therapy may be
enough, although most surgeons will give a short course of oral steroids
before and after surgery. Intraoperatively, patients may receive a periocular
steroid injection, if possible, or an intravenous bolus of steroids or both, if
previous steroids have not been started. Periocular injections may be
delivered to the sub-Tenon’s space. In small children, 20 mg of triamcinolone
acetonide is effective (40). A study published by Cleary et al. (41) has shown
that 4 mg (4 mg/0.1 mL) of intracameral preservative-free triamcinolone
(Triesence, Alcon, Inc., Fort Worth, TX) used at the conclusion of the
cataract surgery can effectively control postoperative inflammation in a safe
manner in the pediatric population. Postoperatively, patients should receive
aggressive anti-inflammatory treatment in the form of aggressive topical
steroid regimens, increased immunosuppression, or oral steroids. The
increase in immunosuppression should remain for at least 3 to 6 months,
though longer if smoldering inflammation appears to persist.
Postoperatively, the eye may appear quiet at first but then may respond
with an intense inflammatory reaction, which typically peaks about 48 hours
after surgery and lasts for 5 to 6 days even in the presence of high doses of
systemic corticosteroids. Postoperatively, children with uveitis should be
examined very closely, every day or every other day, for the first week for
close monitoring (2). Occasionally, a severe dense fibrin that is refractory to
steroids and NSAIDs may occur in the anterior chamber within 6 days after
surgery. Recombinant tissue plasminogen activator (rt-PA) may be used to
treat dense fibrin. A report on 11 pediatric eyes, 3 with uveitis, who
developed dense fibrin clots after cataract surgery that was refractory to
intensive prednisolone acetate and cycloplegic agents showed a benefit of rt-
PA (42). Injection of rt-PA, 100 μL of 10 μg/100 μL, into the anterior
chamber thought a limbal paracentesis, resulted in complete resolution of
fibrin formation in all eyes, except for two eyes in a JIA patient, within 24
hours. The physician may decide to continue corticosteroids at the same time
if there is no improvement in inflammation over the first 5 days
postoperatively, because there can be a delay in treatment response; however,
worsening of inflammation in spite of standard care therapy should prompt
the physician to intensify the treatment or add an additional medication to the
regimen. Aggressive anti-inflammatory treatments, even in the absence of
inflammation, are key to prevent such occurrences, as intracameral t-PA does
require sedation for the child.
The surgery is only one part of the visual rehabilitation in a child.
Postoperative amblyopia therapy under the care of a pediatric
ophthalmologist is equally important for achieving the best possible vision.
The child will require proper postoperative spectacles or contact lenses,
orthoptic evaluation, and likely occlusion therapy. Parents must be educated
and reminded about the critical role of amblyopia therapy. Despite an
excellent anatomic and surgical outcome, visual rehabilitation for an aphakic
eye may be suboptimal because many patients of this age are unable to or
refuse to wear an aphakic contact lens or aphakic spectacles (43). Among
children, contact lens intolerance is reported to range from 17% to 38%
(30,44). The frequent concomitant presence of band keratopathy makes
contact lens fitting difficult. Multiple lost contact lenses carry a substantial
expense for the family. Children often are intolerant of aphakic spectacles
because of the aniseikonia and weight. The age of the child at the time of
surgery, the presence or absence of amblyopia, and the presence of
irreversible retinal or optic nerve pathology are the primary factors
influencing the outcome of cataract surgery in the pediatric uveitic patient.
Vitreoretinal Surgery
Scleral buckle procedures, whether encircling or segmental, may be indicated
to treated tractional retinal detachments, combined RRD/TRD, or to relieve
traction caused by snowbanking. In children under the age of 4, the scleral
buckle, if encircling, may need to be segmented to allow growth of the eye,
usually done 3 months after placement as long as traction is settling and other
methods for inflammatory control have been incorporated (Figure 48-2).
FIGURE 48-2 A, B: Color fundus photographs of a 14-
year-old with par planitis with snowbanking causing a
tractional retinal detachment and progression of fluid to
the macula (blue arrows). C, D: Color fundus photographs
after placement of a scleral buckle and cryotherapy with
the red arrow demonstrating the previous area of traction
and fluid that has now resolved and yellow arrow
displaying the scleral buckle. No obvious breaks were
seen.
Pars plana vitrectomy may be performed for the many vitreoretinal

complications secondary to chronic uveitis. Yet, unlike adults, pars plana
vitrectomies in the pediatric population may be more daunting and done only
if necessary, because creating a posterior hyaloid detachment can cause more
problems than it solves. Removal of the vitreous and inflammatory cytokines
and other mediators of inflammation is postulated to have a curative or
moderating effect on the clinical course in patients with intermediate uveitis,
JIA-associated uveitis, and sarcoidosis, though this is not the mainstay of
treatment in these pediatric patients and should be used judiciously.
Indications for pars plana vitrectomy include persistent vitreous
opacification, medically unresponsive vitritis, tractional or rhegmatogenous
retinal detachment, vitreous hemorrhage (Video 48-2 ), epiretinal

membrane creating significant distortion or heterotropia, cyclitic membrane,
prevention or treatment of amblyopia, and improving the ophthalmologist’s
view of the posterior segment. These patients frequently have cataract,
posterior synechiae, and vitreous opacification that must be addressed either
prior to or at the time of vitreoretinal surgery. Final visual prognosis often is
limited by the presence of macular or optic nerve pathology.
Indications
There are four objectives in pars plana vitrectomy in patients with chronic
uveitis. The first is to remove opacities in the visual axis. The second is to
remove vitreomacular membranes producing retinal traction or epiretinal
membranes causing macular pucker. The third is to remove cyclitic
membranes or thickened anterior cortical hyaloid causing hypotony. The
fourth is to alter the course of vitreous inflammation refractory to medical
therapy or persistent cystoid macular edema.
Removal of opacities in the visual axis

In cases of vitreous inflammation refractory to medical therapy or vitreous
hemorrhage secondary to neovascularization, a pars plana vitrectomy is
performed to clear the visual axis by the removal of media opacities, vitreous
debris, and/or hemorrhage. Typically, the posterior hyaloid cannot be fully
lifted in a child, as it comes off in sheets; a surgeon is more likely to create
vitreous schisis than a true posterior vitreous detachment in a child. Rather,
the goal of the surgery is to clear the visual axis.
Alter the course of vitreous inflammation refractory to medical
therapy
Vitrectomy may alter the course of inflammation and result in stabilization of
the disease by removing immunocompetent cells and inflammatory mediators
from the vitreous cavity (27,28,45). Vitrectomy may potentially reduce
cystoid macular edema either by eliminating the contact between an inflamed
vitreous body and the macula or by allowing better penetration or distribution
of corticosteroids and longer duration of the drug (26,27,46). However,
others have noted that although vitrectomy removes vitreous debris and
appears to allow for easier intraocular penetration of corticosteroid, no
change in the inflammation process was observed postoperatively and can
actually induce more inflammatory changes later (2).
Removal of vitreomacular membranes

Membranectomy may be performed after pars plana vitrectomy to remove
inflammatory membranes from the retina and ciliary body in cases of severe
traction or hypotony. Anteroposterior vitreomacular traction from an
inflamed vitreous body may result in chronic macular edema or a shallow
tractional retinal detachment. Tangential traction caused by epiretinal
membranes may cause decreased vision and secondary macular edema. These
membranes may be removed at the time of vitrectomy by delamination or en
bloc techniques. Traction also may lead to retinal breaks in atrophic retina,
creating a combined tractional–rhegmatogenous retinal detachment, further
complicating the surgical case. These combination cases may be better served
with the addition of a scleral buckle, depending on the child’s age and
location of the pathology, as more peripheral pathology would be better
supported with a scleral buckle.
In cases where chronic macular edema is present and it is difficult to tell
how much is the result of traction, increased immunosuppression may a
better first-line treatment, especially if there is severe photoreceptor
disruption, as visual recovery may be limited. In cases of a shallow
detachment that is purely tractional in nature, using a scleral buckle to relieve
traction may be a better first line of treatment.
Caution should be taken when considering membrane removal as these
membranes tend to be very adherent, and more damage may occur while
attempting to delaminate the membrane. One must also consider the
chronicity of the macula edema and structural integrity of the ellipsoid zone,
as visual function is not always improved with this intervention and,
therefore, not worth the risks.
Removal of cyclitic membranes or thickened anterior cortical

hyaloid
A cyclitic membrane may occur and may result in hypotony. Chronic traction
of the ciliary body from a cyclitic membrane or a thickened anterior cortical
hyaloid may result in ciliary body detachment and reduced aqueous humor
formation. Pars plana vitrectomy and cyclitic membrane removal can be
performed to reattach the ciliary body. A skilled assistant is required to assist
in scleral depression in the region of the ciliary body so that the surgeon can
dissect and remove the anteriorly located membranes or surgically segment
them to reduce traction. Alternatively, endoscopic vitrectomy can be
performed to remove the cyclitic membrane (46).
Procedure
Vitreoretinal surgery is performed using a standard three-port vitrectomy
procedure. A longer infusion tip (4 to 6 mm) may be required in uveitis
patients to accommodate scleral thickening, choroidal edema, or retinal
separation (1). Direct visualization to confirm that the infusion cannula is
within the vitreous cavity is critical before turning on the infusion.
The infusion is turned on and the endoillumination probe and vitrectomy
cutter are introduced. A complete vitrectomy is performed, although the
posterior hyaloid cannot typically be fully lifted in children. Preretinal and
epiretinal membranes are dissected or stripped from the retinal surface using
a combination of scissors, membrane picks, and forceps. If there is vitreous
base exudation or neovascularization, it may be treated with laser scatter
photocoagulation or cryopexy. Scleral depression is used at the completion of
the case to inspect 360 degrees for any untreated breaks or tears. The hyaloid
is often very adherent in these cases, and attempting to lift it from the retina,
especially in thinner, peripheral retina, can lead to retinal breaks. Any breaks
are treated with retinal laser or cryopexy, and all attempts need to be made to
remove traction from these tears, including a scleral buckle element of sorts,
if needed. Long-acting gas, or more likely, silicone oil, for tamponade of at
least 3-month duration may be needed. Aggressive preoperative
inflammatory control is needed. This is mostly achieved with systemic
medications versus local steroids as systemic immunosuppression is likely to
be effective and less likely to cause other possible postoperative
complications, such as glaucoma.
Lasers
In the nonuveitic eye, lasers often are used to perform peripheral iridotomy
and posterior capsulotomy and to treat retinal or subretinal
neovascularization. In a patient with uveitis, however, a laser procedure may
incite extensive inflammation, leading to additional drop out and increased
retinal neovascularization, entering the child into a vicious cycle.
Periprocedural control of inflammation is important, with frequent topical
corticosteroid dosing and consideration of periocular or systemic
corticosteroids.
Surgical Iridectomy Preferable to Peripheral

Iridectomy
Patients with uveitis may develop posterior synechiae sufficient to produce
relative pupillary block with subsequent iris bombe configuration, narrow
angles, and possibly angle-closure glaucoma. Ordinarily, laser peripheral
iridotomy is the therapeutic procedure of choice. In the pediatric uveitic
patient, the positioning of a child at the Nd:YAG laser is difficult.
Additionally, chronic or recurrent inflammation often will result in
subsequent closure of the laser peripheral iridotomy. A surgical iridectomy
performed with the patient under general anesthesia is preferable to laser
peripheral iridotomy in the pediatric uveitic patient.
Posterior capsulotomy versus surgical posterior capsulotomy

Cataract formation is a frequent complication of uveitis. After
phacoemulsification cataract extraction with posterior chamber IOL
implantation, thick perilenticular membranes often form in the pediatric
population, particularly in patients with chronic or recurrent uveitis. These
perilenticular membranes may result in ciliary body dysfunction, ocular
hypotony, and chronic macular edema. The thick inflammatory membranes
often require multiple sessions of Nd:YAG capsulotomy, which may incite
further inflammation, with subsequent reaccumulation. Eventually, a surgical
posterior capsulectomy, retrolental membranectomy, and total vitrectomy
may be necessary. If IOL implantation is attempted in a pediatric uveitic
patient, especially a patient with JIA, a posterior capsulectomy with the
removal of the anterior hyaloid face and a thorough near-total vitrectomy
should be considered in attempts to eliminate the “scaffold” along which
secondary membranes may subsequently form (23). This reduces the
likelihood of subsequent posterior synechiae and cyclitic membrane
formation and decreases the need for postoperative Nd:YAG laser
capsulotomy, which may be difficult in small children (26–29,47).
Panretinal or segmental photocoagulation for retinal

neovascularization
Patients with sarcoidosis may develop large areas of retinal capillary
nonperfusion and secondary retinal or disc neovascularization. Both retinal
and disc neovascularization may regress in patients with sarcoidosis in up to
50% of patients treated by systemic medical therapy alone (48,49). When
persistent or progressive peripheral retinal neovascularization causes
recurrent vitreous hemorrhage, laser photocoagulation to the avascular areas
of retina is indicated. Local grid treatment is applied to the areas of ischemia
adjacent to retinal neovascularization to reduce the neovascular drive. Wide-
field fluorescein angiography can define the areas of ischemia and help guide
photocoagulation. Laser photocoagulation is often accompanied by a
significant increase in cystoid macular edema (48). Steroid therapy and/or
antivascular endothelial growth factor therapy can be used in combination
with laser treatment to blunt the inflammatory response.
Peripheral neovascularization
Patients with intermediate uveitis may develop peripheral retinal
neovascularization. These patients have thick ropy neovascularization in the
far periphery extending over the ora serrata, which often is obscured from
visualization by pars plan exudation and vitritis. In patients with unremitting
chronic pars planitis that is recalcitrant to steroid or nonsteroidal anti-
inflammatory drug (NSAID) therapy, cryotherapy and scatter laser
photocoagulation of the peripheral retina have been reported to be effective in
causing regression of peripheral neovascularization and reduction of
inflammatory activity. This treatment can reduce the frequency of vitreous
hemorrhage and may reduce the severity of the intermediate uveitis and
cystoid macular edema. Cryotherapy is thought to reduce inflammation by
eliminating the inflammation stimulus in the peripheral retinal tissue (50,51).
Reduction in neovascularization may be caused by direct vessel ablation or
by destruction of ischemic retinal tissue (51,52). In more recent times, if
steroid or NSAID therapies are not enough, children are put on systemic
immunosuppression before cryotherapy is done to reduce some of the
inflammation this procedure may cause. Regression of snowbanking and
neovascularization may occur with aggressive systemic immunosuppression
(48,49). If regression is not seen and the pediatric patient is on aggressive
therapies, cryotherapy to the deep retinal neovascularization may be
performed.
In children, cryotherapy is performed under general anesthesia.
Retrobulbar or subconjunctival anesthesia may be given for intraoperative
and postoperative pain control. Cryotherapy is applied directly to the areas of
exudation at the pars plana using a double freeze–thaw technique (29,50).
The ice ball should be seen to cover the exudative area and
neovascularization. Uninvolved pars plana and retina are treated with one
Cryo-tip width beyond the involved area. In areas where the visualization of
the ice ball is precluded, freezing should be continued for a time interval
similar to that required in adjacent areas with adequate visualization.
Periocular steroids may be administered following the procedure. The
procedure may be repeated in 3 to 4 months if there is residual disease.
Cryotherapy was first advocated in 1973 (50) and was later reported to
eliminate inflammation in 78% of 27 eyes (53). Visual acuity improved or
remained unchanged in 89% of eyes and eliminated the need for
corticosteroids therapy in 90% of eyes (53). In the presence of better, more
tolerable, and safer immunosuppression, cryotherapy has become more of a
secondary procedure.
Panretinal scatter photocoagulation has been shown to be effective in the
treatment of peripheral neovascularization associated with intermediate
uveitis. Regression of neovascularization, improvement of cystoid macular
edema, and stabilization of inflammation were reported in 10 eyes with
scatter diode or argon photocoagulation treatment, either alone or in
combination with pars plana vitrectomy (54).
Several rows of photocoagulation are delivered to the inferior retinal
periphery, just posterior to the area of exudation or neovascularization.
Treatment may be carried out with either argon endophotocoagulation or an
indirect ophthalmoscopic delivery system using argon or diode
photocoagulation. Laser photocoagulation appears to be a safe and effective
alternative to cryotherapy and is a useful adjunct during a therapeutic pars
plana vitrectomy.
SLOW-RELEASE STEROID
TREATMENT FOR UVEITIS
Two sustained-release, steroid-based implants or drug delivery devices are
currently are on the market. One device, the Retisert implant, is a reservoir-
based, surgically implanted device containing 0.59 mg of fluocinolone
acetonide (FA); it is designed to release the drug over a 3-year period. The
device initially was developed by Control Delivery Systems (Watertown,
MA) and now is licensed by Bausch & Lomb (Rochester, NY) (Figure 48-3).
The device has been shown to be extremely effective for the treatment of
uveitis, even in patients who have had poor control of their disease with
aggressive systemic therapy (Figures 48-4 and 48-5) (55). A pilot study by
Patel et al. (56) has shown that the Retisert can effectively control posterior
inflammation in the pediatric patients. A total of six eyes of four children
were enrolled in this study. Retisert implants were able to control the
inflammation of all six eyes. The safety profile was very similar to what has
been described in adult patients. One of the six eyes of the pediatric patients
developed visually significant cataract that was surgically removed. Two
patients also developed high IOP, which were treated with glaucoma shunting
procedures. Unfortunately, not much more data is available for the pediatric
population; hence, safety and effectiveness in pediatric patients below the age
of 12 years have not been well established (57).
FIGURE 48-3 FA reservoir device (Retisert, Bausch +
Lomb, Rochester, NY) alongside rulers.
FIGURE 48-4 A: Color fundus photograph 1 year before
FA implant. Optic nerve swelling and peripapillary
necrotizing retinitis with intraretinal hemorrhage are seen.
Image taken with a 30-degree fundus camera. B: Six
months after FA implant. Healed retinitis. Image taken
with a 50-degree fundus camera.
FIGURE 48-5 Fluorescein angiogram taken just before
(A) and 4 months after (B) FA device implantation.
Midarteriovenous phase (A) and late phase (B) are shown.
(Courtesy of Glenn Jaffee, MD.)
Fluocinolone Implantation Procedure

The procedure for surgical implantation of the fluocinolone implant in kids
>6 years of age is similar to that in adults. No implants have been done in
children under the age of 6 years. A limbal-based conjunctival peritomy is
performed in the inferotemporal quadrant using Westcott scissors and 0.3
forceps. The inferotemporal quadrant is not used if there is significant
pathology of the vitreous base in that location; an alternate site is chosen,
with a preference for an inferior-based location. All episcleral vessels are
cauterized using eraser-type cautery. A 4-mm circumferential incision is
performed 3 to 4 mm from the limbus using a 15-degree blade or the
equivalent. The wound is inspected to assure that the uvea has been
adequately incised using a no. 15 blade to create a full-thickness incision
through sclera. The fluocinolone implant, which has a double-armed 9-0
Prolene suture with TG-175 needles that is passed through the eyelet at the
base of the strut, is grasped with nontoothed forceps and inserted through the
incision into the vitreous cavity. All prolapsed vitreous is excised with a
vitrector. The implant is sutured into place by grasping the needle of the
anterior arm of the suture and performing a radial full scleral thickness bite
through the wound emerging through the scleral 2 mm anterior to the wound.
If the eye is not stable from an IOP standpoint, an infusion port may be
placed for the stabilization of the eye.
The process is repeated using the needle attached to the posterior arm of
the suture, going near full thickness through the wound emerging 2 mm
posterior to the wound. The elevated aspect of the implant housing the drug
should face the center of the eye. The implant is tied firmly into place using a
3-1-1 knot, with the ends of the suture left long after the needles have been
removed. The rest of the wound is closed with four radial sutures of 8-0
Prolene (two on either side of the central anchoring suture) tied with a 2-1-1
knot, with the suture ends cut short and the knots rotated into the sclera. The
long ends of the central anchoring suture are tucked under the radial sutures
on either side. The conjunctiva is closed using either plain gut or Vicryl
suture. The implant is inspected with indirect ophthalmoscopy to assure that
it is appropriately placed in the vitreous cavity. Standard subconjunctival
injections of antibiotic and steroid are given at the end of the procedure.
Dexamethasone Implantation Procedure

A second FDA-approved, sustained-release, steroid-based drug delivery is a
dexamethasone bioerodible device called Ozurdex, designed to release drug
over a 6-month period (Ozurdex, Allergan Inc., Irvine, CA) (Figure 48-6). It
is an office-based, 23-gauge injectable implant. Previously, an earlier 20-
gauge version of the dexamethasone implant, which required surgical
implantation in the operating room, was shown to be effective in five boys
with chronic cystoid edema associated with intermediate uveitis (Figure 48-
7). The currently FDA-approved device does not require surgical cutdown
and is injected using a 23-gauge needle through the pars plana
inferotemporally, typically under general anesthesia. The implant contains
0.7 mg of dexamethasone in the Novadur solid polymer delivery system that
is comprised primarily of polylactic–polyglycolic acid. The polymer breaks
down as the dexamethasone is being released. Eventually all the
dexamethasone is released, and the polymer continues to break down to
carbon dioxide and water, with no remaining constituents in the eye. There
are very limited data showing the efficacy and safety in children (58). A
retrospective case series in 14 eyes of 11 children with posterior segment
uveitis showed control of inflammation with the dexamethasone implant and
improvement in CME and visual acuity in all but one eye within a month.
Unfortunately, 31% had a relapse within 6 months with a median time to
relapse at 4 months (59). Another series of pediatric patients with
intermediate or posterior uveitis in which the dexamethasone intravitreal
implant was used demonstrated that, even with repeated injection when
necessary, the treatment was found to be safe and effective (60).
FIGURE 48-6 Dexamethasone bioerodible device
(Ozurdex, Allergan Inc., Irvine, CA).
FIGURE 48-7 Fluorescein angiogram of a 12-year-old
boy with intermediate uveitis with chronic cystoid macular
edema just before (A) and 2.5 months after (B)
dexamethasone bioerodible device implantation.
Though the use of the local steroid implants may appear attractive, caution
must be taken when using these implants in the pediatric population, as
cataracts and glaucoma are real concerns and can complicate an already
complicated eye. Cataract extraction and filtering procedures are already
more complex in the pediatric population, let alone those also with uveitis.
As with most of the studies, surgical results were usually better achieved if
these complex eyes were being treated with systemic immunosuppression.
SURGICAL APPROACH TO UVEITIS BY

DISEASE
This chapter concludes with specific surgical approaches to a few of the more
common pediatric uveitides: ocular toxocariasis, sarcoidosis, intermediate
uveitis, and JIA-associated uveitis. The focus of this section of the chapter is
on the specific surgical approaches for the complications and sequelae of
these uveitides.
Ocular Toxocariasis
Ocular toxocariasis is a monocular disease that affects children.
Pathogenesis
Toxocariasis is an infectious disease caused by the invasion of tissue by the
larvae of Toxocara canis. T. canis is a roundworm of dogs. It is a ubiquitous
worldwide parasite. The incidence of infected puppies has been estimated to
vary from 33% to 100% worldwide (61). Human infection occurs when a
child ingests soil-containing ova. The ova hatch in the small intestine, and the
larvae pass through intestinal wall. The larvae migrate to various tissues,
including the eye, where they may wander about aimlessly for weeks or
months, or the larvae may become encysted by a focal granulomatous
reaction, where they can remain alive for months or years (62). The adult
worms do not develop in humans; thus, Toxocara eggs will not be found in
the stool of an infected person.
Ocular toxocariasis typically affects children at an average age of 7.5
years (range 2 to 31 years); 80% of the cases occur in children younger than
age 16 years (63). Intermediate uveitis from Toxocara is almost always
unilateral, whereas idiopathic intermediate uveitis is bilateral in 70% to 90%
of cases (64); thus, ocular toxocariasis must be suspected in all cases of
unilateral intermediate uveitis, particularly in children.
Clinical Signs
Ocular toxocariasis presents in a variety of clinical patterns. This is on the
differential for leukocoria. A variable amount of vitreous inflammation is
present. The main ocular findings include peripheral retinochoroiditis;
posterior retinochoroiditis, also called a posterior pole granuloma; and
vitritis. Fibrous tractional bands may result in epiretinal membrane formation,
traction retinal detachment, and combined traction–rhegmatogenous retinal
detachment. Vitreoretinal surgery has been shown to be effective in clearing
inflammatory debris from the vitreous cavity and reattaching the macula in
some eyes with retinal detachment (65).
The posterior pole granuloma is a white, elevated, intraretinal or
subretinal mass from 0.5 to 4 disc diameters in size, with tractional bands
from mass to the macula or to the optic disc (Figure 48-8). The granuloma is
the larva of Toxocara surrounded by an eosinophilic abscess with a
granulomatous inflammatory reaction (66). In the periphery, hazy white
inflammatory masses may be seen at the pars plana (Figure 48-9) and often
are associated with retinal folds extending posteriorly toward the optic disc.
Tractional bands from these inflammatory masses to the optic disc may result
in a retinal fold through the macula, macular traction, or macular dragging.
Toxocara endophthalmitis usually presents with a white eye and little pain
but with marked vitreous inflammation, granulomatous keratic precipitates,
and anterior chamber inflammation. A white retrolental mass, which
represents an organization of the inflammatory process into a cyclitic
membrane between a detached retina and lens, may occur and must be
differentiated from retinoblastoma. Amblyopia also may occur as a
complication of toxocariasis.
FIGURE 48-8 Posterior pole of an 8-year-old boy with
inflammatory granuloma caused by T. canis. Note traction
on the surrounding retina. Vitritis has been controlled with
systemic prednisone.
FIGURE 48-9 Fluorescein angiogram of pars plana
inflammatory exudation in an 8-year-old boy with ocular
toxocariasis.
Diagnosis
The diagnosis of ocular toxocariasis usually is presumptive based on the
clinical findings and correlation with serum antibodies to T. canis measured
by serologic ELISA. A 1:8 dilution is considered positive in the presence of
the appropriate clinical findings (62). In cases where the diagnosis is
uncertain, ELISA on aqueous or vitreous biopsy specimens may facilitate the
diagnosis. The presence of eosinophils in aqueous or vitreous biopsy
specimens also suggests the diagnosis of toxocariasis.
Medical Management
Ocular toxocariasis is treated medically. Therapy is directed at the
inflammatory response to prevent inflammation-induced tissue injury and
secondary membrane formation. The inflammation is treated with
corticosteroids, either topically, periocularly, or systemically. If the
inflammation is refractory to steroid treatment, therapy with anthelmintics
directed at the larvae itself may be attempted. Thiabendazole 50 mg/kg/d for
7 days has been proposed if steroid therapy failed (67). There have been
reports of clinical improvement of ocular toxocariasis treated with
thiabendazole (25 mg/kg twice a day for 5 days), albendazole (800 mg twice
a day for 6 days), and mebendazole (100 to 200 mg twice a day for 5 days)
(68).
Indications for Surgery

Intraocular inflammation may lead to macular detachment through either
direct vitreomacular traction or epiretinal membrane formation creating a
macular pucker. Traction also may lead to retinal breaks in atrophic retina,
creating a combined traction–rhegmatogenous detachment. A pars plana
vitrectomy may be beneficial for patients who have not had a satisfactory
response to medical treatment or for those who have marked vitreous fibrosis
and tractional complications. Indications for surgery are tractional retinal
detachment, epiretinal membrane formation creating significant distortion or
heterotropia, impending retinal detachment, or cyclitic membrane. When a
retinal detachment appears inevitable, early vitrectomy to eliminate the
vitreous tractional band from the granuloma has been successful, providing
the preoperative visual acuity was good (69).
Surgical Management
For a tractional macular detachment, pars plana vitrectomy and membrane
peeling may be performed in a manner similar to that for other types of
epiretinal membrane (69). The fibrous membranes located between the
peripheral granuloma and the optic disc usually have extensions into the
underlying retina and need to be carefully lifted off from the retinal surface
before they can be severed. These membranes usually remain tightly adherent
to the optic disc and peripheral granuloma; therefore, they often need to be
circumcised rather than delaminated or peeled. Because the granulomas
appear to be an intimal part of the retina, attempts to extirpate the retinal
granuloma usually are unsuccessful and may cause undesirable
complications. Therefore, the granulomas usually are left in place. In most
cases, macular traction can be released without removal of the granuloma,
although scleral buckling is necessary in some cases (Video 48-3 ).
Outcomes
The use of pars plana vitrectomy for a tractional retinal detachment in cases
related to toxocariasis in the pediatric population has been reported.
Anatomic success for reattachment ranges from 66% to 83% (69–71).
However, the rate of redetachment is fairly high, ranging from 24% to 48%.
Vitreous surgery to treat macular traction due to T. canis was reported in
1977 (72). Later, 12 (71%) of 17 eyes attained retinal reattachment after
vitreoretinal surgery for retinal detachment associated with complications of
toxocariasis (70). Fifteen eyes (88%) had stabilization or improvement in
visual acuity. Redetachment occurred in 4 (24%) of 17 eyes. In a series in
which 10 (83%) of 12 eyes attained retinal reattachment after vitrectomy for
tractional macular disorders associated with toxocariasis, 9 eyes (75%) had
stabilization or improvement in visual acuity (69). Redetachment occurred in
5 (42%) of 12 eyes and was associated with epiretinal membrane
proliferation. The presence of a preoperative tractional fold through the
macula was associated with a poor final visual outcome. In 12 eyes with
toxocariasis-associated complications, only 9 had traction-related macular
problems (71). The other eyes had vitreous opacification alone. Of those with
tractional detachment, 6 (66%) of 9 eyes were reattached, and these cases
improved or were stabilized visually. Redetachment occurred in three eyes
(33%). Recovery of the intraocular larvae usually is usually not possible
because of the small size of the organism (~18 to 21 μm) or because of
destruction of the organism by inflammatory cells. The unintentional
extraction of toxocariasis larvae during vitreoretinal surgery has been
reported (73). Visual prognosis may be guarded in this population, as only
18% of pediatric patients in a series of 45 eyes attained best corrected visual
acuity of 20/50 or better (74).
Postoperative Care
It is essential to remember that a successful operation is only part of the
visual rehabilitation in a child. Postoperative amblyopia therapy in
conjunction with a pediatric ophthalmologist is necessary. The primary
factors influencing the outcome of surgery for the complications of ocular
toxocariasis are factors such as the age of the patient, disease duration before
diagnosis, preexisting amblyopia, and compromise of the macula. The
prognosis for improved visual acuity and normal binocular vision is better
when the onset of the disease occurs in older patients, and the disease is
detected early in its course.
Sarcoidosis
Sarcoidosis is an idiopathic, noncaseating granulomatous inflammatory
disease that may affect any part of the body, most commonly the lungs, but
may include the eye and orbit, lymphatics, heart, kidneys, musculoskeletal
system, and skin (75). The incidence of sarcoidosis is 6 to 10 in 100,000
(76,77). It is 10 times more frequent among African Americans than among
Caucasians (76). Although 75% of affected patients are between 20 and 50
years old, sarcoidosis may occur in children (78).
Children aged 5 years or younger may develop a childhood sarcoid
arthritis (71). The classic triad of symptoms consists of the skin, eye, and
joint lesions; initial pulmonary involvement is rare. It can be easily
misdiagnosed as JIA, which also presents with joint and eye findings (79,80).
However, children with JIA-associated uveitis usually suffer from
pauciarticular arthritis, are antinuclear antibody (ANA) positive, and rarely
develop skin lesions, whereas children with sarcoidosis usually develop
polyarthritis, are ANA negative, often exhibit skin lesions in the form of
erythema nodosum, and have elevated serum angiotensin-converting enzyme
(ACE) (81). One must also consider Jabs-Blau syndrome in patient with
granulomatous involvement of the skin, eyes, and joints in the absence of
lung involvement as this disease has a genetic component, with a known
knockout mutation in the CARD15/NOD2 gene.
Diagnosis
Initial systemic workup consists of a chest radiograph, serum ACE (though
this is may be elevated in children without sarcoidosis), lysozyme, serum and
urine calcium, and liver enzymes (81). If the results of these studies are
negative, high-resolution chest computed tomography and pulmonary
function test should be performed. Gallium scanning may show uptake in the
lacrimal and parotid glands, also known as the panda sign. Sarcoidosis may
first present with uveitis only, with a negative workup, and with extraocular
manifestations evolving slowly over several years (82). Thus, laboratory and
radiologic tests should be repeated periodically in “sarcoid suspects” (81).
The definitive diagnosis of sarcoidosis is a histopathologic one and has
been made by biopsy of the parotid gland, lung, conjunctiva, skin, or lacrimal
gland (81). Conjunctival biopsy is a simple procedure to perform on easily
accessible tissue with a low complication rate. Random conjunctival biopsies
have been reported to be positive in 55% to 71% of patients with biopsy-
proven extraocular sarcoidosis (83,84) and in 28% of patients with suspected
sarcoidosis (83). True sarcoid granulomas may be too small to be seen with
slit-lamp examination, and prominent nodules often turn out to be large
follicles, an ectopic lacrimal gland, or foreign body fibrosis rather than
noncaseating granulomas (85). Because true sarcoid granulomas are often too
small to be seen with slit-lamp examination, random conjunctival biopsy may
be reasonable.
Conjunctival biopsy
The lower fornix is the preferred site for conjunctival biopsy. The lower lid is
retracted and a strip of stretched conjunctiva is excised with Westcott
scissors. The optimal size of biopsy specimen is approximately 1 cm long × 3
mm wide. A topical antibiotic is instilled and pressure is applied for 5 to 10
minutes to prevent hemorrhage and soft tissue edema. No suturing is needed.
There have been no reports of infection or symblepharon formation after
conjunctival biopsy. Multiple sectioning is recommended because the
noncaseating granulomas may be sporadic throughout the conjunctival tissue
(84).
Clinical Signs
The ocular manifestations of sarcoidosis in children are similar to those seen
in adults and include granulomatous anterior uveitis, posterior uveitis, pars
planitis, periphlebitis, macular edema, epiretinal membrane, branch retinal
vein occlusion, retinal neovascularization, tractional retinal detachment,
conjunctival and iris granulomas, and keratoconjunctivitis sicca. Severe
visual loss worse than 20/200 has been reported in 6% to 24% of patients
with ocular sarcoidosis and is more common in patients with chronic
posterior uveitis (82,85).
Medical Management
Sarcoidosis is primarily treated medically. Treatment should be directed
toward absolute control of the inflammation to minimize any potential ocular
complications and structural damage. Mild anterior uveitis is treated with
topical corticosteroids and cycloplegics. Systemic steroids are indicated in
refractory anterior uveitis and in patients with posterior uveitis, retinal
neovascularization, or optic nerve compromise. If inflammation persists with
systemic steroids, the addition of systemic immunosuppression may be
indicated. Immunosuppressive medications such as azathioprine,
cyclosporine, methotrexate, or cyclophosphamide may be required for
refractory disease or as part of a steroid-sparing strategy, as steroid therapy is
not a long-term treatment option for the pediatric population (85–88).
Surgical Indications
Vitreoretinal surgery occasionally may be needed to remove nonclearing
vitreous hemorrhages, peel epiretinal membranes, repair tractional or
rhegmatogenous retinal detachments, or remove inflammatory debris from
the vitreous cavity. Additionally, the complications of chronic uveitis, such as
band keratopathy, glaucoma, and cataract, may need to be treated surgically.
The major cause of poor visual outcome in these patients is macular
pathology, such as cystoid macular edema and epiretinal membrane. Vitreous
hemorrhage secondary to retinal neovascularization may cause decreased
visual acuity as well. Secondary glaucoma may occur as a consequence of
severe anterior segment inflammation. Band keratopathy or cataract may
develop as a consequence of chronic uveitis.
Surgical Management
Iris nodules
Surgical excision of iris nodules may help in the diagnosis and control of
sarcoid uveitis refractory to medical management. The surgical excision of
iris nodules for histopathologic confirmation of the diagnosis of sarcoidosis
in two 10-year-old boys in whom anterior uveitis was refractory to medical
management was reported (89) and resulted in control of inflammation. The
authors hypothesize that the iris granuloma may become a focus of continued
cytokine production and ocular inflammation, and they suggested that total
surgical excision of the iris masses may help in the diagnosis and control of
sarcoid uveitis refractory to medical management.
Cataracts
Cataracts may form as a result of recurrent inflammation from ocular
sarcoidosis. Cataract extraction may be achieved through the limbal approach
with phacoemulsification or through a pars plana approach with lensectomy
in combination with a pars plana vitrectomy. Placement of an IOL in young
patients who will experience future flare-ups of inflammation or have an
anticipated chronic course of inflammation places them at high risk for
developing thick retrolental membranes, severe postoperative inflammation,
iris capture, chronic cystoid macular edema, or hypotony. In a retrospective
analysis of 102 patients with sarcoidosis-associated uveitis (87), the
successful implantation of a posterior chamber lens during cataract surgery
was reported in 19 (90.5%) of 21 adult eyes; however, the success rate is
likely to be much lower in the pediatric eye with sarcoidosis-associated
uveitis. After cataract extraction and IOL implantation, young patients with
sarcoidosis were reported to have a marked postoperative flare-up of
inflammation accompanied by significant fibrin and protein within the eye
that peaked 48 hours after surgery and persisted for 5 to 6 days
postoperatively (2). The consideration of placing an IOL is similar to the
earlier discussion for other uveitic conditions. The removal of the vitreous
scaffold intraoperatively (21), careful control of inflammation with high-dose
steroids during the first postoperative week, and follow-up within 5 to 7 days
after surgery are important (2). Tissue plasminogen activator can be used to
clear severe dense fibrin in the anterior chamber.
Glaucoma
Secondary glaucoma with sarcoid uveitis appears to be a particularly poor
prognostic sign and is associated with severe vision loss (90). Glaucoma
associated with uveitis usually is notoriously difficult to treat. Conventional
filtering surgery has a lower chance of success in these patients with ongoing
uveitis because recurrent inflammation promotes fibrous tissue growth and
subsequent closure of the filtering bleb. Some studies report reasonable
success rates using glaucoma drainage devices, that is, success rates of 77%
to 94% with follow-up from 1 to 10 years, with choroidal effusions,
hypotony, and cystoid macular edema being the most common postoperative
complications (14,17–20).
Vitreoretinal Complications
Retinal manifestations of ocular sarcoidosis include retinal vasculitis,
peripheral retinal and disc neovascularization, choroidal nodules or
granulomas with overlying sensory retinal detachment, subretinal
neovascularization, epiretinal membrane proliferation, and tractional retinal
detachment. Patients with sarcoidosis may develop large areas of retinal
capillary nonperfusion and secondary retinal or disc neovascularization. Both
retinal and disc neovascularization may regress in patients with sarcoidosis in
up to 50% of patients treated by systemic medical therapy alone (48,49).
When persistent or progressive peripheral retinal neovascularization causes
recurrent vitreous hemorrhage, laser grid photocoagulation or cryopexy may
be applied to areas of ischemia as defined by fluorescein angiography. Laser
photocoagulation usually is followed by a significant increase in cystoid
macular edema (48). Steroid therapy should be used in combination with
laser treatment to blunt the anticipated inflammatory response. When disc
neovascularization causes recurrent vitreous hemorrhage, panretinal
photocoagulation may be required to cause regression of the disc vessels.
Vitrectomy has a role in removing nonclearing vitreous hemorrhages that
have result from peripheral retinal or disc neovascularization. Active
neovascularization at the time of surgery may be treated with
endophotocoagulation or peripheral cryopexy. Vitrectomy may be needed for
epiretinal membrane peeling or for repair of tractional or rhegmatogenous
retinal detachments.
Pars plana vitrectomy clears the visual axis by removing media opacities
and vitreous debris in patients with vitreous inflammation refractory to
medical therapy. Vitrectomy may alter the course of inflammation and result
in stabilization of the disease by removing immunocompetent cells and
inflammatory mediators from the vitreous cavity (27,28,45). Vitrectomy may
potentially reduce cystoid macular edema either by eliminating the contact
between an inflamed vitreous body and the macula or by allowing better
penetration or distribution of corticosteroids (26,28,54).
In the pediatric sarcoid-associated uveitis patient, surgery is best
undertaken after the eye has been quiescent for as long as possible but at least
for 3 months. Perioperative inflammation must be rigorously controlled with
aggressive topical, periocular, and possibly systemic corticosteroid use.
Maximum control of inflammation preoperatively, intraoperatively, and
postoperatively allows for the greatest chance of a successful surgery.
Intermediate Uveitis
Intermediate uveitis is a relatively common bilateral disease that affects
otherwise healthy children and young adults. The course is variable, ranging
from a mild self-limiting disease to a chronic disease with exacerbations and
remissions. The main ocular finding is a marked exudative response in the
peripheral retina and inflammation in the anterior vitreous. Vision loss is
most commonly due to vitreous debris, cystoid macular edema, epiretinal
membrane, and cataract. Vitreoretinal surgery may be needed for clearing of
vitreous debris as well as attenuation of inflammation and the exudative
response. Cryotherapy or panretinal scatter photocoagulation may be used for
peripheral neovascularization. Uveitic cataracts may be treated with surgical
intervention.
Intermediate uveitis is an idiopathic intraocular inflammatory syndrome
of the peripheral retina and pars plana, which is known by many names, most
commonly pars planitis and peripheral uveitis. Intermediate uveitis has been
reported in 16% to 33% of all uveitis cases presenting among children (91).
The age of onset ranges from 5 to 65 years, with the mean occurrence in the
third decade of life (29). Severe cases tend to present at an earlier age,
whereas older patients have a milder form of the disease. From 70% to 90%
of cases are bilateral, although the majority of cases are asymmetric (92).
Symptoms and Signs

Patients with intermediate uveitis often present with minimal symptoms,
which may include floaters or blurred vision but no pain, photophobia, or
redness of the eye. A mild anterior chamber reaction may be present or
absent. Posterior subcapsular cataracts are the most common anterior segment
complication, ranging from 15% to 60% (93,94). The incidence of glaucoma
is quite low, reported to be 8% in one large series (93). Vitreous cells are the
most characteristic sign for intermediate uveitis, ranging from mild to severe.
Large gray-white or yellow exudative aggregates, also called snow banks,
may be present during active pars planitis. White collagen bands at the pars
plana may be seen chronically in some patients, even during periods of
quiescence. Vitreous yellow-white aggregates, also called snowballs, may be
seen inferiorly. Vitreous inflammation tends to be mild early in the course of
disease, but the vitreous can later become organized and opacified (26,29).
Peripheral retinal neovascularization may occur, resulting in vitreous
hemorrhage. Cystoid macular edema is the most common cause of permanent
visual loss, and it may occur in 30% to 60% of patients (29,92). Serous,
tractional, rhegmatogenous, and combined retinal detachments occur in 5%
of eyes. Optic disc edema is not uncommon.
The clinical course of intermediate uveitis may range from a mild course
with complete remission to a relentlessly progressive course with severe
exudation, neovascularization, and resistance to therapy, which is seen more
often in children (92,95). Brockhurst and Schepens (91) described four
different groups as defined by their clinical course. A mild course with
complete remission was seen in 31% of patients, a mild chronic course in
49%, a severe chronic course in 15%, and a relentlessly progressive course in
5%. Others (91) found 19% to have mild inflammation, 42% moderate
inflammation, and 39% severe inflammation in their group of patients with
intermediate uveitis who were followed 10 years later.
Diagnosis
The differential diagnosis of intermediate uveitis includes many causes of
vitreous inflammation. It is imperative to exclude infectious causes because
antimicrobial therapy may be curative. Infectious entities in the differential
diagnosis include Lyme disease, toxocariasis, Whipple disease, tuberculosis,
syphilis, human T-cell leukemia virus type 1, Epstein-Barr virus, and cat-
scratch disease. Equally important is the exclusion of intermediate uveitis
associated with underlying systemic disease such as sarcoidosis, Behçet
disease, multiple sclerosis, retinoblastoma, or intraocular
leukemia/lymphoma. Initial laboratory studies include a complete blood
count with differential (myeloproliferative and infectious disease), ACE
(sarcoidosis), chest x-ray film (sarcoidosis and tuberculosis), tuberculin skin
test (purified protein derivative) with anergy panel or QuantiFERON Gold
testing or equivalent, microhemagglutinin Treponema pallidum or fluorescent
treponemal antibody absorption test (syphilis), and ANA (systemic lupus
erythematosus and other connective tissue diseases), with consideration of
Lyme or Toxocara antibody testing (29).
Management
Having excluded treatable infections and noninfectious etiologies, treatment
of intermediate uveitis may begin. Intermediate uveitis is treated medically
and is directed toward the complications and sequelae of intraocular
inflammation such as cystoid macular edema, cataract, vitreous opacity,
peripheral neovascularization, or retinal detachment. A five-step regimen has
been proposed (64) that is a modified approach based on a four-step regimen
initially described in 1984 (44): (a) topical corticosteroid for anterior segment
inflammation with periocular corticosteroid injections; (b) oral NSAIDs
should inflammation recur following the third injection, with topical NSAIDs
in the presence of cystoid macular edema; (c) systemic corticosteroids should
inflammation persist or recur despite the previous interventions; (d)
peripheral retinal cryopexy or indirect laser photocoagulation should pars
planitis recur following the sixth regional steroid injection; and (e) pars plana
vitrectomy versus immunosuppressive chemotherapy should inflammation be
recalcitrant to the preceding modalities. Many antimetabolites and
immunosuppressive drugs have been tried, including cyclosporine,
methotrexate, azathioprine, 6-mercaptopurine, cyclophosphamide, and
chlorambucil (63). With the advent of newer immunosuppressive therapies
that are better tolerated, the use of immunosuppression for treatment is now
stared sooner than later in an attempt to reduce both the topical and systemic
steroid load in order to avoid as many steroid side effects/complications in
these young patients.
Peripheral laser or cryotherapy

In patients with unremitting chronic pars planitis that is recalcitrant to steroid
or NSAID therapy, cryotherapy and scatter laser photocoagulation of the
peripheral retina have been reported effective in causing regression of
peripheral neovascularization and reduction of inflammatory activity (50,51)
and vitreous hemorrhage (51,52). For discussion, see earlier section: “Lasers
—Peripheral Neovascularization (Intermediate Uveitis).”
Vitrectomy
In patients with vitreous opacities impairing vision, causing amblyopia, or
obscuring the ophthalmologist’s view of the periphery, pars plana vitrectomy
is an effective treatment for visual rehabilitation as well as attenuation of
inflammation. Pars plana vitrectomy with or without pars plana lensectomy is
used to treat certain complications of intermediate uveitis, such as vitreous
opacification, tractional retinal detachment, vitreous hemorrhage, and
epiretinal membrane. In cases of vitreous inflammation refractory to medical
therapy, pars plana vitrectomy clears the visual axis by the removal of media
opacities and vitreous debris. Vitrectomy also may alter the course of
inflammation and result in stabilization of the disease by removing
immunocompetent cells and inflammatory mediators from the vitreous cavity
(27,28,45). Vitrectomy may potentially reduce cystoid macular edema either
by eliminating the contact between an inflamed vitreous body and the macula
or by allowing better penetration or distribution of corticosteroids (26,28,54).
General guidelines for vitrectomy for medically refractory intermediate
uveitis include the (a) removal of the posterior hyaloid; (b) removal of as
much anterior vitreous as possible; (c) careful inspection of the periphery,
looking for neovascularization and occult retinal breaks that can be treated
with endolaser photocoagulation; and (d) maximal perioperative control of
inflammation with intravitreal, periocular, and systemic corticosteroid use.
Cataract surgery
Cataract formation is another complication of intermediate uveitis. Cataract
extraction may be achieved through a limbal approach with
phacoemulsification or through a pars plana approach with lensectomy in
combination with a pars plana vitrectomy. If a concurrent vitreous opacity
exists, combined pars plana lensectomy and core vitrectomy is classically the
procedure of choice. A combined limbal cataract extraction and pars plana
vitrectomy approach also can be used.
Implantation of a posterior chamber IOL is dependent on the risk of
future or chronic inflammation. A peripheral iridectomy should be made if
recurrent or chronic inflammation is anticipated. Since the postoperative
course is usually not as problematic, the risk of IOL implantation with
children with intermediate uveitis is not as severe as with children with JIA,
though aggressive care to control inflammation must be taken (96).
In the child with intermediate uveitis, surgery is best undertaken after the
eye has been quiet for at least 3 months. Perioperative inflammation must be
rigorously controlled with aggressive topical, periocular, and possibly
systemic corticosteroid use or steroid-sparing immunosuppression. Maximum
control of inflammation preoperatively, intraoperatively, and postoperatively
allows the greatest chance for a successful surgery.
Juvenile Idiopathic Arthritis

JIA, formally known as juvenile rheumatoid arthritis, is a chronic arthritis of
at least 3 months’ duration that affects children younger than 16 years
(97–99). The estimated prevalence is 113 per 100,000, with an annual
incidence of 14 cases per 100,000 in the United States (5). The course of JIA-
associated uveitis is predominantly chronic, lasting a mean of 19.7 years (92),
with a relapsing and remitting course in 60% of patients and an unremitting
course in 20% of patients. Only 20% of patients have a single episode of
intraocular inflammation (100).
Symptoms and Signs

JIA-associated uveitis is a binocular disease that affects children. The clinical
course is predominantly chronic. The main ocular finding is a chronic
iridocyclitis with associated complications of uveitis, such as band
keratopathy, cataract, secondary glaucoma, amblyopia, and strabismus.
Surgical treatment of refractory glaucoma or cataract may be necessary.
These children are at increased risk for intraoperative complications due to
permanent structural changes from chronic inflammation and are at increased
risk for postoperative complications from an excessive postoperative
inflammatory response and recurrent inflammation.
JIA-associated uveitis is the most common cause of anterior uveitis in
childhood and is associated with significant ocular morbidity (101). The
degree of final visual loss was correlated with the degree of visual loss at
onset, and the risk of ocular complications was correlated with the severity of
inflammation observed at the initial examination as defined by the presence
of posterior synechiae (102). Other factors that correlated with poor visual
prognosis were pauciarticular arthritis (four or fewer joints affected), young
age at onset, female gender, ANA positivity, and rheumatoid factor
negativity. Associated band keratopathy occurs in 6% to 66%, associated
cataract in 18% to 71%, and associated glaucoma in 11% to 38% (22).
Blindness, with a visual acuity worse than 20/200, ranged from 0% to 41%
(22). Chronic or recurrent uveitis eventually caused the complications of
cataract, glaucoma, hypotony, vitreous organization, optic neuropathy, or
macular edema.
Medical Management
JIA-associated uveitis is primarily treated medically. Treatment should be
directed toward absolute control of the inflammation with medications.
Although topical and periocular corticosteroid injections remain the first-line
therapy for anterior uveitis, up to 61% of patients with JIA-associated uveitis
either do not respond to corticosteroid treatment or require prolonged
therapy, with its attendant side effects (102). If inflammation persists with
frequent topical steroid dosing or continues to recur every time
corticosteroids are tapered and withdrawn, the addition of chronic oral
NSAIDs may be useful (103). Refractory inflammation should be treated
with steroid-sparing immunosuppression. In one large study, virtually no
toxicity was found in 127 patients in whom low-dose methotrexate was given
for the management of JIA-associated joint disease (30,104). Other
immunomodulatory agents such as cyclosporine, azathioprine, or
chlorambucil may be considered if continued inflammation with loss of
vision occurs despite these measures (5). Newer modalities include biologic
therapies, such as tocilizumab and adalimumab (105,106).
Surgical Indications
Children with JIA-associated uveitis may require surgical treatment for
refractory glaucoma, cataract, or band keratopathy. These patients are at
increased risk for intraoperative and postoperative complications due to
chronic inflammation. Even if the eye appears clinically quiet preoperatively,
it often responds to surgery with excessive bleeding, postoperative
inflammation, and unexpected postoperative IOP responses such as ocular
hypertension or hypotony. General anesthesia with endotracheal intubation
may be complicated by the presence of cervical arthritis or spondylitis,
temporomandibular joint inflammation with restriction of movement, or
micrognathia. The decision to proceed with surgery for the complications of
JIA-associated uveitis in a child is difficult and must be considered carefully.
Cataract surgery
Cataracts occur in 18% to 71% of patients with JIA-associated uveitis and
can be attributed to chronic inflammation and corticosteroid use (21,22).
Cataract surgery in JIA-associated uveitic eyes is daunting to even the most
accomplished cataract surgeon because of the unknown risks that can occur
intraoperatively and the uncertainty of the postoperative course (3,4). Surgery
is frequently technically challenging because of posterior synechiae, iris
vasculature delicacy, pupillary membranes, miotic pupils, and corneal
opacification (5). The combination of the heightened pediatric immune
response and a uveitic eye can result in violent postoperative inflammation if
proper preoperative care is not taken. Loss of vitreous or retention of cortical
material may exacerbate intraocular inflammation. Indications for cataract
surgery include visual rehabilitation, prevention or treatment of amblyopia,
and enhancement of the ophthalmologist’s view of the posterior segment for
ongoing assessment of the optic nerve and retina. Given the risk of surgery
and the sometimes unpredictable postoperative course, prevention of cataract
in patients with JIA is preferable (1).
The surgical technique for cataract extraction in the child with JIA should
include removal of the cataract, including the posterior capsule, all
retrolenticular and cyclitic membranes, and most of the vitreous. Historically,
pars plana lensectomy with pars plana vitrectomy was the procedure of
choice (26). For the pars plana approach, the lens and posterior and anterior
capsules are cut and aspirated with an ocutome probe. This is followed by a
near-total vitrectomy and removal of the lens capsule, anterior hyaloid, and
vitreous scaffold to reduce secondary membrane formation (21). A surgical
peripheral iridectomy should be performed if recurrent or chronic
inflammation is anticipated.
An alternative approach to cataract extraction may be accomplished by a
combined limbal approach with phacoemulsification and pars plana
vitrectomy. During the limbal approach, synechiolysis, pupillary stretching,
and iris hooks are used as needed for adequate exposure. Capsulorrhexis,
hydrodissection, and phacoemulsification are performed in standard fashion,
and the incision is closed. This is followed by a pars plana approach for
complete posterior capsule excision, followed by an anterior vitrectomy.
Classically, IOL implantation had not been recommended in patients with
JIA-associated uveitis (1,2,21,30). The IOL may provide a stimulus for
intraocular inflammation and structural support for inflammatory membranes,
which may induce irreversible ocular pathology. Thus, patients with JIA
usually have been left aphakic after cataract surgery, necessitating the use of
aphakic spectacles or contact lenses. Despite excellent anatomic outcomes,
visual rehabilitation for aphakic eyes may be suboptimal because many
patients of this age are unable to or refuse to wear aphakic contact lens or
aphakic spectacles (9). Among children, contact lens intolerance is reported
to be 17% to 38% (30,44). The frequent concomitant occurrence of band
keratopathy makes contact lens fitting difficult. Multiple lost contact lenses
carry a substantial expense. Children often are intolerant of aphakic
spectacles because of the weight and aniseikonia.
Cataract surgery outcomes

The results of cataract extraction with or without IOL implantation in patients
with JIA-associated uveitis historically have been suboptimal. In a report of
162 eyes with extracapsular cataract extraction, only 30% of these pediatric
patients had visual acuity of 20/40 or better, and 48% of patients had a visual
acuity of hand motion (HM) or worse (107). With the advent of improved
microsurgical instruments and techniques, seven of eight eyes of adults and
children with JIA-associated uveitis that had standard phacoemulsification
cataract extraction, without a posterior capsulectomy and with posterior
chamber IOL implantation, achieved a final visual acuity better than 20/40
(108). However, all three patients aged 13 years or younger had postoperative
results inferior to those of adult patients at a median follow-up of 14 months.
The children’s eyes developed persistent postoperative inflammation,
posterior synechiae, and pupillary membranes. The authors concluded that in
selected adults with cataracts caused by JIA-associated uveitis, standard
phacoemulsification cataract extraction with IOL implantation may have
excellent results. They cautioned that IOL implantation in children with JIA
merits further investigation.
In another report (96), the results of five eyes of children aged 4 to 8
years who had JIA-associated uveitis after phacoemulsification cataract
extraction, posterior capsulectomy, anterior vitrectomy, and IOL placement
and four eyes of children aged 3 to 8 years after pars plana lensectomy and
vitrectomy without IOL placement were examined. The patients were
followed for 5 years. Visual acuity with IOL was 6/6, 6/60, 6/240, light
perception (LP), and HM, whereas visual acuity with aphakia was 6/9, LP in
two patients, and no light perception (NLP) in one patient. Four of five
patients with IOL implantation required a subsequent surgical retrolental
membranectomy and total vitrectomy. Preoperative topical or systemic
corticosteroids or immunosuppressive treatments were not reported in any
children. After surgery, each child received a retro-orbital injection of
methylprednisolone and an intravenous bolus of hydrocortisone.
Postoperatively, each child received hourly topical corticosteroids for 1 week
that was tapered on an individual basis. Although the outcomes were poor,
the patients were mildly better with IOL implantation.
In 2003, favorable outcomes with IOL implantation with cataract surgery
were reported in children with JIA-associated uveitis (39). Five children (six
eyes) younger than 13 years with JIA-associated uveitis underwent standard
phacoemulsification cataract extraction without posterior capsulectomy and
with posterior chamber IOL implantation. Postoperative visual acuity was
20/40 or better in all six eyes. Median follow-up was 44 months.
Preoperatively, four children (five eyes) were on low-dose methotrexate
immunosuppressive therapy for a median length of 1.25 years before surgery.
Preoperatively, two children (three eyes) were on additional
immunosuppressants or systemic corticosteroids. All eyes received frequent
topical corticosteroids for a median of 2 weeks preoperatively and 8.5 weeks
postoperatively. Five eyes required Nd:YAG capsulotomy. One eye needed a
subsequent surgical posterior capsulectomy and pars plana vitrectomy. The
authors concluded that with adequate long-term preoperative and
postoperative control of intraocular inflammation with systemic
immunosuppressive therapy and intensive topical perioperative corticosteroid
therapy, children with JIA-associated uveitis may have favorable surgical
outcomes after cataract surgery with posterior chamber IOL implantation.
Management of Inflammation
It is critical that cataract surgery with or without IOL implantation be
attempted after absolute and sustained control of uveitis for a minimum of 3
months in patients with JIA. Long-term preoperative systemic
immunosuppressive therapy and aggressive perioperative topical
corticosteroids may vastly improve the outcome of cataract extraction in
children with JIA and may allow for IOL implantation for improved visual
rehabilitation. For aphakic children, vigilant postoperative amblyopia therapy
is critical, and patients should be followed in conjunction with a pediatric
ophthalmologist with proper aphakic contact lens or spectacles, orthoptic
evaluation, and occlusion therapy.
Glaucoma surgery
Elevated IOP frequently accompanies uveitis in patients with JIA. The
incidence of glaucoma in patients with JIA varies between 11% and 38%
(22). The prognosis for patients who develop glaucoma with JIA is poor. One
group (107) reported a 35% incidence of NLP vision in children with uveitis
and glaucoma. Medical therapy often proves inadequate for controlling IOP,
necessitating surgical intervention.
Glaucoma surgery outcomes

Trabeculectomy in children with chronic intraocular inflammation often is
unsuccessful. The increased failure rate of filtration procedures in children
has been postulated to be secondary to decreased scleral rigidity and
accelerated healing of the Tenon capsule in the younger population (2). In
addition, the extensive inflammatory response of the pediatric eye, as well as
possible recurrent or chronic inflammation, results in fibrous tissue growth
and subsequent closure of the filtering bleb over time. Although adjunctive 5-
fluorouracil may keep the filtering bleb open in the initial few weeks or
months after surgery, recurrent inflammation may later close the bleb (2).
Thus, the long-term success rate of filtering surgery has a decreasing chance
of success in these patients over time.
The success rate with glaucoma drainage devices is slightly more
encouraging. Twenty-seven eyes with secondary glaucoma due to JIA had
Molteno implants and were followed for 40 months (19). A successful
outcome with IOP > 5 and ≤23 mm Hg was achieved in 89% of patients. Life
table analysis success rates were 95% after 27 months and 90% after 52
months. Postoperative complications included flat anterior chamber, tube
blockage by iris or vitreous, cataract, cornea–tube touch, choroidal
detachment, corneal edema, and corneal abrasion.
The inflammation should be maximally controlled with corticosteroid or
immunosuppressant medications for as long as possible preoperatively in
patients undergoing nonurgent glaucoma surgery. The glaucoma drainage
device can be implanted in standard fashion between Tenon fascia and sclera
and secured in place. The tube is placed through either the limbus or the pars
plana in vitrectomized eyes. The tube and its insertion site are covered with a
patch of preserved donor sclera. Topical corticosteroids are used
postoperatively. If a glaucoma drainage implant is placed in a vitrectomized,
aphakic, pediatric patient, a pars plana tube placement will better allow
contact lens fitting following surgery (1).
Band keratopathy
Band keratopathy is observed in 6% to 66% of patients with advanced JIA-
associated uveitis (21) and can be removed by EDTA chelation and
superficial keratectomy. For discussion, see “Therapeutic Surgery: Band
Keratopathy” section of this chapter.
Management of inflammation
In the child with JIA-associated uveitis, surgery is best undertaken after the
eye has been quiescent for as long as possible. Good control with systemic
immunosuppressive therapy can help prevent the complications of both
chronic inflammation and chronic corticosteroid use. It also allows an
improved chance of successful surgery with adequate control of
inflammation. Perioperative inflammation must be rigorously controlled with
aggressive topical, periocular, and possibly systemic corticosteroid use.
Maximum control of inflammation preoperatively, intraoperatively, and
postoperatively allows the greatest chance of a successful surgery.
OVERALL
In review, pediatric patients with uveitis require special attention, not only for
treatment but also for surgical intervention. Quiescence is the key, and if
possible, the surgeon should wait for the eye to be inactive for at least 3
months before proceeding with surgery. Proactive and aggressive control of
inflammation, which may require systemic immunosuppression,
preoperatively and perioperatively are essential for good postoperative
outcomes.
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49
Uveitis Masquerade Syndromes
Mikel Mikhail, and Lisa J. Faia
INTRODUCTION
The term masquerade syndrome is typically used in ophthalmology to
describe conditions that include, as part of their presentation, the presence of
intraocular cells but are not related to immune-mediated uveitic entities. The
term was first coined in 1967 to describe a conjunctival carcinoma presenting
as a chronic conjunctivitis (1).
Determining the cause of a masquerade uveitis is a diagnostic challenge,
since a wide variety of conditions present with intraocular cells. These
include immune-mediated processes (whether endogenously or exogenously
triggered), neoplastic conditions, as well as a variety of nonneoplastic
entities. Typically, masquerade syndromes refer to neoplastic entities that
may mimic intraocular inflammation. For the purposes of this chapter, we
have included the most common neoplastic, as well as nonneoplastic
diseases, that may present in the form of anterior or posterior segment
inflammation. Knowledge of these conditions when approaching a child with
a recurrent, chronic, or otherwise atypical uveitis is critical to avoid a delay in
diagnosis and management. Often, such a delay may have a significant
impact on the child’s visual morbidity and potentially, mortality.
PREVALENCE
Although exact numbers regarding the actual prevalence of masquerade
syndromes are difficult to come by in the pediatric literature, these conditions
are thought to constitute <3% of the patient population in subspecialty uveitis
clinics (2–4).
GENETICS AND ENVIRONMENTAL
FACTORS
Most neoplastic masquerade uveitis syndromes in the pediatric population
have an associated genetic component. Understanding of the role of genetics
in these entities is important for establishing a diagnosis, and for predicting
prognosis for visual recovery and/or patient survival.
For example, retinoblastoma is one of the few cancers for which a single
genetic mutation predicts disease development at very high penetrance. In
order for RB to arise, a mutation on both alleles of the RB1 gene is required
(5). A review of the genetics of retinoblastoma is highlighted in (Section III,
Chapter 44). Conversely, ciliary body medulloepithelioma, another neoplastic
entity that can present as a masquerade uveitis, has no identifiable
cytogenetic abnormality and usually presents sporadically (except in the
exceedingly rare case of pleuropulmonary blastoma family tumor and
dysplasia syndrome) (6,7).
Another neoplastic entity that can masquerade as a uveitis in the pediatric
population is leukemia. Acute lymphoblastic leukemia (ALL) is a childhood
disease that occurs as a result of recurrent genetic insults that block precursor
B- and T-cell differentiation and drive the proliferation and survival of
aberrant cells. Approximately 80% of ALL cases involve these chromosomal
abnormalities, which are incorporated into the World Health Organization
classification of acute leukemia (8). These translocations are important for
disease management, because they are also prognostically significant and
guide treatment. For example, the most common genetic lesion occurring in
childhood ALL is a 12;21 translocation (12; 21 (p13.2; q22.1), which is seen
in up to 25 percent of children with the B-lineage disease of ALL (19,22,23).
This translocation predicts a more favorable prognosis. Translocations
involving 11q23.3, although less common, predict a poorer prognosis in both
adults and children (8–11).
While the often cited Philadelphia chromosomal abnormality is common
in adults with ALL, it is only observed in 5% of children (12,13). The T-
lineage ALL are a separate disease entity. T-cell ALL is most commonly seen
in young adult males. 60% of these patients have an abnormal karyotype,
characterized by a pattern of recurrent karyotypic abnormalities involving
both T-cell (TCR) and non-TCR gene loci (14).
Histiocytic disease entities, commonly associated with masquerade
uveitic syndromes, also have a genetic component. For example, somatic
mutations that activate the MAPK signaling pathway are commonly found in
patients with Langerhans cell histiocytosis (LCH) and can help stratify
disease severity (15–18). While juvenile xanthogranuloma (JXG) limited to
the skin does not typically exhibit genetic changes, systemic JXG has been
associated with mutations in several MAPK pathway genes (19,20).
Nonneoplastic diseases that can present as uveitis can also present with
complex gene abnormalities and variable inheritance patterns. For example,
while they rarely masquerade as uveitis, inherited retinal degenerations have
variable genotypic and phenotypic patterns. The examining ophthalmologist
should pay attention to the patient’s family history, as well as associated
systemic features, to be able to identify these masquerade uveitic cases that
may present as retinitis pigmentosa (RP). This is important to confirm the
diagnosis, counsel the patient and his or her family members, and determine
eligibility for clinical trials or approved treatment.
Most masquerade uveitic masquerade syndromes have no identifiable
environmental factors. An exception to this is in cases of vitreous
hemorrhage and intraocular foreign bodies, which are commonly associated
with trauma. An evaluation for nonaccidental trauma in these cases is
important and is thoroughly explored in (Section IX, Chapter 70).
Visual rehabilitation of children presenting with masquerade uveitic
syndromes encompasses the same principles of early intervention and
rehabilitation associated with any cause of vision loss. This is further
explored in Section X. Patients with these disease entities are often diagnosed
late in the disease course, due to the “masquerade” nature of these
pathologies. The treating ophthalmologist should order a thorough diagnostic
workup for these patients, a delay in diagnosis should not impede early
intervention and amblyopia management to avoid permanent functional
impairment.
OTHER HEALTH CARE PROVIDERS
A multidisciplinary approach to the care of patients presenting with
masquerade uveitic syndromes is important to avoid visual and systemic
morbidity. The role of other physicians and other health care providers is
critical in the management of these complex patients. Many of these
masquerade syndromes involve neoplastic entities that are systemic in nature.
If there is clinical suspicion of malignancy, the early involvement of a
specialized oncology team is critical to avoid systemic morbidity or
mortality. In benign conditions, these masquerade uveitic syndromes
commonly present with systemic disease, and the involvement of a
pediatrician and/or subspecialists is important for optimal systemic
management of these patients. These patients can present with vision loss as
their primary complaint, and it is incumbent upon the treating
ophthalmologist to refer to specialty services as appropriate. Genetic
counseling, nursing care, social work, as well as occupational and physical
therapy, ensure adequate support to these patients and their families.
SPECIFIC DISEASE ENTITIES

Neoplastic
The neoplastic disease entities, whether benign or malignant, that commonly
present as a masquerade syndrome in the pediatric population include
retinoblastoma, leukemia, JXG, Langerhans cell histiocytosis, and ciliary
body medulloepithelioma. While common in the adult population, primary
CNS lymphoma, systemic non-Hodgkin lymphoma, lymphoproliferative
disorders, uveal melanoma, and metastatic tumors are rare in the pediatric
population but are nevertheless included in this chapter.
Retinoblastoma
Retinoblastoma is the most common intraocular malignancy in children, with
a reported incidence of 1 in 15,000 (21). While leukocoria and strabismus are
the most frequent presenting signs of retinoblastoma, a sizeable percentage of
cases can present with a masquerade uveitis. In fact, up to 40% of patients
with a final pathologic diagnosis of retinoblastoma were initially
misdiagnosed as having uveitis (22). This “pseudouveitis” is most commonly
due to tumor cells seeding in the anterior chamber (23–26). In some reports,
this has been reported to occur in 0.5% of patients with retinoblastoma, and a
pseudohypopyon has been shown to occur in up to 0.25% of patients (24).
Retinoblastoma can also be confused for vitreous hemorrhage, toxocariasis,
toxoplasmosis, or endophthalmitis (27). Despite the atypical nature of these
presentations, it is critical that the ophthalmologist maintain a high degree of
suspicion for retinoblastoma in a child due to the potential associated
morbidity and mortality.
Retinoblastoma can present in its exophytic, endophytic, or diffuse
infiltrating forms (28). The exophytic pattern occurs when the intraretinal
tumor grows in the direction of the subretinal space. Retinal vessels are
typically seen overlying a subretinal mass. The endophytic pattern occurs
when the tumor grows into the vitreous cavity and invades the inner retinal
layers. These tumor cells can seed the vitreous cavity. Vitreous tumor cells
can masquerade as a vitritis. These cells are typically tumor cells, although
they may rarely be inflammatory in nature, particularly if the tumor is large
and undergoes necrosis. The diffuse infiltrating type is characterized by
diffuse involvement of the retina, resulting in a placoid thickness rather than
a well-circumscribed mass, and it is typically seen in older children (29). The
latter pathology is the most likely type to present as a masquerade uveitis
(Figure 49-1). These have been typically described in unilateral, nonfamilial
cases (30). These lesions also do not typically present with intraocular
calcifications, a hallmark of the disease. Imaging to aid in diagnosis in those
cases may not be particularly helpful, given the fact that intraocular
calcifications are often absent.
FIGURE 49-1 This figure demonstrates a case of an 11-
year-old female who presented with a history of unilateral
leukocoria in the right eye (A). She had an anterior and
posterior uveitis with an ill-circumscribed mass inferiorly
(B). Enucleation revealed a poorly differentiated
retinoblastoma with retrolaminar and choroidal invasion
(C). The patient was treated with systemic chemotherapy
and recovered well.
Ultrasonography or MRI may simply show diffuse retinal thickening. The

clinician has to therefore maintain a particularly high level of suspicion in
order to ascertain the diagnosis. Clues to aid diagnosis include the shifting of
the pseudohypopyon with head movement and the classic white color, as
opposed to the contrasting yellowish hypopyon seen in cases of true
inflammation.
Nevertheless, a diagnostic anterior chamber paracentesis may be required
to establish the diagnosis. This carries the possible risk of tumor seeding
through the needle tract. There is no large-scale, peer-reviewed comparative
study in the literature to address this issue, and most of our knowledge comes
from smaller case series or case reports (31–36). Needle aspiration biopsies
should be reserved for cases where the diagnosis cannot be established
clinically or radiographically and should preferably be performed by an
experienced ocular oncologist. Clear corneal incisions are less likely to result
in tumor seeding, since they are not expected to result in tumor spillage
inside the eye. Cytologic sampling shows hyperchromatic nuclei and scanty
cytoplasm (37).
A detailed discussion of the treatment of retinoblastoma is found in
Chapter 45. Treatment decisions depend on tumor characteristics, such as
laterality, and patient factors, such as hereditary status and overall metastatic
risk. The body of literature is ever-changing, and treatment is often center
dependent. Small tumors detected on examination can be treated with
transpupillary thermotherapy. Larger tumors are often treated by a
combination of carboplatin and transpupillary thermotherapy. For more
advanced lesions, combination chemotherapy (involving two or three drugs),
in addition to local therapy is advocated. Patients with the nonfamilial
bilateral variant are treated with systemic combination therapy and local
therapy. Some teams propose bilateral intra-arterial chemotherapy. The
treatment of unilateral retinoblastoma is also controversial. Depending on the
stage of the tumor and the patient’s risk of metastases, patients can be treated
with enucleation, combination chemothermotherapy, orbital brachytherapy,
conventional external irradiation, or more recently, intra-arterial and/or
intravitreal chemotherapy (38–44). Vitrectomy is not advocated for those
patients due to the risk of systemic dissemination (27), although new
techniques of endoresection in refractory cases may change this paradigm
(45).
Leukemia
The second most common neoplastic entity to present as a masquerade
uveitis is leukemia. Acute leukemia is the most common systemic
malignancy in children, affecting 1 in 30,000 below the age of 14 years. ALL
is the most common type, accounting for 90% of cases. Abnormal
proliferation of bone marrow blast cells displaces the native bone marrow and
results in pancytopenia (anemia, neutropenia, thrombocytopenia).
Chronic leukemia is less common in children compared to adults.
Leukemic cells commonly infiltrate the choroid and the retina, and
children classically present with a leukemic retinopathy or choroidopathy.
These are relatively straightforward to recognize. Oftentimes, however, iris
and anterior segment involvement can be confused with a uveitis.
Leukemic “uveitis” typically presents as a creamy-white
pseudohypopyon. Leukemia can also show a so-called pink hypopyon due to
intermixing of leukemic cells and the presence of an accompanying hyphema.
This layering of leukemic cells typically shifts with a change in head
position. The trabecular meshwork may also be involved, resulting in a
secondary glaucoma. The iris can be thickened and heterochromic due to
diffuse infiltration by leukemic cells, or it can be nodular in appearance due
to cellular clumping.
Eye involvement in children is reported to occur in about 9% of cases
(46). Leukemia can rarely present with anterior chamber inflammation,
composed of malignant leukemic cells. Although rare, refractory uveitis and a
hypopyon have been reported as the first manifestation of ALL (46). More
commonly, however, anterior chamber inflammation is the sole or
accompanying sign of relapse in a child with a history of acute leukemia
(47–50). Relapse of disease is common in leukemia, and anterior segment
relapse accounts for 0.2% to 2% of all cases of leukemic relapse (46,51). A
review of 196 children treated with acute leukemia showed 2 patients with a
bilateral anterior uveitis that was the first sign of relapse (52). The eye is
considered a sanctuary site for leukemic cells, and these cells are routinely
shielded from systemic treatment. Once chemotherapy is stopped, these cells
can often proliferate even in the absence of clinically apparent systemic
disease (52). Anterior chamber relapse has been reported to occur in the first
few months after finishing maintenance chemotherapy in these children. It is
also important to note that even in the absence of hematologic evidence of
relapse, anterior segment involvement should suffice to warrant further
investigation. Out of 11 cases of ocular relapse reported by Bunin et al., only
one child had concomitant hematologic relapse (53,54).
As with lymphoma, it is imperative to diagnose these lesions since
ocular, including anterior segment, involvement entails central nervous
system involvement. Anterior chamber paracentesis is diagnostic, and blast
cells are usually seen. Early diagnosis and therapy gives the child the best
chance at an ultimate cure. Local radiotherapy and/or intraocular injection
therapy is commonly required to treat ocular involvement with leukemia.
There should also be a diligent search for extraocular spread of leukemic cells
in the event a diagnostic tap demonstrates leukemic involvement of the eye.
Collaboration between the ophthalmologist and oncologist is critical in those
cases.
Juvenile Xanthogranuloma
JXG is a benign, non–Langerhans cell systemic histiocytosis that typically
affects children in the first 2 years of life (55). The disease characteristically
involves the skin, resulting in raised, reddish-yellow papules that are most
commonly found on the face. The most common extracutaneous site of
involvement is the eye, although other organs, such as the testes, spleen, lung,
and bone, can also be involved. Ocular complications due to JXG are rare,
occurring in around 0.5% of patients. They mostly occur in the 1st year of
life (56). The most common presenting ocular symptom is eye redness (57).
In the eye, the most common site of involvement is the iris. JXG can present
as an iris mass or a nodule (Figure 49-2). These iris nodules tend to bleed,
resulting in a spontaneous hyphema (57). This can result in glaucoma and
heterochromia (58–60). Choroidal infiltration can also occur and result in a
retinal detachment and a hemorrhagic infarction of the retina (61). JXG can
also rarely involve the eyelid (62).
FIGURE 49-2 This figure demonstrates a case of a 16-
year-old female who presented with an iris mass of the
right eye (A). This was initially thought to be a uveitis,
later believed to be a melanoma, and finally diagnosed as
juvenile xanthogranuloma. There was no hyphema. She
was treated with intensive topical prednisolone drops
every hour for 3 weeks, with subsequent resolution of
uveitis. The left eye was normal (B). An anterior segment
optical coherence tomography image (AS-OCT) of the
right eye clearly demonstrates the deposition of debris on
the anterior iris surface (C). (Courtesy of Carol Shields,
MD.)
Classically, the most common eye masquerade syndrome associated with

JXG is iris infiltration and anterior chamber inflammation (61,63–67).
JXG emphasizes the importance of a thorough systemic evaluation in any
case of pediatric uveitis. The ophthalmologist should thoroughly examine the
skin to check for the characteristic reddish-yellow papules. Cutaneous lesions
have been reported to occur in up to 41% of cases of ocular JXG (1). In other
reports, ocular JXG has been reported to occur more commonly in isolation.
For example, in a series of 31 eyes, cutaneous JXG was only present in three
cases (57). Whereas the presence of skin lesions is helpful, their absence does
not exclude the diagnosis, as the skin lesions may follow the ocular
diagnosis, by as much as 8 to 10 months (56).
When biopsied, these lesions demonstrate large histiocytes with a
characteristic foamy cytoplasm and Touton giant cells. These are typified by
a peripheral wreath of nuclei arranged around an eosinophilic cytoplasm (67).
Interestingly, iris lesions tend to present with fewer Touton giant cells than
skin lesions, which may complicate the histologic diagnosis (55,67,68).
Ocular JXG lesions can be diagnosed by skin biopsy if the characteristic skin
lesions are present, sparing the eye any diagnostic intervention. If the skin
papules are absent, an anterior chamber paracentesis can be performed to
prove the diagnosis by demonstrating the classic foamy histiocytes (69). An
iris biopsy may also be performed if the paracentesis is noncontributory or
equivocal, although this should be considered last resort and reserved for
cases where the iris lesion occupies <25% of the surface area of the iris
(63,67).
Whereas cutaneous JXG can typically be observed, ocular involvement
requires treatment, unless the disease is limited to the epibulbar or eyelid
tissue (70). There are few established protocols for the management of JXG.
Simple iris JXG lesions are typically treated with topical, periocular, or
systemic steroids to reduce their size and prevent ocular complications, such
as glaucoma, hyphema, and amblyopia (71). Topical and systemic ocular
hypotensive agents, including systemic acetazolamide, may be required to
control the intraocular pressure if elevated. Some cases are refractory to
corticosteroids and may require irradiation or immunosuppressive
medications (70,72). Chemotherapy may be used to shrink these lesions,
followed by low-dose radiotherapy, typically 300 to 400 cGy (73). An
excisional biopsy may also be considered if the lesion does not respond to the
aforementioned treatments, although this should generally be avoided, if
possible, given the associated visual morbidity.
Ciliary Body Medulloepithelioma

Whereas melanomas of the iris or ciliary body are rare, ciliary body
medulloepithelioma (also known as diktyoma), is the most common ciliary
body neoplasm in children. These tumors occur in the first decade of life.
They are unilateral, amelanotic, and solitary and arise from the ciliary body
(Figure 49-3).
month-old male who presented with a ciliary body mass
with iris invasion (A, B). Ultrasound biomicroscopy
demonstrated a ciliary body mass with angle and iris
invasion, with characteristic intralesional cysts (C). This
was histologically confirmed as ciliary body
medulloepithelioma. The eye was enucleated. (Courtesy of
Carol Shields, MD.)
The clinical findings of medulloepithelioma are not always readily apparent.

Because the tumor is located in the ciliary body, it can often compress
adjacent intraocular structures prior to its presentation as a visibly evident
ciliary body mass. This compression can result in secondary glaucoma,
hyphema, vitreous hemorrhage, rubeosis, cataract, ectopia lentis, and uveitis
(74–77). A chronic granulomatous inflammatory response can occur in these
cases (76). Also, there may be an underlying history of blunt trauma that may
obscure the initial diagnosis (77). In fact, up to 20% of cases of
medulloepithelioma are misdiagnosed, and patients are surgically treated for
other presumed conditions. Patients often undergo lensectomy for a presumed
idiopathic or traumatic cataract, or a glaucoma drainage procedure for ocular
hypertension, resulting in a delay in diagnosis of up to 1 year (78)
Extraocular extension results in clinically apparent orbital spread, which is a
poor prognostic indicator.
A definitive diagnosis can only be established histopathologically,
although a preoperative ultrasound is helpful. Ultrasonography shows an
irregularly shaped lesion with internal cysts on B-scan and is highly reflective
on A-scan (79,80).
A delay in the diagnosis of a ciliary body medulloepithelioma due to a
masquerade can result in significant morbidity and even mortality to the
patient. A high degree of suspicion should be maintained for a chronic uveitis
unresponsive to typical uveitic therapies, and the clinician should have a low
threshold for imaging those patients (76). Surgical removal provides a
definitive histopathologic diagnosis, and removal of the tumor is often
curative. The decision to enucleate is based on a large tumor size, a painful
eye, or a strong suspicion for malignancy on the basis of rapid growth (77).
These tumors rarely metastasize but can spread locally. They have a high
recurrence rate within the eye if locally resected.
Langerhans Cell Histiocytosis

LCH is a proliferative disease of Langerhans cells, which are histiocytic
dendritic cells, derived from the bone marrow. The disorder is more likely to
occur in the pediatric population and is rare, occurring at a rate of 5 per
million. LCH can occur in any organ system or can manifest as a multisystem
disorder (81). It most commonly occurs in the bone (82). The systemic
features of LCH include patient irritability, fever, weight loss, a lytic bone
lesion, and diabetes insipidus as well as other nonspecific findings (83).
Eye involvement in LCH is rare, and most commonly involves the orbit
in about 25% of cases (82). There are some case reports, however, outlining
intraocular involvement. Aggressive systemic LCH with ocular involvement
can be a devastating disease, with significant ocular morbidity (84).
Choroidal infiltration, vitritis, and iris nodules can occur in cases of LCH
(84–87). These children can present with neovascular glaucoma, macular
involvement, and ensuing amblyopia (83) (Figure 49-4).
year-old male with known Langerhans cell histiocytosis.
He presented with pain and a red eye (A). Clinical
examination demonstrated an iris mass, uveitis, and a
choroidal mass (A, B). Fine needle aspiration biopsy of the
iris mass confirmed Langerhans cell histiocytosis. He was
treated with low-dose plaque radiotherapy, with interval
decrease in the size of the choroidal mass (C). He
recovered well and maintained vision in that eye. (From
Shields CL, et al. Langerhans cell histiocytosis of the uvea
with neovascular glaucoma: diagnosis by fine-needle
aspiration biopsy and management with intraocular
bevacizumab and brachytherapy. J AAPOS
2010;14(6):534–537. Courtesy of Carol Shields, MD.)
Due to the associated visual morbidity, although rare, LCH should be

included in the differential diagnosis of a child with an anterior uveitis and/or
iris or choroidal infiltrative disease. These cases are typically well controlled
with local or systemic corticosteroids. Shields et al. reported the successful
use of brachytherapy as well as intravitreal bevacizumab in a case of LCH
presenting with secondary neovascular glaucoma (83).
Others have reported success with low-dose external radiotherapy or low-
dose, fractionated whole eye external beam radiation (88,89). Since LCH is
commonly associated with multiorgan dysfunction, systemic methotrexate or
other chemotherapeutic agents are commonly used to control the systemic
disease. Stem cell transplantation as well as newer targeted therapies are also
being increasingly used (81).
LYMPHOMA AND POSTTRANSPLANT

LYMPHOPROLIFERATIVE DISORDER
Unlike its adult counterpart, primary CNS lymphoma is exceedingly rare in
the pediatric population, with only 15 to 20 reported cases in North America
on an annual basis (90). Whereas the majority of adult patients presenting
with a masquerade uveitis were later found to have primary CNS lymphoma,
this kind of presentation is exceedingly rare in children (2).
Conversely, systemic non-Hodgkin lymphoma, presenting with uveitis,
can occur in children. Nevertheless, more than half of all patients are above
the age of 60, and the disease entity is also rare in those under 40. Uveal
lymphoma may present with both anterior and posterior segment
inflammation. It can also present with iris nodules that resemble those
commonly seen in ocular sarcoidosis (1). This entity typically occurs in
primary systemic lymphoma; an increasing body of literature has reported
uveitis in the context of patients suffering from a lymphoproliferative disease
after organ transplantation. The underlying cause of this condition, termed
posttransplant lymphoproliferative disease (91), is believed to be related to
Epstein-Barr virus infection, possibly resulting in B-cell proliferation, in the
absence of the regulatory role of T cells due to systemic immunosuppression
(37). These patients typically improve with a decrease in their systemic
immunosuppression.
If there is any clinical suspicion, vitreous biopsy should be performed to
provide a definitive diagnosis and ensure early treatment. Communication
between the retina surgeon and the pathologist is pivotal to ensure an
adequate sample has been obtained for diagnosis. Ideally, during vitrectomy,
a 1-mL undiluted sample should be obtained prior to starting the infusion.
The aspirate should be fixed in 95% alcohol immediately after aspiration,
rather than balanced salt solution, to avoid cellular degradation (37).
Uveal Melanoma and Metastatic Tumors

Uveal melanoma is generally a disease of the elderly, with a median age at
diagnosis of 62 years (92–94). Uveal melanoma, however, can occur at any
age and can even present congenitally (95–98). Younger patients with uveal
melanoma typically have a better prognosis, independent of the size of the
tumor.
Approximately 5% to 10% of adults with uveal melanoma present with a
uveitis. The prevalence of uveitis in pediatric cases of uveal melanoma is less
clear. However, we do know that children have a higher incidence of iris
melanoma compared to adults (94,99–101). This potentially makes them
more likely to present with some cellular reaction in the anterior chamber.
Diffuse, plaque-like melanomas, in particular, may present as anterior or
posterior uveitis, or as posterior scleritis and serous retinal detachment. For
example, Martin described a case of diffuse iris melanoma in an 11-year-old
child who presented with a clinical picture of uveitic glaucoma and iris
nodules (102). After not responding to a glaucoma filtration procedure and
systemic steroid therapy, the patient showed epibulbar extension of the
tumor. The eye was enucleated and found to harbor a diffuse iris melanoma
that metastasized and led to the patient’s death. Harley et al. described a case
of a malignant transformation of multifocal tapioca iris nevus in a 31-month-
old boy, which was successfully treated with plaque radiation (103) (Figure
49-5).
FIGURE 49-5 This figure demonstrates a case of a case of
a 31-month-old who presented with a multifocal iris
nevus. Slit lamp photograph shows multifocal nevus, with
the largest lesion at 3 o’clock (A). Four years later, the
lesion had grown in size, and there were new numerous
satellite lesions (B). This was consistent with malignant
transformation to melanoma, which was confirmed on fine
needle aspiration biopsy. The patient was treated with
plaque radiotherapy. Four years after treatment, slit lamp
photography showed tumor regression (C). Ultrasound
biomicroscopy showed the iris nodule pre- (D) and
posttreatment (E), demonstrating tumor regression.
(Courtesy of Carol Shields, MD.)
Notably, uveal melanoma can masquerade as other more common pediatric

tumors, such as JXG, and should be on the differential diagnosis of any iris or
ciliary body mass lesion, even if it is amelanotic. Iris angiography can prove
useful in differentiating melanoma from JXG. While JXG lesions of the iris
typically present with diffuse hyperfluorescence, focal dye leakage is more
indicative of an iris melanoma or a metastatic lesion (104).
Metastatic disease to the uvea from an extraocular primary site is
exceedingly rare in the pediatric population. Kaliliki et al. presented a case of
a metastatic ciliochoroidal lesion from a primary sarcoma of the lower
extremity (105) (Figure 49-6). Given the rarity of this disease entity in
children, it is unclear how many present with a concomitant uveitis. While
they are unlikely to be encountered by an ophthalmologist in his or her
career, it is important to keep melanoma and metastatic tumors in mind when
examining a child with an iris or ciliary body mass with or without uveitis.
year-old female who presented with painless vision loss of
the left eye from a metastatic ciliochoroidal lesion from a
primary sarcoma of her lower extremity. The left eye
demonstrates loss of the normal red reflex. The right eye
was normal (A). Examination revealed a solitary large
ciliochoroidal amelanotic mass in the nasal quadrant.
Magnetic resonance imaging of the orbit demonstrates a
characteristic high signal T-1 mass lesion nasally (B). C:
The gross globe displayed an ovoid amelanotic
hemorrhagic ciliochoroidal mass measuring 20 × 20 × 18
mm. (From Kaliki S, et al. Ciliochoroidal metastasis as the
initial manifestation of an occult soft-tissue extraosseous
sarcoma in a 10-year-old girl. J AAPOS
2013;17(2):217–220. Courtesy of Carol Shields, MD.)
Nonneoplastic Entities
While the term masquerade syndrome typically refers to neoplastic entities
that masquerade as benign processes, a number of nonneoplastic conditions
can present as a uveitis in a child.
These include vitreous hemorrhage, retinal detachment, and intraocular
foreign bodies. Retinal degenerations, such as RP, iris stromal cysts, and
idiopathic nonspecific orbital inflammatory syndrome (NSOI) can also
present with intraocular inflammation.
Vitreous Hemorrhage
An acute vitreous hemorrhage can be readily identified by the clinician. One
to three weeks after the onset of a vitreous hemorrhage, however, red blood
cells become smaller and tan colored.
These so-called ghost cells may resemble white blood cells. As a result,
an old vitreous hemorrhage can masquerade as a uveitis. These cells can layer
down in the anterior chamber, resulting in a pseudohypopyon. They can also
block the trabecular meshwork, and result in increased intraocular pressure,
an entity commonly described as “ghost cell glaucoma.”
A high degree of suspicion should be exercised by the clinician when
examining a child with vitreous hemorrhage. Ruling out nonaccidental
trauma in the context of unexplained vitreous hemorrhage is critical. In
addition, surgical intervention may be necessary due to the amblyogenic
nature of a vitreous hemorrhage, although a clear time frame as to when to
operate has not been established and is beyond the scope of this chapter (see
Chapter 11 on Amblyopia). Most importantly, ruling out a retinal detachment
or tear is paramount. If the vitreous hemorrhage does not permit a clear view
to the posterior segment and an ultrasound cannot be performed or is
equivocal, early clearing of the vitreous hemorrhage should be encouraged
(Video 49-1 ).
Retinal Detachment
Retinal detachment can present as a masquerade uveitis, particularly in the
pediatric population. Children, particularly preverbal children, are unable to
vocalize loss of vision, or they may not recognize a change in vision.
Vitreous hemorrhage, pigmented cells (referred to as tobacco dusting), and
inflammation can occur in eyes with a rhegmatogenous retinal detachment.
Liberated outer segments and inflammatory photoreceptor cells can
migrate anteriorly and block the trabecular meshwork, resulting in elevated
intraocular pressure, commonly referred to as the Schwartz-Matsuo
phenomenon (106–108). These photoreceptor cells can mimic white blood
cells and can be confused for an anterior uveitis.
Intraocular Foreign Body

Children with occult intraocular foreign bodies are particularly prone to
presenting with a masquerade uveitis. The clinical history is often not
contributory, either because the child does not recall or verbalize the time or
mechanism or injury or for fear of punishment from his or her parents or
guardians. Early recognition is critical to allow for timely intervention and
provides the best prognosis for visual recovery. Removal of the intraocular
foreign body within 24 hours of injury significantly reduces the risk of
infectious endophthalmitis (109–112). A noninfectious endophthalmitis may
also occur in cases of retained metal fragments (113). This has been reported
in the context of copper foreign bodies, plant seeds, or even animal hairs
(113,114).
These cases of noninfectious endophthalmitis should be managed in the
same manner as infectious cases, since it is often impossible to determine if
there is an underlying infection on clinical examination alone.
Retinal Degenerations
Retinal degenerations are typically characterized by a progressive loss of
photoreceptors, nyctalopia, visual field loss, and abnormal electrophysiology.
Most cases are diagnosed in childhood or adolescence. They are a
heterogenous group of disorders and include entities such as RP, Bardet-
Biedl syndrome, Leber congenital amaurosis, Usher syndrome, Batten
disease, and congenital stationary night blindness (CSNB) (see Chapters 24,
26, 27, 29, 32 and 33).
The presence of vitritis in RP in adults is well documented (115,116). As
many as 37% of adult patients with RP have vitreous cells, with a more
significant inflammatory reaction observed in younger (<30 years old)
compared to older RP patients (115). Similarly, Stunkel et al. reported a 38%
incidence of vitritis in pediatric RP patients, with some disease entities such
as Baret-Biedl syndrome, CLN3 Batten Disease, and Leber congenital
amaurosis, reporting a greater number of cells and a higher incidence of
vitritis compared to others such as Stargardt disease or CNSB, in which
vitreous cells are rare (117). Interestingly, it appears that vitritis may be a
predictor of progression and a marker of disease severity. The presence of
vitreous cells may be more apparent in diseases that are typically rapidly
progressive such as Bardet-Biedl, compared to others that are traditionally
stationary, such as CSNB (118). These cells may be truly inflammatory in
nature. Sampling of the aqueous and vitreous fluid in RP eyes has shown an
increased level of proinflammatory cytokines (116). Alternatively, they may
simply represent shed photoreceptor cells, with greater cellular density
implying more progressive retinopathy and loss of neural elements.
Typically, these retinal degenerations present with signs other than
uveitis. These include a posterior subcapsular cataract, optic disk pallor
macular edema, vasculitis, epiretinal membrane, geographic RPE atrophy,
and peripheral pigmentary changes. While these disease entities commonly
overlap, they often have distinct phenotypes that are beyond the scope of this
chapter. A genotypic assessment in those patients is necessary to establish a
diagnosis, guide treatment, and evaluate the patient’s visual prognosis.
Iris Stromal Cyst

Iris stromal cysts are benign translucent masses that typically arise on the
surface of the iris as a result of aberrant implantation of corneal or
conjunctival epithelium inside the eye. They tend to enlarge, occupy the
anterior chamber, and lead to angle occlusion and glaucoma. They may also
leak, resulting in an anterior uveitis.
Whereas most iris cysts are acquired in the adult population, they tend to
be congenital in individuals younger than 20 years (119). Children with
primary iris stromal cysts typically have a more aggressive course than do
adults, as these lesions can obstruct the visual axis, and result in amblyopia
(120,121). They can also cause a secondary glaucoma, corneal edema, and
uveitis, and may require treatment. Current options include aspiration,
cryotherapy, sclerotherapy, or surgical resection (119). It is important to
completely eradicate these cysts to prevent reepithelization and recurrence.
They may require multiple treatments to allow for globe and vision
preservation.
Anterior chamber inflammation can occur due to primary cyst leakage or
secondary to treatment. If the cyst ruptures, severe uveitis, hypopyon, fibrin
formation, and glaucoma can ensue (119,122). In those cases, topical or sub-
Tenon's corticosteroids and/or an anterior chamber washout may be helpful to
control the inflammatory response.
Idiopathic Nonspecific Orbital Inflammatory Syndrome

Idiopathic NSOI (also known as orbital pseudotumor) is an inflammatory
condition of the orbits that often simulates a neoplasm. While it is more
commonly seen in adults, it can also present in the first and second decades
of life. Its presentation in children is different from that in adults. Unlike in
adults, uveitic features are more prominent in the pediatric form of the
disease, and it is reported to occur in up to 35% of patients (123). Most often,
it presents bilaterally and occurs with varied and nonspecific symptoms.
Common early symptoms include eyelid edema, photophobia, proptosis,
pain, and ptosis. The photophobia encountered in idiopathic orbital
inflammation may occur with or without uveitis. While uveitis typically
occurs in the context of proptosis, making the diagnosis relatively
straightforward, uveitis may be the first presentation of the disease, and
patients can often present without orbital signs (123–125). In children with
NSOI, the anterior uveitis tends to be more resistant to treatment and have an
increased incidence of relapse, while the orbital disease tends to be worse
(123).
There is seldom a systemic association with the ocular disease. However,
in a series of 29 pediatric cases, NSOI was associated with peripheral
eosinophilia, raised erythrocyte sedimentation rate (ESR), and a positive
antinuclear antigen (ANA) (126). Whereas a positive ANA may be a sign of
autoimmune dysfunction, the other hematologic findings are likely simply to
be related to the inflammatory orbital response. Whereas there is some
association with thyroid disease in adults, this does not appear to be the case
in children. Nevertheless, there is one case of hypothyroidism associated with
bilateral nonspecific orbital inflammation, posterior scleritis, and anterior
uveitis in a 13-year-old child (127).
While we do not advocate routine imaging for any patient with an
anterior uveitis, if the uveitis is recurrent, is resistant to treatment, or is
associated with orbital signs, diagnostic imaging of the orbit
(ultrasonography, CT, or MRI of the orbits) should be performed to rule out
orbital inflammation as a cause of the disease. Treatment is typically
systemic, with oral steroids as the mainstay, followed by a tapering of the
steroids.
SUMMARY
A number of masquerade entities can present as uveitis in a child. These
conditions include both neoplastic and nonneoplastic diseases, as well as
benign and malignant entities. It is important for the clinician to maintain a
high index of suspicion when examining a child with any presentation of
suspected intraocular inflammation. The ophthalmologist should perform a
complete ocular and systemic history and examination to elicit any symptoms
or signs that may highlight an underlying masquerade syndrome. Masquerade
syndromes should always be considered as part of the differential diagnosis
in any case of pediatric uveitis. This is particularly important in cases of
recurrent, chronic, or atypical uveitis.
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SECTION VIII
50
Worldwide Causes of Childhood
Blindness
Clare E. Gilbert, and Nathalie Lepvrier-Chomette
OUTLINE
This chapter describes what is currently known about the magnitude,
distribution, and causes of blindness in children worldwide and briefly
describes some of the major blinding diseases, concentrating on those that are
avoidable (i.e., preventable or treatable) and emphasizing control at the
population level rather than management of the individual child. More
detailed descriptions of retinal conditions, such as retinopathy of prematurity
(ROP) (see Chapter 52) and toxoplasmosis (see Chapter 47), are available.
INTRODUCTION
Definitions
The following definition of blindness in children are used throughout this
chapter: an individual aged 0 to 15 years (UNICEF definition of childhood)
who has a visual acuity of <6/60 in the better eye or a central visual field of
<10 degrees (the World Health Organization [WHO] categories of severe
visual impairment and blindness) (1). Although this definition can be readily
applied to adults, it is more difficult to measure visual acuity and assess
visual fields in young children, particularly in those with additional
disabilities. This poses particular challenges not only for clinicians but also
for ophthalmic epidemiologists, who may be concerned with identifying
blind children in the community to determine prevalence or who want to
undertake clinical trials or undertake longitudinal studies. This definition also
fails to capture the impact of visual loss on a child’s functional abilities,
quality of life, or sense of well-being.
EPIDEMIOLOGY OF BLINDNESS IN
CHILDREN
Prevalence and Magnitude of Blindness in Children
Population-based, cross-sectional surveys are the gold standard method of
estimating the prevalence of a condition in the population. However,
blindness prevalence surveys of children are difficult to conduct for a variety
of reasons. First, blindness in children is relatively rare, and very large
sample sizes, that is, approximately 30,000 children, are required to give
precise estimates of the prevalence. As there are many different causes, even
larger samples are required to give representative data, as some causes cluster
within families (e.g., congenital cataract and retinal dystrophies). Children of
school age are unlikely to be present at the time the survey team visits the
community, and this leads to an unrepresentative, biased sample, which can
overestimate the prevalence, because visually impaired children are less
likely to be at school. Indeed, in most developing countries, it is estimated
that only 10% of blind children are receiving any form of education. Higher
levels of clinical and diagnostic skills are also needed by team members to
perform surveys on children.
There are also challenges with other sources of data. For example, some
industrialized countries keep registers of the blind and if children are
included would provide population-based data. But to provide reliable
information, the database needs to be obligatory and constantly updated to
remove children who have had sight-restoring treatment or who have died or
who are no longer children. Other forms of registers, such as birth defects
registers, cannot provide blindness data as they do not use a functional
definition. Lastly, information collected from health care facilities, social
welfare organizations, or special education programs are also likely to be
biased as not all the blind children in the relevant population are included.
For example, in some countries, special schools cannot manage children with
other impairments; therefore, blind children with other impairments may not
be represented.
In developing countries, even these potential sources of data are not
available. Other approaches that have been successfully adopted include
training individuals who know their communities well and who then act as
key informants to identify the blind children in their communities. The
children thought to be blind are then examined by a team to confirm the
impairment and determine the cause of blindness. This approach, which was
developed in Bangladesh (2), has been used in a number of low-income
countries (3–7). Advantages of the “key informant method” are that a large
population can be covered relatively quickly, and awareness is created.
Questionnaires to parents and caregivers have also been assessed, and while
this tool has good sensitivity and specificity for identifying children with
physical and cognitive impairments, questionnaires do not reliably identify
children with sensory impairments (8). What is currently recommended is
that a variety of different methods and sources of data be used in the same
population, as has been undertaken in Mongolia (9), Bangladesh (10), Fiji
(11), Vietnam (12), Tanzania (13), and Indonesia (14).
Between the years 2000 and 2018, a literature review identified 22
studies that provided data on the prevalence of blindness in children in 17
countries (5–7,9,11,13–29). The data, together with earlier studies, suggest
that the prevalence of blindness in children is associated with under-five
mortality rates (U5MRs) expressed as deaths per 1,000 live births ranging
from a prevalence of approximately 0.3/1,000 children in countries with low
U5MRs (<10/1,000 live births) to 1.2/1,000 in the poorest countries of the
world in which U5MRs exceed 200/1,000 live births (30,31). Rates are
higher in poor countries because (a) there are potentially blinding conditions
in poor communities, such as vitamin A deficiency, malaria, and the use of
harmful traditional eye medicines, which do not occur in affluent
populations; (b) there is inadequate control of preventable causes of
blindness, including measles infection, ophthalmia neonatorum, and
congenital rubella; and (c) there is inadequate management of conditions
requiring optical or surgical treatment because children are not identified or
referred, parents cannot afford the travel and care, or there are inadequately
trained and equipped eye facilities to evaluate high refractive errors, cataract,
glaucoma, and ROP. Outcomes can also be compromised because children
present very late.
In 1992, there were estimated to be 1.5 million blind children, three-
fourths of whom lived in low-income countries (32). In 2010, the data were
revised to 1.26 million (33) using estimates of the prevalence of blindness in
each country based on their U5MR. For example, country x with a U5MR of
8/1,000 live births was assigned a prevalence of 0.3/1,000, whereas country z
with a U5MR of 112/1,000 was assigned a prevalence of 0.8/1,000. Over the
last few decades, U5MRs have continued to fall in most middle- and low-
income countries apart from the poorest countries in Africa where the decline
is generally slower (34). In 2018, the calculation was repeated, giving a
revised figure of 1.025 million blind children (Table 50-1). This represents a
32% decline from 1992. The group of countries classified as high income by
the World Bank (35) has the lowest number (62,380, 6% of the total), and the
lower middle-income region has the highest (547,640, 53% of the total).
Lower- and middle-income countries have a relatively higher proportion than
the other two regions, and this reflects the higher prevalence in each country
and the larger child population. The following countries have the highest
estimated number of blind children in descending order: India, Nigeria,
China, the Democratic Republic of the Congo, Pakistan, Ethiopia, Indonesia,
and Bangladesh.
TABLE 50-1 Estimate of the number of blind

children in 2018, by World Bank economic region
The main factors leading to the decline in the estimate are improving
socioeconomic development, greater coverage of public health programs for
control of conditions leading to corneal scarring, and, in many countries,
greater access to tertiary-level eye care services, including for ROP.
Given the variation in prevalence as well as differences in population
structure, the number of blind children/million total population ranges from
approximately 60 in high-income settings to 600 in the poorest countries
(36).
Incidence of Blindness in Children

The rate at which new cases of blindness develop in the child population over
time can be estimated from blind registers using new cases as the numerator
and through active surveillance systems, such as that employed by the British
Ophthalmological Surveillance Unit (37). Active surveillance can provide
accurate incidence data, particularly if more than one source of ascertainment
is used, which allows capture–recapture analysis with adjustment of
estimates.
Estimates of the incidence of childhood visual impairment are available
for only a few countries. Using pooled data from the Scandinavian visual
impairment registers, the overall annual incidence of all levels of visual
impairment was reported to be 0.8/10,000 individuals <19 years in 1993. A
more recent national UK study reported annual age group–specific incidence,
which was highest in the first year of life at 4.0/10,000, with the cumulative
incidence (lifetime risk) increasing to 5.3/10,000 by 5 years and to 5.9/10,000
by 16 years of age (38). Given the paucity of incidence data, global estimates
of the number of children becoming blind each year are not possible. Most
studies from low-income settings suggest that the vast majority of blind
children are either blind from birth or become blind before their fifth birthday
(10,39). This is the age group that should, therefore, be targeted for
interventions and services.
Mortality in Blind Children

Many of the blinding eye diseases of childhood are associated with high child
mortality from chromosomal abnormalities, congenital rubella, prematurity,
vitamin A deficiency, measles infection, malaria, meningitis, tumors, or
metabolic diseases. In the 1980s and 1990s before global programs for the
control of vitamin A deficiency were established (see sections below), it was
estimated that 50% to 60% of blind children died within a few years of
becoming blind largely due to the very high mortality rate in children with
keratomalacia due to acute vitamin A deficiency (40). Even in industrialized
countries, blind children have a high mortality rate, recently reported in the
United Kingdom as 10% in the year following the diagnosis of severe visual
impairment or blindness (38). Similar findings have also been reported in
other studies (41,42). No data are available from other regions, but mortality
is likely to be higher particularly among children with other impairments
such as cerebral palsy (CP) (43). This means that prevalence data markedly
underestimate the real magnitude of blindness in children, as prevalence data
can only capture those children who survive.
CAUSES OF BLINDNESS IN CHILDREN

Classification
In 1993, the WHO developed a system for classifying the causes of blindness
in children, which takes account of the underlying cause and the major site
affected in the visual system (i.e., whole globe, cornea, lens, uvea, retina,
optic nerve, glaucoma, and others, which include lesions of the higher visual
pathways) (44). The classification by underlying cause uses the time of onset
of the insult leading to blindness including factors operating at conception
(e.g., chromosomal abnormalities and genetic diseases), factors acting during
the intrauterine period (e.g., teratogens including infections, drugs, alcohol),
factors operating during the perinatal period (e.g., prematurity, ophthalmia
neonatorum), conditions acquired during childhood (e.g., measles, vitamin A
deficiency, harmful traditional eye remedies, malaria), and conditions where
the time of onset cannot be reliably determined. The last group includes
conditions such as microphthalmos, which may have several different causes,
and cataract, in which the underlying cause is often not known. The
classification system can be modified to group conditions into those that are
definitely prenatal and those that occurred after birth. Both elements of the
classification systems are useful particularly in low-income settings, where
there is often limited information on the family history or past medical
history and where facilities for investigation and definitive diagnoses are not
readily available. This classification is currently being updated by the WHO.
Causes of Blindness in Children

Data on the causes of blindness were reported on almost 27,000 blind
children between 2000 and 2018, and most studies used the WHO
classification. Most studies undertaken outside high-income settings entailed
the examination of children in special education, and so the data are subject
to inherent biases as not all blind children are enrolled. However, in several
settings, the causes of blindness in children in special education have been
compared to causes in children identified in the community, and the findings
are broadly comparable (10). However, children with additional disabilities
tend to be underrepresented in special education, likely leading to an
underestimate of visual loss due to lesions of the optic nerve and higher
visual pathways.
Data on the causes of blindness in children by anatomical site have been
analyzed using the World Bank’s economic grouping of countries. This
analysis has been done as the socioeconomic status of a country largely
determines the availability and access to public health interventions, such as
measles immunization and the availability of specialist services. The findings
show that there is considerable variation in the causes by region (Table 50-2).
For example, in high-income countries, lesions of the optic nerve and higher
visual pathways are responsible for almost 40% of blindness and often are
attributed to complications of preterm birth (38). In the poorest countries,
corneal disease, mainly corneal scarring, still accounts for over a third of
blindness. Globally the following conditions account for three quarters of all
blindness: corneal scarring, conditions of the whole globe (such as
microphthalmos and anophthalmos which are relatively common in India),
retinal conditions (mainly dystrophies but also ROP) and disorders of the
lens.
TABLE 50-2 Distribution of the main anatomical

site of abnormality among blind children, by
World Bank economic region
CVI, cerebral vision impairment; Glauc., glaucoma.
In Figure 50-1, data on the magnitude and causes have been combined.
FIGURE 50-1 Estimated magnitude and causes of

blindness in children by cause and World Bank economic
region.
In addition to the 1.025 million children who are blind from eye diseases,
there are estimated to be a further 12.8 million children who are visually
impaired from uncorrected refractive error (89), principally myopia.
Approximately 2 of the 12.8 million children are blind. The largest number of
myopic children and adolescents live in Southeast Asia.
Regional Variation in Causes and Change Over Time
The pattern of causes has changed over time as global initiatives for the
control of vitamin A deficiency and measles have led to a reduction in
corneal scarring, which in 1992 were believed to be the commonest causes in
low-income countries (32). In many low-income countries, cataract is now
the commonest avoidable cause, whereas ROP has increased in importance in
many middle-income countries and is increasing in importance in lower
middle-income countries (90).
Preterm birth, which affects an estimated 15 million infants globally (91),
contributes substantially to blindness in children from lesions of the higher
visual pathways in high-income countries and ROP in middle-income
countries (92).
Cataract and glaucoma are important causes of blindness in children in all
regions but particularly in low-income countries. While the management of
these conditions should be essentially the same throughout the world, in
practice children in low-income settings often present very late which
compromises the visual outcomes of surgery (93,94). Although increasing in
number, specialist eye care centers for children are still not adequate in terms
of numbers or resources to meet the needs of the population (95), and surgery
is often undertaken by general ophthalmologists who are not always aware of
visual development or the specific problems or long-term implications of
operating on children. There are also considerable barriers from the parents’
perspectives, in terms of cost, fear or poor outcomes, the difficulties of travel,
and beliefs concerning the cause of their child’s problem. In Asia, there is
some evidence that girls with bilateral cataract are less likely to access
services than boys, and this probably reflects cultural preferences for boys
(96).
AVOIDABLE CAUSES OF BLINDNESS

IN CHILDREN
The principles of prevention, which can be applied to any impairment, are as
follows:
Primary Prevention
Prevention of occurrence of the disease, leading to a reduction in the
incidence of the disease (e.g., measles immunization)
Secondary Prevention
Treatment of the disease to prevent impairment leading to a reduction in
the incidence of the impairment (e.g., antibiotic treatment of infections,
screening and treatment of type 1 ROP)
Tertiary Prevention
Treatment of the impairment to restore function (e.g., surgery on a
cataract in a blind child), which reduces the prevalence of impairment,
and rehabilitation, which compensates the impairment to reduce
disability (e.g., low-vision or assistive devices).
The available evidence suggests that globally 30% to 40% of children are
blind from conditions amenable to primary and secondary prevention, or
interventions, which restore function from avoidable causes. In high-income
countries, only a few children become or remain blind from avoidable causes,
whereas in low-income settings, at least 50% of blind children have
avoidable causes.
MAJOR AVOIDABLE CAUSES OF

BLINDNESS IN LOW INCOME
COUNTRIES
The major causes of blindness in children in low-income countries are those
that lead to corneal scarring, including vitamin A deficiency disorders
(VADD), measles infection, the use of harmful traditional eye remedies, and
ophthalmia neonatorum. Other causes include congenital rubella and malaria.
Vitamin A Deficiency Disorders
The definition of vitamin A deficiency is not entirely straightforward and has
recently been revised (97) to avoid the term “subclinically deficient,” which
gives the misleading impression that this state is benign. The term VADD is
now being used to reflect the systemic nature and consequences of the
deficiency state. As well as being a cause of blindness in children in
developing countries, clinical trials undertaken over the last 20 years in most
regions of the world have conclusively demonstrated that VADD is also an
important cause of child mortality even in children who do not manifest the
characteristic eye signs (98).
In 2002, it was estimated that there were 140 million preschool-aged
children and more than 7 million women who were vitamin A deficient. More
than 10 of the 140 million children had the clinical signs of VADD (97).
Since then, there have been concerted efforts to scale up supplementation of
high-dose vitamin A with six monthly doses for children between the age of 6
and 59 months. However, coverage remains lower than the 80% target
recommended particularly in East and Southern Africa and South Asia where
coverage is a little over 60% (Figure 50-2) (99). Approximately 46 million
children in the least developed countries are not protected.
FIGURE 50-2 Vitamin A supplementation coverage
(2014) in priority countries. (Reprinted with permission
from Valentine PH, Jackson JC, Kalina RE, et al.
Increased survival of low birth weight infants: Impact on
the incidence of retinopathy of prematurity. Pediatrics
1989;84:442–445. Copyright © 1989 by the American
Academy of Pediatrics.)
Vitamin A is found as retinol in animal food sources, including breast milk,

cheese, fish, liver, and eggs, and as carotenes and carotenoids (provitamin A
compounds) in plant sources, including yellow and red fruits (mango,
papaya), dark green leafy vegetables, and red palm oil. Vitamin A is
necessary for epithelial cell differentiation, growth, gene expression, immune
responses, hemopoiesis, and organogenesis and is an essential component of
rhodopsin. Approximately 95% of total body vitamin A is stored in the liver
as retinal palmitate, and stores are usually sufficient for 6 months. Vitamin A
is released from the liver bound to retinol-binding protein, which is
transported to tissues bound to prealbumin.
VADD results from a combination of poor dietary intake, malabsorption
usually secondary to diarrhea and malnutrition, and increased tissue demand
from any illness, particularly if there is fever. It is useful to think of the
underlying causes of VADD as those that are distal (i.e., the
macroenvironment) and those that are more proximal (i.e., household and
individual factors). Distal causes include all those interrelated factors that
lead to poverty, including inadequate education and health care systems,
political instability, poor roads and communication systems, unfavorable
climate and soil, lack of private landownership, and low prices for products
in the international arena. Under these circumstances, the essential elements
of primary health care are either not available or not accessible on account of
distance, cost, or lack of awareness, and income generation is challenging.
All these factors predispose communities toward poverty and the adverse
health consequences. More proximal causes of VADD at the household level
include low levels of maternal education, inadequate water supplies and
sanitation, overcrowding and large family sizes, and lack of land ownership.
At an individual level, malnutrition, measles infection, and diarrhea can
precipitate a child who is borderline vitamin A deficient into acute deficiency
with keratomalacia or corneal ulceration.
Low dietary intake of vitamin A–rich foods does not necessarily mean
that affordable vitamin A–rich foods are not available; inadequate breast-
feeding, traditional weaning and child-feeding practices and taboos, and
overcooking or sun-drying foods, which reduces the provitamin A content,
are also important factors. Children born in communities where women of
childbearing age also have low vitamin A status, identified by high rates of
night blindness, are also more likely to be deficient. Babies born to women
with adequate vitamin A status will have adequate liver stores, but children
born to deficient women will have deficit stores at birth, and the deficiency
will not be rectified because breast milk levels of retinol are also low.
In VADD, dedifferentiation of conjunctival and corneal epithelia leads to
loss of goblet cells, squamous metaplasia, and xerosis of the conjunctiva and
cornea (known as xerophthalmia). Bacterial action on deposits of keratin in
the conjunctiva leads to accumulation of white, foamy, cottage cheese–like
material. These accumulations, Bitot spots, are usually located at the
temporal limbus and are pathognomonic of VADD (Figure 50-3). Reduced
photoreceptor rhodopsin causes night blindness and, occasionally in adults,
fundus changes. Acute VADD can lead to corneal ulceration and necrosis
(known as keratomalacia) (Figure 50-4).
FIGURE 50-3 Bitot spot in a child with long-standing
VADD. (Courtesy of Clare Gilbert.)
FIGURE 50-4 Keratomalacia in a child with severe, acute
VADD. (Courtesy of Allen Foster.)
Conjunctival xerosis and Bitot spots are signs of long-standing VADD, found
principally in children aged 3 to 8 years. Children most at risk of
keratomalacia are aged 6 months to 4 years with protein energy malnutrition
who may not exhibit other features of xerophthalmia before the onset of
corneal ulceration. Because not all children who are vitamin A deficient
develop eye signs, it is important to be able to identify communities at risk,
because children with xerophthalmia represent only the “tip of the iceberg” of
those who are deficient in vitamin A in the community. Table 50-3 shows the
WHO minimum prevalence criteria for the different ocular signs of VADD,
which indicate that the condition is frequent enough to be a public health
problem. Under-five mortality rates can also be used to assess the likely risk
that VADD is a public health problem (97).
TABLE 50-3 WHO classification and minimum

prevalence criteria for pediatric xerophthalmiaa
and maternal vitamin A deficiency as a public
health problem
aAll stages refer to preschool children except where noted.
bProvisional cutoffs above which community interventions may be warranted. CIC, conjunctival
impression cytology; RDR, relative dose response; MRDR, modified RDR.
Adapted from Sommer A, Davidson FR. Assessment and control of vitamin A deficiency: the Annecy
Accords. J Nutr 2002;132(Suppl 9): 2845S–2850S. Copyright © 2002 The American Society for
Nutritional Sciences, by permission of Oxford University Press.
Primary Prevention for the Control of Blindness Due

to VADD
Poverty alleviation and development.
Improved water supply and sanitation to prevent diarrhea.
Education of women.
Child spacing and family planning programs.
Dietary diversification with health and nutrition education to improve
the intake of vitamin A in children and women of childbearing age, that
is, promotion of breast-feeding, appropriate complementary and
weaning foods, home gardening, and instruction on the preparation and
storage of vitamin A–rich foods. Biodiversity, through cross breeding of
foods to increase their provitamin A content, is also being recognized as
another food-based strategy.
Fortification of commonly consumed foods (an industrial process) and
bio-fortification (by selective cross breeding of plants and crops)
increase pro-retinol content. This approach has been very successful in
the Philippines where monosodium glutamate has been fortified, for
example (37).
Measles immunization.
Distribution of vitamin A to communities at risk (i.e., 50,000 IU every
month for 3 months for infants aged 0 to 5 months; 100,000 IU every 4
to 6 months for infants aged 6 to 11 months; 200,000 IU every 4 to 6
months for children aged 12 months and older; for women, two doses of
200,000 at least 1 day apart postpartum) (16). Distribution can be
through existing health care facilities in conjunction with immunization
programs or as a vertical program. Some countries distribute powders,
which contain additional micronutrients.
Vitamin A prophylaxis of individual children at risk, that is, those with
measles and severe diarrhea from communities at risk.
Many low-income countries are implementing more than one community-

based intervention to control VADD (100).
Vitamin A treatment for all children with signs of xerophthalmia, that is,
three doses given over 2 weeks
Tertiary Prevention
Keratoplasty and optical iridectomy
Corneal grafting in children, particularly penetrating keratoplasty, is always

challenging, but in low-income settings, there are additional obstacles.
Children with corneal scarring usually come from the most disadvantaged
families in the poorest communities, and they cannot afford the surgical,
medical, and optical treatments and regular follow-up visits, which are
essential for a good outcome. Children are also likely to present late and will
be densely amblyopic; good-quality corneal material is rarely available, and
the majority of surgeons are not trained in the surgical techniques and
postoperative care. In addition, surgery is complex, as there are often
adherent leukomas and secondary cataract. Finally, most corneal scarring in
children is associated with vascularization, which leads to very high rates of
graft rejection. For all these reasons, corneal grafting is not recommended for
the control of corneal blindness in children in developing countries (101).
However, optical iridectomy can be very successful for children with dense,
central corneal scarring by restoring sufficient vision for independent
mobility and sometimes for more detailed tasks (102).
Measles
The measles virus is highly contagious, and measles infection can be
associated with high case fatality rates (103). The Expanded Program on
Immunization, which includes measles immunization, has had a dramatic
impact on child deaths: in 1999, there were estimated to be 850,000 measles-
related deaths globally, comprising 10% of all childhood deaths, which had
declined to 89,780 by 2016. Indeed, measles and rubella immunization are
thought to have contributed 23% of the overall decline in under-five mortality
rates between 1990 and 2008 (104). More than 95% of the deaths occurred in
low-income countries where measles immunization rates remain below the
target of 80% coverage (Figure 50-5).
FIGURE 50-5 Measles immunization coverage with first
dose of measles containing vaccines in infants, 2016
(Source: WHO/UNICEF coverage estimates 2016
revision. July 2017. Map production: Immunization
Vaccines and Biologicals, (IVB). Word Health
Organization. 194 WHO Member States. Date of slide: 19
July 2017.)
However, the number of measles cases has recently increased in some high-
income countries as a result, in part, of unfounded fears about the risk of
autism following mumps, measles, and rubella (MMR) immunization
(Figure 50-6) (105).
FIGURE 50-6 Provisional measles cases data (February
2018 to January 2019) based on monthly data reported to
WHO (Geneva) as of March 2019. (From World Health
Organization. Global Measles and Rubella Monthly
Update. March 2019. Available from:
https://www.who.int/immunization/monitoring_surveillance/burden/vpd/s
Malnutrition and overcrowding lead to a higher infecting dose of the virus

and are thought to be major factors contributing to the severity of measles,
particularly in African children (106). Studies in Africa show that
approximately 1% to 3% of children develop corneal ulceration following
measles; approximately a third of children with corneal ulceration give a
history of recent measles, and up to 80% of children with corneal scarring
give a history of measles.
The pathogenesis of measles-related corneal scarring is complex and
multifactorial, and the following mechanisms have been implicated
(107,108): (a) measles keratitis, which reduces corneal epithelium barrier
function; (b) secondary herpes simplex keratitis due to reduced cell-mediated
immunity; (c) secondary bacterial infection; (d) exposure keratitis; (e)
harmful traditional eye medicines, particularly in regions where measles
infection is severe, that is, in sub-Saharan Africa; and (f) acute VADD
leading to corneal ulceration and keratomalacia. As retinol is required for
epithelial differentiation, measles infection imposes a huge demand, as all
epithelial tissues can be affected. Fever also increases the metabolic rate and
demand for retinol at a time when the child is ill and anorexic. Stromal herpes
ulceration can make eating painful, and in many communities, there are
customs and taboos concerning which food sick children should be given.
Sick children are often given watery “soup” made of the local carbohydrate
only, without oil, vegetables, protein, or vitamins. Measles infection of the
gastrointestinal tract leads to malabsorption and protein-losing enteropathy
with loss of retinol-binding protein together with retinol in the feces. Retinol
is also found in the urine of children with measles. So, at the time when the
child most needs high levels of retinol, the intake is low, absorption is
reduced, and there is increased loss in the feces and urine. A child who had
borderline liver stores with normal serum retinol levels before measles can be
precipitated into severe, acute deficiency and lead to keratomalacia and
blindness and death. Repeated doses of high-dose vitamin A should be given
to all children with measles, and this reduces complications, including
mortality (109).
Primary Prevention for the Control of Measles-Related

Blindness
Measles immunization at 9 months, with a second dose at a later date
Health education regarding the use of harmful traditional eye medicines
Prophylactic high-dose vitamin A to all children with measles
Diagnosis and appropriate treatment of corneal ulceration, that is,
repeated high-dose vitamin A, antibiotics, and antiviral agents
Tertiary Prevention
Optical iridectomy
Harmful Traditional Eye Remedies
In animist societies, disease is thought to have several origins, that is, they
may be supernatural, such as being caused by spirits or by angering ancestors
or breaking taboos; they may arise as a result of conflict, tension, jealousy, or
immoral behavior; they may be the result of the “evil eye” or witchcraft; or
they may be passed down within the family through the mother. Other
conditions are due to weakness, eating unclean food, or lack of respect
toward parents or elders. Remedies are based on these understandings. The
WHO has defined traditional healing in the following terms: “the sum total of
all the knowledge and practices, used in diagnosis, prevention and
elimination of physical, mental or social imbalance and relying exclusively
on practical experience and observation handed down from generation to
generation, whether verbally or in writing” (110).
The use of harmful traditional remedies, initiated or used as part of
cultural practices within the family or community, or administered on the
advice of traditional healers, has been an important cause of corneal blindness
in children in low-income settings. This was particularly the case in Africa,
although use of harmful remedies is probably declining as a result of better
levels of education and the decline in measles, which parents feared would
lead to blindness. However, it is important to recognize that traditional
practices are widespread, and although some may be harmful, many are
benign, such as ritual bathing or dances, or may even be of benefit as in the
case of direct application of breast milk into the eye, steam baths, or
inhalations.
There are several mechanisms whereby traditional remedies can lead to
ocular damage and blindness. Firstly, injury to the adnexa from chemical or
thermal burns can lead to exposure keratitis and secondary infection (Figure
50-7). Exposure keratitis can also occur from parents keeping the eyes of the
children held open, a practice believed to prevent blindness in children with
measles in parts of West Africa. Secondly, objects and material inserted into
the eye can cause mechanical damage. Examples include insertion of twigs
and leaves, ground-up cowrie shells, and liquids or sap from plants, which are
either acidic or alkaline. Thirdly, mechanical damage predisposes toward
infection, particularly from fungi if plant material is inserted. Fourthly,
infected fluids can lead to severe infection (e.g., urine from someone infected
with gonorrhea or breast milk that has been expressed into a contaminated
container) (Figure 50-8). Lastly, harmless traditional remedies can lead to
corneal blindness indirectly, as a consequence of delay in seeking more
appropriate treatment.
FIGURE 50-7 Lid burn in a Tanzanian child with measles

as a result of hot oil being administered. (Courtesy of
Allen Foster.)
FIGURE 50-8 Gonococcal keratoconjunctivitis in a girl
who had urine instilled as a treatment for seasonal
epidemic conjunctivitis in West Africa. (Courtesy of Clare
Gilbert.)
Primary Prevention for Control of Blindness Due to

Harmful Traditional Eye Remedies
Good primary health care and primary eye care services that are
accessible and affordable
Health education concerning potentially harmful practices
Training programs for traditional healers, so that they avoid harmful
practices and become part of the primary eye care system (50)
Early diagnosis and appropriate treatment of corneal ulcers
Tertiary Prevention
Optical iridectomy
Ophthalmia Neonatorum
Ophthalmia neonatorum, defined as conjunctivitis within 28 days of birth,
was an important cause of blindness in children at the turn of the century in
Europe. The introduction and widespread use of Credé’s prophylaxis
(cleaning of the lids immediately after birth, followed by instillation of
topical silver nitrate solution) resulted in a dramatic decline in blindness even
before antibiotics were available (111). Current data are not available on the
incidence of vision loss in children from ophthalmia neonatorum.
Ophthalmia neonatorum can be due to a range of organisms, but the most
important pathogens are Chlamydia trachomatis and Neisseria gonorrhoeae.
Transmission of these organisms from infected mother to infant is reported to
be 18% to 50% and 28%, respectively. Despite recent public health measures,
rates of chlamydial infection, particularly among teenage girls, are increasing
in many countries in Europe and North America, whereas rates of sexually
transmitted diseases (STDs) in Africa, particularly due to gonorrhea, remain
high. For example, a recent review of 171 studies in Africa involving over
300,000 pregnant women reported that 4% had gonorrhea and 7% had
chlamydial infection of the genital tract (112).
Approaches to the control of ophthalmia neonatorum still rely on Credé’s
prophylaxis, and several clinical trials have been undertaken in different
settings, which show that 1% silver nitrate solution, 1% tetracycline
ointment, and 0.5% erythromycin are all equally effective against gonococcus
(111). These agents have been recommended by the Canadian Task Force
(113). Studies in Africa have shown that 2.5% povidone–iodine aqueous
solution is also effective (114) and has the advantage of being easy and cheap
to manufacture locally. However, although some argue that tetracycline eye
ointment is preferable (115), a recent systematic review concludes that no one
agent is superior to another and that prophylaxis should only be considered
when rates of STDs are high (116).
Treatment of babies with ophthalmia neonatorum due to gonococcal or
chlamydial infection requires systemic as well as intensive topical treatment,
taking into account the susceptibility of the organism as well as availability
and cost of medication.
The control of gonococcal ophthalmia neonatorum is a challenge in low-
income countries as rates of STDs and infection due to penicillinase-
producing N. gonorrhoeae are high and a high but declining proportion of
births take place outside health care facilities, attended either by trained or
untrained traditional birth attendants or by family members.
Primary Prevention of Ophthalmia Neonatorum

Health education to prevent STDs
Screening and treatment of STDs during pregnancy
Ocular prophylaxis with 1% silver nitrate solution, 1% tetracycline
ointment, 0.5% erythromycin, or 2.5% povidone–iodine
Prompt diagnosis and tropical and/or systemic treatment with antibiotics
appropriate to the microbiology and local susceptibility
Tertiary Prevention
None
Congenital Rubella Syndrome

Rubella infection, which is usually acquired during childhood, is a mild, self-
limiting condition and may be subclinical. However, if a woman becomes
infected during the first trimester of pregnancy, the developing fetus can be
infected, leading to a range of outcomes, including intrauterine death,
premature birth, perinatal death, and structural and/or functional
abnormalities that make up congenital rubella syndrome (CRS). CRS
includes deafness, congenital eye disease, cardiovascular disease,
microcephaly, and developmental delay. If the infection is acquired during
the first 10 weeks of gestation, up to 90% of fetuses are affected, but if
infection occurs after 17 weeks of gestation, about 3% are affected. Without
immunization, CRS is one of the commonest causes of disability in children.
Ocular involvement occurs in approximately 40% of children with CRS with
cataract being the commonest manifestation occurring in approximately 50%
and is followed by microphthalmos, corneal opacity, and glaucoma (117).
Estimates in 1996 suggested that over 100,000 children were born with CRS
in 78 low-income countries that did not include rubella immunization in their
national program. This estimate is likely to be low (118).
Primary prevention of CRS entails rubella immunization. Rubella
vaccine, which uses live attenuated rubella strains, is highly efficacious and
provides protection for up to 20 years (119). Strategies for immunization
include selective targeting of girls and women of childbearing age to protect
future mothers. This strategy does not provide herd immunity and does not
reduce rubella epidemics. Immunization of all children aged 1 to 2 years can
confer herd immunity and prevent epidemics if immunization coverage is
sufficiently high. Another strategy is to immunize all infants and women of
childbearing age who are seronegative. It is very important that the optimum
strategy is adopted, which should be based on local epidemiology. In 2011,
the WHO recommended that when rubella immunization is introduced into
national programs for the first time, an initial campaign should include all
children aged 9 months to 15 years (120) and be supported by surveillance of
rubella infection and CRS cases. In 2012, the WHO launched its Global
Measles and Rubella Strategic Plan 2012–2020 (121). By 2014, 140 countries
included rubella immunization in national programs with Africa being the
region with the lowest coverage where only 15% of countries include rubella
vaccine. Most countries include rubella with the mumps and measles vaccine
(120). Achieving and maintaining high vaccination coverage is critical but
challenging, because there is evidence that low coverage (i.e., where <60% of
infants are immunized) in countries with a high to medium force of infection
can lead to an increase in the number of women of childbearing age who are
susceptible to rubella infection during pregnancy. This scenario has the
potential to increase the number of CRS cases (122), which has occurred in
Greece (123). In 2017, the WHO estimated that 20 million young children
were undervaccinated (124).
A recent review of the impact of rubella immunization on the number of
CRS cases shows that the annual incidence in countries which had not
introduced rubella immunization by the year 2010 remains high, at over one
per thousand live births. Countries that have not introduced rubella
vaccination are mainly in Africa and some in Southeast Asia. In contrast, in
Europe and the Americas, where rubella immunization coverage is high, the
annual incidence is almost negligible (125). Most cases of CRS born in the
United States were due to conceptions that occurred overseas (126).
Primary Prevention of Congenital Rubella Syndrome

Rubella immunization
Early detection, referral, and management of children with cataract and
other eye anomalies
Tertiary Prevention
Rehabilitation and low-vision care
Malaria
Neurologic sequelae following severe malaria are another cause of blindness
in children encountered in low-income settings, particularly in Africa.
Malaria due to Plasmodium falciparum is the most severe form, leading to
severe anemia, hemoglobinuria from massive intravascular hemolysis
(blackwater fever), cerebral malaria and its sequelae, and death. Most
cerebral malaria occurs in children in sub-Saharan Africa, where 575,000
children aged <5 years are estimated to be affected annually. Studies
undertaken in Malawi and West Africa have shown that children with
cerebral malaria can develop characteristic disc and retinal findings, that is,
papilledema, retinal hemorrhages, Roth spots, cotton wool spots, and
extensive areas of retinal pallor and edema (127) (Figure 50-9) (128). Retinal
arterioles and veins can also appear orange in color on account of limited
circulation of red blood corpuscles, which are sequestered in the circulation.
Retinal hemorrhages indicate a poor prognosis. Recent studies of fluorescein
angiography and histopathology confirm that the retinal whitening is due to
cytotoxic edema secondary to acute ischemia (129).
FIGURE 50-9 Malaria retinopathy: a poor prognostic

finding in children with cerebral malaria. (Republished
with permission of Future Medicine Ltd. from Beare NA,
Lewallen S, Taylor TE, et al. Redefining cerebral malaria
by including malaria retinopathy. Future Microbiol
2011;6(3):349–355; permission conveyed through
Copyright Clearance Center, Inc.)
Cerebral malaria has a high mortality rate even with appropriate medical and
supportive treatment, and up to a third of survivors have epilepsy and
neurologic, cognitive, and/or behavioral sequelae that persist longer than 6
months. This includes cerebral vision impairment (CVI). If one assumes that
approximately 10% of children with neurologic sequelae are cortically blind
(130), this represents approximately 1,000 to 2,000 new cases annually.
Many of the children who are blind from CVI have other serious neurologic
problems and, in this setting, are likely to have a very high fatality rate.
Primary Prevention of Malaria

Programs for the control of malaria, with use of impregnated bed nets,
control of mosquito breeding sites, and intermittent treatment of
pregnant women
Prophylactic agents
Diagnosis and treatment of malaria in children with antimalarial drugs to
which the organism is sensitive
Management of children with cerebral malaria, although evidence is
lacking on the most effective treatments
Tertiary Prevention
Rehabilitation
MAJOR CAUSES IN ALL REGIONS

Lesions of the CNS contribute significantly to blindness, particularly in high-
income settings where they are often secondary to prematurity. Fetal alcohol
syndrome (FAS) is another potentially preventable cause. Myopia among
older children and adolescents is increasing in all regions, particularly in East
Asia, and ROP, which only affected preterm infants in high-income countries
until 30 years ago, is now an important cause of blindness in upper and lower
middle-income countries and is becoming a problem in urban centers in low-
income countries.
Lesions of the Central Nervous System

In a national surveillance study in the United Kingdom, 48% of children
newly diagnosed as blind were blind from lesions of the higher visual
pathways, and an additional 13% were blind from optic atrophy (38). Studies
from other European countries report 15% to 54% of children to be blind
from CNS abnormalities. The high figure of 54% was reported from the study
from the Republic of Ireland, which included children identified in
institutions for children with multiple impairments as well as those registered
as blind (131).
In studies from European countries and India, CNS lesions were
attributed to a range of factors including metabolic and genetic diseases, the
effect of teratogens, intrapartum events associated with birth asphyxia, the
consequences of prematurity, and postnatal events including tumors, trauma,
and meningitis (132). A high proportion of children blind from CNS lesions
have additional impairments, most commonly CP, as well as a range of other
ocular conditions. Recent studies have also highlighted that non–visually
impairing lesions of the ventral and dorsal streams are common, particularly
in children who were born preterm, and give rise to a range of functional
difficulties, such as visual guidance of movement, search, and attention (i.e.,
visual agnosia) (133–135). These difficulties are frequently undiagnosed or
misdiagnosed.
Hypoxia resulting from intrapartum events has frequently been recorded
as the underlying cause of both CP and blindness. However, there is a
considerable body of evidence from high-income countries suggesting that
clinical indicators of birth asphyxia (i.e., low Apgar score and low umbilical
pH) are very poor predictors of neurologic abnormality (136). In one
longitudinal study in Australia, only 8% of cases of CP were attributed to
intrapartum events (137). Longitudinal population-based studies have also
shown that the incidence of CP in infants with normal birth weight is
relatively stable despite improvements in obstetric care. The underlying cause
of CP in children with normal birth weights in these settings seems to be due
to ill-defined events or processes occurring before birth, which either cause
cerebral damage or make the cerebrum more susceptible to damage during
delivery. There is also emerging evidence that at least 10% of children with
sporadic CP have likely causative single-gene mutations (138). The situation
is, however, different in low-income settings where a wider range of
conditions lead to CP, including neonatal meningitis, and prolonged or
difficult delivery (139). Whether the pattern of visual impairment is the same
between these two settings is not yet known.
The incidence of CP and visual loss is higher in children with low birth
weight and in those born at or before 32 weeks of gestation (136).
Periventricular leukomalacia and para- and intraventricular hemorrhage are
found in a higher proportion of preterm infants particularly those with birth
weights <1,000 g at birth. Infants at the boundaries of survival, such as those
born <26 weeks of gestational age, are at the greatest risk with up to 37%
having evidence of CNS injury (140). Once again there is debate concerning
the etiologic significance and interrelationship of prenatal, intrapartum, and
neonatal events since preterm birth may itself be a consequence of adverse
intrauterine events and predispose the immature cerebral tissue to damage
during birth and the neonatal period. There is some evidence that the better
survival of extremely low birth weight babies over the last 10 to 15 years has
been at a cost in terms of higher rates of disability (140).
Primary Prevention of Lesions of the CNS

Prevention of preterm birth (see section on ROP below)
Prevention of neonatal meningitis
Good antenatal and obstetric care
Early diagnosis and management of neonatal sepsis
Tertiary Prevention
Rehabilitation
Fetal Alcohol Syndrome
Alcohol consumption during pregnancy can lead to a range of phenotypes,
called fetal alcohol spectrum disorders (FASD), which reflects the range of
structural and functional abnormalities that can result from alcohol exposure
in utero. The full syndrome (FAS) is now one of the most common and most,
preventable causes of cognitive impairment in children. For example, in the
United States, FASD has a birth prevalence of 1.9/1,000 live births
(compared with 1.6/1,000 live births for Down syndrome). South Africa is
reported to have the highest rates of FASD anywhere in the world where it
occurs in 3 to 30/1,000 live births (141).
For diagnostic purposes, the child should have each of the following
abnormalities, although they may be present in varying degrees: prenatal
and/or postnatal growth retardation, structural and/or functional CNS
abnormalities, and a characteristic facial appearance (i.e., microcephaly, short
palpebral fissure, poorly developed philtrum, and a flat maxilla). Up to 90%
of children with FASD have ocular signs and symptoms, including optic
nerve abnormalities, such as optic atrophy or optic nerve hypoplasia (48%),
tortuous retinal blood vessels (50%), short palpebral fissure, broad telecanthic
folds, ptosis (50%), and strabismus (40% to 50%) (142). Less frequent
changes include high refractive errors, microphthalmos, nystagmus, cataract,
and corneal opacity, including Peters anomaly. Affected children can also
develop a range of psychosocial disabilities and behavioral problems (141).
The full syndrome occurs in 30% to 50% of the children born to alcoholic
women. Whether there is a safe limit for alcohol consumption during
pregnancy has not yet been clarified, but the current recommendations are to
abstain.
Primary Prevention
Health education before conception and during antenatal care about the
harmful effects of alcohol during pregnancy
Secondary PreventionNo effective interventions

Tertiary Prevention
Management of ocular features, low-vision care, and rehabilitation
Myopia in Children
Myopia, which is the most common type of refractive error, is increasing
globally, particularly among children in the economically advanced countries
of East and Southeast Asia (143–145). In these countries, myopia can start
early, around the age of 6 or 7 years, and can affect 80% of adolescents. In
these countries, the prevalence is higher in children in urban areas where
myopia can have an earlier age of onset and progress to high myopia
throughout adolescence. High myopia is of particular concern as it increases
the risk of vision impairment from myopic macular degeneration, retinal
detachment, cataract, and glaucoma later in life (146). Indeed in 2015, it was
estimated that 10 million people were visually impaired from myopic macular
degeneration, which is projected to increase to 55.7 million by 2050 (147).
Factors implicated in the increasing incidence and severity of myopia
associated with urbanization include greater educational pressure with
prolonged periods of close work and lack of time spent outdoors, the latter
probably being mediated by lack of exposure to daylight (145,148,149).
Having parents with myopia is known to increase the risk, and interactions
between behavioral and environmental factors and genetic variants are an
active area of research, because on their own, genetic variants only explain
0.6% to 2.3% of the variance in refractive error (150).
Several approaches are being explored to reduce the risk of myopia or to
slow its progression and include optical, pharmacologic,
behavioral/environmental, or surgical approaches (151). Interventions include
topical medication with antimuscarinic eye drops, alternative ways of
correcting myopia with bifocal contact lenses or spectacles, for example,
increasing the amount of time children spend in daylight by being outdoors or
by altering the design of classrooms, or surgery to reinforce the sclera.
There is evidence from clinical trials that antimuscarinic eye drops slow
the progression of myopia, but side effects, such as increased sensitivity to
light, blurred near vision and allergies, lack of availability of the medication,
and the need for prolonged treatment, limit their usefulness (152). There is
increasing evidence that the amount of time children are exposed to daylight
reduces the incidence of myopia, with a greater impact among younger
children and by spending longer times outdoors (153–155). However,
paradoxically, this does not seem to reduce the progression of established
myopia. Myopia control is an active area of research given the impact of
uncompensated myopia on academic achievement (156) and the long-term
complications of high myopia.
Primary Prevention
Increasing time spent outdoors (limited evidence)
Topical muscarinic drugs
Tertiary Prevention
Optical correction
Globally, there are approximately 15 million preterm births, defined as birth
<37 weeks of gestational age (GA), each year, and approximately 2.3 million
are born <32 weeks of GA (157). Ten countries account for 60% of preterm
births: China, India, Nigeria, Pakistan, Indonesia, the United States,
Bangladesh, the Philippines, and the Democratic Republic of the Congo.
Several epidemics of blindness due to ROP have been described (Figure
50-10). The most recent, the third epidemic, started in Latin America and
Eastern Europe in the 1990s, then spread to the Middle East and East and
South Asia, and is beginning to emerge in sub-Saharan Africa (92).
FIGURE 50-10 Epidemics of blindness due to ROP.
(Reprinted from Gilbert C, Malik ANJ, Nahar N, et al.
Epidemiology of ROP update—Africa is the new frontier.
Semin Perinatol 2019;43:317–322. Copyright © 2019
The main driver of the third epidemic is the rapid expansion of intensive
neonatal care as the Ministries of Health strive to reduce neonatal mortality.
However, the level of care can be suboptimal with inadequate equipment to
safely deliver supplemental oxygen from immediately after birth, lack of
qualified staff with high nurse-baby ratios, and overcrowded units, all of
which compromise care. In addition, many countries have an absolute lack of
ophthalmologists to screen and treat ROP, or the number of skilled
ophthalmologists with the requisite time and motivation is low. In addition, in
settings were neonatal care is suboptimal, more mature babies can develop
sight-threatening ROP, which means a high proportion of infants need to be
screened (158).
In 2013, it was estimated that 32,300 infants were becoming blind or
visually impaired from ROP every year (90). All regions are now affected
(Figure 50-11) (159) with studies being reported from sub-Saharan Africa
(160).
FIGURE 50-11 Annual incidence of visual impairment

and blindness from ROP per 100,000 live births.
(Reprinted by permission from Springer: Blencowe H,
Moxon S, Gilbert C. Update on blindness due to
retinopathy of prematurity globally and in India. Indian
Pediatr 2016;53(Suppl 2):S89–S92.)
Many countries, such as those in Latin America and South Africa, now have
ROP programs with some achieving high coverage of screening and
treatment. Programs are scaling up in India (161) and China, and initiatives
are starting in some sub-Saharan countries, such as Nigeria, Kenya (162),
Ghana, and Tanzania, and in Pakistan and Bangladesh. A study in Latin
America highlighted the importance of national guidelines, health
management information systems, and legislation that mandates ROP
screening as being important factors leading to high coverage (163).
However, many countries do not yet have guidelines (164), which need to
vary depending on the population at risk (165). The active engagement of
Ministries of Health is also vital (166).
Reports of the proportion of infants who develop any ROP or “severe” or
“treatment-warranted” ROP varies considerably depending on the case mix,
the definitions used, the criteria for screening, and the expertise of the
examining ophthalmologists. In the United States, in a multicenter cohort of
7,483 infants, 459 (6%) developed type 1 ROP (167); in Switzerland, 1.9% of
6,719 infants developed stage 3 to 5 ROP (168); and in the United Kingdom,
4% of 8,112 infants were treated (169). In these studies, the majority of
infants treated were extremely preterm. In India, the proportion treated is
often similar despite the use of much wider screening criteria. A higher
proportion develops aggressive posterior ROP (APROP), and the birth weight
and GA of affected infants are far higher. For example, in one study, 4.4% of
4,161 infants were treated for ROP, 17% of whom had APROP (170).
The main risk factors for ROP are increasing prematurity, low birth
weight, and intrauterine growth restriction. Postnatal risk factors include poor
weight gain, sepsis, early hypothermia (171), hyperglycemia, and poorly
regulated oxygen leading to hyperoxia or fluctuating hypoxia/hyperoxia
(172). Other potentially modifiable risk factors include transfusion of blood
products and thrombocytopenia (173). There is also evidence that a course of
antenatal steroids for women threatening preterm delivery improves a range
of outcomes, including ROP (174). (See also Chapter 52.)
Neonatologists have to tread a difficult path in relation to the
administration of supplemental oxygen. A recent meta-analysis of the five
multicenter clinical trials compared the outcomes of different oxygen
saturation targets (NeOProM) (175). The lower target oxygen saturation
target range (85% to 89%) had a higher risk of mortality, but a lower risk of
ROP treatment than the higher target range (91% to 95%). There remains
controversy as to the saturation targets recommended, but some reports have
recommended 90% to 94% (alarm limits 89% to 95%) in high-income
countries (176) and 88% to 95% (alarm limits 89% to 96%) in low- and
middle-income countries (177).
Primary Prevention
Prevention of preterm birth
Regular antenatal care to detect and manage complications, which

predispose to preterm birth (e.g., hypertension), and health education
about good nutrition and behavioral factors (e.g., smoking cessation)
Regulation of assisted fertilization to reduce multiple births
Prevention of ROP among preterm infants
A course of antenatal steroids to women threatening preterm delivery as

recommended by WHO guidelines (2015) (178).
High-quality neonatal care immediately after birth, including delayed
cord clamping, gentle respiratory support, careful use of oxygen, and
prevention of hypothermia (179).
High-quality care throughout the neonatal period, including safe
delivery of oxygen, infection control, promotion of weight gain, pain
and temperature control, and supportive care (such as kangaroo mother
care and nesting). Caffeine treatment for apnea also reduces severe ROP
(180).
Screening for ROP of preterm babies at risk, starting 3 to 4 weeks after
birth
Urgent treatment (within 48 hours) of infants developing the sight-
threatening stages of ROP, with close follow-up
Vitreoretinal surgery for stage 4a ROP
Tertiary Prevention
Vitreoretinal surgery for the stage 4b and 5 ROP
Frequent follow-up throughout childhood of babies treated for ROP to
detect and manage complications such as refractive errors and squint
Rehabilitation
CHANGE IN MAJOR CAUSES OVER

TIME
As the causes of blindness in children reflect levels of socioeconomic
development and health care provision, it is hardly surprising that the pattern
of causes of blindness changes over time in response to changes in the wider
environment, medical advances, and provision of services. Some notable
examples include the increase in blindness due to ROP in middle-income
countries (75,76) and the increasingly important contribution of prematurity
and FAS in industrialized countries. In low-income countries, the marked
increase in coverage with measles immunization, together with multisectoral
initiatives to control micronutrient deficiencies, including VADD, means that
blindness due to corneal scarring is declining in many of the poorer regions
of the world.
New conditions that can cause vision impairment and blindness have
emerged over the last decade, such as Ebola virus disease in Africa, which
can lead to chronic uveitis and secondary cataract in adolescents (181), and
Zika infection, which leads to vision impairment among affected children in
Brazil and other countries in the region (182).
UNIVERSAL HEALTH COVERAGE

In 1997, the International Agency for the Prevention of Blindness, an
umbrella organization of over 100 agencies, organizations, and professional
bodies involved in eye care, prevention of blindness, and services for the
blind, together with the WHO launched a global initiative called VISION
2020: The Right to Sight. The control of blindness in children is one of the
priorities of this initiative, which embodies advocacy, resource mobilization,
community participation, human resource development, placement of
appropriate infrastructure and equipment, and implementation of cost-
effective control strategies. The control of blindness in children was included
not on the basis of the numbers affected, which are small in comparison to
the number of blind adults, but because of the long-term impact and
consequences of blindness in children. This initiative is due to end in the year
2020.
Over the last 20 years or so, much has been achieved. The control of
blindness in children is now included in the national prevention of blindness
plans in many countries, and several international nongovernment
organizations (NGOs) have prioritized the control of blindness in children.
However, much more needs to be done: from policy to the development of
comprehensive eye care services that are affordable and child friendly, that
extend from communities to tertiary levels, and that encompass the wide
range of ocular conditions that can affect children.
Universal health coverage, which is being promoted by the WHO, means
that “all people and communities can use the promotive, preventive, curative,
rehabilitative and palliative health services they need, of sufficient quality to
be effective, while also ensuring that the use of these services does not
expose the user to financial hardship.
This definition of UHC has three related objectives:
Equity in access to health services—everyone who needs services

should get them, not only those who can pay for them;
The quality of health services should be good enough to improve the
health of those receiving services; and
People should be protected against financial-risk, ensuring that the cost
of using services does not put people at risk of financial harm (183).”
Ministries of Health are currently identifying their priorities for UHC in their
respective countries, and this provides a unique opportunity for prioritizing
eye care (184). In particular, it is imperative that eye care for children is
integrated into the broader agenda for child health, including for primary eye
care (185,186), the integration of correction of refractive errors into school
health, ROP screening as an integral component of neonatal care, and eye
care as an essential element of tertiary-level health care.
ACKNOWLEDGMENTS
CG Support from The Queen Elizabeth Diamond Jubilee Trust, United
Kingdom, for work on ROP in India and other Commonwealth countries is
acknowledged.
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51
Education and Management of
Worldwide
Tala Al-Khaled, Samir N. Patel, and R. V. Paul Chan
INTRODUCTION
First and Second Epidemic of Retinopathy of
Prematurity
Retinopathy of prematurity (ROP) is a vasoproliferative disorder of the
developing retina, described by Terry in 1942 (1). At that time, ROP was
described as “retrolental fibroplasia.” There were few treatment options, and
a significant number of infants developed total retinal detachments (2).
During the 1940s and 1950s, the “first epidemic” of ROP occurred mainly in
developed countries due to unmonitored use of oxygen in premature babies
(3). A better understanding and restriction of oxygen supplementation
subsequently led to reduced blindness. However, as advances in prenatal and
neonatal care led to the increased survival of smaller and less mature babies
in the 1970s and 1980s, another wave of ROP began, leading to the “second
epidemic” of ROP (4).
Fortunately, a better understanding of the natural course of ROP and
advances in treatment have led to improved structural and functional
outcomes, first with the multicenter prospective trial, Cryotherapy for
Retinopathy of Prematurity (CRYO-ROP) (5), then the Early Treatment for
Retinopathy of Prematurity (ET-ROP) trial (6), and, more recently, the
Bevacizumab Eliminates the Angiogenic Threat of Retinopathy of
Prematurity (BEAT-ROP) study, which has reported the utility of intravitreal
antivascular endothelial growth factor (VEGF) agents in some cases (7) (see
also Chapters 52 and 53).
Third Epidemic of ROP and Global Workforce

Shortage
Despite advances in ROP care, the disease remains a major clinical challenge
in the United States (8). Continued improvements in medical care have
translated into increased neonatal survival rates and, therefore, a growing
number of infants at risk for ROP (9,10). Meanwhile, the number of
ophthalmologists who manage ROP is limited and expected to decrease in
part because of concerns of medicolegal liability and the complexity of
scheduling care (11). Therefore, in the United States, there are increasingly
fewer ophthalmologists willing to meet the growing need for ROP
surveillance.
Meanwhile, from a global perspective, there is resurgence of ROP,
sometimes called the “third epidemic,” mostly in middle-income countries,
and most recently, we are seeing an increase in ROP in sub-Saharan Africa
(12,13). In these countries, premature babies are now surviving at a greater
rate due to medical advances. However, inadequate resources and facilities,
varied ROP screening programs, and a lack of experience with ROP
management are resulting in thousands of children going blind from ROP. In
this chapter, we discuss how emerging treatments, technology, and
educational initiatives relate to both the growing burden of ROP in the United
States and the growing epidemic worldwide.
TREATMENT
Advancements in ROP management were facilitated in the 1960s and 1970s,
when both laser photocoagulation and cryotherapy were shown effective as a
treatment (5,14,15). The CRYO-ROP multicenter trial examined cryotherapy
versus observation in eyes of premature babies with threshold ROP. A
significant reduction in unfavorable structural outcomes (retinal detachment,
macular fold, or retrolental tissue) and visual acuity of <20/200 was found in
eyes undergoing cryotherapy compared with untreated eyes (5) (details in
Chapter 52). These early results would continue to hold true as the 15-year
follow-up demonstrated unfavorable structural outcomes in about 30% of
treated eyes compared to over 50% of control eyes. Additionally, unfavorable
visual acuity outcomes (20/200 or worse) were significantly reduced in
treated eyes (16).
Based on the results of the CRYO-ROP trial and the development of a
risk analysis model, investigators subsequently investigated the role of earlier
treatment. The ET-ROP study was initiated in the early 2000s and was a
multicenter clinical trial that compared treatment for eyes with type 1 “high-
risk prethreshold” disease and type 2 “low-risk prethreshold” disease versus
eyes treated at conventional “threshold” ROP. Results showed significantly
decreased unfavorable visual acuity and structural outcomes (6).
There has been an increase in the use of intravitreal bevacizumab
(Genentech Inc., San Francisco, CA) for the management of ROP. Initial case
reports in 2007 showed use in the management of aggressive posterior ROP
and other cases of aggressive disease (17,18). The BEAT-ROP study, a
clinical trial comparing laser and anti-VEGF (bevacizumab 0.625 mg), found
a very short-term benefit of reduced need for further treatment in advanced
ROP even beyond type 1 ROP or the older classification from CRYO-ROP of
threshold ROP and laser in infants (7). In addition, the use of anti-VEGF
appears to have lengthened the time for recurrence or full vascularization,
which was determined as a safe time to stop weekly eye examinations in
treated ROP (see Chapter 52). In the past, these infants had poor outcomes;
the use of bevacizumab offers hope, but until safety and dosing studies are
performed, anti-VEGF is still of concern for both the developing infant and
eye. Currently, at least two multicenter trials are underway for forms of
severe ROP, one by the Pediatric Eye Disease Investigator Group through the
National Eye Institute to assess the lowest effective dose of bevacizumab
given through intravitreal injections and another through Novartis
(RAINBOW Study: RAnibizumab Compared With Laser Therapy for the
Treatment of INfants BOrn Prematurely With ROP), which compare laser to
two doses of ranibizumab in 0.05 mL (19,20).
BURDEN OF ROP AND ROP

EDUCATION: UNITED STATES
Identifying the Problems
In the United States, ROP continues to be an important problem. Despite
advances in preventative measures and an increased understanding of its
pathophysiology, ROP still occurs in about 15,000 infants per year, with
approximately 500 cases of blindness from advanced disease (8). Moreover,
similar technologies and discoveries in neonatal care have resulted in a
growing number of infants at risk for ROP due to increased survival of less
mature and smaller infants (9,10). In the United States, concerns over
medicolegal liability, poor reimbursement, and the complexity of
coordinating care have led to a limited and decreasing number of
ophthalmologists willing to manage ROP. A 2006 American Academy of
Ophthalmology survey showed that only half of retinal specialists and
pediatric ophthalmologists were willing to manage ROP and about a fifth
planned to stop providing ROP care in the future (11).
Potentially exacerbating this trend are data that suggest inadequate
preparedness for the future workforce responsible for ROP care. Chan et al.
(21) studied the accuracy of image-based ROP diagnosis of retina fellows
compared to expert specialists. Data showed that while retina fellow
diagnosis of treatment-requiring ROP was relatively high, there was more
variability in diagnosis of type 2 prethreshold ROP, in which 47% of eyes
were overcalled as treatment-requiring ROP and 36% were undercalled as
mild to no ROP (Figure 51-1). A follow-up study by Myung et al. (22)
involved pediatric ophthalmology fellows and showed similar results. For the
detection of type 2 or worse ROP, fellows demonstrated a mean sensitivity of
0.527 and mean specificity of 0.938. Figure 51-2 displays examples of
images from one eye that was frequently misdiagnosed by these pediatric
ophthalmology fellows. These studies have significant clinical implications
because accurate diagnosis is crucial for appropriate management of ROP,
especially since anecdotal evidence shows some institutions utilize fellows
for primary evaluation of ROP patients. Overdiagnosis can lead to
unnecessary interventions, whereas underdiagnosis can lead to missing
potentially progressing ROP and blinding disease without appropriate
treatment.
FIGURE 51-1 Mean distribution of ROP diagnoses by
seven retinal fellows. A:Among the 28 eyes with a
reference standard diagnosis of type 2 prethreshold ROP,
the 7 fellows diagnosed type 2 prethreshold ROP in 5
(17%) eyes. B:Among the 15 eyes with a reference
standard diagnosis of treatment-requiring ROP, the 7
fellows diagnosed treatment-requiring ROP in 14 (91%)
eyes. (Reprinted with permission from Chan RVP,
Williams SL, Yonekawa Y, et al. Accuracy of retinopathy
of prematurity diagnosis by retinal fellows. Retina
2010;30(6):958–965.)
FIGURE 51-2 Example of study images frequently

misdiagnosed by fellows. Nasal (A), posterior (B), and
temporal (C) images diagnosed as type 2 ROP by the
expert reference standard and no ROP by two of five
fellows (40%), mild ROP by one of five fellows (20%),
type 2 by one of five fellows (20%), and treatment-
requiring by one of five fellows (20%). (Reprinted from
Myung JS, Chan RVP, Espiritu MJ, et al. Accuracy of
retinopathy of prematurity image-based diagnosis by
pediatric ophthalmology fellows: implications for training.
J AAPOS 2011;15(6):573–578. Copyright © 2011
In an effort to further explore this issue, two web-based surveys of training

programs in the United States have characterized trends in ROP education. In
a survey of ophthalmology residents and residency program directors, Nagiel
et al. (23) showed that two-thirds of responders estimated that residents
performed <20 ROP examinations during their entire residency training.
Also, despite over 80% of resident responders reporting they were not
confident in their examination skills, about one-third of resident examinations
lacked supervision by an attending at the bedside. A separate study of
pediatric ophthalmology and retina fellowship training programs also showed
concerning trends (24). From almost 30% of respondents, up to two-thirds of
ROP examinations performed by fellows were not followed up by an
attending. Therefore, there were a significant number of infants who were
examined exclusively by a trainee. Additionally, there was great variability in
the number of ROP examinations performed by fellows across programs,
with pediatric ophthalmology fellows performing significantly fewer laser
photocoagulation procedures than retina fellows (Figure 51-3). Also, only
7% of respondents reported the use of formal evaluations to assess fellow
competency in ROP management. Moreover, U.S. ophthalmology residents’
competency in ROP diagnosis was assessed by Swamy et al. through a series
of web-based cases (25). Overall, ROP of any category was accurately
diagnosed 48% of the time. In particular, type 2 ROP was only correctly
identified in 34% of cases with a tendency for underdiagnosis, rather than
overdiagnosis, of this category of disease (25). Notably, failure to correctly
assess for the presence of plus disease was a leading cause in the
misdiagnosis of type 2 ROP (25). The survey and study data, therefore,
showed the following regarding ophthalmology training in the United States:
(a) a limited exposure to ROP in ophthalmology residency; (b) lack of direct
attending supervision during ophthalmology residency and fellowship ROP
training; (c) ROP accurately diagnosed by residents in less than half of cases;
and (d) extremely varied ROP curricula and experience across fellowship
programs.
FIGURE 51-3 Attending and fellow responses to the
number of ROP laser photocoagulation procedures
performed by fellows during the course of their training,
stratified by specialty. (Reprinted from Wong RK, Ventura
CV, Espiritu MJ, et al. Training fellows for retinopathy of
prematurity care: a web-based survey. J AAPOS
Association for Pediatric Ophthalmology and Strabismus.
With permission.)
The study results revealed possible areas of refinement in ROP education:

increasing ROP exposure during ophthalmology residency may be beneficial;
having a more robust experience in residency may not only inspire more
physicians to later consider incorporating ROP into their future practice, but
it would also better prepare those who actually pursue subspecialty training in
pediatric ophthalmology or retina; and a greater emphasis should be placed
on attending supervision of trainee ROP examinations at the bedside. As the
data demonstrated, currently many trainee ROP examinations occur without
attending supervision. Having multiple, separate examinations may increase
stress on already sick infants as well as present logistical challenges with
respect to scheduling repeated dilated exams in the neonatal intensive care
unit (NICU) (26). Incorporating supervised instruction may minimize these
issues and also allow for direct and immediate comparison of examination
techniques, clinical findings, and treatment recommendations between expert
and novice. Moreover, supplementing supervision of indirect
ophthalmoscopy with occasional live wide-angle digital imaging may prove
to be a beneficial teaching approach. Several reports demonstrated that the
use of digital imaging during laser treatment helps detect skip areas after
panretinal laser photocoagulation (27). The use of digital images can identify
areas of missed laser by ophthalmologists in training even when the areas
were not able to be detected by indirect ophthalmoscopy (28) (Figure 51-4).
With all concerns taken together, there is an increased need for ROP
surveillance, but there is currently a lack of sufficiently trained
ophthalmologists for examinations, and the need of trained ophthalmologists
appears to be growing.
FIGURE 51-4 A:Digital retinal image (RetCam II, Clarity
Medical Systems, Pleasanton, CA) of skip area
(demarcated in red) adjacent to the ridge found in the 9:00
to 11:00 clock-hour region. B:Digital retinal image of skip
area adjacent to the ora serrata found in the 3:00 to 5:00
clock-hour region. C:Digital retinal image of skip area
adjacent to the ridge found in the 5:00 to 7:00 clock-hour
region. D:Digital retinal image of several skip areas found
in the 11:00 to 1:00 clock-hour region.
Potential Solutions
Standardization of ophthalmology fellowship curricula may also help
improve training in ROP management. Currently, the Association of
University Professors of Ophthalmology (AUPO) Fellowship Compliance
Committee is attempting to establish uniform training standards. For
example, there are suggestions to retina fellowship programs regarding
minimum operative numbers for vitrectomies and scleral buckles. However,
fellowship programs are not required to follow these AUPO guidelines, and
in fact, many reputable fellowships are not in compliance. Moreover, even
among fellowship programs adhering to AUPO standards, there are no
recommendations for pediatric ophthalmology and retina fellows regarding
ROP examinations, ROP laser procedures, or assessment of competency in
ROP management (29,30). Perhaps there is a need to establish a minimum
number of ROP examinations and laser procedures as well as incorporate
other tools such as ophthalmic clinical exercises, 360-degree assessments
(evaluations based on multiple sources, such as supervising faculty, peers,
residents, nurses, technicians, students, and patients), and global faculty
performance ratings for ROP fellow training (31–34). These devices have
proven useful in ophthalmology residency training and may be helpful in
fellowship training as well.
Lastly, an area of needed research and commentary is the current
American Board of Ophthalmology certification process for
ophthalmologists. Board certification, and recredentialing every 10 years
under the American Board of Ophthalmology, is widely sought after among
practicing ophthalmologists; however, no credentialing is offered for
subspecialty care, such as advanced surgeries in cornea, glaucoma, or retinal
disease. The clinical significance of this lack of recredentialing regarding
ROP involves results from Kemper et al. (35), which found that many
ophthalmologists without subspecialty training were performing ROP
screening and treatment. It is unclear if formal training in pediatric
ophthalmology or retina makes a difference in competency of ROP
management, but a credentialing process to ensure certain standards of
training would likely help with public accountability (36). Though an
unproven correlate, studies in intensive care medicine, made up of a wide
range of specialists, including critical care medicine, pulmonology, surgery,
anesthesiology, and neurology, have shown that care led by intensivists with
certification in critical care medicine is strongly associated with improved
patient survival (37).
BURDEN OF ROP AND ROP

EDUCATION: WORLDWIDE
Identifying the Problems
Despite major advances in management, ROP continues to be a devastating
problem worldwide, causing blindness in more than 50,000 children (38). As
expected, high-income and developed countries do not constitute the largest
share of this staggering number, mainly due to high-quality neonatal services,
excellent equipment, evidence-based screening protocols, and a sufficient
access to ophthalmologists trained in ROP management. Perhaps
surprisingly, low-income countries, on the opposite end of the socioeconomic
spectrum, also are not responsible for the majority of these cases. In these
countries, poor survival of preterm infants from a lack of neonatal resources
reduces the number of infants at risk for developing ROP. Rather, the largest
portion of blind children from ROP comes from middle-income countries and
has been called the “third epidemic” of ROP (12).
Although most of the morbidity and blindness are potentially preventable
in middle-income countries, the health care systems may not have the
resources to deliver high-level care necessary to effectively manage children
with ROP. In middle-income countries, rising numbers of children with
blindness from ROP have multifactorial causes (39). First, when compared to
higher-income nations, teenage pregnancy tends to occur more often, and
utilization of prenatal care is less common (40). Second, although neonatal
care is advancing and allowing survival of smaller and less mature babies,
inadequate equipment and insufficient nursing staff do not allow for
continuous supplemental oxygen monitoring and regulation. Lastly, screening
programs tend to vary, and the risk profile of babies at risk for developing
ROP is different than that in high-income countries (38). Studies have shown
that ROP occurs in larger, more mature babies in middle- than in high-
income countries. Therefore, if protocols recommending screening of
younger gestational age and smaller birth weight infants such as in high-
income countries like the United States or United Kingdom are used in
middle-income countries, cases of ROP may be missed (41,42).
Many middle-income countries have an inadequate number of
ophthalmologists experienced in ROP management or lack facilities and
infrastructure to deliver conventional laser therapy. Gilbert et al. (38)
reported that in Lima, Peru, a single ophthalmologist provided ROP care for
the entire city of 8 million people. Anecdotally, although there may be ROP
coverage, some cities in Latin America lack neonatal units equipped with
lasers or the necessary anesthesia services available for sedation or general
anesthesia. Therefore, laser photocoagulation often cannot be offered in these
countries.
Given the above, it is important to understand the level of preparedness
of international ophthalmologists to manage ROP. A recent survey of
international ophthalmologists’ exposure to ROP during training reported
that, for nearly half of the participants, screening for ROP under direct
attending supervision occurred between 1% and 33% of the time during
training and trainees performed zero laser photocoagulation procedures for
treatment of ROP (43).
Furthermore, international ophthalmology trainees participated in a web-
based study that prompted users to classify ROP based on a collection of
wide-field retinal images (44). The structure of this study was identical in
nature to the study directed toward trainees in the United States (25).
Likewise, the findings characterizing the common errors made by
international trainees were consistent with their U.S. counterparts: (a) ROP
was incorrectly diagnosed in more than half of the cases; and (b) treatment-
requiring ROP and type 2 ROP were commonly underdiagnosed due to poor
accuracy in identification of plus disease (44).
In summary, these findings suggest the following in regard to
international ophthalmology training: (a) limited surveillance of trainees
when performing ROP examinations; (b) inconsistent structured feedback
geared toward ROP that is provided to trainees and absence of uniform ROP
curricula in training programs; and (c) competency of ophthalmology
residents in the diagnosis of ROP, which requires significant enhancement,
particularly in the setting of middle- and lower-income countries where
comprehensive ophthalmologists may be the only physicians screening for
ROP. Since this pattern of findings is seen in training programs both in the
United States and on an international level, additional considerations should
be made to reform and standardize current ophthalmology training program
objectives to emphasize competency in screening and treatment of ROP.
Potential Solutions
Preventative Measures
In the context of the “third epidemic” of ROP, there are discussions regarding
fundamental changes in health care practices and policies with the aim of
improving the care of neonates at risk for ROP. The Every Preemie-Scale
initiative funded by the United States Agency for International Development
(USAID) released a Do No Harm Technical Brief with recommendations
summarized as follows: (a) tailor screening criteria by geography, which
refers to modifying gestational age and birth weight cutoffs in light of care
received in the NICU and oxygen usage; (b) educate health care providers on
the appropriate use of oxygen delivery equipment and the importance of
oxygen level monitoring; (c) ensure adequate workforce is available to
provide neonatal care; (d) secure equipment necessary to perform retinal
exams and assess for ROP; and (e) create national policies that mandate
screening for ROP, as appropriate (45). In regard to implementing policy
change, the Economic Model of Retinopathy of Prematurity (EcROP) was
applied to regions in the United States and Mexico and found that a
comprehensive ROP screening program is cost-effective and cost-saving,
indicating the benefits of establishing national protocols for ROP (46).
Currently, there are many groups working together to prevent, screen for, and
treat ROP (45). These include a number of USAID-funded programs in
collaboration with Helen Keller International, Orbis International, the
Retinopathy of Prematurity Network (ROPNET), the Aravind Eye Health
System, and the Global Education Network for Retinopathy of Prematurity
(GEN-ROP), as well as other global and regional programs (45). Notably,
telemedicine has emerged to play a vital role in promoting ROP screenings
(45).
Telemedicine and Tele-Education

Increasing awareness of ROP as a preventable disease and helping to
establish local research in disease trends will both help address the rising
global epidemic of ROP. Experts from developed nations are conducting
international workshops and conferences to educate local clinicians and
improve ROP programs in middle- and low-income countries. Educational
initiatives and aid to these countries also already occur with organizations,
such as ORBIS International (47,48), the International Pediatric
Ophthalmology and Strabismus Council (IPOSC), and Unite for Sight (49).
However, many initiatives may only provide immediate but unsustainable
help, for example, a one-time symposium or a surgical mission to help a
finite number of patients. However, the goal should be to offer a more
sustainable solution, such as training local clinicians to be competent in
screening and treating ROP.
With this initiative in mind, a potential solution to managing the global
burden of ROP is telemedicine, which is the use of information technology to
support health care between entities separated from each other (50).
Telemedicine has already shown promise in other fields. Telementoring has
been widely studied for minimally invasive procedures in general surgery,
neurosurgery, gynecology, and urology (51–53). These programs attempt to
address the dissemination of advanced procedures, such as laparoscopic
surgery, to hospitals usually unfamiliar or inexperienced with them. For
example, Antoniou et al. (54) found that telementored surgical techniques,
such as laparoscopic cholecystectomies performed by trainees with guidance
from remote expert surgeons, were feasible and safely performed. In
radiology, which is heavily reliant on imaging, teleradiology has proven
benefits for consultation from remote experts, establishment of long-distance
education systems, and the creation of digital teaching files (55,56).
Synchronous telemedicine via real-time videoconferencing could allow for
sustainable educational curricula between experts and nonexperts, rather than
a one-time experience as in the past. Tele-education has already been utilized
by many medical schools and training programs for videoconferencing
lectures and has provided a feasible method to disseminate information to
satellite campuses (57,58).
Tele-education has the potential to address some of the challenges in
worldwide ROP education. Widespread retinal imaging of ROP would allow
for the creation of digital libraries of images as teaching files to assess
competency of local ROP screeners in international locations as well as of
ophthalmology residents and fellows in the United States. The GEN-ROP in
collaboration with the Imaging and Informatics for ROP (i-ROP) consortium
recently designed and implemented a tele-education system for
ophthalmologists-in-training to improve their competency in the diagnosis of
ROP (59). The web-based platform allows for users from around the world to
access the teaching program that consists of a pretest, posttest, and training
modules. Trainees utilizing the tele-education system work through 65
clinical cases consisting of retinal images from 36 infants with varying
degrees of ROP (59). The tele-education program was piloted by
ophthalmologists-in-training in the United States and Mexico; using the
program improved abilities of trainees to identify each of the defining
features of ROP, including zone, stage, and plus disease, in order to improve
the diagnostic accuracy of ROP (59,60). Tele-education can be implemented
in ophthalmology training programs to supplement resident and fellow
experiences with ROP screening by providing standardized training material,
practice cases, and feedback. Likewise, health care workers participating in
ROP telemedicine programs can receive training through this program (59).
In parallel to these educational initiatives, new paradigms in computer-
facilitated image analysis of ROP are being developed to improve diagnostic
accuracy among clinicians. Multiple artificial intelligence (AI) algorithms
have been developed for diabetic retinopathy screening, and some have
recently been approved for clinical use (61–64). A series of studies have
demonstrated the efficacy of machine learning in automatizing ROP
diagnosis. Ataer-Cansizoglu et al. developed the i-ROP system, and AI was
applied on a dataset of 77 wide-angle retinal images of infants and compared
to clinical judgment in ophthalmoscopic examinations or three manual
gradings of each image. The i-ROP system achieved 95% accuracy in
classifying ROP versus no ROP, which was comparable to the performance
of three individual experts (96%, 94%, 92% accuracy) (65,66). More recent
studies through the i-ROP consortium have shown that a deep convolutional
neural network detects the presence of plus disease with 93% sensitivity and
94% specificity and accurately selects the category of ROP by assessing
criteria that guide the diagnosis (67,68). The utility of these computer-
facilitated image analysis programs is a pivotal topic in the development of
remote ROP screening programs. Telemedicine also provides potential
benefits to the many logistical challenges of ROP care worldwide. In store-
and-forward programs already implemented in the United States and abroad,
fundus photography of infants can be collected by trained neonatal staff for
subsequent evaluation by a remote ophthalmologist (69,70). This strategy
could improve accessibility to ROP experts in rural areas of the United States
and other international areas usually void of such specialized care. There
would also be fewer travel constraints on ophthalmologists, who not only
usually work in outpatient settings distinct from neonatal units but also may
be responsible for providing ROP coverage for multiple hospitals. Logistical
challenges of coordinating bedside exams with neonatal staff, such as timing
of dilation and presence of a nurse for examination, would also be decreased.
Serial retinal imaging, in contrast to retinal drawings, may provide a faster
and objective way of documenting disease findings and progression (71).
Due to these potential benefits, many clinicians and investigators are
exploring telemedicine in ROP. Many studies have explored the accuracy of
image-based ROP diagnosis compared to reference standard of dilated
ophthalmoscopy. Results have shown good sensitivities and specificities for
the detection of ROP via telemedicine (72–74). There is evidence that
indirect ophthalmoscopy and digital imaging are comparable screening
methods particularly when assessing for advanced types of ROP (75–77). The
e-ROP study is a North American–based project supported by the National
Eye Institute that aimed to evaluate the efficacy and accuracy of remote
reading of retinal images for ROP (78). In a study conducted by Quinn and
colleagues under the umbrella of e-ROP, health care workers were trained to
capture fundus images, and additional staff were trained to remotely read the
images to screen for ROP. When compared to expert ophthalmologists, these
workers identified clinically significant ROP with 90% sensitivity and 87%
specificity (78). In the context of telemedicine, incorporation of fundoscopic
digital imaging, coupled by expansion of the workforce, would allow for
greater populations to be served (77). Additionally, in a cost comparison of
telemedicine versus ophthalmoscopy for ROP management, Jackson et al.
(79) found an advantage to telemedicine, which costs about $3,000 per
quality-adjusted life year (QALY), while traditional management with serial
ophthalmoscopy costs roughly $5,500 per QALY (79).
INTRAVITREAL ANTI-VEGF
THERAPY: GLOBAL IMPLICATIONS
In addition to emerging telemedicine programs, the encouraging results with
anti-VEGF agents for ROP suggest a role for the therapy on the global scale.
As mentioned previously, developing nations are now dealing with increasing
numbers of children with ROP, and clinicians in these countries may lack the
experience, support staff, equipment, or facilities to adequately treat with
laser photocoagulation. Intravitreal anti-VEGF therapy may have advantages
over laser, including ability to administer through opaque media or
significant pupillary neovascularization, ease of use with less time needed for
treatment, and potentially cheaper cost.
Despite its exciting potential, intravitreal anti-VEGF therapy is still not
ready to completely replace laser photocoagulation, which has historically
been the gold standard of treatment. The biggest concerns and unanswered
questions regard the potential systemic and local adverse events of the
antiangiogenic agent. Despite local administration of bevacizumab, it has
been reported to enter the systemic circulation after intravitreal injection in
animal models and has been shown to exhibit a bilateral response after
unilateral administration in adults (80–82). Sato et al. (83) reported levels of
serum bevacizumab, with resultant reduced serum VEGF, in preterm infants
treated with intravitreal bevacizumab. Similarly, Wu and colleagues
identified the persistence of bevacizumab in the circulation of neonates for 8
weeks. The consequential reduction in VEGF levels could carry various
implications, including disruption of normal physiologic development (84).
Although VEGF mediates and drives the ROP disease process, it is crucial
for the survival and growth of many organs, such as the brain, lungs, and
kidneys (85–87). Morin and colleagues identified that neonates treated for
ROP with bevacizumab were at three times greater risk for developing
significant neurologic dysfunction, such as cerebral palsy and loss of hearing
and vision, compared to their counterparts who underwent laser
photocoagulation (88). In the BEAT-ROP study, there was a statistically
nonsignificant increase in mortality in babies treated with bevacizumab
(6.6%) versus those treated conventionally (2.6%) (7). Lepore et al. reported
4-year follow-up data of 21 infants with type 1 ROP who had one eye treated
with 0.5 mg bevacizumab and reported that fluorescein angiography revealed
that all the eyes treated with anti-VEGF therapy had evidence of vascular
irregularities at the peripheral retina and posterior pole (89). Currently, the
functional value to these structural findings is unclear. Additionally, some
circumstantial evidence exists implicating anti-VEGF agents in
thromboembolic events (90). Therefore, there is legitimate concern about the
potential systemic effects and long-term sequelae of an antiangiogenic agent
used in developing neonates. Moreover, despite the “ease” of intravitreal
injections and the potential to train nonophthalmologists in this technique,
local complications occur. Endophthalmitis, retinal hemorrhage, cataract, and
retinal detachment are possible even in trained hands, and specific knowledge
of eye anatomy in a premature baby is critical (91).
Other questions remain regarding the dosing, timing, and frequency of
administration of anti-VEGF agents; management of treatment failure; and
which anti-VEGF agent is most ideal for use in ROP. Initial reports used 1.25
mg bevacizumab, but subsequent studies demonstrated effectiveness with
0.625 mg or even lower dosing (7,17). Recently, a phase 1 dose de-escalation
study found that 0.031 mg bevacizumab, or 5% of the dose used in the
BEAT-ROP trial, was effective for treatment (20). This issue is important
since current reports do not seem to vary dosing based on infant weight,
gestational age, postmenstrual age (PMA), level of supplemental oxygen use,
or nutritional status. Future studies will help answer these important
questions and establish the lowest effective dose that can be used to
neutralize the excess VEGF in the eye without risk to the developing eye or
to systemic organs. The timing of injection seems to be important as well.
Treatment too early may promote significant delay of normal vascularization,
whereas treatment too late may promote retinal detachment in eyes with
preexisting traction from neovascularization (92). Additionally, even though
the response to anti-VEGF agents can be dramatic, with regression of severe
ROP occurring 24 to 48 hours after a single injection, more studies will need
to determine the utility and safety of multiple injections.
Lastly, concerns about disease progression and frequency of follow-up
exist. Vanderveen et al. identified the following trends: (a) recurrence of ROP
is greater following treatment with anti-VEGF compared to laser, which
beckons the need for patients to undergo further interventions; and (b)
revascularization of the retina is hindered with anti-VEGF injections, which
demands more extensive surveillance of patients following treatment (93).
Especially problematic is that disease recurrence following anti-VEGF
treatment seems to occur later in the course of disease than with laser therapy
(even 1 year), which can lead to retinal detachment in eyes that would
normally not have received follow-up after laser treatment based on
conventional standards. For example, Hu et al. reported a review of 17 eyes
with recurrence of ROP after initial bevacizumab treatment as initial
monotherapy. Mean age at treatment-requiring recurrence was slightly <50
weeks PMA, with a maximum at 69 weeks PMA. Of the eyes studied, almost
30% progressed to retinal detachment (94). Given these emerging data,
babies treated with intravitreal anti-VEGF therapy will need longer and likely
more frequent follow-up, which may be problematic in developing countries
with limited resources. See also Chapter 53.
CONCLUSION
Tremendous advances have occurred in ROP management over the past
decades. However, ROP remains a significant clinical challenge in highly
developed countries and a leading cause of preventable pediatric blindness
worldwide. More studies regarding the future ROP workforce in the Unites
States and abroad are needed to ensure an adequate number of high-quality
ophthalmologists capable of managing the rising burden of ROP in middle-
and high-income countries. Additionally, further efforts on spreading
awareness and education of ROP via emerging technologies such as tele-
education need to continue to help address the current “third epidemic” of
ROP globally. The use of pharmacotherapy has profound potential in this
respect as well, but much research is still needed on its potential systemic and
long-term effects before it becomes gold standard.
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52
Clinical Trials and Management of
Julia P. Shulman, and M. Elizabeth Hartnett
PREVALENCE
Blindness from retinopathy of prematurity (ROP) varies from country to
country and depends in large part on the level of socioeconomic development
and, therefore, likelihood that premature infants survive long enough to
develop ROP. Based on 1993 World Health Organization estimates, over
50,000 children are blind from ROP worldwide with many more who have
unilateral loss of vision or visual impairment (1). In the United States, the
incidence of ROP was estimated in the Early Treatment for Retinopathy of
Prematurity (ETROP) study to be 68% among infants of <1,251 g, similar to
the results in the Cryotherapy for Retinopathy of Prematurity (CRYO-ROP)
study conducted 15 years earlier (2,3). This has been attributed to advances in
neonatal care that allow for improved survival of the extremely low birth
weight infants. In more recent studies, the incidence of ROP has been
reported to range from approximately 0.12% to 0.2% corresponding to 1 in
every 820 live births to as frequent as 1 in every 511 births (4,5). The
incidence of ROP increases with decreasing birth weight and has been
reported to be 33.2% for infants with birth weight <1,000 g, which is much
lower than in the ETROP and CRYO-ROP studies (3–5). This difference may
represent a true decrease in the incidence of disease or simply reflect
differences in the methodology used in the studies and whether they were
prospective or retrospective.
Although advances in neonatal care and ROP management have led to
better visual outcomes than those reported in earlier studies, ROP remains a
leading cause of childhood blindness necessitating further research to achieve
effective prevention and treatment. As larger numbers of extremely low birth
weight infants survive, effective management of ROP will play a greater role
in the quality of life of these patients (3).
This chapter will discuss major clinical trials that frame the
understanding, classification, and management of ROP.
Oxygen
High Oxygen
Retrolental fibroplasia (RLF) (Figure 52-1) was first recognized in the 1940s
(6) and likely represented the cicatricial form of the most severe stage of
ROP, stage 5 ROP. The white pupil found in the eyes of a premature infant,
approximately 32 weeks’ gestational age (6), represented a retrolental
fibrovascular membrane behind which existed a total retinal detachment.
Probably the first comparative administered study was performed by Arnall
Patz testing high oxygen administered to all premature infants versus only
those with respiratory distress (7). Scientists were yet to publish their
thoughts on high oxygen in RLF (8). Following strong evidence from this
study for his hypothesis that high oxygen caused RLF, Dr. Patz and Dr.
Kinsey performed a multicenter study to gain greater evidence (9). Once high
and uncontrolled oxygen delivery was recognized as a cause of RLF, efforts
to regulate oxygen were developed and initiated, and, as a result, RLF was
nearly completely eradicated.
FIGURE 52-1 Eye of infant with retrolental fibrovascular
membrane from stage 5 ROP. Total retinal detachment,
determined by ultrasonogram, exists posterior to the white
retrolental membrane.
However, ROP continues to cause significant visual morbidity with increased

survival of young gestational age and very low birth weight premature infants
and with emerging neonatal care throughout the world. We now recognize
that multiple factors are involved in the development of ROP, and the role of
supplemental oxygen appears more complex (Chapter 40). In addition, the
recognition of earlier stages of ROP has both facilitated investigation into the
causes of ROP and led to earlier treatment to reduce the risk of retinal
detachment and blindness (Tables 52-1 to 52-3).
TABLE 52-1 Major clinical trials in retinopathy of

prematurity
a240 had bilateral disease and were treated with cryotherapy (cryo) in one eye and observed in the
fellow eye. In 51 asymmetric eyes, treatment was randomly assigned to either cryo or observation.
bFor the 1- and 3½-year studies, Teller acuity vision was determined. For the 5½-, 10-, and 15-year
outcome studies, testing using the ETDRS visual acuity chart was also performed.
cOf the 401 infants randomized in the high-risk prethreshold group, 317 had bilateral disease and were
treated with either early ablative therapy within 48 hours of the first diagnosis of high-risk prethreshold
or conventionally. In the remaining 88 asymmetric cases, the eye was randomized to either early
treatment or CM. CRYO-ROP, Cryotherapy for Retinopathy of Prematurity; Light-ROP, Light
Reduction on Retinopathy of Prematurity; BW, birth weight; ROP, retinopathy of prematurity; MO,
month; GA, gestational age; STOP-ROP, Supplemental Therapeutic Oxygen for Prethreshold
Retinopathy of Prematurity; NICU, neonatal intensive care unit; RA, room air; Con, conventional
oxygen; S, supplemental oxygen; PMA, postmenstrual age; HOPE-ROP, High-Oxygen Percentage
Retinopathy of Prematurity; ETROP, Early Treatment for Retinopathy of Prematurity; BEAT-ROP,
Bevacizumab Eliminates the Angiogenic Threat of Retinopathy of Prematurity; CM, conventional
management; BCVA, best-corrected visual acuity; ETDRS, Early Treatment Diabetic Retinopathy
Study visual acuity; PUFAs, polyunsaturated fatty acids.
TABLE 52-2 Later studies performed on original

CRYO-ROP cohort
RD, retinal detachment; ROP, retinopathy of prematurity; CRYO-ROP, Cryotherapy for Retinopathy of
Prematurity; Light-ROP, Light Reduction on Retinopathy of Prematurity; ETROP, Early Treatment for
Retinopathy of Prematurity; OU, both eyes; PMA, postmenstrual age (gestational age plus chronologic
age in weeks).
TABLE 52-3 Later studies performed on ETROP-

ROP cohort
ETROP, Early Treatment for Retinopathy of Prematurity; ROP, retinopathy of prematurity; ET, early
treatment; CM, conventional management; OU, both eyes; RD, retinal detachment.
Oxygen Fluctuations
Oxygen fluctuations, as measured by partial pressure of dissolved arterial
oxygen (PaO2), have been associated in animal studies and clinical trials with
the development of ROP (49–51). Studies by Saito, Cunningham, and Penn
demonstrated increased incidence of ROP and severe ROP in infants exposed
to oxygen fluctuation at the same gestational age and birth weight (49–51).
Oxygen fluctuations were measured either transcutaneously (49) or by
measuring the PaO2 in an arterial blood gas (50,51). Of note, transcutaneous
PO2 measurements estimate PaO2; however, due to the S shape of the
oxygen–hemoglobin dissociation curve, increases in PaO2—even at very high
levels—have little impact on saturation when oxygen saturations >90% in
healthy infants (52). So, dangerously high PaO2 may not be reflected in the
oxygen saturation level. In sick neonates, who are prone to episodes of hypo-
and hyperoxemia, transcutaneous PaO2 becomes a less accurate estimate of
arterial PaO2 (53,54).
Animal studies have demonstrated that PAO2 fluctuations in rat pups
played a key role in the development of abnormal retinal vascularization. In
addition, fluctuations in oxygen were more damaging than constant exposure
even if the total oxygen administered was less (50).
Clinical Trials of Oxygen

STOP-ROP (Supplemental Therapeutic Oxygen for Prethreshold Retinopathy
of Prematurity), a large multicenter trial, was designed to test supplemental
oxygen as a strategy to prevent threshold ROP (Table 52-4) and did not show
additional progression of prethreshold ROP at high pulse oximetry values of
96% to 99%, but also did not decrease the need for peripheral retinal ablative
procedures (33). In a post hoc analysis, the study showed reduced progression
to threshold ROP in a subgroup of infants treated with supplemental oxygen
(to achieve SaO2 of 96% to 99%) with prethreshold ROP without plus
disease.
TABLE 52-4 Results from STOP-ROP study

aThe target alpha value was 0.025, deliberately conservative to allow for preplanned sequential testing.
To balance the benefits and risks of supplemental oxygen, a number-needed-to-treat analysis was
performed. This analysis compares the number of infants with prethreshold ROP who would have to be
treated with supplemental oxygen to spare an infant from retinal surgery to the number of infants
treated with supplemental oxygen to be expected to cause one episode of pulmonary exacerbation. The
number requiring supplemental oxygen to prevent an adverse retinal event was 13.2 and to cause a
pulmonary exacerbation was 13.7. Pulmonary events were not lethal, although they did prolong
hospitalizations (33,55).
bThe proportion of infants who experienced any one or more of these adverse pulmonary events by 3
months corrected age was 57% (supplemental) versus 46% (conventional), P = 0.005, and was
unaffected after adjusting for baseline covariates of race, ROP severity, gestational age, and pulmonary
status.
STOP-ROP, Supplemental Therapeutic Oxygen for Prethreshold Retinopathy of Prematurity; ROP,
Retinopathy of Prematurity. NS, not significant; PMA, postmenstrual age (gestational age plus
chronologic age in weeks).
The SUPPORT study (Surfactant, Positive Pressure, and Pulse Oximetry

Randomized Trial) compared target ranges of oxygen saturation of 85% to
89% versus 91% to 95% in 1,316 infants with regard to severe ROP
(threshold disease, need for surgery, or use of bevacizumab), death before
discharge from the hospital, or both. The rates of severe retinopathy or death
did not differ significantly between the low and high oxygen saturation
groups, but death before discharge occurred more frequently in the low
oxygen group; severe retinopathy among survivors occurred less often in the
low oxygen group (56). Similar findings were found in the Benefits of
Oxygen Saturation Targeting Study II (BOOST-II) in the United Kingdom
and Australia, in which similar oxygen saturation targets as SUPPORT were
used to test infants born at younger gestational ages than 28 weeks (57).
A meta-analysis of recent literature revealed that low oxygen saturation
reduces severe ROP by 50%; the relative risk in favor of low oxygen
saturation was 0.42 (95% CI, 0.34–0.51) for severe ROP and 1.12 (95% CI,
0.86–1.45) for mortality (58). This analysis included the STOP-ROP cohort
and evaluated a total of 10 studies.
Oxygen saturation as measured by pulse oximetry has been the main
measure of hyperoxemia, hypercarbia, and low pH (as a measure in the first 3
postnatal days) but has also been shown to be associated with more severe
ROP (59).
These studies demonstrate that the effect of oxygen on the development
and progression of ROP is not as straightforward as whether high or low
oxygen is delivered to premature infants who develop ROP.
Light
Animal Studies
Light was proposed as a causative factor in ROP because light leads to a
release of free radicals that can cause oxidative damage, death of endothelial
cells, and release of angiogenic compounds. Expression of tumor necrosis
factor-α and vascular endothelial growth factor (VEGF) during retinal
neovascularization can be initiated by lipid hydroperoxides (60). However,
the retina is most metabolically active in the dark; thus, the release of
oxidative compounds conceivably might be greater than that released when
the retina is illuminated.
Animal studies have demonstrated that embryonic hyaloid vasculature
regresses by a light-mediated pathway. Pregnant mice dams placed in the
dark at late gestation with pups raised in the dark until 8 days postpartum
showed persistent hyaloid vessels (61). Investigators demonstrated that light
transmitted through a dam’s abdomen has an effect on retinal vascular
development in the pup early in gestation, corresponding to the first trimester
in humans. They identified a fetal light response pathway in mice that
requires melanopsin to regulate retinal angiogenesis (see also Chapter 2).
Dark rearing of mice from late gestation resulted in retinal hypoxia with an
increase in VEGF expression (61,62). These data suggest that insufficient
exposure to light in early gestation may result in the development of severe
ROP (62).
Brain-derived neurotrophic factor (BDNF) plays a role in ganglion cell
maturation and is reduced in mice reared in the dark (63).
Clinical Trials
The Light Reduction in Retinopathy of Prematurity (Light-ROP) study (Table
52-1) was designed to test the effect of reduced light to the preterm infant’s
retinas on the development of ROP. Infants were randomized to either a
group in which infants wore goggles that reduced ultraviolet light exposure
within 24 hours of birth or a group in which infants did not. There was no
evidence that reduced light exposure had an effect on the incidence of ROP
or the severity of ROP when it developed (32).
Based on the animal studies of Lang, Yang et al. evaluated the average
day length for 90 days from estimated date of conception in 684 eyes of 343
premature infants. Each additional hour of average day length was found to
decrease the likelihood of ROP by 28% (62).
Genetics
Studies have reported that ROP occurs in Caucasians more than in African
Americans (64) or Asians (65). A small study reported that native Alaskans
were more susceptible to ROP compared to nonnative Alaskans (66). A study
of monozygotic and dizygotic twins supported the concept genetic
predisposition in ROP (67). Although race cannot be addressed using animal
models, different strains of rats were shown to develop different degrees of
severity of retinopathy under the same conditions (68).
Genetic variants in the wnt signaling pathway genes including NDP,
FZD4, and TSPAN12, which are known to cause familial exudative
vitreoretinopathy (FEVR), have been reported in association with severe
ROP (69). In addition, variants in VEGF, endothelial PAS domain–
containing protein 1 (EPAS 1) and superoxide dismutase (SOD), have been
reported in association with any ROP. Variants in BDNF have also been
associated with severe ROP in extremely low birth weight infants (70), and
clinical studies reported reduced BDNF in extremely low birth weight infants
who developed ROP (71).
The association of ROP with multiple gestation and use of assisted
reproductive technology suggests a genetic link (72). However, ROP is
known to be affected by external factors such as oxidative stress, nutrition,
and others suggesting that epigenetic modifications by perinatal factors play a
role in the ultimate expressed phenotype of ROP. Epigenetic factors may also
help explain the variability in ROP severity among premature infants (73). In
addition, many reports have limited number of infants enrolled and
differences in gestational ages or birth weights of premature infants.
Extremely premature infants may have lacked the genetic variants that
predisposed less premature infants to develop ROP.
Worldwide Impact
ROP is a growing cause of blindness, especially in middle-income
developing countries where neonatal care is now emerging (see also Chapter
50).
Pathophysiology
Two Phases of ROP in the Past (Historical Context)

In models of oxygen-induced retinopathy (OIR) in the mouse, rat, and beagle,
the pathogenesis of ROP appears to be divided into two phases (see also
Chapter 40). The initial phase is characterized by compromise and death of
the normal vascular growth due to the relative hyperoxemia of the
extrauterine environment and delay in normal retinal vascular growth due to
fluctuations in oxygenation (74). Other factors, including neural guidance
molecules, are involved (75). As the metabolic demands of the developing
retina increase but the infant’s circulation and lungs have not developed
sufficiently to maintain less variable oxygenation, and the infant is brought
out of supplemental oxygen, there is a relative retinal hypoxia that triggers
the release of angiogenic factors including VEGF.
In individual preterm infants, unlike experimental OIR models, precise
phases do not occur. OIR phases can be related to human ROP as follows:
Phase I represents stage 1 to 2 ROP and Phase II represents stage 3 ROP with
plus disease (severe or treatment-warranted ROP). Neither the rat nor mouse
OIR model has stages 4 or 5 ROP, in which the retina is detached (see
Chapters 40 and 41). The beagle model develops retinal folds, which
approximate some of the pathologic findings in stage 4 ROP (73).
VEGF Signaling Disorder Developmental

Angiogenesis Causes Intravitreal Neovascularization
and Delays Physiologic Retinal Vascular Development
Vascular Endothelial Growth Factor

VEGF is essential for the development and growth of blood vessels. Down-
regulation of VEGF secondary to hyperoxia is hypothesized to play a role in
the cessation of normal retinal blood vessel growth seen in premature infants
(76). In the proliferative phase of the disease, hypoxia and reactive oxygen
up-regulate VEGF expression (50,77). VEGF signaling through its receptors
leads to abnormal extraretinal vessel growth (74,78,79). Anti-VEGF drugs
may be a promising new avenue of therapy in ROP; however, late
recurrences, persistent avascular retina, unknown optimal regimens, and
unknown systemic effects remain a barrier to their use (see also Chapter 53).
Postnatal Growth
Insulin-Like Growth Factor 1

The lack of IGF-1 has been found to be associated with reduced retinal
vascularization despite the presence of VEGF in animal models (80). IGF-1
levels are inversely correlated with the severity of ROP, the basis for the
WIN-ROP (81) and other protocols that associate infant growth and severity
of ROP (see Chapter 39); in addition, IGF-1 promotes VEGF-induced
neovascularization through stimulation of the MAPK pathway (82).
Oxidative Stress
Vitamin E
Oxidative stress has been linked to ROP through a number of mechanisms
related to oxygen delivery to retinal tissue in the premature infant. The retina
is believed to be susceptible to oxidative damage because of (a) its abundance
of polyunsaturated fatty acids, the double bonds of which are vulnerable to
peroxidation reactions (83); (b) its high metabolic rate and rapid rate of
oxygen consumption (84); and (c) through its vulnerability to light toxicity.
In addition, the premature infant has an inability to induce oxidative
scavenging mechanisms during times of oxygen stress (85).
A number of early studies tested the use of antioxidants, such as vitamin
E, to reduce ROP in infants. A meta-analysis of six of these studies (36)
found a 52% overall reduction in the incidence of intravitreous
neovascularization in ROP when infants were given vitamin E; however, the
study was limited by the inclusion of only six trials in the meta-analysis
(Table 52-1). Vitamin E administration was accompanied by lower incidence
of ROP in a retrospective study of 402 infants (86). Sepsis or late-onset
necrotizing enterocolitis reported in earlier vitamin E studies did not occur in
a later study performed by the same group and was attributed to improved
immunologic function with ongoing development of the preterm infant (87).
Vitamin E therapy has also been reported in association with a greater
incidence of intraventricular hemorrhage in the premature infant (17). A
Cochrane review in 2003 demonstrated that vitamin E supplementation
reduced the risk of intracranial hemorrhage and increased the risk of sepsis
but reduced the risk of severe ROP and blindness (88). The review did not
support the routine use of vitamin E supplementation.
Lutein
Lutein may reduce oxidative stress and prove a source of macular pigment.
However, randomized trials have shown conflicting results in terms of
incidence and severity of ROP (89,90).
Neurovascular Effects
Sapieha et al. have demonstrated that retinal ganglion cells sense ischemic
stress and regulate production of various antiangiogenic factors (75).
However, it remains early for translation of the findings into clinical
management.
There are no symptoms of acute ROP, nor can a specific visual behavior in
the preterm newborn herald a concern for ROP. Therefore, effective
screening is essential for diagnosis.
Age of Onset
Most ROP manifests at about 32 weeks of postmenstrual age (PMA). PMA
equals gestational age plus age after birth in weeks (27,91). The definition of
“treatment-warranted ROP” has evolved over the years and differs between
the United States and Europe (92,93). CRYO-ROP defined treatment-
warranted ROP as threshold disease, in which there was a 50% risk of a poor
outcome (Table 52-5). Threshold ROP peaked at approximately 37 weeks of
PMA (19,33). The ETROP defined treatment-warranted ROP as a less severe
form, type 1 ROP, mainly because of poor outcomes in zone I threshold
ROP. Type 1 ROP was defined as a ≥15% risk of a poor outcome and peaked
in infants at approximately 35 weeks PMA (37). Eyes that rapidly progress to
prethreshold ROP have a greater risk of developing threshold ROP (19,29)
and will develop threshold disease at an earlier PMA. When infants achieve
45 weeks’ PMA without developing prethreshold ROP, they have a low risk
of developing treatment-warranted ROP or of having a poor outcome (27).
Infants treated with anti-VEGF agents can have an altered natural history
with recurrences of severe ROP reported beyond 1 year of age (91,94–96).
TABLE 52-5 Unfavorable retinal outcomes
ROP, retinopathy of prematurity.

Course of Retinopathy of Prematurity
Most ROP regresses; however, the disease typically progresses before
regressing. Rapid progression from type 2 prethreshold ROP to type 1 ROP
occurred in 22% of infants with half of these occurring within a week in the
ETROP study (42). In about 6% of infants born weighing <1,251 g, threshold
ROP developed (11), and unfavorable structural outcomes occurred in 52%
of untreated eyes and 30% of treated eyes with unfavorable visual outcomes
in 64% of untreated and 45% of treated eyes at 15 years (12). Early treatment
for type 1 ROP reduced poor visual and functional outcomes (97).
Definitions of Treatment-Warranted Retinopathy of

Prematurity
Based on the CRYO-ROP study, treatment-warranted ROP was defined as
threshold ROP at which the risk of an unfavorable anatomic outcome
approached 50% (10,11). Threshold ROP was diagnosed by the presence of
five contiguous or eight total clock hours of stage 3 ROP in zone I or II with
plus disease (Figure 52-2). Because a high frequency (87%) of zone I eyes
had an unfavorable outcome in the CRYO-ROP study, later studies such as
STOP-ROP (33) and High-Oxygen-Percentage Retinopathy of Prematurity
(HOPE-ROP) (34) recognized the need to closely monitor threshold ROP in
zone I eyes (Table 52-6). In the ETROP trial, prethreshold ROP was further
subdivided based on the risk of an unfavorable outcome determined by a risk
analysis program based on natural history data from the CRYO-ROP study,
the RM-ROP2. Eyes with high risk were defined as having ≥15% risk,
whereas eyes at low risk had <15% risk of an unfavorable outcome. An
unfavorable outcome was primarily based on one of four categories of visual
function determined with Teller Acuity Cards (37). Secondary outcomes were
based on unfavorable structure outcomes including a retinal fold or
detachment involving the macula or a retrolental opacity blocking the visual
axis (Table 52-5). Type 1 ROP was defined as zone I, any stage ROP with
plus disease; zone I, stage 3 ROP without plus disease; and zone II, stage 2 or
3 with at least two quadrants of plus disease based on the standard
photograph from the CRYO-ROP study (see Table 52-6) (97). Treatment of
the peripheral retina with laser for type 1 ROP was found to decrease
unfavorable structural outcomes from 15.6% to 9.0% at 9 months (37,97) and
from 15.2% to 8.9% at 6-year follow-up (40,41). Although a statistically
significant benefit in Teller grated acuity was seen at 9 months in the early
treatment group compared to conventional management, the difference was
not statistically significant by 6 years of age (41). However, type 1 eyes
receiving early treatment had a significantly lower rate of unfavorable
outcomes when compared to eyes treated conventionally (41). Based on the
ETROP results, treatment for type 2 ROP (defined as zone I, stages 1 and 2
without plus disease, or zone II, stage 3 without plus disease) was
recommended only if it progressed to type 1 ROP or threshold (97).
FIGURE 52-2 Artist rendition of threshold disease

defined in the multicenter trial of CRYO-ROP study as
five contiguous or eight total clock hours of stage 3+ in
zone I or II.
TABLE 52-6 Definitions of prethreshold threshold

ROP and treatment-warranted ROP
aBased on standard photograph used in CRYO-ROP studies; requires two quadrants of plus disease
(similar to criteria used in STOP-ROP study).
bThe definition of threshold ROP was different for the CRYO-ROP and STOP-ROP studies: (a) For
STOP-ROP, plus disease was defined as two quadrants of posterior pole dilation/tortuosity, whereas in
CRYO-ROP it was defined as about four quadrants of posterior pole dilation/tortuosity, and (b) for
STOP-ROP, a less stringent definition of threshold disease was given for zone I eyes that included any
ROP with plus disease or any stage 3 with or without plus disease.
ROP, retinopathy of prematurity; ETROP, Early Treatment for Retinopathy of Prematurity.
In the CARE-ROP and RAINBOW trials testing anti-VEGF agents in ROP,

the definition of treatment-warranted ROP was introduced and each trial
defined it slightly differently (Table 52-6).
These definitions provide guidelines for managing infants with acute
ROP and are summarized in Table 52-6 (92,93).
DIAGNOSIS AND DIAGNOSTIC

STUDIES
Indirect Ophthalmoscopy
The diagnosis of ROP has also evolved. The gold standard of using indirect
ophthalmoscopy is important to determine treatment-warranted ROP, to
determine when screening examinations can be stopped, and when to perform
the next screening examination for ROP. Images obtained with contact
cameras can be used to document and provide additional information to aid in
management of infants seen with indirect ophthalmoscopy. In addition,
screening using telemedicine approaches has been advocated because of
insufficient numbers of skilled ophthalmologists to perform indirect
ophthalmoscopy to meet the guidelines for optimal screening of premature
infants. Camera-based imaging can be considered for referral-warranted ROP
and is based on identifying one of three features of ROP: any ROP in zone I,
any stage 3 ROP, plus disease (98). However, the e-ROP multicenter study
found sufficient concern of missing infants with treatment-warranted ROP
suggesting that use of telemedicine alone was not sufficient to prevent
missing any eye with treatment-warranted ROP (99).
Indirect ophthalmoscopy can be performed at the bedside or clinic. Once
the infant’s eyes are fully dilated and the infant is swaddled, a drop of topical
anesthetic is instilled into one eye and a lid speculum is inserted. Initially no
scleral depression is performed and the retinal vessels are assessed for
vascular dilation and tortuosity in each of the four quadrants starting around
the optic nerve (Figure 52-3) and moving to the periphery (Figure 52-4).
Scleral depression can be useful to determine the zone of intraretinal
vascularization. Determination of zone I is made either by estimating the
circle surrounding the optic nerve having a radius of twice the distance from
the optic nerve to the fovea or by viewing the image of the fundus through a
28-diopter lens with nasal edge of the optic nerve at one edge of the field of
view; the limit of zone I is at the temporal field of view (100). If the retinal
vasculature does not extend beyond the image size of the 28-diopter lens,
then vessels are defined as being within zone I, but it is recognized that there
are differences between zone I in one clock hour versus all clock hours
falling within zone I. Zone II is retinal vascularization beyond zone I but not
within one disc diameter of the nasal ora serrata at the two clock hours
abutting the nasal horizontal meridian, for example, 9 o’clock OS or 3
o’clock OD (100). Zone III is the remaining avascular temporal area of the
retina (Figure 52-5).
FIGURE 52-3 Moderate plus disease present posteriorly
as dilated, tortuous vessels around the optic nerve.
FIGURE 52-4 Peripheral plus disease noted by dilated
and tortuous retinal arterioles and veins in the peripheral
retina extending into the ridge.
FIGURE 52-5 Circular zones I (A), II (B), and III (C)
drawn onto schematic of fundus. Zone I is the least mature
and encompasses the smallest area of the retina with
developed retinal vasculature.
There can be difficulties examining the lightly pigmented, blonde fundus or

deeply pigmented fundus. In the blonde fundus (Figure 52-6), the choroidal
vessels are apparent, and the examiner may mistake these for retinal vessels.
In the deeply pigmented fundus (Figure 52-7), it can be difficult to visualize
the retinal vessels against the dark, pigmented background. In both of these
cases, the 20-diopter lens can improve diagnostic ability.
FIGURE 52-6 Blonde fundus permits visualization of
deeper choroidal vessels (blue box) and in avascular retina
can be mistaken for retinal vessels.
FIGURE 52-7 Pigmented fundus can reduce the ability to
distinguish retinal vessels in the periphery, noted
especially superiorly temporally in left of image.
Classically, at the junction of vascularized and avascular retina, the examiner

determines the stage of ROP. The clock-hour extent of each stage is not used
currently to determine treatment-warranted ROP (13,37) but are still
important in determining eyes at risk of progressive stage 4 ROP after laser
treatment (101).
The same procedure is performed in the fellow eye.
CONTACT CAMERA DIAGNOSIS

Portable contact cameras allow for photo-documentation of the fundus
examination after pupillary dilation as well as the performance of fluorescein
angiography in some models. The images can be obtained by a physician,
nurse, or medical assistant who have had training in the use of the camera.
The preparation for imaging with the camera is similar to that for an
examination. After instillation of anesthetic drops and placement of a lid
speculum, a coupling agent such as goniosol or artificial tear gel is used on
the surface of the cornea, tip of the camera lens, or both to obtain retinal
images.
Multicenter Studies (ICROP, e-ROP)
International Classification of Retinopathy of

Prematurity Characterizes Progression of Retinopathy
of Prematurity
The International Classification of ROP was developed to describe the levels
of severity of ROP based on several parameters: zone, stage, extent of stage,
and presence of plus disease (102). This classification was updated in 2005 to
reflect changes in the understanding of the disease since the initial
publication in 1984 (100). The zone of ROP refers to one of three areas that
best describes the retinal area that has apparent intraretinal vascularization
(Figure 52-5). Normally, in development, vascularization of the retina starts
posteriorly with early vascularization around the optic nerve disputed to be a
result of angioblast growth or vasculogenesis. From these early vessels,
further vascular proliferation and migration extend vascularization toward the
ora serrata.
The stage of ROP defines the severity of the ROP and in stages 1 to 3
describes clinical appearance of the retina at the junction of the vascularized
retina and the avascular area. ROP was described by zone of disease (see
above), stage, and plus disease. There are five stages. In stage 1, a line is
apparent (Figure 52-8). In stage 2, there is a ridge having obvious volume
(Figure 52-9). In stage 3 ROP, there is neovascularization growing onto the
vitreous at the ridge (Figure 52-10). Stage 4 ROP is partial retinal
detachment, 4A without macular involvement and 4B with macular
involvement (Figure 52-11). Stage 5 ROP is a total retinal detachment and is
described as closed if the retina is adherent to itself or open if it is not. In
some cases of stage 5 ROP, there is a peripheral attached avascular retina
causing the appearance of a peripheral retinal trough.
FIGURE 52-8 Stage 1 ROP seen as a discontinuous

white, tortuous line between vascularized (posterior) and
avascular (peripheral) retina temporally.
FIGURE 52-9 Stage 2 ROP with a prominent ridge at the
junction of the vascular and avascular retina.
FIGURE 52-10 Image of the eye treated with laser to
avascular retina (upper right of field). Note stage 3
neovascularization extending into vitreous. Some blood is
seen within the neovascular tissue.
FIGURE 52-11 Early stage 4A ROP with exudation
posterior to the ridge. The elevated retina extending
superiorly and inferiorly from area of exudation at 9
o’clock appears out of focus.
The extent of ROP refers to the number of clock hours of the highest stage.
Plus disease refers to dilation and tortuosity of the retinal arterioles and veins
in the posterior pole (Figure 52-3) and was based on a standard photograph
in multicenter trials (10,100). Pre-plus disease was defined as abnormal
dilation and tortuosity of the posterior pole vessels that was insufficiently
severe for the diagnosis of plus disease but was not normal and was defined
after the CRYO-ROP study (100). Aggressive posterior ROP (AP-ROP) is an
aggressive form of posterior retinopathy observed commonly in zone I or
posterior zone II with dilation and tortuosity of the arterioles and veins in all
four quadrants out of proportion to the peripheral retinopathy. AP-ROP does
not usually progress through the classic stages, extends circumferentially, and
can be accompanied by a circumferential vessel. Flat neovascularization at
the junction of the vascularized and nonvascularized retina is different in
appearance from zone II and stage 3 ROP, and this aggressive form of ROP
progresses to stage 5 if not promptly treated (100).
e-ROP
The use of telemedicine in the screening of ROP has been validated in a
multicenter trial from the NEI (e-ROP), which compared screening results
from wide-field retinal images obtained by non–physician certified ROP
imagers using a standard protocol with conventional retinal examinations by
trained ophthalmologists (99). For neonatal care centers utilizing remote ROP
screening, it is recommended that indirect ophthalmoscopy be performed at
least once by a qualified ophthalmologist prior to treatment of ROP or
discontinuation of acute phase screening (91).
The differential diagnosis of ROP includes FEVR, Norrie disease,
incontinentia pigmenti, congenital retinal fold, Toxocara canis infection, and
other causes of leukocoria (Table 52-7). A white pupil, referred to as
leukocoria, can occur from media opacities at any level from the cornea to
the retina. A history of prematurity and low birth weight are the most helpful
historical data in the differential diagnosis of ROP.
TABLE 52-7 Differential diagnosis of leukocoria
CMV, cytomegalovirus; PFV, persistent fetal vasculature; ROP, retinopathy of prematurity.

MANAGEMENT
Screening
The screening protocol at each neonatal intensive care unit (NICU) should be
based on published recommendations and preferences of the screening
ophthalmologists, neonatologists, and NICU nurses. All at-risk infants should
be identified and receive adequate dilated retinal evaluations at appropriate
times. The Joint Statement of the American Academy of Pediatrics, the
American Association for Pediatric Ophthalmology and Strabismus, and the
American Academy of Ophthalmology (updated in 2018) provided the
following guidelines for infants who should be screened for ROP: (a) all
infants born weighing ≤1,500 g and/or ≤30 weeks’ gestational age and (b)
infants born weighing between 1,500 and 2,000 g or a gestational age >30
weeks who experience an unstable clinical course (e.g., infants requiring
extensive oxygen supplementation or infants with hypotension requiring
inotropic support) at the discretion of the neonatologist or pediatrician. The
first examination should be performed prior to hospital discharge, at 4 weeks
after birth, or at 31 weeks’ PMA depending on the gestational age (91).
Examinations should be repeated periodically, for example, every 2
weeks if there is no ROP or with unequivocally regressing ROP, weekly with
any ROP, and more frequently with type 2 ROP or suspected presence of AP-
ROP. If intraretinal vascularization proceeds toward the ora serrata,
examinations are performed less frequently. Signs indicating that the risk of
visual loss from ROP is minimal include PMA of 45 weeks, intraretinal
vascularization into zone III without previous zone I or II ROP, and complete
intraretinal vascularization (91). When any of these signs is present,
screening can be discontinued and the focus of care changed to that of visual
rehabilitation. However, these findings are only for infants who have not
received anti-VEGF treatment (more on treatment and follow-up after anti-
VEGF treatment below).
Clinical Studies (WIN-ROP, G-ROP, etc.)

Several additional screening algorithms that take into account postnatal
weight gain such as WIN-ROP and CHOP-ROP have been developed
(103,104) (see also Chapter 39). The WIN-ROP algorithm correctly predicted
treatment-warranted ROP based on postnatal weight gain and serum insulin-
like growth factor 1 (IGF-1) and IGF binding protein 3 levels in 79 preterm
infants (103). The WIN-ROP algorithm was further validated with postnatal
weight gain measurements only in a U.S. cohort of 318 infants with 100%
sensitivity in identifying infants who developed severe ROP (105).
The Postnatal Growth and ROP (G-ROP) study was a multicenter
retrospective cohort study of infants in 30 North American Hospitals during
2006–2012. 7,483 infants were included in the analysis validating a postnatal
weight gain–based protocol (CHOP-ROP) regarding ability to predict Type 1
ROP (106). A cohort of 367 infants who participated in the Premature Infants
in Need of Transfusion (PINT-ROP) followed for weight gain demonstrated a
decrease in required examinations in infants with BW <1,000 g by 30% and
identified all but one infant who required laser treatment (104) (see also
Chapter 39).
TREATMENT
According to ETROP, once type 1 ROP develops, treatment should be
considered within 48 to 72 hours (91,107). Based on current guidelines,
treatment is laser to the avascular retina or anti-VEGF treatment for selected
cases (91).
Following laser, treated eyes are monitored 1 week later and then weekly
for regression of vascular activity and assurance that no fibrovascular
traction, repeat plus disease, recurrent neovascularization, or vitreous haze
occurs. These features are concerning for need for additional laser
(neovascularization or plus disease) or monitored for possible lens-sparing
vitrectomy (vitreous condensation, ridge thickening, and retinal elevation).
Following anti-VEGF, treated eyes are monitored often 1 day after
injection and then weekly. During follow-up, eyes are evaluated for
regression of vascular activity as defined by plus disease, or stage 3 ROP, and
progressive stage 4 ROP, or full vascularization. Progressive stage 4 ROP
manifests differently in eyes treated with laser compared to those treated with
anti-VEGF. In eyes treated with laser, most of the time progressive stage 4
ROP manifests as thickening of the ridge, vitreous condensation, or
organization at the ridge or optic nerve, plus disease, or vitreous hemorrhage,
particularly over the ridge. Following anti-VEGF treatment, these features
exist, but there can also be vitreous organization and condensation over the
area of the previous ridge especially in eyes with AP-ROP or condensation at
the optic nerve (Figure 52-12). Full vascularization of the retina to the ora
serrata should be confirmed in eyes treated with anti-VEGF since recurrences
even after 2 years of anti-VEGF have been recorded (95). Current guidelines
recommend continued examination until retinal vascularization extends to the
ora serrata or until PMA of at least 65 weeks if anti-VEGF medications were
used (91). Since ROP has been shown to recur past 65 weeks PMA (94–96),
clinicians may monitor for longer durations in anti-VEGF–treated infants.
Some clinicians advocate laser to the peripheral avascular retina in anti-
VEGF–treated eyes that have not had full vascularization. Harper et al.
demonstrated that only 50% of infant achieved vascularization into zone III at
150 weeks PMA and most eyes had vascular abnormalities of fluorescein
angiography after treatment for Type 1 ROP with anti-VEGF agents (108).
FIGURE 52-12 Detached retina in anti-VEGF–treated eye

that occurred months following anti-VEGF and then laser
treatment with apparent quiescence of disease. (Images by
Glen Jenkins, CRA, OCT-C.)
Laser
Laser to the avascular retina is the mainstay of treatment for ROP. Current
treatment recommendations are based on the ETROP study, which defined
Type 1 ROP (see Table 52-1) as necessitating ablative therapy to improve
structural and visual outcomes (37). Although techniques for laser
administration vary widely, successful laser ablative therapy is administered
after adequate anesthesia and analgesia have been accomplished and extends
from the vascular–avascular junction to the ora serrata 360 degrees. Near-
confluent burns or painting of the peripheral retina is utilized. As the
intravitreal neovascularization improves after initial treatment, laser can be
added posteriorly to parts of the retina that were not previously accessible or
visible, as occurs following regression of flat neovascularization in AP-ROP.
If there is inadequate regression of disease, laser can be repeated, especially
in any skip areas. Anti-VEGF agents can be considered and have been used
in cases of incomplete response to laser; however, this remains controversial
without knowledge of optimal dosing and full understanding of side effect
profiles (see below).
Anti-VEGF Studies
The Bevacizumab Eliminates the Angiogenic Threat of Retinopathy of
Prematurity (BEAT-ROP) study demonstrated a reduction in the rate of
recurrence of neovascularization in infants with stage 3, zone I ROP treated
with intravitreal bevacizumab (0.625 mg in 0.025 mL) and followed until 54
weeks’ PMA compared to eyes treated with laser. The interval for disease
recurrence was 19.2 ± 8.8 weeks with bevacizumab compared to 6.4 ± 6.7
weeks with laser for zone I disease, introducing the need for longer-term
follow-up to assure no disease recurrence. No difference between the
treatments was found statistically for zone II ROP (39). A follow-up study of
infants in the original BEAT-ROP cohort reported recurrences in 8.3% with
the average peak occurring at 16 weeks (±4 days) after injection (109). See
also Chapter 53.
Late recurrences of ROP after treatment with intravitreal bevacizumab
have been reported (94,95), including a report of a patient with recurrence at
3 years of age (96).
Reduced serum VEGF levels in infants treated with a single intravitreal
bevacizumab injection have been reported, but long-term systemic effects
remain unknown (110). More recent studies investigated the efficacy of
ranibizumab, which has a shorter half-life and reduces serum VEGF for a
shorter duration than bevacizumab (111). When used at the dose of 0.25
mg/0.025 mL (half the standard adult dose), ranibizumab resulted in higher
rates of recurrence than bevacizumab with an overall reactivation rate of 33%
at an average of 8.3 ± 2.7 weeks (111).
The Ranibizumab Compared With Laser Therapy for the Treatment of
Infants Born Prematurely with ROP (RAINBOW) study prospectively
compared treatment with intravitreal ranibizumab at two doses (0.2 and 0.1
mg of 10 mg/mL ranibizumab) compared to laser. Up to two retreatments
were allowed per eye in the ranibizumab groups after 28 days and up to 24
weeks, and re-treatment with laser was allowed. A total of 224 infants was
enrolled in the study, and the primary outcomes were absence of active ROP
and unfavorable structural outcomes in both eyes at week 24. The primary
outcome measure was met in 80% of the ranibizumab 0.2 mg group, 75% of
the ranibizumab 0.1 mg group, and 66.2% in the laser group (111). The
success rate of the laser group was much lower than what was seen in the
ETROP study; however, enrolled infants had more severe treatment-
warranted ROP than type 1 ROP.
To address the optimal dosing of anti-VEGF agents, the Pediatric Eye
Disease Investigator Group (PEDIG) studied de-escalating doses of
bevacizumab in the treatment of ROP. One eye was treated with 0.25 mg in
10 μL intravitreous bevacizumab for type 1 ROP. If an improvement was
seen in plus disease of zone I stage 3 ROP by 5 days or sooner without
recurrence for 4 weeks, the next group of infants received a lower dose of
0.125 mg, then 0.063 mg, and finally 0.031 mg, all in 10 μL volumes (112).
Even the lowest dose of 0.031 mg, which represents 5% of the dose used in
the original BEAT-ROP study, was effective in 9/9 eyes at 4 weeks. Serum
VEGF was reduced and bevacizumab was detected in the serum even at the
lowest doses (112). At 6 months follow-up, 41% of the 61 study eyes
received additional treatment for either early recurrence within 4 weeks of the
initial treatment (5%), late recurrence after 4 weeks (18%), and for persistent
avascular retina (18%). One patient developed a stage 5 retinal detachment, 4
infants died, and 56 of 61 had regression of ROP with normal posterior poles.
These data suggest good structural outcomes with low-dose bevacizumab;
however, additional treatment may be needed (113).
The Comparing Alternative Ranibizumab Dosages for Safety and
Efficacy in ROP (CARE-ROP) study compared 0.12 mg of intravitreous
ranibizumab (20 μL) with 0.2 mg (20 μL) in type 1 ROP in 19 infants and
found the two groups equivalent in terms of not requiring rescue therapy at
24 weeks follow-up (92). This study allowed for reinjection of ranibizumab
after at least 28 days and did not demonstrate plasma VEGF level alteration.
A summary of the key anti-VEGF therapy trials for ROP is presented in
Table 52-8.
TABLE 52-8 Anti-VEGF therapy trials in the

management of ROP
Z, zone; S, stage; +, plus disease; AP-ROP, aggressive posterior ROP; BEAT-ROP, Bevacizumab
Eliminates the Angiogenic Threat of ROP; RAINBOW, Ranibizumab Compared with Laser Therapy
for the Treatment of Infants Born Prematurely with Retinopathy of Prematurity; PEDIG, Pediatric Eye
Disease Investigator Group; CARE-ROP, Comparing Alternative Ranibizumab Dosages for Safety and
Efficacy in Retinopathy of Prematurity.
Although clinical trials have shown efficacy with lower doses of

bevacizumab and with ranibizumab, there are several considerations. There is
difficulty in assuring dose accuracy because of the dilution, small volumes
used in infant eyes, and the lack of optimal syringes and needles. (All trials
used doses that were specially mixed by compounding pharmacies in sterile
hoods.) Injections need to be carefully given to avoid injuring the retina or
the lens. The pars plana is not developed in the premature infant so the safe
zone in which to enter the eye is often <1.0 mm from the limbus. Recent
reports recommended the use of short 32G 4 mm needles on low-volume
syringes to avoid contralateral retinal injury and assure delivery of the correct
concentration of drug (108).
The need for reinjection is common with ranibizumab and persistent
avascular retina appears common. However, the time for reduced serum
VEGF is longer with bevacizumab. Both drugs appear to change the natural
history of ROP and require longer times of follow-up. Additional questions
persist (see also Chapter 53).
Predicting Progressive Stage 4 Retinopathy of

Prematurity Requiring Surgery
Few studies have provided information regarding risk factors associated with
progressive stage 4 ROP requiring surgical intervention. Clinical trials have
not been funded to pursue this question. To address this question, we studied
features of 72 eyes of 36 infants who had been treated with laser for threshold
ROP. Features were abstracted from examinations made 1 week prior to the
development of stage 4 ROP that was found to progress over time or 2 weeks
after laser in eyes that had regression of threshold ROP. Eyes were stratified
into two groups based on whether progressive stage 4 ROP or regressed stage
3 ROP occurred. A generalized estimating equation model was used to
account for within-subject variability and determine predictive features of
progressive stage 4 ROP. Features assessed included clock hours of ridge
elevation, quadrants of plus disease, quadrants of neovascularization, and
vitreous state. Ridge elevation was defined as white thickened tissue at the
junction of vascularized and avascular retina. Absence of ridge meant that no
or only a line denoted the junction. Plus disease was defined as dilated and
tortuous retinal arterioles and veins. The presence of only dilated veins in a
quadrant was counted as a half quadrant. The vitreous state was defined as
hazy, with hemorrhage, or vitreous condensation permitting visualization of
the regressed primary vitreous structures. The following features predicted
progressive stage 4 ROP: ≥6 clock hours of ridge elevation (P = 0.0248), ≥2
quadrants of plus disease (P = 0.0490), and vitreous haze (P = 0.0039) (101).
Neovascularization did not predict progressive stage 4 ROP; however, in a
separate analysis, it was associated with a poor surgical outcome (114).
Prevention
Prevention of ROP would be accomplished by preventing or reducing
premature births and through good prenatal care (115), reduction in teenage
pregnancies (116), and avoidance of illegal drug use (117).
To reduce the incidence of ROP, in the 1960s, NICUs began to monitor
oxygen delivery to the preterm newborn, particularly in the perinatal period
so as to avoid high oxygen levels, such as those used when RLF was first
recognized (6). NICUs now have the technology to maintain oxygen within
predetermined “safe” limits, defined by the Fetus and Newborn Committee of
the American Academy of Pediatrics and American College of Obstetricians
and Gynecologists (118). However, several studies provide evidence that not
only the concentration but also the variability of oxygen plays a role in the
development and severity of ROP (49,119) (see Chapter 40).
To prevent blindness from ROP in the preterm infant, screening is
important. Beginning with the CRYO-ROP study (11), it has been shown that
laser treatment or cryotherapy (Table 52-9) delivered to the avascular zones
of eyes with threshold ROP within 72 hours reduced unfavorable outcomes
and vision loss (11,14,33). The ETROP trial refined the definition of eyes at
risk of threshold ROP and an unwanted outcome: all zone I eyes with any
ROP and plus disease, zone I eyes with stage 3 ROP without plus disease,
and zone II eyes with stages 2 or 3 ROP and plus disease (Table 52-1, see
also Definitions of Treatment-warranted ROP) (2,37,121). If treatment is
adequate but ROP progresses to stage 4 ROP with certain retinal features (13)
or in eyes with stage 5 ROP, vitreoretinal surgery is considered. Finally,
visual rehabilitation is necessary to treat amblyopia and myopia that occur
more commonly in prematurely born children and even more often in those
who had ROP compared to infants born full term (Tables 52-2 and 52-3)
(22,122). Segmentation of encircling elements used to treat stage 4 ROP or
rhegmatogenous retinal detachment reduces optical irregularities, permits
growth of the globe, reduces myopia, and may improve visual outcomes
(123,124). Further discussion on amblyopia is provided in Chapter 11.
TABLE 52-9 Laser compared to cryotherapy for

treatment of threshold ROP
Laser-treated eyes 5.2 times more likely to have 20/50 visual acuity (VA). Linear regression analysis
revealed poor correlation (r = 0.36, P = 0.14, n = 18) in best-corrected visual acuity (BCVA) between
eyes of the same infant with bilateral disease and had one eye treated with laser and the fellow eye with
cryotherapy. For laser-treated eyes that lacked retinal dragging, BCVA was predicted to be 20/32, but
for cryotherapy-treated eyes (119), BCVA was predicted to be 20/50.
The thickness of the lens was most strongly correlated to refractive outcomes in both laser-treated (r =
0.885, P < 0.001) and cryotherapy-treated eyes (r = 0.591, P = 0.026) (120).
ETDRS, Early Treatment Diabetic Retinopathy Study visual acuity testing standards; VA, visual acuity.
Vision rehabilitation is important in all premature infants with or without
ROP. Premature infants are more likely to require strabismus surgery, to be
treated for amblyopia, and to be myopic than full-term infants (Tables 52-2
and 52-3). These conditions are greater in premature infants with more severe
levels of ROP than those with milder ROP (22,47,121,121). Infants with
macular heterotopia may have potential for visual acuity (125), so correction
of refractive errors and treatment of amblyopia are indicated. The ETROP
found patients without obvious macular heterotopia had moderate visual loss
(126). For the aphakic infant who had retinal detachment repair, visual
rehabilitation with special adjustments for low vision is recommended. Even
when grating acuity cannot be measured, low vision can be stratified into
levels characterized by the presence of light perception in different fields of
gaze (126) and may be important to infant and child development. In the
child with macular acuity in one eye and aphakic amblyopia in the fellow
eye, spectacle wear also provides protection against ocular trauma. We
recommend that infants be managed by a pediatric ophthalmologist for visual
rehabilitation starting at an earlier age than that recommended for full-term
infants and that children be enrolled in an early intervention program that
enhances the use of all of the senses.

A team approach and effective communication are important in the care of
the premature infant. During the time that the infant is in the NICU, the core
caregivers include the neonatologist, the screening ophthalmologist, and the
nursing staff. Often a nurse manager or ophthalmic technician can facilitate
communication between the ophthalmologist and the NICU to assure that eye
drops are given at the appropriate time, consultation reports are available for
the examining ophthalmologist to fill out, and the individual infant’s nurse is
available to assist and monitor for apnea and bradycardia during the
examination. We prefer to have a nurse, fellow physician, or trained
technician assist by securely holding the swaddled infant during the
examination. It is important to provide a written consultation for all infants
and to communicate with the neonatologist, especially about infants with
progressing ROP. Ongoing communication with the parents helps to facilitate
understanding of the condition, coordinate follow-up, and assure compliance
with follow-up after discharge.
Once the infant is discharged from the NICU, follow-up examinations for
the retina and later for visual rehabilitation are important. Often, a written
information booklet is given to parents or guardians with emphasis placed on
the importance of the timing of appointments. A system is required to
reschedule infants as soon as possible if appointments are missed and
requires training of the office staff to assure infants needing urgent
appointments are examined. When repeated appointments are missed, we
send a certified letter to the parents or guardians. Once the risk of ROP is
reduced, follow-up with a pediatric ophthalmologist and in an early
intervention program is important for visual rehabilitation.
It is also important to communicate about examinations, diagnosis, and
treatments for ROP when an infant is transferred to another hospital. This is
especially important if anti-VEGF has been given since recurrences can occur
at later time points than the natural history. In our center, the ROP
coordinator communicates directly with the ROP coordinator or their
designee at the accepting hospital. This is done in addition to any physician-
to-physician sign-outs to create redundancy in the information.
Ethical questions arise when working in neonatal intensive care. Each
question is taken on an individual basis, accounting for the infant’s general
condition and fellow eye, and is addressed by a team that may include the
neonatologist, parents, and ophthalmologists. For example, it is universally
agreed that stage 5 ROP should be prevented, and efforts are made to
accomplish this. Careful monitoring of eyes after treatment for ROP is
essential to diagnose progressive stage 4 ROP that might be repaired with a
lens-sparing vitrectomy (127,128). However, cases of stage 5 ROP can occur
when an infant is too sick to undergo treatment, when treatment does not
prevent retinal detachment, or when infants are lost to follow-up despite
efforts to prevent this. The question may arise as to whether heroic surgery
should be performed to reattach the retina in an eye with stage 5 ROP in a
preterm infant with multiple medical conditions. In addition, visual prognosis
in stage 5 disease even with partial retinal reattachment (which is considered
anatomical success) is very poor (114,129,130). Evidence exists that low
vision can be stratified in infants with repaired retinal detachments from stage
5 ROP (129) and can be useful in children when at a level believed to have
little value in adults (131). Still, long-term benefits of low vision on child
development and quality of life are difficult to study and remain largely
unknown.
FUTURE STUDIES
Erythropoietin Derivatives (PENUT)
Early studies of erythropoietin (epo) found an association with severe ROP;
however, later studies suggested that timing of epo administration made a
difference in outcome (73). Erythropoietin has anti-inflammatory and
antiapoptotic effects and promotes neurogenesis and angiogenesis, which will
be studied in the PENUT (Preterm Epo Neuroprotection Trial) in extremely
low gestational age neonates (72). The study will enroll 940 patients and
follow infants to 22 to 26 months corrected age. Safety of epo will be
assessed as well as effects on inflammatory cytokines, brain injury, as well as
clinical outcomes.
RISK FACTORS
Severe (Treatment-Warranted) Retinopathy of
Prematurity
The greatest single risk factor for developing ROP is being born prematurely
(64,132–144). However, we now know much more about the risks of
progression of the disease. The CRYO-ROP study provided data from the
natural history cohort (Table 52-1) on ocular and systemic characteristics
associated with increased risk of ROP, progression to threshold ROP, and
development of an unfavorable macular outcome (Table 52-5) (19). An
important ocular risk factor was zone I. If, at 32 weeks PMA, incomplete
vascularization in zone I was present, 32.8% of eyes developed threshold
ROP, compared to 9.3% with incomplete vascularization in zone II. Eyes
with zone I ROP also were at high risk of developing threshold ROP. Other
ocular risk factors for developing threshold ROP included plus disease, stage
3 ROP (19), ≥6 clock-hour stage 3 (18), and iris vessel dilation (31).
Nonocular risk factors associated with the development of threshold ROP
included young gestational age, multiple births, out-of-nursery birth, low
birth weight (19), and Caucasian race (Table 52-1) (19,28). Once
prethreshold ROP developed, the risk of developing threshold ROP was
about the same given these factors. For each 100-g increase in birth weight,
there was a 27% decrease in the percentage of infants who developed
threshold ROP. Black race was associated with a 6/5% lower chance of
developing threshold ROP compared to Caucasian race if any ROP was
present and a 51% decreased chance of developing threshold if prethreshold
ROP developed. Once threshold ROP developed, the risk of an unfavorable
anatomic outcome (Table 52-5) was about the same in blacks as for
Caucasians (19). Factors associated with increased risk of prethreshold
progressing to threshold ROP included lower PMA at the diagnosis of any
ROP, ROP in zone I at the first screening examination, rapid progression of
ROP to prethreshold characteristics, plus disease at the first prethreshold
examination, and white race (19,29).
UNFAVORABLE MACULAR OUTCOME
In the initial natural history report, ocular characteristics associated with an
unfavorable macular outcome included zone I ROP, plus disease, and stage 3
ROP (Table 52-1). Eyes with zone I ROP had an odds risk of 8.24 toward
developing an unfavorable outcome compared to zone II ROP (19). For each
clock hour of stage 3 ROP >5 clock hours, there was a 26% increased risk of
an unfavorable macular outcome (19). Stage 3 ROP with plus disease in zone
II was associated with a 62% risk of an unfavorable result compared to stage
3 ROP without plus disease in zone II in which the risk was 3%. In the 5.5-
year report from the natural history cohort of CRYO-ROP (18), an
unfavorable anatomic outcome occurred in 62.5% of zone I eyes compared to
44.2% of zone II eyes. All unfavorable anatomic outcomes in this cohort
from CRYO-ROP occurred in eyes with 6 or more clock hours of stage 3
ROP with plus disease in zone I or II (18).
In the ETROP study, at 6-year follow-up, the difference in unfavorable
outcomes for all high-risk prethreshold eyes was 8.9% for early-treated eyes
versus 15.2% for conventionally managed eyes (P < 0.001). The greatest
benefit of early treatment was seen with zone I, stage 3 disease, with or
without plus (unfavorable outcomes were doubled for these eyes in the
conventionally treated group), and zone II, stage 3 disease with plus (40).
EFFECTS OF ENVIRONMENTAL
FACTORS AND NONOCULAR
TREATMENTS ON RETINOPATHY OF
PREMATURITY
Oxygen
The relationship of oxygen to the development of ROP is complex and
incompletely understood. Metabolic and oxygen needs increase during the
development of the retinal vasculature and the maturation of the neural retina
and photoreceptors (145). Early retinal vessels are also sensitive to
fluctuations in outside oxygen delivery. A number of factors also affect
oxygen delivery to the retinal vasculature and neural retina in the premature
infant, including poor blood oxygenation secondary to immature lungs and
respiratory disease, anemia of prematurity, and changes in the ratio of fetal to
adult hemoglobin affecting oxygen affinity to hemoglobin.
Steroids
The role of steroid administration and its effect on ROP remain incompletely
understood. A meta-analysis published in 2018 demonstrated that antenatal
steroids reduced the risk of ROP development and progression to severe ROP
(146).
Prenatal steroids administered to women in preterm labor have resulted in
reduced infant mortality and morbidity, primarily through improved lung
function. There is evidence from several studies that prenatal steroids are
protective for the development of ROP (143,147–149). Later administration
of steroids, for example, when treating lung disease, has been reported to
have an adverse effect (150,151) or no effect (152–154) on the incidence of
ROP. Adverse effects appear to be more likely when steroids are given more
than 3 weeks postnatally (151) and less likely when used within 2 weeks of
birth (152,153).
In a rat model of OIR, an angiostatic steroid, anecortave acetate, believed
to prevent endothelial cell migration, greatly reduced intravitreous
neovascularization (155) with little to no effect on normal retinal vascular
development. Dexamethasone inhibited both normal retinal growth and
pathologic intravitreous neovascularization in rabbits (156). Intravitreous
triamcinolone was found to have dose-dependent reductions in intravitreous
neovascularization (157). Further investigation into possible effects of
steroids on ROP is warranted.
Surfactant
Surfactant therapy that has increased survival in very low birth weight
premature infants initially appeared to be beneficial in reducing the incidence
of ROP (158). However, follow-up studies have failed to confirm this
(18,144,159,159). Studies found a reduced incidence of severe ROP in
preterm infant treated with surfactant therapy (160); however, studies used
historic controls (161). In a recent meta-analysis, inositol, an essential
nutrient and important in maturation of surfactant, was shown to significantly
reduce the incidence of ROP (38,162).
Carbon Dioxide
Clinical studies that analyzed intermittent blood gas samples found that both
hypocarbia (132) and hypercarbia (38,133,163) were associated with the
development of ROP. The only study that analyzed continuous
transcutaneously monitored CO2 showed no association between absolute
blood CO2 level or variability and the subsequent development of ROP (164).
The evidence from animal models, however, is unequivocal. Hypercarbia
alone (165) and metabolic acidosis (166) have induced retinopathy in rats.
When hypercarbia is combined with variable oxygen (165), the severity of
disease is increased.
CO2 affects the retinal vasculature by causing dilation of vessels in part
through relaxation of retinal pericytes (167). With vessel dilation,
oxygenation (166) and blood flow (168) are increased to the retina. The
damaging effect of CO2 in animal models has been speculated to be through
increased delivery of oxygen to the retina, followed by a reduced stimulus for
the release of hypoxia-induced angiogenic factors, such as VEGF, and, in
turn, delay retinal vascular development. Retinal VEGF mRNA down-
regulation, followed by up-regulation prior to the appearance of maximal
neovascularization in hypercarbic OIR, has been demonstrated (169).
A small trial in ventilated preterm infants that allowed the level of carbon
dioxide to passively rise to between 45 and 55 mm Hg PaCO2 showed no
difference in the incidence of ROP (170) compared to infants who were
ventilated to maintain PaCO2 levels of 35 to 45 mm Hg. Infants exposed to
PCO2 in the highest quartile for GA on at least 2 of the first 3 days of life
were found to have an increased risk of severe ROP (59).
Other Risk Factors and Treatments

Other small studies have described additional risk factors for ROP, including
intraventricular hemorrhage, likely via shared risk factors (4,171); systemic
infection and inflammation (172–174); respiratory distress syndrome;
bronchopulmonary dysplasia; and patent ductus arteriosus (143). Bilirubin,
through its antioxidant qualities, was proposed as a protective compound
against ROP; however, elevated bilirubin was not found to be protective
against ROP (103) and was speculated to be a risk factor in one study (81).
Elevated glucose levels in the 1st week of life were found to be associated
with the development of ROP (175).
D-Penicillamine, used to prevent or treat hyperbilirubinemia, was found
to be associated with reduced ROP (176). A recent randomized, double-blind,
placebo-controlled trial of 88 infants showed no difference in the incidence of
ROP with and without oral D-penicillamine administration (177). Iron, a
known oxidant, has been implicated indirectly through studies of blood
transfusions, anemia, and erythropoietin use. Blood transfusions have been
associated with increased risk of ROP development (140,178), as has the use
of erythropoietin (179).
Propranolol, a nonselective beta-blocker, used to reduce the growth of
infantile hemangiomas, is hypothesized to reduce VEGF levels, and its safety
as well as efficacy in reducing the progression of ROP will be evaluated in
the PROP-ROP study (180).
There was no association between mean arterial blood pressure
measurement and ROP reported in an observational study in newborn
preterm infants over the 1st week of life (181).
Several studies have compared the incidence in ROP between twins and
singletons and have found no difference in risk once birth weight was taken
into account (182,183).
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53
Anti-VEGF Treatment in Retinopathy
of Prematurity
Anna L. Ells, Wei-Chi Wu, and Darius M. Moshfeghi
INTRODUCTION
Retinopathy of prematurity (ROP) affects up to 16,000 premature infants in
the United States per year (1,2) and an estimated 184,700 globally (3), and is
a leading cause of childhood blindness (4–10). Incomplete retinal vascular
development predisposes the premature infant to a vasoproliferative
retinopathy characterized first by slowed or even arrested blood vessel
growth followed by abnormal vascular development into the vitreous. When
retinal vascular development restarts, it may proceed as ordered intraretinal
angiogenesis or transition into pathologic pathways with abnormal
angiogenesis followed by gliosis, vitreoretinal traction, and retinal
detachment (11).
To accurately and reproducibly describe the vascular pathology
associated with ROP, the International Classification of Retinopathy of
Prematurity (ICROP) has defined ROP characteristics such as location
(zone), extent of neovascularization (clock hours), severity (stage), and
modifiers (presence of plus, pre-plus, and aggressive posterior disease)
(12,13). (Information on diagnosis, screening parameters, and clinical trials is
found in Chapter 52.)
However, it should be remembered that countries around the world have
different appearances of ROP with larger and older infants than in the United
States and Canada at risk for severe ROP; therefore, many countries have
developed their own recommended screening guidelines (see Chapter 50).
PATHOGENESIS OF ROP
The pathogenesis of ROP is related to interruption of the normal pattern of
retinal vascular development with ensuing pathologic processes. Most studies
performed on normal retinal vascular development have been done in
animals, but there are differences between humans and other species (see
Chapter 5). In normal human retinal development, the initial vasculature is
believed to occur by vasculogenesis beginning in the 16th week of gestation
and continuing through at least 22 weeks of gestation (14). From that time,
how vessels develop in the human infant is believed to occur by a process of
angiogenesis based on animal models.
During vasculogenesis, it is believed that growth factors increase as a
result of relative intrauterine hypoxia and increased metabolic activity
associated with neuronal retinal differentiation and include stromal-derived
factor (14) and its receptors CXCR-4 and vascular endothelial growth factor
(VEGF), produced from retinal ganglion cells (RGCs) and astrocytes (15).
The third trimester is believed to be associated with a transition from
vasculogenesis to angiogenesis for retinal vasculature development.
Angiogenesis is characterized by endothelial sprouting from an existing
vessel and proliferation and is often stimulated by VEGF, as well as
recruitment of mural cells from other factors during blood vessel creation
(16). In humans, angioblasts migrate radially from the optic nerve to the ora
serrata, expressing VEGF in relationship with local tissue hypoxia gradients,
whereas astrocytes have been found important in this process in experimental
animal models (17). Endothelial tip cells direct vessel development (18),
utilizing tractable filopodia, which in turn are patterned by the response of the
numerous guidance receptors they express, including VEGF receptors,
Unc5b, and neuropilin 1 (NRP1) (19–21). Trailing endothelial stalk cells then
establish a fledgling vascular network. VEGFC and VEGFR3 have important
roles in the process of endothelial tip to stalk cell conversion through
regulation of notch signaling (22). Retinal vascular maturation is modulated
by a complex interaction of mural cells, which mature into pericytes and
smooth muscle cells, and apoptosis. Ultimately, it is believed that the dual,
simultaneous process of vascular maturation and involution results in the
development of the retinal vessels (23,24).
The disruption of normal retinal vascular development by ROP has been
characterized by two phases (25): (a) phase I, delayed physiologic retinal
vascular development and vasoattenuation due to oxygen stresses (see also
Chapter 38), and (b) phase II, vasoproliferative intravitreal
neovascularization. It is believed that when a preterm infant is born, there is
insufficient insulin-like growth factor 1 (IGF-1) and other factors that are
usually supplied through the placenta (15). Low IGF-1 is associated with
slow retinal vascular development (see Chapter 39). In addition, a number of
events related to birth and to postnatal insults, including oxygen fluctuations,
inflammatory stimuli, and oxidative stress occur, delaying physiologic retinal
vascular development further and causing aberrant growth of blood vessels
into the vitreous. Birth also may expose the infant to relative extrauterine
hyperoxia (26,27) (see Chapter 40). The oxygen concentration in the tissue is
complicated, because fetal hemoglobin has a greater affinity for oxygen than
does adult hemoglobin, and there can be variable effects based on shunting
and dilution of blood. Early experimental models suggested that high oxygen
injured newly formed retinal capillaries (28–32) in phase I and did not
promote physiologic retinal vascular development. More relevant and recent
experimental models suggest that oxygen stresses of preterm infants today
cause other events that lead to increasing levels of VEGF with delayed
expression of VEGFR2, which lead to first delay in physiologic retinal
vascular development and then, once VEGFR2 is present and activated in
endothelial cells, disordered divisions of endothelial cells that have access to
outside the retinal plane where they proliferate as intravitreal angiogenesis in
phase II. The disordered proliferation of retinal endothelial cells into the
vitreous does not allow them to extend as intraretinal neovascularization in
physiologic retinal vascular development (33,34). The ischemia from the
peripheral avascular retina, then combined with the increasing metabolic
demand of the developing retina, results in tissue hypoxia that induces cells,
including glia, astrocytes, and neurons (35,36) to overexpress angiogenic
factors and trigger angiogenic pathways involving, for example, VEGF and
erythropoietin (15,37,38). Stage 3 ROP occurs at about 32 to 34 weeks of
gestational age and is characterized by pathologic intravitreal
neovascularization (28–35). Extrapolating, it has been postulated that zone I
disease may stem from aberrant vasculogenesis, whereas zone II disease is
likely angiogenesis dependent (39,40). Compared to control eyes undergoing
surgery for congenital cataract, vitreous samples from eyes with stage 4 ROP
requiring surgery demonstrate a nearly 50-fold increase in VEGF
concentrations with the highest levels in eyes with plus disease (41). It is
clear that retinal vascular maturation and the pathogenesis resulting in ROP
have numerous steps involved in interruption of normal development
followed by ROP. Therefore, there may be more than one treatment required.
TREATMENT
The goals of treatment in ROP are to preserve vision potential, preserve
retinal architecture, and limit local and systemic complications related to
therapy. The past decade has included numerous case series and a few
clinical trials that support pharmacotherapy using anti-VEGF drugs.
However, due to limited strong evidence, laser photocoagulation of the
avascular retina remains the gold standard treatment of ROP requiring
therapy (42) based on the results of two large National Eye Institute (NIH)
studies: The Cryotherapy for Retinopathy of Prematurity (CRYO-ROP) trial
and the Early Treatment for Retinopathy of Prematurity (ETROP) trial
(10,43). The conclusions of these studies are ablative therapy for type 1
ETROP, zone I, stage 3 disease; zone I, plus disease; and zone II, stage 2 or 3
with plus disease (43,44), replacing the earlier threshold criteria from the
CRYO-ROP trial (see Chapter 52).
Scientific Premise and Rationale for Anti-VEGF

Therapies
As noted above, immature retinal vascularization at the time of birth is driven
by angiogenesis, and VEGF is a dominant angiogenic factor (31). Inhibition
of VEGF has been successful in other adult retinovascular diseases associated
with aberrant angiogenesis, including age-related macular degeneration
(AMD), diabetic retinopathy, and retinal vascular occlusions (45–59). The
initial published report of efficacy of anti-VEGF for ROP involved
bevacizumab (60,61).
Historically, the management of ROP has changed over time, initially
with observation, followed by cryotherapy and then laser. Both cryotherapy
and laser photocoagulation have several limitations (62–64), including the
need for infrastructure and equipment investment, some form of anesthesia,
and mentored training. In addition, there is a moderate risk of adverse events
and inescapable side effects, including decreased night and peripheral vision
and increasing evidence for the propensity for high myopia.
Furthermore, the results are not universally encouraging, particularly
across regions having insufficiently skilled ophthalmologists to screen,
diagnose, and treat ROP (Table 53-1) (43,66–68). Even in regions with good
outcomes, about 20% to 35% of infants with posterior ROP progress to
adverse anatomic outcomes and/or visual disability/impairment
(43,65–67,70), suggesting that retinopexy is not uniformly successful across
diverse populations of treaters (64). The authors believe that this is due to
lack of a uniform screening strategy coupled with lack of training in proper
application of retinopexy in type 1 ETROP infants, resulting in delayed or
late treatment and lower rates of success (64).
TABLE 53-1 Retinopexy success rates
aCRYO-ROP evaluated initial anatomic success of cryotherapy versus observation at 3 months

posttreatment.
bWills evaluated anatomic success at 3 months for cryotherapy versus laser photocoagulation (Argon
1991, Diode 1992).
cPrimary outcome in ETROP was related to visual function. A key secondary outcome was anatomic
outcomes at 9 months reported herein. Additionally, treaters in ETROP had the option of using either
cryotherapy or laser photocoagulation, with all but one infant receiving laser photocoagulation.
dBEAT-ROP primary outcome was the rate of recurrence at 54 weeks of postmenstrual age. The results
reported herein are for the combined pool of zone I and aone II posterior ROP. For zone I-only eyes,
the results were substantially worse—58.0% success (n = 19 of 33 eyes).
eRAINBOW, despite using two doses, defined primary outcome as superiority of the 0.2-mg
ranibizumab dose relative to laser at 24 weeks with respect to absence of active ROP or absence of
unfavorable structural outcomes.
The use of anti-VEGF has made accessible the treatment of ROP, particularly
posterior ROP, in that success routinely exceeds 75% at preventing adverse
anatomic outcomes and visual acuity loss regardless of training of the
physician, anti-VEGF medication choice, dosage, or geographic location
(Table 53-2) (68,73–81). Hence, we enter the fourth era of ROP treatment.
TABLE 53-2 Anti-VEGF for ROP
aBEAT-ROP primary outcome was rate of recurrence at 54 weeks of postmenstrual age. The results
reported herein are for the combined pool of zone I and zone II posterior ROP. For zone I-only eyes,
the results were substantially worse—58.0% success (n = 19 of 33 eyes).
bAngiographic and anatomical outcomes, mortality.
cPrimary outcome of bevacizumab versus laser is defined as persistence or recurrence at 54 weeks of
postmenstrual age.
dRAINBOW, despite using two doses, defined primary outcome as superiority of the 0.2-mg
ranibizumab dose relative to laser at 24 weeks with respect to absence of active ROP or absence of
unfavorable structural outcomes.
Possible Anti-VEGF Candidates for ROP

Anti-VEGF agents for treating ROP include pegaptanib (Macugen; Eyetech
Inc., Cedar Knolls, NJ, United States), bevacizumab (Avastin; Genentech
Inc., San Francisco, CA, United States), ranibizumab (Lucentis; Genentech
Inc., San Francisco, CA, United States), and aflibercept (Eylea; Regeneron
Pharmaceuticals, Tarrytown, NY, United States). Each has different
pharmacokinetic effects, molecular sizes, structures, and half-lives (Table 53-
3).
TABLE 53-3 Pharmacodynamics and

pharmacokinetics of the three most commonly
used anti-VEGF agents for ROP
aEstimated half-life based on mathematical modeling showed aflibercept to have an intravitreal half-
life of 7.13 days as compared to the intravitreal half-life of ranibizumab of 4.75 days and bevacizumab
of 8.25 days (88–91).
bHuman data not available. The half-life in aqueous humor of nonvitrectomized macaque eyes was 2.2
days for aflibercept and 2.3 days for ranibizumab (92).
ROP, retinopathy of prematurity; VEGF, vascular endothelial growth factors.
Pegaptanib is a single-strand nucleotide that is believed to specifically bind

the splice variant, VEGF165. Pegaptanib was the first VEGF inhibitor
approved by the U.S. Food and Drug Administration (FDA) for the treatment
of neovascular AMD in December 2004 (49,93). However, it is rarely used in
most countries now because this drug was shown to be less effective than the
other anti-VEGF agents.
Bevacizumab is a full-length monoclonal antibody (149 kDa), which
binds to human VEGF-A (94). It was originally developed for the treatment
of metastatic colorectal cancer (94) and has been used to treat many
retinopathies with good results. Bevacizumab is by far the most widely
studied anti-VEGF agent in ROP, and good responses have been
demonstrated especially in severe zone 1 or posterior zone II ROP
(68,95–98).
Ranibizumab is a smaller humanized monoclonal antibody Fab fragment
(48 kDa), which binds to VEGF-A and was specifically designed for
intraocular use (99). Without the Fc antibody region, it is rapidly eliminated
from the bloodstream with a systemic elimination half-life of approximately 2
hours (86,100). Because of its rapid clearance, ranibizumab is believed
potentially to have less of an effect on developing organs that require VEGF
in preterm infants. Ranibizumab was shown to be effective for type 1 ROP
(81,101,102) but with somewhat conflicting results due to higher incidence of
nonresponse and recurrence of ROP in several series and a clinical trial
(69,75,103–105).
Aflibercept is a humanized recombinant fusion protein (115 kDa), which
binds to multiple VEGF family members, including VEGF-A, VEGF-B, and
placental growth factor (PlGF) (106). Compared to bevacizumab and
ranibizumab, aflibercept has a greater binding affinity for VEGF165 and a
longer estimated intraocular half-life (107,108). Few studies have been
performed testing aflibercept as the initial treatment for ROP, but some
investigators reported good responses in treatment-requiring posterior ROP
(76,109,110).
The Rationale for Each Drug

All anti-VEGF agents are used off-label, because they are not FDA approved
for intraocular use in children. (Ranibizumab is FDA approved for use in
ROP in Europe, as of this writing). When selecting the appropriate treatment
for infants with ROP, evidence, side effects, potential complications, and cost
are often taken into consideration. Reactivation or recurrence of ROP can
occur even a year after treatment and even after intraretinal vascularization
has extended into zone III. Before considering treatment with an anti-VEGF
agent, it must be ascertained that the patient will have regular follow-up so
that if ROP recurs, treatment can be performed.
Bevacizumab is the most widely investigated anti-VEGF drug, and its
effect in ROP was reported in a prospective randomized multicenter trial
(68). Organ systems in the preterm infant require VEGF, and, therefore,
safety concerns exist particularly since bevacizumab lowers serum VEGF for
over a month (111). Several studies tested neurodevelopment scores in
infants treated with anti-VEGF or laser and reported conflicting results (see
below), although studies varied in design and limitations (112–115). A
considerable limitation regards neurodevelopmental testing, which often
relies on tasks requiring vision in young children. Long-term follow-up will
be important to assess neurodevelopment. In addition, most infants at risk of
treatment-warranted ROP are also at risk of neurodevelopment delay from
extreme prematurity. Other studies reported lower myopia and better vision at
5 years (68,116) (see Chapter 52).
Ranibizumab and aflibercept were designed for intraocular use, and both
have greater affinity and potency than bevacizumab (107,117,118).
Ranibizumab reduces serum VEGF for a shorter duration than bevacizumab
or aflibercept but also may require additional injections. Published studies
report more nonresponders and higher recurrence rates when reviewing
treatment with ranibizumab to studies using other agents (69,75,103,104).
The binding affinity of aflibercept to VEGF receptors is almost 100 times
greater than that of ranibizumab and bevacizumab (Table 53-3). Aflibercept
binds and inhibits VEGF-A, PlGF-1 and PlGF-2, and VEGF-B, which have
been implicated in pathologic vascular remodeling (119). The intermediate
size of aflibercept (115 kDa compared to 48 kDa for ranibizumab and 148
kDa for bevacizumab) results in longer intravitreal half-life and a duration of
clinical action potentially as long as 2.5 months (Table 53-3) (88). The
potency and long duration of aflibercept may lead to greater complications by
removing VEGF, which is important in retinal development and is a survival
factor. However, if it is shown to be safe in experimental studies, aflibercept
may result in fewer injections and potentially fewer recurrences. Current
reports of small studies have not reported problems (76,109,110). Future
prospective, randomized trials with larger enrollment will be needed to
confirm its effects and its safety. In addition, all anti-VEGF agents require
additional evidence from large-scale clinical trials followed with sufficient
follow-up to assess reliable neurodevelopmental scores.
Long-Term Safety and Neurodevelopmental Outcomes

VEGF plays an important role in neurogenesis in embryos and preterm
newborns. Blocking VEGF-A expression was reported to impair brain
vascularization (120) and lead to neuronal apoptosis in the retina (121). In
addition, factors induced by hypoxia, including VEGF, were lower in preterm
compared to term rabbit pups, and reduced VEGF has been postulated as a
cause of neurodevelopmental delay and poor growth of the cerebral cortex in
premature infants (122). Significant neurogenesis continues in preterm
infants until the third trimester (122), so the concern that deprivation of
serum VEGF in preterm infants may have long-term effects on the
development of the central nervous system and other systems exists.
In newborns, Sato et al. (123) found that systemic VEGF levels were
depressed for at least 2 weeks after the administration of either 0.25 or 0.5
mg intravitreal bevacizumab (IVB) in patients with stages 3, 4, and 5 ROP.
Kong et al. (124) demonstrated that serum free VEGF levels decreased 2 days
following treatment with each group, 0.25 or 0.625 mg IVB or laser
photocoagulation, and the reductions were more significant in both the IVB-
treated groups. They also found the clearance of bevacizumab from the
bloodstream took at least 2 months after IVB (124). Wu et al. (125,126)
further showed that serum VEGF levels in type 1 ROP infants were
significantly decreased for up to 12 weeks after a single administration of
0.625 mg IVB.
Compared to bevacizumab, ranibizumab has a much shorter half-life of 2
hours in plasma in adults (100). Zhou et al. (127) demonstrated that in ROP
infants, plasma VEGF levels were reduced following intravitreal ranibizumab
(IVR) but returned to normal 1 week later. Wu et al. (126) also reported that
IVR caused almost no detectable suppression of systemic VEGF at 2 months
after injection.
Huang et al. (128) reported that intravitreal aflibercept (n = 5 infants) or
IVB (n = 9 infants) caused serum VEGF suppression for up to 3 months after
a single injection, noted as greater in the group that received IVB.
Concerns about the potential adverse effects of long-term reduction in
serum VEGF on neurogenesis in the preterm infant were raised
(115,129,130). The current largest retrospective series was reported by Morin
et al. (115). The Bayley-III scoring system was used to compare
neurodevelopmental outcomes of preterm infants at 18 months of corrected
age following previous treatment with IVB (n = 27) or laser ablation (n = 98).
The study showed higher odds of severe neurodevelopmental disabilities in
patients receiving IVB compared to laser ablation. However, whether the two
groups in this study could be directly compared was brought into question
(131). The study included sicker infants in the IVB group, and some of the
infants in the laser group had milder ROP that did not meet the current
treatment criteria (131). Moreover, more patients in the laser group were
excluded from the study due to the inability to perform the Bayley-III exam.
Using the Bayley-II exam, Lien et al. (112) retrospectively compared
neurodevelopmental outcomes of preterm infants treated with IVB only (n =
12), laser treatment only (n = 33), or a combination of laser and IVB
treatment (n = 16). They reported the neurodevelopmental scores were
similar at 2 years after treatment with IVB-only or laser-only group, but the
study was retrospective and also had bias as to the treatment choice for
individual infants.
In a prospective noncomparative case series, Martinez-Castellanos et al.
(113) evaluated ROP patients 5 years after IVB injections using the
standardized Denver Developmental Screening Test II. All except one
critically ill patient had normal neurodevelopmental scores. However, the
study had no control group, and the patient number was relatively small (n =
13).
Kennedy et al. (114) reported a randomized prospective study comparing
neurodevelopmental outcomes at the corrected age of 18 to 22 months in
infants from the BEAT-ROP trial. The Bayley-III neurodevelopmental scores
and the growth indices (weight, length, and head circumference) were not
significantly different between the IVB (n = 7) and laser (n = 9) treatment
groups. However, the study was limited by the relatively small patient
number, which was not powered to identify small but important differences
that may have been detected in a sufficiently powered study.
Apart from the limited number of patients, most of the above studies
reported neurodevelopmental outcomes in bevacizumab-treated premature
infants before 2 years of age (112,114,115), and only one study reported 5-
year outcomes (113) (Table 53-4). Many developmental deficits in cognition,
emotional and behavioral development, and social adaptive functioning can
emerge at older ages in the absence of obvious neurodevelopmental
impairment in toddlerhood (132,133). Therefore, a longer follow-up of these
anti-VEGF–treated infants is needed to fully comprehend their late
neurodevelopmental outcomes.
TABLE 53-4 Published studies reporting

neurodevelopmental outcomes in premature
infants treated with bevacizumab for ROP
Anti-VEGF ROP Trials

The anti-VEGF literature is replete with small trials, poorly conceived, with
inability to determine safety (68,72,75–81,95,104). However, it remains clear
that anti-VEGF is a successful treatment to reduce vascular activity in
posterior ROP (Table 53-2).
We have outlined the major prospective randomized controlled trials
using anti-VEGF for ROP (Table 53-2). Chapter 52 will discuss the major
anti-VEGF clinical trials in more detail.
Technique of Intravitreal Anti-VEGF Injection for

ROP in an Infant
Intravitreal injections for ROP in preterm infants are performed at the bedside
in the neonatal intensive care unit. There are differences in techniques used in
the literature, and there is no consensus as to a standardized approach.
However, because the anatomy of the premature infant eye differs from the
adult, there is potential risk of damage to the lens or retina during the
injection (134–137). The lens of the premature infant is relatively larger
compared to the adult. The pars plana has not completely developed in the
preterm infant such that the safe zone to enter the vitreous is narrow. The
anteroposterior diameter of the globe is about two-thirds that of the adult.
Given that injections are performed on awake infants, it is important that the
ophthalmologist be experienced in delivering intravitreal injections. Sedation
may be useful to calm the infant and is recommended under the supervision
of the attending neonatologist and neonatal staff.
After an informed consent, the procedure is performed using sterile
technique. A mask and sterile gloves are recommended. Appropriate direct
light is important. A sterile speculum is used such as an Alfonso pediatric
speculum. Topical anesthetic is applied. A 10% ophthalmic preparation of
Betadine is applied to the periorbital skin and lids. Five percent povidone
iodine is placed into the conjunctival cul-de-sac and onto the cornea. A small-
needle and small-volume syringe is recommended such as the 32 G on a 1.0-
cc syringe. Using calipers to measure, the needle is inserted at 0.5 mm
posterior to the limbus parallel to the visual axis. If using a 4-mm needle, it
may be inserted to the needle hub; however, if a 12.5-mm needle is used (1/2
inch), then the needle should only be inserted approximately 4 mm into the
vitreous cavity. A scleral depressor or pair of forceps can be used to stabilize
the eye if necessary. Immediately following the injection, the fundus should
be examined with indirect ophthalmoscopy to ensure that the drug is in the
vitreous and that the central retinal artery is patent. Topical antibiotics may
be recommended for 3 to 7 days.
RECURRENCE AND RETREATMENT
OF ROP AFTER ANTI-VEGF THERAPY
From our experience and published studies over the last decade, it is clear
that anti-VEGF therapy for ROP can result in excellent initial regression of
ROP; however, recurrence of ROP is common and can occur late in the
course. It is a fine balance to pharmacologically inhibit pathologic intravitreal
neovascularization and allow more normal intraretinal vascularization of the
premature retina to occur. Appropriate dosing and agents remain
undetermined. Furthermore, delivery systems are not consistently accurate to
deliver small volumes and dilute concentrations of drugs often formulated for
adult eyes. To complicate the pharmacologic treatment paradigm, a single
injection of anti-VEGF agent does not always result in permanent reversal of
ROP, and a second injection 4 to 8 weeks following the initial injection may
be required (104). The factors that may be related to recurrence of ROP
following anti-VEGF are discussed additionally in Chapter 38.
Most anti-VEGF drug efficacy studies in the literature use recurrence rate
requiring treatment as the primary outcome. In the BEAT-ROP study, the
eyes treated with bevacizumab with zone I ROP had a 6% recurrence rate,
and those eyes with zone II ROP had a 5% recurrence rate within a
postmenstrual age (PMA) of 54 weeks (68). In their second follow-up report,
the authors reported an overall recurrence rate of 8% after treatment with IVB
monotherapy. Other studies reported a higher recurrence rate of ROP
following IVB. Hwang et al. reported a 14% recurrence rate following IVB
and only 3% in the laser group within a follow-up of approximately 24 weeks
in a retrospective review (95). Karkhaneh et al. reported a 10.5% recurrence
rate in the IVB group and only 1.4% in the laser group (72). Toy et al.
reported a 91% recurrence rate at a mean interval of 15-week postinjection in
their retrospective series of IVB-treated infants (138). In the dose study of
bevacizumab for ROP (73), the overall recurrence rate was 41%. Recurrence
rates appear to be the same or higher in those premature infants treated with
IVR. The Comparing Alternative Ranibizumab Dosages for Safety and
Efficacy for ROP (CARE-ROP) study reported a 21% recurrence rate after
IVR (74); Ells et al. reported a 29% recurrence rate in their series of infants
treated with IVR, at a mean PMA of 72 weeks (102); and Lyu et al. reported
a 64% recurrence rate at a mean PMA of 43 weeks after IVR (139) (Table
53-5).
TABLE 53-5
In the many published case series and clinical trials using IVB and IVR, there
is no standardized terminology used for “recurrence” of ROP that results in
another treatment. In reviewing the published studies referenced above, there
are three clinical scenarios that result in retreatment; early failure of ROP to
regress within the first 4 weeks after initial anti-VEGF injection; recurrence
of ROP (stage 2 or 3) following initial good regression after initial anti-
VEGF injection; and persistent avascular retina in zones II and III following
good regression of ROP after initial anti-VEGF injection. In the case of early
failure, type I ROP does not regress, and a second anti-VEGF injection or
laser is performed depending on the PMA of the infant, location of disease,
and the whether or not there is fibrovascular or cicatricial ROP present. Anti-
VEGF drugs are typically not used in the presence of fibrovascular ROP to
avoid the phenomena known as “crunch” in which the drugs may accelerate
the contraction of the fibrotic extraretinal neovascularization resulting in a
tractional retinal detachment (140). In the more common scenario of
recurrence or reactivation of ROP after good initial regression of severe
disease, ROP may slowly recur weeks, months or even over 2 years after the
anti-VEGF treatment (141). Hu et al. reported a series of infants requiring
retreatment at a range of 36 to 60 weeks of PMA (142), and Karkhaneh et al.
reported a recurrence with retreatment up to 90 weeks of PMA (72). Ells et
al. reported a retreatment range of 46 to 120 weeks of PMA (102), and
Yonekawa’s group reported recurrence as late as 5 years after anti-VEGF
injection for type I ROP. These studies reinforce the importance of
meticulous long-term follow-up of infants treated with anti-VEGF to prevent
undetected cicatricial ROP and associated vision loss or blindness.
Given the high reported rate of recurrence of ROP following anti-VEGF
treatment for ROP, it may be beneficial to perform laser photocoagulation to
persistent avascular retina in zones II or III, at a PMA greater than 55 weeks
for most infants. There are, currently, no standardized protocols for follow-
up, the need for fluorescein angiography, and retreatment or treatment of
persistent avascular retina for these infants; however, we expect to see
published standardized terminology of recurrence and retreatment protocols
in the near future.
CONCLUSION
In summary, we live in an anti-VEGF era in which the treatment has great
promise for primary treatment for type I ROP. These drugs minimize the
permanent destruction of the retina, promote relatively normal angiogenesis,
and act extremely rapidly to induce regression of VEGF-mediated ROP.
Anti-VEGF treatment can be performed safely by an experienced
ophthalmologist at the bedside of an infant in the NICU. However, we still
have many unanswered questions regarding long-term neurodevelopmental
effects of these drugs due to systemic drug exposure and potentially
prolonged suppression of serum VEGF levels in these premature infants. We
need further investigation and understanding for type of anti-VEGF drug for
the premature infant, and we also await further dose-related studies to
determine the fine balance of promoting normal angiogenesis after regression
of ROP while preventing local retinal neural–vascular toxic effects. Given the
high rate of recurrence of ROP following an anti-VEGF injection, potentially
for months to years, standardized terminology of recurrence is needed, and
standardized protocols for follow-up examinations and possible retreatment
with both laser or anti-VEGF drugs are required.
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54
Evolution of Stage 4 Retinopathy of
Prematurity
Antonio Capone Jr, and Michael T. Trese
The acute vasoproliferative manifestations of retinopathy of prematurity

(ROP) are generally managed with ablation of the peripheral retina or
pharmacotherapy with anti-vascular endothelial growth factor (VEGF)
agents. In the early treatment for retinopathy of prematurity (ETROP) study,
ablative treatment resulted in unfavorable structural outcomes in 9.1% of
eyes at 9 months (1) and 2 years (2). In general terms, 1 in 10 infants treated
for ROP with laser developed advanced ROP (stage 4A, 4B, or 5). In the
CRYO-ROP study, some eyes continued to progress to an unfavorable
anatomic outcome 15 years after treatment (3). Accumulating evidence
suggests that pharmacologic treatment of ROP with anti-VEGF drugs may
achieve similar therapeutic effects as laser photocoagulation with fewer
cicatricial complications such as retinal detachment or macular dragging (4).
(See also Chapters 52 and 53.)
The current paradigm for the management of advanced ROP remains
primarily one of incisional surgery (generally vitrectomy, occasionally scleral
buckle) to relieve traction in eyes that progress to retinal detachment. This
chapter addresses the evolution to retinal detachment and serves as a
foundation for understanding the surgical management of the advanced stages
of ROP.
PATHOGENESIS OF LATE STAGES OF

The inability to obtain tissue specimens as stages 4 and 5 ROP evolve limits
basic science studies of the late stages of ROP. We speculate on possible
mechanisms for the two most commonly seen configurations of retinal
detachment in ROP. Evidence supporting these speculations is drawn from
histologic, biochemical, and clinical observations of eyes affected with
advanced ROP (stages 3, 4, and 5), from the oxygen-induced retinopathy and
variable oxygen models of ROP (5) and with the morphologically similar
retinal detachment of familial exudative vitreoretinopathy.
Retinal vascular assembly is carried out in utero in a relatively hypoxic
setting, in which VEGF is believed to drive developmental angiogenesis
(6,7). Growth factors responsible for pathologic vascular development may
also be crucial for physiologic vascularization of the retina, albeit with
different timing and at different levels (4).
The hyaloid vasculature or primary vitreous fills the developing vitreous
cavity with blood vessels that initially have many attachments to the
developing retina. During development, the attachments involute. The last of
these vascular structures to regress is the hyaloid artery. Involution of the
hyaloid vasculature and tunica vasculosa lentis occurs through apoptosis and
is generally completed by 40 weeks’ postmenstrual age. However, the
primary vitreous can persist, with a wide spectrum of clinical features
previously referred to as persistent hyperplastic primary vitreous and more
recently referred to as persistent fetal vasculature syndrome (8). (See also
Chapter 2.)
In the full-term infant, secondary vitreous consists of a dense collagen gel
that replaces the primary vitreous, except in the areas of the vitreous canals
where the former primary vitreous attachments to the retina and optic nerve
existed. The best known of these is Cloquet canal. There is an evidence that
the retinal vascular assembly and involution of the hyaloid system and tunica
vasculosa lentis by apoptosis are partly affected by VEGF (5,9) and
influenced by WNT signaling (Figures 54-1 and 54-2) (10,11). The
production and assembly of collagen gel might be a result of several cells,
including hyalocytes, and influenced by growth factor signals and
extracellular matrix mediators (12).
FIGURE 54-1 Effect of VEGF in normal vascular
development.
FIGURE 54-2 ROP vascular phases and effusive retinal
detachment phase biochemically.
In the fetus, wound healing responses and scar formation differ depending on
the developmental time in utero. For example, fetal regenerative wounds heal
without scar formation in the second and early third trimesters (13).
Transforming growth factor-β1 (TGF-β1) has been widely studied regarding
its effects on wound healing and in the eye. TGF-β1 leads to excessive scar
formation in fetal wounds and contributes to hyaluronic acid synthesis (14). It
differentially affects adult and fetal fibroblast migration and hyaluronan
synthesis (15). In addition, it down-regulates VEGF in cell culture (12).
Although direct evidence is lacking, TGF-β is present in fetal development
and conceivably could affect collagen structure, vascularity, and wound
healing in the eye. These processes could participate in the development of
retinal detachment seen in stages 4 and 5 ROP.
CLINICAL FEATURES OF RETINAL

DETACHMENT IN RETINOPATHY OF
PREMATURITY
The International Classification of ROP describes retinal detachments as
stages 4A, 4B, and 5 (16). Others have also used the term predominantly
effusive and predominantly tractional to further classify these detachments.
The predominantly effusive retinal detachment has a characteristic shape.
The retina is convex toward the examiner and smooth with fluid extending
primarily posterior to the ridge detaching the ridge and macula. This
detachment begins at the ridge. Some of the components of this subretinal
fluid are blood products (17). Accordingly, subretinal fluid may vary from
relatively clear to turbid or bright red in color. Effusive detachments are less
common now that cryotherapy is used less frequently than laser (Figures 54-
3 and 54-4).
FIGURE 54-3 Schematic drawing showing the area of
highest VEGF concentration anterior to the advancing
retinal vascular ridge. The inset shows the relationship of
developing intraretinal and extraretinal vascularization
contributing to the formation of subretinal blood (effusive
retinal detachment) and the areas of cellular elements
(primary vitreous and tunica vasculosa lentis) contributing
to predominantly tractional retinal detachment.
FIGURE 54-4 Artist’s drawing showing the
predominantly effusive 4B retinal detachment of ROP
extending from the ridge to include the center of the fovea.
Subretinal fluid may be clear to turbid.
Predominantly tractional detachment (Figure 54-5) is characterized by

peaked retinal folds pulled toward the center of the eye. The retinal ridge may
circumferentially contract and pull the retina toward the center of the eye.
Frequently, prior to detachment, a central stalk and scaffolding of spokes
extending to the retina become apparent. The network can be predominantly
posterior when associated with regression of posterior hyaloid structures,
predominantly anterior when associated with regressed structures of the
tunica vasculosa lentis, or both. This tractional form of retinal detachment is
common after laser treatment has reduced the vascularity, producing a
“quiet” eye with less effusion into the subretinal space.
FIGURE 54-5 An infant with ROP born at 24 weeks

gestational age and 430 g birth weight received a half dose
of intravitreal bevacizumab at 38 weeks PMA for stage 3
ROP with plus disease. Retinal detachment 1 week later
was characterized by circumferential hyaloidal
contraction. (Reprinted with permission from Yonekawa
Y, Wu WC, Nitulescu CE, et al. Progressive retinal
detachment in infants with retinopathy of prematurity
treated with intravitreal bevacizumab or ranibizumab.
Retina 2018;38(6):1079–1083.)
Accelerated contraction of fibrovascular proliferation, termed “crunch,” with

progressive retinal detachment is a phenomenon most commonly encountered
in eyes managed with anti-VEGF therapy. It has been described in
proliferative diabetic retinopathy (18) as well as in ROP (19). Two
progressive TRD configurations unique to acute ROP treated with anti-VEGF
agents have been reported: one relatively peripheral with a circumferential
configuration (“circumferential”) (Figure 54-5) and one very posterior with
prepapillary contraction (“prepapillary”) (Figure 54-6).
FIGURE 54-6 An infant with ROP born at 33 weeks

gestational age and 1,800 g birth weight received
intravitreal bevacizumab at 39 weeks PMA, for zone I,
stage 3, with plus disease. The patient developed
prepapillary contraction of fibrous proliferation with
associated TRD. (Reprinted with permission from
Yonekawa Y, Wu WC, Nitulescu CE, et al. Progressive
retinal detachment in infants with retinopathy of
prematurity treated with intravitreal bevacizumab or
ranibizumab. Retina 2018;38(6):1079–1083.)
MECHANISM OF PREDOMINANTLY
EFFUSIVE RETINAL DETACHMENT
Predominantly effusive 4B retinal detachment is believed to occur as a
consequence of vascular leakage into the subretinal space. Sang et al. (20)
observed vessels in the subretinal space of humans in a histologic study;
however, the pathogenesis of the subretinal neovascularization is enigmatic.
Okamoto et al. (21) developed a transgenic mouse model in which VEGF
expression was linked to the rhodopsin promoter and observed intraretinal
and subretinal neovascularization as a result of increased expression of
VEGF in the outer retina. Although direct clinical evidence is lacking, this
model supports the notion that VEGF expressed in the retina can lead to
subretinal neovascularization, leakage of fluid into the subretinal space, and
thus retinal detachment, characteristics clinically noted in the predominantly
effusive form of stage 4B retinal detachment (Figure 54-4).
MECHANISM OF PREDOMINANTLY
TRACTIONAL RETINAL
DETACHMENT
The predominantly tractional retinal detachment of ROP involves one of the
most interesting features of retinal detachment evolution in the premature
child’s eye. The involuting hyaloid system and tunica vasculosa lentis play an
important role. Eyes with predominantly tractional retinal detachments
generally develop sharply peaked retinal folds. These folds are attached to
spokes connecting to the central stalk, best appreciated with endoillumination
during vitreous surgery. The stalk always extends to the disc and can have
other direct isolated strands connecting to the retina at or posterior to the
ROP ridge. The anterior aspect of the stalk often connects to the posterior
lens surface. In eyes with profoundly immature vascular architecture, as seen
in some eyes with aggressive posterior ROP, the retina may detach flatly as a
consequence of diffuse hyaloidal contraction (22).
The traction generally has a dominant direction first toward the center of
eye and then posteriorly toward the optic nerve or anteriorly toward the lens
(Figure 54-7). With progression in the weeks prior to the infant’s original
due date and thereafter, the vitreous organizes and the circumferential folds
are often drawn centrally. Clear to turbid subretinal fluid can be present.
FIGURE 54-7 Artist’s drawing of a predominantly

tractional 4B retinal detachment of ROP with
asymmetrical traction by posterior cellular elements
(hyaloid vasculature), the symmetrical traction by
predominantly posterior cellular elements, and the
configuration resulting by traction of predominantly
anterior cellular elements (tunica vasculosa lentis). Upper
row shows appearance with indirect ophthalmoscope.
Lower row depicts cross section of condition in upper row.
In the case of crunch TRDs following anti-VEGF pharmacotherapy,

circumferential detachments likely occur as a consequence of flat
midperipheral neovascularization resulting in a taut circular detachment. For
the prepapillary configuration, the anti-VEGF agents likely induce
contraction of immature prepapillary vascular precursor cells, resulting in
traction detachment centered over the disc. The former detachments occur
primarily in eyes vascularized more anteriorly before the development of
acute ROP, the latter in eyes with more posterior disease (19).
A possible scenario is outlined. An infant is examined a few weeks after
premature birth during the period of slowed retinal vascular development and
active apoptosis of the hyaloid system and tunica vasculosa lentis. At that
time, only the hyaloid vessel or a few persistent vessels at the lens may be
visible. The continued retinal ischemia from avascular retina drives an
increase in angiogenic activity through increased VEGF action, with a
reduction in apoptosis of the hyaloid endothelium resulting in clinically
visible hyaloid vasculature and rubeosis iridis. In addition, an increase in
proteolytic enzymes might release or activate growth factors within
extracellular matrix that result in fibrosis. One possible candidate is TGF-β1,
which aggravates the scarring process in fetal tissue and allows formation of
hyaluronic acid. Increased concentration of hyaluronic acid in the secondary
vitreous then would promote liquefied vitreous noted in eyes with late stages
of ROP. This liquefied vitreous would provide less internal tamponade and
allow the retina to be pushed forward from subretinal exudate or blood
(effusive) or pulled (tractional) as RPE tries to resorb fluid. The increased
scar tissue may account for the cicatricial phase seen in the classic
“retrolental fibroplasia” of late stage 5 ROP (Figures 54-8 and 54-9).
Surgical specimens of infant eyes operated on for ROP provide evidence for
regressing vascular elements within the stalk likely from the hyaloid
vasculature (Figures 54-10 to 54-12).
FIGURE 54-8 ROP tractional retinal detachment phases.
Shown is the biochemical cycle leading to the cicatricial
stage of ROP (4–5) predominantly tractional retinal
detachment.
FIGURE 54-9 Retrolental fibroplasia in an eye with
advanced ROP.
FIGURE 54-10 Transmission electron micrograph of the
stalk material with vascular elements consistent with
hyaloid vascular structures (magnification ×4,000).
FIGURE 54-11 Transmission electron micrograph
showing oriented dense collagen around the regressing
vascular structure of the central stalk (hyaloid vasculature)
(magnification ×5,280).
FIGURE 54-12 Light micrograph showing apoptotic cells
(arrows) in a vitreous stalk removed from a patient
undergoing surgery for ROP. Cells were stained using an
ApopTag (Oncor Inc.) kit, which labels the 3'-OH DNA
ends generated by DNA fragmentation during apoptosis
(magnification ×220).
It has been our clinical observation that eyes that show more prominent
clinical evidence of the regressed hyaloid structure following laser often
develop retinal detachment. In a study of late ROP, vitreous haze and the
appearance of the vitreous organization in front of the lens in eyes treated for
threshold ROP predicted progression of stage 4A ROP (23,24). We speculate
that this is the clinical correlate of reduced apoptosis and activated TGF-β1
supporting hypocellular gel contraction. In addition, prominent or persistent
iris vessels at 34 to 35 weeks’ postmenstrual age were associated with
increased risk of threshold ROP and a higher risk of retinal detachment (25).
We speculate that this increase in iris vessel activity reflects a resurgence of
the VEGF activity.
REFERENCES
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2. Good WV; Early Treatment for Retinopathy of Prematurity Cooperative Group. The Early
Treatment for Retinopathy of Prematurity study: structural findings at age 2 years. Br J
Ophthalmol 2006;90(11):1378–1382.
3. Palmer EA, Hardy RJ, Dobson V, et al. 15-year outcomes following threshold retinopathy of
prematurity: final results from the multicenter trial of cryotherapy for retinopathy of
prematurity. Arch Ophthalmol 2005;123(3): 311–318.
4. Kang HG, Choi EY, Byeon SH, et al. Intravitreal ranibizumab versus laser photocoagulation for
retinopathy of prematurity: efficacy, anatomical outcomes and safety. Br J Ophthalmol
2019;103(9):1332–1336. pii: bjophthalmol-2018-312272.
5. Stahl A, Connor KM, Sapieha P, et al. The mouse retina as an angiogenesis model. Invest
1995;15:4738–4747.
7. Chan-Ling T, Gock B, Stone J. The effect of oxygen on vasoformative cell division: evidence
that ‘physiological hypoxia’ is the stimulus for normal retinal vasculogenesis. Invest
8. Goldberg MF. Persistent vasculature (PFV): an integrated interpretation of signs and symptoms
associated with persistent hyperplastic primary vitreous (PHPV). LIV Edward Jackson
Memorial Lecture. Am J Ophthalmol 1997;124:587–626.
9. Penn JS. Variable oxygen exposure causes preretinal neovascularization in the newborn rat.
10. Lobov IB, Rao S, Carroll TJ, et al. WNT7b mediates macrophage-induced programmed cell
death in patterning of the vasculature. Nature 2005;437:417–421.
11. Liu C, Nathans J. An essential role for frizzled 5 in mammalian ocular development.
Development 2008;135:3567–3576.
12. Mandriota SJ, Menoud PA, Pepper MS. Transforming growth factor B1 down-regulates
vascular endothelial growth factor receptor 2/flk-1 expression in vascular endothelial cells. J
Biol Chem 1996;271:11500–11505.
13. Longaker MT, Whitby DJ, Adzich NS, et al. Studies in fetal wound healing. VI. Second and
early third trimester fetal wound demonstrate rapid collagen deposition without scar formation.
J Pediatr Surg 1990;25:63–69.
14. Krummel TM, Michna BA, Thomas BL, et al. Transforming growth factor beta (TGF-beta)
induces fibrosis in a fetal wound model. J Pediatr Surg 1988;23:647–652.
15. Ellis IR, Schor SL. Differential effects of TGF-β1 on hyaluronan synthesis by fetal and adult
skin fibroblasts: implications for cell migration and wound healing. Exp Cell Res
1996;228:326–333.
16. International Committee for the Classification of Retinopathy of Prematurity. An international
classification of retinopathy of prematurity. II. The classification of retinal detachment. The
International Committee for the Classification of the Late Stages of Retinopathy of Prematurity.
Arch Ophthalmol 1987;105(7):906–912.
17. Trese MT. Surgical results of stage V retrolental fibroplasia and timing of surgery.
18. Arevalo JF, Maia M, Flynn HW, et al. Tractional retinal detachment following intravitreal
bevacizumab (Avastin) in patients with severe proliferative diabetic retinopathy. Br J
Ophthalmol 2008;92:213–216.
19. Yonekawa Y, Wu WC, Nitulescu CE, et al. Progressive retinal detachment in infants with
retinopathy of prematurity treated with intravitreal bevacizumab or ranibizumab. Retina
2018;38(6):1079–1083.
20. Sang DN, Hirose T, Soque J. Histopathology and immunopathology of retinal detachment in
retinopathy of prematurity. In: Shapiro MJ, Biglan AW, Miller MM, eds. Retinopathy of
prematurity. Chicago, IL, 1993:219–222.
21. Okamoto N, Tobe T, Hackett SF, et al. Animal model. Transgenic mice with increased
expression of vascular endothelial growth factor in the retina. A new model of intraretinal and
subretinal neovascularization. Am J Pathol 1997;151:281–291.
22. VinekarA, Trese MT, Capone A Jr; Photographic Screening for Retinopathy of Prematurity
(PHOTO-ROP) Cooperative Group. Evolution of retinal detachment in posterior retinopathy of
prematurity: impact on treatment approach. Am J Ophthalmol 2008;145(3):548–554.
2004;24:237–241.
24. Coats DK. Retinopathy of prematurity: involution, factors predisposing to retinal detachment,
and expected utility of preemptive surgical reintervention. Trans Am Ophthalmol Soc
2005:103:281–312.
25. Kivlin JD, Biglan AW, Gordon RA, et al. Early retinal vessel development and iris vessel
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Chapter 55
Treatment of Stages 4 and 5
Antonio Capone Jr, Michael T. Trese, aand M. Elizabeth Hartnett
As discussed in Chapters 52 and 54, although retinal ablation is effective in a

majority of cases of retinopathy of prematurity (ROP) requiring treatment,
approximately 10% of treated eyes progress to retinal detachment. In eyes
treated with peripheral retinopexy, detachment typically occurs either in eyes
with “skip areas” of incomplete peripheral ablation or those with aggressive
posterior ROP located in posterior zone 2 or zone 1. In eyes managed with
anti-VEGF therapy, accelerated contraction of fibrovascular proliferation
(“crunch”) and consequent traction retinal detachment occur in two unique
progressive configurations: one relatively peripheral with a circumferential
configuration (“circumferential”) (Figure 54-5) and one very posterior with
prepapillary contraction (“prepapillary”) (Figure 54-6).
Retinal detachment in infancy profoundly impacts visual development.
The untreated natural history for eyes with even minor degrees of detachment
is poor (1). The goal of surgery in infants with ROP-related retinal
detachment is to normalize anatomy insofar as is possible in the interest of
maximizing visual potential. Data from some series report poor outcomes
following attempted surgical repair (2,3). Conversely, centers dedicated to
and experienced in the management of ROP-related retinal detachment report
consistent and positive results, particularly for stage 4A detachments (4–7).
The advanced stages of ROP (stages 4A, 4B, and 5) are often poorly
understood. Common misconceptions are that (a) macula-sparing (stage 4A)
partial retinal detachments are benign, (b) surgery should be deferred until the
macula is detached, and (c) eyes with complex detachments do not attain
meaningful levels of visual acuity. Whether to perform surgery for stage 5
ROP remains a topic of controversy. The argument is that the anatomic and
visual outcomes are often so poor that such interventions are not “worth it.”
Admittedly, stage 5 ROP is daunting. The surgical learning curve is long and
steep. It is zero-error tolerance surgery, as a single iatrogenic retinal break
may mean failure. Complete retinal attachment and restoration of near-
normal posterior pole anatomy are uncommon. Neurologic comorbidity may
limit vision even when surgery is successful. However, poor visual outcome
is not invariable, and good visual outcomes are not unheard of (8–12).
Children having even limited vision make use of their vision remarkably well
(9).
Once a stage 4A detachment is noted, surgery should be performed
promptly to prevent macular detachment, particularly if it is located
temporally. More advanced detachments are surgical candidates as well,
because visual development occurs in infants with even minimal retinal
reattachment. The ambulatory vision threshold of 5/200 for adults does not
seem to apply to children deprived of vision in early infancy, as many
ambulate successfully with significantly lower levels of vision. Children with
only light perception vision object emphatically to occlusion (13). It is only
logical to assume that novel vision restorative technologies will be available
during the lifetime of an infant born today. It is reasonable to expect more
options will await a sighted child, or even one with restored retinal anatomy,
than one who does not perceive light at all.
BASIC CONSIDERATIONS
Surgical Anatomy of ROP-Related Retinal Detachment
Fibrous proliferation and contraction of neovascularization along the ridge
and onto the overlying vitreous precede traction retinal detachment.
Condensation of vitreous into sheets and strands acts as a scaffold for further
extension of the fibrovascular tissue. Traction along the retinal surface and
contraction of the posterior hyaloid face contribute to distortion of the
posterior pole architecture. The configuration of the retinal detachment in
ROP depends primarily on the location of the ridge and the orientation of
vectors of vitreoretinal traction. Tractional forces are exerted in the following
ways:
1. Intrinsic to retina (Figure 55-1). These forces consist of proliferation
intrinsic to the ridge itself causing a tractional vector that cannot be
removed surgically. Contraction of this circumferential vector results in
a radial fold. It can be addressed by scleral buckle alone when it is the
sole tractional vector located near or anterior to the equator.
2. Ridge to lens (Figure 55-2). The most common and easily
conceptualized traction vector, and the most important to interrupt
surgically, often extends circumferentially from the ridge toward the
midperipheral lens.
3. Ridge to ridge (Figure 55-3). This vector extends as a sheet across the
“mouth” of the developing funnel-shaped retinal detachment.
4. Ridge to ciliary body (Figure 55-4). This vector extends from the ridge
anteriorly toward the ciliary body.
5. Ridge to retina. This traction extends from the elevated ridge back
toward the portion of the retina just anterior to the original location of
the ridge.
6. Disc stalk. There are three variants of the proliferative tissue extending
from the optic disc: (a) a typically very adherent epiretinal sheet
extending along the retinal surface to the ridge often seen in eyes in
which the ridge is located posterior to the equator, (b) a transvitreal stalk
extending to the ridge usually seen in eyes with an equatorial ridge, and
(c) a transvitreal sheet extending from ridge-to-ridge sheet usually seen
in eyes with a ridge located anterior to the equator (Figure 55-5).
FIGURE 55-1 Contraction of proliferation intrinsic to the
circumferential ridge (arrow), resulting in a radial fold.
Some or all of these components are present in all ROP-related retinal

detachments. The configuration of the detachment is determined by the
overall tractional contribution of the multiple force vectors.
FIGURE 55-2 Proliferation extending anteriorly from the
ridge toward the lens in an anteriorly open–posteriorly
open funnel detachment. Traction originates at the ridge in
a circumferential, purse-string pattern that draws the retina
anteriorly and centrally (arrow).
FIGURE 55-3 Sheet of proliferation originating from the
ridge and extending across the “mouth” of the funnel-
shaped retinal detachment toward the contralateral ridge
(arrow).
FIGURE 55-4 Sheet of proliferation originating from the
ridge extends anteriorly toward the ciliary processes
(arrow).
FIGURE 55-5 Central “T-shaped” stalk extending
transvitreally from the optic disc (out of focus) anteriorly
to join with the ridge-to-ridge sheet (arrow).
Timing of Surgery
ROP-related detachments may appear stable in the first few weeks or months
after peripheral retinal ablation. Yet neither the stability of the partial
detachment nor visual acuity is predictable from the retinal appearance in
infants with ROP. This is particularly true for untreated eyes or those with
incomplete peripheral retinal ablation. ROP-related retinal detachments can
progress—often very gradually—for years. As the eye grows, the cicatricial
proliferation does not, and retina tethered to the eye wall is vulnerable to
stretching to the point of tractional detachment or development of a retinal
break.
Visual outcome of eyes with even partial ROP-related retinal detachment
is generally poor by age 4.5 years. In the cohort of 61 eyes from the
Multicenter Trial of Cryotherapy for Retinopathy of Prematurity study with
partial retinal detachment 3 months after threshold, only six eyes had vision
of 20/200 or better at age 4.5 years (1,14). The difficulty exists in
determining whether an eye with residual or recurrent features of threshold
ROP will develop progressive stage 4 ROP or show regression after laser
treatment. One retrospective study of retinal features in eyes treated with
laser for threshold ROP determined that ≥6 clock hours of ridge elevation, ≥2
quadrants of plus disease, and vitreous haze were predictive of progressive
stage 4 ROP and that these features should indicate to the ophthalmologist
that this patient likely will require surgery to prevent stage 4B ROP (15).
Although valuable, the study had the limitation of analyzing both eyes of
some infants who underwent bilateral surgery without accounting for
correlation between eyes. Others have found the importance of even a clock
hour of bleeding or fibrovascular proliferation associated with the ridge as
causative harbinger of progressive retinal detachment and vision loss.
In general, earlier surgical intervention is better, as the prognosis for
vision is best when the detachment is least extensive. In eyes with stage 4
detachments and thorough peripheral retinal ablation, surgery typically is
done between 38 and 42 weeks’ postmenstrual age. An exception to this rule
is the eye with stage 4 ROP in a vascularly active eye with marked plus
disease that lacked prior peripheral retinal laser ablation. In such eyes, the
risk of uncontrollable bleeding is much greater, and it is better to treat the
avascular peripheral retina first and delay surgery. Similarly, if the eye has a
stage 5 retinal detachment with significant vascular activity, one may need to
wait until 48 to 52 weeks’ postmenstrual age for the vascular activity to quiet
(16).
Equipment
Standard vitrectomy equipment of any gauge can be employed for surgery on
eyes with advanced ROP. Instrument availability and flex characteristics of
the vitrector and light pipe are the primary reasons that 23-gauge is our “go-
to” platform. Infusion port aside, we do not often use a cannula for the other
two instrument ports, or a short 2-mm cannula is used. For complex stage 4B
and most all stage 5 detachments, dissection will often be carried out with a
23-gauge or 20-gauge Trese spatula (Figures 55-6 and 55-7).
FIGURE 55-6 20-gauge Trese spatula.
FIGURE 55-7 23-gauge Trese spatula.
PROCEDURAL TECHNIQUES
Scleral buckle and vitrectomy, alone and in combination, have been used to
manage ROP-related retinal detachments. The specific interventions vary
depending on the location and type of traction causing retinal detachment.
Successful surgery in ROP is rooted in visualization, but the anatomic
nuances are sometimes not clearly apparent with the standard diffuse en face
lighting of the operating microscope. Oblique illumination with a standard
endoilluminator positioned transcorneally or intracamerally is often effective
for highlighting subtle tissue planes. The endoillumination probe is
positioned obliquely with respect to the surgeon’s viewing axis through the
operating microscope, directed at the surgical plane of interest. The probe is
held by an assistant when bimanual dissection is required and the tissue of
interest visualized using an unlit operating microscope (17). Endoscope-
assisted surgery has been described to improve visualization of vitreous
planes (18).
Scleral Buckle
We rarely perform scleral buckle for ROP-related retinal detachments, as
scleral buckle procedures do not address transvitreal traction as effectively as
vitrectomy and have several disadvantages described below. Encircling
scleral buckle may be useful when the ridge is located near or anterior to the
equator and for rhegmatogenous detachments. Radial scleral buckle elements
can be placed to relax radial folds running from the disc to the ridge as well
as for focal support of retinal breaks.
The element of choice for encircling scleral buckle in infants is a 40, 41,
or 240 silicone band. If possible, the band is placed to support the ridge on
the anterior portion of the element, placed along the greater curvature of the
eye in order to minimize migration of the band. Sutures are imbricated to
provide height. The anterior chamber is tapped with a 30-gauge needle to
normalize intraocular pressure (IOP). In eyes with substantial subretinal fluid,
drainage through an external sclerotomy, using a scleral cut down, diathermy,
and release of subretinal fluid, can be carried out to permit tightening of the
band, IOP reduction, and reattachment of the retina.
Disadvantages of scleral buckle for stage 4A ROP are the dramatic
anisometropic myopia (19) and the second intervention required for scleral
buckle transection or removal so that the eye can continue to grow. More
importantly, however, not all tractional forces can be alleviated with scleral
buckling alone. In a retrospective comparison of stage 4 ROP treated with
scleral buckle or lens-sparing vitrectomy, progressive stage 4 ROP was
prevented after one procedure in 7 of 11 eyes treated with lens-sparing
vitrectomy compared to only 5 of 16 eyes treated with scleral buckle (P =
0.03, exact chi-square) (20). Sears and Sonnie (21) found that the addition of
scleral buckling to vitrectomy did not improve outcomes.
Lens-Sparing Vitrectomy
Lens-sparing vitrectomy is our preferred approach for most stage 4A
detachments, many stage 4B detachments, and some open-funnel stage 5
detachments. Lens-sparing vitreoretinal surgery is uniquely suited to the
management of stage 4 ROP. Maguire and Trese (22) reported the technique
initially for eyes with subtotal posterior retinal detachment involving the
macula.
A light pipe or pic, vitreous cutter, and membrane peeler cutter scissors
are used in the procedure. The eye is entered through the pars plicata/anterior
pars plana at a clock hour advantageous to approaching the existing traction.
A core vitrectomy is performed addressing the organized vitreous in four
planes: transvitreal ridge to ridge, ridge to periphery, ridge to lens, and
tentacles from the central stalk of organized vitreous extending from the optic
nerve head to the ridge. It is helpful to plan the approach so as to relieve as
much traction as can safely be performed. Releasing traction from the ridge
to lens and then ridge to peripheral retina before releasing the circumferential
ridge to ridge transvitreal traction reduces the potential problem of the ridge
being pulled anteriorly toward the ciliary body once the transvitreal traction
is released. The ridge to peripheral vitreous traction then can be difficult to
address safely. When this dissection is complete, a fluid–air exchange is
performed. During the fluid–air exchange, the retina is not forced to reattach
as subretinal fluid will be reabsorbed over time once the transvitreal traction
has been removed or reduced. The sclerotomies are closed and the infant
positioned such that the air bubble displaces subretinal blood from the
posterior pole. Subretinal fluid may continue to be reabsorbed over weeks in
the postoperative period.
Infants with ROP often have multiple life-threatening comorbidities and
are the retina surgeon’s highest anesthesia risk population as described in
Chapter 57. Furthermore, ROP is a rapidly progressive bilateral
vitreoretinopathy, where both eyes commonly require prompt surgical
intervention. This had traditionally required separate visits to the operating
room for each eye.
One way to minimize the anesthesia risks for the infant, as well interval
progression of the ROP-related tractional retinal detachment, is to operate on
both eyes consecutively during the same anesthesia session. This has been
termed immediate sequential bilateral pediatric vitreoretinal surgery (ISBVS)
(23). Each eye is treated as a completely separate procedure with reprepping,
redraping, rescrubbing, new instruments, medications, and intraocular fluids.
Given the reported incidence of postvitrectomy endophthalmitis of 0.03% to
0.08%, the risk of bilateral endophthalmitis as separate unrelated events
would be assumed to be 1 in 500,000 to 10,000,000 simultaneous vitrectomy
procedures (24,25). In comparison, the risk of anesthesia-related death in
children is as high as 1 in 10,000, and all-cause perioperative mortality is 1 in
100 to 1,000 in children, even higher in neonates, and even higher in
premature infants (26,27). We do not advocate ISBVS for all infants. The
majority of infants can be treated with staged bilateral surgery. However,
ISBVS can be a powerful option for appropriately selected cases.
Lensectomy and Vitrectomy

In some stage 4B and most stage 5 ROP–related retinal detachments, there is
substantial retrolenticular fibrovascular proliferation, and the lens may be
displaced anteriorly. Such eyes require combined lens removal and bimanual
vitrectomy. The two approaches to advanced ROP-related detachments
requiring lensectomy are the lensectomy–vitrectomy technique and, less
often, the open-sky technique.
In the lensectomy–vitrectomy technique, infusing instruments, such as an
irrigating light pipe, light pic, or spatula, may be used in a two-port approach.
Alternatively, an infusion is placed through the limbus inferiorly for a three-
port approach. A portion of the iris often is sacrificed to allow full
visualization of the fibrous proliferation at the mouth of the funnel and to
minimize reproliferation along the posterior iris. Bimanual dissection
techniques are used to free the retina from traction as much as possible.
In the open-sky technique, the cornea is trephined and placed into media.
Bimanual dissection is used to circumcise the retrolental preretinal membrane
under direct visualization through the operating room microscope. Continued
dissection of membranes within the funnel of the retinal detachment is then
completed. A viscoelastic substance may be used to spread tissue planes
using a pedal-operated method to inject the substance under low pressure. A
glass coverslip can be used to provide visualization during dissection. A light
pipe may be placed on the glass coverslip to direct light to the region of
interest.
In both approaches, drainage of subretinal fluid can be performed to
evacuate blood-laden subretinal fluid and to allow the retina to settle back.
Care is taken not to forcefully attempt to reattach the retina intraoperatively
so as to avoid iatrogenic breaks. The retina is permitted to reattach over
weeks or months in the postoperative period.
ENZYMATIC VITREOLYSIS
Separation of the vitreoretinal interface during vitrectomy is challenging in
complex pediatric diseases such as ROP. Sebag has demonstrated
histopathologically that the adhesion between vitreous cortex and internal
limiting membrane of the retina is stronger in humans aged 9 months to 20
years, as compared to those 21 years or older (see also Chapter 4). The
stronger vitreoretinal adhesion in children precludes induction of posterior
vitreous detachment (PVD) during vitrectomy in most infants and young
children. Although a Weiss ring (epipapillary hyaloidal condensation) may be
apparent after attempted mechanical hyaloidal separation in children, the
separation is likely to be lamellar with residual outer cortical vitreous
remaining on the retinal surface.
In proliferative vitreoretinopathies such as ROP and familial exudative
vitreoretinopathy, adequate removal of epiretinal membranes enmeshed
within layers of formed vitreous is crucial to achieve retinal reattachment.
Autologous plasmin enzyme–assisted vitrectomy is well described (28) in a
range of pediatric and adult vitreoretinal diseases with supporting level II and
III published evidence in a variety of pediatric conditions, namely, traumatic
macular hole (29,30), stage 5 ROP (31,32), congenital X-linked retinoschisis
(33), and combined hamartoma of the retina and retinal pigment epithelium
with associated macular-involving epiretinal membrane (34).
Both human-derived plasmin enzyme (both autologous and heterologous)
and ocriplasmin (Jetrea; Thrombogenics, Leuven, Belgium) have been used
as pharmacologic vitreolysis agents to facilitate the induction of PVD during
vitrectomy in ROP (35). Ocriplasmin is a serine protease and truncated form
of plasmin that is active against substrates, such as fibronectin and laminin,
and cleaves the vitreoretinal interface to treat vitreomacular traction in adults
(36–39).
The use of plasmin enzyme to assist surgery has been reported in the
ROP population (32).
The data regarding plasmin was retrospective in nature and lacked control
arms, but it did demonstrate an acceptable safety profile and appeared to
augment vitreous separation. Further studies are warranted here. Ocriplasmin
demonstrated slightly less encouraging results in a prospective study of
pediatric vitreoretinopathies, excluding ROP (40). Although the data
presented in that study do not show a clear-cut benefit to using ocriplasmin as
an adjunct in pediatric retinal surgery, the complexity of the surgeries and the
small sample size limited the analysis, necessitating the need for future
studies and pharmacologic agents. The investigators did note that vitrectomy
appeared to be easier in the treatment arms after the unmasking. In general,
the overall ability to dissociate the hyaloid from the retina with less
mechanical force and improved visualization of surgical planes are
advantageous (see also Chapters 3 and 4).
POSTOPERATIVE CARE
Attention to the emotional needs of a child and their family is essential to a
successful outcome. This includes thorough explanation to family members
and encouragement to patients. Controllable postoperative complications
must be avoided. Ports should be closed with sutures in all but the most
straightforward core vitrectomies in children. Peripheral iridectomies should
be made routinely in aphakic and pseudophakic eyes, generally inferiorly for
safety against iris bombe and angle closure. The trust that a child has in their
retinal specialist depends upon the avoidance of inflicting pain; alienating a
child for the benefit of a better office examination is not a victory.
Postoperative day 1 evaluation consists largely of assessing for infection and
IOP. More detailed examination is obtained in the ensuing visit at 1 week
when it will be better tolerated by the child. If a child is adamant about
refusing dilated fundus examination, usually an office B-scan can suffice for
the short term until a repeat EUA.
COMPLICATIONS
The most important complications of vitreous surgery are endophthalmitis,
rhegmatogenous retinal detachment, and development of cataract. Although
endophthalmitis is potentially devastating, it is rare after vitreous surgery.
The incidence of rhegmatogenous retinal detachment in eyes of infants
undergoing lens-sparing vitrectomy for 4A ROP is low in the setting of
extensive peripheral retinopexy. Cataract, a common complication of vitreous
surgery in adults, is seen in only approximately 16% of children at 2.4 years
(41). Glaucoma is a lifelong risk as well (42). Some of the complications
encountered during follow-up, such as corneal opacity, glaucoma, and
phthisis, are more common in eyes that have had a complicated surgical
course.
OUTCOME EXPECTATIONS
The surgical goal in eyes with macula-sparing ROP–related traction retina
detachments (stage 4A ROP) is an undistorted or minimally distorted
posterior pole, total retinal reattachment, and preservation of the lens and
central fixation vision (Figures 55-8 and 55-9). Surgery for tractional retinal
detachments involving the macula (stage 4B ROP) is performed to minimize
retinal distortion and prevent total retinal detachment (Figures 55-10 and 55-
11). The practical surgical goal is to reattach as much of the retina as is
possible without causing a retinal break. Partial residual retinal detachment
and retinal drag/distortion are common in such eyes. The functional goal for
stage 4B eyes is ambulatory vision.
FIGURE 55-8 Nasal stage 4A ROP–related tractional
retinal detachment located in zone I in a 38-week-old
infant 1 month following laser.
FIGURE 55-9 Resolved nasal stage 4A ROP–related
tractional retinal detachment seen in Figure 55-8
following lens-sparing vitreous surgery. Straightening of
the vessels inferonasal to the optic disc is the only
residuum of the tractional detachment following
vitrectomy.
FIGURE 55-10 Stage 4B ROP–related tractional retinal
detachment prior to surgery. The red reflex visible
superiorly is an area of attached retina.
FIGURE 55-11 Attached posterior retina with residual
subretinal lipid in the eye seen in Figure 55-8 following
lensectomy and vitrectomy.
Vitreous surgery can interrupt progression of ROP from stage 4A to stage 4B

or 5 by directly addressing transvitreal traction resulting from fibrous
proliferation and may also reduce dragging of the macula. In experienced
hands, lens-sparing vitrectomy has been reported with primary retinal
reattachment in 90% of eyes with stage 4A ROP (4–10). Early visual
outcomes are encouraging with prompt diagnosis and management (43). In a
cohort of 11 consecutive eyes of children able to cooperate with Teller visual
acuity testing (mean age 2.5 years), Snellen equivalent was 20/80 or better in
8 eyes (73%) (5). Similar results have been reported by others for stage 4A
ROP (44), with positive outcomes also reported for eyes with stage 4B
detachments (45).
The surgical goal for stage 5 ROP is to reattach as much of the retina as
possible (Figures 55-12 and 55-13). As with stage 4B detachments, partial
residual retinal detachment is common. Form vision can be preserved
following vitrectomy for stage 5 ROP (11,46). Lower levels of vision can be
measured by detecting the perception of light in different fields of vision as
determined in different levels of illumination (13). This measurement permits
one to stratify low levels of vision and then follow low vision in infants
treated for stage 5 ROP. Maximal recovery of vision following the insult of
macula-off retinal detachment and interruption of visual development in
infants may take years.
FIGURE 55-12 Stage 5 ROP–related tractional retinal

detachment prior to surgery demonstrating retrolental
fibroplasia. The red reflex visible superiorly is an area of
attached retina.
FIGURE 55-13 Attached posterior retina with residual
subretinal lipid in the eye seen in Figure 55-12 following
lensectomy and vitrectomy.
When to Consider Another Procedure

Some stage 5 ROP eyes with advanced pathology with shallowing of the
anterior chamber have lens-cornea touch and central corneal opacification.
These cases often present late but are indeed amenable to surgical
intervention. The surgical approach involves entering through the limbus
once the anterior chamber has been deepened inferotemporally to allow
infusion cannula placement. A temporal wound is made at the horizontal
meridian with a naked trocar blade, then a vitreous cutter is inserted into the
crystalline lens, and lens material is aspirated from within the capsular bag to
deepen the anterior chamber. The iris can then be mechanically swept from
the posterior corneal surface with a cyclodialysis spatula in a side-to-side
motion or viscodissected, after which the cutter is again used to remove the
fibrotic pupillary margin, enlarge the pupil beyond the margins of the central
corneal opacity, and perform a surgical iridectomy if required. The lens
capsule is then meticulously excised. Topical cycloplegics, steroids, topical
ocular hypotensive, and hypertonic sodium chloride drops are prescribed. In
most cases, the cornea will clear sufficiently to allow posterior segment
surgery within weeks.
Reoperation is rarely necessary for eyes with stage 4A ROP. Eyes with
more severe retinal detachment (e.g., stages 4B and 5 ROP) are candidates
for reoperation. The initial procedure in such eyes continues until traction is
relieved to the point that the retina is highly mobile. Several weeks later, once
subretinal fluid is resorbed, the eye is reevaluated to determine if there is
merit to repeat intervention. Approximately half of stage 5 eyes require repeat
surgery.
CONCLUSION
Surgical intervention offers the potential for preservation of vision for eyes
with ROP-related retinal detachment, particularly if performed prior to
macular distortion or detachment (i.e., stage 4A). Surgery for traction retinal
detachments involving the macula (stage 4B ROP) is performed to minimize
retinal distortion and prevent total detachment (stage 5) with the goal of
providing ambulatory vision. Maximal recovery of vision following the insult
of macula-off retinal detachment and interruption of visual development in
infants may take years.
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2004;122(12):1816–1818.
7. Lakhanpal RR, Sun RL, Albini TA, et al. Anatomic success rate after 3-port lens-sparing
vitrectomy in stage 4A or 4B retinopathy of prematurity. Ophthalmology 2005;112(9):
1569–1573.
8. Hirose T, Katsumi O, Mehta MC, et al. Vision in stage 5 retinopathy of prematurity after retinal
reattachment by open-sky vitrectomy. Arch Ophthalmol 1993;111(3):345–354.
9. Seaber JH, Machemer R, Eliott D, et al. Long-term visual results of children after initially
successful vitrectomy for stage V retinopathy of prematurity. Ophthalmology
1995;102(2):199–204.
10. Fuchino Y, Hayashi H, Kono T, et al. Long-term follow up of visual acuity in eyes with stage 5
retinopathy of prematurity after closed vitrectomy. Am J Ophthalmol 1995;120(3):308–316.
11. Mintz-Hittner HA, O’Malley RE, Kretzer FL. Long-term form identification vision after early,
closed, lensectomy-vitrectomy for stage 5 retinopathy of prematurity. Ophthalmology
1997;104(3):454–459.
12. Kono T, Oshima K, Fuchino Y. Surgical results and visual outcomes of vitreous surgery for
advanced stages of retinopathy of prematurity. Jpn J Ophthalmol 2000;44(6): 661–667.
13. Hartnett ME, Rodier DW, McColm JR, et al. Long-term vision results measured with Teller
Acuity Cards and a New Light Perception Projection Scale after management of late stages of
2004;24:237–241.
16. Hartnett ME. Features associated with surgical outcome in patients with stages 4 and 5
retinopathy of prematurity. Retina 2003;23(3):322–329.
17. Wong SC, Capone A. Illumination Techniques for Complex Pediatric Anterior Retinal
Detachment and Associated Retrolental Plaque. Retina 2015;35(9):1905-1907.
18. Yeo DCM, Nagiel A, Yang U, et al. Endoscopy for pediatric retinal disease. Asia Pac J
Ophthalmol (Phila) 2018;7(3): 200–207.
19. Chow DR, Ferrone PJ, Trese MT. Refractive changes associated with scleral buckling and
division in retinopathy of prematurity. Arch Ophthalmol 1998;116:1446.
20. Hartnett ME, Maguluri S, Thompson HW, et al. Comparison of retinal outcomes after scleral
buckle or lens-sparing vitrectomy for stage 4 retinopathy of prematurity. Retina
2004;24(5):753–757.
21. Sears JE, Sonnie C. Anatomic success of lens-sparing vitrectomy with and without scleral
buckle for stage 4 retinopathy of prematurity. Am J Ophthalmol 2007;143(5): 810–813.
22. Maguire AM, Trese MT. Lens-sparing vitreoretinal surgery in infants. Arch Ophthalmol
1992;110:284–286.
23. Yonekawa Y, Wu W-C, Kusaka S, et al. Immediate sequential bilateral pediatric vitreoretinal
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24. Oshima Y, Kadonosono K, Yamaji H, et al. Multicenter survey with a systematic overview of
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25. Govetto A, Virgili G, Menchini F, et al. A systematic review of endophthalmitis after
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26. Flick RP, Sprung J, Harrison TE, et al. Perioperative cardiac arrests in children between 1988
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27. van der Griend BF, Lister NA, McKenzie IM, et al. Postoperative mortality in children after
101,885 anesthetics at a tertiary pediatric hospital. Anesth Analg 2011;112(6): 1440–1447.
28. Trese MT. Enzymatic vitreous surgery. Semin Ophthalmol 2000;15:116–121.
29. Margherio AR, Margherio RR, Hartzer M, et al. Plasmin enzyme-assisted vitrectomy in
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30. Wu WC, Drenser KA, Trese MT, et al. Pediatric traumatic macular hole: results of autologous
plasmin enzyme-assisted vitrectomy. Am J Ophthalmol 2007;144:668–672.
31. Tsukahara Y, Honda S, Imai H, et al. Autologous plasmin-assisted vitrectomy for stage 5
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33. Wu WC, Drenser KA, Capone A, et al. Plasmin enzyme-assisted vitreoretinal surgery in
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34. Cohn AD, Quiram PA, Drenser KA, et al. Surgical outcomes of epiretinal membranes
associated with combined hamartoma of the retina and retinal pigment epithelium. Retina
2009;29:825–830.
35. Wong SC, Capone A Jr. Microplasmin (ocriplasmin) in pediatric vitreoretinal surgery: update
and review. Retina 2013;33(2):339–348.
36. de Smet MD, Gandorfer A, Stalmans P, et al. Microplasmin intravitreal administration in
patients with vitreomacular traction scheduled for vitrectomy: the MIVI I trial. Ophthalmology
2009;116:1349–1355, 1355.e1–e2.
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38. Benz MS, Packo KH, Gonzalez V, et al. A placebo-controlled trial of microplasmin
intravitreous injection to facilitate posterior vitreous detachment before vitrectomy.
39. Stalmans P, Benz MS, Gandorfer A, et al. Enzymatic vitreolysis with ocriplasmin for
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2012;16:606–615.
40. Drenser K, Girach A, Capone A. A randomized, placebo-controlled study of intravitreal
ocriplasmin in pediatric patients scheduled for vitrectomy. Retina 2016;36(3): 565–575.
41. Nudleman E, Robinson J, Rao P, et al. Long-term outcomes on lens clarity after lens-sparing
vitrectomy for retinopathy of prematurity. Ophthalmology 2015;122(4):755–759.
42. Nudleman E, Muftuoglu IK, Gaber R, et al. Glaucoma after Lens-Sparing Vitrectomy for
Advanced Retinopathy of Prematurity. Ophthalmology 2018;125(5):671–675.
43. Prenner JL, Capone A Jr, Trese MT. Visual outcomes after lens-sparing vitrectomy for stage 4A
retinopathy of prematurity. Ophthalmology 2004;111(12):2271–2273.
44. Lakhanpal RR, Sun RL, Albini TA, et al. Visual outcomes after 3-port lens-sparing vitrectomy
in stage 4 retinopathy of prematurity. Arch Ophthalmol 2006;124(5):675–679.
45. El Rayes EN, Vinekar A, Capone A Jr. Three-year anatomic and visual outcomes after
vitrectomy for stage 4B retinopathy of prematurity. Retina 2008;28(4):568–572.
46. Trese MT, Droste PJ. Long-term postoperative results of a consecutive series of stages 4 and 5
SECTION IX
Principles of Surgery for Pediatric
Retinal Conditions
56
General Surgical Considerations and
Preoperative Management in Infants
and Children
Michael T. Trese, and Antonio Capone Jr
INFORMED CONSENT
Informed consent in the United States constitutes permission from the parent
or legal guardian for the physician to perform both examination and surgical
therapy. This consent should be obtained prior to any intervention. In the
circumstance in which parental consent is impossible, a court order can be
obtained to perform the needed interventions. In the age of antivascular
endothelial growth factor (VEGF) therapy for pediatric retinal diseases,
consent for off-label use must be recorded and institutional review board
(IRB) and investigational new drug (IND) approvals obtained.
There are many psychological issues that enter into both the consent and
the need for surgical intervention in infants and children. Parents often feel
both a responsibility to protect the child and a concern that they somehow
may be responsible for whatever problem has affected the child. Most parents
of children with known or established medical problems have learned to deal
with these issues by the time they reach the vitreoretinal surgeon, but
occasionally parents lack sensitivity to these issues. It is rare that parents
require psychological counseling, but this should be considered if the family
is having difficulty coping with the issues of a child with severe ocular
problems.
SURGICAL ANATOMY AND THE

VITREORETINAL INTERFACE
The surgical anatomy relative to each vitreoretinal disease is addressed in the
respective disease management sections in this textbook. However, some
general considerations are important. Customarily, the size of the eye gives
some insight into the severity of the intraocular disease. It is often true in
bilaterally affected individuals that the smaller eye is the worse eye. Even
eyes of 6-mm corneal diameters rarely can achieve visions as good as 20/200.
However, it also is possible for small eyes to achieve visual acuities that are
unexpected in eyes of that size. It often is helpful to advise parents of the
possibility that one eye may be smaller than the other as the child ages.
Another issue relative to the anatomy of the pediatric retina is that, in several
diseases, the retina is larger than the retinal pigment epithelial area. This
anatomic occurrence leads to retinal folds that are radial or circumferential
and that can be attached to the lens.
One of the most significant steps in vitreoretinal surgery in infants and
children is selecting the location of entry into the eye. In eyes of children
younger than 6 months postterm, particularly those with retinopathy of
prematurity (ROP), the pars plana ciliaris may not be developed and,
therefore, anterior entry is advisable (1). When performing lens-sparing
vitrectomy in ROP eyes, we enter approximately 0.5 mm posterior to the
limbus. In ROP eyes in which lensectomy is being performed, we enter
through the iris root and into the lens anterior to the lens equator to avoid
injury to any retinal fold in the posterior chamber. This also is true in
diseases, such as persistent fetal vasculature syndrome, in which the
periphery may not be seen well and peripheral anatomic anomalies are often
present. It is wise to err on the side of anterior entry in all surgical
interventions involving infants and young children to avoid inadvertent
retinal damage. We prefer using two-port vitrectomy in children and entering
in the horizontal meridians at 3 and 9 o’clock. This allows good ability to
view the retina superiorly and inferiorly, even in very small orbits. Entry into
these locations does not cause damage to the long posterior ciliary nerves or
vessels.
One of the most challenging parts of vitrectomy in infants and children is
management of the vitreoretinal interface. The vitreoretinal interface consists
of a very adherent posterior hyaloid that frequently, if not uniformly, cannot
be easily peeled or perhaps peeled at all from the anterior retinal surface.
The vitreous of many dystrophic eyes, as well as eyes with ROP, is
different than the solid vitreous of the term infant. The vitreous in ROP and
dystrophic eyes is characterized by layers of solid and liquid sheets. These
sheets can be separated from each other and frequently give the surgeon the
false impression that they have peeled the hyaloid, when in reality the hyaloid
remains along the retinal surface, or a schisis of the hyaloid occurs. It often is
necessary to leave cortical vitreous along the retinal surface when performing
vitreoretinal surgery in children and infants. In one series, it was shown that,
after removal of subretinal neovascular membranes in younger individuals,
redetachment rates and surgical results were not negatively impacted by
leaving the cortical hyaloid in that setting (D’Amico, personal
communication, 2003). In conditions characterized by progressive retinal
dragging, such as ROP and familial exudative vitreoretinopathy, it is our
impression that removing the hyaloid may be desirable to minimize posterior
segment distortion. However, hyaloid removal in children has increased risk
of retinal tears, which are difficult to treat successfully making ocriplasmin-
assisted surgery appealing (see also Chapter 4). Used in infants and children
to enzymatically cleave the vitreoretinal juncture, ocriplasmin is being tested
currently but may make primary dissection easier and safer and reduce the
late complications of postvitrectomy detachment in a somewhat opaque
hyaloid (2,3).
SURGICAL TEAM
In dealing with extremely small infants, who often have young parents and
come from poor socioeconomic settings, it is very important to adopt a team
approach to their care. This frequently requires not only skilled retinal
personnel but also an anesthesiologist who is comfortable with the care of
pediatric patients, particularly with very low-birth-weight premature infants.
It also involves interaction with the pediatrician, neonatologist, social
services, the pediatric ophthalmologist, vision therapist, vision teachers, and
the school system. These patients are a challenge preoperatively,
intraoperatively, and postoperatively, and all of the care must be coordinated
in order to realize maximal development of the visual system. It is important
to remember that these patients are “learning to see” at the time they are
receiving surgical intervention. Care is taken to avoid adding to the
amblyogenic issues, particularly with unilaterally affected infants. In
addition, it is necessary to document that the patient’s family has been told
that they must interact with other members of the team caring for the child.
This requires the surgeon interacting with the other members of the support
staff by providing good communication of the child’s structural findings and
encouraging aggressive refractive and amblyopic care.
Frequently, the surgeon is asked to predict the visual potential of the
child. This is an extremely difficult question to answer and is difficult to
predict based on anatomic results alone. We have seen several children with
extraordinarily distorted retinas, who had visual acuities between 20/20 and
20/60 (4). Conversely, we have seen children with excellent anatomic
outcomes following surgery, who have had no light perception vision. This
supports the philosophy of aggressive refractive and amblyopic care until the
child’s visual potential can be determined, at approximately age 4 or 5 years.
In some older children with anisometropia, vision can improve at older ages.
It is helpful to work with pediatric ophthalmologists who strive to seek the
best visual potential. Working hard for low levels of vision is extremely
important to the child and family as the child develops. The child with low
levels of vision (20/1,900 or less) usually is able to visually ambulate and
function at a much higher level than the adult with the same level of vision
EXAMINATION UNDER ANESTHESIA

Evaluation of the child may be very difficult in an office setting. Examination
under anesthesia (EUA) is extremely important for complete retinal
evaluation of the far periphery or even the posterior pole in a child who
cannot be examined adequately in the office setting. This may be due to the
child’s immaturity, mental status, or combative nature. One also must
determine the urgency of evaluation, depending on the tempo of the
particular disease. It is best to perform EUAs in the hospital or in an
ambulatory surgical center that is comfortable with pediatric anesthesia so
that appropriate treatment (cryotherapy, laser, or more sophisticated
vitreoretinal surgical interventions) may be applied, if necessary, without the
need for a second anesthetic procedure.
INTRAOCULAR GAS AND

INHALATION ANESTHESIA, PATIENT
POSITION, AND POSTOPERATIVE
CARE
The surgeon who cares for infants and children must realize that there are
several anesthesia concerns unique to infants and children with vitreoretinal
problems that require a different mindset than caring for adults. The use of
expanding gases poses a higher risk in children. For that reason, in children,
if a sustained gas is needed, we usually use 10% C3F8 well below the
isoexpansion percentage of 14% (5). Most often, we use air. Nitrous oxide
anesthesia is avoided since it can cause unexpected expansion and then
shrinkage of the air bubble. This also is true of air travel that allows the
bubble to expand as the air atmospheric pressure is reduced. The family
should be cautioned in this regard. The child should wear a bracelet (Figure
56-1) indicating whether air or gas is in the eye and that nitrous oxide
anesthesia and air travel should be avoided until all gas is gone from the eye.
FIGURE 56-1 Image of bracelet worn by infants and
children when air or gas is in the eye, indicating nitrous
oxide anesthesia and air travel should be avoided until all
gas is gone from the eye.
Predictably, postoperative positioning is much more difficult in children. We

often position children facedown the first night after surgery. They can be
held face down on a parent’s lap. On the second postoperative day, we have
the child remain elevated in an infant seat for all activities for 2 weeks unless
otherwise indicated to prevent the child from bumping his or her head, hitting
his or her eye, or burrowing into the mattress or pillow. Children also can
badly injure the eye with a flailing arm or fist after vitreous surgery. For this
reason, we use elbow restraints and a Fox shield to protect the eye
postoperatively. We also are concerned about amblyopia that has been
encouraged by patching and postoperative medications. In unilateral cases,
we use atropine drops in both eyes for 5 days to try and reduce the chance of
amblyopia by atropinizing just the postoperative eye.
REFERENCES
1. Hairston RJ, Maguire AM, Vitale S, et al. Morphometric analysis of pars plan development in
humans. Retina 1997;17:135–138.
detachment in the rabbit. Arch Ophthalmol 1993;111:849–854.
traumatic pediatric macular holes. Ophthalmology 1998;105:1617–1620.
4. Ferrone PJ, Trese MT, Williams GA, et al. Good visual acuity in an adult population with
marker posterior segment changes secondary to retinopathy of prematurity. Retina
1998;18:335–338.
5. Chang S, Lincoff H, Coleman DJ, et al. Perfluorocarbon gases in vitreous surgery.
57
Anesthesia for Infants and Children
P. Anthony Meza
INTRODUCTION
Most of this chapter will focus on anesthesia for the premature infant. This
chapter is no replacement for a textbook of pediatric anesthesiology, but it
does provide a description of our anesthetic techniques and the reasons we
use these techniques when caring for children and infants with retinopathy of
prematurity (ROP) and other eye diseases.
For older infants and children, standard pediatric anesthesia techniques
work well for eye surgery. Even though many of these children with a history
of extreme prematurity and very-low-birth weight (VLBW) have some
diminished pulmonary function on to adulthood, they almost always tolerate
general anesthesia very well. Most of these general anesthetics can be done
with a laryngeal mask airway (LMA).
TOXIC EFFECTS OF ANESTHETICS

AND SEDATIVES ON THE
DEVELOPING BRAIN
In the developing brain of young animals, there is known neuronal cell death
after exposure to every current anesthetic or sedative. This toxicity is
dependent on dose and exposure time and worsened by the combination of
several anesthetics. In monkeys, the maximum sensitivity to anesthetics
correlates with that of a 26-week gestation human fetus (1). Even when
extensive neuronal degeneration is detected after neonatal anesthetic
exposure in animals, several studies show no impairment in behavior, spatial
memory, or cognition of the animals in adulthood.
Most recently, research at the Mayo Clinic has shown that repeated
exposure to anesthesia and surgery in children younger than age 2 was a
“significant risk factor for the development of learning disabilities.” The
study concluded that the possibility that multiple exposures to anesthesia and
surgery may adversely affect neurodevelopment cannot be excluded (2).
Sevoflurane is the most commonly used inhalational anesthetic used for
pediatric patients. In rats, sevoflurane, with or without tissue injury, causes
long-term impairment in spatial short-term memory, but “environmental
enrichment” reduces the occurrence of long-term neurocognitive impairment
(3). However, the relevance of this research to neonates remains unclear.
Current research in animal and human studies has not been definitive in
showing whether anesthesia in infants and children should or should not be
used. A recent extensive review of this issue concluded that “There is very
little if any human evidence to support a recommendation that anesthesia and
surgery at a particular age is either safe or unsafe” (4).
For the above reasons every ophthalmologist caring for infants and very
young children should seek to limit and, when possible, delay exposure of
these patients to general anesthesia as much as their eye diseases allow.
ANESTHESIA FOR THE PREMATURE

INFANT
The premature or VLBW infant afflicted with ROP is extremely delicate and
requires great consideration and attention when undergoing general
anesthesia. An experienced practitioner in pediatric anesthesiology best meets
these requirements (5,6).
The following is an overview of the concerns of general anesthesia based
on the neonatal infant’s anatomy and physiology as organized by systems. (a)
The central nervous system has lower anesthetic requirements in neonates, an
immature blood–brain barrier, and an immature central respiratory drive with
the potential for apnea. (b) Within the cardiovascular system, cardiac output
is rate dependent; therefore, bradycardia must be treated aggressively. There
may be patency of the ductus arteriosus and foramen ovale; thus, air bubbles
must be avoided. (c) In the respiratory system, one must consider the
anatomic features of difficult intubation (relatively large head and short neck
in relation to the rest of the body) and short trachea, with risk of
endobronchial intubation increased. The possibility of damaged lungs may be
present, resulting from respiratory distress of the newborn, leaving the
VLBW infant with bronchopulmonary dysplasia (BPD). (d) Metabolic
concerns include relatively high oxygen consumption and rapid desaturation
with apnea. (e) Hematologic issues requiring attention include larger blood
volume per kilogram in relation to other age groups and increased amounts of
fetal hemoglobin, requiring higher hematocrit for adequate oxygen delivery.
(f) Pharmacologic points to remember include the possibility of immature
drug metabolism and elimination (narcotics are metabolized more slowly)
and immature hepatic enzyme function. (g) The renal system is unable to
concentrate or dilute urine; therefore, excessive sodium or water infusions
should be avoided. (h) Temperature regulation requires adequate body
warming. This is due to a high neonatal body surface area to volume ratio,
which leads to rapid heat loss. An inability to shiver contributes to this
problem. (i) Endocrine system management necessitates glucose
supplementation (7). This typically is given as glucose infusion at 4 mL/kg/h
D10W (6 to 7 kg/min).
A preoperative hemoglobin level should be near 10 g/dL. Some
premature neonates at about age 2 months will reach their physiologic nadir
of hemoglobin around 8 g/dL and are at risk of postanesthetic apnea because
of low hemoglobin levels. Many of these infants with significant airway
disease may be on diuretic therapy. If a blood specimen for electrolytes is
obtained by heel stick or other peripheral means, it may hemolyze and give a
falsely elevated potassium level. If a blood sample for potassium level is
essential, then a venous sample should be obtained.
The first and foremost concern for the anesthetist is about the degree of
respiratory compromise experienced by the VLBW infant. The degree of
respiratory compromise may be mild, in which case the child requires no
supplemental oxygen. In this scenario, the anesthetist must be aware of the
VLBW infant’s decreased functional residual capacity and increased
compliance of the neonatal chest wall. In addition, there is decreased oxygen-
carrying capacity due to residual fetal hemoglobin, which binds oxygen with
high affinity, thus reducing that available to the tissues. Physiologic anemia is
generally encountered at about age 2 months in the VLBW infant when the
hemoglobin level may be as low as 8 g/dL. Regardless of the fitness of the
VLBW infant presenting for general anesthesia, the anesthetist must always
be concerned about the possibility of postanesthetic apnea. In a recent study,
postanesthetic apnea occurred in 26% of infants younger than 44 weeks of
postconceptual age (PCA) and 3% of infants older than 44 weeks of PCA. All
VLBW infants younger than 60 weeks of PCA should be observed and
monitored for apnea for at least 12 hours after general anesthesia. All infants
older than 60 weeks of PCA with a history of prematurity who are still on
apnea monitors at home will also require apnea monitoring after general
anesthesia. Recent refinements suggest that a 35-week ex premature infant
may be sent home to an unmonitored setting when the infant has reached a
PCA age of 54 weeks and 34-week ex premature at 56 weeks PCA (8).
However, these guidelines differ among institutions, some requiring
monitoring for infants under 60 weeks of PCA.
The VLBW premature infant can present with severe respiratory
compromise, as with moderate to severe BPD. In BPD, there is a loss of the
small airways and alveoli and an increase in airway and pulmonary arterial
smooth muscle. These factors make reactive bronchoconstriction and
pulmonary arterial vasoconstriction more likely to occur. Pulmonary
ventilation–perfusion mismatch, leading to hypoxemia, can occur with any
stress, especially those brought on by general anesthesia. The lungs of the
infant with BPD will also have a variable amount of destructive fibrosis
affecting both the airways and the pulmonary vasculature. The VLBW infant
may have lung disease that ranges between mild BPD and BPD so severe that
continuous mechanical ventilation is required.
Because the VLBW infant usually will require endotracheal intubation
and mechanical ventilation early in life, the incidence of subglottic stenosis is
increased. Care must be taken in placing appropriately sized endotracheal
tubes so that subglottic stenosis is not exacerbated and subglottic edema is
avoided.
RAE oral endotracheal tubes (endotracheal tubes with a premolded
curvature at the standard lip level, named after the inventors Ring, Adair, and
Elwyn) are best not used in premature VLBW infants because the molded
curves usually place the tip of endotracheal tube below the carina, resulting in
endobronchial intubation and one-lung ventilation.
Mechanical ventilation with anesthesia ventilators can be difficult for
VLBW infants. The best anesthesia ventilators have settings for a minimum
body weight of 3 kg. These infants, who may weigh less than this, are best
ventilated in the pressure control mode. If the anesthesia ventilator has only a
volume ventilation mode available, it may be impossible to use for the
VLBW neonate. Care must be used in setting the peak inspiratory pressure
(PIP). The PIP must be set high enough to ensure adequate ventilation but
low enough to prevent barotrauma and the catastrophic complications of
pneumothorax, which can easily lead to death in the VLBW infant. When
mechanical ventilation is not easily accomplished, even if pressure control
ventilation is available, manual ventilation by hand with close attention to
PIP is recommended.
The neonate should be transported from the neonatal intensive care unit
to the operating room in an isolette so that adequate body temperature can be
easily maintained. Supplemental oxygen delivery is used as needed when
transporting the VLBW infant. Monitoring by pulse oximetry and
electrocardiography also should be available during transport of the neonate
to the operating room.
Some neonates will arrive in the operating room breathing room air;
others will arrive intubated, requiring complete ventilatory assistance.
The operating room temperature should be warmed to 27°C (80°F) prior
to arrival of the neonate. Warm air blankets are used on the operating room
table on which the neonate will be placed. Maintaining normothermia for the
neonate is essential.
After routine monitoring of the electrocardiogram, the noninvasive blood
pressure cuff, pulse oximetry probe, temperature probe, and a neonatal-sized
precordial stethoscope are placed. Then, induction of general anesthesia may
begin. We have found carefully performed general anesthesia to be safe for
the infant with ROP. A 1997 British study found that infants with ROP
undergoing cryosurgery had fewer and less severe complications receiving
general anesthesia compared to those who received only topical anesthesia
for cryosurgery (9).
When no intravenous (IV) access is available prior to anesthetic
induction, we prefer an anesthetic face mask inhalation induction with
sevoflurane, oxygen, and air mixture. Sevoflurane has the advantages, such
as rapid uptake and less noxious odor, over the currently available
inhalational anesthetic agents. Nitrous oxide, if used at all, must be
administered with extreme caution. Nitrous oxide increases the pulmonary
artery pressure in adults and older children and may increase the risk of
hypoxemia. Nitrous oxide also must be discontinued 15 minutes prior to the
injection of air into the eye.
Prior to placing the anesthetic face mask on the neonate, a roll of surgical
towels is placed under the shoulders of the neonate to extend the neck
slightly. This maneuver helps maintain a patent airway by aligning the oral,
pharyngeal, and laryngeal axes and facilitates orotracheal intubation. An
appropriately sized oral airway may be placed at this time to help maintain
the airway. After general inhalational anesthesia has been established, IV
access is sought. The level of inhalational agent (usually sevoflurane) will be
decreased to avoid hypotension during the search for a suitable vein for
establishing IV access. For infants weighing 3 kg or less who must have their
airway secured quickly, an awake intubation may be performed with or
without IV access.
Establishing IV access can be difficult, even for the most experienced
practitioner. Recommendations for success in establishing IV access in these
neonates are basic. The anesthetist managing the airway cannot be the person
obtaining IV access. Attention to the airway of the neonate requires the full
attention of that anesthetist. Adequate lighting must be available.
Appropriately sized equipment, such as a small tourniquet and small IV
catheters (24 and 22 gauge), are necessary. After a target vein is identified
and the tourniquet placed, the skin surrounding the target vein is held slightly
taut. This will help prevent movement of the target vein when the skin is
pierced with the tip of the IV cannula. The skin should be pierced 1 to 2 mm
away from the desired point of entry of the target vein to avoid having the IV
cannula pass through the target vein when the skin is pierced. Once the skin
is pierced, the target vein should be entered with a deliberate 1- to 2-mm
thrust of the IV cannula tip so that the target vein will not be pushed aside
with the movement of the IV cannula. The plastic IV cannula is advanced
when it is estimated that 1 to 2 mm of the IV cannula is in the lumen of the
target vein. It is helpful to have a T piece with Luer-Lok connected to the hub
of the IV cannula in order to allow injection of medications without using
large IV fluid boluses. If medications are administered through the injection
ports along the IV tubing, they may not reach the neonate for a long period of
time due to the low flow rate of the IV fluid required by the neonate. When
IV access must be established quickly to administer emergency medication, it
is recommended that a 25- or 23-gauge butterfly be placed in a scalp vein or
other easily visualized vein. A plastic IV cannula then can be placed after the
clinical situation is stabilized. As soon as IV access is established, we
recommend the administration of IV atropine 0.02 mg/kg to prevent
bradycardia from laryngoscopy and other causes. For example, if
laryngospasm occurs, succinylcholine can be administered intravenously
without the concern of it inducing bradycardia if the infant has been
premedicated with atropine (Figures 57-1 through 57-4).
FIGURE 57-1 Skin of the area of the target vein is held

taut as the tip of the IV cannula enters the skin 1 to 2 mm
away from the target vein.
FIGURE 57-2 Target vein is entered with a deliberate 1-
to 2-mm thrust of the IV cannula into the vein.
FIGURE 57-3 Intravenous catheter is connected to the IV

tubing with a Luer-Lok T piece.
FIGURE 57-4 Medications are administered directly at

the site of the T piece.
Only preservative-free saline can be used as a flush solution for these

neonates, because flush solutions containing benzyl alcohol can precipitate
acidosis in small babies.
For short procedures that allow easy access to the neonate’s airway, the
LMA is ideal (10). The LMA has been shown to provide a satisfactory
airway in infants with BPD during minor surgical procedures and results in
fewer respiratory problems compared to an endotracheal tube. Procedures
such as extended eye examination under anesthesia and laser surgery allow
for easy placement of an LMA and rapid reinsertion of the LMA, should it
become dislodged. We do not recommend the use of the LMA for open-eye
procedures where the patient’s entire head and neck are covered by surgical
drapes and easy access to the LMA would not be available. We have had one
extreme situation of a neonate with severe reactive airway disease
accompanying BPD. In this situation, the bronchospasm was precipitated by
intubation. An LMA was used successfully for an open-eye procedure, but
great care was taken by the surgical and anesthesia teams to secure and not
dislodge the LMA during the procedure. The use of the LMA for an open-eye
procedure is the rare exception and is not recommended unless the infant
does not tolerate intubation well.
When we are certain we have adequate ventilation via mask airway, have
established IV access, and plan to proceed with oral endotracheal intubation,
we administer IV rocuronium, 1 mg/kg, for muscle relaxation.
Prior to commencing anesthetic induction, the three sizes of uncuffed
endotracheal tubes (2.5-, 3.0-, and 3.5-mm internal diameter) used in
neonates should be available, with stylets in place. For most neonates, a
laryngoscope fitted with a Miller 0 laryngoscope blade is the instrument of
choice for intubation. The anesthetist should hold the laryngoscope handle
close to the laryngoscope blade to allow use of his or her fifth finger to apply
external pressure over the neonate’s cricoid cartilage. This pressure pushes
the glottis into view when the tip of the laryngoscope blade is pulling up on
the neonate’s epiglottis. During laryngoscopy, this maneuver will maximize
the chances of good visualization of the glottis of the neonate for successful
intubation (Figures 57-5A and B and 57-6).
FIGURE 57-5 Laryngoscope is held close to the
laryngoscope blade. (A) Grip of laryngoscope prior to
insertion. (B) Grip of laryngoscope during intubation.
FIGURE 57-6 Fifth finger of the anesthetist’s hand

holding the laryngoscope is used to place pressure over the
infant’s cricoid cartilage.
For maintenance of general anesthesia, we generally change over from

sevoflurane to isoflurane because isoflurane is less toxic when given for a
long period of time. We rarely administer any narcotic to these neonates for
ROP surgery because of their decreased ability to metabolize narcotics and
our desire to establish a normal respiratory pattern as soon as possible after
anesthesia to facilitate extubation. For neonates who will remain intubated for
a long period after surgery with general anesthesia, we titrate small amounts
of IV fentanyl.
We have encountered a few neonates who have severe reactive airway
disease and do not tolerate more than moderate to low inspired concentrations
of sevoflurane or isoflurane. In these cases, we have used low concentrations
of sevoflurane or isoflurane augmented with propofol infusion. We begin the
propofol infusion at 100 μg/kg/min and titrate up or down as the clinical
situation requires. We use IV rocuronium for muscle relaxation as needed.
CRISIS AND TREATMENT OF

BRONCHOSPASM
One of the most difficult anesthetic complications to deal with when
anesthetizing the VLBW infant for ROP surgery is bronchospasm. Although
rare, bronchospasm occurs in the VLBW infant with moderate to severe BPD
at about 1 to 3 months after birth. The bronchospasm, which probably is a
combination of airway smooth muscle and arterial smooth muscle
contraction, can be of sudden onset and lead to rapid oxygen desaturation.
Bronchospasm can be very difficult to treat. If it does not abate with positive
pressure via face mask or endotracheal tube, we administer IV epinephrine in
1-μg boluses until the bronchospasm improves or the heart rate is >215
beats/min. Sometimes, increasing the sevoflurane concentration helps relieve
the bronchospasm; at other times, the sevoflurane must be discontinued to
stop bronchospasm. It is important to emphasize that this type of sudden
severe bronchospasm is rare.
Because of limited respiratory reserve, the VLBW infant with BPD
should not receive elective general anesthesia when the infant has an upper
respiratory infection (URI). Infants and children with a URI are more likely
to have respiratory complications. These patients already have significant
respiratory compromise and should not be placed at greater risk than
necessary when receiving general anesthesia for ROP surgery. These infants
may require a full 6 weeks to recover from a URI to be suitable for elective
general anesthesia.
EXTUBATION IN OPERATING ROOM

VERSUS TRANSPORT TO NEONATAL
INTENSIVE CARE UNIT WITH SLOW
EXTUBATION
Most VLBW infants aged 4 months or older with mild to moderate lung
disease will tolerate extubation in the operating room after the ROP eye
surgery is completed. Some of the VLBW infants in this age group with
severe lung disease will require postoperative ventilatory support. Most of the
VLBW infants will require postoperative apnea monitoring, and many of
these infants will be on continuous apnea monitoring at home.
CONCLUSION
Anesthetic care of the VLBW infant for ROP surgery requires a trained
pediatric anesthetist. These patients require extra time and consideration
because of the fragility of their physical condition caused by the multiple
medical problems of extreme prematurity.
Tables 57-1 through 57-8 list the guidelines for medications, endotracheal
tubes, and LMAs we use at William Beaumont Hospital, Royal Oak,
Michigan.
TABLE 57-1 William Beaumont Hospital basic

pediatric anesthesia drugs
STP, sodium pentothal.
TABLE 57-2 Endotracheal tube size and length by

weight (wt) and age
Depth of insertion for the neonate = wt in kg + 6.0 cm. This gives length from distal tip to corner of
mouth. External diameter of the endotracheal tube approximates the size of the patient’s little finger.
Cuffed endotracheal tubes are recommended after age 6 years. Tube tip should be halfway between
thorax inlet and carina. All tubes are checked for placement by auscultation and if necessary by x-ray
film. When ventilating for life support at usual pressures, the uncuffed endotracheal tube should leak at
20-cm H2O inspiratory pressure. The general tube size rules used after age 1 year:
TABLE 57-3 LMA patient selection guidelines
LMA, laryngeal mask airway.
TABLE 57-4 MAC concentrations of common

anesthetic agents in children
MAC, minimum alveolar concentration; N2O, nitrous oxide.

TABLE 57-5 Pharmacologic therapy for acute
intraoperative bronchospasm
TABLE 57-6 Treatment of pain in the

postanesthesia care unit
PO, by mouth.
TABLE 57-7 Pediatric resuscitation medications
aMay also give endotracheally.

IO, intraosseous.
TABLE 57-8 Cardiovascular drips
aFor isoproterenol, norepinephrine, and epinephrine, 1.0 mL/h provides 0.1 μg/kg/min.
bEach I mL/h infused will provide the same number of μg/kg/min of dopamine or dobutamine (or 1.0
mL/h provides 1.0 μg/kg/min).
REFERENCES
1. Istaphanous G, Loepke A. General anesthetics and the developing brain. Curr Opin Anesthesiol
2009;22: 368–373.
Randall P, et al. Cognitive and behavioral outcomes after early exposure to anesthesia and
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surgery. Pediatrics 2011;128:e1053–e1061.
3. Shih J, et al. Delayed environmental enrichment reverses sevoflurane-induced memory
impairment in rats. Anesthesiology 2012;116:586–602.
4. Davidson AJ, Sun LS. Clinical evidence for any effect of anesthesia on the developing brain.
Anesthesiology 2018;128:840–853.
5. Gregory GA, ed. Pediatric anesthesia, 3rd ed. New York: Churchill Livingstone, 1994.
6. Cook DR, Marcy JH, eds. Neonatal anesthesia. Pasadena: Appleton Davies, Inc., 1988.
7. Seefelder C. Challenges in neonatal anesthesia. Presented at Practical Aspects of Pediatric
Anesthesia, Harvard Medical School, The Children’s Hospital, Massachusetts General Hospital,
May 22–24, 2002.
8. Cote CJ, Lerman J, Anderson BJ, eds. A practice of anesthesia for infants and children, 6th ed.
Philadelphia, PA: Elsevier, 2019:64–65.
9. Haigh P, Chiswick M, O’Donoghue E. Retinopathy of prematurity: systemic complications
associated with different anaesthetic techniques at treatment. Br J Ophthalmol
1997;81:283–287.
10. Ferrari L, Goudsouzian N. The use of the laryngeal mask airway in children with pulmonary
dysplasia. Anesth Analg 1995;81:310–313.
58
External and Minor Surgical
Procedures
Emmanuel Y. Chang, Antonio Capone Jr, and Michael T. Trese
INTRODUCTION
A broad range of noninvasive and minimally invasive surgical procedures is
necessary in the course of managing children with pediatric vitreoretinal
pathology. This chapter details the pertinent aspects of these interventions.
Improved imaging modalities and novel therapeutic targets have expanded
our understanding of the timing and roles of specific therapeutic interventions
in pediatric retinal diseases.
Many of these procedures such as laser retinopexy, cryoretinopexy, and
intravitreal therapy are performed routinely on adults in the clinical setting
with topical, subconjunctival, or retrobulbar anesthesia. However, infants and
children will usually require general anesthesia or monitored sedation for
analogous procedures. Children may also require repeated examination under
anesthesia to determine follow-up care.
EYE EXAMINATIONS IN THE

NEWBORN INTENSIVE CARE UNIT
Eye examinations of premature infants at risk of developing retinopathy of
prematurity (ROP) are routine events in neonatal intensive care units
(NICUs). The healthcare team for ROP management consists of
ophthalmologists, neonatologists, NICU nurses, and the parents. The
neonatologist determines whether an individual infant can be safely examined
in view of potential systemic complications of the examination. Nurses not
only administer mydriatics and assist with the examination but also often play
a critical role in assembling the infant’s healthcare team. Parental
involvement in keeping the examination schedule once infants are discharged
from the hospital is critical.
Scleral depressed fundus examination is performed following dilation of
the pupils, generally employing a lid speculum and a Flynn lens loop style
depressor. Prolonged examination may increase the systemic instability of the
infant. Combination cyclopentolate 0.2% and phenylephrine 1% eye drops
(Cyclomydril) are administered to each eye with a second application 2 to 5
minutes after the first. When adequate dilation is not obtained, phenylephrine
2.5% (except in the setting of systemic hypertension) and either
cyclopentolate 0.5% or tropicamide 1% may be instilled twice, 2 to 5 minutes
apart. Premature infants may be very sensitive to these topical medications
resulting in systemic hypertension and tachycardia if overadministered.
Special attention should also be given to the possibility of significant
bradycardia, arrhythmia, asystole, hypoventilation, apnea, or aspiration
during examination in particular with less stable infants. It is imperative to do
thorough but expedient depressed fundus examinations. Infants are
appropriately monitored with examinations attended by a member of the
NICU nursing staff. The examination is interrupted if the infant is not stable
and is not resumed until the infant is again stable. The examination is
terminated if recommended by the NICU nursing staff and physicians. Some
nurses will increase the oxygen flow rate on premature infants during ROP
examinations; it is important for surgeons to be aware of this increase as
significantly increased oxygen flow may sometimes mask vascular dilation
and tortuosity. It is also imperative that the nurses remember to return the
oxygen flow levels to previously established target oxygen saturation for
premature infants.
Equipment required for the eye examination includes an indirect
ophthalmoscope with condensing lenses (20D, 28D, or 30D), an infant lid
speculum, and a scleral depressor. Sterile instruments are used for each
examination. Outbreaks of epidemic keratoconjunctivitis within the neonatal
ICU have been reported from ROP rounds (1). Diligent hand washing, use of
antiseptic lotions, and changing of gloves between infants are necessary to
avoid the transmission of infectious disease from one infant to another. The
infant is swaddled to minimize body movement with an assistant holding the
infant to minimize head movement. Topical ophthalmic anesthetic (e.g.,
proparacaine HCl 0.5%) is used in both eyes. The lid speculum is placed, and
the right eye is examined first by convention. An anterior segment evaluation
is performed to evaluate both the development and the appearance of the iris
and iris vasculature, and the fundus examination of each clock hour to the ora
serrata is performed. The findings are recorded using the International
Classification of ROP nomenclature (2–4).
DIGITAL FUNDUS IMAGING IN THE

NEWBORN INTENSIVE CARE UNIT
The healthcare team behind a telemedicine approach to ROP surveillance is
the same as that described above for monitoring with binocular indirect
ophthalmoscopy (BIO). However, in this paradigm, trained NICU nurses
typically assume image acquisition responsibilities. The team concept is
particularly important in a telemedicine paradigm, because the
ophthalmologist is no longer on-site for the evaluation program. Those who
acquire images (NICU nurses or other trained personnel) make up the front
line of the evaluation program. Nurse team members and other interested
NICU nurses require training in telemedicine techniques.
Several retinal imaging cameras are available now that we will discuss
their strengths and weaknesses later in this section. All current fundus
cameras require well-dilated eyes to obtain the best imaging; the dilation
protocol is the same as for BIO.
Infants should be closely monitored for bradycardia, apnea, and
tachycardia after dilation and during the imaging process. There is no
evidence to suggest that the examination with retinal imaging is harmful to
the infant or more stressful than BIO (5) and some evidence to suggest that it
may be less stressful (6). A drop of topical anesthetic is applied to each eye.
A wire speculum (Alfonso or pediatric Barraquer) is used to open the eyelids
and then a methylcellulose coupling agent is applied to the cornea or lens
surface since all current neonatal digital fundus cameras are corneal contact-
based cameras.
Digital fundus imaging should be performed on all infants at risk for ROP
on a dedicated day once weekly as outlined in the joint statement on ROP
from the American Academy of Pediatrics, American Association for
Pediatric Ophthalmology and Strabismus, and American Academy of
Ophthalmology (7). This allows for continuous monitoring of longitudinal
changes and limits the possibility that a child will inadvertently be lost to
follow-up. Morning scheduling of imaging sessions allows problems in
image acquisition/transfer to be addressed prior to a change of shift. Also, a
confirmatory BIO examination by the ophthalmologist responsible for
oversight of ROP screening can be performed the same day, if warranted.
Six standard images (iris image, disc-centered image, and four additional
fundus peripheral images with the disc located at the 3 o’clock, 6 o’clock, 9
o’clock, and 12 o’clock positions) of each eye are recommended (8).
Following dilation, the iris is imaged first to document the extent of dilation.
This image is useful in determining whether poor image quality is due to poor
dilation or photographer skill/technique.
The lens surface is cleaned thoroughly after each patient with an alcohol
swab and allowed to air-dry prior to a subsequent examination (9). It is
imperative to ensure good sterilization measures in ROP tele-screening to
minimize the incidence of an adenovirus outbreak in this at-risk population
where their immunity is still developing. The camera should be shut down if
not in use for extended periods of time to maximize the working life of the
light source in non–LED-based lighting systems. Software updates from the
manufacturer should be installed promptly. A maintenance contract may be
recommended.
A repeat examination within 24 hours is recommended for incomplete
image sets particularly when the retinal examination suggests the presence of
ROP, images characterized by poor focus/contrast/exposure, or images
inadequate for interpretation. If a NICU is providing consistently inadequate
images, it may be necessary to schedule a repeat training session or replace
personnel. If an infant cannot be effectively imaged, urgent BIO is indicated
to rule out the possibility that the inadequate images are not a consequence of
ROP, such as for poor dilation due to iris vascular congestion, as opposed to
operator error or insufficient instillation of drops to dilate the pupils. Infants
for whom images adequate for remote management cannot be obtained,
including consistently poor pupillary dilation, images where the retinal
vasculature cannot be appropriately discerned, or media opacity require BIO.
There are now four FDA-approved retinal imaging cameras on the market
and available for telemedicine screening of ROP: (a) Retcam3 (Natus,
Pleasanton, CA), (b) Icon (Phoenix Technology Group, Pleasanton, CA), (c)
Panocam (Visunex Medical Systems Co. Ltd, Fremont, CA), and (d) 3nethra
Neo (Forus health, San Leandro, CA). Only the Retcam and Icon cameras
currently have fluorescein angiography capabilities. All cameras currently
available are mydriatic and contact-based fundus imaging systems.
Retcam is now currently in its third generation with Retcam3 camera. It
is a contact-based imaging system providing a 130-degree field of view with
interchangeable lenses for higher magnification. The Retcam3 has a
fluorescein angiography module unit using a 471-nm excitation Arc-Xenon
light source. It is important to turn off the light source when not in use to
maximize bulb life, a consideration less important for LED-based light
sources. Natus also sells a more affordable version known as the Retcam
shuttle for fundus photography without fluorescein angiography capability.
Retcam3 is a Windows PC-based operating system that acquires 1.9
megapixel images and short 2-minute videos at VGA resolution levels. It
utilizes a delicate glass fiber optic cable that can break if bent at acute angles
or stepped upon and needs to be handled with care. The illumination light
source is housed in the main unit and transmitted by fiber optic to the
handheld wired device. The Retcam3 does very well acquiring images in
light-colored fundi; however, darkly pigmented fundi are more difficult to
discern vessels from background due to poorer contrast and glare. Images are
viewed on the base console unit, which has a built-in computer and monitor.
The Icon camera by Phoenix Technology Group is also a contact-based
imaging system. It provides a 100 plus degree field of view with
interchangeable lenses for higher magnification. The angle of illumination in
the Icon camera is tighter than that of the Retcam, resulting in a brighter
illumination field and improved contrast in dark fundi. Instead of a halogen
and arc-xenon–based light source for excitation, a LED light source is used
for illumination and excitation in angiographic applications. The advantage of
the LED is the significantly extended life expectancy compared to that of
filament-based bulbs. LED light sources, however, do generate slightly
cooler toned fundus images compared to filament-based bulbs. Due to the
energy efficiency and compact nature of LED light sources, the light source
is imbedded in the handheld device, which eliminates the need for a costly
fiber optic cable. It is still a wired handheld-based system connected to a
main console unit. Similar to the Retcam3, the Icon camera has fluorescein
angiography capabilities and the ability to capture video for 2 minutes. The
images acquired are at a higher resolution than the Retcam3 with a 3
megapixel image sensor and 8 MB raw TIFF image acquisition. The base
console unit also has a built-in computer and monitor similar to the Retcam3
system.
Panocam by Visunex Medical Systems is a portable contact-based
cordless handheld imaging system. It offers a 130-degree field of view with
an interchangeable higher magnification view as well. Illumination is
provided by a LED light source; however, the Panocam does not have
fluorescein angiography capability. Currently, the Panocam provides the
highest image resolution at 13 megapixels with an 8 MB image file. Since the
device is cordless, it is very portable and does not require a base console
system to be shuttled with it as do the Retcam or Icon systems. The handheld
camera device weighs approximately 1.4 pounds and has an operating run
time of approximately 3.5 hours with various configurations for docking
stations that include a base station or a portable dock station weighing
approximately 19 pounds. The operator is able to visualize and confirm the
acquired image immediately after capture due to a build-in LCD screen on
the device.
The remaining fundus camera system to be discussed is 3nethra neo by
Forus Health. It is a portable contact-based imaging device with a fixed 120-
degree field of view. There is no option for interchangeable lenses or to
create a higher magnification view. The 3nethra neo is also a LED-based
imaging system with a camera sensor that allows 4 megapixel image
acquisitions. It does not have fluorescein angiography capability. This system
is not as portable as the Panocam system; however, it has a smaller footprint
than the Retcam3 and Icon imaging systems. This system requires a portable
notebook connected to the device to view, acquire, and review images.
The device most often employed and studied the most for routine ROP
screening is the RetCam (Clarity Medical Systems, Pleasanton, CA) family
of cameras. For ROP, a wider 100- to 130-degree lens is preferred for wide-
angle visualization of the fundus. The external lens is employed for iris and
anterior segment imaging. High-magnification imaging for optic nerve or
macular pathology is performed with either a 30D or an 80D lens. Many
viewing softwares allow for image processing capability, including
manipulation of contrast, brightness, and color saturation.
Not only does digital fundus imaging provides exact documentation of
retinal pathology and findings but also it allows physicians to detect subtle
minute serial changes that may occur that can be overlooked from a fundus
drawing. It reduces the chance of missing occult disease progression and
decreases medicolegal risk. Digital photography enables physicians to consult
and discuss unique cases with other medical specialists. Lastly, it helps
parents to better appreciate and understand the importance of the retinal
examination and their infant’s or child’s risk of serious vision threatening
conditions.
Software algorithms have also facilitated better screening and diagnostic
capacity in analyzing digital fundus images, such as the ROP tool to measure
retinal vascular dilation and tortuosity in ROP and the previous FocusROP
system now called Phoenix Connect (Phoenix Technology Group,
Pleasanton, CA), which accurately overlays and indicates zone 1 and offers
clinicians additional parameters to assess the overall state of the retina (10).
The advent of deep learning algorithms has accelerated additional aids in
tele-screening/telemedicine for retinal diseases in diabetes as well as ROP.
The Imaging and Informatics in Retinopathy of Prematurity (i-ROP)
Research Consortium developed and trained a neural network that achieved a
sensitivity and specificity of 100% and 94% for detection of pre-plus disease
or worse (11). Fully automated algorithms such as this may help improve
cost-efficiencies and offer improved baseline standardized screening
practices, especially in hospital areas where the lack of subspecialty expertise
may be present. As the ophthalmic field is rapidly developing the use of tele-
screening, one can learn from radiology colleagues how their use of
automated reading algorithms for radiographic images have helped to
augment their accuracy and efficiency in ensuring problematic disease areas
are not overlooked when evaluating hundreds of images throughout their day.
This technology can help focus attention on abnormalities identified from the
algorithm.
LASER PERIPHERAL RETINAL

ABLATION FOR VASCULAR/VEGF
MEDIATED RETINOPATHIES
Peripheral panretinal scatter laser retinopexy is performed for a variety of
retinal conditions with associated ischemia that induces neovascularization
(i.e., hypoxia-induced angiogenesis) from multiple growth factors and most
notably, vascular endothelial growth factor. Conditions in which retinal
neovascularization grows into the vitreous include ROP (see Chapter 52),
familial exudative vitreoretinopathy (FEVR) (see Chapter 6), nonaccidental
trauma (see Chapter 70), persistent fetal vasculature syndrome (see Chapters
36 and 65), and incontinentia pigmenti (see Chapter 66). Laser ablative
therapy may also be utilized in exudative retinopathies such as Coats disease.
In addition to the equipment needed for routine eye examinations, which
includes a sterile lid speculum and scleral depressor, and an indirect
ophthalmoscope with a condensing lens, balanced salt solution, artificial
tears, or methylcellulose eye drops are useful to maintain corneal clarity.
Various ophthalmic lasers (diode and argon and frequency doubled) are
available today for use in peripheral retinal ablation. The choice of the
optimal wavelength of laser depends on the application and desired treatment
target in the retina or choroid. For example, yellow wavelength is
preferentially absorbed by hemoglobin and could be advantageous to treat
and ablate Coats aneurysms (12) but highly disadvantageous in an eye with
persistent tunica vasculosa lentis, which would be cataractogenic. For the
same reason, a less scattering/less absorptive red/near-infrared wavelength
laser would be beneficial in an eye with an active persistent tunica vasculosa
lentis to minimize cataract formation.
Because of the remote possibility of laser damage to assistants or others
in the laser treatment area, special laser precautions as mandated by
Occupational Safety and Health Administration (OSHA) and facility
standards are required. Protective eyewear should be supplied to anyone in
the same room in which the laser procedure is being done. For ROP, it is
often recommended that the treatment be done in a separate area within the
NICU or in the operating room. Otherwise, all personnel in the treatment
room must wear the appropriate protective goggles for the specific laser
wavelength, and all other infants in the room must have a physical barrier to
stray laser light. In addition, if viewing windows into the nursery exist, no
observers in the hallway or adjacent rooms should be able to observe through
the windows.
Laser treatment for high-risk ROP using a binocular indirect
ophthalmoscope is still standard care in the United States despite rapid rise in
the use of intravitreal anti-VEGF therapy. Laser treating ophthalmologists
must be adequately trained and experienced, or proctored by experienced
ophthalmologists. The treatment goal is the obliteration of the
nonvascularized peripheral retina by placing photocoagulation burns that are
near confluent spots or confluent spots using a painting technique. Spots are
placed on the avascular retina from the ridge extending anteriorly to the ora
serrate without treating the ridge. The number of spots required will depend
on the area of avascular retina as well as on the optics of the condensing lens
used. (The lower diopter power will give a smaller spot size due to greater
magnification.) Scleral depression is used as necessary. The power and
duration of the laser burns should be titrated to give a photocoagulation burn
without penetrating Bruch membrane to avoid immediate hemorrhage at the
laser spot. The intense grade 1 (faint, gray-white) to grade 2 (white with gray
periphery) retinal burns are made accurate to avoid delivering excessive
energy. Increased spacing of laser is recommended along the long posterior
ciliary arteries at 3 and 9 o’clock to decrease the risk of anterior segment
ischemia. Frequent changes in laser power settings are often needed during
treatment. The type of laser, number of spots, and laser power range
employed are recorded. Subtenon or subconjunctival lidocaine (2% solution)
may be administered at the conclusion of the procedure to mitigate
discomfort induced by scleral indentation in the course of the procedure.
Topical steroid drops administered for 1 week following the laser procedure
may be beneficial to treat postprocedural inflammation. Improvement in
features of vascular activity of type 1 ROP is seen within 1 week, although
for eyes with more severe type 1 ROP (threshold characteristics) may show
no improvement. However, if no worsening is seen, an examination in
another week is performed and should show improvement in plus disease and
extent of neovascularization. If no improvement is seen after 2 weeks post
laser treatment, a thorough examination should be promptly repeated to look
for skip areas with strong consideration of fluorescein angiography to
evaluate any nonperfused retina that may require additional laser. If persistent
neovascular activity remains despite confirmation of complete ROP laser,
systemic factors such as ongoing sepsis, difficult oxygenation
status/excessive fraction of inspired oxygen (FiO2), elevated intracranial
pressure, and other multi-organ system issues are considered and may need to
be assessed. For very posterior ROP disease in zone 1, it is important to apply
laser into the temporal avascular notch if possible. These posteriorly placed
laser spots in the temporal macula location migrate anteriorly to the posterior
equatorial location as infants grow older, and the macula is spared (13).
CRYOPEXY FOR PERIPHERAL

RETINAL ABLATION AND LATTICE
DEGENERATION
Cryotherapy has been employed to achieve peripheral retinal ablation for
ROP since the 1970s (14) but has largely been supplanted by laser (15).
Indications for utilizing cryotherapy include poor fundus visibility, lack of
laser availability, and physician’s unfamiliarity with indirect laser techniques.
As with laser retinopexy, choices for anesthesia include local anesthetic,
sedation, general anesthesia, or some combination. In general, conjunctival
incision is not required. Sterile technique is employed in instances requiring
incisional cryopexy. A standard retinal cryoprobe or pediatric cryoprobe may
be employed. While visualizing the peripheral retina with BIO, a row of
applications is placed starting at the ora serrata and extending contiguously
back to the fibrovascular ridge. Post-procedural anesthesia and anti-
inflammatory agents are routinely employed due to the discomfort and
inflammation that occur as a consequence of cryopexy. Significantly greater
post-operative inflammation is seen with cryopexy compared to laser
retinopexy. Although laser retinopexy is now the preferred choice over
cryopexy for peripheral retinal ablation and barricade, cryopexy still offers
significant advantages in treating the peripheral retina in instances where
direct illumination by the laser to the retina is limited/impeded in the setting
of preretinal hemorrhage/vitreous hemorrhage and poor pupillary dilation.
FOCAL RETINOPEXY
Focal retinopexy is applied with laser or a cryoprobe depending on the lesion.
Both techniques can be used to treat peripheral retinal breaks, telangiectatic
vascular pathology in Coats disease, retinal vascular hemangiomas, and
peripheral vascular nonperfusion in intermediate uveitis, as well as small (3
mm in basal diameter and 2 mm in thickness) equatorial and peripheral
retinoblastoma lesions (see also Chapters 44 and 45). When treating larger
vascular lesions as in von Hippel-Lindau syndrome or Coats disease, a low-
power long-duration laser treatment is necessary to slowly blanch the
vascular lesion to a white appearance. Retreatment may need to be carefully
applied since larger lesions often times obtain a red appearance again despite
initial treatment in the same treatment session. If the laser power is too
intense, there is the risk of creating an iatrogenic vitreous hemorrhage or
retinal break from overlying hyaloidal contracture. Feeder vessels may also
be treated to accelerate the closure of large vascular retinal lesions. It is
important to explain to family members that a staged multiprocedural
approach in the treatment of large retinal lesions may be necessary.
In general, the techniques of application are similar to those described for
peripheral retinal ablation just above. There are two notable exceptions to this
statement: a triple freeze–thaw technique is employed in treating pars plana
fibroglial inflammatory aggregates (16) (“snowbanks”) and retinoblastoma
(17) when employing cryopexy. Also, laser-induced hyperthermia
(transpupillary thermotherapy) has generally replaced traditional
photocoagulation for the treatment of retinoblastoma (18).
LASER RETINOPEXY FOR ABNORMAL

VITREORETINAL
INTERFACE/LATTICE
DEGENERATION
Extensive peripheral laser retinopexy may also be performed prophylactically
in children who have conditions with a high incidence of rhegmatogenous
retinal detachment (19) (Stickler syndrome (20), pathologic myopia, and
juvenile retinal dialysis). Dynamic ophthalmoscopy is critically important to
identify the abnormal vitreoretinal adhesions and interfaces that may be
present in these types of eyes. Many of these vitreoretinal dystrophies that
predispose to rhegmatogenous retinal detachments have both abnormal
vitreoretinal adhesions as well as abnormal retinal tissue consistencies.
Since the retinal tissue is commonly thinner than a normal healthy eye in
patients with optically empty vitreous cavity–associated vitreoretinal
dystrophies, careful attention to the intensity and confluence of the laser spots
is very important to minimize the risk of laser-induced retinal necrosis.
Excessive laser power can create iatrogenic retinal breaks and tears, leading
to subsequent postprocedural complications, such as rhegmatogenous retinal
detachments during the first few weeks after treatment. Varying techniques
and practice patterns exist when prophylactically treating optically empty
vitreous cavity vitreoretinopathies to prevent retinal detachments such as: (a)
encircling laser with two to three rows around all lattice degeneration, (b)
diffuse PRP-scatter style laser to the entire vitreous base or eight rows of
laser from the ora serrate to about the equator, and (c) PRP-scatter style laser
to the posterior vitreous base with scattered anterior radialized rows of laser
every 2 to 3 clock hours. Most important when prophylactically treating the
lattice degeneration in these eyes is identifying and treating any areas of
retinal traction, retinal breaks, and atrophic holes that may have been difficult
to identify during the office examination due to poor patient cooperation and
limited scleral depression in children.
INTRAVITREAL INJECTION OF
PHARMACOLOGIC AGENTS
Intravitreal pharmacotherapy is becoming a rapidly used modality, especially
in the use of anti-VEGF drugs for ROP (21). Although anti-VEGF therapy
reduces vascular activity in severe ROP (see Chapters 52 and 53), many
systemic risk factors and unique considerations of anti-VEGF therapy use
need to be carefully considered.
Injecting a pharmacologic agent directly into the vitreous cavity is not a
trivial procedure, even though it is a quicker procedure than laser or
cryotherapy. Serious complications related to the injection procedure have
occurred in <0.1% of intravitreal injections, including endophthalmitis,
rhegmatogenous retinal detachments, and iatrogenic traumatic cataracts.
Many of the latter issues arise due to a lack of understanding of pediatric
anatomy because neonatal/infant eyes have much larger lenses relative to the
globe size than in adult eyes and the location of the pars plana/ora serrata
varies based on age of the patient. Furthermore, in the very young neonatal
eye or microphthalmic eye, the pars plana may have yet to develop, and
injections are performed through the pars plicata. It is important to use age-
appropriate locations when determining the location to perform an intravitreal
injection (Table 58-1) (22).
TABLE 58-1 Sclerotomy site placement for infants

and children based on disease type and age
aDetermination of ora-limbus distance is based on regression analyses of control and disease subjects,
as measured from the limbus, permitting ≥1 mm from the ora serrata; we recommend nasal
sclerotomies be placed 0.25–0.5 mm shorter than the recommended location.
bWe recommend transillumination-guided identification of the ora serrata, given the variability in ora-
limbus distance and possible anterior–posterior contraction among infants.
FEVR, familial exudative vitreoretinopathy; PFV, persistent fetal vasculature; ROP, retinopathy of
prematurity.
Reprinted from Wright LM, Harper CA, Chang EY. Management of infantile and childhood
retinopathies: optimized pediatric pars plana vitrectomy sclerotomy nomogram. Ophthalmol Retina
2018;2(12):1227–1234. Copyright © 2018 by the American Academy of Ophthalmology. With
permission.
Intravitreal injection can be performed in infants and adolescents with topical

and/or subconjunctival anesthesia. Older infants and juveniles will likely
require sedation at a minimum or a brief general anesthetic to safely perform
the injection. Volume of injection and infant systemic blood pressure are
important considerations in smaller neonatal/infant eyes because of the easier
risk to compromise retinal perfusion with intravitreal injection.
An intravitreal injection requires sterile equipment, including lid
speculum, toothed forceps, 5% povidone–iodine, balanced salt solution or
saline drops, topical anesthetic eye drops, and 2% lidocaine for
subconjunctival injection. A binocular indirect ophthalmoscope with an
appropriate condensing lens should be available to inspect post-injection if
any concerns are present. Topical anesthesia is applied to the cornea prior to
treatment (tetracaine, proparacaine, or 4% lidocaine). The lids and
conjunctival fornices are treated with topical 5% Betadine. A sterile lid
speculum is placed to hold the lids open. The conjunctiva may be retreated
with topical 5% Betadine. The drug is injected through the temporal sclera
into the vitreous cavity with a 30-gauge needle at the appropriate age-based
distance from the limbus. A sterile cotton swab is used to stem reflux of drug
while removing the needle. Normal intraocular pressure or confirmation of
retinal perfusion is important after injection. A topical antibiotic, such as a
broad spectrum antibiotic like Polytrim, may be given three times daily for 3
days. Examinations are performed within 3 to 7 days later to rule out post-
injection complications, such as infection.
Injections in neonatal or infant eyes are performed with the axis of the
needle more parallel to the pupillary-optic nerve axis than in adult injections
to avoid iatrogenic lens injury. If using a 0.5-inch needle, it is important not
to inject to the hub of the needle in order to avoid iatrogenic retinal injury. A
SAFER protocol has been suggested using a shorter injection needle (23). A
general guideline for the distance from the limbus for intravitreal injection is
0.5 mm during the neonatal period, 1.0 mm at age 6 months, 1.5 mm at 1
year, 2 mm at 2 years, and 3 mm for children 4 years and older. Recently, a
published nomogram is available based on age and disease entity (19).
An area of special attention with the rapid adoption of intravitreal therapy
is awareness of nongood manufactured practices (non-GMP) sourced drugs
such as bevacizumab. Bevacizumab is separately compounded for use, and
awareness of sterility and storage handling of the drug is important to ensure
safety and bioefficacy of the drug. Physicians should ensure appropriate
sterility testing measures were completed by the compounding pharmacy
prior to using the drug. Inappropriate storage temperature, incorrect solvent
diluents, and delayed use may also affect the biologic efficacy. Also, it is
advisable to use different lots when performing bilateral intravitreal
injections of pharmacy-compounded drugs to minimize the risk of bilateral
endophthalmitis. Additional considerations are the effects of dilution and
consistency of drug dose when injecting small volumes into an eye. Improved
injection systems for drug delivery are important for intravitreal therapies in
infant eyes.
SUMMARY
As is true of adults, children with retinal pathology will often require
noninvasive or minimally invasive interventions. For premature infants,
examination alone is a minor procedure. While anesthesia requirements
generally differ from those for analogous treatments in adults, the procedures
described above are highly effective approaches to disease management and
are standard tools in the armamentarium of the pediatric retinal specialist.
REFERENCES
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detachment. The International Committee for the Classification of the Late Stages of
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Mehta M, Adams GG, Bunce C, et al. Pilot study of the systemic effects of three different
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Association of Certified Orthoptists. Screening examination of premature infants for retinopathy
of prematurity. Pediatrics 2013;131:189.
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803–810.
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retinopathy of prematurity. Biomed Res Int 2015;2015:808639.
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59
Anterior Segment Surgery
Combinations With Posterior Segment
Diseases
Mark K. Walsh
It is not infrequent that pediatric retinal surgeons encounter anterior segment

pathology that is best addressed at the time of posterior segment surgery.
These combined surgeries most commonly entail corneal, lenticular, and/or
glaucoma surgery. Depending on the individual surgeon’s comfort level,
concomitant corneal or glaucoma surgery often is performed by an anterior
segment subspecialist colleague during the same surgery that addresses the
surgical retinal issue. One reason to perform these surgeries in a combined
fashion is to try to minimize general anesthesia exposure to pediatric patients.
In this chapter, we will briefly discuss common combined lenticular surgery
with retinal surgery, such as in the setting of persistent fetal vasculature
syndrome (PFVS), retinopathy of prematurity (ROP), and trauma. We will
also address combined corneal surgeries with retinal surgery, including
temporary keratoprosthesis (KPro) followed by retinal surgery and
penetrating keratoplasty (PKP) or the Boston KPro all in one surgical setting.
We will also briefly discuss glaucoma tube shunt procedures in conjunction
with retinal surgery. Finally, we will discuss at some length (with many
operative pearls for success) intraocular lens (IOL) implantation in pediatric
eyes that lack sufficient capsular or zonular support, for example, in eyes that
have been left aphakic from previous surgery for congenital cataract or
PFVS, or subluxed crystalline lenses, such as in Marfan syndrome.
COMBINED LENTICULAR AND

RETINAL SURGERY
Severe pediatric retinal detachments often present with a dense white
fibrovascular plaque adherent to the posterior lens, as in stage 4b/5 ROP,
Norrie disease, and severe combined anterior and posterior PFVS. To
approach the retina safely, one must use an anterior approach usually through
the limbus versus the less frequently performed open-sky technique. The
limbal method can either be done with two ports—infusion in one hand and
vitrector in the other—or three ports with an infusion line placed into the
anterior chamber through the limbus. The lens is then aspirated with the
vitrector, and the lens capsule is gently removed with forceps. Depending on
the pathology, iris hooks can be used to increase the exposure/view or in
some cases, a partial iridectomy is performed. Once exposure and visibility
are optimized, the retinal detachment is addressed, and the eye is typically
left aphakic. Future secondary IOL placement can be considered in rare
instances, for example, mild PFVS with an epipapillary traction retinal
detachment that resolves after the initial surgery without significant macular
disease sequelae. Of note, even if there is no retinal detachment detected on
ultrasound, if there is no view of the pars plana in the setting of PFVS due to
an opaque retrolental plaque, one should still perform the lensectomy and
removal of the plaque via the limbal route and not the pars plana. The pars
plana can be nonexistent or rudimentary in PFVS with the retina inserting in
some areas directly onto the pars plicata. Therefore, placing sclerotomies
without direct visualization of the pars plana creates high-risk situations for
iatrogenic retinal breaks. In some cases, for example, traumatic cataract, an
anterior chamber maintainer/infusion can be placed and the lens then
aspirated through a limbal paracentesis using the vitrector. At that point, if
the view posteriorly is good enough to visualize the pars plana, the case can
then be converted to a pars plana vitrectomy (PPV) in order to optimize
visualization to address the posterior pathology, such as vitreous hemorrhage
or retinal detachment.
COMBINED CORNEAL AND RETINAL

SURGERY
Corneal opacities that preclude a sufficient posterior view for retinal surgery
may necessitate combined corneal PKP with vitrectomy. This is most often
encountered in the setting of traumatic corneal scarring, perhaps after initial
penetrating repair, with subsequent nonclearing vitreous hemorrhage or
retinal detachment. After placement of vitrectomy trocars in the pars plana,
the cornea can be trephined, and an Eckardt (silicone) temporary KPro
(DORC) or Landers (polymethylmethacrylate or PMMA) temporary KPro
(Ocular Instruments) is sewn in place with several interrupted 10-0 nylon
sutures. The KPro should afford a clear view to the posterior segment
pathology to allow surgical treatment. Finally, a donor corneal button is
sutured in place of the temporary KPro. Central corneal opacities can also be
seen in the setting of severe pediatric retinal detachments, such as stage 5
ROP, due to a flat anterior chamber and lens–corneal touch. Staged surgery
with initial lensectomy, partial iridectomy, and reformation of the anterior
chamber can sometimes lead to corneal clearing and permit retinal surgery 2
to 4 weeks later without the need for a corneal transplant (1).
In adults, Descemet stripping automated endothelial keratoplasty
(DSAEK) and Descemet membrane endothelial keratoplasty (DMEK) have
largely supplanted PKP for corneal endothelial disease causing corneal
clouding because of improved outcomes versus PKP. Instead of replacing the
full thickness of the cornea, DSAEK and DMEK are partial-thickness
transplants that replace primarily the corneal endothelium. Because pediatric
PKP success rates are estimated to be only approximately 50% (range, 15%
to 100%) and postoperative care can be challenging and often requires many
examinations under anesthesia, DSAEK and DMEK have been performed in
pediatric patients recently (2–4). Combined DSAEK with PPV and IOL
implantation has been recently reported for adult patients with bullous
keratopathy and aphakia or with dislocated crystalline lens (5). In these cases,
the cornea is cloudy, but the view is sufficient enough to perform vitrectomy
followed by endothelial keratoplasty. Thus, it may also be possible to
perform combined endothelial keratoplasty with PPV in select pediatric
patients.
KERATOPROSTHESES
The Boston type 1 KPro has been used in select high-risk children with
severe corneal disease given the high rate of PKP rejection/failure in pediatric
patients and the potential for rapid visual recovery and low concern for graft
rejection. The KPro is an artificial cornea consisting of a PMMA optic and
back plate with an intervening corneal graft ring. The first cases in children
were reported in 2006 (6). Placement of the KPro is often accompanied by
lensectomy due to the short distance between the lens and cornea in children,
tube shunt placement because of a high risk of glaucoma in anterior segment
dysgenesis disorders, at least an anterior vitrectomy, and sometimes PPV, if
there is additional posterior pathology. Unfortunately, severe postoperative
complications are much more frequent in children with KPros than adults
with a recent large series reporting no light perception (NLP) vision in 45%
of children at last follow-up (7) due to high rates of endophthalmitis, retinal
detachment, and KPro extrusion. Thus, it has been suggested that without
significant advances to improve KPro outcomes in children, the risks of
placing KPros in children currently outweigh the potential benefits (7,8),
although this viewpoint is controversial with some still advocating for KPro
placement in select pediatric eyes.
COMBINED GLAUCOMA DRAINAGE

DEVICE IMPLANTATION AND
VITRECTOMY
Complicated pediatric retinal disease is frequently accompanied by
glaucoma. Often, glaucoma develops after primary vitrectomy (9) and is
subsequently treated surgically, but on occasion, glaucoma drainage device
implantation is performed in combination with a primary vitrectomy. If the
plan is to place the tube shunt into the pars plana instead of into the anterior
chamber, then a vitrectomy and lensectomy, unless the eye is already aphakic
or pseudophakic, are performed either before tube shunt placement or
together in a combined procedure. There is some argument that pars plana
placement of tube shunts should actually be preferred over anterior chamber
placement in small pediatric eyes, which have shallow anterior chambers that
increase the risk of tube–corneal endothelium contact and subsequent corneal
failure (10). In the most complicated cases, glaucoma drainage device
implantation and vitrectomy are also combined with corneal surgical
placement of either a PKP or KPro.
COMBINED SUTURELESS
INTRASCLERAL INTRAOCULAR LENS
FIXATION AND PARS PLANA
VITRECTOMY
Eyes that lack sufficient capsule to support an in-the-capsular bag or sulcus-
placed IOL or with significant zonular dehiscence present unique surgical
challenges. Historically, in these instances in adults, IOLs have either been
sutured to the iris or sclera or an anterior chamber IOL (ACIOL) is placed.
For multiple reasons, these techniques are not ideal for pediatric eyes. Sutures
have limited life spans and can eventually break or erode, leading to late IOL
dislocation (11–13), which is especially troubling in children who need
decades of IOL stability. ACIOLs can lead to corneal decompensation and
uveitis–glaucoma–hyphema syndrome, complications, which are likely more
common in smaller pediatric eyes with shallow anterior chambers; therefore,
ACIOLs have largely been avoided in children. Recently in adults, sutureless
intrascleral (SIS) fixation of three-piece IOLs has gained popularity given
their potentially lower risk of side effects and decreased complexity and
surgical time relative to scleral sutured IOL techniques. Additionally, SIS
IOL fixation techniques are more versatile than ACIOLs or iris-fixated IOLs
in that they can be performed in eyes with severely damaged or even absent
irides. There are several surgical variations of SIS IOL fixation (14–16), but
they all involve fixating the haptics of a three-piece IOL into the sclera
without the requirement of sutures to hold the haptics in place. For a variety
of reasons, aphakic children may not always be tolerant of aphakic contact
lenses, and aphakic spectacles create image distortion and prismatic effects
(17). Thus, since 2011, the author has been performing SIS IOL fixation in
children who are aphakic and contact lens intolerant without good capsular or
zonular support or in children who have subluxed crystalline lenses with
zonular dehiscence, such as in Marfan syndrome (18). Fortunately, these
IOLs have been remarkably well tolerated without any significant
postoperative complications such as IOL dislocation, endophthalmitis, haptic
erosion, or retinal detachment with up to 8 years of follow-up (Walsh MK,
unpublished data, 2020). Recently, the author has moved to a novel SIS IOL
fixation technique that is transconjunctival in which the scleral tunnels for the
haptics are made with 27-gauge valved trocar cannulas with subsequent
melting of the tips of the haptics to form bulbs or flanges to theoretically
further decrease the risk of haptic slippage (19,20). As an added benefit,
because this surgery is transconjunctival, the surgery itself is quicker, and
postoperatively patients are more comfortable and tend to heal faster.
Flanged Sutureless Intrascleral Fixation of IOL

Flanged SIS IOL fixation was first described by Yamane and colleagues (21)
in adults. It involves fixating the haptics in the anterior chamber in 30-gauge
needles and then simultaneous externalization of the haptics. The author
found it more challenging to work in the smaller anterior chamber space than
posterior to the iris and thus modified this flanged technique by making the
scleral tunnels with 27-gauge valved trocar cannulas (see supplemental
Surgical Video 59-1 with voiceover annotation), which also allowed for
externalization of one haptic at a time (19). The author typically performs a
complete 25-gauge PPV in these cases, so three 25-gauge valved cannulas are
placed to start the case. Then before placing the infusion line, the scleral
tunnels are made using 27-gauge valved trocar cannulas (Figure 59-1). It is
easier to make the scleral tunnels with the eye firm at the beginning of the
case versus after connecting the infusion line as fluid can escape the eye
through the infusion line, thus softening the eye when applying pressure to
the eye during 27-gauge cannula placement. Typically, the scleral tunnel
openings are made at 12:00 and 6:00 for ergonomic reasons but also because
the corneal white-to-white distance is typically shorter vertically than
horizontally, so the IOL can be safely placed more posterior to the limbus if
the haptics are oriented in the vertical meridian. However, other meridians
can be used. The 12:00 and 6:00 clock hours at the limbus are marked with
an eight-ray corneal marker to ensure the scleral tunnel openings are exactly
180 degrees apart. 14 mm is then measured with a caliper at 12:00 and 6:00,
which usually is about 2 mm posterior to the limbus in pediatric eyes. This
measurement is done to estimate the furthest posterior the IOL can be safely
placed to decrease iris–IOL contact. Most three-piece IOLs have haptic-to-
haptic diameters of 13 mm, but stretching them to 14 mm does not seem to
place undue stress on them. The superior tunnel is made with the 27-gauge
valved trocar cannula blade heading to the surgeon’s left (if seated at the top
of the patient’s head) and limbus parallel and obliquely at approximately 20
degrees to make a tunnel of approximately 2 mm in length. The 27-gauge
valved trocar cannula is placed after displacement of the conjunctiva both
anteriorly and distally with a cotton swab to decrease the risk of
postoperative haptic erosion or extrusion through the conjunctival opening in
the early postoperative period. The inferior scleral tunnel is made in similar
fashion but directed to the surgeon’s right. If working on the right eye,
rotating the eye slightly temporally will help facilitate trocar cannula
placement if the patient’s nose gets in the way. Additionally, if working on
the patient’s left eye, then placing the infusion line in the inferonasal
quadrant instead of the typical inferotemporal quadrant decreases the chance
that the infusion line interferes with the 27-gauge trocar cannulas when
making the scleral tunnels, but the inferotemporal quadrant works well too. In
aphakic eyes or eyes with dislocated IOLs (Figure 59-2), the author then
performs a complete transconjunctival 25-gauge PPV to decrease the risk of
iatrogenic retinal breaks given that much of the work will be done in the
anterior vitreous space. In cases of subluxed crystalline lenses, as in Marfan
syndrome, a pars plana lensectomy is performed prior to a PPV typically just
using the 25-gauge vitrector to remove the native lens.
FIGURE 59-1 Flanged IOL fixation cannula positions.
The teal colored cannulas are typical 25g valved cannulas
set up in a standard 3-port vitrectomy fashion with the
infusion line placed in the inferonasal cannula (top right
of the drawing) in this left eye. The purple colored 27g
cannulas are used to create the scleral tunnels for haptic
fixation and are positioned exactly 180 degrees apart and
are directed limbus parallel and in opposite directions.
FIGURE 59-2 Dislocated three-piece intraocular lens in a
14-month-old boy with history of aniridia, congenital
cataract, glaucoma, contact lens intolerance, and bilateral
amblyopia. This IOL was successfully repositioned via
sutureless scleral tunnel fixation.
FIGURE 59-3 Handshake maneuver with 2 forceps. A
25g broad-based platform forceps is placed through the
temporal limbus in this left eye and grasps the trailing blue
haptic which is then handed to the 27g broad-based
platform forceps in the superior scleral tunnel 27g purple
cannula. The end of the haptic is grasped with the 27g
forceps, then the purple cannula is slid up the shaft of the
27g forceps before gentle externalizing that superior
haptic.
FIGURE 59-4 Flanged IOL fixation haptic melting to
form bulbs or flanges. While holding the haptic tip with a
smooth forceps, the haptic is gently lifted off the surface
of the conjunctiva then melted to form a bulb or flange
using disposable thermal cautery. The haptic is not
touched directly but simply bringing the cautery tip close
to the haptic tip will melt it. Care is taken not to touch
ocular structures or the drape with the cautery. The haptics
are then pushed back under the conjunctiva and into the
mouth of the scleral tunnel.
After the vitrectomy and scleral tunnels have been completed, attention is
then turned to the IOL. In the case of a dislocated three-piece IOL, it is
elevated into the anterior vitreous cavity with a soft tip cannula using the
extrusion function. The soft tip cannula can be rotated to expose the tip of
one of the haptics into or just posterior to the pupillary plane. A broad-based
platform forceps (e.g., 27-gauge Grieshaber Maxgrip, Alcon) is then inserted
into one of the 27-gauge scleral tunnel cannulas, and just the tip of the haptic
is grasped. A 0.3-mm forceps is then used to slide the 27-gauge cannula up
the shaft of the Maxgrip forceps before gently externalizing the haptic. The
second haptic can sometimes be easily visualized posterior to the iris and can
be externalized in similar fashion, but more frequently, it is easier to use the
posterior visualization lens (e.g., BIOM) with a light pipe in one hand and the
broad-based platform 27-gauge forceps in the other hand to grasp and then
externalize the second haptic. Depending on the angle between the haptic and
forceps shaft, on occasion, the haptic will slip out of the tunnel when trying
to externalize it. Typically, it is simple just to place the forceps right back
into the scleral tunnel to attempt externalization again without having to
reinsert the 27-gauge cannula.
Flanged SIS IOL Fixation With IOL Exchange or

Secondary IOL Placement
In an aphakic eye or dislocated one-piece acrylic IOL, a beveled 3-mm clear
corneal wound (CCW) is created at the limbus at approximately 10:30 with a
3-mm keratome. A temporal limbal paracentesis is created with a 15-degree
blade. A cohesive viscoelastic followed by a dispersive viscoelastic is
injected into the anterior chamber through the paracentesis to protect the
corneal endothelium. If residual lens capsule is noted in the pupillary axis, it
is removed with the vitrector. It is not necessary to remove all capsular
remnants and may theoretically be advantageous to leave some peripheral
lens capsule to buffer the anterior IOL surface from the posterior iris. If there
is a one-piece acrylic IOL in the eye, it will need to be exchanged. It can be
elevated into the anterior vitreous using a soft tip cannula on extrusion. Using
IOL forceps through the CCW, one haptic can be externalized and grasped
with a 0.3-mm forceps, and then the IOL forceps are placed into the anterior
chamber to span the optic at which point it will deform and slide out of the
CCW without having to cut the IOL. Alternatively, IOL scissors can be used
to cut the IOL optic approximately 80% of its width and then rotate it out of
the CCW. A three-piece acrylic IOL is then loaded into an injector cartridge
and injected through the CCW with the leading haptic posterior to the iris.
The trailing haptic is placed into the CCW or anterior chamber with
McPherson forceps. Using a handshake maneuver (Figure 59-3) with a 25-
gauge blunt broad-based platform forceps (e.g., Alcon Grieshaber Maxgrip)
in one hand through the temporal paracentesis, the trailing haptic is gently
grabbed, and the tip of the haptic is directed to the 27-gauge forceps in the
superior 27-gauge valved cannula. The superior haptic followed by the
inferior haptic are then externalized as described above.
Flanged SIS IOL Fixation With One-Piece PMMA

IOL
In the case of a dislocated one-piece PMMA IOL, one can consider
exchanging the IOL but that requires at least a 6-mm incision as this IOL
cannot be cut or folded due to its rigid nature. One could also suture-fixate it
to the sclera using 9-0 polypropylene suture (Prolene, Ethicon), which
probably has a longer life span than 10-0 polypropylene. The author has
successfully fixated one-piece PMMA IOLs using the flanged SIS IOL
fixation technique in adults but not in children. Caution should be used if
attempting flanged SIS fixation of a one-piece PMMA IOL, because these
haptics do not tend to form bulbs when heated but instead melt into more
irregular ribbon-like structures; thus, care should be taken not to make these
flanges too large. Additionally, both the haptics and optic–haptic junction are
relatively stiff; thus, the haptic-to-haptic diameter is fixed.
Forming the Haptic Bulbs or Flanges and Finishing the

Surgery
After successful externalization of both three-piece IOL haptics, a disposable
handheld fine tip high heat thermal cautery pen (Bovie) is used to melt the
distal 1 to 2 mm of the haptic to form bulbs or flanges (Figure 59-4).
Alternatively one can melt each haptic as soon as they are externalized. Care
is taken not to touch the tip of the haptic, which leads to haptic distortion, but
instead get close to the tip so that it melts into a nice bulb/flange. Often, the
tip of the haptic is sitting in a thin layer of balanced salt solution (BSS) on the
surface of the eye, which acts as a heat sink. Thus, with one hand, the haptic
tip is elevated off the conjunctiva and BSS with a smooth forceps (e.g.,
McPherson or tying forceps) to facilitate melting. Care is taken not to touch
the cornea or drape with the hot cautery to mitigate surgical site fire risk. The
haptic tips are then gently pushed under the conjunctiva and into the mouth of
the scleral tunnel with the McPherson forceps. The author prefers to place the
flange/bulb just into the mouth of the tunnel so that a thin layer of sclera
covers the bulb but the blue of the haptic is still visible. Postoperatively, one
should see some of the blue of the haptic through the conjunctiva and scleral
roof (Figure 59-5). Pushing the haptics too deeply into the tunnel runs the
risk of the haptic completely slipping out of the tunnel and back into the eye.
If the haptics have not been bent and the tunnels are exactly 180 degrees
apart and the same distance from the limbus, the IOL should be well centered
without tilt every time.
FIGURE 59-5 Ideal postoperative positions of the distal

flanged blue haptic ends. They are easily visualized under
the conjunctiva and are covered with the thin scleral roof
of the scleral tunnel mouth/opening. The blue of the
haptics is still visible indicating the haptic has not been
pushed too far into the scleral tunnel which can lead to
higher risk for IOL dislocation.
At this point, a 10-0 polyglactin (Vicryl) suture is placed in the CCW and the
knot rotated and/or buried posteriorly, and viscoelastic is removed from the
anterior chamber through the temporal paracentesis using the vitrector. The
author prefers absorbable 10-0 polyglactin, which is monofilament and thus
less irritating to the eye versus larger polyglactin sutures, over 10-0 nylon so
that the corneal suture does not need to be removed postoperatively. The
posterior visualization lens (e.g., BIOM) is then used to facilitate gentle 360-
degree scleral depression to ensure the absence of any iatrogenic peripheral
retinal breaks. Often times, the irides are poorly dilating in these cases, but if
still widely dilated, then acetylcholine chloride intraocular solution (Miochol-
E, Novartis) is injected into the anterior chamber to cause miosis. The
vitrector is then placed through the temporal paracentesis, and a small
peripheral iridectomy is created usually at 12:00. This has eliminated the risk
of postoperative reverse pupillary block. The three 25-gauge cannulas are
then removed, and unlike in adults, in children, these are sutured
transconjunctivally either with 6-0 plain gut, or small conjunctival openings
are made over the sclerotomies and are sutured with 7-0 polyglactin followed
by conjunctival closure with 6-0 plain gut. If the IOP does not seem to be
maintained, a sterile filtered air bubble can be injected into the anterior
chamber followed by an air bubble injected posteriorly on a 30-gauge needle
through the pars plana. The air bubbles help to seal any occult leakage sites.
If an air bubble has been injected, the patient is asked to remain supine until
the first postoperative day. Postoperatively, dilation is avoided indefinitely as
long as the postoperative pupil is at least 3 mm in diameter and the peripheral
retina can be well visualized. This decreases the risk of postoperative iris–
IOL capture, which can happen if the pupil is widely dilated and the
peripheral iris is not adequately supported by residual lens capsule.
Fortunately, the author has not seen this complication in his pediatric series.
Although any three-piece IOL should work for SIS fixation, the author
has the most experience with the Alcon MA50BM, which has a 6.5-mm
optic, which is larger than most optics, and PMMA haptics. The Zeiss CT
Lucia IOL has polyvinylidene fluoride (PVDF) haptics, which appear to be
stronger and upon melting form larger and more uniform flanges,
characteristics that may offset the slightly smaller 6.0-mm optic. The author
has also used the Abbott AR40e three-piece IOL as it has a rounded anterior
optic edge which may theoretically decrease the risk of postoperative cystoid
macular edema. In terms of picking the specific IOL power, the author
always enlists the help of the referring pediatric ophthalmologist as it is a
balancing act between leaving the child too hyperopic initially versus future
eye growth leading to myopia later in life.
CONCLUSIONS
As pediatric retinal surgeons, our patients often have multiple ocular
problems that need to be addressed surgically, including corneal and
lenticular disease, as well as glaucoma. Combined anterior and posterior
segment surgery can theoretically limit the risk of repeated general anesthesia
needed for separate surgeries to address anterior and posterior disease
sequentially. Additionally, eyes that need surgery for secondary IOL
placements in the absence of capsular/zonular support or have subluxed
crystalline lenses due to zonular dehiscence are becoming more commonly
seen in vitreoretinal practices. Newer flanged SIS IOL fixation techniques
combined with PPV offer new hope for aphakic children, who are contact
lens intolerant and at risk for amblyopia.
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2006;141:308–312.
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60
Surgical Approaches to Infant and
Childhood Retinal Diseases: Scleral
Buckles
Jessica G. Lee, and Philip J. Ferrone
SURGICAL APPROACHES TO INFANT

AND CHILDHOOD RETINAL
DISEASES: SCLERAL BUCKLES
Scleral buckling is generally preferred as the first-line approach to treat most
pediatric retinal detachments. The surgical management of infant and
childhood retinal diseases often involves the placement of a scleral buckle
either alone or combined with vitreous surgery. Over the past two decades,
advances in microsurgery have popularized pars plana vitrectomy over scleral
buckles for the treatment of retinal detachments in adults. According to
Medicare data, scleral buckling has sharply declined, and the preference for
retinal detachment repair has shifted further toward vitrectomy with a
distribution of 83% vitrectomy, 5% scleral buckling, and 12% pneumatic
retinopexy in 2014 (1,2). Despite this trend that favors vitrectomy over
scleral buckling for the treatment of retinal detachments in adults, in the
pediatric patient population, scleral buckling remains an important and
valuable surgical procedure. Scleral buckling continues to be the most
prevalent (61% to 76%) initial procedure in pediatric retinal detachment cases
without advanced proliferative vitreoretinopathy (PVR) (3).
Rhegmatogenous retinal detachment (RRD) is a major cause of vision
loss in adults. In children younger than 18 years of age, RRD occurs much
less frequently and accounts for only 2% to 6% of all cases of retinal
detachment (4–7). Compared to adults, children present more often with
macula-involving RRD, PVR, longer duration of retinal detachment, and
worse visual acuity (8). The anatomy of the pediatric eye and the clinical
features and etiologic factors of pediatric retinal detachments are different
from adult retinal detachments; thus, the surgical approach to retinal
detachment repair needs to be appropriately tailored to the pediatric patient
(9,10). Scleral buckling is preferred as the primary way to treat pediatric
retinal detachments, because it avoids having to induce a posterior vitreous
detachment in young eyes with a highly adherent vitreoretinal interface and
the potential complications associated with vitrectomy, such as the risk of
creating iatrogenic breaks during intraocular surgery and complications
related to gas or silicone oil. It can also help avoid the need for cataract
surgery in young eyes that have good accommodation. Scleral buckling can
also avoid the need to require young and potentially noncompliant patients
from having to maintain a head position after vitrectomy.
INDICATIONS
In the pediatric age range from birth to 18 years of age, there are vitreoretinal
diagnoses requiring retinal surgery that are unique and more common to the
pediatric patient population, especially during the newborn and infant phase
of life. Newborns and infants may present with retinal detachments due to
retinopathy of prematurity (ROP), persistent fetal vasculature (PFV), familial
exudative vitreoretinopathy (FEVR), and other congenital anomalies. With
increasing age, children can present with retinal detachments often related to
trauma with giant retinal tears and/or dialyses, myopia, uveitis, previous
intraocular surgery, or retinal detachments related to Stickler syndrome
(9,10). The surgical management of these retinal detachments can often
involve the placement of a scleral buckle alone or in conjunction with
vitreoretinal surgery.
BASIC CONSIDERATIONS REGARDING

TECHNIQUE
The placement of a scleral buckle in a pediatric eye can present with issues
that are not present in the adult eye. In young children who present with a
RRD, performing a complete scleral depressed exam is often not well
tolerated in the office. If examining the peripheral retina is difficult to
complete in the office, there should be a low threshold for performing an
examination under anesthesia. Localization of a retinal break is paramount
when placing a primary scleral buckle to ensure that the retinal break is
closed. Access to imaging technology such as color fundus photography,
fluorescein angiography, ultrasonography, and optical coherence tomography
(OCT) can be very useful for diagnosis and documentation when examining
children. In children who have a retinal detachment associated with
peripheral ischemia or abnormal retinal vascular findings such as in ROP,
FEVR, or Coats disease, fluorescein angiography allows ischemic retinal
vascular abnormalities to be better identified and treated with appropriate
laser photocoagulation, as needed.
The small size of a pediatric eye and the unique characteristics of the
pediatric ocular anatomy warrant several particular considerations when
performing retinal surgery. At birth, the axial length of an infant’s eye is
approximately 66% of an adult eye and measures approximately 16.5 mm
(11). At 2 years of age, the axial length is approximately 21 mm and
increases by another 2 mm over the next 11 years of development (11). The
shorter axial length and the larger lens relative to the size of the globe can
affect surgical planning. In the pediatric eye, especially in infants, placement
of a scleral buckle may necessitate a second future surgery to divide the
buckle to permit eye growth and reduce anisometropic amblyopia (12). The
conjunctiva is much thicker, and there is greater redundancy of Tenon
capsule in children than in adults (13). In pediatric eyes, after a 360-degree
conjunctival peritomy is done, the thicker conjunctiva and robust Tenon
capsule can require additional careful dissection to isolate the rectus muscles
compared to an adult eye. Scleral thickness is age-dependent, with infants
having significantly thinner scleras than older children. In the first 6 months
of life, the average scleral thickness is approximately 0.4 mm. In a 2-year-old
child, the sclera doubles in thickness to approximately 0.8 mm. When passing
partial-thickness scleral sutures in infants, thinner suture material can be
considered as well as the use of flat spatula needles (14). The vitreous is
firmly adherent to the retina in pediatric eyes and can make vitreous surgery
more challenging than in adults. If there is PVR, all of the epiretinal
proliferation may not be able to be removed with vitrectomy alone in these
young eyes (15); thus, placement of a scleral buckle in conjunction with
vitrectomy can be useful to help relieve tractional forces secondary to PVR.
In approximately 40% of cases, trauma has been identified as a causative
factor of pediatric retinal detachments (8,9,16). Children can present weeks
or months after the inciting trauma with a chronic macula-off RRD
complicated by PVR. Oftentimes in children, the PVR is exacerbated by the
adherent nature of the posterior hyaloid and the chronicity of the detachment,
which can lead to significant subretinal bands and membranes along with
epiretinal PVR. Evidence of PVR even with subretinal bands does not
automatically warrant a vitrectomy approach in children. Depending on the
degree of PVR and the location of retinal breaks, a primary scleral buckle
alone or with external drainage can be used successfully to seal the retinal
breaks and address the vitreoretinal tractional forces of PVR to reattach the
retina (Figure 60-1). In cases of chronic macula-off RRDs with severe PVR,
children may also have a traumatic cataract, posterior synechiae, and anterior
cyclitic membranes causing hypotony. In such severe cases of PVR,
combining vitrectomy with scleral buckling may be necessary. When
examination of the fundus is limited, either due to a patient’s limited
cooperation or due to opacified media, a B-scan ultrasonography should be
performed. In traumatic pediatric retinal detachments, if a retinal break
cannot be found, a retinal dialysis should be suspected. A retinal dialysis
most commonly occurs after trauma in younger patients (17,18). Retinal
dialyses are most commonly found in the inferotemporal and superonasal
quadrant of the retinal periphery (19). A retinal dialysis with little to no
subretinal fluid can be treated with photocoagulation or cryopexy. A retinal
dialysis causing a retinal detachment with no PVR can be treated successfully
with a scleral buckle. Several studies show that a single scleral buckling
surgery is successful in approximately 87% to 95% of cases of retinal
detachment associated with dialysis (17,19). Successful treatment of retinal
dialyses with segmental scleral buckling has also been reported (14).
FIGURE 60-1 Wide-field fundus photo of a 9-year-old
girl after an airbag injury. She was repaired with a scleral
buckle 220/240 and peripheral cryotherapy for a retinal
dialysis with retinal detachment due to her trauma. We see
pigmentary changes in the posterior pole from old
commotio retinae, a choroidal rupture, and peripheral
cryotherapy scars along with a scleral buckling effect.
Stickler syndrome is the most common cause of inherited and childhood

retinal detachment (20). Stickler syndrome is an autosomal dominant
disorder, caused by mutations in COL2A1, COL11A1, or COL11A2, with
characteristic ophthalmologic and systemic manifestations, primarily
orofacial, auditory, and articular involvement (21,22). Ocular features of
Stickler syndrome are characterized by abnormalities of the vitreous, high
myopia, paravascular pigmented lattice degeneration, cataracts, and retinal
detachment (Figure 60-2).
FIGURE 60-2 Top Row Left:Wide-field fundus photo of
macula-off rhegmatogenous retinal detachment in a 14-
year-old boy with Stickler syndrome, monocular due to
total retinal detachment in the other eye. Top Row
Right:OCT macula demonstrating subretinal fluid.
Bottom Row Left:Postoperative day 1 wide-field fundus
photo after repair with 220/240 scleral buckle. Bottom
Row Right:Postoperative day 1 OCT macula showing
resolution of subretinal fluid.
Patients with Stickler syndrome are at a high risk of vision loss from RRD
caused by a giant retinal tear (see also Chapter 37). Due to the inherited
nature of Stickler syndrome and high rate of bilateral spontaneous retinal
detachments, even “unaffected” family members without a history of a retinal
detachment should be examined. There is some controversy regarding the
best way to prophylactically manage these patients. Some experts advocate
for prophylactic circumferential cryotherapy or laser at the ora serrata, laser
therapy around areas of paravascular pigmented lattice degeneration (23,24),
or the placement of a prophylactic scleral buckle. Retinal detachments caused
by giant retinal tears can be managed with primary vitrectomy with gas or
silicone oil tamponade or combined scleral buckle-vitrectomy. The use of
perfluorocarbon liquid optimizes the management of giant retinal tears by
helping to avoid issues with retinal slippage. In cases where there is a giant
retinal tear with PVR, often a scleral buckle combined with pars plana
vitrectomy using direct perfluorocarbon–silicone oil exchange may be
helpful. Placement of a scleral buckle can be useful to support the vitreous
base (Figure 60-3) (25).
FIGURE 60-3 Top:Wide-field fundus photo of retinal
detachment due to giant retinal tear in a 12-year-old boy.
Bottom:Wide-field fundus photo after repair with pars
plana vitrectomy, scleral buckle (42/72) with silicone oil.
Retinal detachments due to ROP, FEVR, and PFV tend to be more tractional
and often require surgical management with vitrectomy either with or without
a scleral buckle (see also Chapter 66). In the treatment of retinal detachments
due to ROP, the goals of scleral buckling are to reattach the retina and to
reduce the progression from stage 4A or 4B to stage 5 ROP (12,26). A
circumferential scleral buckling procedure is a reasonable approach if the
detachment already has the fibrovascular ridge dragged to the pars plicata
with significant retinal dragging (14). After retinal reattachment, the
encircling scleral buckle should be divided at about 3 months after the scleral
buckling surgery to prevent anisometropia, which can range from 5 to 9 or
more diopters. Division of the scleral buckle reduces the risk of
anisometropia and allows for eye growth (14). If the tractional retinal
detachment involves the macula (stage 4B), a lens-sparing vitrectomy (LSV)
is a more favored approach (27–29). The surgical approach for stage 5 ROP
typically involves lensectomy, pars plicata vitrectomy, and rarely scleral
buckling (14,30).
FEVR is a hereditary disorder characterized by abnormal retinal
angiogenesis that leads to abnormal or incomplete vascularization of the
peripheral retina (see also Chapters 61 and 66). The fundus changes are
similar to those seen in ROP but affects children who were born full-term
with normal birth weight. In severe manifestations of FEVR, the areas of
peripheral avascularized retina can lead to retinal detachments caused by
vitreoretinal traction. Similar to retinal detachments caused by ROP,
depending on the configuration and macular involvement of the retinal
detachment, vitrectomy with or without scleral buckling can be considered. If
the detachment is primarily exudative, scleral buckle alone or with external
drainage can be used to successfully reattach the retina. If the detachment is
primarily tractional, a vitrectomy is often necessary to relieve the
vitreoretinal traction. If there is a rhegmatogenous component to the
tractional detachment, the placement of a scleral buckle can help support the
vitreous base and peripheral ischemic retina and breaks (31).
PFV is a congenital disorder in which the fetal hyaloid vasculature fails
to regress as it should (see also Chapter 65). It usually presents unilaterally
without associated systemic findings in a normal full-term infant. PFV has a
wide spectrum of associated clinical manifestations including cataract,
microphthalmia, optic nerve hypoplasia, glaucoma, dysplastic retina, retinal
fold, and tractional retinal detachment. The severity of the microphthalmia
and the degree of dysplastic retina largely determines the visual prognosis. In
cases of posterior PFV with a vitreous stalk, a vitrectomy with transection of
the stalk can resolve the retinal traction. In such cases, the role of a scleral
buckle is minimal and is not recommended. In cases of tractional retinal folds
or tractional retinal detachment, membrane peeling is often necessary, and a
vitrectomy approach is preferred (32).
EQUIPMENT
In children, over the age of 10 years, the eye is nearly fully grown and is
essentially the same size as an adult eye (11). Commonly used buckle
elements are the 3.5-mm solid silicone band (Type 41), 4-mm band (Type
42), or a 6-mm band (Type 220) with a 2.5-mm encircling band (Type 240).
These different silicone band elements can be used alone or in combination
with pars plana vitrectomy. For younger patients with a smaller eye, smaller
elements such as the 2.5-mm encircling band (Type 240) or 3.5-mm
encircling band (Type 41) may be preferred. The choice of band is often
determined by the size of the eye and the localization of the retinal pathology
on the sclera. There may be a preference for a larger buckle in children
especially if the pathology is inferior, because the development of PVR can
be more aggressive following rhegmatogenous detachment in children (15).
Regarding additional instrumentation besides the buckle elements, varying
sutures and needles can be used depending on surgeon preference. For infants
and young children with thin sclera, spatulated needles are often preferred to
reduce the risk of entering the globe. The authors include a list of preferred
instrumentation for scleral buckling (Table 60-1).
TABLE 60-1 Pediatric scleral buckle instrument

list
The same general principles of scleral buckle placement for retinal
detachment repair in adults apply: localizing break(s) on the sclera, choosing
and placing an appropriate-sized buckling element to support the breaks,
treatment with cryotherapy or laser to create a permanent chorioretinal
adhesion, and release of subretinal fluid for a good contour of the buckle and
the break, if needed. However, in pediatric patients, there are some special
considerations.
For an encircling element, a 360-degree conjunctival peritomy is
performed, and the muscles are isolated. For segmental elements, the
quadrant localized to the retinal defect can just be isolated. The muscles are
isolated, and traction sutures are placed beneath the insertion of each rectus
muscle. Tenon capsule is cleaned from the scleral surface by lysing its
delicate adhesions with a cotton-tipped applicator. In young patients with
robust Tenon capsule, it may be necessary to use forceps to grasp the
intermuscular septum adjacent to each muscle to strip it posteriorly. The
sclera is cleaned in all four quadrants and examined to check for any areas of
marked scleral thinning. Extreme care is used when manipulating areas of
marked scleral thinning to avoid scleral perforation (33). Correct localization
of the retinal break on the sclera is important to enable the scleral buckle to
support the break adequately. Treatment around the retinal break can be
performed either with cryotherapy or with indirect laser photocoagulation if
the retina is apposed to the retinal pigment epithelium. The appropriate-sized
buckling element should be chosen and placed so that it is approximately 2
mm beyond the anterior and posterior extent of the break localized on the
sclera (15). To allow for proper imbrication of the buckle, intrascleral
mattress nylon sutures are placed 1- to 2-mm farther apart than the width of
the buckling element (15). Tightening the band circumferentially usually
induces more axial elongation of the globe rather than imbrication and is
minimized. During the process of manipulating the encircling band, the
intraocular pressure of the eye should be frequently checked with palpation.
If the pressure is too high after appropriate imbrication of the buckle, the
height of the subretinal fluid can be checked to see if external drainage
should be performed. If the buckle appears to be in good position and there is
a minimal amount of subretinal fluid and the intraocular pressure is high, the
pressure can be lowered with an anterior chamber paracentesis. One method
to perform a paracentesis is with careful removal of aqueous fluid using a 30
gauge needle on a TB syringe that is inserted into the anterior chamber
avoiding damage to the lens.
Depending on the amount of subretinal fluid, external drainage can be
considered if all of the vitreoretinal traction has already been relieved. In
cases of RRDs with persistent bullous retina with an extensive amount of
subretinal fluid, partial, slow external drainage can be done. In cases of ROP
and FEVR, external drainage is sometimes performed to allow the retina to
settle back from the lens unless there are areas of residual traction and
redundant retina that can lead to retinal breaks or retinal incarceration during
the external drainage process (15). There are various techniques to externally
drain the subretinal fluid. Classically, a 2- to 3-mm radial scleral cut down is
performed with a 64 blade, and diathermy is applied to the sclera and
extruding choroid. Subretinal fluid can be released with electrodes from
TR4000 (MIRA), if possible, or with a 30 g needle. In cases with some
residual subretinal fluid remaining that is neither bullous nor high enough to
allow for safe external drainage, an intravitreal injection of air or short-acting
gas like SF6 (100% concentration, 0.4 mL) can be considered. If the buckle
needs to be additionally tightened, a paracentesis can be done to normalize
the intraocular pressure.
In children younger than 3 years of age, the encircling band is cut
approximately 3 months after the scleral buckling operation to allow for eye
growth once stable retinal reattachment has been achieved (14). The buckle
element is usually not removed entirely; rather, it is severed so that the
encircling element that has become encapsulated can continue to support the
retina. The encircling band is typically divided in the area where the retina is
flat without any evidence of traction. Westcott scissors or a 57-degree blade
can be used to make an incision through the conjunctiva and then the
pseudocapsule, in the direction along the length of the band. If the
pseudocapsule overlying the band can be identified, a 64 blade can be used to
carefully create an incision along the length of the band and then cut. The
band can then be lifted from the sclera and cut with Stevens scissors (15).
Chandelier-assisted scleral buckling has emerged as a new technique that
may potentially offer some benefits over traditional scleral buckling
techniques. Although scleral buckling remains the preferred method to treat
pediatric retinal detachments due to its declining popularity, many
vitreoretinal surgery trainees do not feel as comfortable with scleral buckling
compared to prior generations. Chandelier-assisted scleral buckling takes
advantage of the modern-day wide-angle viewing systems and uses oblique
endoillumination to help identify peripheral breaks.
There are different types of chandelier systems including the single
valveless pars plana 25-gauge cannula/chandelier system (Alcon, Fort Worth,
TX), the dual 29-gauge chandelier/cannula system (Synergetics, O’Fallon,
MO), and the twin uncannulated 27-gauge chandeliers (Eckard TwinLight
Chandelier; DORC International, Zuidland, The Netherlands). These
chandelier systems can be combined with any type of different wide-field
viewing systems such as the noncontact BIOM (OCULUS Surgical Inc., Port
St. Lucie, FL), noncontact Resight (Carl Zeiss Meditec AG, Jena, Germany),
or contact wide-field lens (Volk, Mentor, OH). The chandeliers can be
positioned 180 degrees away from the primary retinal break, 90 degrees away
from it or adjacent to the break to optimally visualize the break. While the
chandelier is in the eye, the retinal break can be marked and treated with laser
or cryotherapy under direct visualization. Some potential disadvantages of
using chandelier-assisted scleral buckling include the risks of lens damage,
light toxicity, endophthalmitis, and PVR redetachment. Upon removal of the
chandelier cannula, it is important to check if there is a vitreous wick and to
trim it with scissors to minimize any vitreous traction (34).
POSTOPERATIVE CARE
At the end of the surgery, a peribulbar block (0.75% Marcaine + 2%
lidocaine; 1:1) is given to reduce postoperative pain after the patient awakens
from general anesthesia. In the immediate postoperative period after scleral
buckling, children can present with more periorbital edema and conjunctival
edema than adult patients. If there are no medical contraindications, in older
children, a Medrol dose pack can be given along with the standard
postoperative steroid eye drops, which can help minimize postoperative
swelling. The advantage of scleral buckling is that it can avoid the need to
require strict head positioning in young patients. However, if head
positioning is required, parents can be asked to encourage children to
participate in activities that require a head down position such as coloring,
working on puzzles, or using their tablet.
It is imperative that pediatric patients are closely comanaged with
pediatric ophthalmologists for amblyopia prevention and follow-up of any
refractive concerns. With the thinner profile scleral buckles that are typically
used today, the myopic shift is usually only a few diopters after scleral
buckling. However, in children that require placement of a larger element or
sponge, large refractive shifts can occur and should be managed closely by
the pediatric ophthalmologist. Infants may also require close follow-up with
pediatric ophthalmologists or optometrists for contact lens fitting if they are
aphakic.
In the long term, children should be examined at least annually if not
more frequently, depending on their ocular status and risk of retinal
detachment. If a child is monocular, follow-up every 6 months may be more
appropriate and monocular precautions need to be emphasized with
protective eyewear recommended. Retinal detachment precautions should be
reviewed with both patient and parents.
COMPLICATIONS
Complications following scleral buckles can include changes in refractive
error, diplopia, strabismus, infection and extrusion of the scleral buckle,
anterior segment ischemia, and choroidal detachments (14). Scleral buckling
can change the shape of the globe and lead to refractive changes with or
without astigmatic changes. In most studies, astigmatic changes are
infrequent but may often be more associated with radial buckles, anteriorly
located buckles, and high buckles (35,36). The most common refractive
change with scleral buckling is increased myopia due to increase in the axial
length of the eye. In the adult population, Smiddy et al. reported on the
refractive outcome of 75 eyes of 69 patients and showed that encircling
elements induced an average of 2.75 diopters of myopia (37). Sato et al.
looked at the refractive change after scleral buckling in 35 eyes of 35
pediatric patients and found that the treated eyes became less myopic than the
fellow eyes and concluded that scleral buckling may be impeding ocular
growth (38). The mean refractive change 1 to 4 years after the scleral
buckling was 0.6 diopters of myopia in the eye with the scleral buckle
compared to the mean change of 1.3 diopters of myopia in the fellow eye.
Diplopia after scleral buckling is usually transient and resolves
completely in about a week; however, rarely it can be permanent and has
been found to occur about 4% of the time postoperatively (39). Long-term
diplopia after retinal detachment surgery has been attributed to the scleral
buckle itself, scarring in the orbital space, or myoscleral adhesions (40,41).
To minimize scarring around the muscles, the rectus muscle sheaths should
be preserved as much as possible. Smaller and thinner silicone elements have
less tendency to cause heterotropia compared to larger sponge elements (39).
Infections after scleral buckling are rare, and the reported incidence
varies from 0.2% to 5.6% (42–44). Many of the previous studies looked at
silicone sponge exoplants. In a 2013 study by Chhablani et al. (44), the
infection rate of solid silicone exoplants was 0.2% compared to 0.5% to 5.6%
in silicone sponges (42,43). The most common causative organism was
coagulase negative staphylococci (44).
Extrusion of the silicone band is also another rare complication. Silicone
band extrusion through the conjunctiva may be seen incidentally on follow-
up examination or may present with eye redness, pain, and irritation.
Depending on the degree of extrusion, the buckle may need to be removed or
resutured with proper closure of Tenon fascia and conjunctiva over the
buckle.
Anterior segment ischemia has been described as a rare complication,
especially in patients with sickle cell hemoglobinopathy. Placement of a high
scleral buckle possibly with the use of excessive cryotherapy may increase
the risk of anterior segment ischemia by causing limited perfusion due to
compromise of the anterior and posterior ciliary arteries (36,45). Anterior
segment ischemia from an encircling band may be prevented by ensuring that
the band is not overly tight around the eye or excessively high.
Serous choroidal detachments can develop shortly after scleral buckling
and usually spontaneously resolve within a few weeks after surgery. They
occur at a rate of 23% to 44% after scleral buckling (36,46,47). Risk factors
that can lead to serious choroidal detachments are excessive photocoagulation
or cryotherapy along with postoperative hypotony and high myopia (48).
Prior to surgery, it is crucial to have a discussion with the patient’s parents so
that they have a good understanding of their child’s ocular condition, need for
surgery, and have realistic expectations regarding their child’s postoperative
visual potential. In children with ROP, FEVR, PFV, Stickler syndrome, or
other congenital anomalies, parents will need to have a good understanding
of the extent of their child’s ocular disease and the limited visual potential
that their child may have. Wide-field fundus photos, B-scans, OCT, and any
other imaging modalities that can be shown to the parents can be helpful to
explain the complex ocular condition their child may have. If the ocular
condition only affects one eye, comparing the photos of the normal eye to the
affected eye can help parents appreciate the extent of ocular disease their
child may have and the need for surgical repair.
There is a high likelihood that multiple surgeries may be needed for
successful retinal detachment repair in the pediatric population. Children
have a higher risk of PVR, which can require additional retinal repair surgery.
The single-procedure anatomical success rate for pediatric retinal detachment
is reported to range from approximately 50% to 90% (15,49,50). Even if the
retinal detachment is repaired with a single surgery, the child may have
concomitant ocular conditions that need to be addressed surgically. In
traumatic retinal detachments, if the patient sustained a central corneal
laceration, a traumatic cataract, or developed glaucoma in addition to having
a retinal detachment, the patient will require surgeries from other
subspecialties. Combined surgeries with different subspecialties can help ease
the burden of having to undergo general anesthesia multiple times for the
patient. Most importantly, it should be emphasized to the parents that the
recovery process will be a long journey that requires close follow-up and
possibly multiple surgeries. The need for follow-up, patching, contact lenses,
and/or glasses should be continually emphasized. Parents will need to be
reminded to follow-up with the pediatric ophthalmologists or optometrists to
ensure any refractive issues or amblyopia concerns are being properly
addressed. If the visual potential is extremely poor, discussing a low vision
consultation and advising of social work assistance to help navigate different
social and school aid networks to help with vision and activities may be
helpful.
WHEN TO CONSIDER ANOTHER

PROCEDURE
Sometimes scleral buckling alone is not sufficient, and it either needs to be
combined with pars plana vitrectomy, or a primary vitrectomy needs to be
performed. Indications for primary vitrectomy in pediatric eyes include dense
vitreous hemorrhage, severe PVR, RRDs with posterior retinal breaks, and
primarily tractional retinal detachments such as in ROP, FEVR, and PFV
(3,29). Scleral buckling remains the preferred method for repairing most
pediatric retinal detachments, because it avoids intraocular surgery and the
need to manipulate a highly adherent vitreous. However, there are times
when a primary scleral buckle is not the ideal treatment regimen. In tractional
retinal detachments, a vitrectomy is necessary to release the tractional forces
to treat the retinal detachment. Thus, severe PVR cases and retinal
detachments associated with ROP, FEVR, and PFV are usually primarily
tractional and require a vitrectomy to successfully reattach the retina.
Depending on the severity of the tractional detachment, if there are any
associated rhegmatogenous breaks, a supporting scleral buckle combined
with a vitrectomy may be necessary to reattach the retina.
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pediatric rhegmatogenous retinal detachment in Taiwan. Am J Ophthalmol
2005;139(6):1067–1072.
50. Wang N-K, Tsai C-H, Chen Y-P, et al. Pediatric rhegmatogenous retinal detachment in East
Asians. Ophthalmology 2005;112(11):1890–1895.
61
Surgical Approaches to Infant and
Childhood Retinal Diseases:
Intravitreal Surgery
Jon athan E. Sears
INTRODUCTION
Surgical management of pediatric retinal disorders using vitrectomy can be
among the most challenging interventions in patients with vitreoretinal
disease for many reasons. First, diagnosis may require an examination under
anesthesia (EUA). Second, pediatric vitreoretinal disease often accompanies
genetic disorders, and, therefore, surgical intervention is designed to repair or
stabilize the anatomy of tissues that have a molecular basis to their
dysfunction that is not addressed surgically. Finally, the ocular anatomy of
children is different from adults. The vitreous is more adherent to the retina,
and the vitreoretinal interface is different in children than adults. Children
often develop retinal detachment from not only rhegmatogenous origins but
also simultaneous tractional and exudative causes. The following chapter will
outline systematic techniques in the practice of vitrectomy in children.
PREOPERATIVE ASSESSMENT
One of the most critical components of successful surgical outcomes is a
clear surgical plan. Patients that are too young to comply with adequate
preoperative evaluation require an EUA. Although the safety of anesthetic
gases in infants and toddlers has not been established, at no point should an
EUA be avoided if an office visit yields inconclusive data (1). EUAs can be
extended to surgical intervention provided detailed informed consent and
open dialogue with parents and legal guardians is obtained. All EUAs should
involve careful anterior and posterior ophthalmic examination including
cycloplegic retinoscopy, intraocular pressure measurement, measurement of
corneal diameter, and imaging studies as necessary including widefield
photographs, fluorescein angiography, ocular coherence tomography, and
ultrasound as necessary. Perioperative assessment with fluorescein
angiography yields informative data especially relevant to the multiple causes
of peripheral avascularity common to shaken baby syndrome, Coats disease,
familial exudative vitreoretinopathy (FEVR), combined retinal pigment
epithelium (RPE) hamartoma, and retinopathy of prematurity (ROP) (2).
In order to be certain that the safety and efficacy of surgical intervention
is maximized, all children require a detailed preoperative physical
examination from a pediatrician and the knowledge that 23-hour observation
is necessary for any child born prematurely and <1 year old.
WIDE-ANGLE VIEWING SYSTEMS

Although the Avi wide-angle viewing system includes a smaller pediatric
version of a handheld lens, most pediatric vitrectomy cases benefit from
noncontact lens systems, because the absence of a contact lens enables the
use of cannulas in a small eye, which in company with handheld systems
restricts movement of instruments within the eye (Figure 61-1A and B) (3).
Figure 61-1 Contact versus noncontact wide-angle
viewing systems. A:Wide-angle handheld contact lens is
small enough to use for even ROP surgery but is limited
when cannulas are used simultaneously. B:Noncontact
lens can be used with cannulas without limitation but has
more edge distortion than wide-angle contact lenses.
PORT PLACEMENT
Although transconjunctival surgery is safe and effective, all beveled pediatric
self-sealing incisions should be closed with a 7-0 Vicryl (4). Therefore, at the
outset, one should consider conjunctival peritomy to facilitate closure of the
sclerotomies but transconjunctival sutures if placed correctly are believed to
be safe. Any delay in sclerotomy closure once the cannula is removed can
cause hypotony and devastating intraocular hemorrhage; the surgeon should
decide on the quickest technique of port closure. Often a preplaced suture at
the infusion site is advantageous.
The first decision in considering vitrectomy is whether entry sites should
be placed at the limbus or pars plana, and whether cannulas for
microincisional vitrectomy should be used at all. There is little doubt that
current 25G instrumentation is superior to larger gauge instrumentation at the
time of this writing. However, the location of port placement depends on the
age and diagnosis of the child. For example, over half of cases involving
persistent fetal vasculature (PFV) involve media opacity that precludes a
view of pars plana. In patients with this disorder, pars plana might be absent,
that is, the retina inserts directly into pars plicata, therefore, risking a
devastating retinal break from an entry site tear (5). This scenario is also true
for stage 5 ROP in which a lensectomy is necessary. In these cases, limbal
ports with iris hooks facilitate lens extraction and necessary posterior
vitrectomy that is safe from creating retinal tears. Limbal incisions are
difficult to make with the standard trochar/cannula set, especially if a contact
lens is required because it will ride on the cannulas. Therefore, one could
consider not using cannulas, such as in the 25G short pack designed to rely
on a shorter, stiffer vitrectomy probe and a sutured infusion cannula with
naked incisions through pars plana or the limbus (Figure 61-2A). On the
other hand, if there is a view of pars plana, placing ports through pars plana
naturally makes posterior manipulation easier. Valved cannulas are a
significant advantage to surgery because the eye remains pressurized as
instruments are exchanged, and in cases that are vascularly active, delay in
closure and hypotony can be disastrous.
Figure 61-2 A:25G short pack relies on a shorter
vitrectomy probe and sutured infusion cannula in the
absence of superior cannulas. This is helpful is smaller
eyes. B:45-degree light pick is helpful for a 3-port
vitrectomy to assist in elevating the posterior hyaloid.
C:Chandelier light source facilitates bimanual surgery.
In all cases other than PFV and stage 5 ROP, sclerotomies can be placed
through pars plicata or pars plana. In all cases, scleral depressed examination
is necessary to ensure that these areas are free from retina. For stage 4A ROP
surgeries, ports can be placed in a phakic eye 1.5 mm from the surgical
limbus (6–8). In most cases and cases that require contact lens viewing, the
superior sclerotomies are recommended without cannulas. In some cases,
end-irrigating light sources can reduce incisions from three sites to two sites.
With the advent of 25G instrumentation, a challenge to these infusion sources
is whether or not adequate volume/eye pressure can be maintained during
vitrectomy. In older children (>2 years), without anomalous anatomy such as
microphthalmia, ports can be placed at 3 mm from the limbus.
PHAKIA VERSUS PSEUDOPHAKIA

VERSUS APHAKIA
Often the first decision in treating vitreoretinal disease is whether to leave a
child phakic or pseudophakic or aphakic. There are three considerations
important to this decision. First, what is the status of the companion eye, that
is, which refractive challenge minimizes amblyopia? For example, bilateral
pathology that requires bilateral lensectomy might be safer with aphakia.
Second, what is the choice of tamponade? If silicone oil is used, the child’s
lens can stay clear for at least 3 months, and if there is a posterior subcapsular
cataract that develops, cataract surgery can be timed with silicone oil
removal. Often children with silicone oil tamponade and pseudophakia
develop posterior synechiae that can cause pupillary block even in the
company of a PI. This rarely occurs in the phakic child. Finally, in cases of
self-abuse/trauma, aphakia is preferred because the risk of lens
dislocation/anterior segment disorganization is minimized by aphakia. A
proper basal PI inferiorly prevents silicone oil migration into the anterior
segment.
INTRAOPERATIVE CONSIDERATIONS
Posterior Hyaloid Separation and Vitrectomy
Elevation of the posterior hyaloid is the first step in all vitrectomy cases.
Although separation of the cortical vitreous is essential to complete
vitrectomy, it is often very challenging to accomplish because of the firm
attachment of the posterior hyaloid. In order to facilitate induction of a
posterior vitreous detachment (PVD), a 45-degree light with active suction
from the vitrectomy probe under high magnification can be useful (Figure
61-2B). Staining of the vitreous with intraoperative triamcinolone acetonide
can be valuable, as can the use of plasmin, cautery, or a flex loop in select
cases (9, 10). There are cases where complete hyaloid removal is difficult,
such as in stage 4A ROP or Stickler’s associated detachment and yet
successful outcomes can still be achieved. Vitreous removal is better safe
than complete. In general, separation of the hyaloid to ante-equatorial
position is sufficient, but naturally elevation of the posterior hyaloid to the
vitreous base is the gold standard and easiest opportunity to provide a
complete vitrectomy.
Membrane Stripping
Membrane stripping often leads to retinal breaks, which are disastrous to the
pediatric patient due to risk of proliferative vitreoretinopathy (PVR).
Therefore, it is favorable to approach membrane stripping with a bimanual
approach that uses a chandelier light source to facilitate the simultaneous use
of forceps and horizontal scissors (Figure 61-2C). Alternatively, a 45-degree
light pick can be useful but does pose a risk for retinal penetration when
applying counter traction.
Drainage of Subretinal Fluid

Perhaps an undisputed statement in adult and pediatric vitrectomy is that
retinal breaks should be avoided. Although this sounds very obvious, the
particular relevance to pediatric vitrectomy lies in the nature of tractional and
exudative causes of pediatric detachment. Adherent hyaloid and stretched and
displaced retina often forces the surgeon to finish the repair with the retina
still detached but without breaks. Recently, perfluoro-n-octane (Perfluoron,
PFO, Alcon)–assisted external drainage has become essential to reattaching
retina to facilitate placement of thermal treatment necessary to prevent
redetachment. For example, exudative processes such as Coats disease with
retina to the lens (Figure 61-3A) is best managed by external drainage while
infusing BSS into the anterior chamber first (Figure 61-3B), placement of
pars plana ports when the retina relaxes to provide sufficient room second
(Figure 61-3C), separation of the posterior hyaloid and complete vitrectomy
(Figure 61-3D), and then complete removal of subretinal fluid (SRF) using
PFO to cause the SRF to migrate anteriorly toward an external sclerotomy
created posterior to the muscle insertions (Figure 61-3E and F). Once the
retina is attached and under PFO, thermal treatment can be used to
thoroughly ablate the origin of exudation (Figure 61-3G). This technique has
worked well in other disorders such as stage 5 ROP in which a complete
vitrectomy was possible, combined retinoschisis/rhegmatogenous retinal
detachment (RRD) in X-linked retinoschisis (XLRS), and especially FEVR.
Therefore, in contrast to adult RD repair, internal drainage through a
retinotomy should be avoided.
Figure 61-3 Internal PFO-assisted external drainage of
subretinal fluid prevents making retinal breaks. A:The
retina is against the lens in a stage 5 Coats patient;
therefore, an (B) external drainage while infusing anterior
chamber fluid allows retina to relax enough to (C) place
ports through pars plana. D:After complete vitrectomy,
(E) residual fluid can be driven anteriorly by PFO (F) and
subretinal fluid further evacuated as before externally.
This allows placement of (G) thermal treatment to treat the
origin of subretinal fluid in a completely attached retina.
This procedure is effective in Coats, ROP, FEVR, and X-
linked schisis with retinal detachment.
Choice of Tamponade
There is no “set rule” for the choice of tamponade. Certainly, the silicone oil
study has instructed us that in terms of long-term tamponade (C3F8 vs.
silicone oil), both gas and oil are equally effective (11). But in children,
silicone oil creates less acute cataracts than gas and naturally bears less
attention to meticulous face down positioning. Silicone oil also enables vision
through the tamponade and hence is less amblyogenic and easier to examine
in the acute postoperative period. On the other hand, for superior breaks in
rhegmatogenous detachments or pediatric macular holes, SF6 is a good
choice. Therefore, oil is favorable for long-term tamponade and SF6 or gas
for short-term tamponades. There is no evidence of advantages of heavier
centistoke silicone oil versus lighter oil (5,000 vs. 1,000).
Adjuvant Scleral Buckle

In general, if one considers using a scleral buckle (SB), it is a good idea to
use one. For certain cases, combined SB and pars plana vitrectomy (PPV) is
essential such as Stickler, trauma, and FEVR (12, 13). But in children with
ROP, an SB does not increase successful outcomes (14). Additionally, an SB
can be growth restrictive and needs to be severed or removed if placed
around an eye less than 2 years old after an appropriate duration (15).
POSTOPERATIVE EVALUATION
Attention to the emotional needs of a child and their family is essential to a
successful outcome. This includes thorough explanation to family members
and encouragement to patients that make them feel proud, hopeful, and
trusting. Therefore, all controllable postoperative complications must be
avoided. Ports should be closed with sutures. Peripheral iridectomies should
be made routinely in phakic and pseudophakic eyes, generally inferiorly, to
reduce the risk of iris bombe and angle closure. Intraocular pressure should
be adjusted carefully in the operating room and measured to prevent
tamponade overfill before leaving the OR. Either retrobulbar or peribulbar
anesthesia is essential, and especially if marcaine is used, appropriate dosing
to prevent cardiac arrhythmia should be discussed with pediatric anesthesia
personnel. Finally, the trust that a child has in their retinal specialist depends
upon the avoidance of inflicting pain; alienating a child for the benefit of a
better office examination is not a victory. Small children that can be held
safely can be readily examined with a sterile disposable speculum and scleral
depressor. Postoperative day 1 generally involves making sure there is no
infection, normal intraocular pressure, and a good red reflex. A more detailed
examination could be obtained in the ensuing visit at 1 week. If a child is
adamant about refusing dilated fundus examination, usually an office B-scan
can suffice for the short term until a repeat EUA is considered the only
option.
CONCLUSION
Vitrectomy for children can restore both the normal anatomy and function of
the retina in cases where blindness is a certain outcome. Specific surgical
pearls associated with particular diagnoses are listed in Table 61-1 as a help
to the reader about to intervene in behalf of a child with vitreoretinal disease.
Careful preoperative evaluation by an examination under anesthesia can
direct a successful surgical plan. Port placement is essential to effective
intervention and is dependent on the preoperative diagnosis. Separation of the
posterior hyaloid is necessary but not at the expense of creating devastating
posterior breaks. Internal PFO-assisted external drainage is useful in
completely reattaching retinas intraoperatively without risking conversion of
a tractional/exudative detachment into a combined rhegmatogenous one and
facilitates thermal treatment. Choice of tamponade must consider amblyopia,
visual function during rehabilitation, and particular challenges of the patient.
A kind and caring approach for these vulnerable patients and their families
enriches the lives of caregivers, families, and children.
TABLE 61-1 Surgical tips for vitrectomy

REFERENCES
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markers of brain development during sevoflurane anaesthesia in children up to 3 years old. Br J
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coherence tomography with fundus photographs, fluorescein angiography, and histopathologic
analysis in assessing Coats disease. JAMA Ophthalmol 2019;137(2):176–183.
3. Inoue M. Wide-angle viewing system. Dev Ophthalmol 2014; 54: 87–91.
4. Gonzales CR, Singh S, Schwartz SD. 25-Gauge vitrectomy for paediatric vitreoretinal
conditions. Br J Ophthalmol 2009;93: 787–790.
5. Stark WJ, Fagadau W, Lindsey PS, Taylor HR, Michels RG. Management of persistent
hyperplastic primary vitreous. Aust J Ophthalmol 1983;11:195–200.
6. Machemer R, deJuan E. Retinopathy of prematurity: approaches to surgical therapy. Aust N Z J
Ophthalmol 1990;18:47–56.
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8. Prenner JL, Capone A Jr, Trese MT. Visual outcomes after lens-sparing vitrectomy for stage 4A
9. Cernichiaro-Espinosa LA, Berrocal AM. Novel surgical technique for inducing posterior
vitreous detachment during pars plana vitrectomy for pediatric patients using a flexible loop.
10. Wu WC, Drenser KA, Lai M, Capone A, Trese MT. Plasmin enzyme-assisted vitrectomy for
primary and reoperated eyes with stage 5 retinopathy of prematurity. Retina 2008; 28:S75–80.
11. The Silicone Study Group. Proliferative vitreoretinopathy. Am J Ophthalmol 1985;99:593–595.
12. Fivgas GD, Capone A Jr. Pediatric rhegmatogenous retinal detachment. Retina
2001;21:101–106.
13. Hocaoglu M, Karacorlu M, Sayman Muslubas I, Ersoz MG, Arf S. Anatomical and functional
outcomes following vitrectomy for advanced familial exudative vitreoretinopathy: a single
surgeon's experience. Br J Ophthalmol 2017;101: 946–950.
14. Sears JE, Sonnie C. Anatomic success of lens-sparing vitrectomy with and without scleral
buckle for stage 4 retinopathy of prematurity. Am J Ophthalmol 2007;143:810–813.
15. Hinz BJ, de Juan E Jr, Repka MX. Scleral buckling surgery for active stage 4A retinopathy of
prematurity. Ophthalmology 1998;105:1827–1830.
62
Endoscopic Vitreous Surgery in
Sui Chien Wong, and Thomas C. Lee
INTRODUCTION
Achieving consistent surgical outcomes in complex pediatric
vitreoretinopathies such as familial exudative vitreoretinopathy (FEVR) and
retinopathy of prematurity (ROP) remains one of the most difficult
conundrums for experienced pediatric retinal surgeons. In acute ROP with
stage 5 disease or total tractional retinal detachment (TRD), one of the most
surgically challenging vitreoretinal conditions, success following vitrectomy
as defined by partial or complete anatomical reattachment varies widely from
28% (1) to 69% (2) in large retrospective studies. In children who develop
traumatic retinal detachment (RD) following open globe injury, retinal
reattachment rates following vitrectomy similarly vary from 36% (3) to 53%
(4,5), albeit in a more heterogenous disease. In persistent fetal vasculature
(PFV), previously termed persistent hyperplastic primary vitreous, the risk of
iatrogenic rhegmatogenous RD following vitrectomy varies from 9% (6) to
21% (7) in recent studies.
While part of the variability in surgical success rates may be explained by
case selection bias and the very nature of the pathology in these
vitreoretinopathies, in the authors’ experience, the difference between
surgical success and failure is often determined by how well surgical planes
are delineated and appropriately manipulated and dissected. The challenge of
optimal visualization of surgical planes stems from the anteroposterior
orientation of TRD in many of these cases coupled with the suboptimal top-
down perspective through the cornea of microscope-based (conventional)
contact or noncontact viewing systems. Endoscopy offers the surgeon a
unique intraocular perspective, conferring several advantages over current
contact and noncontact viewing systems in certain scenarios, specifically in
complex pediatric vitreoretinopathies. This chapter explores the role and
relevance of endoscopic vitrectomy in children.
Endoscopic Vitreoretinal Surgery
Instrument Development
The term endoscope is derived from two Greek words—endon, meaning
within or inside, and skopin, meaning to examine or view. The first
ophthalmic endoscope was developed by Thorpe (8) in 1934. This had an
integrated forceps for removing nonferromagnetic foreign bodies. A separate
illumination was required. Later that same year, Thorpe (9) developed an
endoscope incorporating illumination measuring 6.5 mm in width. Monocular
visualization was achieved through an eyepiece attached to the proximal end
of the endoscope. It was over 40 years later before Norris and Cleasby (10)
developed a markedly smaller 1.7-mm-diameter endoscope for intraocular
and orbital surgery. In 1990, Volkov et al. (11) described the use of a 20-
gauge endoscope through the pars plana with image projection on an
electronic monitor in 23 patients with anterior segment opacities.
Present-day endoscopes are available in 19, 20, or 23 gauge and
incorporate up to three functions in a single instrument, namely, a fiberoptic
video camera, a xenon light source, and an endolaser (12,13). A 23-gauge
endoscope has the advantage of being able to fit through standard 23-gauge
microcannula systems. Although there is a trade-off with a slightly narrower
field of view (FOV) of 90 degrees and the number of fibers that can be
incorporated into a smaller diameter instrument, the image quality with the
10,000 pixel resolution is certainly adequate for surgery for rhegmatogenous
RD, proliferative vitreoretinopathy (PVR), endophthalmitis, and nonclearing
vitreous hemorrhage. With 20-gauge endoscopes, resolution is at 10,000
pixels but with a wider FOV of approximately 110 degrees. With a 19-gauge
endoscope, a 160-degree FOV is attainable at a resolution of 17,000 pixels. In
practical terms, this translates to being able to resolve detail down to 20 μm
or less at an optimum working distance of 3 to 4 mm. Taking into account
that the diameter of a first-order retinal arteriole is approximately 125 μm, the
endoscope allows intraoperative visualization of fine retinal capillaries,
useful for delineating retinal tissue from fibrovascular membranes in these
complex cases. The image resolution with the endoscope is, however, lower
than the natural surgeon’s human retina when seen directly with a microscope
and is akin to the difference between a video filmed at standard definition
versus high definition, respectively. This resolution trade-off between the
endoscope and microscope view is compensated by the ability to access areas
that cannot normally be visualized intraoperatively (see below). Conversely,
with the wide FOV, a panoramic and less magnified view of the retina is
possible by illuminating from the mid-vitreous cavity. Endoscopes are
available in straight or curved configurations. A curved probe can be very
useful for reaching diametrically across the posterior pole of the lens while
avoiding iatrogenic lenticular touch, enabling visualization of very anterior
structures such as the vitreous base, ciliary body, or posterior iris. However,
in the authors’ experience, curved probes can be more difficult to work with
due to the additional axis of rotation compared with a straight instrument.
Clinically Relevant Optical Properties

The endoscope has two surgically relevant properties, namely unique (a)
optical property and (b) perspective (angle of surgical visualization),
conferring advantages on microscope-based viewing systems in certain
clinical scenarios.
Optical property
In pediatric vitreoretinopathies such as advanced ROP (stage 4A, 4B, or 5)
and FEVR, anterior retinal pathology is one of the hallmarks of disease.
Being able to visualize and dissect the anterior hyaloid face and vitreous
cortex, whether it is enmeshed within an opaque retrolental fibrovascular
plaque or remains relatively transparent, is particularly important. Pars plana
vitrectomy is most commonly performed using microscope-based contact or
noncontact viewing systems, where light from an endoilluminator is first
reflected off vitreoretinal tissue, subsequently transmitted through the
patient’s pupil plane and cornea, before being seen by the surgeon at the end
of the eyepieces of an operating microscope. Using this technique, it is
difficult to make out a distinct anterior hyaloid face as light is transmitted
through it. In contrast, with the endoscope, the source of endoillumination
and surgeon’s view are coaxial, as the visualization of anatomical structures
is based on the capture of reflected rather than transmitted light. To
demonstrate this, we used lightly frosted cellophane tape to represent the
anterior hyaloid face (Figure 62-1) and placed this over printed paper to
signify the retina. Evidently, visualization of an almost transparent anterior
hyaloid face is better with reflected than transmitted light.
FIGURE 62-1 Simulated view of the vitreous during pars
plana vitrectomy. The anterior vitreous is represented by
lightly frosted cellophane tape. The retinal surface is
represented by alphabets printed on a sheet of reddish-pink
paper. A:With standard microscope (conventional) wide-
angle viewing systems (e.g., BIOM), illumination is
separated from the surgeon’s view through the operating
microscope. In the image below, the cellophane tape is
barely visible overlying the alphabets. B:With endoscopy,
illumination and the surgeon’s view are coaxial. In the
image below, the same frosted tape is now almost opaque
and highly visible, completely obscuring the underlying
alphabets.
Perspective
The anatomical perspective of the endoscope is unique. Microscope-based
(conventional) contact and noncontact viewing systems enable visualization
of posterior segment structures through a bird’s-eye or top-down view
through the pupil, with the most anteriorly visible structure being the vitreous
base and occasionally the pars plana. Even with deep scleral indentation,
anatomical structures anterior to that, that is, the pars plicata, ciliary body,
lens equator, and posterior iris surface, are almost impossible to directly
visualize and manipulate surgically. As ophthalmologists, the bird’s-eye view
is the anatomical perspective with which we are most familiar and
comfortable based upon our experience using direct and indirect
ophthalmoscopes. In contrast, the endoscope provides an unfamiliar side-on
perspective relative to the patient’s visual axis, which is an angle of
visualization approximately perpendicular to the microscope-based bird’s-eye
surgeon’s viewing axis. Through a pars plana sclerotomy, the vitreous base,
pars plana, pars plicata, ciliary body, posterior lens surface, and retroiridial
surface can be directly visualized without distortion from spherical aberration
at the edge of a lens or deep scleral indentation (Figure 62-2). This is
advantageous in pediatric vitreoretinopathies in which adequate dissection of
fibrovascular membranes involving the aforementioned structures can be the
difference between surgical success and failure.
FIGURE 62-2 Intraoperative view through a 20-gauge
higher resolution endoscope. A:This is a child with
aphakic glaucoma and rhegmatogenous RD. Normal
anterior anatomical structures are clearly visualized. C,
ciliary processes; P, pars plana; H, hyaloid (anterior
hyaloid face). Note that the anterior hyaloid face is seen
very clearly, due to the use of reflected rather than
transmitted light. B:This is a neonate with stage 4B ROP.
Anterior traction RD is present. Endoscopy enables direct
visualization and guidance of a 20G MVR blade through
the pars plicata for safer sclerotomy formation. There is
much less space than normal for safe sclerotomy formation
as the anterior TRD is in close proximity to the pars
plicata. Endoscopic guidance reduces the risk of iatrogenic
lens or retinal injury. L, lens; M, MVR blade (20 gauge);
C, ciliary processes; R, retina.
The unique anatomical perspective of the endoscope comes into its own,
when an unhindered direct view behind the iris or under an immobile opaque
tissue plane is required. For instance, in anterior TRD in stage 4 ROP or
FEVR, the RD can be drawn close to the lens or pars plicata, precluding safe
trocar insertion during sclerotomy formation, due to the risk of retinal injury
from an unseen trocar tip entering the eye. The endoscope enables
unhindered direct visualization of the pars plicata and its surrounding areas
normally obscured by iris, thus facilitating safer trocar insertion under direct
internal visualization. For opaque retrolental plaques, the surgeon can
alternate the position of the endoscope between visualizing above and below
opaque fibrovascular membranes. Instead of relying on indirect clues, direct
visualization of otherwise inaccessible safe preretinal dissection planes is
possible, reducing the risk of iatrogenic retinal break formation. Minimizing
the risk of complications is critical for surgical success in these zero error
tolerance eyes. In ROP, unlike rhegmatogenous RD or diabetic TRD in
adults, an iatrogenic retinal break often portends a poor prognosis (14).
Learning Curve
There are several key aspects to the learning curve of endoscopic vitrectomy
(15). Firstly, the surgeon’s view is derived from a video feed projected onto a
monitor, rather than through the eyepieces of the microscope. This is a two-
dimensional image, necessitating the greater use of nonstereoscopic clues
such as shadows and inferring object distance by recognizing the size of
anatomical landmarks relative to the known instrument gauge. It is
noteworthy that intraocular shadows formed with an endoscope provides a
different perspective from conventional viewing systems due to the differing
angle of reflected light captured, that is, coaxial versus noncoaxial
illumination, respectively. Secondly, the intraoperative view is derived from a
live video feed to a monitor, dissociated from the surgeon’s manipulation of
surgical instruments. This requires some relearning of hand-to-eye
coordination and feedback. Thirdly, the advantage of being able to resolve
detail down to at least 20 μm equates to a highly magnified surgical
perspective. It is essential that greater care is taken to maintain awareness of
relative instrument positions within the narrow FOV to avoid inadvertent
retina or lens touch. Fourthly, the endoscope adds rotation to the x-, y-, and z-
axes of movement of conventional viewing systems. Fifthly, there is a need
for regular adjustment of illumination levels. The optimum luminosity
required to illuminate anatomical structures can vary significantly depending
on the distance from the endoscope. The closer the endoscope is to the object
of interest, the less light is required to avoid whiteout. This is the result of the
narrower dynamic range of a digital image capture system such as the
endoscope, relative to the human eye, predisposing to loss of some detail in
the high or low extremes of illumination. Taking these factors into account,
the initial experience with the endoscope is unfamiliar and can be
disorientating. In the author’s experience, the use of a wet lab can be very
useful for accelerating the familiarization process. Having a microscope
simultaneously set up enables the surgeon to switch readily between both
viewing systems, and this is useful for maintaining endoscopic orientation.
The additional surgical time that is initially required is time well invested, as
the ability to directly visualize surgical planes in complex surgical scenarios
pays dividends in terms of reduced complications and improved clinical
outcomes.
Clinical Application
In 1981, the first endoscopy-assisted vitreous surgery was described (16). In
adults, there is good evidence for its use in rhegmatogenous (17) and TRD
(18), PVR (19), endophthalmitis (20,21), penetrating eye injury (22),
intraocular foreign body (22), retained lens fragments (23), and sutured
posterior chamber intraocular lens placement (24). There is growing evidence
of its use in children (24,25). At the time of writing, the authors have
submitted their surgical outcomes of endoscopic vitrectomy in children for
publication. This would likely represent the largest endoscopic surgical series
in children to date. The first unpublished study from Children’s Hospital Los
Angeles (Lee & Wong) present preliminary surgical outcomes data on 210
endoscopic vitrectomies in 154 eyes, with the most common indications
being ROP (20%), trauma (20%), and pars planitis (6%). Across all RD eyes
(tractional and/or rhegmatogenous), 63% retinal reattachment was achieved.
In a second unpublished study by one of our authors from Great Ormond
Street Hospital in London (Wong), 57 consecutive eyes with stage 4A or 4B
had endoscopic vitrectomy. None of the eyes (0%) required primary
lensectomy (or developed treatment-requiring cataract) within 12 months
postoperatively, with a 83% overall primary retinal reattachment rate, highest
in stage 4A (92%). These data support the authors’ clinical experience of
endoscopic vitrectomy in complex pediatric vitreoretinopathies, particularly
ROP traction RD, where a high level of lens preservation can be achieved
with good anatomic reattachment rates.
The optical properties and anatomical perspective of the endoscope and
microscope-based wide-angle viewing systems are complementary. As such,
the authors utilize endoscopy as an adjunct to microscope-based contact and
noncontact wide-angle viewing systems in almost all pediatric vitrectomies.
In experienced hands, pediatric vitrectomies can be performed exclusively
with endoscopy where necessary. Examples of the authors’ indications for
endoscopy are described hereinafter.
The most obvious advantage of endoscopy is its ability to circumvent any
anterior segment opacity precluding visualization of the posterior segment
with conventional (transcorneal) viewing systems. The clinical utility is
relevant to both children and adults with vitreoretinal disease. Although a
temporary keratoprosthesis (26) is an alternative by enabling visualization of
the posterior segment with conventional viewing systems, the patient is
committed to having a penetrating keratoplasty with active ocular disease,
possibly increasing the risk of graft failure. Another option is a permanent
keratoprosthesis, which has significant risks. A retrospective review of 35
cases revealed a 23% vitreoretinal complication rate, which included
intraoperative choroidal hemorrhage, rhegmatogenous RD, vitreous
hemorrhage, and endophthalmitis (27).
Sclerotomy placement in adults is generally straightforward with a few
exceptions such as penetrating trauma, hypotony, and choroidal detachment.
In contrast, in complex pediatric vitreoretinopathies such as advanced ROP,
FEVR, and PFV, atypical anatomical landmarks are commonplace. The retina
may be drawn anteriorly to the posterior lens capsule and, rarely, the
posterior iris surface, precluding safe pars plana sclerotomy placement. The
anatomy of the pars plana itself may be abnormal. The endoscope can be
helpful for internally directing transscleral incisions, minimizing concerns
over safe entry.
One of the major limiting factors of anatomical success in children with
TRDs in diseases such as ROP, FEVR, and PFV is the adequacy of
membrane dissection in the difficult to access area encompassing the anterior
retina, ciliary body, posterior lens capsule, and anterior hyaloid face.
Visualization of these structures is suboptimal and at times impossible with
conventional wide-angle viewing systems. It is not widely appreciated that
the quality of the surgical view with the endoscope is equally good whether at
the macula or pars plicata, because image degradation or distortion at the
edge of the lens of conventional viewing systems does not apply. In complex
pediatric vitreoretinopathies dominated by anterior pathology, the ability to
maintain good visualization in difficult-to-access surgical spaces is highly
advantageous.
In PFV, a stalk typically extends anteroposteriorly toward the posterior
lens surface. Microscope-based viewing systems provide a bird’s-eye view,
through the cornea and lens, of the top of the stalk. This enables an accurate
assessment of the size of the retrolental (anterior hyaloid) plaque, its position
relative to the visual axis, and thus the amblyogenic impact and potential
benefit of surgery. In PFV cases with significant posterior involvement, the
retina may be drawn up along the length of the stalk. The presence and extent
of this is potentially difficult to assess with microscope-based viewing
systems, where the surgeon’s line of sight is roughly parallel to the length of
the stalk. The anteroposterior position at which the stalk is transected is
critical to avoid an iatrogenic retinal break and rhegmatogenous RD. The
endoscope provides a view of the side rather than top of the stalk, thus
enabling the surgeon to track retinal vessels up along the length of the stalk
until it terminates below a safe surgical transection zone.
In children, the primary retinal reattachment rate following pars plana
vitrectomy for rhegmatogenous RDs with or without associated trauma is
lower than adults (15,28). This is partly due to the more aggressive PVR. The
optical properties and unique perspective of the endoscope can aid better
delineation and more complete removal of hyaloidal tissue and pre- and
subretinal membranes, potentially reducing the risk of PVR formation and
reproliferation. The ability to visualize below a tissue plane with the
endoscope can similarly be applied to the retina for removal of subretinal
membranes and bands.
There is a risk of vitreous incarceration in sclerotomy ports during
vitrectomy in pediatric retinal diseases such as ROP and FEVR, due to the
anterior nature of these diseases. Resultant unrecognized postoperative
vitreous traction and retinal break formation can lead to a poor outcome.
Endoscopy enables direct visualization of the internal aspect of the
sclerotomy facilitating adequate vitreous clearance (29).
CONCLUSIONS
The endoscope has unique optical properties that are complementary to
microscope-based contact and noncontact viewing systems. The ability to
visualize and surgically manipulate difficult to access vitreoretinal surgical
planes from a different perspective can be the difference between success and
failure in selected pediatric vitreoretinopathies. The endoscope has become
an indispensable part of the authors’ surgical armamentarium. Its utility value
will be even greater in the future when high-definition stereoscopic
visualization becomes a reality.
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(Phila) 2018;7(3): 200–207.
SECTION X
Management of Vitreoretinal
Conditions of Infants and Children
63
Pediatric Rhegmatogenous Retinal
Detachment
Lejla Vajzovic
INTRODUCTION
Pediatric rhegmatogenous retinal detachments (PRRDs) are complex, rare
occurrences and are often related to trauma, congenital abnormalities, genetic
syndromes, high myopia, and prior ocular surgery. This is in contrast to the
most common causes of rhegmatogenous retinal detachment (RRD) in adults,
which is related to retinal breaks that develop during a posterior vitreous
detachment (PVD).
EPIDEMIOLOGY
In the general population, RRD has an annual incidence of approximately 10
cases per 100,000 population (1). PRRD is much less common with an
annual incidence of only 0.38 to 0.69 per 100,000 in the population under age
20 years (2) but makes up 3% to 13% of all retinal detachments (1,3–8). The
average age at presentation of PRRD is between 7 and 13 years of age
(1,2,9,10). Trauma is the most common cause of PRRD and accounts for
about 40% of all RRD in children compared to 11% of RRD in the adult
population. There is a significant male preponderance of PRRD, especially in
trauma-related cases (up to eighty percent), but even in nontraumatic PRRD
(1,2,9,10).
Congenital abnormalities are also common causes of PRRD and account
for 35% to 56% of all cases in Western populations and 12% to 17% in case
series from East Asia. The incidence is even higher among younger patients
<10 years of age. Geographic differences in rates of PRRD secondary to
congenital abnormalities can be explained by regional differences in other
conditions that predispose to RRD, such as the high incidence of high myopia
in Asia (11–13) and/or of atopic dermatitis in Japan (14). Intraocular surgery,
most often cataract surgery in eyes without trauma, presents with high
incidence of PRRD (15–17).
INDICATIONS
Children often do not recognize or report symptoms of retinal detachment.
Thus at presentation, the pediatric pathology is typically advanced often with
macular involvement, proliferative vitreoretinopathy, chronic duration, and
poor visual acuity. Often there is a high incidence of bilateral pathology.
These features pose significant management challenges. Children with PRRD
may require multiple surgeries and may have ocular malformations, including
associated hereditary and congenital nonhereditary pathologies. For all of
these reasons, visual potential after repair of PRRD may be limited even after
successful surgical repair because of chronicity and the effects of deprivation
amblyopia (2,18–20).
Although PRRD is generally repaired successfully with newer
vitreoretinal surgical techniques, it may require multiple surgical procedures
due to late and advanced presentation. Therefore, there needs to be a clear
understanding that the decision for surgery may incur significant external and
emotional pressures on the family and the patient. Despite these challenges,
in many instances repaired eyes have good vision, and at times better vision
than the fellow eyes; therefore, surgical management is often recommended.
The possibility of bilateral ocular disease also supports surgical intervention
in the presenting eye as well as consideration of therapy for the fellow eye.
However, at times surgical management may not be the best option if PRRD
is too advanced with no light perception vision and secondary sequela such as
neovascular glaucoma, hypotonus, and phthisical eye. Comfort care and a
referral to ocularist might be recommended in these instances. It is always
important to perform a careful examination and potential prophylactic
treatment of the fellow eye.
CLINICAL PRESENTATION
Poor visual acuity on routine screening, strabismus, or leukocoria are
common presenting symptoms. Because PRRD may be chronic due to limited
detection, the timing of retinal detachment can range from several weeks to
many months. In studies that include all age groups, the mean time to RRD
presentation is usually <30 days (1) but is longer in children and can be
difficult to determine. The visual acuity difference is often picked up when
the fellow eye becomes involved. Hence, pediatric patients typically present
with poor visual acuity, greater delay in diagnosis, and with a higher
incidence of macular involvement in 75% to 85% as opposed to 50% in
adults (2,10,18,21–23). Also, pediatric patients also have much higher rates
of PVR, between 30% and 60%, on presentation and with 10% to 20% with
at least PVR grade C (2,9,10). Lastly, they more frequently present with
bilateral retinal detachment than adults (24). Up to 25% of affected pediatric
patients have a diagnosis of bilateral retinal detachment on presentation or
during follow-up period (Figure 63-1) (9). This percentage is higher in
patients with congenital or hereditary abnormalities. When traumatic
etiologies are excluded, pediatric patients with unilateral PRRD have retinal
tears and lattice degeneration.
FIGURE 63-1 Stickler syndrome type 1 and bilateral
retinal detachments. A 10-year-old female presented with
sudden loss of vision (20/200) in both eyes. On clinical
examination, dense vitreous veils, macula involving retinal
detachment in both eyes was noted with superotemporal
giant retinal tear in the right eye and posterior
superotemporal retinal tear in the left eye. Widefield
fundus photo and optical coherence tomography
demonstrate these findings in the right eye (A) and left eye
(B). She underwent sequential retinal detachment repair a
week apart with scleral buckle, vitrectomy,
perfluorocarbon, laser photocoagulation, and long-acting
gas tamponade in both eyes. Visual acuity improved to
20/80 in the right eye (C) and 20/50 in the left eye (D).
The PRRD appearance is typically with demarcation lines, subretinal PVR,

and retinal microcysts indicating chronic process. Although a single retinal
break or a tear is commonly the cause of PRRD, giant retinal tears (GRTs)
(Figure 63-2) are also reported in up to 15% in some case series (2,10).
PRRD are generally seen associated with syndromes such as Stickler
syndrome type 1 and in high myopes (25) (Figure 63-1). On the other hand,
retinal dialysis occurs as a consequence of trauma (nonsurgical and surgical)
(2,24) and is typically located inferotemporally (Figure 63-3).
FIGURE 63-2 Traumatic retinal detachment with giant
retinal tear. A 12-year-old female presents with hand
motion vision 2 days after trauma to the right eye.
A:Vitreous hemorrhage and superior retinal detachment
secondary to giant retinal tear from 8 to 3 o’clock was
noted on widefield fundus photograph. B:B-scan
echography demonstrated total retinal detachment with
more bullous superior portion. C:Patient underwent scleral
buckling with broad anteriorly placed, low lying buckle
and 25 gauge vitrectomy with perfluorocarbon use,
endolaser photocoagulation, and longer-acting gas
tamponade. Visual acuity improved to 20/50
postoperatively, and retina is shown attached on widefield
fundus photograph. D: The retinal layers, including
ellipsoid zone, are present across the macula after
reattachment, despite the limited foveal pit.
FIGURE 63-3 Retinal detachment associated with
inferotemporal traumatic retinal dialysis. A 10-year-old
male presents with peripheral visual field loss and 20/20
visual acuity several months after trauma to the left eye. A,
B:Widefield fundus photograph and optical coherence
tomography demonstrate inferotemporal retinal
detachment secondary to retinal dialysis that is not seen in
these images. C, D:Patient underwent scleral buckling
with broad anteriorly placed, low lying buckle and
cryotherapy to dialysis edges. The retina reattached, and
he regained peripheral vision. Widefield fundus
photograph of the attached retina (C) demonstrates a
curved demarcation line near the macula and consistent
with chronicity, inferotemporal cryotherapy scar, and
scleral buckle indentation. Optical coherence tomography
(D) demonstrates resolution of subretinal fluid with retinal
reattachment and good foveal contour. Loss of outer
retinal layers temporally (on OCT) in the area previously
detached is also consistent with a chronic detachment.
ETIOLOGIES
Many of the conditions associated with PRRD are covered in depth elsewhere
(see Chapters 37, 60 and 61). The vasoproliferative retinopathies, particularly
retinopathy of prematurity, persistent fetal vasculature syndrome, and familial
exudative vitreoretinopathy, are generally associated with traction-exudative
retinal detachment and may present with a combined mechanism (see
Chapters 55, 65 and 66).
TRAUMA
Trauma, nonsurgical or surgical, is a leading cause of PRRD and often occurs
late due to the support of a well-formed vitreous. Because spontaneous RRDs
in a child with normal ocular anatomy and without hereditary or congenital
association are very rare, trauma should be considered regardless of history
(9,10,26,27). Nonsurgical traumatic PRRD may present with vitreous
hemorrhage limiting posterior pole visualization and if dense typically
requires earlier pars plana vitrectomy (PPV) to improve vision and reduce
amblyopia and to address retinal traction and repair PRRD. This is often in
combination with a scleral buckle. Traumatic injury to other tissues, such as
cornea, iris root, lens, zonules, and optic nerve, should be considered in
surgical assessment and planning. PRRD has been reported in up to 15% of
cases of traumatic pediatric cataract and should be looked for in the
preoperative evaluation (27). Surgical management of posterior segment
trauma is detailed in Chapter 69.
AFTER OCULAR SURGERY

There are reports of PRRD following ocular surgical procedures (28), most
commonly pediatric cataract extraction (9,15,16,29,30); however, good
incidence data are lacking for pediatric and even for iatrogenic adult RRDs
following ophthalmic surgeries. Published pediatric series demonstrate a high
rate of PRRD after cataract extraction whether the eye was left aphakic (3.2%
at 7 years (30)) or had modern surgery with an intraocular lens placed within
the capsular bag (5.5% at 10 years (16) and 7% at 20 years (15)). The onset
of postcataract RRD is delayed in children compared to adults, with median
onset of detachment in children at 5 to 9 years after the surgery (15,16,30).
Several series show the increased risk of PRRD after cataract surgery for
children with intellectual disability: 23% rate of PRRD (15) or 12 times
higher risk (16). An increased risk of 16% with PRRD was also reported for
children with other ocular or systemic anomalies (15). Eyes progressing to
PRRD following ophthalmic surgical procedures, for example, cataract,
strabismus, corneal transplantation, or glaucoma tube shunt often fare
considerably worse than previously unoperated eyes presenting with PRRD.
INHERITED
Several inherited disorders primarily connective tissue disorders are
associated with RRD.
Stickler syndrome is an autosomal dominant genetic condition that
presents in 1:7,500 live births and is the most frequent inherited connective
tissue disorders. It is the most common cause of inherited, childhood retinal
detachments, but it continues to be widely underdiagnosed. At least 6
subgroups have been characterized according to genetic abnormalities of type
II, IX, or XI collagen. These structural proteins are principally and
collectively expressed in the eye and in articular and hyaline cartilage. Thus,
affected patients may present with premature arthropathy and classic
orofacial, auditory, and ocular features (25,31) (see Chapter 37).
If Stickler-related PRRD is suspected, the presence of a GRT is likely.
PPV and/or scleral buckle should be placed especially if there is an inferior
GRT component. If a PPV is planned, the vitreous traction is removed as
carefully as possible and the devitalized retina anterior to the GRT is
removed to decrease a scaffold for future anterior PVR or ischemic signal. In
young children, the vitreous can be very adherent to the highly mobile retina
and may be difficult to completely remove. Nonetheless, it is important to
remove the vitreous traction as much as possible. Perfluorocarbons are used
to help unfold and flatten the retina. Endolaser is performed with 2 to 3 rows
along the posterior, side horns and anterior edges of the GRT. If the anterior
horns cannot be treated adequately, cryotherapy may be placed at the anterior
horns adjacent to the ora serrate. Long-acting gas or silicone oil is used as a
tamponade agent depending on the location of the GRT, positioning
requirements and maturity of patient in maintaining position. Attention to
fellow eye and prophylactic treatment with laser, cryotherapy, or scleral
buckling is recommended (32), but evidence as to the best approaches
remains scarce.
Marfan syndrome is an autosomal dominant genetic disorder that results
in weakening of connective tissue in the musculoskeletal, cardiovascular, and
ocular organ systems. It is the second most common inherited connective
tissue disorder with an incidence of between 1/5,000 and 1/20,000. The
abnormality in over 80% of Marfan patients involves defects in the protein
fibrillin 1 (FBN1) on chromosome 15, a structural component of microfibrils
found in connective tissue throughout the body. More than 500 different
mutations in the FBN1 gene have been identified. The major ocular
abnormality in Marfan syndrome is ectopia lentis or lens subluxation or
dislocation. Clinically, ectopia lentis is bilateral in 60% to 87% of Marfan
patients. The most common direction of dislocation on exam is
superotemporal. Retinal tears and detachments are also quite common in
patients with Marfan syndrome due to high myopia, excess lattice
degeneration, vitreous liquefaction, choroidal and scleral thinning, and
vitreous traction from ectopia lentis. Each of these changes is thought to be
related to fibrillin alterations. Retinal detachment occurs in 5% to 26.5% of
patients with Marfan syndrome and is bilateral in 30% to 42% of these cases
(33,34).
Therefore, when addressing PRRD in patients with Marfan syndrome,
lens status and its stability need to be taken into careful consideration
preoperatively. The subluxed lens is typically addressed at the time of PRRD
and removed through a pars plana approach. The patient is left aphakic with a
surgical peripheral iridotomy to avoid pupillary block secondary to
tamponade agent. Scleral thinning secondary to Marfan syndrome should also
be noted preoperatively and sclerotomy size and sites carefully chosen and
sutured at the end of procedure. Use of small gauge instrumentation is often
important. In rare extreme scleral thinning cases, scleral patch grafts may be
required to cover thin sclera and sclerotomy sites. Use of scleral buckles in
the setting of diffuse scleral thinning should be avoided as there is a risk of
buckle intrusion into the globe.
Ehlers-Danlos syndrome (EDS) is also an inherited connective tissue
disorder caused by defective collagen synthesis, the main features being
hyperelasticity and vulnerability of the skin, recurrent bleeding from fragile
blood vessels, and secondary deformities of the joints. Ocular involvement is
a rare occurrence and may include corneal and scleral rupture from minor
blunt injury, lens displacement, and RRD (35,36).
The surgical approach in EDS patients is similar to Marfan syndrome
described above where subluxation and scleral thinning may add to
complexity of PRRD repair. These features should be carefully considered
preoperatively, discussed with the patient and family and addressed at the
time of PRRD repair.
It is often important to impose more frequent examinations to assess for
retinal pathology and prevent retinal detachment and vision loss in patients
with the above diseases. It is also important to teach and reinforce the
symptoms associated with retinal detachment to family members and the
patients. It is important to perform retinal examinations on both eyes
especially while the patient is under anesthesia. It is also important to
examine family members.
NONTRAUMATIC INFEROTEMPORAL
RETINAL DIALYSIS (“JUVENILE
DIALYSIS”)
A retinal dialysis is a circumferential break occurring at the ora serrata as
opposed to the more typical posterior margin of the vitreous base in giant
tears. Dialyses, in general and following trauma, are more common in the
inferotemporal quadrant (Figure 63-3). However, most dialyses detected
nasally or superiorly are associated with a history of preceding blunt trauma.
Idiopathic retinal dialyses accounted for approximately 2% of retinal
detachments in one series and typically occurred in the inferotemporal
quadrant of young adult eyes. Retinal detachment associated with retinal
dialysis often is subclinical without vision loss or visual symptoms.
Associated findings include demarcation lines, retinal cysts, peripheral
microcystoid degeneration, and yellow-white vitreous opacities (Figure 63-
3A). PVR is rarely seen, likely due to the overlying vitreous base serving as
tamponade limiting access of cells to the central vitreous cavity (37).
Idiopathic retinal dialyses typically present in emmetropic males in the
second decade of life. They may be bilateral and may involve multiple family
members (38–41). Therefore, in patients with bilateral nontraumatic retinal
dialyses, examination of family members is indicated. Depending on the
extent of retinal detachment, these detachments typically are addressed with
scleral buckling and/or laser or cryoretinopexy.
ATOPIC DERMATITIS
Atopic dermatitis, a chronic pruritic skin condition, is an allergic form of
eczema primarily affecting infants, children, and young adults. The onset
usually is in infancy, often with a positive family history of atopic disease.
The prevalence is on the order of 10% in the United States and up to 20% in
Japan. The incidence of atopic dermatitis is thought to be rising with retinal
detachments representing approximately 3% of RRDs seen in Japan. RRD
typically occurs in early adulthood but may be seen in late adolescence
(14,42).
Eyelid dermatitis, blepharitis, conjunctivitis, cataract, and keratoconus
are recognized anterior segment associations. Retinal detachment has been
reported in 6% to 10% of Japanese patients. The peripheral fundus is
characterized by a “fluffy” appearance with signs of vitreous base
contraction, including wrinkling of the pars plana epithelium and oral
dialyses. Retinal breaks often are multiple and located at the ora serrata or in
the nonpigmented ciliary body epithelium. Retinal detachments are generally
low lying and asymptomatic, although GRTs occur as well. Intense pruritus
prompts frequent eye rubbing and face slapping, which are believed to play a
role in causing retinal or ciliary body breaks. Scleral buckle with thorough
peripheral retinopexy is advocated. The presence of dense anterior chamber
angle pigmentation in patients with atopic dermatitis can be a sign of breaks
of the peripheral retina or ciliary body epithelium and should be an indicator
to perform a meticulous peripheral retinal examination (14,42–47). In cases
where retinal detachments involve GRTs, the surgical approach is similar to
above described approach for GRT associated with Stickler syndrome.
EXAMINATION AND DIAGNOSTIC

STUDIES
In-office clinical examination and visual acuity determination are often
challenging in young patients but are important as a starting point. Ancillary
testing such as widefield fundus photography with or without fluorescein
angiography (Figure 63-1), optical coherence tomography (OCT) (Figure
63-1), and B-scan ultrasonography (Figure 63-2) are often critical for better
posterior segment visualization and assessment of PRRD before surgery.
Thorough history should include past birth history, past medical history,
family history, and review of systems to determine potential clues as to the
etiology of the PRRD. Clinical examination of parents and siblings including
widefield imaging with or without fluorescein angiography and OCT may be
indicated to help detect familial exudative vitreoretinopathy,
oculoarthropathies, or retinal degenerative conditions, as examples.
In the operating room, a more thorough examination is performed under
anesthesia and includes external evaluation, including facial features,
dermatologic abnormalities, joint and digital appearances, intraocular
pressure measurement, anterior segment examination and corneal diameter
measurements, extended ophthalmoscopy, widefield fundus imaging with
fluorescein angiography, and OCT. A and B-scan ultrasonography is
performed where indicated. Thorough dilated and scleral depression
examination of the companion eye is critical as pathologic findings tend to be
bilateral. It is not uncommon to focus treatment initially on the better seeing
eye. It is also important to consider timing for prophylactic intervention in the
fellow eye if no obvious abnormalities are found on examination under
anesthesia (EUA).
GENERAL CONSIDERATIONS FOR

SURGERY
Pediatric patients especially newborns, infants, and toddlers have much
smaller eyes with different surgical landmarks compared to adult eyes. In
addition, they are not just smaller adult eyes, but they have different
anatomical considerations, dense vitreous gel, increased vitreoretinal
adhesion, and increased propensity for membrane proliferation. All of these
need to be taken into account when performing pediatric retinal detachment
repair. However, the treating principals are the same as in adults: the goal is
to identify, treat with retinopexy, support all retinal breaks, and release
traction. Single-procedure anatomical success rates range from approximately
50% to 90% (48) due to advanced presentation. Additionally, silicone oil is
commonly used as a tamponade agent in cases with PVR and requires a
subsequent surgery for removal. Hyaloidal contraction and fibrovascular
proliferation leading to traction is much more likely to be a contributing
factor in PRRDs than in retinal detachments in adults. Although scleral
buckle alone is the preferred surgical approach in pediatric patients (Figure
63-3), substantial traction may require a combined approach with scleral
buckle and/or vitrectomy (Figure 63-2). In addition, lens sparing techniques
are preferred to optimize visual rehabilitation and limit amblyopia, but the
lens may need to be removed to facilitate thorough vitreous removal. The
ultimate goal is to repair the retinal detachment and minimize recurrences
and, therefore, aphakia with thorough vitreous and membrane removal may
be the better option. In some cases, phacoemulsification with intraocular lens
placement may be of benefit at the time of vitrectomy.
Preoperatively and intraoperative, surgeon should plan for equipment
needed including a scleral buckle tray, type of scleral buckle, small gauge
vitrectomy system (if vitrectomy considered), adjuvant materials for
vitrectomy (perfluorocarbon liquid, triamcinolone, gas, oil), and retinopexy
with cryotherapy or laser (see Chapters 60 and 61).
SURGICAL TECHNIQUES
Scleral Buckling
A scleral buckle is often used as a primary cerclage or at times as an
encircling band in combination with vitrectomy. It is important to precisely
localize break(s) on a buckle or encircling band in PRRD. The buckling
effect should be broad and low, and there should be adequate treatment with
laser or cryotherapy to create a permanent chorioretinal adhesion. It is
important not to cause significant distortion of the globe, which may further
open the retinal break, creating a “fishmouth” effect allowing vitreous fluid to
enter the subretinal space. When performing scleral buckling in infants or
children <2 years of age, thinner and narrower elements are used but the
same principles apply. The band often ends up being placed just anterior to
the equator. It is important to cut the encircling band in infants or small
children after 3 to 6 months to allow for eye growth. Refractive errors are
also treated aggressively and continuously reassessed in the postoperative
period to maximize visual outcomes. Further specifics about scleral buckling
are discussed in Chapter 60.
Vitrectomy
Today, vitrectomies are mostly performed in small gauges using noncontact
and contact wide-angle viewing systems to visualize the peripheral retina;
some of these lenses are smaller in lateral dimension to allow access to the
infant eye, but adult lenses can also be helpful depending on the size of the
eye. Pediatric patients have lower systolic blood pressure compared to adults
and perfusion of the central retinal artery should be monitored during surgery
to assure that infusion pressure or scleral indentation is safe. In the anterior
trans-limbal vitrectomy approach, the infusion is provided with an anterior
chamber maintainer. Creation of a PVD is an important step, if it can be
safely performed, in the successful management of a retinal detachment
during vitrectomy. This may be essential in managing RRD in older children.
Diluted, sterile triamcinolone can highlight vitreous to help distinguish it
from retina and reduce the creation of a vitreous schisis so that vitreous
removal is achieved. In eyes with proliferative vitreoretinopathy, it is
preferable to perform segmentation instead of delamination when removing
preretinal membranes due to the firm vitreoretinal attachments in children. A
bimanual approach with use of a chandelier illumination system is helpful.
Use of heavy liquid, perfluorocarbon, is preferred in pediatric retinal
detachments to flatten the retina and avoid posterior drainage retinotomies.
Retinotomies or additional breaks are best avoided, because extensive fibrous
proliferation can occur postoperatively and lead to recurrent retinal
detachment. If a retinotomy is unavoidable, it is usually recommended to
create one superiorly and as close to the ora serrata as possible. In the case of
retinal detachment associated with coloboma, there is also an increased risk
of subretinal movement of perfluorocarbon liquid through the colobomatous
defect. Therefore, use of perfluorocarbon liquid should be avoided in these
cases (Figure 63-4B). For vitreous tamponade, the use of long-acting
intraocular gas, such as C3F8, is ideal but may not be feasible if the patient is
unable to be compliant about positioning or there is difficulty removing all of
the cortical vitreous. Therefore, silicone oil is often used except in
colobomatous defects to reduce the risk of silicone oil in the subretinal space
or intracranial cavity. In children, silicone oil emulsifies much more quickly
and an “inverse hypopyon” may be seen (Figure 63-4). Further specifics
about vitrectomy are discussed in Chapter 61.
FIGURE 63-4 Pediatric patients after retinal detachment
repair demonstrate: a pseudohypopyon secondary to
emulsified silicone oil (A, slit-lamp anterior segment
photograph) and central and inferior corneal endothelial
droplets secondary to retained perfluorocarbon liquid (B,
Retcam anterior segment image during exam under
anesthesia).
Inherited Conditions: Stickler Syndrome, Marfan

Syndrome, and Ehler-Danlos Syndrome
PRRDs commonly present bilaterally in inherited disorders and are treated
aggressively in both eyes with scleral buckle alone, vitrectomy alone, or a
combination. Patients with Stickler syndrome may benefit with a combined
buckle/vitrectomy, but this decision is also dependent on presentation,
presence of vitreous hemorrhage, and GRT location. Perfluorocarbon liquid
is helpful in GRTs. Patients with Marfan syndrome and Ehler-Danlos have
very thin sclera making placement of scleral buckle difficult secondary to the
risk of a scleral rupture. Therefore, a vitrectomy may be decided to repair
PRRD.
In patients with genetically confirmed Col2A1 mutation in Stickler
syndrome type 1, prophylactic treatment of the fellow eye with laser
photocoagulation, cryotherapy, or scleral buckling have been considered
(25,49,50). The protocol for cryotherapy entails 360 degrees of cryotherapy
just posterior or bridging the ora serrata. In the retrospective comparative
case series review of 487 patients with type 1 Stickler syndrome, the bilateral
control group had a 7.4-fold increased risk of retinal detachment compared to
bilateral cryoprophylaxis group and the unilateral control group had a 10.3-
fold increased risk of retinal detachment compared to the unilateral
cryoprophylaxis group (50). Data regarding prophylactic laser and outcomes
are more limited (25,51). Stickler syndrome is addressed in greater detail in
Chapter 37.
Traumatic or Nontraumatic Retinal Dialysis

Nontraumatic retinal dialysis is commonly known as idiopathic “Juvenile
Dialysis.” These breaks are typically asymptomatic unless the macula is
involved and present in inferotemporal quadrant. Asymptomatic retinal
detachments secondary to retinal dialysis can be repaired with scleral buckle
or stabilized with retinopexy. Repair of symptomatic retinal dialysis typically
is accomplished by a scleral buckling procedure (Figure 63-3). Light
cryopexy is applied along the dialysis edge. Subretinal fluid drainage usually
is not necessary for recent-onset retinal detachments from dialyses but is
helpful in chronic retinal detachments from dialyses with viscous subretinal
fluid. Subretinal bands are seen in chronic retinal detachments from dialysis,
but PVR is not typically noted. Subretinal bands often stretch after repair and
allow retinal reattachment; therefore, they typically do not need to be
removed or cut (52). Overall, anatomic and visual outcome for detachments
from traumatic and idiopathic juvenile retinal dialysis are typically quite good
absent macular involvement (44,45,53,54).
Postoperative Care
In pediatric patients, postoperative care is important and involves compliance
with the regiment of postoperative medications, maintaining the correct
position, and visual rehabilitation. Deprivational amblyopia often limits
visual potential in this age population and can be the biggest battle.
Postoperatively, it is important to aggressively treat amblyopia with
correction of refractive errors in conjunction with pediatric ophthalmologists
to aid in maximal visual potential recovery. In children with poor visual
prognosis despite treatment and surgery, ophthalmologists should be aware of
the community resources that are available for the parents to assist with their
child’s integration into society. Useful resources include parent support
groups, schools for children with special needs, national societies for the
visually handicapped for access to visual aids, and learning of life skills, for
example, reading and writing braille.
For monocular pediatric patients or patients having a high risk of retinal
detachments, children and parents must be counseled about full-time
protective eyewear including with child-friendly frames and polycarbonate
lenses. Counseling should also address risks of injury in contact or
noncontact sports and the importance of protective eyewear such as sports
goggles to avoid accidental trauma to either eye.
COMPLICATIONS OF PRRD SURGICAL

REPAIR
Scleral Buckling
Scleral buckling is the preferred method to repair PRRD when possible.
However, any procedure comes with potential intraoperative and
postoperative risks. Intraoperatively, recti muscle injury or transection,
scleral perforation during suture passes or belt loop creation, and subretinal
hemorrhage during external subretinal fluid drainage can be encountered.
Postoperatively, recurrent detachment due to unsupported or missed breaks,
inadequate or misplaced buckles, and inadequate retinopexy requires further
surgical interventions. Additionally, choroidal congestion and choroidal
effusions can be noted with high scleral buckles in the immediate
postoperative period and typically resolve spontaneously. Ocular motility
restrictions and diplopia are known complications of buckling procedures and
often improve spontaneously as postoperative periocular tissue swelling
subsides. Residual diplopia requires referral for strabismus evaluation and
treatment. Late buckling complications include scleral band exposure, which
require buckle removal to avoid or treat periocular infection.
Vitrectomy
PPV for PRRD is often performed as a primary procedure in PRRD with
vitreous hemorrhage and GRT and is combined with scleral buckle in PRRD
with substantial vitreous tractional components. Any intraocular procedure,
particularly PPV with use of tamponade agents, has increased risk of cataract
formation and elevated intraocular pressure requiring additional treatments
with medication or surgery. Glaucoma is also increased with the use of
postoperative topical steroids in children. Silicone oil tamponade presents
with the risk of silicone oil emulsification and oil movement into the
periocular or retrobulbar tissues.
OUTCOMES AND WHEN SHOULD

ADDITIONAL PROCEDURES BE
CONSIDERED
Single-procedure anatomical success rates range from approximately 50% to
90% (48) often due to late presentation. PRRD repair often requires complex
and multiple surgical procedures and techniques including step-wise
approaches, such as scleral buckling followed by later vitrectomy and
tamponade surgery if buckling alone was not successful. Silicone oil
tamponade is often recommended for advanced cases especially as pediatric
patients are unable to position. However, even with silicone oil, positioning is
important. Waiting about 3 to 6 months in between repeat vitrectomy
procedures is not unusual especially with peripheral detachments, because
that period allows for proliferative vitreoretinopathy membrane formation to
mature, inflammation to subside, and for subsequent more successful
membrane removal and tamponade. Anatomic success of PRRD surgical
repair can approach 75% to 80%; however, final VA >20/200 is less likely
achieved due to late presentation and need for multiple surgical interventions.
It is critical that the patient and the patient’s family are counseled on this
range of potential surgical procedures needed and outcomes. They should
also be counseled on the need for close follow-up, monitoring, and safety
precautions for this and the fellow eye. This is important as the young child
grows older and is exposed to sports activities or other potential traumas.
CONCLUSIONS
The incidence of PRRDs is low and is often associated with inherited
conditions or trauma. Bilateral vision threatening pathology is common, and
thorough examination and prophylactic treatment of the fellow eye are
important. Counseling of patient and parents is critical. Repair of PRRDs
with current advanced vitreoretinal surgical techniques is successful, but
multiple surgical interventions are often indicated.
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64
Coats Disease
J. Michael Jumper
INTRODUCTION
Coats disease is a form of idiopathic retinal telangiectasia most often
occurring unilaterally in male children and young adults. In 1908, George
Coats described three subtypes of retinal disease characterized by massive
retinal exudation and varying degrees of vascular abnormalities (1). He
subsequently revised his classification after concluding that two of the three
subtypes were manifestations of a single clinical condition (the third was
recognized as a separate entity previously described by von Hippel) (2). In
1956, Reece further refined the description and definition to what we now
consider to be Coats disease (3). Recent literature has sought to clarify the
definition, specifying that the term “Coats disease” should refer only to
idiopathic congenital retinal telangiectasia with intraretinal or subretinal
exudation and without appreciable vitreoretinal traction (4). This strict
definition has been applied to distinguish true Coats disease from the myriad
of conditions that result in either retinal telangiectasia or retinal exudation.
Since the original description more than a century ago, much has been
learned regarding the natural history and reasons for vision loss in Coats
disease. We are now better able to preserve vision and prevent the globe-
threatening complications of this disease. Despite this, the cause of Coats
disease remains elusive.
EPIDEMIOLOGY (PREVALENCE,
ENVIRONMENTAL FACTORS,
WORLDWIDE IMPACT)
Coats disease is rare, and prevalence data are lacking. A population-based
study in the United Kingdom estimated an incidence of 0.09 per 100,000 (5).
Coats disease can present at any age and has been diagnosed at 1 month and
79 years of age (6–8). In a tertiary care case series, the median age of
presentation is 5 years with the majority of patients presenting before age 10
(6). The age of presentation in the population-based study was slightly older
at 8 years (5). In all series, the earlier age of presentation is associated with
increased disease severity and accelerated progression (5,6,8). In a series of
advanced Coats disease reported by Haik, 40% presented before 2 years of
age (8). There is no known racial or ethnic predilection and the demographics
and outcomes are similar in all populations studied (9).
The majority (~95%) of Coats cases is clinically unilateral, and the
presence of bilateral disease should raise suspicion of another cause of
exudation. In the cases of bilateral Coats disease, the findings are often
asymmetrical and asynchronous (6,8,10–13). For unknown reasons, 67% to
90% of patients Coats disease are male (6,9). No environmental factors have
been shown to cause or influence the course of Coats disease. Because Coats
disease is rare and clinically unilateral, the worldwide health impact is
minimal.
GENETIC ASSOCIATION
The molecular mechanism underlying the vascular changes in Coats disease
is currently unknown. A genetic cause has been proposed by several
investigators (14,15). Cytogenetic studies of individual cases of Coats disease
have revealed a partial deletion of chromosome 13 in 1 patient and pericentric
inversion of chromosome 3 in another (16,17). Due to the sporadic nature of
Coats disease, linkage analyses have not been possible. A somatic mutation
in the genes encoding proteins involved in vasculogenesis was proposed by
Black and coworkers who reported a mother with Coats disease whose male
offspring had Norrie disease. Both had a missense mutation in the Norrie
Disease (NDP) gene (18). An NDP mutation was then found in one of nine
tissue samples of eyes enucleated with Coats disease (18).
Other studies were unable to find a link between Coats disease and
mutations in the FZD4 gene that is associated with familial exudative
vitreoretinopathy (19). Genes associated with retinitis pigmentosa and a
Coats-like phenotype have been studied including the crumbs homologue 1
(CRB1) gene but no causative association has been made (20–25).
PATHOLOGY/PATHOPHYSIOLOGY
Coats disease is the second most common disease mistaken for
retinoblastoma and is found in 7% of enucleation specimens where the
clinical diagnosis was retinoblastoma. It is important to understand the
distinguishing pathologic findings of Coats disease in order to differentiate it
from retinoblastoma (26). Coats’ description of enucleated eyes demonstrated
the characteristic features of the disease: vessel thickening and hyalinization
alongside endothelial cell thinning and loss; subretinal exudate with
prominent cholesterol crystals and a mononuclear infiltrate (1,27,28).
Electron microscopy confirms thickening of the retinal capillary adventitia
with loss of pericytes and endothelial cells (29,30).
It is believed that the endothelial changes lead to breakdown of the
blood–retinal barrier, fluid movement into the vessel walls that leads to
dilation, telangiectasia, and leakage into and under the retina. It is at this
point when the characteristic deposits of lipid, hemorrhage, fibrin, and
inflammatory cells are seen. The inner retinal layers often contain lipid-laden
“ghost cells” that likely represent transformed retinal pigment epithelial cells
(31,32).
Detailed biochemical analysis of the subretinal fluid in Coats disease
suggests that the elevated cholesterol levels are due to oversaturated and not
dysfunctional retinal pigment epithelial cells (33). Little is known about the
fibrotic nodule that forms in the macula and is a common cause of vision loss
in Coats disease. Jonas and Holbach reported a clinicopathologic correlation
in a patient who underwent surgical removal of “subretinal tissue
proliferation,” at the time of traction retinal detachment repair 8 years after
the macular fibrosis was first identified (34). The cellular mass contained
lipid/hemosiderin-laden macrophages as well as retinal pigment epithelial
cells with fibrous metaplasia. No vascular elements were described. This does
not exclude the possibility that neovascularization was once present.
Pathologic analysis of disciform scar in age-related macular degeneration
failed to identify vascular elements in 25% of cases, presumably because the
vessels were obliterated during the scarring process (35). Ong et al.
demonstrated histopathology of capillaries and retinal vessel ingrowth into
select cases of Coats fibrotic nodules (36). In a histopathologic analysis of 62
eyes with Coats disease, Chang et al. described the accumulation of lipid
exudate in all layers of the retina as well as in the subretinal space. Pigment-
laden macrophages were also present in the subretinal exudate. Subretinal
pigment neovascularization was observed in four eyes in this study (28).
Calcification is uncommon and can be used to distinguish Coats disease
from retinoblastoma except rarely in late, advanced cases of Coats where
osseous metaplasia can occur (37).
CLINICAL FEATURES AND

SYMPTOMS AND SIGNS
The most common presenting signs of Coats disease are decreased visual
acuity, strabismus, and leukocoria. The visual acuity is highly variable, but
the majority of cases have a presenting visual acuity of 20/200 or worse, and
the presenting visual acuity appears to correlate strongly with long-term
prognosis (6,38). Examination rarely reveals anterior segment findings,
although severe cases may show corneal edema, anterior chamber
shallowing, cholesterolosis, or neovascular glaucoma (6).
Posterior segment findings range from mild peripheral telangiectasia of
the retinal vasculature to florid exudation with total retinal detachment
(Figure 64-1). The telangiectatic vessels are rarely located in the macula, and
more than half the telangiectasias are found anterior to the equator (6). For
unclear reasons, the telangiectatic vessels appear to have a predilection for
the temporal and inferior quadrants (6). The telangiectasias may be
accompanied by grape-like clusters of microaneurysms, fusiform dilation of
retinal arterioles (“light bulb aneurysms”), sheathing of retinal vessels, or
venous beading. While the peripheral retinal changes are largely unilateral,
widefield angiography may demonstrate peripheral retinal changes in the
unaffected contralateral eye in some cases of Coats (39). A multicenter,
retrospective study of 350 eyes of 175 Coats patients found widefield
angiographic evidence of peripheral vascular abnormalities in 18.8% of
fellow eyes; none of which progressed over time (40).
FIGURE 64-1 The spectrum of Coats disease ranges from
mild peripheral retinal telangiectasia (A, B) to more
pronounced vascular changes (C) that can lead to total
exudative retinal detachment (D).
Despite the peripheral location of the telangiectasias, the intraretinal and

subretinal exudate often migrates toward the macula, leading to severe visual
loss despite the large distance between the primary pathology and the fovea.
The reason for this macular accumulation of exudate is not known, but it has
been hypothesized that the retinal pigment epithelium in this area pumps
more actively, acting as a sump within the eye and drawing the fluid and
exudate into the posterior pole. The exudate that accumulates in Coats disease
contains unusually high levels of cholesterol and protein, and these
components are concentrated in the macula as free water is extracted by the
retinal pigment epithelium (33). In cases of massive exudation, a dense
nodule may form in the macula, which is associated with a particularly poor
prognosis (41). The mechanism for formation of the nodule is not fully
understood. Studies of patients with macular fibrosis indicate that this
represents a form of intraretinal neovascularization at the point of lipid
accumulation (28,34,42,43) (Figure 64-2).
FIGURE 64-2 Lipid accumulation in the central macula
can lead to the formation of a fibrous nodule (A).
Angiography reveals the presence of a retinal–retinal
anastomosis (B, Arrow) with intraretinal
neovascularization causing late leakage (C).
Other macular changes that have been described in Coats disease including
vitreomacular traction, epiretinal membrane, and macular hole are all likely
due to exudative abnormalities in the vitreoretinal interface (44–47) (Figure
64-3).
FIGURE 64-3 Vitreomacular traction causing macular
detachment as a complication of Coats disease. A: Color
fundus image and B: corresponding OCT image of
vitreomacular traction causing macular detachment as a
complication of Coats disease.
Patients typically present with varying degrees of exudative retinal

detachment, and the majority of patients will develop a significant retinal
detachment if left untreated (11). The detachment usually begins in the area
of the telangiectasia, subsequently involving the macula as a result of
exudation, and eventually progressing to total retinal detachment. Advanced
cases may develop a secondary glaucoma as a result of forward displacement
of the lens–iris diaphragm or neovascularization of the iridocorneal angle (4).
SYSTEMIC ASSOCIATIONS AND

There is no known association between true Coats disease and any systemic
abnormalities. The term “Coats-like reaction” has been applied to exudative
retinopathies that occur with systemic conditions such as
facioscapulohumeral dystrophy (48–50), Senior-Loken Syndrome (51),
Turner Syndrome (52), Alport syndrome (53), and others. Ocular conditions
that have been associated with a Coats-like reaction include retinitis
pigmentosa (25), branch retinal vein occlusion (54), and toxoplasmosis (55).
Unlike Coats disease, this type of exudation is more often bilateral when
related to systemic conditions and results from a loss of vascular integrity
that may be directly attributed to the underlying condition (4).
Advanced Coats disease must be differentiated from other causes of
nonrhegmatogenous retinal detachment including retinoblastoma, persistent
fetal vasculature, retinopathy of prematurity, familial exudative
vitreoretinopathy, retinal capillary hemangioma, retinal cavernous
hemangioma, incontinentia pigmenti, Norrie disease, and toxocariasis (8).
DIAGNOSTIC STUDIES
The diagnosis of Coats disease is primarily based on clinical characteristics
alone and examination under anesthesia (EUA) is often required in children
who are unable to tolerate a careful fundoscopic examination. Ancillary
testing is sometimes helpful in determining the severity of the disease or in
differentiating it from other causes of leukocoria and vision loss in children.
In eyes with clear media, fluorescein angiography will highlight areas of
telangiectasia and may also show microaneurysms, areas of capillary
nonperfusion, or retinal neovascularization. Areas of anomalous vasculature
will leak in the late phases of the angiogram (56). Handheld, widefield
fundus photography and angiography have become an important tool in
documenting findings, diagnosing the disease, directing treatment, and
assessing treatment response (57) (Figure 64-4). When the diagnosis is
unclear, ultrasonography and/or computed tomography (CT) can be used in
advanced cases of Coats disease to assess the eye for an intraocular mass,
extent or retinal detachment, and presence of calcifications suggestive of
retinoblastoma. Optical coherence tomography (OCT) can be used to evaluate
the extent of intraretinal and subretinal fluid, exudate, outer retinal atrophy,
or fibrosis in the macula (Figure 64-5).
FIGURE 64-4 Handheld, widefield fundus angiography
(A) done at the time of examination under anesthesia helps
to identify the vascular abnormalities and direct therapy.
The widefield photo (B) taken immediately after laser
treatment shows the treatment pattern and intensity applied
to the affected area.
FIGURE 64-5 OCT is used to identify the location of
exudate and fluid in an eye with Coats disease (A, B).
Serial OCT images (C, D) show resolution of the exudate
with time.
INDICATIONS FOR SURGERY

Because Coats disease is so rare, prospective, randomized trial data to
determine the optimal therapy are not available. Treatment options are best
tailored to disease severity. Observation may be appropriate in some patients
with stage 1 (telangiectasia without exudation) Coats disease, especially if 15
years of age or older and if follow-up can be assured (58). Treatment should
generally be applied at the first signs of exudation.
PROGNOSIS
Untreated, active Coats disease commonly progresses to total retinal
detachment and, in a third of cases, neovascular glaucoma. In one study, 20
of 25 (80%) patients with untreated advanced Coats disease went on to
phthisis, neovascular glaucoma, or both, and only 2 eyes had spontaneous
regression of the total retinal detachment (8). Only rare cases of spontaneous
regression of Coats disease have been reported (59). The younger the age of
diagnosis, the greater the risk of progression (11,60). Along with an early age
at diagnosis, other features that have been identified as risk factors for a poor
outcome include (a) more extensive telangiectasia (3 or more quadrants); (b)
the presence of retinal macrocysts; (c) diffuse exudation; and (d) macular
involvement (13,58).
The Shields and associates classification scheme for Coats disease
correlates the stages of the disease with known indicators of long-term
prognosis (58) (Table 64-1). The visual prognosis varies with the stage of the
disease (58). This system utilizes five stages, representing the spectrum from
isolated telangiectasia to end-stage neovascular glaucoma with phthisis. Stage
1 disease consists of telangiectasia only, stage 2 indicates that both
telangiectasia and exudation are present (2A, extrafoveal exudation; 2B,
foveal exudation), stage 3 indicates retinal detachment (3A, subtotal retinal
detachment; 3B, total retinal detachment), stage 4 consists of total retinal
detachment and secondary glaucoma, and stage 5 indicates end-stage disease
approaching phthisis. Poor visual outcome (20/200 or worse) is rare in stage
1 disease but occurs in half of patients with stage 2 disease and nearly 75% of
those with stage 3 disease, even when those patients receive aggressive
treatment.
TABLE 64-1 Classification for Coats disease

Adapted from Shields JA, Shields CL, Honavar SG, et al. Classification and management of Coats
disease: the 2000 Proctor Lecture. Am J Ophthalmol 2001;131(5):572–583.
Daruich et al. have proposed a modification to the Shields classification

based on their findings that a subfoveal nodule has a significant impact on
visual prognosis by adding 2B1—“without subfoveal nodule” and 2B2
—“with subfoveal nodule” (41). Macular features on OCT have been
associated with visual acuity, with either macular fibrosis or outer retinal
atrophy associated with poorer visual acuity (36). Patients with stage 4 or
stage 5 disease invariably have a poor visual prognosis (58).

TECHNIQUE AND WHEN TO
CONSIDER ANOTHER PROCEDURE
Treatment in children is most commonly performed at the time of EUA.
Options for treatment are addressed below and include ablative therapy,
pharmacotherapy, and intraocular surgery of some combination of these.
Laser ablation is often the first line of treatment. Antivascular endothelial
growth factor (VEGF) therapy may be used as an adjunct described below.
Cryopexy may have a role as an ablative therapy but has been associated with
worsening retinal detachment and its timing and application should be
carefully considered. Intraocular surgical intervention may be required for
stage 3 or greater Coats disease.
EQUIPMENT
Laser system with visible (532 nm or 577 nm) or near-infrared (810 nm)
beam delivered via the indirect ophthalmoscope.
Retinal cryopexy system with external contact probe.
Ablative Therapies
Supporting Evidence and Procedural Technique

Successful ablation of the leaking, abnormal vessels in Coats disease was
first reported using diathermy by Guyton and McGovern in 1943 (61). Xenon
arc photocoagulation was later used but found to be difficult to focus,
required long exposure times, and was often painful (62). This was replaced
by laser photocoagulation, which was advocated by Pesch and Meyer-
Schwickerath (63) and remains a mainstay of therapy. Laser treatment is
directed to the telangiectatic vessels, microaneurysms, and ischemic retina.
Frequency-doubled Nd:YAG (532 nm), dye yellow (577 nm), and diode (810
nm) lasers are now most commonly employed in treating Coats disease.
Green and yellow wavelengths are adequately absorbed by intravascular
chromophores and can be used to treat abnormal vessels in detached retina
(64,65). The slightly longer yellow wavelength has a theoretical advantage of
reduced scatter by ocular structures and deeper penetration requiring less
power (66–68). Scatter laser photocoagulation to the area of telangiectasia
with direct treatment to the abnormal vessels and adjacent retina is preferred
if the retina is attached. Long, intense burns directed at the vascular
abnormalities will often cause a color change to the vessel. Repeat treatment
later in the same treatment session may be useful to adequately treat a large
vascular abnormality. Laser to the surrounding retina should be with
moderately intense burns. In detached retina, treatment with long, intense
laser can often be directed to the vascular abnormalities. This can eventually
lead to reattachment that can allow for further laser in another session. Long-
term follow-up shows good anatomic results in select cases of Coats disease
with laser therapy alone (69). The use of intraoperative widefield
angiography to identify the full extent of the vascular abnormalities may
reduce the number of retreatments needed (70). Schefler et al. reported good
anatomic results with repetitive aggressive diode laser therapy alone. In their
study, a median of 4.8 treatment sessions performed approximately every 2
months was needed to control exudation (71).
Cryotherapy has been traditionally used for stage 3 or 4 disease in which
the peripheral telangiectasias are not accessible with laser. Cryotherapy is
directed to the telangiectatic vessels and microaneurysms in detached retina.
Studies have shown an increased risk of traction retinal detachment after
cryotherapy (72). Cryotherapy is believed to damage broad areas of RPE and
reduce the pump action important to reattach retina once telangiectasias are
closed.
Postoperative Care
Multiple treatment sessions may be needed to completely ablate the leaking
vessels and repeat treatments should generally be spaced at least 3 months
apart. This also allows adequate time for the resorption of subretinal fluid and
provides a more accurate assessment of treatment efficacy (58). Macrocysts
persist in 10% to 15% of eyes with Coats disease.
Recurrence of disease after initially complete treatment has been
described in up to 7% of Coats patients. Mean time to recurrence in one study
was 10 years highlighting the need for long-term follow-up of Coats patients
(58).
Complications
Overly aggressive cryotherapy may worsen the retinal detachment (58).
Cryotherapy along with intraocular pharmacotherapies may also worsen
detachment as described under pharmacotherapies below. Subfoveal fluid and
macular exudation have been seen to worsen after laser or cryotherapy in
some patients, and it has been suggested that this may be reduced or
prevented by placing several rows of barrier laser posterior to the area of
telangiectatic vessels in a region of attached retina. This barrier is intended to
prevent posterior migration of fluid after subsequent treatment of the
telangiectasias themselves (38).
Intraocular Pharmacotherapies
Equipment
Pharmacotherapeutic: triamcinolone, pegaptanib, bevacizumab, ranibizumab,
or conbercept; syringe with small gauge needle; prep solution such as
betadine; sterile lid speculum; calipers.

The rationale to use intraocular drug therapy is based, in part, on the
discovery that angiogenic and inflammatory bioactive factors are found in
higher levels in the aqueous humor and subretinal space in the affected eye of
Coats patients including VEGF, angiogenin, nitric oxide, monocyte
chemoattractant protein 1 (MCP-1), IL-6, and IL-1b (73–77).
Intravitreal triamcinolone (IVT) has been shown to reduce vascular
leakage and proliferation in conditions including pseudophakic cystoid
macular edema, diabetic macular edema, and proliferative diabetic
retinopathy (78,79). Reports on the use of IVT alone in Coats disease have
been mixed. Jarin and coworkers reported an adult with Coats unresponsive
to conventional laser, who responded to a 25-mg IVT injection with
reduction in macular edema (80). Jonas treated 2 adult patients with the same
dose of IVT with no effect (81). IVT has also been used in combination with
ablative therapy, although worsening detachment may occur (see the
Complications Section below).
Intraocular VEGF levels in Coats patients are significantly higher than
controls and increase with stage of the disease (73). VEGF inhibition has
been studied as single therapy (82,83) and as an adjunct to steroid (84) or
ablative therapy (85,86). Case reports and small, nonrandomized series
indicate an effect of various anti-VEGF agents including pegaptanib (87),
bevacizumab, ranibizumab, and conbercept (88–90).
These studies suggest that anti-VEGF therapy can, at least temporarily,
reduce or eliminate exudation and, in turn, improve vision.
Complications
Bergstrom and Hubbard performed cryotherapy 1 to 4 months after a 1-mg
IVT injection in 5 patients with total bullous retinal detachment due to Coats
disease. Cryotherapy was performed over 2 to 3 sessions. Several
complications occurred including elevated intraocular pressure in 4 of 5
patients, severe cataract in 3 of 5 patients, and inoperable rhegmatogenous
retinal detachment with proliferative vitreoretinopathy in 3 of 5 patients.
Time from IVT injection to developing retinal detachment ranged from 4 to
24 months. The authors advised avoiding the combination of IVT and
cryotherapy in pediatric patients with Coats disease (91).
As with IVT, cases of subretinal bands, vitreoretinal fibrosis, and traction
retinal detachment have been encountered when anti-VEGF injection therapy
was combined with ablative therapy, especially cryotherapy (84,92).
Intraocular Surgery
Equipment
For drainage of Coats detachment fluid, the equipment includes intraocular
infusion fluid and delivery needle/cannula, scleral cut down blade and/or
external drainage needle, and laser and/or cryopexy device. For more
complex vitreoretinal procedures for recalcitrant detachment, this may also
include vitrectomy system, endolaser, buckle, long-acting gas, or silicone oil.

Patients with stage 3 or greater Coats disease may require intraocular surgery
to reattach the retina and allow ablative therapy. In their large consecutive
series of Coats patients, Shields and coworkers reported that retinal
detachment repair (including ablative laser or cryotherapy combined with
either subretinal fluid drainage alone or in combination with vitrectomy) was
required in 52 of 124 (17%) eyes. Another 14 eyes (11%) required
enucleation (58). Even with advanced disease and a poor visual prognosis,
surgery may reduce the risk of neovascular glaucoma and phthisis. In a case
series of 13 eyes with total retinal detachment from Coats disease, 4 of the 6
untreated eyes went on to neovascular glaucoma or phthisis. Neovascular
glaucoma did not develop in any of the 7 eyes treated with intraocular
infusion, subretinal fluid drainage, and cryotherapy or laser treatment (93).
More recent studies confirm these findings (94).
Many different techniques have been described to surgically reattach the
retina in Coats disease; these include some variation of infusion via the
limbus or pars plana followed by subretinal fluid drainage and an ablative
procedure to the reattached retina. Drainage techniques are multiple but often
include scleral cutdown with choroidal penetration or 27-gauge needle
penetration without cutdown (95,96).
More invasive therapy for recalcitrant detachment in Coats disease has
been described including endolaser via a two-port, pars plana, nonvitrectomy
approach (97), as well as vitrectomy with or without scleral buckling,
lensectomy, endodrainage, gas tamponade, or silicone oil (98–100). Based on
their long-term experience in treating patients with stage 3B Coats disease, Li
and coauthors concluded that early intervention may prevent neovascular
glaucoma/phthisis and reduce the risk of vitreoretinal fibrosis (100).
In a pooled analysis of treatments for Coats disease, other, rare
indications for surgery were identified including epiretinal membrane in 2%,
macular hole in 0.17%, and presenting traction retinal detachment in 0.44%
(71). Epiretinal membrane and macular hole have been described in both
children and adults with Coats disease and may form as a result of the disease
process or secondary to treatment (44–47,101). The surgical techniques
described are similar to conventional surgery with addition of laser ablation
of the peripheral telangiectasia.
Complications
Although no large study has specifically addressed complications of
intraocular surgery for Coats disease, the complications described are similar
to those for other forms of pediatric vitreoretinal surgery. With both drainage-
only and vitrectomy procedures, intraocular hemorrhage, lens damage, and
retinal breaks can occur. Care must be taken when placing intraocular
infusion and surgical instruments into the eye to avoid contact with the lens.
The surgeon should be familiar with the anatomical difference with age in the
pediatric population and plan instrument placement accordingly (see also
Chapter 61). Because phthisis and neovascular glaucoma are common
outcomes of advanced Coats disease, it is difficult to know if this is a surgical
complication when it occurs postoperatively. As with any intraocular surgery,
infection is possible.
The primary factors determining final visual outcome in patients with Coats
disease are the presenting visual acuity, extent of retinal detachment, and the
extent of macular exudation. As previously mentioned, cases with florid
macular exudation may develop subfoveal fibrosis, which is almost
invariably associated with a poor visual prognosis (4). Stage 1 disease rarely
results in severe visual loss, but stage 2 disease will progress to a final acuity
of 20/200 or worse in half of patients, even with aggressive treatment.
Treatment in stage 3 and 4 disease is primarily aimed at anatomic correction
of the retinal detachment, lowering of intraocular pressure, and prevention of
phthisis. Stabilization or improvement of visual acuity is generally seen in
patients with fewer than 4 clock hours of involved retina (38). It has been
noted that visual outcomes are particularly poor in Coats patients referred
with an initial diagnosis of retinoblastoma, presumably due to the extent of
retinal detachment (10). Long-term follow-up of patients with Coats disease
has shown that the condition can recur after long periods of quiescence, and it
should be emphasized that periodic examination is important even in patients
who have been stable for decades (102).
SUMMARY
More than a century after it was originally described, Coats disease remains a
cause of severe vision loss in the young. Slit-lamp examination and indirect
ophthalmoscopy alone can often differentiate this disease from other causes
of retinal detachment. When necessary, ultrasonography, fluorescein
angiography, and CT can assist in making the diagnosis and ruling out other
disorders such as retinoblastoma.
We now enjoy greater diagnostic accuracy and are able to salvage more
eyes with Coats disease, but the rates of central vision loss remain high due to
complications such as retinal detachment, macular fibrosis, and amblyopia.
Future research in this disease should focus on earlier detection and methods
to maintain macular function.
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65
Persistent Fetal Vasculature Syndrome
Supalert Prakhunhungsit, and Audina M. Berrocal
INTRODUCTION
Persistent fetal vasculature syndrome (PFVS), previously known as persistent
hyperplastic primary vitreous (PHPV), is a spectrum of conditions that are
characterized by failure of involution of the primary hyaloidal vasculature
system, incomplete ocular neurovascular development, or both (1,2). The
term PHPV does not optimally describe the pathologic findings in this
condition, because it fails to acknowledge complete intraocular vasculature
involvement found in this disorder. Therefore, PFVS is more widely used and
accepted as the term to describe this spectrum of diseases.
GENETICS
PFVS is most commonly sporadic. However, there are reports of inheritable
transmission in some families. An autosomal dominant pattern in an Egyptian
family was reported (3). Many other studies demonstrated cases with
autosomal recessive pattern and, normally, they are associated with other
ocular abnormalities (4–8). Furthermore, the ATOH7 gene mutation has been
detected in PFVS cases with autosomal recessive inheritance mapped on
10q21.3 location (9–12). Apart from those two genes, a novel mutation found
in the COX15 gene, a cytochrome c oxidase assembly protein on
chromosome 10, has also been reported in a case with bilateral PFVS (13).
Patients with bilateral PFVS may benefit from a referral to a geneticist for
evaluation of possible syndromic associations and their potential implications
on future offspring (14).
PATHOPHYSIOLOGY
From an embryologic perspective, the intraocular vasculature starts to form
as early as the 3rd week after fertilization. The hyaloid artery enters and
transverses anteriorly through the vitreous compartment branching into
multiple channels making up the vasa hyaloidal propria (1,2,14). The
posterior tunica vasculosa lentis is then formed at the posterior surface of the
developing lens anastomosing with iridohyaloid or capsulopupillary vessels.
As the secondary vitreous forms at the beginning of second trimester, the
involution of vessels starts posteriorly, first in the hyaloid artery, then in the
vasa hyaloidal propria and iridohyaloid vessels, and finally in the tunica
vasculosa lentis. The regression of the physical barrier caused by the
iridohyaloid vessels allows the zonular ligament and lens to develop normally
(1). The failure of normal developmental regression causes the varied degrees
of PFVS phenotypes.

Clinically, PFVS is approximately 90% unilateral and has a variable degree
of clinical findings depending on the degree of defects in the involution of the
vasculature during embryogenesis. The incompletely regressed hyaloidal
stalk is integrated into the posterior lens surface and forms a retrolental
membrane that can lead to a cataract and posterior lenticonus (Figure 65-1).
The ciliary processes can be centrally dragged and detached resulting in
ocular hypotony (Figure 65-2). Moreover, the traction from the retrolental
membrane can also exert a radial force on peripheral retina and lead to the
tractional retinal detachment and, sometimes, a rhegmatogenous retinal
detachment (14). The tractional force can also cause anomalies in the
posterior segment of the growing eye that include vitreous hemorrhage and,
occasionally, retinal dysplasia and retinal detachment. Microcornea,
microphthalmos, and swollen lens, causing shallowing of the anterior
chamber secondary to the anterior movement of the lens–iris diaphragm, can
also be observed (2,15,16).
FIGURE 65-1 Combined type PFVS. The persistent stalk
extending from the optic nerve head to the posterior lens
capsule. Posterior retinal traction and fold were noted
(Left). Large vessels were visible within the connecting
stalk (Right).
FIGURE 65-2 A 3-month-old boy with elongated ciliary

body processes and central lens opacity in PFVS. The
vitreous fiber is exerting force radially to the ciliary body
also visible at the anterior hyaloid surface.
Bilateral PFVS is a rare clinical phenotype that is observed in approximately

<10% of all PFVS cases (2,14,17,18). Bilateral cases were found to be more
commonly associated with combined anterior and posterior PFVS (1,2), and
these cases have a higher risk of posterior retinal involvement (1). Recently, a
newly identified syndrome of bilateral microcornea, posterior
megalolenticonus, persistent fetal vasculature, and chorioretinal coloboma
(MPPC) was reported (19–22) (Figure 65-3).
FIGURE 65-3 A 5-year-old boy with chorioretinal

coloboma and PFVS. The persistent stalk extended from
the coloboma to the posterior surface of the lens.
Typically, the characteristic findings of PFVS only occur in the patient’s eyes
as described earlier. When the systemic abnormalities are discovered, the
associated syndromes should be aware of. Trisomy 13 (Patau syndrome)
should be suspected in children with congenital brain, heart, or spinal cord
abnormalities (23). Muscle weakness and wasting starting very early in their
childhood are seen in PFVS patients with Walker-Warburg syndrome
(24–27). Pediatric patients with intellectual or learning disabilities are found
in Dandy-Walker syndrome with cerebellar maldevelopment resulting in
hydrocephalus and in Angelman syndrome with laughter, hand-flapping
movements, and microcephaly (28). Some congenital infections, such as
congenital CMV (27) and congenital rubella (14), are also found associated
with PFVS in the eyes. Glucose 6-phosphate dehydrogenase (G6PD)
deficiency can be found in PFVS patients with a history of hemolytic anemia.
Other reported medical conditions associated with PFVS include pulmonary
stenosis, cerebral palsy, and facial clefting syndrome (14). Maternal cocaine
use during pregnancy has also been reported to be associated with bilateral
PFVS (27,29).
TYPES OF PFVS AND

NOMENCLATURE
PFVS has many different phenotypes. PFVS cases are normally classified
into three separate groups according to the pathologic location. The clinical
findings of an anterior PFVS include a shallow anterior chamber, engorged
iris vessels, cataract, retrolental fibrovascular membrane, and/or elongated
ciliary processes (30). A posterior PFVS is characterized by the presence of
one or more of the following clinical features: elevated vitreous membrane or
stalk from the optic nerve, preretinal membranes, retinal fold or retinal
dysplasia, retinal detachment, or optic nerve hypoplasia. If the features of
both an anterior and a posterior PFVS are observed, the PFVS is classified as
a combined PFVS (2,18,27). Microphthalmia can be present in both anterior
and posterior PFVS (30,31). However, one study reported that
microphthalmic eyes demonstrate more severe posterior disease (27).
DIAGNOSTIC STUDIES
Fundus Photography
Preoperative or intraoperative fundus photography is recommended for
documentation of these cases. These photographs can also be used to
demonstrate the ocular abnormalities to parents and to discuss treatment
options and further plans for the patients. The anterior chamber angles can be
imaged with a digital camera. Often, the angle of the major vessels of the
retinal arcades can be evaluated preoperatively and compared to subsequent
images. This can document change and may indicate that posterior traction is
pushing the detachment anteriorly and starting to occlude the angle. This may
preempt the development and management of angle-closure glaucoma.
Ultrasonography
Ultrasonography is a useful investigative tool to evaluate the posterior
compartment of the eye, especially when a cataract or media opacity obscures
the posterior segment. The efficacy of ultrasonography for evaluating cataract
cases with either anterior or posterior PFVS was studied. The result was a
high true positive rate, up to 92%, which supports the important role of
ultrasonography for preoperative evaluation in PFVS cases (32). A unilateral
cataract in a newborn infant should trigger potential PFVS, and
ultrasonography should be performed. However, ultrasonography can be very
challenging in pediatric eyes. In some cases, the thin stalk that connects the
optic nerve head to the posterior surface of the lens capsule can easily be
overlooked and be later discovered intraoperatively (33). In such cases, the
cataract surgery alone may not be sufficient for clearing the visual axis, and a
vitrectomy might be needed (33). In cases of complex cataracts, a pediatric
retina specialist is often the person to perform surgery and the pediatric
ophthalmologist the one to manage the patient’s visual rehabilitation.
Ultrasonography reveals details concerning axial length and retinal
involvement that relate to the postoperative visual prognosis in the long term.
When eyes are smaller than 15 mm or when there is a disparity between the
two eyes that is larger than 3.5 mm, best vision outcome is rarely better than
light perception in the shorter eye even with surgical intervention (14).
In addition to ultrasonography to diagnose PFVS, it is also valuable to
distinguish PFVS from other mimicking diseases. Most importantly, the
detection of calcification within intraocular masses or tumors in young
children should heighten suspicion of the diagnosis of retinoblastoma (34,35).
Therefore, a more detailed investigation is needed before considering
surgery.
Widefield Photography With Intravenous Fluorescein

Angiography
Fluorescein angiography (FA) enhances the presence of persistent
vasculature in PFVS patients along Cloquet canal, which may be difficult to
visualize ophthalmoscopically. FA focusing on the posterior portion of the
lens or on the retrolental tissue can enhance radially oriented vasculature in a
brittle star (Figure 65-4). More posterior focus toward the optic nerve head
may demonstrate the hyaloid artery and its branches when present. In
complicated cases, FA can also be used (36–38). Recently, FA was used to
evaluate full-term patients who were diagnosed with unilateral PFVS. FA
showed detail of the peripheral retinal avascularity presenting in both the
affected and unaffected eyes (39) (Figure 65-5). This information could be
useful for following patients in the long term and for better understanding
this disease in the future. It is also important to assess the presence of
avascular retina in the fellow eye, which may portend the diagnosis of
familial exudative vitreoretinopathy (FEVR) even in the presence of
microphthalmia and retinal detachment in the obviously affected eye (40).
FIGURE 65-4 Left: A 3-month-old boy noted to have a
thin whitish retrolental membrane with eccentric lens
opacity. The ciliary body was also elongated.
Right:Fluorescein angiography showed the presence of
the persistent hyaloidal vessels connecting to the posterior
lens capsule in a brittle star pattern. Left:Interestingly, the
vessel was hardly seen in the color photograph.
FIGURE 65-5 Left: Fluorescein angiography showed
peripheral avascular retina seen in an eye with combined
PFVS. Right: Abnormal foveal circulation was also noted
on the macula.
From a surgical perspective, FA can show the extent of the vasculature

extending into the vitreous cavity, and this can provide intraoperative
guidance during vitrectomy for the pediatric retina surgeon. Furthermore, the
foveal circulation detected by FA is a good predictor for postoperative foveal
hypoplasia and poor central vision (41) (Figure 65-5).
Optical Coherence Tomography

Macular imaging could yield clinical value. A previous optical coherence
tomography (OCT) study of posterior type PFVS demonstrated
microstructural abnormalities that included posterior hyaloidal organization,
vitreoretinal traction, and disruption of ellipsoid zone. All of these features
were correlated with poor visual outcome in posterior PFVS (42). Currently,
with the aid of intraoperative OCT (Resight, Carl Zeiss Meditec,
Oberkochen, Germany), the anatomical status of the fovea can be assessed at
the time of the vitrectomy and can be useful in evaluating outcomes from
visual rehabilitation and patching. Moreover, macular structure can provide
clues into the pathophysiology of the disease and facilitate management
discussions with patients and families.
Other Imaging Modalities

Visual evoked potential (VEP) is a noninvasive and objective approach to
assess visual functions in infants and young children, which can be
challenging in a newborn. The visual potential of the diseased eye can be
evaluated by VEP and be used to make a decision whether surgery is feasible
for the affected eye (27,43). VEP was also used in follow-up to evaluate the
potential of postoperative visual improvement (43).
Magnetic resonance imaging (MRI) is helpful with suspected
intraocular tumors. Retinoblastoma is a crucial diagnosis to rule out
especially in young patients with leukocoria especially before proceeding to
surgery (44,45). CT scan is not routinely recommended due to the amount of
radiation exposure but can be valuable in some cases, in which an ocular
oncologist is best consulted (see also Chapters 44 and 45).
Color Doppler imaging uses ultrasound energy to derive characteristic
flow patterns in combined PFVS that can be used for both screening and
diagnosis. This novel technique may provide new and important information
to diagnose and potentially guide treatment of PFVS; however, the method
requires further validation (46).
When patients with PFVS present with leukocoria, it is important to rule out
other causes of leukocoria, including retinoblastoma, retinopathy of
prematurity, FEVR, Norrie disease, incontinentia pigmenti (2,34), Toxocara
endophthalmitis, scar tissue surrounding an occult intraocular foreign body,
X-linked retinoschisis, and retinal detachments from other causes (2). In
patients younger than 3 years of age, retinoblastoma must be considered until
proven otherwise. Unlike PFVS, microphthalmos is rarely seen in
retinoblastoma unless the affected eye(s) is/are in late stage and phthisis bulbi
occurs. PFVS is generally unilateral, whereas FEVR, Norrie disease, and
incontinentia pigmenti are normally bilateral disorders. However, FEVR can
be asymmetric and an eye with retinal detachment can be smaller than the
fellow eye. However, a clue is that conditions are not PFVS is that they do
not show a connecting trunk and rarely affect the inferonasal aspect of the
posterior lens capsule as PFVS does, even though retinal dragging and
folding can occur.
In cases of bilateral PFVS, Norrie disease or FEVR must be ruled out
(47,48) (see also Chapters 35 and 66). Bilateral cases of PFVS might be
difficult to differentiate from Norrie disease since there are no
pathognomonic features to differentiate one from the other. Patients with
bilateral FEVR rarely present with bilateral retinal detachments. The family
history of inheritance pattern, systemic associations, and especially genetic
testing can be helpful in these cases (48).
Complete history taking, ocular examination, and systemic review as well
as investigations, such as ultrasonography, MRI, and FA, are helpful in
distinguishing other diagnoses from PFVS.
MANAGEMENT
PFVS management is based on the pathologic extent of the anterior or
posterior segment, and this should be assessed (49). Management of PFVS
includes the following objectives: (a) to save vision by eliminating the
amblyogenic media opacities or, in posterior or combined PFVS, by releasing
the retinal tractional forces from multiple directions that cause retinal
abnormalities; (b) to preserve globes and to prevent complications, such as
secondary glaucoma and eventual phthisis bulbi (50,51); and (c) to improve
patient cosmetic appearance in advanced cases (14,18,52). The heterogeneity
of clinical presentations of PFVS makes it especially challenging to manage
surgically. Prior to surgery, the postoperative lifelong risks of retinal
detachment, glaucoma, and other associated complications should be
disclosed and made clear to the parents.
SURGICAL INDICATION
The management of PFVS varies by PFVS phenotype. Indications for
surgical management in PFVS have changed in recent years due to
improvements in knowledge about the disease, surgical techniques, and
surgical instrumentation. In unilateral PFVS cases in which the fellow eye is
normal, the value of surgery continues to be debated since the visual results
are often poor (27). Generally, surgical indications include media opacity,
vitreoretinal traction, retinal detachment (14), progressive shallowing of the
anterior chamber from swollen cataractous lenses, traction on ciliary
processes, recurrent severe vitreous hemorrhage, and uncontrolled intraocular
pressure from secondary glaucoma (2,50).
Surgery is usually not indicated in cases with a clear visual axis,
nonprogressive anatomical pathologies, and an unaffected anterior chamber
(2,53). Similarly, severe microphthalmia (54,55), severe retinal or optic nerve
hypoplasia, and severe associated ocular anomalies or phthisis resulting in
dense afferent pupillary defects or no light perception may be considered as
contraindications for surgery by some surgeons in patients with unilateral
PFVS (14,27). These factors in patients with bilateral PFVS are also
considered to be relative contraindications for surgery since the visual
potential is often poor (14).
TIMING OF SURGERY
If surgical management is considered, the ideal timing should be as early as
possible before the development of deprivational amblyopia or secondary
complications (14,27,56). The delay in surgery in indicated cases could lead
to biologic consequences that develop within weeks to months, such as
retinal detachment and secondary glaucoma. The critical period of visual
development plays an important role in the optimal timing of surgery, and it
facilitates early postoperative visual rehabilitation. Many studies reported the
benefit of early surgery relative to postoperative visual improvement
(27,52,56). Even in bilateral cases, early surgical intervention was found to
increase the potential of achieving functional vision (48,56). In addition to
the visual aspect, Federman et al. reported that early surgical intervention
facilitated better cosmesis and long-term eye growth compared to
nonoperated eyes (57). Conversely, a deferral in surgical intervention could
lead to poor final visual outcome despite adequate anatomical success (49).
From a histologic study of the retrolental fibrovascular stalk—irreversible
retinal folds, macular dragging, and tractional retinal detachment caused by
anteroposterior traction could occur if the glial and vascular components are
left to mature in the eye (56,58). A significant retinal reattachment with
reversal of retinal folds and macular dragging was reported in a patient that
underwent surgery prior to the age of 13 months (56).

TECHNIQUE
Prior to starting the surgical intervention, an examination under anesthesia is
beneficial to comprehensively assess the eyes and obtain additional
information. The treatment for an individual patient can be simultaneously
planned. Ocular imaging, FA, and ultrasonography (both A-scan and B-scan)
can be repeated for more accurate information and documentation. These
investigations can clarify the preoperative anatomy of individual patients so
that the safest surgical approach can be planned, since the creation of
intraoperative retinal breaks can lead to a considerably worse prognosis with
an unacceptably high rate of recurrent retinal detachment (14,59). For
example, if the eye appears to be almost normal with no anterior traction
noted and the anterior retina is intact with pars plana or pars plicata
presented, then the posterior surgical approach is the preferred option. If,
however, it appears that the ocular anatomy has been adversely affected, then
a limbal approach is indicated.
EQUIPMENT
Trocars and cannulas with vitrectomy system for limbal approach
Pediatric (short) small gauge trocars and cannulas with vitrectomy system for
3-port posterior surgery
End-irrigating light pipe and vitrectomy system for 2-port posterior surgery
Endodiathermy
Transillumination (optional)
Vitreoretinal scissors, forceps (as needed)
Endoscope (as needed)
SURGICAL INCISIONS
Preference of surgical incision location varies among surgeons, with some
preferring the limbal approach and others opting for the pars plicata or iris
root approach. Since PFVS has dramatically variable ocular presentations and
severity among patients (1), an incision through displaced ocular structures,
such as anteriorly pulled retina or centrally dragged ciliary process, or
abnormal vasculature can cause unfavorable consequences and a worse
prognosis (14,57). Transillumination to locate the ora serrata can be
performed before performing a sclerotomy, but it is often unsuccessful (57).
LIMBAL APPROACH
Limbal incision has been advocated by some retina specialists (1,57,60,61)
since it is associated with the following advantages (Figure 65-6):
FIGURE 65-6 Limbal sclerotomies in an eye with

combined PFVS and microphthalmia.
1. Manipulation of the vitreous base is generally avoided (27);

2. Peripheral retina that is anteriorly and centrally dragged by the retrolental
membrane is untouched (27,48); and,
3. In some cases, with a narrow or nonexistent pars plana, the retina inserts
directly into the pars plicata or ciliary body (48,57). Thus, sclerotomies
through the limbus are usually harmless.
PARS PLICATA APPROACH

Some surgeons reported a preference for the pars plicata (also called
posterior) approach (56,61–63) because it usually reduces traction on the
vitreous during lens surgery, and it allows early removal of the persistent
hyaloidal stalk—both of which could minimize the rate of rhegmatogenous
retinal detachment (33,64). Moreover, the pars plicata approach leaves the
cornea and anterior chamber angle less disturbed, and it permits better
visualization during removal of lens material and the capsular bag
(33,65–69). The location of posterior incision varies from 0.5 mm through
the iris root (70) to 1 to 2 mm posterior of the limbus depending on the
patient’s age and the extent of the microphthalmia of the affected eye (56).
The age-based method of sclerotomy placement is a useful guideline for
placing sclerotomies in pediatric cases; however, these data were based on
normally developed eyes (71,72) (see Chapter 61).
The pars plicata approach is also beneficial in cases in which lens
extraction is not required, such as predominately posterior component PFVS
with nonaxial lens opacity. In these particular cases, a lens-sparing
vitrectomy using a pars plicata approach could be employed (70,73).
Sisk et al. compared these two approaches and found similar visual and
anatomic outcomes between groups and no significant difference between
groups for complications or visual prognosis (14). Perhaps now that we have
better instrumentation for pediatric vitrectomy, outcomes may improve.
THE VITRECTOMY
Anterior PFVS
When the lens opacity or Mittendorf dot is eccentric to the visual axis, it
should be typically left untouched. Rarely, they will progress to
complications such as secondary angle closure from anterior rotation or
swelling of the lens. Cases with purely anterior PFVS with significant
cataract, lensectomy with anterior vitrectomy is usually performed (16,27).
The surgery aims to get rid of the amblyogenic factor from the media opacity.
During the lensectomy, it is crucial to remain inside the capsular bag at all
times to avoid additional vitreous traction, which can cause a retinal tear.
Afterward, the remnant of the lens material and capsular bag should be
adequately eliminated. A peripheral iridotomy is recommended to prevent
shallowing of the anterior chamber and postoperative glaucoma.
Furthermore, sub-Tenon’s steroids will reduce the potential for significant
postoperative inflammation. However, intraocular pressure must be
monitored since infants are prone to respond to steroids with high intraocular
pressure.
Posterior and Combined PFVS

The surgical techniques to manage PFVS have changed considerably over the
years. Initially, Reese described a two-step approach that consisted of
needling of the lens followed by dissection of the retrolental membrane (1).
There was a variation in the two-step techniques reported, including open-sky
dissection to minimize the risk of intraocular bleeding (50,74–76). Closed
system vitrectomy is now available due to advancements in vitreoretinal
surgical techniques.
There are two types of modern vitrectomy—two-port and three-port
vitrectomy. Two-port vitrectomy requires a special instrument (i.e., irrigating
end light pipe) in one sclerotomy and a high-speed vitreous cutter in the other
(27,56,70,73). In phakic patients, contact between the trocar and the
crystalline lens can be avoided by keeping the instruments parallel to the
visual axis as they pass through the pars plicata (70,73). The anterior portion
of the fibrovascular membrane is separated before being cautiously dissected
with a vitreous cutter (56) or with the membrane-peeler-cutter scissors
(27,77) (Figure 65-7). Dissection is then performed anterior to any large
visible vessels to prevent bleeding in order to release the traction from the
ciliary body or anteriorly dragged retina. The removal of ciliary traction
makes the risk of postoperative hypotony or phthisis less likely (27,48). In
lens-sparing vitrectomy, immediate division of the connecting stalk could
reduce the movement of the stalk, and the damage to the posterior lens
capsule, and this results in less postoperative cataract progression in cases
with nonaxial lens opacity (70,73). After the stalk is divided, endodiathermy
can be applied when patent vessels are visible (27,77,78). However,
endodiathermy can also be applied before cutting the stalk.
FIGURE 65-7 Intraoperative view after lensectomy was
done showing the posterior lens capsule and radially
oriented vitreous fibers. These were cut and separated
from the retrolental membrane.
Complete vitrectomy was performed when posterior segment traction was

present. In phakic patients, the range of instrument movement should be
limited to the postequatorial area to avoid lens injury (70,73). Then, the
removal of the connecting vitreous sheet and hyalocytes around the residual
stalk is performed (56) (Figure 65-8). The posterior vitreous stalk is then cut
and endodiathermy applied. Membrane peeling is performed if tractional
epiretinal tissue or tractional retinal detachment is present (14,27). In most
cases, laser is not applied to the peripheral avascular area since there was a
low risk of neovascularization (48). In some cases where there is peripheral
view through the cataract and avascularity is noted with mild leakage by FA,
laser is recommended. Surgical peripheral iridotomy should be considered in
most cases to prevent secondary angle closure with or without glaucoma.
Then, it is important to remove any clotted blood and/or vitreous from the
peripheral iridotomy. In the end, a fluid–air (27,48,78) or fluid–viscoelastic
(48) exchange is performed before closure. Limbal or scleral sutures are
required in all cases. For bilateral PFVS eyes usually associated with a
combined anteroposterior type, the removal of the lens to approach the
retrolental membranes is recommended. Therefore, the tendency to develop
shallow anterior chamber and secondary glaucoma may be reduced (48).
FIGURE 65-8 The vitreous around the stalk was removed

to release the tractional forces extending to the stalk and
surrounding retina.
In our experience, we prefer a limbal approach, placing trocars thru the

limbus (if the size of the eye allows or simply making limbal incisions with
the needle of the trocar), filling the anterior chamber, and deepening it with
viscoelastic to stabilize it. We then remove the lens in the bag with aspiration
and open the anterior and the posterior capsule. It is important to understand
where the stalk inserts to start cutting the posterior capsule around the stalk.
Once the stalk is released, then a vitrectomy is performed. It is important to
understand the radial insertion of the vitreous and reduce all traction around
the stalk so it falls back freely. If the stalk has retina in it, it is important not
to cut it and add a rhegmatogenous component to the complex retinal
detachment. An air–fluid exchange in these eyes is recommended making
sure that the stalk does not fall over the macula. The baby should be
positioned based on the treatment done in the eye.
Eyes treated for PFVS do not benefit from intraocular lens implantation
at the time of surgery. If the eye is very similar in size to the other unaffected
eye and the macula is normal, leaving anterior capsule for future sulcus lens
implantation can be considered.
Sub-Tenon’s steroid injection and subconjunctival antibiotics may be
applied at the end of the surgery. Anti–vascular endothelial growth factor
(anti-VEGF) medication may be considered to aid in the reduction of anterior
or posterior bleeding and postoperative inflammation, but evidence is limited.
The first follow-up visit is scheduled the day after surgery. Postoperative
drops include topical antibiotic eye drops and topical steroid eye drops.
Cycloplegics after surgery are recommended to avoid posterior synechia
formation.
ADVANCEMENT IN PEDIATRIC
RETINAL SURGERY FOR PFVS
Intraoperative Optical Coherence Tomography
(Intraoperative OCT)
Intraoperative OCT enables surgeons to visualize the posterior portion of the
lens (Figure 65-9). With intraoperative OCT, it is possible to determine how
the retina and fibrovascular tissue attached to the posterior lens surface and
whether it is necessary to sacrifice the lens. There are PFVS cases in which
the lens is almost normal with only slight opacity. Intraoperative OCT can
determine whether the opacity extends only to the anterior face of the hyaloid
or if it involves the lens resulting in the need for lensectomy procedure. This
is a very important difference to the development of the postoperative vision
in these eyes. Moreover, this information could lead to appropriate surgical
procedures to treat the lens and/or retrolental fibrovascular tissue. In posterior
PFVS, intraoperative OCT can be used to evaluate the extent of the epiretinal
membrane-like structure, vitreomacular traction, and other vitreoretinopathies
that require membrane peeling. Moreover, the intraoperative OCT can be
used to evaluate the macular anatomy in terms of visual prognosis assessment
(Figure 65-10). All of this information can be derived from the real-time use
of intraoperative OCT.
FIGURE 65-9 The intraoperative OCT through the lens

opacity showing the stalk attached to the posterior lens
capsule. There was a hyperreflective thin line indicating
the anterior hyaloidal face of the hyaloid that could
potentially be separated from the posterior lens capsule.
FIGURE 65-10 The intraoperative OCT of a 5-month-old
boy with combined PFVS. The intraoperative OCT shows
the loss of physiologic foveal depression over the entire
macula.
Small-Gauge Surgery
Small-gauge surgery for pediatric cases has many advantages. Pediatric eyes
have a shallower anterior chamber, a relatively larger lens thickness, a shorter
axial length, and less vitreous volume compared to adult eyes (79).
Moreover, affected eyes with PFVS are often smaller than those from age-
matched children. Thus, the instruments used for adult cases are not
appropriate for use in pediatric surgery. Accordingly, pediatric surgical
instruments were designed and introduced to improve surgical outcomes.
Short 25-G vitrectomy designed specifically for pediatric eyes is now
commercially available, with vitrector and light pipe lengths that are
approximately two-thirds the length of their adult-sized counterparts (80).
The reduced length of the short 25G makes instrument manipulation easier in
limited spaces.
The recent development and introduction of 27-G vitrectomy may also
have a potential role in pediatric vitrectomy. Smaller sclerotomies facilitate
better self-sealing wounds, which reduces the risk of postoperative hypotony,
bleeding, and endophthalmitis (81). However, the use of more flexible
instruments and reduced flow rate may be of concern when used in pediatric
cases, especially when membranes are tough or thick (81,82).
Endoscopic Vitrectomy for PFVS

There is good evidence to support the efficacy of endoscopic vitrectomy in
many adult retinal diseases. However, the use of endoscopic vitrectomy is
often limited in pediatric retinal disorders due to the narrow space in smaller
eyes, the differences in ocular anatomy (83,84), and the nature and
complexity of pediatric retinal diseases. The unfamiliar side-on perspective of
the viewing system might influence some surgeons to refrain from using it;
however, this system is helpful in many aspects. First, it can be used as an
adjunct to wide-angle viewing systems in pediatric vitrectomies, especially
when significant media haziness is present that prohibits the use of a
conventional viewing system (84–86). Secondly, at the start of a surgical
procedure, it can directly visualize the eye for sclerotomy placement over a
safe area (87). Endoscopic vitrectomy also facilitates visualization in difficult
to access surgical spaces and visualization of anterior pathologies (86–88).
Unfortunately, we do not have 25- or 27-G endoscopes available specifically
for pediatric eyes.
In PFVS, the above view produced by a conventional noncontact viewing
system provides a perfect view of the surgical area in these media opaque
patients. However, the parallel perspective of the fibrovascular stalk makes it
difficult for surgeons to perform an optimal dissection without going too far
posterior into the persistent vasculature and dragged retina. At this point in
the procedure, the side-on view produced by the endoscope could help to
perpendicularly visualize the fibrovascular stalk, which would make it easier
to precisely identify the running vasculature to the anterior membrane (87).
Additionally, endoscopic vitrectomy could have a role in special situations,
such as microcornea, posterior megalolenticonus, PFVS, and coloboma
syndrome (86).
NONSURGICAL TREATMENTS
When a patient is not considered a good surgical candidate, conservative
treatment is an alternative. Also, the nonsurgical option is beneficial in mild
PFVS cases with the preservation of normal visual axis, so the surgical
intervention is not fully indicated (Figure 65-11). Some studies reported that
eyes not treated by surgery could survive to adulthood (89–91). Therefore,
conservative treatment may be a reasonable treatment option in selected cases
of PFVS. For example, cases with spontaneous lens resorption that allowed
the anterior chamber to remain deep enough even when contracture of the
retrolental membrane caused the lens to move forward (2). If conservative
treatment is the strategy of choice, the follow-up visit should be scheduled for
the near term to facilitate early detection of possible complications, including
progression of cataract in mild disease and any forms of glaucoma like lens-
inducing glaucoma (92).
FIGURE 65-11 The nonvisually significant stalk
disconnected itself and was floating independently in front
of the optic nerve in a 5-year-old boy. The other end of the
stalk attached to the posterior lens capsule eccentrically. In
this case, the conservative treatment was recommended.
However, the severe forms of PFVS are known to have progression of

disease. The reported rate of eventual blindness (no light perception) ranged
from 27% to 70% in unoperated eyes (14,48,52,69), whereas 30% of patients
retained at least light perception (48). It is valuable to preserve this
rudimentary level of vision, especially in bilateral patients, because it allows
patients to maintain a normal sleep–wake cycle (93,94).
POSTOPERATIVE CARE
The treatment of amblyopia, either from anisometropic or refractive
component, could be augmented by the combined efforts of pediatric
ophthalmologists and optometrists. Concerning amblyopia treatment—
refractive correction, occlusion therapy, or pharmacologic atropinization in
the better eye, and aphakic contact lens fitting in the surgical eye is advised to
achieve the best visual outcome (14). Lastly, protective polycarbonate glasses
should be prescribed to prevent accidental trauma to the remaining eye.
Patients and parents are always informed to observe for abnormal ocular
signs, and they are advised to return to the clinic as soon as possible if any of
those signs appear. In patients with severe microphthalmia that have no light
perception, a referral to an ocularist for a prosthesis to encourage normal
orbital development is recommended.
COMPLICATION AND MANAGEMENT

Although surgical intervention for PFVS yields several benefits, there are still
some potential associated risks. The most commonly observed postoperative
complications are retinal detachment and postoperative glaucoma. The retinal
tear can take place during any phase of surgery. Posterior placement of a
sclerotomy incision in a patient with a preoperative tractional retinal
detachment could result in a retinal tear that progresses to a catastrophic
retinal detachment (33,77). Moreover, the retinal break could produce a
potentially high risk of recurrent detachment due to the common nature of
proliferative vitreoretinopathy in pediatric patients (14). The avoidance of
complications can be enhanced by a complete and careful preoperative
assessment of the eyes and by identifying a safe site to perform the
sclerotomy, especially in these small eyes that frequently have a poorly
developed pars plana area (57).
Postoperative patients are also at risk of developing glaucoma in their
lifetime, with incidence rates that ranged from 11% to 36%
(14,52,56,77,78,95–97). The variation in reported incidence may be
attributable to factors that include age at surgery, coexisting ocular
anomalies, microcornea, aphakia, and the length of the follow-up period
(96,98–101). The mechanism of glaucoma in these patients could be angle
closure, neovascular, or mixed mechanism (14). Patients who had surgery at
a younger age were more likely to progress to glaucoma (98,102), with
glaucoma onset that ranged from 2.6 to 22 years after PFVS surgery (98).
Intraocular pressure could be controlled either by medications or glaucoma
surgery (56). Therefore, lifelong surveillance for the development of
postoperative glaucoma is usually recommended (96,98,100).
Other postoperative complications include intraocular hemorrhage, such
as hyphema and vitreous hemorrhage (77); progression of lens opacity
ranging from 5.1% to 32% (56,77,78,96); and strabismus that may require
subsequent surgery (77,97).
The visual outcomes after surgical management are varied due to many
factors. Eyes with chiefly anterior disease produce better visual outcome
compared to eyes with posterior or combined type (14,16,27,60,67,103). Age
at presentation was also reported to be one of the proven prognostic factors in
many studies (52,60,103). The earlier the age of presentation, the better the
visual prognosis. Karr and Scott found a visual outcome better than 20/200 in
children who presented earlier than 2.4 months versus 20/300 or worse in
patients who presented at 4.3 months (60). The visual result is mostly poor in
cases with bilaterality, glaucoma (14,103), concomitant congenital anomalies
that affect the optic nerve or macula (2), severe microphthalmia in which the
axial length is <15 mm or more than 3.5 mm disparity between eyes (14),
massive subretinal blood (48), and severe retinal dysplasia or detachment
(2,64).
Concerning the surgical intervention, the objective of early surgery is to
preserve the globes or to prevent subsequent complications, especially in
patients with posterior disease. Functional vision was only barely obtained
(50,103). However, improvements in instrumentation and surgical technique
in the current vitrectomy procedure, early diagnosis, and intervention
followed by aggressive postoperative measures, such as amblyopia treatment,
significantly improve the visual outcome (27,33,52,56,57,60,97). Sisk et al.
reported that approximately two-thirds of patients who underwent surgery
developed form vision, counting fingers, and fix and follow or better for at
least 6 months postoperatively (14). It is known that bilateral disease is more
associated with the combined type, and the visual prognosis in these cases
might be worse when the patient has two affected eyes. However, Walsh et
al. found that nearly 70% of bilateral cases that had vitrectomy in at least one
eye had at least light perception, and there were only 11% of cases that were
phthisical at the last follow-up. Therefore, surgery in bilateral cases could
restore or maintain vision and avoid a phthisical event (48).
LONG-TERM MANAGEMENT AND

VISUAL REHABILITATION
Postoperative aphakic correction and amblyopia therapy are usually
performed in patients who undergo lensectomy. Visual rehabilitation in these
patients will be guided by pediatric ophthalmologists and optometrists. Every
child with unilateral PFVS should undergo a trial of occlusion therapy after
removal of the opacity (33). Visual improvement should be observed in most
cases, even in cases with retinal detachment (60). However, in cases with
advanced associated anomalies, such as severe retinal dysplasia or optic
nerve abnormalities, the occlusion therapy should not be overly aggressive.
Moreover, the treatment should be stopped without delay in these patients if
there is no improvement after a short trial to avoid undesirable psychosocial
impact (104). In bilateral patients, occlusion therapy is rarely performed as a
result of the symmetric severity between eyes (48).
SUMMARY
PFVS is a congenital condition that needs to be detected early in life for the
diagnosis and intervention. The successful management of PFVS starts from
the correct diagnosis and the careful assessment of the anatomical
involvement. The detailed history taking, comprehensive ocular examination,
and investigations play crucial roles for making the correct diagnosis and
differentiating PFVS from mimicking but critical diseases. The treatment
options of PFVS are varied depending on the extent of the pathologic
phenotypes. Fortunately, the advancement of the surgical techniques and
instrumentations improves the surgical and visual outcomes in the recent
years. However, the patients and family members also need to be counseled
preoperatively for the possible treatment options as well as the potential
complications that need to be monitored lifelong. The importance of the
postoperative follow-up and managements such as amblyopia treatment
should be emphasized for the successful PFVS management.
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66
Pediatric Vitreoretinopathies: Familial
Exudative Vitreoretinopathy, Norrie
Disease, and Incontinentia Pigmenti
Lauren M. Wright, and Yoshihiro Yonekawa
INTRODUCTION
Many pediatric vitreoretinopathies have retinal vascular origins of disease,
and the most well known are retinopathy of prematurity (ROP) and familial
exudative vitreoretinopathy (FEVR). Earlier stages of the vitreoretinopathies
are usually characterized by vascular anomalies and peripheral avascular
retina, which lead to neovascular proliferation and subsequent traction retinal
detachment. The goal of management is to first make the correct diagnosis
since there may be systemic implications. Then, we must identify the eyes at
high risk for, or in the earliest stages of, neovascular states, and attempt to
stop the progression. If eyes have progressed to retinal detachment, the goal
is to identify if, when, and how to intervene, in order to maximize the visual
potential of the child. This chapter will provide an overview of the surgical
management of FEVR, Norrie disease, and incontinentia pigmenti (IP). Other
vitreoretinopathies with surgical indications also include ROP, persistent fetal
vasculature, and X-linked retinoschisis, and these are covered in other
chapters. Coats disease is considered more of a retinopathy rather than a
vitreoretinopathy. Therefore, the surgical approaches are distinct and also
covered elsewhere (see Chapter 64).
FAMILIAL EXUDATIVE
VITREORETINOPATHY
Genetics
FEVR is a heritable retinal vascular condition marked by anomalous
vasculature and failure of the retinal vessels to develop into the periphery.
The biologic basis of the disease involves defects in the Wnt signaling
pathways. Many gene mutations have been implicated, and these are
discussed in Chapter 42.

FEVR is marked clinically by areas of avascular retina, often in the far
periphery, with vascular buds at the junction of the avascular and
vascularized retina. The disease presents variably with dragged vessels,
retinal folds, and intra- and subretinal exudation. The pathophysiology of
FEVR is discussed in Chapter 42. Exudative, tractional, and rhegmatogenous
retinal detachments (ERD, TRD, RRD, respectively) may occur, and stages
of disease are defined by extent of retinal involvement (see Table 66-1) (1).
FEVR may present bilaterally in as many as 85% of cases (2) but may be
markedly asymmetric with variable effects on visual development.
TABLE 66-1 Clinical classification of familial

exudative vitreoretinopathy
RD, retinal detachment.
Careful examination is crucial in making the diagnosis of FEVR, as the

presentation may mimic other vitreoretinal disorders, most notably ROP and
persistent fetal vasculature (PFV), see also Chapter 65. Distinguishing FEVR
from ROP and PFV is important, because FEVR is a life-long disease
requiring relatively more diligent long-term follow-up with genetic
implications. The surgical anatomy is often different as well, which
influences the surgical approach and goals as described below.
A history of prematurity and classic ROP findings in the neonate reliably
distinguishes ROP from FEVR in the majority of patients. Some cases can be
challenging to distinguish between the two, such as relatively older children
who present without known birth histories or children born only slightly
prematurely with vitreoretinopathy out of proportion to their relatively benign
gestational age (3,4). Family history, fluorescein angiography, genetic
testing, and examination of the fellow eye and family members can be
instrumental in such cases.
PFV is usually a distinct diagnosis from FEVR, but there can be
significant overlap in phenotype, and FEVR is not uncommonly accompanied
by elements of persistent fetal vasculature (3,5–7). Similar to above,
angiography, genetic testing, and examination of family members and the
status of the fellow eye will provide clues.
Diagnostic Studies
For many years, fluorescein angiography has remained an important ancillary
test in diagnosing and staging FEVR (7,8). Wide-field angiographic
capabilities has further enhanced our appreciation of the peripheral vascular
anomalies that exist in FEVR (1,2,9) and facilitates precise image-guided
laser treatment and identification of peripheral vascular activity. Ultra
widefield photography alone has been shown to be effective also (10), but
widefield fluorescein angiography is the most sensitive imaging study.
Optical coherence tomography (OCT) has revealed details of the pathologic
microanatomy of the disease, including posterior hyaloid
contraction/organization, vitreomacular traction (VMT), macular edema
(ME), intraretinal exudation, and subretinal lipid aggregation, and outer
retinal changes (11). When possible, OCT obtained in the clinic may help
identify active disease, guide surgical management, and inform postoperative
outcomes. Intraoperative OCT may offer similar utility in operative guidance.
Careful multimodal preoperative assessment can help guide treatment course
and provide prognostic information to families of affected children. In eyes
with hazy cornea or lens/media opacities, B-scan ultrasonography can help
identify the presence of retinal detachment and direction of tractional vectors,
which are useful for planning surgical approach and sclerotomy placement.
For female patients or carriers of inherited vitreoretinopathies like FEVR,
prenatal ocular ultrasound can also serve as a sensitive, valuable tool for
antenatal assessment of ocular pathology (12).
Treatment Indications
Indications for Laser and Anti-VEGF

Laser photocoagulation remains the mainstay of treatment for FEVR.
Addressing the avascular peripheral retina with laser photocoagulation is
recommended for stage 2 disease (retinal neovascularization) and can be
considered for stage 1 disease (avascular peripheral retina without
neovascularization) depending on the severity and other considerations
(presence of leakage, severity of nonperfusion, availability for follow-up,
fellow eye status, among others) (see Table 88-1). Since FEVR is more
progressive during the first few years of life, and examinations are
challenging or require general anesthesia in this age range, the threshold to
treat younger patients tends to be lower. Laser treatment is aimed not only at
preventing retinal detachment but also to avoid worsening exudation. Laser
becomes less effective after subretinal fluid and exudation develop, and the
macula can become compromised as well with progressive disease, leading to
poor visual outcomes. In general, cryotherapy is used less in recent times
mainly due to theoretically higher likelihood of hyaloidal contraction,
inflammation, and worsening exudation (13).
The role of intravitreal antivascular endothelial growth factor (VEGF)
injections is not well established in FEVR management. Its efficacy as
primary treatment has not been impressive in limited case series, and
management usually requires laser or surgical intervention (14). It may be
used preoperatively for eyes with severe vascular activity, but the eyes must
be monitored carefully for postinjection hyaloidal contraction (15,16). Anti-
VEGF treatment can also be used for the treatment of ME associated with
FEVR (11). Histopathologic studies have shown that an inflammatory
component is present in eyes with advanced FEVR (17). Localized
inflammation of endothelial cells likely contributes to earlier stages of disease
development as well, leading to capillary drop out. Supporting the
inflammatory role of the disease, a report has suggested that adjunctive
steroid treatment (topical and intravitreal injection) may decrease edema in
eyes with FEVR and may theoretically serve as a means to prevent capillary
dropout and advancing disease (18). Further studies are necessary to evaluate
the use of local corticosteroid treatment in FEVR.
Indications for Surgery

One of the most important considerations for managing retinal detachment in
FEVR is determining when not to operate, as not all retinal detachments
require surgery (Figure 66-1). For macula-sparing detachment (stage 3), the
goal is to prevent progression to a macula-involving detachment (stage 4). If
the tractional retinal detachment only involves the far peripheral retina, the
patient can undergo conservative management with consideration of only
laser photocoagulation of the avascular retina, especially if there is vascular
activity, seen as neovascularization, exudation, and/or leakage on fluorescein
angiography. Surgery can be considered if there is any progression on follow-
up examinations. With any retinal detachment in FEVR, but especially in
stage 5 (total detachment), chronicity of the retinal detachment is important in
determining whether surgery is indicated, as well as the status of the fellow
eye. Older children with stage 5 disease will have thin, atrophic retina with
dense membranes that may not benefit from intervention, and these eyes are
at very high risk for complications. Infants, however, may benefit from
surgery especially if there is bilateral disease and little to lose, but this of
course also needs to be balanced with the risks of surgery.
FIGURE 66-1 Retinal folds in familial exudative
vitreoretinopathy that do not require surgical intervention.
The first patient has a tight radial retinal fold (A) without
fluorescein leakage (B). The second patient also has a tight
radial retinal fold that inserts very anteriorly involving the
lens capsule outside the visual axis (C) without significant
fluorescein leakage (D). Such eyes usually do not require
surgical intervention.
Basic Considerations Regarding Technique

Any complication from vitrectomy in children tends to be less forgiving than
in adult eyes, so scleral buckling can be considered as first-line treatment for
peripheral progressive retinal detachment in eyes with FEVR. Supporting the
traction externally has been shown to decrease vascular activity and
progression (19). However, when tractional vectors of organized vitreous can
be easily appreciated and accessed and there is clear anterior-posterior
traction, vitrectomy may be the preferred technique. This is especially true if
connections to the posterior lens capsule are present, which is more often
seen in stage 4 disease with macular involvement.
Prior to consideration of vitrectomy, the extent and nature of contractile
tractional vectors should be assessed. Macula-involving TRD in FEVR is
often characterized by radial retinal folds with overlying organized hyaloid.
Tight, “dry” folds with retina–retina adhesion and no associated subretinal
fluid flanking the fold usually signifies a quiet eye and less likely to progress.
Surgery is usually not recommended in such configurations. The fold may or
may not settle with surgery, and there is often no change in visual outcome
based on our prior experiences. OCT can be useful in detecting whether there
is fluid around the fold or not (11) and for evaluating tractional vectors (20),
but OCT in eyes with poor vision can be challenging. The insertion of the
retinal fold also tends to be anterior and difficult to access with vitrectomy
without removing the crystalline lens in these eyes. A clear lens should be
kept in place as much as possible in a young child, and the complications
associated with aphakia usually outweigh the minimal, if any, improvement
in visual potential gained from operating on tight dry folds.
Vascularly active folds with associated fluid and exudation tend to be
broader in configuration with leakage on fluorescein angiography. These eyes
tend to be more amenable to improvement after surgical intervention. Laser
photocoagulation alone can be attempted in milder cases to see if nonsurgical
intervention can settle the retina and reattach the macula, or scleral buckle
alone if there is a focal peripheral knot of vascular activity or peripheral
traction anterior to the equator. However, as described above, vitrectomy is
often required in eyes with active stage 4 disease, wherein release of posterior
traction is the goal, as well as clearing of vitreous hemorrhage if present
(Figure 66-2).
FIGURE 66-2 Surgical management of familial exudative
vitreoretinopathy with traction retinal detachment and
vitreous hemorrhage. A 1-month-old girl presented with
vitreous hemorrhage (A). The fellow eye had classic
peripheral vascular findings of familial exudative
vitreoretinopathy (FEVR), and B-scan ultrasonography
showed a retinal detachment. A 25-gauge “short” system
is used where an infusion is sewed in place, and
sclerotomies are made with trocars without cannulas. This
allows sclerotomy creation without penetrating too deep
into the eye, where there is risk for retinal penetration in
severe traction retinal detachments. Also note that the
incisions are made 1 mm posterior to the limbus (B).
Vitrectomy is first performed in areas where we are certain
that there is only vitreous hemorrhage, and not retina.
Then, the vitreous cutter is gradually moved toward the
retinal detachment, making sure not to create iatrogenic
retinal breaks (C). The base of a broad retinal fold
becomes apparent (arrows, D). The fold inserts relatively
posteriorly, and there is a surgical space anteriorly that can
be dissected (E). Most of the hemorrhage and sheets of
hyaloid are removed, and there is an underlying broad
retinal fold (arrows, F). Air fluid exchange is performed
using the vitreous cutter to prevent vitreous incarceration
into the sclerotomies (G). The surgery was followed by an
examination under anesthesia several weeks later with
fluorescein angiography and indirect laser
photocoagulation. Six months after the surgery, the fold
has mostly resolved (H).
Selection of surgical approach based on degree of pathology was put forward

by Pendergast and Trese’s landmark article that defined stages of FEVR (2),
see also, Chapter 42. Based on their surgical outcomes, they suggested that
stage 3 eyes with predominantly exudative changes respond favorably to
scleral buckling alone, whereas stage 3 eyes with predominantly tractional
forces and stages 4 and 5 are likely to benefit from vitrectomy to directly
address the traction. More recently, Yamane et al. (19) expanded on surgical
outcomes with scleral buckle versus vitrectomy for progressive traction
retinal detachment in a series of 31 eyes. In this study, scleral buckling
resulted in cessation of fibrovascular proliferation and successful retinal
reattachment when performed in eyes with fibrovascular proliferation in the
peripheral retina extending circumferentially <2 quadrants (in 12 of 13 eyes).
Vitrectomy with or without lensectomy was performed for posterior
proliferation or proliferation in 2 or more quadrants of the periphery.
Reattachment was achieved in 7 of 7 eyes with vitrectomy alone and 7 of 11
eyes that required lensectomy–vitrectomy. In the latter four cases wherein
retinal reattachment was not achieved, all subjects were <15 months old and
progressed rapidly to total retinal detachment, highlighting the more
aggressive vascular proliferation and leakage that can occur in young patients
with FEVR. Nonetheless, Yamane’s study shows that anatomical success can
be achieved with carefully selected buckling in cases of peripheral traction
and with vitrectomy in many situations of posterior or extensive peripheral
pathology (see outcomes).
Equipment
To choose the most appropriate plan of care for the patient, thorough
preoperative assessment is necessary. An examination under anesthesia
(EUA) is often required to assess the extent and character of pathology in
children with FEVR. If scleral buckling or vitrectomy is being considered,
equipment necessary for these surgeries is ideally available at the time of
EUA when possible to minimize anesthesia exposure. For scleral buckling
and vitrectomy, elements and instrumentation vary depending on surgical
goals. Surgical equipment and instrumentation for pediatric retina surgery is
discussed in detail in Chapters 60 and 61.
Given the predominance of peripheral and anterior pathology in FEVR,
wide-field viewing is essential. The overall view and the visualization of
tractional vectors are usually superior during indirect ophthalmoscopy and
will likely be more of a challenge intraoperatively. Therefore, planning the
surgical maneuvers during the EUA first is useful. Large-scale studies have
not evaluated the safety and efficacy of endoscope-assisted vitrectomy for
eyes with stage 3, 4, and 5 FEVR, but the coaxial illumination and
visualization afforded by the endoscope is particularly useful to visualize
sheets of vitreous that require segmentation and may be an adjunctive option
as well. Choice of instrument gauge depends on surgeon preference and
anatomic goals. 25-gauge and 23-gauge systems are most commonly used.
Many cases can be performed with the vitreous cutter alone, such as to
address posterior tractional vectors and vitreous hemorrhage, but denser
membranes will require bimanual dissection, whether with picks and forceps
for blunt dissection, or with MVR blades, scissors, or needles for sharp
dissection.
Procedural Techniques
Retinopexy and Therapeutic Injections

There are no standardized guidelines yet on how to deliver the laser using the
laser indirect ophthalmoscope delivery system. In our practice, in most
circumstances of moderate to severe vascular activity, we prefer to apply
fluorescein-guided near-confluent laser spots across the areas of avascular
retina. The goal is to apply laser burns of moderate intensity. Neovascular
fronds are not treated directly, but laser immediately anterior to the
neovascularization is important. However, care must be taken to assure that
there is no focal retinal detachment—these areas should be avoided, as one
may unknowingly increase the laser power due to the lack of uptake from the
retinal detachment, which can lead to iatrogenic breaks. Guidance of laser
placement with the use of fluorescein angiography is helpful, and application
of fluorescein angiography is discussed more in Chapter 17.
Techniques and protocols for improved safety in intravitreal injections in
the infant and pediatric population have been described in ROP (21,22) (see
Chapter 53), and these recommendations are similarly applicable in FEVR.
Most importantly, a careful preinjection of EUA is required to assure that the
pars plana is available. Younger children have smaller available pars
plicata/plana anatomy (23). Eyes with FEVR may also have retina drawn up
anteriorly, and these areas should be avoided.
Vitrectomy and/or Scleral Buckle

When vitrectomy is pursued, the first intraoperative goal is not to create any
iatrogenic breaks, as the retina cannot be completely flattened in these
pediatric eyes with traction retinal detachment and, therefore, cannot be
retinopexied. The second goal is to transect the sheets of tractional organized
hyaloid to allow the retina to settle over time. If possible, carefully separating
the posterior hyaloid off the retinal surface is ideal, but not the primary goal,
as the posterior hyaloid is extremely adherent in children (24), and there is
increased risk for complications with aggressive lifting of the hyaloid.
Whether to completely release the posterior hyaloid or not depends on the
configuration of the vitreous traction and retinal detachment, and amount of
posterior pathology. The majority of eyes with stage 3 and 4 retinal
detachment can be repaired successfully by transecting the organized sheets
of vitreous, using the vitreous cutter. This can be accomplished without
aggressive membrane peeling, especially in young children with earlier
retinal detachment (Figure 66-3). If fibrotic membranes are directly
distorting the posterior pole, gentle membrane peeling can be considered, and
the hyaloid can be peeled off the macula. Hyaloidal removal to the periphery
would be ideal for more extensive fibrotic peripheral membranes, but the
hyaloid is usually very tightly adherent and, furthermore, this is over
peripheral atrophic retina increasing the risk of iatrogenic break formation. If
satisfactory membrane removal is not possible, the area can be supported by a
buckle (25). Posterior hyaloid separation and peeling tends to be easier in
older children and adults with FEVR, but great care must be taken especially
in younger patients to avoid iatrogenic breaks during such maneuvers.
Enzymatic vitreolysis, such as with autologous plasmin, has been used
adjunctively to facilitate hyaloidal separation (26), although this is not readily
available.
FIGURE 66-3 Vitrectomy for familial exudative
vitreoretinopathy with traction retinal detachment. A five-
month-old girl presented with a relatively posteriorly
inserted radial retinal fold (A). There is leakage noted on
the widefield fluorescein angiography (B), and peripheral
vascular changes consistent with familial exudative
vitreoretinopathy (FEVR).FIGURER 66-3(Continued) 25-
Gauge vitrectomy is performed, but note that the incisions
are made 1 mm posterior to the limbus, and the
superotemporal sclerotomy is placed relatively superiorly
to avoid the retinal detachment (C). The organized sheets
of hyaloid connecting the temporal aspect of the fold to the
crystalline lens capsule are first dissected to create space
to maneuver (arrows, D). The tractional vectors
connecting the temporal aspect of the fold to the eye wall
are then dissected (E). There is also a sheet of organized
hyaloid running along the retinal fold and inserting
anteriorly, which is dissected (arrows, F). Endolaser is
applied to the avascular retina (G). Eight months
postoperatively, the retina is mostly reattached (H) and the
leakage has resolved (I).
We generally follow the adage of “less is more” for these surgeries. Because
children can develop relentless proliferative vitreoretinopathy, retinectomy is
generally not recommended as well. Unlike surgery in adults, the goal is not
to have the retina flat at the end of the case. The goal of surgery should be to
cut tractional vectors to release the traction and watch the retina gradually
reattach over time by the pumping mechanism of the retinal pigment
epithelium (27). This is a similar concept in ROP surgery as well (28).
The above tenants are particularly relevant in surgery for stage 5 FEVR
(total retinal detachment) (25,29). Anterior entry into the eye is often
required, such as through the corneal limbus or via iris root sclerotomies.
Unlike stage 5 ROP where there is often a peripheral trough, especially if the
eye was previously lasered, eyes with stage 5 FEVR tend to have very
anterior adhesions to the eye wall. Lensectomy may serve to release
retrolental traction and/or provide access to peripheral anterior membranes
(2,25,30–32). With removal of the crystalline lens, the capsule must be
removed in its entirety, as residual capsule can behave as a scaffold for
cellular proliferation and result in persistent or worsening retinal detachment.
The lensectomy/capsulectomy alone will shift the retinal detachment complex
more posteriorly. If there was corneal compromise from a shallow anterior
chamber, this step also helps to allow the cornea to clear over time. A staged
surgery can be performed in such cases, where the surgeon can return to the
operating room several weeks later to proceed with the membrane dissection
now with a more clear view.
Careful membrane dissection is performed in a variety of ways depending
on how dense the preretinal membranes and plaques are. Bimanual
techniques are usually required where one hand secures the tissue and the
other hand moves in the other direction to initiate the surgical plane and
undermine the membranes. Sharp or blunt dissection can be performed
depending on the density of the membranes. The retina may become more
mobile as the dissection progresses, which is an encouraging sign, but also
increases the risk for iatrogenic break formation. There is little to no harm in
returning several weeks later to complete the dissection, compared to an
iatrogenic break, which may signify the end of treatment. If the subretinal
fluid resorbs and some of the retina can reattach, that will make for easier
subsequent dissection. Persistent subretinal fluid can also be drained
externally, which may help expedite the process, especially if there is
subretinal hemorrhage.
Tamponade is not required unless there is a rhegmatogenous component
to the retinal detachment. If iatrogenic breaks are created, the
recommendation is to relieve the traction over the break as much as possible
and use silicone oil tamponade. A scleral buckle to support the area may be
beneficial also. Repairing a rhegmatogenous component is exceedingly
difficult in stage 5 retinal detachments, but possible in stage 3 or 4, and
placement of oil may also reduce the stimulus for vascular leakage (33,34).
Primary RRD can occur in FEVR also. Interestingly, FEVR-related
primary RRD appears to be more common in Asia (25,31,35–43). Peripheral
vascular abnormality and atrophic changes may predispose to retinal holes
and breaks, as seems to be the case in ROP (44,45). Myopia may contribute
added risk (42). One series showed the most common etiology of FEVR-
RRD to be retinal holes within lattice, but horseshoe tears and giant retinal
tears also occur (43). In fellow eyes of FEVR-RRD cases, lattice
degeneration may be found as well as peripheral retinal fibrovascular
membranes, vitreous traction, and retinal breaks. These changes may
predispose to later onset detachment compared to TRD, as FEVR-RRD
frequently presents in the second to third decade (43).
Rhegmatogenous detachments in eyes with milder stages of FEVR can be
treated relatively similar to conventional RRD. However, the hyaloid is more
adherent and broadly inserted, so primary scleral buckling can often be
considered over vitrectomy (39). For combined rhegmatogenous–tractional
retinal detachment in eyes with more advanced stages, more aggressive
surgery may be required with vitrectomy with or without scleral buckling. As
described above, since there is already a retinal break, all traction needs to be
released from the break, treated with retinopexy and tamponade.
Postoperative Care
Wide-field angiography remains important during follow-up examinations
with or without laser treatment or surgery. Oral fluorescein may be useful as
an alternative to intravenous fluorescein for angiography performed in the
office setting for children who are old enough for imaging, but too young to
tolerate intravenous needles (27,46,47). There may be long periods of
inactivity in FEVR with late reactivation particularly in the nonvitrectomized
eye; therefore, lifelong screening is recommended. For individuals who have
undergone vitrectomy with long-term silicone oil placement, monitoring for
oil migration or emulsification, glaucoma, and cataract is important. Visual
outcomes in these eyes may be poor, and surgical aphakia is common. FEVR
tends to be asymmetric, so monocular precautions for the better seeing eye
are paramount.
Complications
Complications after Laser Treatment

Complications of laser ablative treatment in FEVR are similar to those in
ROP. Immediate complications of the laser procedure can include lens or iris
damage, vitreous hemorrhage, inflammation, elevated IOP, hypotony, and
anesthesia-related complications. Failure to completely ablate the avascular
retina can result in progressive angiogenic stimulus, causing continued
vascular activity and proliferative complications. Recurrent or persistent
exudation can also occur despite laser treatment.
Complications after Vitreoretinal Surgery

Postsurgical complications in FEVR may include infection, bleeding,
cataract, corneal decompensation, and hypotony or elevated IOP. Intra- or
post-op development of a rhegmatogenous break can occur and can be
addressed as described above. When use of a tamponade is indicated,
postoperative positioning can be challenging, and oil emulsification,
particularly with 1,000 cs silicone oil may occur. Unrelenting PVR in the
event of a retinal break can lead to phthisis and blindness. Redetachment and
progression can also occur from recurrence of proliferative and tractional
disease, so the patients require continued monitoring.
Visual outcomes in FEVR vary widely given variable penetrance of the
disease and the wide range of phenotypic changes that are observed
clinically. In general, stage of disease correlates with visual outcomes
(2,30,38,48). This may not always be the case, as peripheral pathology
without detachment may cause macular dragging, other posterior vitreoretinal
interface abnormalities, or outer retinal abnormalities (11). Even in cases of
seemingly mild disease, optic nerve hypoplasia has been found, which may
have variable effects on visual development (49). In cases of advanced
disease with retinal detachment, poor visual prognosis without intervention is
certain. Success rates for reattachment are variable and depend on many
factors, including disease chronicity and stage. Rate of retinal reattachment
with vitrectomy in published series including stage 5 disease ranges from
41% to 86% (2,25,30,37,38,48), and improvement in visual acuity may occur
in over 70% of cases (25). Even in select advanced cases, intervention can be
considered to restore as much visual function as possible (32).

Patients with FEVR require ongoing surveillance to monitor disease activity.
Serial laser treatments may be necessary, and the retina can still detach
despite laser treatment and may require surgery, as discussed above.
NORRIE DISEASE
Genetics
Norrie disease is a rare X-linked recessive disorder with invariably severe
clinical manifestations marked by hearing loss, cognitive delay, and blindness
in affected patients. It is caused by severe mutations/deletions in NDP (MIM
310600) (50,51), a gene also implicated in FEVR (52–54); therefore, Norrie
disease may be considered as an extreme phenotype of FEVR. See also
Chapters 35 and 42 on Norrie and Wnt signaling and related diseases.
Retinal manifestations are often evident at birth. The hallmark of Norrie
disease is severely dysplastic retina in both eyes, accompanied by
aggressively progressive retinal detachment and in many cases with vitreous
and subretinal hemorrhage (29,55,56). Eyes with Norrie disease classically
present with a posterior “pumpkin” lesion (57), which represents a
disorganized mass of retina-derived glial tissue (58,59). There is extensive
retinal nonperfusion, and TRD usually progresses quickly from contraction of
the primitive vitreous and tissue proliferation.
Indications
Most individuals with Norrie disease will unfortunately develop phthisis
bulbi, but early surgical intervention can be beneficial (57). Given the visual
potential with early intervention, it is ideal for patients with Norrie disease to
have a pediatric retinal examination as soon as the diagnosis is considered,
ideally soon after birth, so that the vitreoretinopathy can be detected and
treated. Prenatal diagnosis of Norrie-based maternal ultrasound has been
described (60,61) and such early recognition may help in timely perioperative
planning. Those with family history of Norrie disease will also be aware of
the possible diagnosis. However, a thorough informed consent process with
the patient’s guardians is recommended to ensure that all parties are aware of
the generally poor visual prognosis in Norrie disease. There are currently no
guidelines dictating at which age or chronicity of the retinal detachment that
surgery becomes futile. The decision to intervene will vary from patient to
patient.

Although Norrie disease is commonly accompanied by vitreous and
subretinal hemorrhage, the eyes usually do not develop retinal
neovascularization. One thought that has not been verified is that the retina is
too dysgenic to secrete proangiogenic cytokines. Practices vary, but some
defer laser photocoagulation for this reason and opt for early surgical
intervention if possible. Furthermore, patients with Norrie disease often
present with complete retinal detachment in both eyes and laser is not even an
option.
Equipment
Equipment is as described above for FEVR, though scleral buckle is
uncommon due to severe posterior involvement.
Early retinal detachment in Norrie disease is characterized by a traction
retinal detachment surrounding the posterior glial mass. The goal of
vitrectomy is to transect the tractional hyaloid 360 degrees (Figure 66-4). At
this early stage, membrane peeling is usually not necessary. The retina will be
able to settle and reattach. For eyes that have progressed to funnel retinal
detachment, open funnel configurations will yield superior outcomes
compared to closed funnel retinal detachments, so intervention should not be
significantly delayed. Complete retrolental retinal detachment can be
operated on, especially if the funnel closure has been recent, but the surgeries
are challenging and high risk with guarded prognosis. The approach is similar
to stage 5 FEVR surgery as outlined above. The main difference is that the
retina is even more dysplastic than typical eyes with severe FEVR or ROP,
and it can be difficult to distinguish retinal tissue from organized hyaloid and
proliferative tissues.
FIGURE 66-4 Early vitrectomy for traction retinal
detachment in Norrie disease. A 2-week-old boy with a
family history of Norrie disease was screened soon after
birth and noted to have bilateral traction retinal
detachment with vitreous and retinal hemorrhage (A).
Fluorescein angiography shows extensive nonperfusion of
the retina (B). A 23-gauge system is used. The infusion
cannula is placed in the anterior chamber because there is
an anterior retinal detachment in the vicinity. The
superotemporal cannula is placed in the pars plicata as the
retina is confirmed to be attached in that quadrant. The
retinal detachment is very anterior and attached to the lens
capsule nasally, so a limbal incision is made superonasally
(C). Lensectomy is performed because of the insertion of
the retina to the posterior lens capsule. The nasal cannula
is subsequently removed due to the lack of surgical space.
The capsule is removed carefully in its entirety (D).Figure
66-4(Continued) Vitrectomy is performed with the goal of
transecting the hyaloidal face and other hyaloidal
insertions that connects the retina to itself and the eye wall
(arrows, E). This process is continued 360 degrees to
release the retina from progressive hyaloidal traction
(arrows, F). Air fluid exchange is performed to prevent
vitreous incarceration (G). On post-op month 1, the retina
is starting to reattach as seen on ultrasonography (H). The
view is limited from vitreous hemorrhage. On post-op
month 8, the retina is now completely reattached (I) and
confirmed on ultrasonography (J), and the patient retains
brisk light perception. The fellow eye had similar
presentation and management, but developed progressive
vitreous and subretinal hemorrhage, after undergoing
conservative management.
Lensectomy is often required for “stage 5” Norrie detachments, and again,

complete capsulectomy is a key step. Iridectomy may be a prudent procedure
also, as retina adhered to iris after a lensectomy is almost impossible to
dissect. Single stage repair of total funnel retinal detachment in Norrie
disease has been described in detail with translimbal iridectomy, lensectomy,
capsulectomy, and vitrectomy with careful dissection of the retrolental
membranes to allow opening of the funnel and reattachment of the posterior
pole (29).
Postoperative Care and Complications

These are comparable to those for FEVR, with higher risk of phthisis bulbi in
Norrie disease.
Given the rarity of the disease, and tendency toward no light perception
vision early in life, there are limited reports on surgical interventions for
Norrie disease. However, the literature and our experiences suggest a benefit
to early surgical treatment. In the largest surgical series wherein 14 boys with
definite Norrie disease had vitrectomy with or without lensectomy in at least
1 eye prior to 12 months of age, half of the boys maintained at least light
perception visual acuity (57). The best outcomes are achieved when
intervention is possible prior to advanced retinal detachment.
INCONTINENTIA PIGMENTI
Genetics
Incontinentia pigmenti (IP), or Bloch-Sulzberger syndrome, is a rare X-linked
dominant ectodermal condition caused by mutations in the IKBKG gene. It is
mainly seen in females as it is usually fatal in males. See also Chapter 36.

Patients with IP can experience vision-threatening disease associated with
abnormal retinal perfusion that can potentially cause aggressive traction
retinal detachment. Unlike ROP and FEVR, which stem from incomplete
development of vascularization, the retinal vasculopathy in IP appears to be
also caused by occlusion of previously formed vessels. Severity is often
asymmetric. Exudation, hemorrhage, and hyaloidal contraction can occur
leading to TRD (62). Other causes for vision loss can be from macular
vascular anomalies, retinal artery occlusions, optic neuropathies, and central
nervous system involvement.
Diagnostic Testing
Wide-field fluorescein angiography is an important diagnostic tool and can
reveal nonperfused retina, an avascular retinal periphery, arborization,
vascular loops, and leakage of fluorescein from incompetent vasculature and
neovascularization (63–65). Consistent with vascular occlusive disease, OCT
shows abnormal inner retinal structures that can involve the fovea in
symptomatic or asymptomatic eyes (66,67). Ultrasound is helpful in
evaluating the configuration of retinal detachment through vitreous opacities.
Indications
The retinal vascular disease in IP is the most active and progressive the first
several weeks and months of life. Examination as soon the diagnosis is
suspected is recommended, since early treatment can halt progression to
blinding sequelae.
Wide-field fluorescein angiography-guided laser photocoagulation of the
avascular retina is recommended in eyes with retinal neovascularization to
prevent progression to retinal detachment. There are no established guidelines
for treatment of avascular retina without neovascularization; some
recommend laser of all avascular retina, whereas others recommend
observation (68–70). High-risk angiographic signs, such as vascular leakage
and severe nonperfusion, may warrant early treatment. There may be lower
thresholds for treatment in monocular patients and those who may not be able
to regularly maintain follow-up. The tempo of progression in young children
can be very rapid, so diligent follow-up is recommended, regardless of
treatment strategy.
Vitreous hemorrhage is a common finding in severe IP-related
retinopathy. Media opacities will require B-scan ultrasonography to identify
an underlying retinal detachment. The threshold to surgically clear the
vitreous hemorrhage even in the absence of a retinal detachment should be
low in infants with IP, because of the high likelihood of active neovascular
disease that can rapidly progress, and which may be halted with vitrectomy
and laser, in addition to the risks of deprivation amblyopia.
Similar to late presentation of advanced closed funnel retinal detachment
in the other vitreoretinopathies, and perhaps more so in IP, aggressive surgery
should be weighed against the possibilities for quick deterioration with
surgical sequelae. IP can be asymmetric and, if the fellow eye sees well,
conservative management of delayed presentation of complete cicatricial
retinal detachment is a reasonable option. Monocular precautions are of
course emphasized (Figure 66-5).
FIGURE 66-5 Management of incontinentia pigmenti. An
11-year-old monocular boy with genetically confirmed
incontinentia pigmenti (note that incontinentia pigmenti is
rare in boys, as it is usually lethal in males) presented with
floaters in the right eye. He has mild retinal and vitreous
hemorrhage (A) and retinal neovascularization (B). His
fellow eye had undergone surgical interventions as a
newborn for vitreous hemorrhage and subsequent retinal
detachment but was deemed inoperable and now has dense
band keratopathy and corneal blood staining.
Ultrasonography shows a funnel retinal detachment (C).
The right eye was treated with laser photocoagulation for
mild, yet progressive, retinopathy in a monocular patient.
Most progression normally occurs in the neonatal period.
Equipment
Equipment is as described above for FEVR.
In IP, the overall surgical goal is similar to traction retinal detachment
surgery in young children with ROP, FEVR, and Norrie disease, which is to
clear the media and transect the tractional vectors without creating iatrogenic
breaks.
Postoperative Care
Similar to most conditions with avascular peripheral retina, IP is a lifelong
condition. “Reactivation” of neovascular disease is possible but less likely
with age. Rather, the avascular retina predisposes these eyes to
rhegmatogenous complications. As shown in patients with ROP and FEVR,
untreated avascular retina is prone to retinal breaks in teenage years and
young adulthood, as well as when posterior vitreous detachments occur later
in life (69,71). Lasered peripheral retina may be protective in this regard.
Complications and Outcome Expectations

Retinal detachment in IP particularly portends a poor prognosis due to the
vascular activity of the eyes, which may present with intense
neovascularization and hemorrhagic sequelae (72). Similar to other
vitreoretinopathies, laser intervention may halt the progression to retinal
detachment (73), and if retinal detachment does occur, ideally it can be
addressed prior to posterior involvement (62,63,74). The first successful
vitreoretinal surgery for retinal detachment in IP resulted in partial
reattachment in one of three eyes and complete reattachment in another of
those eyes (71). From a review of 25 patients with IP followed up to 23 years,
a bimodal distribution of retinal detachments was observed (69). Most
tractional detachments (7 eyes) occurred by age 2.5 years, and most
rhegmatogenous detachments (4 eyes) occurred in adults. Three eyes of
young patients (≤2.5 years) developed tractional detachment, despite
prophylactic ablation in 4 eyes.
If patients do develop RRD in older age, the anomalously inserted posterior
hyaloid needs to be appropriately addressed, which usually implies that a
scleral buckle can be considered as part of the surgery.
SUMMARY
FEVR, Norrie disease, and IP are among pediatric vitreoretinopathies with
significant vasculogenic pathology and potentially devastating visual
outcomes. Identification of ischemic and preproliferative stages of disease
and prompt treatment when indicated is paramount to achieving hopeful
outcomes. When necessary, surgical management must be approached with
care, and avoidance of surgery may at times be in the child’s best interest
depending on disease extent and visual potential, and largely dependent on
the status of the fellow eye and systemic considerations. There are windows
of best opportunity for surgical intervention, so appropriate surgical timing is
an important consideration. Goals of surgery and technical approach will
differ based on diagnosis and stage of disease as described above. We hope
this chapter offers an organized approach to surgical care in these conditions.
Please note that there are many ways to approach these surgeries, and
different surgeons will have different techniques, but we tend to share general
philosophies of treatment. General principles of pediatric retinal surgery are
discussed in Chapters 56-62. We hope that further innovations in surgical
instrumentation, techniques, imaging and visualization, together with
advances in our understanding of the pathophysiology, will continue to
enhance our ability to optimize outcomes for these children.
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67
Epiretinal Membrane and Combined
Hamartoma of the Retina and Retinal
Pigment Epithelium
Cynthia A. Toth
INTRODUCTION
An epiretinal membrane (ERM) with nonvascularized cellular proliferation
along the surface of the retina is rare in infants and children, and when it
occurs, is usually unilateral (1). Whereas contracted preretinal tissue is found
in combined hamartomas of the retina and RPE (CHRRPE) (2–4), pediatric
ERM is more commonly idiopathic or secondary to other conditions
including after trauma (such as after penetrating trauma or intraocular foreign
body, or after nonaccidental trauma with premacular hemorrhage (5) or
without pre- or submacular hemorrhage (6)); in chronic retinal detachment
with proliferative vitreoretinopathy; or secondary to inflammation, retinal
vascular disease (Coats), neovascular disease (retinopathy of prematurity,
familial exudative vitreoretinopathy), or toxocariasis (7,8). Pediatric ERM
that is not a result of another ocular condition, or CHRRPE, may be
associated with neurofibromatosis type 2 and is sometimes the earliest
indicator of the more severe phenotype of the disease (9–13). This chapter
will focus on idiopathic ERM and CHRRPE, although the perioperative
considerations and techniques presented here apply in general to the
management of epiretinal proliferations in children.
CLINICAL FEATURES AND NATURAL

HISTORY
Indications
Depending on the age of the child and the location and extent of proliferative
tissue and deformation, a pediatric ERM may result in pronounced vision loss
in the involved eye. ERMs are not commonly identified in very young
children; the majority of pediatric cases are found in children of school age
and older (1,7,8,14–16). Unlike adults, children rarely complain of symptoms
from ERM and, therefore, these are most commonly identified because of
poor visual acuity or an abnormal light reflex or fundus appearance. New
onset of strabismus or even leukocoria may be the first sign of these retinal
conditions (1,4). In chronic macula-involving ERM in infants and very young
children, prolonged deprivation of the retina from sharp formed vision may
result in additional visual limitation from amblyopia. Determining the impact
of the lesion itself and additional impact of amblyopia can be confounded by
the lack of good-quality past ocular history. In children, the timeline of
decline in visual acuity may be lacking, and there may be limited information
on whether and how the configuration of the ERM and retina has changed
over time. Several reports of pediatric ERM note the lack of need for surgical
intervention in numerous cases and the stability and/or improvement in acuity
in cases without surgery. Spontaneous separation of pediatric ERM from the
retina can also occur (17,18).
Epiretinal Membrane
Idiopathic pediatric ERMs have been found centered on retinal vasculature,
extending from the region of the optic nerve head, within the macula, or in
more peripheral locations (1,14,15) (Figure 67-1). The most notable
distinction between epiretinal proliferation in the child versus in the adult is
the persistent adhesion/attachment of the posterior vitreous even in the
presence of condensed posterior hyaloid and cellular proliferation (Figure
67-2). The structure of the vitreous and the vitreoretinal interface in children
is distinct from adults; vitreous liquefaction is atypical under the age of 20
even after trauma or with retinal detachment and progresses with increasing
age (19). The adhesion between the vitreous and retina across the posterior
pole persists into the third decade of life (20). Despite this, in children, glial
proliferation across the retinal surface may result in axial and lateral traction
to the retinal surface resulting in partial-thickness deformation, full-thickness
folds, schisis, and even retinal detachment and choroidal neovascularization
(21).
FIGURE 67-1 Vasocentric epiretinal membrane in an 11-

year-old child with progressive central vision loss. Infrared
imaging on Spectralis (top left) highlights the extent of
gliosis. OCT structural imaging (top far right)
demonstrates partial PVD adjacent to thick preretinal
tissue, and intraoperative OCT shows that same location
before (top middle) and after (middle row middle) removal
of the contracted tissue. This required sharp dissection at
the arcade. The shift in location of the superior arcade
several months after surgery is visible in the repeat scan
location on the infrared image in the middle row, far right,
as is the focal recurrent membrane at the margin of the
vessel. Preoperative fluorescein angiogram (bottom row)
demonstrates a lack of leakage at the site of the ERM and
within the macula. Best corrected visual acuity improved
from 20/126 before surgery to 20/25-1 two years later.
FIGURE 67-2 A macular epiretinal membrane resulted in
new onset of vision loss in a preteen child who had
peripheral laser treatment as an infant for retinopathy of
prematurity. The epiretinal membrane is associated with
dragged vessels, darker retinal appearance, retinal
thickening, and late fluorescein leakage within the macula
and along inferior arcade. This ERM, which appears to be
secondary to prior ROP treatment, bears some similarity to
CHRRPE, and the earlier arteriovenous phase of the
angiogram (lower left) demonstrates one small focus of
tortuous small vessels with leakage along the
inferotemporal arcade (lower right) as well as in the fovea.
The intraoperative OCT before ERM removal reveals the
ERM closely approximated to the retinal surface (middle
left); this lacks the prominent gliosis that is more typical of
CHRRPE, and after ERM removal reveals the bare retinal
surface and residual elevated ILM at the margins of the
peeled area (middle right).
ERMs documented in children are typically not of the thin transparent

“cellophane” type; the majority of pediatric ERMs appear as thick white
contractile premacular fibrosis with retinal striae (1,16,22) (Figures 67-1 and
67-6). In support of the increased contractility of ERM in children, Smiddy et
al. compared histopathology of 11 pediatric and young adult ERMs to reports
for adult ERMs and found more new collagen formation, myofibroblasts, and
myoblastic differentiation of fibrous astrocytes or retinal pigment epithelial
cells in the pediatric tissue (22).
Combined Hamartoma of the Retina and Retinal

Pigment Epithelium
In 1973, Gass described CHRRPE and pointed out features that distinguished
these lesions from malignant melanoma or retinoblastoma (2). CHRRPE are
generally unilateral. The Gass definition included (a) a slightly elevated black
or charcoal gray mass involving the pigment epithelium, retina, and overlying
vitreous (b) extending in a fanlike projection toward the periphery, (c) a base
sheet of flat gray tissue, (d) thickened gray-white retinal and preretinal tissue,
(e) contraction of the inner surface of the hamartoma, (f) absence of RPE or
choroidal atrophy, and (g) absence of retinal detachment, exudation,
hemorrhage, and inflammation (2) (Figures 67-3 and 67-4). The early
definition was augmented by the addition of a review of 60 cases (mean age
15 years) gathered from members of The Macula Society in 1984 (4). In that
review, Schachat et al. described the spectrum of predominant tissue
contributions in different CHRRPE lesions, ranging from melanocytic to
vascular or to glial tissue, and discussed whether idiopathic epiretinal
proliferations might be included at one end of this broad spectrum of
“combined hamartoma.” A series of 77 cases of CHRRPE in a younger group
(median age 7.5 years) demonstrated poorer visual acuity with macular
involvement (23). Another series of 53 patients documented growth in six
patients (24). CHRRPE can occur on or near the optic nerve, in the macula,
or in the periphery (Figures 67-3 to 67-5). In contrast to macular lesions,
CHRRPE around the optic nerve head are more likely to display severe
pigmentary changes and choroidal neovascularization (25) (Figure 67-5).
Figure 67-3 Combined hamartoma of the retina and retinal
pigment epithelium in a 2-year-old child. The lesion
combines hyperpigmentation and epiretinal membrane and
is extramacular and not affecting visual acuity. It has been
monitored during outpatient visits with Optos imaging for
documentation and close-up examination.
Figure 67-4 A 15-month-old with CHRRPE who was
referred for evaluation for elevated retinal lesion, optic
nerve hypoplasia, and history of “eye turn” for several
months. Although the retinal vessels and macula are
dragged toward the superonasal lesion (top left) with retina
over the optic nerve (SDOCT top right), the microvascular
changes on fluorescein angiography (lower panels), flat
gray pigmentary changes beneath the fan-shaped gliosis
and contracted retinal surface of the lesion all point to the
CHRRPE diagnosis. He had surgery with release of
traction and removal of some of preretinal gliosis. After
surgery, the infant had patching of fellow eye and
refractive correction, and he was able to identify a 6-mm
object using this eye.
Figure 67-5 Color, fluorescein angiogram, infrared

images, and OCT in an 8-year-old child with peripapillary
CHRRPE and choroidal neovascularization with subretinal
fluid, hemorrhage, and vision loss. The CNV and intra-
and subfoveal fluid (upper OCT) respond to a series of
anti-VEGF injections and a verteporfin photodynamic
therapy with anti-VEGF injection under anesthesia. The
epiretinal gliosis was not treated. The child had resolution
of the subretinal fluid (lower OCT) and remained
treatment free for >5 years with visual acuity of 20/30.
SYSTEMIC ASSOCIATIONS AND
Although most patients with CHRRPE do not have other systemic diseases,
unilateral CHRRPE have been associated with neurofibromatosis type 1 and
type 2, tuberous sclerosis, Gorlin syndrome, and juvenile nasopharyngeal
angiofibroma (2,9,17,26–33). Pediatric ERMs that are not a result of another
ocular condition may also be associated with neurofibromatosis type 2
(9–13), and in one case, the OCT pattern of thick ERM with an elevated
rolled edge was used as an indicator, which led to a later diagnosis of
neurofibromatosis type 2 (12). Rare bilateral CHRRPE are associated with
neurofibromatosis type 1 or more commonly type 2 (27,34,35) and may be
the first indicator of the disease (28). Prevalence of CHRRPE and ERM in
neurofibromatosis type 2 increases with mutations associated with more
severe systemic disease (36). Thus, evaluation for neurofibromatosis in cases
of CHRRPE or ERM may allow for timely management of other
manifestations of the disease.
Although many of the lesions remain stable, progressive retinal traction
and ERM contraction may result in vision loss especially when the macula is
involved (23) (Figure 67-4). Complications of retinal, vitreous, or subretinal
hemorrhage; macular hole; choroidal neovascularization or exudation; and
lesion growth may occur (4,23,24,37).
DIAGNOSTIC STUDIES
Multiple modes of imaging may be useful in the diagnosis of ERM and
CHRRPE. Fluorescein angiography is important in the evaluation to
differentiate idiopathic ERMs from those associated with retinal vascular
diseases, CHRRPE, other tumors, or inflammation. The characteristic
fluorescein angiographic findings in CHRRPE may vary but include blocked
choroidal fluorescence due to pigmentation, straightening of surrounding
retinal vessels from traction, and tortuous retinal vessels with retinal capillary
dilation telangiectasia shunts and late leakage within the lesion (2,4) (Figures
67-4 and 67-5). Autofluorescence imaging also reveals the pigment blockage
and faint hyperautofluorescence in areas of CHRRPE (25,38).
Especially in peripapillary CHRRPE, the fluorescein angiogram can
reveal associated choroidal neovascularization (Figure 67-5). Optical
coherence tomography (OCT) is well tolerated by children and used to
identify characteristics of the vitreoretinal interface and retinal and subretinal
structures. OCT provides information on the extent of localized vitreous
bands or vitreous separation and of the extent of preretinal tissue and retinal
contact, retinal deformation and folds, schisis, or retinal detachment
associated with the ERM or associated retinal condition (7,8) (Figures 67-3
and 67-5). In contrast to adults, in children, retinal dragging and full-
thickness deep “taco folds” of the retina (Figure 67-6) are more common as
are broad swaths of adhesion between preretinal tissue and the retina (7)
(Figure 67-2). Analyses may also be confounded on OCT imaging by deeply
folded retina in which the layers may appear disorganized due to the atypical
plane of OCT sectioning across steeply folded retina. Similar to adults,
evidence of photoreceptor integrity on OCT appears to be predictive of better
visual outcomes, although dense preretinal tissue and thick retinal folds may
shadow and prevent such a preoperative assessment (7,15).
FIGURE 67-6 A macular epiretinal membrane in a child
with a prominent deep “taco fold” of the retina on OCT
imaging (A, B).
Structural OCT and OCT angiography imaging have further refined our
understanding of CHRRPE. An ERM and retinal adhesion or separation
along with vitreous attachment and retinal deformation and folds is delineated
on the OCT (18,38,39). Inner retinal changes involve thickening, cystoid
spaces, and disorganization of the inner retinal layers and may include
hyperreflective foci, schisis, and striae and either stop at the outer plexiform
layer or involve the deeper layers including the RPE (3,25,39–41).
Hyperreflective, fine sawtooth changes or larger omega-shaped changes in
the outer plexiform layer may occur in lesions in which the outer retinal
changes are less severe (3,39). There is great variability in abnormalities from
mild to severe, including subretinal fluid and RPE alterations, in the outer
retina and RPE (42). Peripapillary CHRRPE demonstrate more severe full-
thickness involvement on OCT compared to macular lesions (25), and
choroidal neovascularization and subretinal fluid may be evident on OCT in
CHRRPE or an ERM (Figure 67-7). In such cases, OCT is a valuable tool for
monitoring response to therapy. Because other ocular or systemic factors
such as brain injury may contribute to vision loss in a child with an ERM (6),
the impact of other disease processes within the eye and of other systemic
health factors should be considered in the evaluation of pediatric ERM,
particularly when considering surgery for visual improvement. On OCTA of
macular CHRRPE, the vessel density in superficial and deep retinal layers
and choriocapillaris has been found to be decreased compared to images of
healthy eyes of the same age (39), or vessels deformation is evident from
traction or schisis (40).
FIGURE 67-7 An epiretinal membrane from proliferative

vitreoretinopathy in an 11-year-old child who was referred
after multiple surgeries for rhegmatogenous retinal
detachment repair. Inner retinal folds, loss of foveal
contour, and retinal schisis are visible on preoperative
OCT (upper right) while gliosis and deformation of the
superior arcade are visible on the infrared image (upper
left). After surgical removal of the ERM and internal
limiting membrane in the macula, the retinal vessel
deformation and inner retinal schisis are nearly resolved.
Old laser scars are visible in the temporal periphery. The
visual acuity improved from 20/250 before surgery to
20/60 several months later.
CLASSIFICATION
In light of the range of presentations of retinal involvement with CHRRPE,
Dedania et al. suggested a classification by location (zones 1 to 3 for
posterior, mid periphery, and far periphery), traction (stages 1 to 3 for no
retinal traction, retinal traction/schisis, and retinal detachment), and retinal
components involved based on OCT (A epiretinal component only, B partial
retinal involvement, C complete retinal and RPE involvement) (43). This
classification does not address vascular involvement and leakage and does
not include whether central macula is or is not involved with lesion/ERM
(23), which would be useful to include for studies of interventions or of
functional outcomes.
EQUIPMENT
Small-gauge vitrectomy system.
Fine pick or barbed needle.
Vitreoretinal forceps.
Vitreoretinal scissors may be used for sharp dissection.
Triamcinolone may be used to mark vitreous or membrane.
Indocyanine green or other dye may be used to stain internal limiting
membrane.
Intraoperative OCT imaging, handheld or in the microscope may aid in
viewing tissues.
Choroidal neovascularization in peripapillary CHRRPE may involve the
macula and cause vision loss. Treatment with anti–vascular endothelial
growth factor can arrest the vascular leakage and improve visual acuity
(Figure 67-5). Response to such therapy can be monitored with fluorescein
angiography and OCT imaging.
Techniques for ERM surgery include vitrectomy (general principles
addressed in Chapter 61) with posterior hyaloid and membrane removal. The
clear vitreous is readily visualized when marked with triamcinolone. This is
quite useful before separating the posterior hyaloid in the posterior pole by
engaging the vitreous and hyaloid using suction from the vitreous cutter,
grasping with forceps, or snagging with the rough flexible loop (Finesse loop,
Alcon). Recognizing the involvement of condensed posterior hyaloid and
often dense adherence of the vitreous to a pediatric ERM, the epiretinal tissue
may peel from the retina during elevation of the hyaloid, or the hyaloid may
be separated along with removal of a broad sheet of ERM. This step should
be managed to control the direction of the traction relative to the retinal
surface to avoid retinal damage. Thus, membrane and hyaloid removal is
generally from posterior to outside of arcades and with traction at an oblique
angle rather than perpendicular to the retinal surface.
Membrane peeling may be initiated with a barbed needle (e.g., a 27-
gauge needle pressed gently at an oblique angle against a flat metal surface to
create a fine barb), which is often effective to engage and separate smooth
adherent macular membranes in children. The pinch-and-peel technique with
forceps or scratching (with the rough flexible Finesse loop) techniques is also
useful at sites of irregular ERM attachment to initiate the peel. As with adult
surgery, attention is given to limit pressure against the retina and to determine
the extent of the peel to avoid compressing the retina or engaging tissue of
the retinal nerve fiber layer, which may appear whitened in the area of
membrane removal. Focal firm adhesions are more common between an
ERM and retinal surface in children, often over retinal vessels (15,16). Thus,
it is not uncommon in pediatric ERM surgery to both peel epiretinal tissue
and to cut the membrane at a very adherent site, rather than risk tearing the
retina (22).
OCT imaging has been useful to visualize such tissue relationships before
and during surgery (5,7). Recurrence of an ERM in children has been noted
in multiple cases; however, as shown with preoperative OCT imaging, this
may also occur due to removal of a single layer of gliotic tissue in an
unrecognized multilayered membrane (5). Intraoperative OCT is useful to
determine the location and extent of residual ERM and scrolled ILM after
peeling (Figure 67-2). Some surgeons have advocated internal limiting
membrane (ILM) staining to guide ILM peeling in pediatric cases with a goal
of diminishing recurrences (Figure 67-1). The peripheral retina is carefully
inspected; however, peripheral vitreous removal is not pursued unless this is
required to release traction from associated pathology. The surgeon thus
avoids working in close proximity to the crystalline lens.
Surgical considerations and preoperative evaluation of CHRRPE may be
more complex than in idiopathic pediatric ERM, and the severity of the
retinal-RPE involvement may limit visual recovery. CHRRPE has the added
complexity of the unknown extent and depth of involvement, including outer
retinal, vascular, and retinal pigment epithelial components. CHRRPE lesions
are more likely to have firm ERM–retinal attachments and may require sharp
dissection to cut the attachment (15,22). Despite this uncertainty,
histopathology of surgically removed ERM in CHRRPE has not shown
retinal components within the membrane (44).
POSTOPERATIVE CARE
Postoperative considerations include managing intraocular pressure and
monitoring for response of macular morphology and for retinal detachment or
bleeding. In children, ocular ultrasound, Optos wide-field imaging, and OCT
imaging are often tolerated better in the clinic than extensive indirect
ophthalmoscopy and thus provide valuable postoperative information. Other
pediatric eye care team members are important in the postoperative period to
manage refractive status, assist in the need for eye protection, and evaluate
and treat amblyopia in younger children.
COMPLICATIONS
Complications have included retinal breaks, vitreous hemorrhage, retinal
detachment, macular edema, transient and permanent elevated intraocular
pressure, and cataract; and ERM recurrence rates have varied widely
(4,8,14–16,22,38). Improved technology, such as OCT imaging for younger
children, has improved our ability to identify, evaluate and monitor the
progression of ERMs in these patients. Intraoperative OCT enables the
surgeon to assess the retinal surface once membrane removal is thought to be
complete and may reveal sites of subclinical persistent membrane or retinal
injury (5,45).
To date, surgical guidance for pediatric ERMs is limited as reports are from
small series of cases with multiple etiologies including CHRRPE and from
children of widely varying ages (from 4 months to adulthood)
(4,7,8,14,15,46–48). Surgical interventions in appropriate cases with
decreased visual acuity have resulted in improved visual acuity in the
majority of patients, although many of these studies include not only
idiopathic ERM but also a range of underlying etiologies (1,7,8,15,16).
Visual potential may be limited due to chronicity of a lesion in early
childhood; nevertheless, numerous reports of pediatric ERM cases and series
have demonstrated improvement in visual acuity, which may take over 6
months after surgical ERM removal (4,7,8,14,15,46–48). There are also many
reports of visual acuity improvement after the vitrectomy and ERM removal
in CHRRPE (4,8,15,22,44,46,47). Although the recurrence of ERM may be
more common with CHRRPE than in other ERMs, visual acuity may still
improve (8,46).
SUMMARY
Pediatric ERMs or CHRRPE lesions are often unilateral and may be observed
when they do not involve the macula or cause vision loss. Neurofibromatosis
type 2 may be associated with either lesion. When the macula is involved,
surgical assessment and perioperative planning should consider the age and
visual development of the child. Surgeons should be prepared for
vitreoretinal attachments that may require sharp dissection. OCT and
fluorescein angiographic imaging are useful in the diagnosis, therapeutic
planning, and monitoring of these conditions.
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68
Surgery for Gene Therapy
Jonathan F. Russell, Albert M. Maguire, and Stephen R. Russell
INTRODUCTION
Gene therapy has been broadly defined as “a pharmacological approach
whereby exogenous genes or nucleic acids are introduced into host cells to
effect local production of a therapeutic protein or halt expression of a disease-
associated protein” (1). A more strict definition is “a treatment method [that]
involves replacing, inactivating, or editing diseased genes” (2). For
autosomal recessive, loss-of-function, disease-causing mutations, adding to
the cells one or more functional copies of the mutated gene, a strategy known
as gene augmentation, holds great promise. For example, in Leber congenital
amaurosis (LCA) associated with biallelic RPE65 mutations, gene
augmentation of the mutated RPE65 retinal trans-isomerase that normally
resides within the retinal pigment epithelium (RPE) has been shown to
functionally restore recycling of retinal vitamin A (3). The first human gene
therapy clinical trials began in 1981; since then, nearly 2,000 test cases have
been performed. In December 2017, a successful phase 3 clinical trial of
voretigene neparvovec-rzyl (VN, Luxturna, Spark Therapeutics, Inc.) for
LCA due to biallelic RPE65 mutations culminated in the first U.S. Federal
Drug Administration (FDA) approval of a gene therapy for an inherited
genetic disease (4,5).
SURGICAL APPROACHES FOR

POSTERIOR SEGMENT GENE
THERAPY
We will detail three gene therapy approaches for inherited retinal diseases:
transvitreal subretinal delivery, transchoroidal subretinal delivery, and
intravitreal delivery. Preclinical testing has demonstrated that direct contact
with the transfection target (in the case of RPE65-associated LCA, this is the
RPE apical surface) maximizes transfection rates for adenovirus-associated
virus (AAV)-based agents (6). For these reasons, when targeting the outer
retina or RPE, a transvitreal approach with transretinal injection into the
subretinal space, heretofore referred to as the transvitreal subretinal approach,
remains the current preferred and only FDA-approved approach. This
approach also has a well-known and acceptable adverse event profile. The
transvitreal subretinal approach has been evaluated using both AAV and
lentiviral vectors in humans in multiple RPE65-associated LCA trial groups
and in trials for choroideremia (7), achromatopsia (8), Usher syndrome type
1B (9), Stargardt disease (10), and advanced exudative age-related macular
degeneration (10,11).
Alternative surgical approaches to gene therapy include transchoroidal
subretinal delivery, which avoids vitrectomy and may avoid a retinotomy.
However, current transchoroidal methodology has been associated with high
complication rates (12). Another alternative approach is intravitreal delivery,
which may be advantageous especially when targeting inner retinal cells such
as retinal ganglion cells, administering small intranuclear interfering
oligonucleotides, or performing gene augmentation of a secreted protein such
as sFlt gene augmentation–related VEGF suppression (13). However, as of
2019 there are no FDA-approved therapies utilizing transchoroidal or
intravitreal delivery.
TRANSVITREAL SUBRETINAL
DELIVERY
Transvitreal approaches to the subretinal space were first developed in the
1990s for removal of subfoveal choroidal neovascularization (14,15) and for
subretinal injection of tissue plasminogen activator to treat massive macular
subretinal hemorrhage (16). Transvitreal subretinal delivery for gene therapy
was developed in animals, pioneered in adults, and then adapted to children.
In pediatric patients, a number of special considerations apply. Pediatric
patients differ from adults in the dosage of immunomodulation, perioperative
management, and surgical maneuvers.
Basic Considerations for Gene Therapy

Potential candidates for gene therapy must first be rigorously evaluated for
eligibility for treatment. Since neparvovec-rzyl (VN) gene therapy is the only
FDA-approved treatment currently, it will be used as an example in this
chapter. Candidates for voretigene neparvovec-rzyl (VN) gene therapy must
have documented biallelic causative RPE65 mutations, and must possess
sufficient viable retinal/RPE cells to provide a likely target for gene
transfection and augmentation to occur. To meet the first criterion, candidates
must undergo genetic testing of the RPE65 locus that demonstrates two
disease-causing mutations or homozygosity of the same mutation. To confirm
that the mutations are on separate alleles, a first-degree relative, preferably
one who is unaffected, should also be tested to demonstrate independent
assortment of the mutations (i.e., proving that each mutation is on a separate
allele, or the candidate is “biallelic”). Testing of the second family member or
retesting a homozygous patient is referred to as two-sample confirmation. In
addition to confirming biallelic mutations, two-sample confirmation assures
that sample switching or mislabeling did not occur on initial patient testing.
For these reasons and because it provides a measure of duplication, two-
sample confirmation is superior to whole genome sequencing even with long
reading frames.
Determination of the presence of “sufficient viable retinal/RPE cells” is a
clinical decision based upon the presence of detectable outer retinal layers or
outer retinal layer function. Structurally this is best determined via macular
optical coherence tomography (OCT), but it can be inferred by the presence
of some intact visual field by confrontation, standard perimetry, or
microperimetry in a child old enough to perform a visual field test (Figure
68-1). The pivotal RPE65-associated LCA phase 3 clinical trial mandated a
central macular thickness (CMT) on macular OCT of at least 100 microns for
inclusion (5), which may differ from and is generally larger than the center
point thickness.
FIGURE 68-1 Representative preoperative color
photographs and OCT of a 5-year-old girl with biallelic
heterozygous mutations in RPE65 causing Leber
congenital amaurosis. Preferential subretinal injection sites
for voretigene neparvovec-rzyl are denoted by yellow
ovals for the right (A) and left (B) eyes. Note the central
choroidal preservation on color photos (A, B) and outer
retinal thinning (yellow arrows) with central islands of
outer retinal preservation on OCT (C, D).
Assessment of Benefits, Risks, and Potential

Complications
Once basic eligibility as described above has been determined by the surgeon
or surgical team, which may include an inherited retinal disease specialist
and/or genetic counselor, the risks versus benefit balance for the patient
should be qualitatively assessed before proceeding. For example, regarding
benefit of VN, the phase 3 trial demonstrated functional visual improvement
in the majority of treated patients (27 of 29, or 93%) (5). Patients with visual
acuity or visual field limited to an extent that they were not ambulatory, i.e.,
could not pass the mobility test described below at a light level of 400 Lux,
were excluded from the phase 3 study. Therefore, it is not known whether
such patients will benefit from VN gene therapy, and the potential for no
improvement should be discussed with the patient and parents of the pediatric
patient. Regarding risks, the treatment profile to date for VN appears largely
or entirely related to the risks of vitrectomy, iatrogenic posterior vitreous
detachment (PVD), and subretinal injection (see Table 68-1). Although the
study allowed treatment of children as young as age 3, the youngest recruited
subjects that could meet the entry criteria, which included passing the
mobility course, were 4 years old. Thus, there is no safety information to
assist the surgeon’s assessment of risk below age 4. Based upon the
developmental globe size and formative pars plana in the youngest
candidates, a higher frequency of vitrectomy-related complications could be
anticipated below a developmental age of 3 or so (17). This should be
considered since the FDA has approved VN for use in children of at least 1
year of age.
TABLE 68-1 Ocular adverse reactions following

treatment with voretigene neparvovec-rzyl in
patients with RPE65-associated Leber congenital
amaurosis
These data include subjects treated in both the phase 1/2 and phase 3 trials (3–5).
aTransient appearance of asymptomatic subretinal precipitates inferior to the retinal injection site 1–6
days after injection.
Immunosuppression is recommended by the manufacturer based upon its

application in the phase 3 trial and in other trials utilizing AAV vectors. The
purpose of perioperative immunosuppression is to minimize sensitization and
reduce potential for vector inactivation. The recommended regimen is to
begin prednisone or prednisolone at 1 mg/kg/d (up to a maximum of 40
mg/day) given as a single oral dose beginning 3 days prior to surgery, on the
day of surgery, and for 3 days thereafter. The dose is then reduced to 0.5
mg/kg/d for 5 days, followed by 0.5 mg/kg/d on alternate days for the next 5
days. For older children capable of swallowing tablets, prednisone tablets
may be prescribed. However, for younger children prednisone (5 mg/5 mL)
or prednisolone (5 mg/5 mL) syrup may be used. Prednisolone syrup
(Orapred, Concordia Pharmaceuticals Inc., Ontario, Canada) tastes better and
is preferred by most patients. Oral steroids are typically used in an
overlapping schedule, because treatment of the second eye with VN is
recommended within 6 to 18 days of the first eye. This interval between VN
treatments was chosen to minimize the likelihood of serial immunogenic
sensitization as T-cell sensitization peaks after 4 weeks (18). Superimposing
the immunosuppression schedule for the first eye on the schedule for the
second eye provides a comprehensive dosage schedule for the patient and
family. To avoid untoward cross reactivity, routine immunizations especially
using attenuated live viruses should be avoided during the perioperative
period. Immunosuppression should not be initiated if the child is clinically ill,
has unexplained fevers, or has any unstable medical conditions. Furthermore,
if the child has recently contracted an upper respiratory or other infection that
could theoretically complicate or cross-react with AAV, consideration should
be given to delay administration of VN.
Equipment for Transvitreal Subretinal Delivery

Sterile hood and trained, certified pharmacy personnel to handle the gene
therapy agent
Gene therapy agent
Small-gauge pars plana vitrectomy system
Transscleral cannula (nonvalved) through which the subretinal cannula will
fit
Tano scratcher or equivalent
Syringe, tubing, and fine cannula (e.g., 39 of 41 gauge)
Subretinal injection system (if approved/compatible for use with the gene
therapy agent)
Intraoperative OCT imaging (less useful for VN injection but may be
especially useful for choroideremia and other disorders with thin retinas and
choroidal layers)
Procedural Technique for Transvitreal Subretinal

Delivery
Specific details of operating room scheduling may need to be explicitly
affirmed in advance, because VN must be administered within 4 hours of
dilution and other gene therapies will likely require similar considerations.
Therefore, arrangements must be made to avoid significant delays,
overscheduling of emergency or unexpected cases, or cancellation of surgery.
Because of the short interval of known efficacy of the diluted VN and its high
cost, it is recommended that the frozen vial not be removed from the −80°F
(−65°C) freezer until the day of surgery when the child has been cleared for
surgery by the anesthesiologist. Preparation of the dilution requires a sterile
hood and trained and certified pharmacy personnel. The gene therapy agent
should be properly diluted and readily available to the surgeon prior to
initiating anesthesia.
Although vitrectomy and subretinal injection can be performed under
retrobulbar anesthesia with monitored anesthesia sedation in most adults,
pediatric patients, especially those with nystagmus, which is manifest in
nearly all LCA patients, will require general anesthesia. Following induction
with general anesthesia, either a laryngeal mask airway (LMA) or
endotracheal tube may be placed. Unless complicated, the procedure can be
completed within 60 to 75 minutes. Following antiseptic application of 5%
Betadine to lids, lashes, and conjunctiva, surgical draping is performed and
sclerotomies are placed in the superonasal, superotemporal, and
inferotemporal quadrants. The infusion line is connected to the
inferotemporal infusion cannula at a pressure of 30 mm Hg or higher. The
phase 3 trial for VN was performed utilizing 20-gauge instrumentation to
remain consistent with prior preclinical studies. In 2018, the majority of
surgeons performing pediatric vitrectomies for indications other than
retinopathy of prematurity use 23-, 25-, or 27-gauge cannula systems. For
VN treatments, each surgeon is encouraged to use whichever 23- or 25-gauge
system he/she prefers. However, the 23- or 25-gauge cannula system should
be checked prior to surgery to ensure that the subretinal injection cannula,
which has an outer diameter of 25 gauge, can easily pass through the
sclerotomy cannulas. As an additional consideration, valved sclerotomy
cannulas are generally discouraged for the injecting port because the fragile
25 (outer gauge) to 39 (inner gauge) or 25- to 41-gauge polyamide injection
cannula tip can be easily damaged upon insertion through a valve. Bending
the cannula tip beyond 45 degrees or so can result in luminal closure and
malfunction during attempted bleb formation.
Because all sclerotomies should be sutured closed in children, performing
a conjunctival fornix–based peritomy facilitates closure. Sterile calipers are
then used to mark the cannula sites, at an appropriate distance from the
limbus based upon age for infants and toddlers (see Table 68-2), and at 3 mm
(aphakic), 3.5 mm (pseudophakic), or 4 mm (phakic) eyes of older children
or adults.
TABLE 68-2 Recommended sclerotomy position

by developmental age
Adapted from Lemley CA, Han DP. An age-based method for planning sclerotomy placement in
pediatric vitrectomy: a 12-year experience. Retina 2007;27(7):974–977.
Small children have high scleral pliability such that any interruption of
infusion increases the risk of globe collapse, catastrophic retinal damage, and
retinal tears. An intraoperative retinal tear in an adult may be managed by the
addition of laser demarcation but in a young child represents potentially a
vision- or even organ-threatening complication. When performing transvitreal
subretinal gene therapy, it is best to avoid any unnecessary or prolonged
vitreous manipulations such as vitreous base shaving. The goal of vitrectomy
is only to create a vitreous-free pathway for safe and reliable delivery of the
vector to the subretinal space. To ensure this is accomplished, an iatrogenic
PVD is completed beyond the site of the anticipated macular injection sites.
In contrast to normal pediatric vitreoretinal adhesion, in RPE65-associated
LCA, the vitreoretinal adhesion is remarkably weak. An iatrogenic PVD can
typically be induced with a silicone-tipped cannula or vitrectomy cutter. The
operating surgeon should then assemble the injection apparatus following the
Luxturna package insert which currently includes a pharmacy-supplied 1 mL
Becton Dickinson syringe containing 0.8 mL of diluted VN, a VN-verified
compatible 6-inch extension tubing with small 0.6- to 0.8-mm lumen, a 39- or
41-gauge injection cannula, and a list of compatibility-tested injection system
components in the VN package insert (19). VN-verified compatibility has
been performed by the manufacturer to assure minimal luminal VN
adsorption and biocompatibility. If alternative components are substituted,
compatibility cannot be assumed. In other trials, injection systems such as the
MedOne injection system (MedOne Microdose Injection Kit, MedOne
Surgical Inc.) have been used, but no safety or comparative studies are
available.
The initial site of subretinal injection is chosen based on the disease and
the preferred location for vector delivery. Dog and primate studies have
demonstrated that RPE transfection with AAV-based vectors only occurs
within the region of exposed apical RPE over the region of a subretinal bleb
(20). Because RPE65-associated LCA is a rod-mediated disease and evidence
suggests that cones may possess a supplemental mechanism for all-trans
retinal trans-isomerization (21), it is unclear whether visual benefit is
conferred with detachment of the fovea. Thus, in RPE65-associated LCA, the
initial site of subretinal delivery is typically chosen along or just outside the
superior arcade (Figures 68-1 and 68-2) in order to treat the central and
inferior macular field needed for ambulation and most near visual tasks. In
addition, this region is thicker than more peripheral retina or the central fovea
and is a good compromise. The closer the injection site is to the fovea, the
greater the risk of inducing a full-thickness macular hole. The further the
injection site is from the fovea, the less control one has over the direction of
subretinal dissection, because the lowest resistance pathway of separation
will determine the direction that the injected subretinal fluid spreads, and this
may not be toward the macula. Generally, the injection site should be no
closer than two disk diameters from the fovea. Occasionally the injected
subretinal fluid will dissect around the disk rather than extend to the macula.
Just prior to introducing the injection cannula or prior to injection, the
intraocular pressure should be reduced to 10 mm Hg to increase the pressure
differential between the injection pressure and the subretinal potential space.
FIGURE 68-2 Transvitreal subretinal injection procedure
for voretigene neparvovec-rzyl: choice of injection site.
Note that this is the surgeon’s view. (Image reproduced
with permission from Spark Therapeutics, Inc.)
The preferred method used to inject into the subretinal space and raise a
retinal bleb is to advance the 39- or 41-gauge injection cannula until the
retina is gently indented (Figure 68-3). Then, the injection elevates the retina
around the statically held cannula (Figures 68-4 and 68-5). It is crucial that
the resulting retinotomy size be minimized to prevent loss of vector into the
vitreous cavity, which will also reduce bleb pressure and/or reduce bleb
enlargement. Since ocular inflammation appears to be related to exposure of
vector capsid to the anterior vitreous cavity structures such as the ciliary body
and iris (22), vitreous exposure should be minimized. Following slow
subretinal injection of 300 μL of VN solution, the injection pressure should
be maintained for at least 5 seconds to allow the bleb pressure to equilibrate
with that in the syringe prior to removing the cannula from the retinotomy. If
the surgeon is unable to complete subretinal injection of 300 μL, the assistant
reports high resistance to injection. If the subretinal elevation extends in an
undesirable direction, alternative additional bleb sites can be chosen and
injected up to the dosage limit total of 300 μL. As with all injection site
choices, the surgeon should be cognizant of the retinal thickness and the
proximity to the fovea at the chosen site to minimize surgical complications.
Injection by an assistant is useful to provide interactive verbal
communication to the operating surgeon that will allow enhanced
troubleshooting of injection difficulties. Maintaining stability of the small
cannula within the subretinal space without stretching the retinotomy is a
remarkably difficult challenge even for experienced surgeons. Thus,
delegating the monitoring of injection resistance and injection volume is
recommended. In the phase 3 VN study, no effort was made to create a “pre”-
bleb by injecting subretinal saline to create a subretinal pocket into which VN
could be injected. However, in other ocular gene therapy trials for
choroideremia, achromatopsia, and others, “pre”-blebs have been utilized
(7,23).
for voretigene neparvovec-rzyl: position of subretinal
injection cannula. Note the placement of fine silicone
cannula against inner retina. (Image reproduced with
permission from Spark Therapeutics, Inc.)

for voretigene neparvovec-rzyl: assembly and positioning
of injection apparatus. See text for details. (Image
reproduced with permission from Spark Therapeutics,
Inc.)
for voretigene neparvovec-rzyl: formation of subretinal
bleb. See text for details. (Image reproduced with
permission from Spark Therapeutics, Inc.)
Within the phase 1 (3), phase 1 follow on (24), and phase 3 VN studies (5),
no dyes or vitreous marking agents such as indocyanine green or
triamcinolone acetonide (Kenalog) were used or recommended. To date, no
stains or marking agents have been tested for biocompatibility with VN, and
hence the potential effects on efficacy are not known. Therefore, they are
generally avoided unless required for a compelling reason. For postmarketing
VN treatments, vitreous marking agents have been reported to be compatible
by some surgeons (25,26).
During the VN phase 3 trial, intraoperative OCT was unavailable.
However, since this study was completed, a number of investigators have
reported that intraoperative OCT may be especially useful when inherited
retinal diseases, such as choroideremia, have led to pronounced retinal and/or
choroidal thinning. In these situations, it may be difficult or impossible for
the surgeon to intraoperatively distinguish subretinal from sub-RPE or
suprachoroidal injection (23). In studies of transvitreal subretinal bleb
formation in model eyes, OCT has been shown to provide a more accurate
assessment of subretinal volume delivery compared to the surgeon (27).
Because central retinal thickness is generally preserved for RPE65-associated
LCA, intraoperative OCT is not necessary for VN delivery. Several surgeons
have noted that intraoperative OCT may facilitate identification of foveal
bulging that precedes development of an intraoperative full-thickness macular
hole or macular dehiscence, thereby warning the surgeon to slow the rate of
injection or change to an alternative retinal injection site.
After subretinal injection, a thorough peripheral retinal examination using
a wide-field inverting lens or indirect ophthalmoscopy with scleral
indentation is then performed to search for any evidence of retinal tear or
dialysis. If retinal defects are found, they are treated promptly using standard
cryotherapy or retinopexy techniques. An air–fluid exchange is then
performed, both to remove the majority of vitreous remnants of viral
genomes to lower the risk of inflammation and sensitization and in an effort
to “spread” the bleb to cover more surface area. All efforts should be made to
assure retention of the maximum dose of subretinal VN, so aspiration during
the air–fluid exchange should not be near the retinal injection site(s). All
sclerotomies should be sutured closed and the overlying conjunctiva sutured
to cover the sclerotomies.
Postoperative Care
Postoperatively patients should remain supine for at least 4 hours to minimize
exposure of viral capsid to anterior vitreous structures and to encourage bleb
“spread.”
Complications
As noted above, the safety profile of VN is favorable, and appears to be
largely or entirely related to the standard risks of pars plana vitrectomy,
iatrogenic PVD, and subretinal injection (Table 68-1). Ocular inflammation
appears to be minimal with systemic immunosuppression, and full-thickness
macular holes are uncommon when the surgical technique described above is
utilized. Other potential complications from transvitreal subretinal gene
delivery include chemosis, corneal abrasion, vitreous hemorrhage,
rhegmatogenous retinal detachment, inadvertent subretinal injection of air
bubble, sensitization to the vector, or endophthalmitis. For gene therapies, in
general, there is potential risk of off-target mutagenesis which may vary by
vector.
One of the most challenging aspects of gene therapy trials has been
demonstrating clinical benefit. In the VN phase 3 trial, two types of
assessments were performed (5). The first type was conventional in-office
testing, otherwise known as visual function testing, including visual acuity;
OCT; Goldmann visual field; red, blue, and white full-field light sensitivity
(FST); and pupillometry. The second type of assessment was functional
visual testing, which attempts to simulate and measure performance of a
function of daily living that requires an integration of visual abilities. The
latter was the favored assessment by the FDA and was the primary endpoint
of the trial. The functional visual test that was used was the multiluminance
mobility test (MLMT), which is an obstacle course performed under
controlled lighting and timed conditions. The trial demonstrated marked
improvement in the MLMT in those patients treated with VN (5). As of 2018,
the resulting improvements have persisted at least 4 years after treatment (4).
In the nontrial clinical environment, functional visual testing remains too
time consuming for routine clinical use. Simpler, widely available visual
function tests include visual acuity, visual fields, and FST. Because of
concerns on the part of payers regarding the degree of effectiveness and
durability of treatment with VN, the manufacturer of VN (Spark
Therapeutics, Inc., Philadelphia, PA) has negotiated contracts with some
providers to reimburse payers for the cost of VN in the event that a treated
patient were to not reach a prespecified performance threshold. For most
contracts, two thresholds have been agreed to, an early postoperative measure
to prove effectiveness and a later measure to assure durability. The threshold
is an improvement of FST by 0.3 log units from baseline measured at 60 to
90 days following treatment that remains improved at 3 years (Figure 68-6).
If such a payer provision is in place for an individual patient, the surgeon
should perform baseline testing or, for uncooperative younger patients,
document the inability to perform FST and perform any additional supportive
baseline testing available.
FIGURE 68-6 Full-field light sensitivity outcomes in

phase 3 clinical trial of voretigene neparvovec-rzyl. The
chart depicts changes in FST from baseline (open circles)
to 1-year follow-up (closed circles), for those randomized
to initial intervention (green) and to delayed
intervention/controls (blue). Note the large increases in
FST in the majority of treated patients. In the 4-year-old
patient (far left), there was no change in FST from
baseline to follow-up.
INTRAVITREAL DELIVERY
Basic Considerations for Intravitreal Delivery
Intravitreal delivery is an appealing route for delivery of gene- or cell-based
therapies, and a number of early and late phase clinical trials of gene therapy
utilize intravitreal injection rather than transvitreal subretinal injection
(28–30). Intravitreal delivery reduces the risk of procedure-related
complications and may be preferred when administering a small diffusible
molecular agent, including a gene or RNA-interfering oligonucleotide, when
multiple administrations are needed or when inner retinal delivery is
sufficient to address the disease mechanism. Compared to subretinal injection
using either transvitreal or transchoroidal approaches, intravitreal injections
are quick, inexpensive, easily repeatable, lower risk, and commonly
performed and are usually done with topical or subconjunctival anesthesia
(except in children, see below). As opposed to localized distribution within or
near a subretinal bleb, intravitreal injection may achieve distribution of the
therapeutic agent across the entire retinal surface. Also, the vitreous gel likely
serves as a reservoir for sustained release of therapeutic agents, since
intravitreally injected medications have shorter half-lives and more rapid
clearance in vitrectomized eyes (31).
However, there are some potential drawbacks to intravitreal delivery.
Distribution across the entire retinal surface may not be necessary if the
disease pathology is relatively localized. For example, retinal disorders
primarily affecting cones may be best served by restricting therapeutic
delivery to the macula where cones are most dense (32). Similarly, diffuse
distribution may be irrelevant depending on the stage of disease and its
temporal–spatial progression. Advanced retinitis pigmentosa with only
central macular sparing would likely benefit little from pan-retinal
therapeutic distribution, and efforts would be better spent preserving the
remaining viable macula. Even the potential of utilizing the vitreous gel for
sustained release may actually be disadvantageous for gene therapy, because
it is possible that sustained release could exacerbate neutralizing immune
responses and hence limit transduction of target cells.
Agents dispersed into the vitreous chamber may quickly come into
contact with the ciliary body, lens capsule, zonules, iris, and in fact the entire
anterior segment (22). In contrast, subretinal delivery is thought to restrict the
agent to contacting only retina or RPE; there is the theoretical concern that
agents could reflux out of the retinotomy site (33), but in our experience, with
proper transvitreal surgical technique (see above), reflux does not occur or is
minimal. The widespread ocular distribution resulting from intravitreal
injection can result in persistent transgene expression in anterior segment
tissues (22). Expression in nontarget tissues carries the theoretical risk of off-
target mutagenesis or other unknown deleterious effects, and it is likely
related to the strong immune responses that are induced by intravitreal
injections but less so by subretinal injection (22,34,35). Increased immune
responses to intravitreal injections may limit the efficacy of intravitreal gene
therapy, particularly when a patient has preexisting neutralizing antibodies to
adenovirus-associated vector (AAV) (36). In contrast, even preexisting
neutralizing antibodies to AAV present in intraocular fluids do not limit the
expression of AAV if the vectors are injected subretinally (37).
Perhaps the greatest disadvantages of intravitreal delivery vis-à-vis
subretinal delivery are lower transduction efficiency and a diminished ability
to transduce cells of the outer retina and RPE (38,39). Lower transduction
efficiency necessitates higher doses, which are in turn prone to inducing
stronger neutralizing immune responses. Transduction of the outer retina and
RPE after intravitreal injection is thought to be anatomically constrained by
the internal limiting membrane (ILM) (39,40). Similarly, the ILM may serve
as a barrier against the spread of agents from the subretinal space into the
vitreous and ocular fluids. A number of approaches have been tested in
preclinical studies to facilitate intravitreal transduction of outer retinal cells,
including viral vector modification/mutation (41–45), laser photocoagulation
of the ILM (46), surgical ILM peeling (47), and enzymatic lysis (40). One
group found that vitreous aspiration followed by intravitreal injection resulted
in pan-retinal, full-thickness transduction of AAV2/8 in mice (48). If vitreous
aspiration improves transduction, perhaps pars plana vitrectomy with or
without ILM peel could achieve diffuse retinal transduction, but this is
conjecture. In some inherited retinal diseases, such as juvenile X-linked
retinoschisis, the ILM is thought to be pathologically porous, which may
enable viral vector penetration without any adjunctive measures (29). It is
unclear if this is true for other diseases.
Intravitreal delivery was first performed in 1911 with the injection of air
into the vitreous chamber for pneumatic retinopexy to treat rhegmatogenous
retinal detachments (49). Intravitreal delivery of medications began with
penicillin for endophthalmitis (50). With the advent of anti–vascular
endothelial growth factor (VEGF) agents for a variety of indications
including neovascular age-related macular degeneration (51) and diabetic
macular edema (52), intravitreal injections have become the most commonly
performed procedure in ophthalmology (53). The technique is also being
tested and used in preterm infants for the treatment of retinopathy of
prematurity with anti-VEGF therapy (54). Performing intravitreal injections
for gene therapy in children, especially in those with congenital nystagmus
such as in trials for CEP290-associated LCA (28), impose an increased level
of complexity, as all but the most mature children will require general
anesthesia, albeit brief, because of the need for compliance and suppression
of nystagmus.
Equipment for Intravitreal Delivery

therapy agent
Gene therapy agent
Intravitreal injection system approved for use with the gene therapy: syringe
and needle
Procedural Technique for Intravitreal Delivery

In older infants and children, conscious sedation or general anesthesia may be
required, and occasionally in adults with severe nystagmus. In our
experience, children are unlikely to tolerate in-office intravitreal injection
until the teenage years. As the field progresses, ideally children with inherited
retinal diseases will be identified in infancy and promptly treated in order to
minimize the extent of retinal degeneration. If so, in the future, it may be
possible to administer gene therapy intravitreally in small infants without
general anesthesia as is currently done with intravitreal anti-VEGF injections
for some neonates with retinopathy of prematurity. However, many children
with inherited retinal disease have nystagmus, which may necessitate
systemic anesthesia for safety of intravitreal injection. For intravitreal
injection, only a few minutes of systemic anesthesia is required, whereas a
pars plana vitrectomy for subretinal injection requires around one hour; there
is concern that prolonged general anesthesia in infants can deleteriously
affect the developing central nervous system (55).
The procedural details for intravitreal injection have been reviewed in
many excellent reviews and instructional videos (54,56–58). We briefly
summarize our technique in the anesthetized pediatric patient (Figure 68-7).
First, informed consent is obtained and documented. This must include a
thorough discussion of the potential benefits and risks of intravitreal gene
therapy, which are largely unknown as of 2019. The eyelids and lashes are
sequentially swabbed with povidone–iodine (PI) swabs. A sterile lid
speculum is inserted using a sterile cotton swab. Then a sterile cotton swab
soaked in 5% PI solution is applied with moderate pressure 1 to 3 mm from
the inferotemporal limbus for approximately 10 seconds. A drop of 5% PI is
applied to the indented site. Sterile calipers are used to mark the injection site
within the area of indented, anesthetized sclera, at an appropriate distance
from the limbus based upon age for infants and toddlers (see Table 68-2), and
at 3 mm (aphakic), 3.5 mm (pseudophakic), or 4 mm (phakic) eyes of older
children or adults. Another drop of 5% PI is applied to the marked site. Then
the medication is injected using a 30- or 32-gauge needle and syringe
approved for that vector or agent, oriented orthogonal to the scleral surface
without a scleral tunnel. A sterile cotton swab is immediately applied to the
injection site after needle removal and held in place for approximately 10
seconds to minimize reflux. Perfusion of the central retinal artery should be
ophthalmoscopically confirmed or the intraocular pressure should be
monitored until within an acceptable range. If the central retinal artery is not
perfused or the intraocular pressure fails to spontaneously fall within 1 to 3
minutes, a drop of 5% PI is applied and an anterior chamber paracentesis is
immediately performed (in our experience, this is rarely necessary). The
speculum is then removed, and the ocular surface and periorbita are rinsed
with sterile saline and wiped with a sterile gauze pad.
FIGURE 68-7 Intravitreal approach to gene therapy. See
text for details. As noted in the text, the surgeon must
adjust the entry site based upon age/development of the
eye (Table 68-2). (From Ochakovski GA, Bartz-Schmidt
KU, Fischer MD. Retinal gene therapy: surgical vector
delivery in the translation to clinical trials. Front Neurosci
2017;11:174. doi: 10.3389/fnins.2017.00174. Copyright ©
2017 Ochakovski, Bartz-Schmidt and Fischer.
https://creativecommons.org/licenses/by/4.0/.)
Postoperative Care
We do not routinely prescribe antibiotics after injection. The parents are
counseled that they may apply sterile artificial tears for a few days if needed
for foreign-body sensation, that the child should avoid eye rubbing for one
day, and to call or return immediately upon severe pain, nausea and vomiting,
or significantly decreased vision.
Complications
Intravitreal injection is a familiar and low-risk procedure. Potential adverse
effects include injection site discomfort, subconjunctival hemorrhage,
chemosis, corneal abrasion from speculum insertion/disinsertion, cataract,
zonular instability, transient or sustained increase in intraocular pressure,
vitreous hemorrhage, rhegmatogenous retinal detachment, and
endophthalmitis. The same set of adverse effects can result from pars plana
vitrectomy with subretinal injection, although cataract is more common after
pars plana vitrectomy (59) and endophthalmitis is probably more common
after intravitreal injection (60). In children especially, intravitreal delivery is
likely safer and easier than transvitreal subretinal delivery, because surgical
induction of a PVD to access the subretinal space can be challenging in
children. PVD induction is relatively straightforward in children with RPE65-
associated LCA treated with VN (see above), presumably because of an
abnormal vitreoretinal interface, but this may not be the case for other
inherited retinal diseases. Also, many inherited retinal diseases feature
atrophic, fragile retinas, making surgical manipulation of the subretinal space
more prone to complications such as iatrogenic retinal breaks than would be
expected with intravitreal delivery.
TRANSCHOROIDAL SUBRETINAL
DELIVERY
Given the disadvantages of intravitreal delivery and surgical challenges of
transvitreal subretinal delivery, multiple investigators have pursued
transchoroidal subretinal delivery for gene and cell-based therapies. This
approach holds special appeal for therapeutic delivery in children because of
the technical challenges and complications inherent to pars plana vitrectomy
and PVD induction in infants (see above). Also, as for transvitreal subretinal
delivery, with transchoroidal delivery, the therapeutic agent can be placed in
direct apposition with the retina and RPE, which are typically the sites of
dysfunction. Presumably the immune responses to transchoroidal subretinal
delivery would be similarly mild like those induced by transvitreal subretinal
delivery, but this is unknown.
Equipment for Transchoroidal Subretinal Delivery

therapy agent
Gene therapy agent
Transscleral, transchoroidal subretinal injection system approved for use with
the agent
Intraoperative OCT imaging (may be helpful)
Procedural Technique for Transchoroidal Subretinal

Delivery
Transchoroidal subretinal delivery would require general anesthesia for use in
children. The delivery is performed by incising the sclera anteriorly, typically
near the equator. Then, the choroid and RPE are incised and a catheter is
passed in the subretinal space posteriorly as shown in mice (61) and in a
human study (12). Alternatively, the catheter is passed in the suprachoroidal
space posteriorly and once the catheter is in position, the choroid and RPE
are traversed with an extensible microneedle (porcine study) (62). In either
technique, the catheter is passed under ophthalmoscopic or microscopic
guidance until the tip is at the preferred location within or adjacent to the
macula, at which point the therapeutic agent is injected into the subretinal
space. Some authors first raise a subretinal bleb and then inject therapeutic
agent into the bleb (62), whereas others do not (61).
The number of published studies utilizing transchoroidal subretinal
delivery in humans is sparse. Ho et al. (12) performed transchoroidal
subretinal injection of human umbilical tissue–derived cells in eyes with
geographic atrophy from age-related macular degeneration. After sclerotomy
and choroidotomy, the authors created an anterior subretinal bleb and then
used sodium hyaluronate viscoelastic (Healon) and a laser diode–based
illuminated catheter tip to facilitate passage of the catheter posteriorly (12).
There was a 37% rate of retinal perforations and 17% rate of rhegmatogenous
retinal detachment, and the authors concluded that the surgical approach
needs modification (12). Another study of transchoroidal subretinal delivery
in pigs observed retinal perforations and subretinal hemorrhage in 15% and
4% of pig eyes, respectively (62). There is also concern that transchoroidal
subretinal delivery may be more prone to reflux of the therapeutic agent out
of the subretinal space and into the sub-RPE or suprachoroidal space, since
the same study in pig eyes observed partial collapse of the subretinal bleb in
22% of eyes (62). Therefore, current transchoroidal subretinal approaches
may be more prone to surgical complications than transvitreal subretinal
delivery methods. Since complications like retinal perforation can be sight
threatening in children, transchoroidal subretinal approaches will require
convincing safety data prior to their adoption for gene therapy in inherited
retinal diseases.
An alternative to transchoroidal subretinal delivery is suprachoroidal
delivery (Figure 68-8), in which therapeutic agents are injected into the
suprachoroidal space (63). A number of current clinical trials are
investigating suprachoroidal delivery of various pharmaceuticals (64). In one
preclinical study, suprachoroidal delivery of AAV vector achieved robust,
full-thickness retinal transduction in rabbits (65). If agents placed in the
suprachoroidal space diffuse across the choroidal circulation to the RPE and
retina (66), this approach has the advantage of obviating choroidotomy and
the attendant risk of retinal perforation; however, by introducing agents into
the choroidal circulation, this approach may be more prone to inducing strong
immune responses. Perhaps immune responses can be avoided by
suprachoroidal delivery of plasmid DNA or nanoparticles without an
encapsulating viral vector, followed by placement of electrical current to
promote transduction (67,68), an approach that remains experimental.
FIGURE 68-8 Transchoroidal subretinal approach to gene
therapy. This figure depicts an anterior sclerotomy and a
catheter in the suprachoroidal space injecting material that
could diffuse across the choroid and retinal pigment
epithelium into the subretinal space. As explained in the
text, alternative transchoroidal approaches to gene therapy
include either an anterior or posterior choroidotomy with
injection of material directly into the subretinal space.
(Image reproduced with permission from Stout JT, Francis
PJ. Surgical approaches to gene and stem cell therapy for
retinal disease. Hum Gene Ther 2011;22(5):531–535. doi:
10.1089/hum.2011.060.)
FUTURE DIRECTIONS
Other potential approaches to therapeutic delivery for inherited retinal
diseases that also affect the central nervous system (e.g., neuronal ceroid
lipofuscinoses) are intracerebroventricular or intrathecal routes. Whiting et al.
(69) carried out intracerebroventricular gene replacement in a canine model
of CLN2, which delayed neurologic deterioration but seemed to have
minimal effect on retinal degeneration. Thus, effective treatment of the retinal
component of these disorders will likely require therapeutic delivery to the
intravitreal or subretinal space.
Much is unknown. Will the gains of VN and other gene therapies last a
lifetime? Could eyes be re-treated? Could eyes be treated in many more
retinal sites, or with diffuse subretinal delivery? Moving forward, for those
providing pediatric eye care and advice to families it will be imperative to
distinguish between FDA-sponsored trials and therapies, and rogue clinics
that are pursuing dubious therapies with sometimes devastating results (70).
Ultimately, it will probably be necessary to individualize delivery methods
for gene therapy in various disorders and even in individual patients based on
the primary site of retinal dysfunction, that is, inner versus outer retina,
central versus peripheral retina, integrity of the ILM, retinal fragility, and
other factors. Ideally, children with inherited retinal diseases will be
identified in infancy, receive a rapid genetic diagnosis, and undergo prompt
individualized gene therapy to arrest or reverse their retinal degeneration.
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69
Childhood Ocular Trauma
Mrinali P. Gupta, and Philip J. Ferrone
INTRODUCTION
Ocular trauma is a significant cause of retinal pathology among pediatric
patients. Moreover, pediatric patients in particular are at increased risk of
ocular trauma with approximately 34% of blinding eye injuries in the United
States Eye Injury Registry occurring in patients aged 0 to 19 years of age (1).
A thorough understanding of the retinal manifestations of ocular trauma and
of key differences in epidemiology, diagnosis, management, and prognosis in
children, as compared to adults, is essential to mitigate the risk of vision loss
in this population.
As with any case of eye trauma, there are individual aspects that make it
impossible to provide a definitive flowchart relevant to that patient. This is no
different for pediatric patients; however, there are considerations to be made
in the pediatric age group (ages 0 to 18 years) that include vulnerability of
children and infants and need to gently assess them; challenges obtaining
history in children; need to assure optimal visual outcomes by addressing
amblyopia; need to work in teams including pediatrics, pediatric
anesthesiology, and pediatric ophthalmology; and recognition and addressing
the fact that the eye continues to grow and that trauma and certain
procedures, such as scleral buckling, may impair that growth. This chapter
provides an overview of the epidemiology, evaluation, and management of
the pediatric ocular trauma patient.
STANDARD DEFINITIONS IN OCULAR

TRAUMA
Use of a standardized, unambiguous, common language is essential for
effective communication regarding the nature and extent of ocular injury both
for clinicians providing care for trauma patients as well as for research and
dissemination of knowledge. This chapter will employ the Birmingham Eye
Trauma Terminology System (BETTS), a standard nomenclature endorsed by
the International Society of Ocular Trauma, the American Academy of
Ophthalmology, the American Society of Ocular Trauma, the United States
Eye Injury Registry, and the World Eye Injury Registry. According to the
BETTS, eye wall is defined as the cornea and sclera. Ocular injury is divided
into “closed globe injury” and “open globe injury.” In a closed globe injury,
there is an absence of a full-thickness wound in the eye wall but involves a
partial-thickness wound (“lamellar laceration”) or no wound in the eye wall
(“contusion”) (Figure 69-1). A closed globe lamellar laceration is due to an
injury from a sharp object that results in a defect in the ocular tissues but does
not involve the entire depth of the eye wall—for example, a conjunctival
laceration or a partial-thickness corneal laceration. The mechanism of a
closed globe contusion injury involves transmission of kinetic energy from
the offending object to the eye (2).
FIGURE 69-1 Ocular trauma definitions. A flow chart

showing standard definitions for ocular trauma, according
to the Birmingham Eye Trauma Terminology System.
Open globe injury involves a full-thickness wound in the eye wall. If the
open globe injury is due to a blunt force, the open globe injury is referred to
as a “rupture.” The mechanism for a rupture is transmission of kinetic energy
from the offending object to the eye, resulting in a transient elevation of the
intraocular pressure, which exceeds the resistance or elasticity of the eye wall
and results in a full-thickness rupture. Open globe injuries generally occur in
the regions where the scleral is weakest—for example, concentric to the
limbus, behind the insertion of extraocular muscles, at the equator, and/or in
areas of prior injury or surgery (e.g., prior extracapsular cataract surgery
wounds). If the open globe injury is due to a sharp object, the injury is
referred to as a “laceration.” A laceration, in turn, may be “penetrating,” in
which there is an entry wound, “intraocular foreign body” (IOFB) in which
there is an entry wound and a retained IOFB, or “perforating,” in which there
is both an entry and an exit wound (Figure 69-1). A combination of injury
types may be present. For example, an explosion may result in several sharp
projectile bodies that cause penetrating injuries into the eye, but only some
may be retained in the globe as an IOFB. Moreover, some of the projectiles
may strike the eye without causing a full-thickness wound but inflict
contusive injuries to the eye (2).
The Ocular Trauma Classification Group has definitions for ocular
trauma that mirror those of the BETTS, except for the addition of a subgroup
that exists within the closed globe injury, called “superficial foreign body,” in
which there is neither a lamellar laceration nor blunt force contusion but
rather a foreign body that does not penetrate the eye wall. An example of this
is a corneal foreign body. In addition, the Ocular Trauma Classification
Group defined three distinct zones of open globe injury. Zone 1 injury
involves the cornea; zone 2 injury involves the region from the corneoscleral
junction (limbus) to 5 mm posterior to the limbus; and zone 3 injury involves
a wound posterior to zone 2. For closed globe injuries, zone 1 involves
external injuries limited to the bulbar conjunctiva, sclera, or cornea, including
conjunctival laceration or corneal abrasion; zone 2 involves injuries to
anterior segment structures up to the posterior lens capsule and includes the
pars plicata, including hyphema or traumatic cataract; and zone 3 injuries
involve the remaining posterior segment structures, including injuries to the
retina or choroid (3).
PREVALENCE OF CHILDHOOD
OCULAR TRAUMA
Much of the data regarding the incidence of serious ocular trauma among
pediatric patients is based upon evaluation of the records of hospitalized
patients. These studies suggest an incidence of 6.8 to 15.2 cases per 100,000
individuals per year (4–7). In the United States, the risk of hospital admission
from eye injury was 6.8 per 100,000 individuals per year for patients aged 18
years and younger (7). The true incidence, however, is likely significantly
higher, since the majority of ocular trauma does not require hospitalization.
Overall, ocular trauma afflicts male patients more than females, and the
disparity increases during the pediatric years and approaches a 4- to 6-fold
higher risk in males by young adulthood (1,4–8). The rate of injuries is higher
later in the pediatric years with a notable increase after age 10 (1,6,7).
Common causes of eye injury among pediatric patients include projectiles,
such as compressed air powered guns such as BB guns and paintball air guns,
fireworks, or thrown projectiles; sports-related injuries; airbag and seatbelt
injuries; bungee cord injuries; and inflicted injury, such as nonaccidental
trauma (see also Chapter 70) (9).
Pediatric open globe injuries are most prevalent between ages 7.7 and
11.6 years of age and exhibit a strong male predilection. In the United States,
pediatric open globe injuries most commonly occur at home and from sharp
objects, such as knives, whereas in developing countries, outdoor injuries are
more common. According to the United States Eye Injury Registry,
approximately 34% of blinding ocular injuries in the database occurred in
patients aged 0 to 19 years, which broke out as 12% in patients aged 0 to 9
years and 22% in patients aged 10 to 19 years. Visual outcomes in the
registry were better among pediatric patients with only 23.5% and 20.5% of
eye injuries in patients ages 0 to 9 years and 10 to 19 years with legal
blindness (<20/200) compared to 31% in patients over 20 years of age.
Among all patients surveyed, the rates of blindness from ocular injuries were
23.1% from contusion, 60% from ruptured globe, 23% from penetrating
trauma, 25% from IOFB, and 64% from perforating trauma (1).
CLINICAL EVALUATION OF THE

CHILDHOOD OCULAR TRAUMA
PATIENT
History
Obtaining an excellent history is central to appropriate diagnosis and
management of ocular trauma. History taking can be particularly challenging
in pediatric patients. Young children may not have the capability to recall or
recount what happened. Moreover, inability to distinguish between relevant
and irrelevant details of the history, being too scared or upset to participate in
history taking, and/or fear of punishment may limit the pediatric patient’s
ability to provide a history. Witnesses to the injury may likewise obfuscate
the history due to a desire to avoid personal responsibility.
In addition to obtaining a complete past medical history, past ocular
history, medication, and allergy history, the following are key questions to
address when obtaining the history of present illness:
When and at what time did the injury take place?

Where did the injury take place?
What happened?
What was the object that struck the eye? If the object can be reproduced,
it should be evaluated.
Was protective eyewear or eyeglasses being worn at the time of the
injury?
Did anyone else witness the injury? Such individuals may provide
useful, additional information.
What part(s) of the body were injured? Consideration should be given
that the other eye or nonocular structures may also be injured.
Questions regarding symptoms (ocular and nonocular): location, onset,
quality, duration, change over time, aggravating/alleviating factors,
associated symptoms.
What, if any, treatments were rendered already?
What was the nature and time of the patient’s last intake of fluids or
food? This will help determine surgical timing.
If relevant based upon the mechanism of injury, what is the patient’s
tetanus immunization status?
Ocular Examination in Acute Pediatric Eye Trauma

The physical examination should begin with a general assessment of the
patient’s overall medical health and well-being. Appropriate
nonophthalmologic care providers should be involved if there is multisystem
trauma. When an adequate examination cannot be achieved at the bedside, an
examination under anesthesia (EUA) should be considered. It is important
when approaching the child to be gentle, not to hurt or frighten the child, and
not to force an examination especially if it is clear an EUA will be needed to
obtain a complete evaluation or perform a possible procedure. Below is a
brief overview of considerations for evaluation and management of
nonretinal traumatic ocular injuries and the indications for surgery. A detailed
treatment of pediatric retinal trauma and its management follows.
External Evaluation
The periocular tissues should be evaluated for evidence of facial or orbital
trauma including evidence of foreign bodies, lacerations, fractures, or
periorbital or orbital swelling. The position of the globe should be assessed to
evaluate for exophthalmos, enophthalmos, hypoglobus, etc. Evidence of
orbital stepoffs, crepitus, or point tenderness to palpation may suggest an
orbital fracture. Ocular motility should be assessed to evaluate for limitations
in motility consistent with orbital pathology or entrapment of muscles in an
orbital fracture site. Evidence of orbital fracture with entrapment should
prompt an immediate oculoplastics evaluation for early intervention. The lids
should be evaluated for evidence of lacerations, and when appropriate, the
lacrimal system must be evaluated to assess for associated injury to the
lacrimal system, which generally would require repair at the time of the lid
laceration repair. In the setting of full-thickness lacerations, an underlying
associated injury to the globe should be considered. When appropriate, the
lids should be everted to evaluate for foreign bodies. In the setting of an open
globe injury, repair of adnexal injuries should occur after repair of an open
globe injury to avoid undue pressure on eye (10,11).
Visual Acuity, Pupillary Reflex, and Intraocular

Pressure
Visual acuity should be measured. The pupils should be evaluated for
response to light and presence or absence of an afferent pupillary defect, as
the latter is a significant prognostic indicator in the setting of ocular trauma.
Intraocular pressure should be measured, except in cases of a known open
globe injury, since manipulation of the globe may promote further extrusion
of ocular tissues. Intraocular pressure may be acutely elevated in the setting
of a hyphema or severely elevated in cases of suspected retrobulbar
hemorrhage and orbital compartment syndrome, an ophthalmic emergency
requiring prompt canthotomy/cantholysis. Decreased intraocular pressure
may reflect open globe injury, cyclodialysis cleft, ciliochoroidal effusion,
and/or retinal detachment (10,11).
Conjunctiva
The conjunctiva should be evaluated for lacerations and hemorrhages. The
length and location of conjunctival lacerations should be noted. Small
conjunctival lacerations generally self-resolve and need not require surgery,
whereas surgical closure with absorbable suture should be considered in
larger conjunctival lacerations (10,11).
Sclera
In the presence of a conjunctival laceration, the underlying sclera should be
carefully evaluated for scleral wounds suggestive of an open globe injury.
Bullous or widespread subconjunctival hemorrhage may suggest an
underlying scleral injury. The decision to surgically explore these cases
depends on the ability to obtain a sufficient examination, including dilated
fundus examination, the mechanism of injury, and other findings noted on
clinical examination or imaging. For example, an exploration may not be
necessary for a low-risk mechanism of injury with normal intraocular
pressure, an unrevealing dilated fundus examination, and a formed globe on
imaging tests. On the other hand, an exploration would be indicated in the
setting of a high-risk mechanism of injury, such as an injury from a sharp
object concerning for a penetrating injury with bullous subconjunctival
hemorrhage and an associated vitreous hemorrhage (10,11). In such cases, the
sclera should be carefully inspected to evaluate for scleral wounds suggestive
of open globe injury. The presence of such a wound should prompt surgical
repair under anesthesia as described below.
Cornea
The cornea should be evaluated for abrasions, lacerations, and foreign bodies.
Fluorescein staining may reveal corneal abrasions, which may be managed
conservatively with antibiotic drops and cycloplegics with or without a
bandage contact lens. Superficial corneal foreign bodies may be removed at
the bedside, although doing so may require sedation or anesthesia in younger
patients. Deeper foreign bodies may be more difficult to remove and deeply
embedded inert and clean foreign bodies may be left intact, especially if
outside the visual axis. Antibiotic drops and cycloplegia may be prescribed
for the associated epithelial defect (10,11).
Corneal lacerations should be evaluated to determine whether the defect
is lamellar or full thickness. Seidel testing with or without gentle pressure on
the globe may be performed to assess for a full-thickness defect. Partial-
thickness lacerations may be treated with antibiotic drops and cycloplegia,
whereas full-thickness lacerations require treatment. Treatment options for
such lacerations include conservative management with a rigid eye shield,
bandage contact lens, antibiotics, and aqueous suppressants for small and/or
self-sealing wounds. Most full-thickness lacerations, however, require repair
either with glue or surgically. Please see below for a detailed discussion of
surgical technique for repair of zone 1 open globe injuries, that is, corneal
lacerations.
Anterior Chamber
Anterior chamber depth should be assessed. A shallow anterior chamber may
suggest an open globe injury or posterior pressure due to anteriorly dislocated
lens or choroidal hemorrhage or effusion. An excessively deep anterior
chamber may suggest a posterior ruptured globe or posteriorly dislocated lens
(10,11).
The anterior chamber should also be evaluated for presence of cells
suggestive of traumatic iritis or microhyphema. Presence or absence of a
layering hyphema and its vertical height should be noted. The annual
incidence of hyphema is approximately 17 per 100,000 individuals (12).
Two-thirds occur in the setting of blunt trauma, while one-third occur in the
setting of penetrating or perforating injuries (13,14). The most common cause
of pediatric traumatic hyphema is sports-related injury (13,14). In the setting
of hyphema, a medical history or family history of sickle cell should be
elicited, and a sickle cell prep or hemoglobin electrophoresis obtained in
patients is at risk. The risk of rebleeding is highest in the first 4 to 7 days
after injury, and studies have associated rebleeding with worse prognosis
(15). Some clinicians, therefore, recommend restriction of activities,
binocular patching, and/or placement of a shield over the eye to reduce ocular
motility and to reduce the risk of activity- or repeat trauma-induced rebleed.
However, there remains no evidence to support these interventions (15).
Management of hyphema involves elevation of the head of the bed,
cycloplegia, and topical steroids with close monitoring and management of
the intraocular pressure. Indications for anterior chamber washout for
hyphema include uncontrolled intraocular pressure despite maximal medical
therapy (>50 mm Hg for 5 days or >35 mm Hg for 7 days; in patients with
sickle cell disease, >25 mm Hg for over 24 hours), total hyphema for more
than 5 days, corneal blood staining, and/or large clots that persist for more
than 10 days (16). General principles for anterior chamber washout include
irrigation of the anterior chamber through a single paracentesis or placement
of an anterior chamber infusion with irrigation or removal of hemorrhage
using a vitrector through a two paracentesis approach. The latter is
particularly effective in cases of clotted hemorrhage. Patients with traumatic
hyphema are at risk for angle recession and glaucoma. Thus, gonioscopy
should be performed after clearance of the hyphema (12,17,18). Hyphema
and the associated inflammation can also result in posterior or peripheral
anterior synechia, especially in hyphemas of longer duration.
The presence of a foreign body and/or lens material concerning for
anterior capsular injury should be noted. The repair of such injuries as
outlined in further detail below.
Iris
The shape and position of the pupil should be noted. Presence of iridodialysis
or cyclodialysis should be noted. A peaked pupil may suggest a penetrating
or perforating injury. Retroillumination should be performed to assess for iris
transillumination defects, especially in cases where an IOFB is suspected
based on the mechanism of injury. For example, small foreign bodies may
result in self-sealing corneal wounds that are difficult to identify on
examination. The only evidence of a foreign body on clinical examination
may be indirect signs, such as the presence of hyphema, an iris
transillumination defect, and/or focal cataract or anterior capsular injury. As
noted above, iris prolapse noted during repair of an open globe injury is best
managed by repositing the tissue using a blunt instrument such as a
cyclodialysis spatula and viscoelastic, as needed. This may be accomplished
with less trauma by sweeping the iris out of the wound internally through a
separate paracentesis wound rather than externally from the wound itself. If
the IOP is too high, the iris may not reposit until the pressure is reduced. Iris
injury is generally not addressed during primary repair but may be
subsequently performed for repair of visually significant iris defects in eyes
with reasonable visual potential (10,11).
Lens
The position, clarity, and stability of the lens should be evaluated. The
presence of anterior or posterior capsular defects, focal or diffuse cataract,
and/or intralenticular foreign body should be evaluated. An anteriorly
subluxed lens generally requires surgical removal to avoid corneal
decompensation or IOP elevation. A posterior subluxed lens with intact
capsule may be left intact or may be removed in a nonurgent fashion. A
traumatic cataract without capsular violation may be scheduled for surgical
removal by a pediatric ophthalmologist or by a vitreoretinal surgeon if there
is concern for zonular weakness necessitating a posterior approach. The
timing of such intervention is dependent upon amblyopia considerations
based on the age of the patient and severity of the cataract. Cases with
capsular violation require more urgent management since the exposed lens
particles incite a robust inflammatory phacoanaphylactic or phacoantigenic
response. When present, such inflammation should be controlled with
aggressive topical steroids and cycloplegia until the lens is removed. In the
setting of an open globe injury that necessitates repair, the open globe injury
may be addressed prior to removal of the traumatic cataract. Anterior
capsular defects with cataract may be addressed through an anterior limbal
approach with anterior vitrectomy and posterior capsulotomy. A lens injury
with posterior capsular defects is best addressed through a posterior approach
using vitreoretinal surgery. The decision regarding whether or when to place
an intraocular lens depends on the age of the patient and the extent of other
ocular injuries. As with all pediatric ocular diseases, refractive and amblyopia
management are central to maximizing visual potential in eyes with lens
injuries (10,11).
Diagnostic Studies
Ancillary imaging studies may be useful to evaluate the pediatric patient with
an ocular injury. B-scan ultrasonography should be avoided in cases of a
known open globe injury, as pressure on the globe may promote further
extrusion of intraocular tissues. Ultrasound may be useful in closed globe
injuries or in cases of small eye wall wounds with low risk of tissue
extrusion. B-scan ultrasonography may reveal the presence of lens
dislocation, IOFB, vitreous hemorrhage, or retinal tear or detachment in eyes
in which media opacity limits the ability to use indirect ophthalmoscopy. B-
scan alone should not be relied upon to rule out IOFB. CT scan should be
considered in known or suspected open globe injuries, potential intraocular or
intraorbital foreign bodies, or in cases where orbital pathology is suspected.
Metal and glass are hyperdense on CT scan, while wooden and organic
matter are hypodense (10,11). The sensitivity of CT scan for identification of
IOFB is high, approximately 94.9% in one study (19). MRI may also be
considered but should be avoided in the setting of potential metallic or
ferromagnetic foreign bodies. MRI can be useful for confirming the presence
of a hypodense IOFB such as wood, which can be mistaken for air on CT
scan (20). Additional retinal imaging studies, such as optical coherence
tomography (OCT), fluorescein angiography, indocyanine green
angiography, and fundus autofluorescence, are generally not used in acute
settings of trauma and are detailed under the relevant posterior segment
manifestations below.
POSTERIOR SEGMENT
MANIFESTATIONS AND TREATMENT
OF PEDIATRIC OCULAR TRAUMA
Except in the case of a known ruptured globe, a dilated fundus examination
with indirect ophthalmoscopy should be performed to evaluate for vitreous
hemorrhage, IOFB, retinal hemorrhage, retinal dialysis, breaks, or
detachment, commotio retinae, traumatic macular hole, choroidal rupture,
and/or traumatic chorioretinal rupture. Except in the case of a suspected open
globe injury or hyphema, a scleral depressed examination should be
considered to evaluate for anterior retinal pathology, such as retinal dialysis,
breaks, or detachments. An EUA should be considered in cases in which a
sufficient evaluation cannot be performed at the bedside. If an open globe
injury is suspected, early management includes placement of a protective
rigid eye shield and limitation of activity. The tetanus vaccination status
should be determined, and a tetanus shot or booster should be administered
based on the vaccination status and mechanism of injury. Diagnostic studies
and systemic and anterior segment considerations are addressed above.
Please see below for a detailed discussion regarding the diagnosis and
management of specific posterior segment manifestations of trauma.
Management of Acute Open Globe Injury

Management of the suspected open globe injury includes placement of a
protective rigid eye shield, limitation of activity, and administration of a
tetanus shot or booster based on the vaccination status and mechanism of
injury. The patient should be made NPO (nil per os, nothing by mouth) in
preparation for surgery, and surgery should be performed within 24 hours of
the injury whenever possible. The surgical goals for open globe injury
include restoration of globe integrity through emergent closure of the primary
wounds and minimization of endophthalmitis risk through closure of the
primary wounds and removal of any IOFBs (10,11).
No randomized, controlled trials have demonstrated efficacy of systemic
antibiotic therapy in reducing endophthalmitis rates after all open globe
injuries, and retrospective studies are inconclusive. Some clinicians initiate
systemic antibiotic therapy for all cases of open globe injury, whereas others
reserve such therapy for high-risk injuries such as penetrating injuries,
perforating injuries, or intraocular foreign bodies compared to ruptured globe
injuries from blunt trauma. There remains no consensus regarding the optimal
regimen, route, frequency, or duration. A broad-spectrum antibiotic regimen
with adequate ocular penetration, such as intravenous vancomycin and
ceftazidime, may be considered. Although fluoroquinolones may be used in
adult patients, their use in the pediatric population is limited due to the
potential adverse effects of destructive arthropathy.
Prophylactic scleral buckle surgery may be considered at the time of the
open globe repair especially in eyes with complex mechanisms involving
blunt force, zone 3 injury, or significant vitreous loss. Disruption of the
vitreous in posterior ruptured globe injuries can result in a cycle of
hemorrhage, inflammation, and vitreous organization, which predisposes to
future retinal breaks, retinal detachment, and proliferative vitreoretinopathy
(PVR). Placement of a scleral buckle may support the anterior vitreous and
reduce the risk and/or severity of subsequent retinal complications. Also,
buckle placement may be more facile at the initial surgery as a peritomy,
dissection of Tenon’s capsule, and isolation of muscles will have already
been performed, whereas placement of a buckle at a later time, such as at the
time of a subsequent vitrectomy for retinal detachment repair, may be more
challenging in the setting of conjunctival and Tenon’s scarring from the prior
open globe repair. However, placement of such a buckle may be challenging
in the setting of the primary open globe repair and may cause further undue
manipulation and pressure upon the freshly repaired globe. In these cases,
sometimes a buckle is best not performed. In cases of injuries that incurred
minimal blunt force even in the setting of perforation, IOFB and vitreous
hemorrhage, a scleral buckle may not be needed. The literature on
prophylactic scleral buckling in the setting of primary open globe repair is
quite limited, with only a few small studies demonstrating mixed results
(21–25). In addition, in young children, segmentation of the buckle may be
considered as the eye grows (see also Chapter 60).
Equipment
Extraocular tray
Relevant sutures (e.g., 10-0 nylon for zone 1 injuries, 9-0 nylon for zone
2 injuries, and 8-0 nylon for zone 3 injuries; 2-0 silk for hooking
extraocular muscles; 6-0 Vicryl sutures for removal and replacement of
extraocular muscles; 7-0 Vicryl or 6-0 plain gut sutures for conjunctival
closure)
Viscoelastic material as needed for globe reformation
Vitrectomy system (in the setting of IOFB)
Foreign body forceps, rare earth magnet, Finesse flex loop (Alcon, Ft.
Worth, TX), or other instruments as needed for removal of an IOFB
Intraocular gas or oil tamponade as needed for IOFB cases
Scleral buckle and tray, if indicated
Subconjunctival or intravitreal (e.g., vancomycin 1 mg in 0.1 cc and
ceftazidime 2.25 mg in 0.1 cc) antibiotics
Procedural Techniques for Acute Open Globe Repair

Open globe injuries include those from rupture or laceration. Surgical
principles of zone 1 open globe injuries, that is, corneal lacerations, include
use of interrupted 10-0 nylon sutures oriented perpendicular to the wound and
at approximately 90% of corneal suture depth. Anatomical landmarks such as
the limbus or a pigmentation line may be approximated first to facilitate
proper alignment of tissues. The length of the sutures as well as the distance
between sutures may be modified to minimize induced astigmatism.
Specifically, suture passes may be longer and wider apart more peripherally
and shorter and closer to one another in the central cornea. Extruded uveal
tissue should be reposited whenever possible using a blunt instrument such as
a cyclodialysis spatula as well as viscoelastic as needed. This may be
accomplished with less trauma by sweeping the iris out of the wound
internally through a paracentesis wound rather than externally from the
wound itself. The anterior chamber may be reformed during suturing using
viscoelastic, balanced salt solution, or filtered air, with attention to the effect
of intraocular pressure on the tightness of sutures. Overfilling the eye will
result in overly loose sutures, whereas suturing in a soft eye will result in
overly tight sutures with the ensuing challenges, respectively, of water tight
closure or induced astigmatism. Suture knots should be buried and
fluorescein should be used to perform a Seidel test to confirm water-tight
closure. Purse string sutures or adjuvant agents, such as glue, may be required
to achieve a water tight closure in irregularly shaped wounds (10,11).
Basic principles of repair of open globe injuries in zone 2 and/or zone 3
include creation of a peritomy and careful inspection to identify all scleral
injuries and their full extent. Since the sclera is particularly weak near the
insertion of muscles, the extraocular muscles may be hooked to adequately
evaluate for scleral wounds in this area and to achieve rotation of the globe to
expose wounds and facilitate their closure. In some cases, the muscles may
need to be secured by suture, as in strabismus surgery, and disinserted to
enable sufficient closure of the open globe injury before resuturing the
muscle to its insertion. Generally, 9-0 nylon suture is appropriate for zone 2
scleral wounds, while 8-0 nylon may be used for more posterior scleral
wounds. Partial-thickness scleral suture passes should be made perpendicular
to the wound at approximately 70% to 90% depth, and suture knots should be
buried. Seidel testing should be performed to confirm a water-tight closure.
Extruded vitreous may be excised using scissors, taking care to avoid traction
on the vitreous. Anatomical landmarks such as the limbus may be
approximated first to facilitate proper alignment of tissues. Anteroposterior
scleral wounds should be first reapproximated anteriorly before addressing
the posterior extent of the wound. Rather than promote tissue extrusion by
manipulating the globe to reach the posterior extent of wounds far posterior
to the equator, these far posterior injuries may be left unsutured, as they
generally self-seal. Extruded uveal tissue should be reposited, if possible, or
excised with care taken to minimize traction on the tissue (10,11).
After repair of the open globe injury and restoration of globe integrity,
subconjunctival antibiotics or intravitreal antibiotics may be considered.
There remain no large prospective studies demonstrating optimal antibiotic
regimen or route in eyes with open globe injury—that is, systemic antibiotics,
subconjunctival antibiotics, intravitreal antibiotics, or a combination of these.
Some clinicians administer a standard regimen of antibiotics for all cases,
whereas others tailor the decision for antibiotics and the route of
administration based on risk factors, such as penetrating injuries, perforating
injuries, or IOFBs compared to ruptured globe injury from blunt trauma.
Procedural Techniques for Removal of an IOFB

In some cases of ocular trauma, the cornea may be so violated that the goals
to restore globe integrity and remove the IOFB are more important than
attempting a keratoprosthesis to provide visualization in order to use standard
vitrectomy techniques. The IOFB may be removed from an enlarged (>5
mm) circumferential incision at the pars plana by having the assistant apply
gentle pressure on the posterior wound while elevating the anterior wound
and with irrigation identify the IOFB that can be manually extracted with
forceps. The wound is closed and the secondary procedure of
keratoprosthesis and vitrectomy is performed later.
In the setting of an IOFB in which removal can be performed using a
vitrectomy, the open globe injury must be closed with a water-tight closure
first. Placement of cannulas prior to adequate closure will result in further
extrusion of intraocular contents. If vitreous hemorrhage obscures
visualization of the infusion cannula, the cannula may be placed first into the
anterior chamber, especially in aphakic or pseudophakic eyes, until the media
are cleared sufficiently to visualize the posterior infusion cannula. A
preoperative B-scan is helpful to rule out choroidal detachment in the area of
the posterior cannula. A vitrectomy should be performed to remove the
vitreous and hemorrhage. If the IOFB violated the lens capsule, a lensectomy
may also be performed using the vitrector. The decision regarding whether or
not to induce a posterior vitreous detachment (PVD) depends upon the age of
the patient and potential complications related to PVD induction in younger
eyes with highly adherent hyaloid. Release of the vitreous attachments
around the IOFB and removal of the IOFB may be sufficient for the
procedure. In some cases, perfluoro-octane (PFO) heavy liquid may be
placed over the macula prior to IOFB removal to float the IOFB away, if
possible, or keep it from delicate retinal structures and the macula.
Depending upon the size and composition of the IOFB, standard
microvitrectomy forceps, foreign body forceps, rare earth magnet, and/or a
Finesse loop (used as a snare) can be used to retrieve the foreign body. The
foreign body can be removed through the pars plana by removing the cannula
and enlarging the sclerotomy tangential to the limbus or, in aphakic eyes,
through a corneal wound. Strike sights are considered for laser treatment and
a scleral depressed examination should be performed to evaluate for other
IOFB material and retinal breaks, which should be treated. An intraocular
tamponade, such as gas or oil, should be placed in eyes with retinal breaks
and considered in eyes with strike sites. Intravitreal antibiotics should be
administered, and the dosage may be reduced in eyes with intraocular
tamponade. There remains no consensus on the optimal dilution in gas or oil-
filled eyes, but some advocate reducing by one-half.
Postoperative Care
After surgery, the eye should be dressed with antibiotics and a sterile patch
and shield. During the postoperative period, the patient is monitored to
evaluate for leaks from wounds and for secondary complications, such as
retinal detachment and/or infection. A protective shield should be kept on the
eye for at least the 1st week to reduce the risk of pressure on the globe that
might result in a wound dehiscence and fluid or ocular contents leaking from
the open globe injury site. A standard regimen of topical steroids,
cycloplegics, and antibiotics should be administered. There is no consensus
on systemic antibiotic therapy after open globe repair. The fellow eye should
be monitored in the long term for development of sympathetic ophthalmia.
Amblyopia should be managed in collaboration with a pediatric
ophthalmologist. Monocular precautions and protective eyewear should be
recommended to protect the fellow eye. Subsequent EUAs may be required
to adequately evaluate the eye postoperatively and/or to remove sutures.
Complications
Complications of open globe injury include anterior segment complications,
such as corneal scar or cataract, endophthalmitis, sympathetic ophthalmia,
nonclearing vitreous hemorrhage, choroidal hemorrhage, retinal tear, retinal
detachment, and PVR.
The incidence of endophthalmitis following open globe injury in patients
of all ages ranges from 0% to 16.5%, with increased risk associated with
penetrating injury, IOFB, delay in wound closure over 24 hours from the time
of injury, injury in a rural setting due to the microbes inoculated, and
ruptured lens capsule (26). Commonly associated microbial agents include
Staphylococcus, Bacillus, Streptococcus, Clostridium, Pseudomonas,
Candida, Aspergillus, Paecilomyces, Fusarium, and dematiaceous fungi
species and polymicrobial infections may occur (26).
Studies of pediatric patients with open globe injuries have described an
incidence of retinal detachment of 25% to 31% (27,28). Risk factors for
retinal detachment after an open globe injury in the pediatric patient are
similar to those in adults and include higher zone, that is, more posterior
injury, longer wound length, vitreous hemorrhage, rupture versus laceration,
and retained IOFB (27). In addition to vitreous prolapse and retinal
incarceration, other important mechanistic factors for retinal detachment after
open globe injury include vitreous hemorrhage, secondary inflammation, and
vitreous scaffolding and traction (29).
Studies suggest that 54% to 56% of pediatric open globe injuries achieve a
final vision of 20/40 or better (30–32). Studies have previously reported
scoring systems to predict visual outcome after ocular injury. The ocular
trauma score (OTS), based on data from the United States Eye Injury
Registry, identified poor initial visual acuity, ruptured globe, perforation,
endophthalmitis, retinal detachment, and presence of afferent pupillary defect
as poor prognostic indicators (33). More recently, a pediatric OTS (POTS)
score was reported, in which the impact of visual acuity on score was de-
emphasized in patients in whom visual acuity could not be obtained. Other
factors included in the POTS score include age (younger age portending a
worse prognosis), zone of injury (higher zone associated with worse
prognosis), presence or absence of unclean injury, delay in surgery >48
hours, and presence or absence of associated ocular findings including iris
prolapse, hyphema, traumatic cataract, vitreous hemorrhage, retinal
detachment, and/or endophthalmitis (34). A toddler/infant OTS (TOTS),
which does not include visual acuity given the difficulty of assessing this
metric in infants and toddlers, included only wound size >6 mm and presence
or absence of hyphema, cataract or lens damage, retinal detachment, and
choroidal detachment in scoring and produced a sensitivity of 81% in
predicting visual outcome (35).

In addition to repair of the ruptured globe and removal of an IOFB, if present,
subsequent vitreoretinal surgery may be required in the setting of vitreous
hemorrhage, retinal detachment, and/or endophthalmitis. Please see below a
detailed discussion of surgical considerations for these clinical scenarios.
Vitreoretinal Surgery for Complications Following
Initial Repair of Open Globe Injuries (Vitreous
Hemorrhage and Retinal Detachment)
Indications
Vitreous hemorrhage and/or retinal detachment after an open globe injury
usually warrants vitrectomy surgery with or without scleral buckle. Vitreous
hemorrhage has been noted in 3.3% to 34% of pediatric open globe injuries
(36). As with adult patients, vitreous hemorrhage associated with open globe
injury, especially that involving zone 2 or zone 3, has a higher risk of
posterior segment pathology, which can result in subsequent retinal
detachment. Thus, these cases often require urgent surgical vitrectomy.
Retinal detachment after blunt injury can occur from a dialysis, retinal
tear, and development of PVR. Depending on the reason, retinal detachments
are treated with a scleral buckle or vitreoretinal surgery. Traumatic macular
hole may also warrant vitreoretinal surgery.

When possible, a vitrectomy is generally performed as a secondary procedure
after repair of the open globe injury to restore globe integrity. In such cases,
the appropriate time for intervention varies (23,24,37–40). Some advocate for
early intervention within 3 to 4 days, citing the potential for reduced
inflammation and reduced formation of tractional membranes with prompt
removal of damaged vitreous and hemorrhage (37,39). Many clinicians
advocate for later vitrectomy after 10 to 14 days, pointing to the reduced
theoretical risk of massive hemorrhage from congested uveal tissue, more
stable globe anatomy after primary repair of the open globe injury, and
increased chance of liquefaction of hemorrhage, as in cases of hemorrhagic
choroidal or retinal detachment, and creation of a PVD in adults (23,40), but
this may not occur as readily in infants and young children.
Equipment
Vitrectomy system
Triamcinolone acetonide injectable suspension, if needed to confirm
facilitate PVD induction (e.g., Triesence, Alcon, Ft. Worth, TX)
Additional instrumentation as needed for to induce and carry out the
PVD (pick, bent microvitreoretinal [MVR] blade, a Finesse flex loop
[Alcon, Ft. Worth, TX], and/or forceps)
Endolaser probe
Internal diathermy
PFO heavy liquid, if needed
Tamponade: intraocular gas (SF6, C3F8) or silicone oil
Scleral buckle and tray, if indicated
In cases of vitreous hemorrhage after open globe repair, small-gauge pars
plana vitrectomy may be performed to evacuate the hemorrhage and address
underlying retinal pathology. A PVD may be induced if readily able or if
necessary to address concurrent underlying retinal pathology, such as
tractional/rhegmatogenous retinal detachment. A scleral depressed
examination should be performed to evaluate for peripheral breaks, and if
found, they should be treated with endolaser and tamponade, as necessary. A
prophylactic scleral buckle surgery may be considered particularly in
conditions with some aspect of blunt force. In perforating injuries in eyes of
infants or young children, in which an IOFB is in the orbit and is rendered
too traumatic to remove, release of tractional vitreous forces between the
anterior and posterior wounds can be helpful to reduce later complicated
tractional retinal detachments. Management of amblyopia is important.
In cases of retinal detachment following open globe injury, the central
principles are (a) to identify and treat retinal breaks and (b) to relieve vitreous
traction on the breaks. Disruption of the vitreous or hemorrhage in an open
globe injury predisposes to later inflammation with vitreous organization and
contracture, retinal breaks, detachments, and PVR. A scleral buckle should be
considered prior to vitrectomy. By supporting the vitreous base, a scleral
buckle can relieve tractional forces at the anterior vitreous. Small-gauge
vitrectomy should be performed to remove vitreous hemorrhage and
organized vitreous associated with retinal detachments after open globe
injury. A posterior cortical vitreous separation may be induced as a PVD, if
possible, and vitreous traction must be relieved from the retinal breaks.
Maneuvers that can facilitate PVD induction in the pediatric population
include vitrectomy with triamcinolone acetonide injectable suspension to
stain the vitreous and make it easier to identify the vitreous face to create a
PVD without vitreous schisis. Additional instruments such as a pick, bent
MVR blade, a Finesse flex loop, and/or forceps may be used to create an
initial opening in the hyaloid to facilitate its separation from the retinal
surface. Scleral depression can be used to identify and relieve traction on all
retinal breaks and areas of concern. If significant anterior membranes are
present, lensectomy should be considered to enable adequate relief of anterior
vitreous traction. Subretinal fluid should be drained from existing internal
breaks and the breaks treated with laser. Intraocular gas, generally C3F8, or
silicone oil tamponade should be placed depending on the location of the
breaks, presence or absence of PVR, and the patient’s age and ability to
position. Silicone oil tamponade is often required in younger patients in
whom positioning may be difficult. An inferior surgical iridotomy should be
performed in eyes receiving silicone oil that were rendered aphakic from the
open globe injury or surgically. Retention sutures (9 or 10-0 Prolene) may be
considered in aniridic eyes undergoing placement of silicone oil. There are no
studies comparing outcomes with and without retention sutures in aniridic
eyes with silicone oil tamponade.
Postoperative Care
Postoperative care focuses on monitoring for retinal detachment, PVR,
infection, and complications of silicone oil. A postoperative regimen of
topical steroids, cycloplegia, and antibiotics should be administered. Based
upon the patient’s age and ability to position, positioning should be
recommended after retinal detachment repair to provide optimal tamponade
at the retinal breaks and to reduce complications of intraocular tamponade.
Amblyopia should be managed in collaboration with a pediatric
ophthalmologist. Monocular precautions and protective eyewear should be
recommended to protect the fellow eye. Subsequent EUAs may be required
periodically to properly examine the young patients.
Parents should be counseled regarding the guarded anatomic and functional
prognosis for retinal detachment repair after open globe injury in the pediatric
population. Studies indicate a significantly lower success rate compared to
retinal detachment after closed globe injury, with anatomic success rates in
open globe associated retinal detachments of 21% to 46.7% (27,41–43).
Anatomic failure in these studies was generally due to new breaks or PVR
(41,43). The risk of PVR is significant with some studies reporting up to 64%
of eyes exhibiting grade C or worse PVR at initial presentation. PVR remains
a major risk factor for anatomic failure (43,44). As with all pediatric ocular
pathology, amblyopia may significantly impact final visual outcome in open
globe injury.
Management of Ocular Sequelae From Closed Globe

Injury
Vitreous Hemorrhage
Pediatric patients may also present with vitreous hemorrhage without an open
globe injury. In a large series of pediatric vitreous hemorrhages at a single
center, 73% were attributable to trauma, including 29.5% from
nonpenetrating trauma, 24.7% from penetrating trauma, 8.6% from
nonaccidental trauma, 5.4% noted postoperatively, and 4.8% associated with
birth trauma (45). Vitreous hemorrhage may occur in isolation or may
commonly be associated with other findings such as hyphema, traumatic
cataract or lens dislocation, and/or open globe injury with or without IOFB.
Vitreous hemorrhage or hemorrhage into a schisis cavity may also occur in
the setting of mild trauma in eyes with retinoschisis. Evaluation of these
patients includes appropriate history and evaluation of the eye and ocular
adnexa for associated injuries. Dilated fundus examination with scleral
depression should be performed to assess for associated retinal pathology
such as retinal tears, retinal dialysis, or retinal detachment, and an EUA
should be considered in patients in whom a sufficient bedside examination
with scleral depression is not possible. When vitreous hemorrhage obscures
the retinal evaluation, B-scan ultrasonography should be employed to
evaluate for evidence of retinal tears and detachments.
Vitreous hemorrhage in the absence of open globe injury or associated
retinal pathology may be observed for several months for clearance in older
pediatric patients before pursuing surgery. Due to the formed nature of the
pediatric vitreous, the hemorrhage may be less likely to disperse and is
slower to resorb than in adults. However, it should be noted that vitreous
hemorrhage may result in development of an epiretinal membrane, and
organization of the vitreous hemorrhage over time may produce secondary
vitreous traction and retinal detachment. Earlier intervention should be
considered in cases with high suspicion of associated retinal pathology
warranting treatment based on clinical exam and mechanism of injury. Due to
amblyopia considerations, earlier intervention should also be performed in
pediatric patients age 6 to 8 years and younger, especially those with dense
vitreous hemorrhage or hemorrhage involving the visual axis. Specifically,
extrapolating from the congenital cataract data, irreversible damage from
occlusive amblyopia may occur as early as 6 to 8 weeks in young infants
(46,47). Moreover, occlusion by vitreous hemorrhage may also induce a high
degree of anisometropic amblyopia in infants, which can persist even after
the vitreous hemorrhage clears or removed (48,49). Given the inability to
determine precise onset of media opacity in older patients, there are limited
data on the precise timeline for irreversible occlusive amblyopia in older
patients. Surgical techniques are similar to those described above under
vitrectomy for vitreous hemorrhage from open globe injury.
Retinal Break, Retinal Dialysis, and Retinal

Detachment From Remote Trauma
Ocular trauma is the most common cause of pediatric retinal breaks, retinal
dialysis, and retinal detachment, accounting for 36% to 61% of detachments
in this population (50–56). In one series, among pediatric retinal detachments
associated with trauma, 22% had round holes, 9% had round holes within
lattice degeneration, 19% had flap tears, 0.9% had flap tears within lattice
degeneration, 19.9% had retinal dialysis, 3% had giant retinal tears, 3% had
macular holes, 5.8% had more than two kinds of break, and 16% had
undetected breaks (54).
Pediatric patients with contusive ocular injury, especially those with
symptomatic flashes, floaters, or vision change, should undergo dilated
fundus examination with scleral depression to evaluate for peripheral retinal
breaks and/or dialyses. EUA should be considered for patients unable to
tolerate a bedside scleral depressed examination if the mechanism of injury or
associated ocular findings, such as vitreous hemorrhage, suggest a high risk
of peripheral retinal pathology. Retinal breaks may be repaired by laser or
cryotherapy at the bedside or under anesthesia. Young patients old enough to
undergo an office procedure but too young to withstand a longer procedure
may tolerate cryotherapy better than laser. Rhegmatogenous retinal
detachment warrants intervention.
Indications and basic considerations regarding surgical technique

As with all retinal detachment repairs, the key goals of the surgery include
treatment and support of all retinal breaks. Due to the frequent absence of a
PVD, the risks associated with inducing a PVD (including iatrogenic break),
and the high rate of PVR in the setting of retinal breaks, repair with scleral
buckle alone is preferable.
Retinal dialysis
Retinal dialysis is the most common type of retinal break in pediatric patients
and has been implicated in 14% to 33% of all pediatric retinal detachments
(50,54). While most are associated with trauma, in some cases a definitive
trauma history cannot be elicited. A dialysis is a circumferential disinsertion
of the retina at the ora serrata. A dialysis may be differentiated from a giant
retinal tear by the location relative to the vitreous base: a dialysis is a defect
anterior to the posterior insertion of the vitreous base, whereas a giant retinal
tear is a defect posterior to the posterior insertion of the vitreous base. Most
retinal dialyses are located inferotemporally. Additional breaks may be
present. In the pediatric patient with formed vitreous and absence of a PVD,
progression of the dialysis can be slow with delayed onset of development of
the dialysis-associated retinal detachment (57,58). In one study, 41% of
retinal detachments were diagnosed more than 1 year after the trauma (59).
Associated findings include demarcation lines and/or retinal cysts. PVR is
rarely encountered. Scleral indentation examination of the trauma patient is
essential for detecting a retinal dialysis. Management of retinal dialysis
generally involves surgical repair with scleral buckle, generally with
cryopexy along the dialysis edge (57,58,60) (Figure 69-2A). Segmental
elements can treat the dialysis effectively, while averting some of the
concerns associated with encircling scleral bands, such as myopic shift with
anisometric amblyopia and/or need to divide the buckle in a subsequent
surgery to enable growth of the globe in infants and very young children.
Subretinal fluid drainage may not be necessary but is considered for larger
detachments and those with chronic, viscous fluid. PVR is rare, and single-
surgery success rates are high (59,61–63).
FIGURE 69-2 Previous commotion retinae, repaired

traumatic retinal dialysis with retinal detachment, and
choroidal rupture. A:Ultra widefield fundus photograph of
the left eye after repair of traumatic dialysis-related retinal
detachment with scleral buckle and cryotherapy. The
retina is attached with buckle effect and temporal
cryotherapy scars. There is retinal pigment epithelium
(RPE) atrophy with areas of RPE clumping
superotemporal to the optic disc from prior commotio
retinae and an inferotemporal, posterior curvilinear yellow
line with tapered edges at the site of a choroidal rupture.
B:Fundus autofluorescence reveals hypoautofluorescence
in the area of RPE disruption. The choroidal rupture is
hypoautofluorescent with a few areas of
hyperautofluorescence at the margin of the rupture site.
C:Optical coherence tomography reveals an epiretinal
membrane and a discontinuity in the RPE–Bruch
membrane complex corresponding to the choroidal rupture
site.
Giant retinal tear
A giant retinal tear is a neurosensory retinal break that extends
circumferentially for three or more clock hours. They are most commonly
found posterior to the vitreous base, although rarely they may be found at or
posterior to the equator. As noted above, the giant retinal tear, unlike a
dialysis, occurs posterior to the posterior insertion of the vitreous base (64).
Trauma accounts for 32% of giant retinal tears in the pediatric population,
whereas hereditary vitreoretinopathies, such as Sticker syndrome, account for
a majority of the remainder (65). In a study of traumatic retinal detachments
in pediatric patients, the rate of giant retinal tear was similar in those with
open (22%) and closed (17%) globe injuries.
Repair of giant retinal tear-associated retinal detachment requires
vitrectomy. Opinions differ regarding the benefit of adjunctive scleral buckle.
A buckle can provide additional support of the anterior vitreous and vitreous
base, especially in pediatric patients or phakic patients in whom vitrectomy
can be limited anteriorly. On the other hand, there is a theoretical increased
risk of retinal slippage in the setting of a scleral buckle. If used, an overly
high buckle should be avoided and one may consider waiting until the retinal
edge has been sufficiently dried before the buckle is tightened. During
vitrectomy, attention should be taken to achieve a PVD, to carefully trim the
vitreous base, and to remove vitreous adhesions to the edge of the giant
retinal tear and the anterior horns of the tear. Perfluorocarbon (PFO) heavy
liquid should be used to flatten the retina and facilitate unfurling of any
folded retinal edge with care to avoid PFO entering the subretinal space. If a
fluid–air exchange is performed, in order to avoid slippage of the retinal
edge, drainage should be done slowly with attention taken to first remove the
meniscus of fluid at the edge of the tear between the PFO bubble and the eye
wall posteriorly before further drying of the retinal edge and finally, careful
removal of the PFO itself. Alternatively, a direct PFO–silicone oil exchange
may be performed. To perform a direct PFO–silicone exchange, the infusion
cannula is replaced with a silicone oil infusion line. Using the light pipe in
one hand and the extrusion cannula in the other, the silicone oil is injected
into the eye using the foot pedal while the PFO is removed by passive
extrusion. Alternatively, an assistant can manually inject the oil through the
oil infusion line. Initially, the extrusion cannula tip should be placed in the
BSS layer between the silicone oil above and the PFO layer below. When the
BSS layer has been removed, the extrusion cannula tip is advanced
posteriorly into the PFO to passively drain the PFO while silicone oil is being
infused. Care must be taken to ensure appropriate oil fill and IOP, as well as
to ensure complete removal of the PFO. Laser retinopexy, or rarely cryopexy,
should be applied to the edge of the tear. Long-acting C3F8 gas or silicone oil
tamponade may be used. In younger patients in particular, limitations in the
ability to position generally necessitate use of silicone oil (66,67). When
possible, the oil should be removed by 1 year.
Traumatic Macular Hole

The first reports of macular hole by Herman Knapp and Noyes were
traumatic in origin. Traumatic macular holes (Figure 69-3A–C) have been
reported in 1.4% of closed globe injuries and 0.15% of open globe injuries
and often occur in association with other posterior segment manifestations of
trauma, including commotio retinae, choroidal rupture, traumatic
chorioretinal rupture, and/or retinal breaks or detachments (1). The
pathophysiology of traumatic macular hole involves blunt force resulting in a
sudden axial deformation of the globe in which there is a reduction in the
anteroposterior diameter and expansion of the equatorial diameter of the
globe. This results in the transmission of horizontal forces to the posterior
pole that split the retinal layers in the fovea. Varying degrees of vitreous
traction may also contribute to traumatic macular hole formation (68–70).
Thus, unlike the slow, degenerative process of vitreoretinal traction at the
fovea implicated in idiopathic macular hole, traumatic macular holes occur
from a sudden structural change and variable vitreoretinal traction. The
progression of and visual outcomes from traumatic macular hole is, therefore,
somewhat unpredictable. Unlike idiopathic macular holes, traumatic macular
holes occur in younger patients and occur more commonly in males as with
most traumatic ocular injuries. OCT is useful in the evaluation of traumatic
macular holes, the vitreoretinal interface, and adjacent injury of the retina and
RPE. On OCT, traumatic macular holes tend to exhibit larger basal diameters
and thinner average retinal thickness than idiopathic macular holes. They are
more often eccentric, elliptical, or irregular in shape than idiopathic macular
holes. Most notably, a PVD and opercula are typically absent in traumatic
macular hole (71).
FIGURE 69-3 Commotio retinae and traumatic macular
hole. A:Fundus photograph of a 10-year-old boy, 4
months after soccer ball injury to the right eye, reveals
retinal pigment epithelium (RPE) atrophy superior to the
optic disc from resolved commotio retinae and a traumatic
macular hole. B:Fundus autofluorescent imaging reveals
hypoautofluorescence with a few punctate foci of
hyperautofluorescence in the area of resolved commotio
retina as well as hyperautofluorescence at the macular
hole. C:Optical coherence tomography reveals a full-
thickness macular hole with associated cystic changes.
Indications and basic considerations regarding surgical technique

Spontaneous closure of traumatic macular holes has been described within
days or months of the injury. Rates of spontaneous closure are variable and
range from 10% to 44% (68,72). The rate of closure is considered higher
among pediatric traumatic macular holes (68,73,74). One series noted a
spontaneous closure rate of 50.0% in children 18 years and younger
compared to 28.6% in adults (73). As with idiopathic macular hole,
spontaneous closure is more likely with smaller traumatic macular holes (75).
The absence of a PVD was also frequently noted in eyes that experienced
spontaneous closure. However, given that vitreous detachment is an age-
related phenomenon, it remains unclear if the likelihood of a PVD has impact
on improved outcomes seen in pediatric compared to adult patients.
Based on the high rate of closure for pediatric traumatic macular holes, a
period of observation may be considered, especially in older pediatric
patients with small holes, good visual acuity, and/or posterior vitreous
adhesion to the edges of the hole. Although most previously reported cases of
closure occurred within 3 months of the injury, there are no clear guidelines
regarding the optimal duration of observation in these patients. In younger
pediatric patients, considerations regarding deprivational amblyopia may
prompt earlier intervention.
Equipment
25G vitrectomy set (consider 25G small instrumentation)
Triamcinolone acetonide injectable suspension if needed to confirm or
facilitate PVD induction (e.g., Triesence, Alcon, Ft. Worth, TX)
Indocyanine green or brilliant blue stain
Internal limiting membrane (ILM) forceps
Additional instrumentation as needed for PVD induction or ILM peel
initiation (pick, bent MVR blade, Finesse flex loop [Alcon, Ft. Worth,
TX], and/or forceps)
Tamponade: intraocular gas (SF6, C3F8) or silicone oil
Procedural techniques
Surgical management of idiopathic macular holes has been highly successful
and centers upon addressing the pathophysiologic anteroposterior and
tangential tractional forces on the retina through pars plana vitrectomy with
removal of the posterior hyaloid and in most cases, the ILM (76). In contrast
to idiopathic macular holes, the role of vitreous traction in traumatic macular
hole is unclear and likely variable. Nonetheless, a similar surgical approach
to idiopathic macular hole surgery involving pars plana vitrectomy with
removal of the posterior vitreous, gas tamponade, and prone positioning has
been employed for traumatic macular hole with success rates ranging from
45% to 100% (median 92.5%) in small case series. Although most of these
series did not include pediatric cases exclusively, the mean ages in these
series was low, ranging from 10 to 32 years (77).
Surgical principles for traumatic macular hole are similar to those for
idiopathic macular hole. Pars plana vitrectomy with complete removal of the
posterior hyaloid is critical. Pediatric patients pose a particular challenge for
PVD induction due to the well-formed vitreous humor and the strong
adhesion of the posterior hyaloid. Complete removal of the vitreous can be
challenging, in some cases with an apparent Weiss ring noted but only a
lamellar separation of the vitreous with a residual outer layer of cortical
vitreous remaining on the retinal surface. Moreover, complications during
PVD induction can result in iatrogenic retinal breaks, retinal trauma, and/or
hemorrhage. In contrast to adult patients, retinal breaks in pediatric patients
may incite intense PVR with potentially catastrophic ocular outcomes.
Maneuvers that may facilitate PVD induction in the pediatric population
include vitrectomy using triamcinolone acetonide injectable suspension to
stain the vitreous and make it easier to identify the vitreous face to create a
PVD without vitreous schisis. In addition to the vitreous cutter, additional
instruments such as a pick, bent MVR blade, Finesse flex loop, and/or
forceps may facilitate creation of an opening in the hyaloid from which to
carry out the PVD induction.
In light of the difficulties achieving PVD induction in the pediatric
population, surgeons have also used adjunctive agents including autologous
plasmin and ocriplasmin (Jetrea, ThromboGenics, Iselin, NJ) for enzymatic
vitreolysis. Plasmin is a protease that can facilitate PVD by lysing fibronectin
and laminin, two components of the vitreoretinal attachment (78). Margherio
et al. reported four cases of pediatric traumatic macular hole closure after 0.4
international units of autologous plasmin-assisted pars plana vitrectomy with
C3F8 gas and face-down positioning for 2 weeks (79). In this study and
others, surgeons have reported improved ease of PVD induction and high
rates of closure of pediatric traumatic macular hole in plasmin-assisted pars
plana vitrectomy (79–81). One disadvantage of plasmin is the technical and
logistical limitations of generating autologous plasmin from the patient’s
blood. Ocriplasmin is a recombinant truncated form of plasmin with
proteolytic activity against fibronectin and laminin that was FDA approved
for the treatment of adult patients with vitreoretinal traction and/or macular
hole. In a single-center, randomized, place-controlled, double-masked phase
2 study, ocriplasmin (175 μg) versus placebo was administered to pediatric
patients prior to vitrectomy. Of the 22 patients included, one patient
randomized to the ocriplasmin group underwent surgery for traumatic
macular hole. There was no significant difference noted in rates of total
macular PVD, vitreous liquefaction grade, adverse effects in ocriplasmin
versus placebo-treated eyes, or a difference in the duration of vitrectomy. The
study was limited by the heterogeneous nature of the study population and
small sample size. Of note, one patient in the ocriplasmin group developed
zonular dehiscence with lens subluxation (82). While the patient had anterior
segment dysgenesis at baseline, other reports have demonstrated zonular
instability after ocriplasmin. Other ocular adverse effects noted in the adult
ocriplasmin population include retinal tear, retinal detachment, retinal
vascular findings, electroretinogram abnormalities, and abnormalities in the
ellipsoid zone (83). Overall, the use of ocriplasmin for enzymatic vitreolysis
for pediatric vitreoretinal surgery is limited by preliminary data
demonstrating efficacy and safety.
Consistent with the management of macular holes at the time, early
reports of surgery for pediatric traumatic macular hole did not involve ILM
peeling per se. However, the surgical technique frequently involved removal
of “epiretinal membranes” using tools such as the diamond-dusted membrane
peeler (80,84). Such maneuvers may have removed not only residual cortical
vitreous but also some of the ILM. ILM peeling has been shown to
significantly improve the closure rate and visual outcomes in idiopathic
macular holes (85–87). ILM peeling may reduce tangential traction on the
macular hole and may further confirm complete removal of cortical vitreous
and/or epiretinal membranes from the retinal surface. Moreover, ILM peeling
may increase retinal compliance and/or stimulate glial cell proliferation.
Indeed, ILM peeling has become a routine technique for macular hole
surgery as has the use of adjuvants for staining the ILM, such as indocyanine
green, triamcinolone acetonide, and brilliant blue (88–90). ILM peeling is
also routinely employed for surgical repair of traumatic macular holes, and
successful closure of traumatic macular holes, including pediatric macular
holes, has been described with ILM peeling (91,92) and inverted ILM flap
technique for large traumatic macular holes (93).
Intraocular gas tamponade using air, SF6, or C3F8 with prone positioning
has also become standard in surgical repair of idiopathic and traumatic
macular holes. The surface tension of the gas is hypothesized to provide a
seal that prevents reaccumulation of intraretinal fluid as the hole closes as
well as to provide a scaffold for hole closure. There remains debate regarding
optimal tamponade (none, air, SF6, C3F8) and the required duration of
positioning (86,94–101) although the general practice is to use gas
tamponade with a few days or more of prone positioning. Silicone oil
tamponade may also be used, especially in young children in whom
positioning can be difficult. Although studies in idiopathic and traumatic
macular holes suggest worse outcomes with silicone oil tamponade than with
gas tamponade, these studies are limited by their retrospective nature and
potentially worse patient and ocular factors at baseline (102,103). Based on
these considerations, potential side effects from oil, and the need for a second
surgery for removal of the oil, the use of silicone oil is generally reserved for
cases such as large holes, recurrent holes, holes that previously failed closure
after surgery, or in patients such as young children who cannot position
effectively.
Early series describing surgery for traumatic macular holes employed
adjunctive agents including transforming growth factor β (TGF-β) and
platelet concentrates, such as autologous platelet rich plasma (92,104,105).
Autologous platelet rich plasma is obtained by centrifuging the patient’s own
blood and collecting the platelet-rich plasma supernatant, which is applied
into the macular hole using a soft-tip cannula. These adjunctive agents are
thought to facilitate hole closure by stimulating fibroglial proliferation at the
edge of the macular defect. These agents may also provide intrinsic adhesion
support similar to that achieved by the tamponade with facedown positioning.
Subsequent studies demonstrated successful closure of traumatic macular
holes using standard techniques, such as pars plana vitrectomy with or
without ILM peeling and prolonged gas tamponade with face down
positioning (80,84). Although some surgeons report successful closure of
macular holes, including pediatric traumatic macular holes, using autologous
platelets or autologous platelet rich plasma (106,107), there are no studies
comparing surgical outcomes with versus without use of these adjunctive
agents. Nonetheless, these agents may be considered for chronic, large, or
recurrent holes or in pediatric patients who are unable to position.
Postoperative care
A postoperative regimen of prone positioning and topical steroids,
cycloplegia, and antibiotics should be administered. There remains debate
regarding optimal duration of positioning (86,94–101) although the general
practice is to employ a few days or more of prone positioning. In children,
the use of cell phone and iPad video games may help them maintain their
eyes in a facedown position. The patient should be monitored for closure of
the macular hole as well as potential complications such as retinal
detachment, infection, and when applicable, complications of silicone oil
tamponade.
Outcome expectations
Traumatic macular holes may close spontaneously in 10% to 44% in all ages
(68,72–74) and up to 50% in pediatric patients (73). There are limited data on
outcomes in pediatric traumatic macular holes specifically. However, the
literature of surgical outcomes after traumatic macular hole surgery, in which
a majority of subjects were children, suggests anatomic closure rates ranging
from 45% to 100% (70,73,77,79,80,84,91,104,105). Since traumatic macular
holes are frequently encountered with other posterior segment manifestations
of trauma, the contribution of other pathologies that may impact visual
prognosis must be considered. Depending on the duration of the macular hole
prior to closure, amblyopic considerations may also affect prognosis.
When to consider another procedure

A subsequent vitreoretinal surgery may be required in the setting failure of
closure or recurrence of the macular hole. A subsequent vitrectomy may
reveal residual posterior hyaloidal tissue or ILM. If not, advanced techniques
discussed above including ILM free flap, inverted ILM flap (93), or use of
amniotic membrane tissue or autologous platelet-rich plasma (106,107) may
be considered. Repeat vitrectomy is required for removal of silicone oil and
may also be required for other complications such as postoperative retinal
detachment.
Commotio Retinae
Commotio retinae, also known as Berlin edema, was first described by Berlin
in 1873 as transient opacification of the outer retina following blunt ocular
trauma. Patients may be asymptomatic or report visual impairment at
presentation (108,109). Histopathologic and ultrastructural analysis of
commotion retinae have demonstrated that areas of opacification most
commonly found contrecoup or opposite from the site of impact correspond
to fragmented photoreceptor outer segments and damaged photoreceptor cell
bodies (110–112). Initially, commotio retinae presents clinically with a gray-
white opacification of the retina, sometimes with associated RPE mottling
(110–112). Involvement of the foveal region may manifest with a cherry red
spot. The OCT at initial diagnosis can reveal intraretinal edema and
hyperreflectivity of the inner segments, outer segments, and/or RPE. In some
cases, the intraretinal edema and hyperreflectivity also extends into the inner
retinal layers. Subsequently, outer receptor damage promotes RPE migration
and phagocytosis of damaged photoreceptors, leading to the clinical
appearance of RPE atrophy and areas of hyperpigmentation after resolution
of the retinal whitening (Figures 69-2A,B and 69-3A,B). OCT can
demonstrate areas of increased hyperreflectivity or disruption of the
photoreceptor inner and outer segments and RPE, and in some cases, retinal
thinning. There is no known treatment. The degree of long-term impact to the
visual acuity and visual field depends on the region of retina involved and the
degree of photoreceptor damage that ensues (108,109).
Choroidal Rupture
Choroidal rupture is a break in the choroid, Bruch membrane, and the RPE
due to trauma. Choroidal ruptures typically occur from blunt globe trauma
but have also been reported in the setting of open globes. In the largest
reported series of choroidal ruptures, including 111 cases at a single
institution, 72% occurred from closed globe injuries (113). Patients with
angioid streaks may be at increased risk for traumatic choroidal rupture
owing to the calcified and weakened Bruch membrane (114).
The predominating theory regarding pathophysiology of choroidal
rupture involves an indirect injury in which traumatic anteroposterior
deformation of the globe results in expansion of the eye equatorially. This
creates a shear force that is centered at, and radiates concentrically from the
optic disc. This results in rupture of the choroid, Bruch membrane, and RPE
that is most commonly crescent shaped, posterior, and concentric to the optic
disc. Rarely, a rupture may occur due to direct injury at the site of contact
with the globe. These ruptures are anterior and parallel to the ora serrata. The
retina is spared due to its elasticity, whereas the sclera is spared due to its
rigidity (108,115). Rupture of the retina in addition to the choroid, Bruch
membrane, and RPE is a distinct entity known as chorioretinitis sclopetaria
(see below). While most patients present with a single choroidal rupture,
multiple choroidal ruptures have been noted in 18% to 39% of cases
(113,116,117).
Choroidal rupture may be obscured at the time of the trauma due to
overlying subretinal hemorrhage. As the hemorrhage resorbs, the choroidal
rupture may be noted as a curvilinear, crescent-shaped yellow line, often with
tapered edges (Figure 69-2A). As noted above, the ruptures may be
concentric to the optic disc or may be found peripherally, parallel to the ora
serrata. In rare cases, a rupture may occur radially in the horizontal meridian.
With time, the ruptures may evolve into white streaks with RPE hyperplasia
at the margins. Fluorescein angiography reveals early hypofluorescence in
the area of rupture due to disruption in the choroidal signal. The normal
choriocapillaris at the edge of the margin may leak into the scar resulting in
late hyperfluorescence and staining (109,118). Indocyanine green
angiography may demonstrate rupture sites obscured by hemorrhage and
reveals hypocyanescence in all phases (118). Fundus autofluorescence will
reveal hypoautofluorescence in the region of the rupture due to the absence of
RPE in this area (Figure 69-2B). The margin of the rupture may reveal
hyperautofluorescence due to RPE hyperplasia at the edges (Figure 69-2B)
(119,120). OCT through the choroidal rupture may reveal the defect (Figure
69-2C). Two OCT patterns have been described, including a disruption in the
RPE or the RPE–choriocapillaris associated with (a) a forward protrusion of
the RPE–choriocapillaris layer resulting in a pyramid or dome shape or (b) a
posteriorly concave defect with loss of the overlying ellipsoid zone and
external limiting membrane (121). The extent of visual impairment after
resolution of hemorrhage correlates strongly with the location of the defect
and whether if it is foveal or extrafoveal (109,113,116).
Management of traumatic choroidal rupture centers upon monitoring for
and prompt treatment of secondary choroidal neovascularization (CNV).
CNV has been reported in 10% to 20% of choroidal ruptures, and the risk has
been reported to be higher among those with longer choroidal ruptures
(113,116). CNV may be evident by clinical evidence of a greenish-gray CNV
membrane (CNVM) under the retina or by associated intraretinal or
subretinal fluid or hemorrhage. Evidence of a membrane or intraretinal or
subretinal fluid on OCT or leakage consistent with CNV on fluorescein and
or indocyanine green angiogram supports the finding of CNV. Historically,
laser photocoagulation or photodynamic therapy was employed for CNV
from choroidal rupture (122). More recently, case reports in adults and in
children suggest efficacy of intravitreal anti–vascular endothelial growth
factor (VEGF) therapy for CNV in choroidal rupture, often with only a single
or few injections (123–126). Anti-VEGF therapy for CNV secondary to
traumatic choroidal rupture is off label and has certain challenges including
the need for anesthesia in some cases and lack of long-term safety. It is also
important to be aware of the risks associated with pregnancy in young girls.
Moreover, as discussed in Chapter 53, while anti-VEGF injections have been
employed for retinal vascular disorders in children (e.g., retinopathy of
prematurity), their use in the pediatric population exhibits certain challenges,
including the necessity in some cases for anesthesia and the lack of
established data on the long-term systemic safety of these agents. The
discussion with the parents should include these considerations.
Traumatic Chorioretinal Rupture

Traumatic chorioretinal rupture, also known as chorioretinitis sclopetaria or
sclopetaria, is a full-thickness break of the choroid and retina due to a high-
velocity projectile such as a bullet or BB striking or passing adjacent to, but
not penetrating, the globe. There is a rapid deformation of the globe either by
a high velocity object striking the eye or by the shock wave of the passing
high velocity projectile. Since the ocular tissues are viscoelastic materials,
they are subject to stress/strain rate dependence, whereby higher rates of
deformation due to high velocity projectiles or their shock waves result in
particularly elevated levels of stress on deformed tissues. The rupture occurs
in areas where the resulting shear stress exceeds tensile strength. It is
postulated that in the setting of high velocity projectile injury, the retina and
choroid deform and retract and thus rupture as a single unit (127).
Clinical features of traumatic chorioretinal rupture include extensive
choroidal and retinal hemorrhages due to rupture of the choroidal and retinal
vessels. The vitreous may be clear or hemorrhagic. A retinal and choroidal
defect is present, although it may be obscured by hemorrhage, and the sclera
is intact (Figure 69-4). Over time, a white fibrous membrane develops. At
the time of injury, the dramatic contrast between the chorioretinal defect and
hemorrhage and the adjacent edematous retina can mimic a retinal
detachment. Ultrasound can be instructive and usually confirms presence of
attached retina. In fact, the risk of retinal detachment is thought to be low
during the acute period after traumatic chorioretinal rupture, perhaps due to
the firm adhesion between the retina and choroid in the area of the defect,
thus resulting in their rupture as a single unit, presence of a formed posterior
vitreous, and/or a protective effect from the fibrous scar commonly found at
the edge of the defect (127). However, there are limited studies with long-
term follow-up, and retinal detachments have been reported after traumatic
choroidal rupture, both within weeks of the injury (128) and years later (127).
The mechanism for such retinal detachments includes retinal breaks at the
edge of the rupture site, retinal breaks secondary to traction from the
proliferative tissue at the rupture site, or peripheral retinal breaks distant from
the rupture site, which may or may not be related to the traumatic
chorioretinal rupture (127,128).
FIGURE 69-4 Traumatic chorioretinal rupture. Fundus

photograph of traumatic chorioretinal rupture reveals
extensive retinal hemorrhages and a retinal and choroidal
defect revealing underlying intact sclera superotemporally.
CONCLUSION
Ocular trauma disproportionately affects the pediatric population with over
one-third of blinding eye injuries in the United States Eye Injury Registry
occurring in the first two decades of life. Children pose unique challenges to
medical history taking and ophthalmic examination. Moreover, the pediatric
ocular anatomy is distinct from that of an adult and has significant
implications for the clinical manifestations, management, surgical approach,
and prognosis of these patients. Finally, amblyopia is a significant
consideration requiring timely intervention of the acute injury to prevent
irreversible occlusive amblyopia and upon appropriate long-term
management of refractive error and amblyopia to optimize visual outcomes.
ACKNOWLEDGEMENTS
MPG is supported by an unrestricted departmental grant from Research to
Prevent Blindness.
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70
Nonaccidental Head Trauma
George Caputo, and Wei-Chi Wu
DEFINITION
In 1971, Guthkelch reported the cases of two infants with subdural hematoma
suggesting for the first time that babies could be injured by shaking (1). Prior
to this report, in 1968, Ommaya was the first to identify cerebral lesions
induced by a whiplash mechanism on nonhuman primates (2).
Nonaccidental head trauma (NAHT) is also called nonaccidental head
injury (NAHI), or shaken baby syndrome (SBS), but this last term does not
account for all the mechanisms implied in the observed lesions and does not
clearly identify the fact that these injuries result from child abuse. The
specificity of the syndrome is a triad involving the brain, subdural space, and
retina; acceleration and deceleration with or without impact are the main
factors explaining the variety and severity of neurologic damage.
NAHI is the leading cause of head injury in the United States accounting
for 80% of head injury in children under 2 years. It is characterized by the
severity of the outcome; 30% of victims die and two-thirds of survivors
suffer from permanent neurologic damage, including blindness and cognitive
and motor impairment (3–7). The ophthalmologist plays a key role in the
diagnosis that can lead to social and forensic consequences.
PREVALENCE
NAHI has become the leading cause of death by brain injury in children in
the United States with an incidence of 24 to 30/100,000 children leading to
about 1,200 to 1,400 cases per year.
The median age encountered in NAHI is 3.9 to 4.6 months, and most of
the children are <1 year in age with exceptional cases in children up to 5
years of age (4,8). The age range corresponds to the peak age when infants
cry the most (Table 70-1).
Table 70-1 Main NAHI case studies
RH, retinal hemorrhages; RD, retinal detachment.
PATHOPHYSIOLOGY OF BRAIN
LESIONS
Brain lesions consist of marked ischemia-inducing cerebral edema and
subdural hemorrhage (SDH); the latter finding is present in 90% of the cases
and is highly specific (10) (Figure 70-1). Diffuse axonal damage in the upper
spinal cord in the cervical region is probably responsible for apnea in these
infants (11,12). Hypoxia–ischemia and secondary brain swelling are then
induced and constitute the main abnormalities found on postmortem
histopathologic examination of the most severe cases. Acceleration and
deceleration by a whiplash mechanism is, on the other hand, the main cause
of subdural bleeding postulated to be caused by rupture of the bridging veins
between the dura and the brain (10).
FIGURE 70-1 Baby girl, 1 month old. Nonaccidental
head trauma. A:CT scan. B:MR sagittal slice, T1-weighted
sequence. C:MR axial slice, diffusion-weighted sequence.
Diffuse subdural and subarachnoidal hematomas (arrows)
located into the tentorium, in the interhemispheric space,
and over the left hemisphere (A, B). Parenchymal anoxic
ischemic injuries in the left frontal and occipital lobes (C).
(Courtesy of Pr C. Adamsbaum.)
The conjunction of hypoxia, venous congestion in immature vessels,

intracranial hypertension, and general hypertension can explain the subdural
and retinal hemorrhages.
SDH is reported to be a thin layer at the surface of the cortex probably
secondary to ischemia or a thicker collection when bridging veins rupture
(12,13).
Finite mathematical models of the eye and its motion show that shaking
at a rate of 4 cycles/s applies a 50-fold increased force to the retina because
of inertia and resonance compared to a single trauma from a 45-cm fall onto
hard ground (13). Diffusion-weighted magnetic resonance imaging (MRI)
studies have provided evidence to support the importance of ischemia in
NAHI by a hyperintense signal from brain hypoxia early in the evolution
(14,15).
OCULAR LESIONS
Diagnostic Studies
If available, a fundus wide-field photograph is taken with a portable retinal
camera, such as RetCam, or a detailed drawing is performed after the
examination is completed. Ultrasound can be helpful in distinguishing retinal
configuration, whether from a retinal detachment, traction, or premacular
hemorrhage beneath a vitreous hemorrhage. Fluorescein angiography is
useful to evaluate for retinal nonperfusion (Figure 70-7B) (16).

At first examination, the child can be hypotonic, with a history of vomiting,
irritation, or seizures, or have severe airway obstruction and benefit from
resuscitation maneuvers with tracheal intubation in the emergency room. Less
than 50% of the perpetrators admit the abusive act (3–5). The father is more
likely to confess than the babysitter. The history told is often incompatible
with the severity of the lesions present. Shaking typically occurs in the setting
of inconsolable crying, and there is no clear interval between symptoms and
abuse. Direct trauma or other fractures are seen in 30% to 50% of the cases
(4,17–19). After stabilizing the clinical status of the infant, a general
examination of the baby is performed to record bruises, signs of trauma, and
the status of long bones prior to performing MRI.
Ophthalmologic assessment is required promptly to help confirm the
diagnosis, and general evaluation is performed to rule out other medical
causes. The eye examination should be conducted as soon as possible by a
trained ophthalmologist using a speculum and topical anesthesia. Facial and
lid bruises are carefully reported. Visual acuity assessment of the ability to
fix and follow is determined. Anterior segment status is also recorded, and
fundus examination is performed using indirect ophthalmoscopy with a 28-
or 30-diopter lens following pupil dilation when possible.
Retinal Hemorrhages
Retinal hemorrhages are the most specific and common findings of NAHI.
They are present in 70% to 93% of the cases (3–7,9,11,18). Unilateral retinal
hemorrhages are reported in 12% to 40% of the cases (6,20) Multiple studies
support the finding of hemorrhages specifically in NAHI. Pierre-Kahn et al.
compared the presence of intraocular hemorrhage in a series of 241
consecutive infants hospitalized for an SDH across three groups based on the
degree of certainty that they had been shaken (7). Intraocular hemorrhage was
seen in 77.5% of infants presumed to have been shaken, in 20% of infants
with signs of head trauma without a relevant history of accidental trauma, and
in none of the infants with proven severe accidental trauma. The authors
concluded that intraocular hemorrhages are suggestive of SBS. A review by
Vinchon et al similarly reported retinal hemorrhages in 53% to 80% of
children with NAHI, but in only 0% to 10% of children following accidental
trauma (21). From a prospective analysis of 150 cases of head trauma, the
authors calculated a sensitivity of 75%, specificity of 93.2%, positive
predictive value of 89.4%, and negative predictive value of 82.9% for retinal
hemorrhage and child abuse (21).
Differential Diagnosis for Retinal Hemorrhages in

Infants and Young Children
In 182 cases of children under 2 years of age admitted for a first episode of
seizures, Curcoy reported that only two had retinal hemorrhages (22). In
accidental brain injury with direct brain trauma, the proportion of children
presenting with retinal hemorrhages was <10% (11,14) and usually existed
from very specific trauma with a predominant crush factor. Convulsions and
vomiting are not recognized to induce retinal hemorrhages. Other causes of
retinal hemorrhages in infants and young children are rare and are listed in
Table 70-2 (23).
Table 70-2 Causes of intraretinal hemorrhages in

young children
Histopathologic studies of postmortem eyes have shown that retinal

hemorrhages in the most severe NAHI are diffuse and extend from the
posterior pole to the far periphery (Figure 70-2). They mainly exist in the
superficial layers under the inner limiting membrane (30% to 38%) or the
nerve fiber layer, but have been reported to be intravitreal and retrohyaloidal
(47%) (Figure 70-3). More rarely, the hemorrhage can be intraretinal or
subretinal (10%). In 15% of cases, hemorrhages present as Roth spots with
fibrin in the centers of the hemorrhages (Figure 70-4). Recent OCT studies
have confirmed in vivo pathologic findings, such as localized detachments of
the posterior hyaloid at the posterior pole, double-layered epiretinal
membranes, macular holes, retinal folds, and retinal schisis in the area of
hyaloidal adherence that exerts vitreous traction, as part of the
pathophysiology (24,25).
FIGURE 70-2 Two-month-old infant with NAHI: diffuse

hemorrhages in all retinal fields and subhyaloidal posterior
hematoma. Circular retinal fold around the posterior pole.
FIGURE 70-3 Mild form of NAHT with localized
macular retrohyaloidal hematoma.
FIGURE 70-4 Fundus photograph of a 3-month-old baby
who developed retinal hemorrhages from NAHI, showing
the representative finding of Roth spots, with fibrin in the
centers of the hemorrhages.
Schisis
Retinoschisis due to NAHI has been described in severe cases and has been
shown in OCT studies of affected babies (24,26). The features reported are
multilayered retinoschisis affecting areas where the vitreous remains
attached. Vitreous traction could explain the mechanism of these lesions.
Macular Changes
Besides epiretinal or subhyaloid hemorrhages, macular holes and macular
pseudoholes have been reported in rare cases and are distinguished by OCT
(24,27). Epiretinal membranes are a common finding in survivors of NAHI
(28) and often present as a two-layered structure, corresponding to a
proliferating structure and the internal limiting membrane. Also, bilateral
traumatic macular holes were found as a complication of NAHI (Figure 70-
5).
FIGURE 70-5 A 7-month-old infant with NAHI presented
with bilateral large macular holes formation. Both right
eye (A) and left eye (B) showed diffuse retinal
hemorrhages with large traumatic macular hole.
Neovascularization and Retinal Ischemia

In 1999, Brown reported the first case of optic disk neovascularization after
NAHI, which developed 4 months after the shaking episode. Initially the 4-
month-old baby had diffuse extensive retinal hemorrhages and intravitreal
hemorrhage and, after 4 months, had documented florid optic disk
neovascularization on angiography performed under anesthesia (29). We
reported three cases of tractional retinal detachment due to extensive
neovascular proliferation in infants having a history of NAHI (30) (Figure
70-6). Wide angle angiography performed in patients after NAHI have shown
wide areas of peripheral retinal ischemia (31) raising the necessity of long
term follow-up and consideration of laser retinal ablation in these cases
(Figure 70-7). A shearing mechanism due to vitreous traction or secondary
retinal ischemia due to hypoxia may contribute to these findings (16,26).
FIGURE 70-6 Total retinal detachment with diffuse

neovascularization and ridge-like structure in the mid
periphery, in a 7-month-old NAHT baby.
FIGURE 70-7 A:Fundus wide-field image of a 6-month-
old baby, 4 months after initial NAHI episode: major
retinal fold with severe contracted neovascularization.
B:Fluorescein angiogram of the same patient showing
large areas of nonperfusion and vascular buds at the
periphery.
Retinal Folds
Retinal folds have been described concentric to the posterior pole (Figure 70-
8) with vitreous being adherent to the top of the fold and to the vitreous base
(32). These findings initially reported in histopathologic studies have been
confirmed by in vivo OCT imaging (26).
FIGURE 70-8 Fundus photograph of a 4-month-old infant
with NAHI, showing diffuse retinal hemorrhages with
representative circular hypopigmented retinal fold.
Retinal Detachment
Retinal detachment has been reported and occurs in the most severe cases. In
a histopathologic study of 16 infants that died of NAHI, Green found 10
cases of retinal detachment (18).
Retinal detachment as reported above can be related to a tractional
mechanism; a rhegmatogenous cause is involved in cases of direct ocular
blunt trauma with a high proportion being from retinal dialyses or giant
retinal tears (33).
Correlation to Severity of Neural Damage

The extent of retinal hemorrhages and associated ocular lesions has been
correlated to the severity of NAHI and to mortality. The presence of a
subdural hematoma is also linked to the presence of retinal hemorrhages (11).
Mills reported a higher mortality in patients presenting with retinoschisis or
retinal detachment (34). Wu et al. identified several independent risk factors
of mortality including respiratory distress or apnea, subarachnoid
hemorrhage, vomiting, and macular retinoschisis in an Asian population.
More specifically, the retinal finding of retinoschisis may increase the
likelihood of mortality with an odds ratio of 8.9 (9).
PATHOPHYSIOLOGY AND
SPECIFICITY OF OCULAR LESIONS IN
NAHI
The role of the vitreous has been emphasized, and the lesions observed have
been attributed to the whiplash mechanism of acceleration–deceleration
present following shaking. The strong adherence of the vitreous body to the
retina in babies and the repeated violent movements of the vitreous are held
responsible for shearing of the superficial retinal vessels predominantly
resulting in diffuse hemorrhages. The hypothesis of an associated retinal
ischemic mechanism in NAHI is supported by the fact that chest compression
alone does not explain diffuse retinal hemorrhages as seen in NAHI. In a
prospective trial evaluating retinal hemorrhages after resuscitation maneuvers
inducing chest compression in 43 patients, Odom et al reported bilateral
multiple small punctate retinal hemorrhages that appeared different from
those in NAHI (35).
Based on histopathologic study of brains of infants who died of NAHI,
Geddes in 2001 proposed that the primary feature of brain lesions in NAHI
was anoxia secondary to brainstem lesions in the cervical region. The latter
results in ischemic cerebral edema and intracerebral infarction and bleeding
as well as subdural hematomas. At the retinal level, prolonged tissue
ischemia could be implicated in enhancing vascular extravasation. A sudden
increase in retinal venous pressure and arterial blood pressure due to cerebral
ischemia and brain edema could lead to vascular leak such as what occurs in
a Vasalva-type mechanism. This pathology would be enhanced by hypoxic
lesions in the infants’ immature vessels. Infants suffer from shaking early in
the first year of age at a time when presumably the retinal vasculature is
particularly fragile and immature. More recently, a comparative
histopathologic study of 18 cases of SBS showed diffuse orbital shearing
lesions from acceleration followed by deceleration and confirmed the effect
from whiplash (10).
The ophthalmologist plays a key role in the diagnosis and management of
NAHI by detecting ocular involvement and, above all, retinal hemorrhages.
Nonophthalmologists have an 87% detection rate of retinal hemorrhages
making the recourse to the specialist mandatory (36,37).
Retinal hemorrhages resolve in 2 to 4 weeks (Figure 70-9), and visual
gain or maintaining fix and follow vision is seen in 75% of the surviving
patients (7,38). This outcome points to planning for vision rehabilitation in
follow-up, especially in cases with asymmetric vitreous hemorrhage. Kivlin
reported good visual outcome in 68% of survivors, although 19% didn’t
fulfill follow-up. Visual impairment can be severe with the presence of
extensive retinal hemorrhages, retinal folds, retinal scarring, retinal schisis,
and retinal detachment, and is seen in 10% of the patients. Visual cortex
involvement or optic atrophy also explains severe visual loss in the remaining
neurologically impaired infants (5). In infants with subhyaloid hemorrhages
involving the macula, awake time can be spent upright such as in a jumper to
allow settling of the blood out of the visual axis.
FIGURE 70-9 Fundus wide-field image of a child who
sustained retinal hemorrhages from NAHI, demonstrating
the appearance of the fundus several weeks later with
resolution of the hemorrhages.
Indications
In case of intravitreal hemorrhage or subhyaloid lasting more than a few
weeks, a lens-sparing vitrectomy can be performed. The surgical procedure
will be anticipated in case of bilateral involvement.
Secondary epiretinal membrane formation requires surgical removal after
carefully assessing retinal and subretinal changes by repeated OCT
examinations. It appears more judicious to delay the surgical removal to
allow safer dissection with less risk of retinal injury during the procedure
because of a better plane separation after several weeks’ evolution. Wide-
field retinal angiography allows identification of peripheral retinal ischemia
that should be addressed by photoablation of the ischemic retina by indirect,
external, or endocular laser during surgery if performed, to avoid extensive
neovascular proliferation and secondary retinal detachment. Prolonged
follow-up is necessary to address these secondary complications.

Vitrectomy is performed in case of long-lasting intravitreal hemorrhage,
secondary epiretinal membrane, or more rarely retinal detachment once
neurologic status is settled and stabilized and usually several weeks after the
initial episode. Lens-sparing vitrectomy is done through sclerotomies
positioned according to the child age (see Chapter 58). Transvitreal
vitrectomy is most commonly used, and a posterior vitreal detachment is
performed at the posterior pole when a membrane is peeled. In the case of a
severe intravitreal hemorrhage, after ruling out a retinal detachment by
preoperative ultrasound examination, the vitrectomy is conducted centrally
and continued posteriorly until a retinal plane is recognized before enlarging
the removed vitreal cone. It is recommended to remove inner limiting
membrane when peeling the epiretinal membrane, as the latter is due to
fibrosis of the hematic content. Because hemorrhages can occur in the
preretinal, intraretinal, and/or subretinal space, secondary fibrosis may also
occur in any of these locations.
Equipment
For evaluation during examination: wide angle fundus camera, fluorescein
angiography system, ultrasound
Small-gauge vitrectomy system
Vitreoretinal forceps for membrane peeling in the case of ERM
Indirect, external microscope adaptor or endolaser
Results after vitrectomy are unpredictable because of the varying underlying
retinal and CNS neurologic lesions (39). Epiretinal membrane peeling is also
achievable, when necessary, but may have limited functional outcomes.
NAHI is a devastating disease, and primary prevention is a main goal in
the future to avoid induced impairment. Primary prevention strategies have
proven effective. Dias set a milestone in a very well-designed prospective
study in 2005, in which there was a 47% reduction in the incidence of SBS
after parents of newborns were provided information on SBS following birth
of their infant and prior to discharge from hospital. The parents were given a
brochure, watched a videotape, and signed a commitment statement with only
minimal demands on hospital staff resources (40).
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ATLAS A
Hemorrhages
George Caputo
Hemorrhages within the posterior segment of the eye can involve the
vitreous, subhyaloid space, inner retina, subretinal space, and choroid. The
determination in children is not always easy or as helpful as it is in adults.
History and specific patterns of hemorrhage become essential in determining
the diagnosis. Long-standing hemorrhage can become khaki colored as
hemoglobin is lost.
Table A-1 shows diagnoses responsible for hemorrhages, while the
following figures are examples of conditions with various hemorrhages in
infants and children. The reader is referred to specific chapters for greater
detail.
TABLE A-1 Diagnoses responsible for retinal and

vitreous hemorrhagesa
aThis table reflects the most common presentations and associations for the listed conditions. There are
always exceptions, and as we learn more about the genetic and phenotypic associations, these tables
may change.
NAHI, nonaccidental head injury; XLRS, X-linked juvenile retinoschisis; ROP, retinopathy of
prematurity; FEVR, familial exudative vitreoretinopathy; RD, retinal detachment.
FIGURE A-1 A 5-month-old male baby with
nonaccidental head trauma presenting with a subhyaloidal
subfoveal hemorrhage expanding into the vitreous (see
also Chapter 70).
FIGURE A-2 Retinal hemorrhages with central whitening
in nonaccidental trauma (see also Chapter 70).
FIGURE A-3 Severe nonaccidental trauma in a 3-month-
old baby, with diffuse hemorrhages in all retinal fields and
circular retinal fold (see also Chapter 70). (Courtesy of Pr
Speeg, Strasbourg.)
FIGURE A-4 Resolution of retinal hemorrhages in a 4-
month-old nonaccidental head injury, showing preretinal
hemosiderin deposits (see also Chapter 70).
FIGURE A-5 Panoramic view of a collected
retrohyaloidal hemorrhage in a 16-year-old boy due to a
Valsalva maneuver.
FIGURE A-6 Noncollected preretinal hemorrhage in an
8-year-old child due to Valsalva mechanism.
FIGURE A-7 Preretinal hemorrhage in a 10-week-old
baby with aggressive posterior retinopathy of prematurity.
Note vessel tortuosity and dilation in all quadrants around
the optic nerve (see also Chapters 52–54).
FIGURE A-8 A:Reabsorbing retrohyaloidal hemorrhage
in a 17-year-old boy secondary to dorsal trauma. B:Optical
coherence tomography of the same patient showing
preretinal condensation and constitution of a preretinal
membrane (see also Chapter 69).
FIGURE A-9 A:A 6-year-old child presenting an
intravitreal hemorrhage secondary to XLRS. B:Diagnosis
was confirmed by the presence of typical lesions in the
fellow eye associating inferior and macular schisis (see
also Chapter 34).
FIGURE A-10 A–C:A 4-year-old boy with acute
leukemia presenting long-lasting intravitreal hemorrhage
treated by 25-G vitrectomy after six cycles of
chemotherapy and biologic remission. Hemorrhage is
dense, and macular scarring is visible. Visual recovery was
20/40. (See also chapters 47 and 49).
ATLAS B
Macula
George Caputo
Macular lesions can result from a very wide variety of diseases ranging from
retinal degeneration to vascular lesions and folds. Table B-1 summarizes the
main differential diagnoses and classification of the different pathologies
observed, while illustrated by clinical cases that appear below. The reader is
referred to specific chapters for greater detail.
TABLE B-1 Ocular conditions involving the

maculaa
may change.
FEVR, familial exudative vitreoretinopathy; ERM, epiretinal membrane; RPE, retinal pigment
epithelium; PR, photoreceptor; ROP, retinopathy of prematurity.
CONGENITAL MACULAR
DYSTROPHIES
FIGURE B-1 Albinism. A, B (OD, OS):A 15-year-old
boy with ocular albinism. Fundus appearance is
hypopigmented throughout allowing abnormal visibility of
the choroidal vascular meshwork. Visual acuity is 20/80.
C (OD), D (OS):Autofluorescence picture and late-phase
indocyanine green (ICG) angiogram OS show small fleck
pattern hyperfluorescence in the foveal area. E (OD), F
(OS):Macular hypoplasia is confirmed by optical
coherence tomography (OCT): partial loss of central
foveal depression and heterogeneity of the outer nuclear
layer in the foveal region (see also Chapter 23).
FIGURE B-2 Best dystrophy. A 15-year-old boy with
Best dystrophy. VA 20/20 in both eyes. A:OD presents
with typical egg yolk, yellowish material starting to
precipitate. B:OS presented 2 years before with choroidal
neovascularization treated by surgical excision: fibrotic
nonactive, nonevolving remnant in upper nasal aspect of
the fovea in OS. C:OCT scan OD showing sub-RPE
material and normal retinal layers. D:OCT scan OS with
stable sub-RPE deposition and possible subretinal fluid
FIGURE B-3 Cone dystrophy. A 17-year-old patient with
cone dystrophy. A, B (OD,OS):Autofluorescence images
show perifoveal hyperfluorescence and foveal mottling.
OCT shows atrophy of the external retinal layers in the
foveal region (see also Chapter 26). Green line through
macular image at left represents location of axial OCT
scan demonstrated at right.
FIGURE B-4 Hypermetropia. A 12-year-old boy with
nanophthalmos and 14 diopters of hypermetropia; vision is
20/100 in both eyes. A, B:Right eye and left eye showing
macular fold. C, D:OCT scans confirm the fold and
anatomical preservation of outer layers like the IS/OS line.
Microcavities are visible in the outer nuclear layer. Green
line through macular image at left represents location of
axial OCT scan demonstrated at right.
EXUDATIVE MACULOPATHIES
FIGURE B-5 Coats. A–D:A 6-year-old boy presenting
with extensive subretinal exudates at the posterior pole,
secondary to Coats disease; vascular dilations are present
in midperiphery. The subretinal deposits extend under the
fovea; well-applied laser treatment resulted in resorption
of exudates 3 months later. E, F:Fundus photograph of a
9-year-old boy with fibrotic macular scarring. Fluorescein
angiography shows peripheral leakage at the posterior
margin of laser scars (see also Chapter 64).
FIGURE B-6 IRVAN. An 8-year-old girl with IRVAN
syndrome: idiopathic retinitis vasculitis, aneurysm, and
neuroretinitis. A:Posterior pole on initial presentation:
extensive posterior exudation is evidenced. B:Subretinal
neovascularization developed 6 months after treatment.
C:Wide-angle angiography shows peripheral retinal
ischemia treated with laser.
FIGURE B-7 Neuroretinitis. A:An 8-year-old girl
presenting unilateral neuroretinitis: papilledema and vessel
congestion are present. B:Three weeks later, small
circinate exudates appeared after resorption of the edema.
Patient was treated for cat scratch disease due to
Bartonella (see also Chapter 47).
PRERETINAL MACULOPATHIES
FIGURE B-8 Epiretinal membrane. A, B:Wide-field view
of an epiretinal membrane in a 10-month-old baby with a
history of intravitreal hemorrhage. B:Late-phase
angiography shows no leakage in the macular area.
C:Idiopathic epiretinal membrane in a 7-year-old boy;
converging folds of the inner limiting membrane (see also
chapter 67). D:Outer retinal layers are preserved as seen in
OCT. E:Thick central ERM in a 5-year-old boy. F:OCT
confirms the retracted fibrotic aspect of the membrane,
which appeared fibrovascular during surgery (see also
Chapter 67). Green line through macular image at left
represents location of axial OCT scan demonstrated at
right.
FIGURE B-9 Hamartoma. A:Red-free photograph of a
combined hamartoma of the RPE and choroid around the
optic nerve in a 9-year-old boy. B:Vision is 20/200, and a
thick epiretinal membrane is seen on the OCT scan (see
chapters 46 and 67). C, D:After vitrectomy and membrane
peeling, the hamartoma around the optic disc is more
clearly visible and macula is less ectopic: vision improved
to 20/60 (see also Chapter 46). Green line through macular
image at left represents location of axial OCT scan
demonstrated at right.
FIGURE B-10 Macular hole. Color SLO wide-field
picture of a macular hole in an 18-year-old boy after ball
closed trauma. The hole is wide, oval shaped, with
underlying pigment epithelial and atrophic changes, and
the small operculum appears green because of its
intravitreal position (see also Chapter 69).
FIGURE B-11 Folds. A, B (OD,OS):A 4-month-old girl
presenting retinal dysplasia in both eyes. Pigment
epithelial changes and vessels are visible in retinal fields
outside the fold. C, D:Fold due to persistent fetal
vasculature; vitrectomy was performed to release traction
from the fibrous stalk. Lens was aspirated and implant
placed. (Diagnosis based on unilateral condition.)
E:Retinal fold believed to be due to ROP based on
medical history. F, G:Retinal fold due to familial
exudative retinopathy (FEVR) in a 4-year-old boy with no
history of prematurity. Fellow eye has no major vascular
changes, but wide-field angiography shows anterior
nonperfused retinal areas (see also Chapter 65).
INFLAMMATORY MACULOPATHIES
FIGURE B-12 Toxocariasis. A 2-year-old boy with

toxocariasis: severe posterior vitreoretinal changes
associated with subretinal cyst localization (see also
Chapter 47).
FIGURE B-13 Toxoplasmosis. A:A 17-year-old boy
presenting foveal scar due to toxoplasmosis. B:OCT scan
shows central atrophy of all retinal and pigment epithelial
layers. C:Two years later, reactivation of the scar showing
marked edema and exudation. D:OCT scan confirms
serous retinal detachment and subretinal material around
the atrophic scar. Green line through macular image at left
right. E:After complete healing, the atrophic scar appears
larger (see also Chapter 47).
FIGURE B-14 Solar retinopathy. An 18-year-old girl with
solar retinopathy. A:Mild central pigment epithelial
change. B:OCT scan shows very localized central defect
in pigment epithelial explaining the microscotoma. Green
axial OCT scan demonstrated at right.
DEGENERATIVE MACULOPATHIES
FIGURE B-15 Retinitis pigmentosa. A 13-year-old boy
presenting with retinitis pigmentosa. Vision is 20/40 OU,
and no major pigmentary changes are visible. Foveal
reflex is absent (A,B), and autofluorescence of both eyes
shows mild hyperreflective ring around the fovea (C,D).
OCT scan shows mild atrophy and changes in external
nuclear layers and outer segments (see also Chapters 26).
Green line through macular image at left represents
location of axial OCT scan (E,F) demonstrated at right.
FIGURE B-16 X-linked retinoschisis. An 8-year-old boy
with bilateral 20/200 vision. A, B:Color photograph of the
posterior pole showing reflect alterations and central
foveal cyst. C, D:OCT scans show cystoid spaces in outer
and inner nuclear layers and large central space (see also
Chapter 34). Green line through macular image at left
right.
MACULAR HOLES
FIGURE B-17 Traumatic macular hole in a 10-year-old
boy following recent blunt trauma. A:Infrared image of
right eye shows disruption of pigment (left) and OCT
(right) shows full-thickness macular hole. Visual acuity is
20/150. B:Two weeks following ocriplasmin injection in
right eye 30 minutes before pars plana vitrectomy with
posterior hyaloidal removal and 80% fill of 10% C3F8 gas
followed by postoperative face down positioning. Visual
acuity improved to 20/100 and inner retina appears to be
bridging hole. C:Eight months after surgery, visual acuity
is 20/50 showing small defect in foveola and less
disruption of ellipsoid zone (see also Chapter 69).
(Courtesy Mike Trese.)
ATLAS C
Pigmentary Changes
George Caputo
Pigmentary changes can be seen in a wide spectrum of diseases, and it is

important to consider degenerative conditions. Family history can be helpful
when determining the diagnosis in some cases. The reader is referred to
specific chapters for greater detail.
FIGURE C-1 Albinism. Optomap wide-field fundus
photography of a 14-year-old girl with ocular albinism.
Note the visibility of the choroidal vascular meshwork (see
also Chapter 23).
FIGURE C-2 Choroideremia. A–E:Central and peripheral
fundus photography of a 15-year-old boy suffering from
late-stage choroideremia: choroidal atrophy sparing a
small area of the macula larger in OD (A, B) than in OS
(C–E). Vision is 20/200 OD and 20/400 OS. F–K:Fundus
pictures of the 8-year-old sibling of the above patient
presenting an early stage of choroideremia: color pictures
show mild pigmentary changes and macular
hyperpigmentation (F, G), best seen in red photographs
(H, I), red-free fundus pictures enhancing the scattered
choroidal vascular meshwork (J, K) (see also Chapter 26).
FIGURE C-3 Fundus albipunctatus. A 17-year-old patient
with fundus albipunctatus. Patient suffers from night
blindness. A1 (OD), A2 (OS):Fundus color photographs
showing white flecks sparing the posterior pole. B–
D:Autofluorescence of the white spots, also seen around
the optic nerve (see also Chapters 26 and 29). E, F:Optical
coherence tomography (OCT) scans of the macula show
preservation of all the retinal layers. Green line through
scan demonstrated at right. G:Peripheral OCT scan in the
zone of the white dots shows mottling in the outer
segments of the photoreceptor with a heterogeneous
appearance of the inner segment/outer segment (IS/OS)
line (see also Chapters 26).
FIGURE C-4 Leber congenital amaurosis. A, B:Wide-
field fundus photograph of a 2-year-old boy with
congenital amaurosis. Posterior pigmentary changes and
honeycomb-like aspect of the peripheral retinal fields,
narrowing of the vessels, and optic nerve edema are
clearly visible in this severely affected patient (see also
Chapters 27 and 28).
FIGURE C-5 Retinitis pigmentosa. A, B:An 18-year-old
girl suffering from severe retinitis pigmentosa associated
with macular edema and peripheral pigmentary
osteoclasts. OS (B) presents with macular pigmentary
changes due to a full-thickness macular hole. C, D:A 12-
year-old boy with retinitis pigmentosa without osteoclast
formation. Vision is 20/200. E, F:OCT scans of the
maculae show loss of the photoreceptor layer and
disorganization in the foveal layers. Green line through
scan demonstrated at right. G, H:Autofluorescence of the
macular area shows a mild hyperautofluorescent area
around the fovea in both eyes (see also ebook Chapter 4).
FIGURE C-6 Stargardt. A, B:Red-free photograph and
OCT scan of an end-stage 20-year-old patient with
Stargardt disease showing diffuse atrophy of the outer
retinal layers and retinal pigment epithelium. Large
scattered choroidal vessels are visible on the scans. Green
axial OCT scan demonstrated at right. C,
D:Autofluorescence imaging shows hypoautofluorescence
of the atrophic areas and hyperautofluorescence of the
pisciform subretinal deposits in the retinal fields (see also
Chapter 30).
FIGURE C-7 Toxoplasmosis. A:Color fundus photograph
of a chorioretinal scar in a 3-year-old boy. Note the
atrophic borders. B:Fluorescein angiography shows mild
hyperfluorescence by window defect and no late staining
FIGURE C-8 Congenital pigment epithelial hypertrophy.
A 2-year-old pseudophakic boy presenting with large
homogeneous areas of pigment epithelial hypertrophy.
ATLAS D
Posterior Segment Masses
George Caputo
Table D-1 presents differential diagnoses for posterior segment masses, while
illustrated case studies follow below. The reader is referred to specific
chapters for greater detail.
Table D-1 Diagnoses responsible for segment

massesa
may change.
FEVR, familial exudative vitreoretinopathy.
FIGURE D-1 Coats. A:Fundus image of a 4-year-old boy
presenting with a retinal detachment due to Coats disease
with massive subretinal exudation (stage 4 Coats disease).
B:The same patient following two sessions of laser
photocoagulation to dilated vessels followed by external
subretinal drainage. Subretinal fibrosis secondary to
condensation of the exudation is molded by the retina (see
also Chapter 64).
FIGURE D-2 Retinal dysplasia. A:A 6-month-old infant
with bilateral retinal dysplasia. Fibrous stalk, partial retinal
detachment, and intravitreal hemorrhage are present in this
eye. B:The same patient after vitrectomy and surgical
ablation of the fibrovascular remnants: retina has flattened
back, leaving the pigmented demarcation line of the
previous detachment and an avascular dysplastic retina.
Fixation is present (see also Chapters 35 and 65).
FIGURE D-3 Combined hamartoma of pigment
epithelium and retina. A:Fundus photograph of a 7-year-
old boy presenting a combined hamartoma of pigment
epithelium and retina complicated by an epiretinal
membrane. Note the peripheral traction and the macular
fold and traction at the limit of the lesion. B:Three months
after vitrectomy and membrane peeling, the macular area
is flatter and peripheral traction less. Vision improved
from 20/400 to 20/200 (see also Chapters 46 and 67).
FIGURE D-4 Angioma. Fundus photograph of a
peripheral fibrotic angioma in a 12-year-old boy.
Exudation is surrounded by mild atrophy and pigment
epithelium changes (see also Chapter 46).
FIGURE D-5 Choroidal hemangioma. A:Color
photograph of a hemangioma in a 20-year-old man: a light
orange mass is clearly visible on the upper area of the
posterior pole. B:Blue filter picture showing mild
epiretinal changes. C:Red photograph showing the light
mass. D:The red-free photograph is out of focus in the
area of the lesion. E:Optical coherence tomography scan
of the lesion shows the thickness of the mass and
subretinal exudation causing reduction in visual acuity
(20/40) (see also Chapter 46). Green line through macular
FIGURE D-6 Retinoblastoma. A:Anterior segment
photograph of a 6-month-old boy presenting leukocoria for
1 month. A posterior vascularized ill-defined mass is
clearly visible in the posterior segment. B:Fundus
photograph showing total retinal detachment due to a large
retinoblastoma (see also Chapters 44 and 45).
FIGURE D-7 von Hippel disease. A:Color fundus
photograph of a 12-year-old girl presenting a large
superior peripheral retinal hemangioblastoma responsible
for posterior exudation and retinal detachment. Dilated
afferent and efferent vessels irrigating the mass are clearly
visible. General assessment includes cerebrospinal
magnetic resonance imaging and ultrasound abdomen
examination to rule out other localization and
pheochromocytoma. B:Fellow eye is normal with no
objective lesions (see also Chapter 46).
ATLAS E
Flecks and Spots
Michael T. Trese, Antonio Capone Jr, and M. Elizabeth Hartnett
The following are examples of flecks or multiple white spots. The reader is
FIGURE E-1 Acute posterior multifocal placoid pigment

epitheliopathy in a 17-year-old boy with antecedent viral
upper respiratory infection (see also Chapter 47).
FIGURE E-2 Retinitis punctata albescens (see also
Chapter 29).
FIGURE E-3 Pisciform flecks in Stargardt flavimaculatus
FIGURE E-4 Fundus albipunctatus (see also Chapter 29).
FIGURE E-5 Congenital rubella retinopathy (see also
Chapter 47).
FIGURE E-6 Candida chorioretinitis (see also Chapter
47).
FIGURE E-7 Example of sarcoid choroiditis (see also
Chapter 47).
ATLAS F
Abnormal Retinal Vasculature
George Caputo
Table F-1 presents various differential diagnoses for abnormalities in

vascular patterns, while illustrated case studies appear below. The reader is
TABLE F-1 Differential diagnoses for abnormal

vasculaturea
may change.
ROP, retinopathy of prematurity; APROP, aggressive posterior ROP; WGA, week gestational age;
FEVR, familial exudative vitreoretinopathy.
FIGURE F-1 Coats disease. A:Retinal angiography of an
8-year-old boy presenting mild Coats disease: dilation of
the capillary meshwork. B:Fundus image; after laser
treatment of vessel dilations (B1, B2). Laser was also
applied in a grid pattern in anterior region of avascular,
and presumed hypoxic, retina fundus appearance (B3) and
angiographic aspect (C1, C2) after laser scarring. (see also
Chapter 64).
FIGURE F-2 Familial exudative vitreoretinopathy
(FEVR). A:Fundus image of a 9-year-old boy presenting a
total retinal detachment OD secondary to FEVR (stage
5B). The detachment is both tractional with severe
preretinal proliferation and vessel dragging, and
rhegmatogenous. Surgery was unsuccessful. B:Fundus
image of the fellow eye showing mild temporal peripheral
changes and suspicious-looking poorly vascularized retina
confirmed by fluorescein angiography. (C, D) (stage 2B).
Avascular retina was treated with laser photocoagulation.
Neovascular buds are present temporally. E:Right eye of a
4-year-old boy treated with vitrectomy and scleral buckle
for stage 5B FEVR. A persistent retinal fold remains with
regression of subretinal exudation. F:The fellow eye has
stage 3B FEVR with peripheral tractional retinal
detachment due to active neovascularization. This eye was
treated with peripheral laser ablation and scleral buckle
(see also Chapters 42 and 66). G, H:Wide-field fundus
image of right and left eyes of a 13-year-old girl following
treatment for FEVR following some resolution of
exudation. Macula is ectopic, and retinal vessels are
dragged. Peripheral pigment epithelial changes are due to
laser photocoagulation and resolution of exudation (see
FIGURE F-3 Incontinentia pigmenti. A, B:Fundus image
of an 8-month-old infant with retinal detachment OS due
to incontinentia pigmenti. Severe preretinal fibrovascular
proliferation and exudation are apparent in the temporal
retina in association with surrounding avascular retina.
Vitrectomy has a poor prognosis in these advanced cases.
C:Retinal vascularization of the fellow eye is normal (see
also Chapter 36).
FIGURE F-4 Aggressive posterior retinopathy of
prematurity with plus disease. A, B:Aggressive posterior
retinopathy of prematurity in zone 1 in an infant of 33
weeks’ postgestational age (GA) (infant born at 25 weeks’
GA and 520 g birth weight). The ridge is very wide and
the neovascular meshwork mildly elevated. C:Detail at
higher magnification showing vascular dilatation and
tortuosity of vessels at the optic disc in all quadrants.
D:Detail of the “ridge” (junction of vascularized and
avascularized retina) at higher magnification shows
vascular activity (see also Section VIII, Chapters 33–49).
FIGURE F-5 Sickle cell disease. A:Fluorescein
angiogram of a 13-year-old girl with SC sickle cell
anemia. Posterior filling is normal against a very dark
background. B:Interruption of peripheral retinal
vascularization and neovascularization with a “sea fan.”
C:Peripheral avascular retina is surrounded by vascular
buds and thin anastomoses between peripheral vessels.
FIGURE F-6 Vessel tortuosity. Wide-field color fundus
image of a 12-year-old girl with congenital vessel
tortuosity. Visual acuity is normal.
FIGURE F-7 Retinal cavernous hemangioma. Fundus
image of a 12-year-old girl presenting retinal cavernous
hemangioma picked up on routine screening. Vision is
20/20 (see also Chapter 46).
ATLAS G
Retinal Detachment and Schisis
George Caputo
Table G-1 summarizes the main examples of retinal detachments and

retinoschisis, while illustrated clinical cases appear below. The reader is
TABLE G-1 Mechanism of retinal detachmentsa
may change.
FEVR, familial exudative vitreoretinopathy; XLRS, X-linked juvenile retinoschisis.
FIGURE G-1 Coats disease. Fundus image of a 4-year-
old boy presenting with leukocoria and a total retinal
detachment. Vessel and capillary dilatations are clearly
seen on the surface of the retina. Exudation gives it a
yellowish appearance, whereas with retinoblastoma, the
appearance can be whiter (see also Chapter 64).
FIGURE G-2 Coloboma. Wide-field fundus image of a
10-year-old boy presenting with a chorioretinal coloboma
complicated by a retinal detachment. The macular pigment
is visible at the edge of the coloboma in this amblyopic
eye (see also Chapter 22).
FIGURE G-3 Giant tear. A:Fundus image of a 12-year-
old highly myopic boy presenting with a total retinal
detachment secondary to a 180-degree giant temporal
retinal tear. B:Shallow retinal detachment in a 12-year-old
boy with a 100-degree temporal retinal tear without
inversion (see also Chapter 63).
FIGURE G-4 Retinoblastoma. A:Anterior segment image
of a 6-month-old infant with total retinal detachment and
leukocoria. The retinal surface has no vascular changes.
B:Fundus image of the same patient showing the
posteriorly located mass of an endophytic retinoblastoma
stage D (see also Chapters 44 and 45).
FIGURE G-5 Rhegmatogenous retinal detachment. A,
B:Total retinal detachment in a 9-month-old infant. The
presence of proliferative vitreoretinopathy and preretinal
pigmented proliferation confirms the rhegmatogenous
component. The infant sustained blunt ocular trauma (see
FIGURE G-6 Retinopathy of prematurity. A:Stage 4 B
retinal detachment with tractional component, plus
disease, and temporal neovascularization. B:Total
tractional retinal detachment due to stage 5 retinopathy of
prematurity (see also Chapter 55).
FIGURE G-7 Stickler syndrome. A:Color fundus
photograph of a 12-year-old girl with Stickler syndrome.
The vitreous bands prevent focusing on the underlying
retina. B:Wide-field fundus image of the same highly
myopic patient showing numerous vitreous veils and
bands (green appearance). The patient had a retinal
detachment treated with an encircling scleral buckle (see
FIGURE G-8 Sturge-Weber syndrome. A:Total serous
retinal detachment in a 4-year-old girl with Sturge-Weber
syndrome. No proliferative vitreoretinopathy or retinal
vascular anomalies are visible on the retinal surface.
Ultrasonography showed diffuse thickening of the choroid
due to hemangioma. B:Fellow eye fundus image showing
a reddish fundus reflex suspicious of diffuse quiescent
choroidal hemangioma (see also Chapter 46).
FIGURE G-9 X-linked retinoschisis. A 5-year-old boy
with diffuse temporal retinoschisis. A large hole in the
inner retinal wall is visible inferotemporally. No
pigmentation or subretinal proliferation is visible (see also
Chapter 34).
ATLAS H
Optic Nerve
George Caputo
Table H-1 lists conditions responsible for optic nerve conditions or

abnormalities. Illustrated case studies appear below. The reader is referred to
specific chapters for greater detail.
TABLE H-1 Optic nerve abnormalitiesa
may change.
FIGURE H-1 Optic atrophy. A, B:Fundus image of a 12-
year-old boy presenting with bilateral optic atrophy
secondary to a surgically resected craniopharyngioma (see
also Chapter 22).
FIGURE H-2 Coloboma. A, B:Central and peripheral
fundus images of a 20-year-old patient with a chorioretinal
coloboma. C, D:Left eye of the same patient. In both eyes,
there is no macular involvement, and visual acuity is
20/25. E:Wide-field fundus image of a 6-month-old male
infant with very large chorioretinal coloboma involving
the macula. F:Left eye of the same patient presenting with
a shallow retinal detachment involving the macula: visual
prognosis is poor. G:Large central optic nerve coloboma
involving the macula with persistent hyaloid artery
attached to the lens in a 2-year-old boy. Vision is limited
to light perception. The appearance is similar to morning
glory syndrome. H:Deep optic nerve staphyloma with a
tilted optic disc in a 2-year-old girl, corresponding to a
form of coloboma (see also Chapter 22).
FIGURE H-3 Dragged optic disc. A, B:Fundus
photograph of a 12-year-old boy with familial exudative
vitreoretinopathy. Mild dragging of the temporal retinal
vessels due to neovascular peripheral proliferation is
visible (see also Chapter 66).
FIGURE H-4 Optic disc edema. A, B:Mild optic nerve
head edema OU from idiopathic normal pressure
hydrocephalus in a 13-year-old girl (see also Chapter 22).
FIGURE H-5 Leber neuroretinitis. A:Unilateral optic
nerve head edema and serous macular involvement in a
10-year-old girl who suffered vision loss from 20/20 to
20/50. B:The same patient, 3 weeks later with small
stellate exudates from resorption of macular fluid
following regression of the disc edema. Serology was
positive to Bartonella henselae, and specific antibiotic
treatment was administered (see also Chapter 47).
FIGURE H-6 Morning glory anomaly. A:Colobomatous
morning glory anomaly in a 3-year-old girl. The central
optic nerve excavation is deep with steep borders, and the
retinal vessels have a radial configuration. B:Morning
glory anomaly with central glial tissue in a 1-year-old
infant showing radial folds involving the macula (see also
Chapter 22). C:Chronic retinal detachment with subretinal
proliferation in an 18-year-old patient with a
colobomatous morning glory anomaly. D:Fundus
photograph of a 12-year-old girl following surgery for
retinal detachment from a deep colobomatous morning
glory anomaly. Silicone oil tamponade was used and was
complicated by subretinal migration and emulsification
due to the presence of a hole on the steep border of the
central excavation (see also Chapter 22).
FIGURE H-7 Persistent fetal vasculature. A:Wide-field
fundus image of a 5-year-old boy following surgery for
posterior persistent fetal vasculature. He initially presented
a peripapillary retinal fold that resolved after cutting the
fibrous stalk surgically. The fibrosis out of the retinal
plane has a green appearance with this scanning laser
imaging technique. B:Bergmeister papilla with central
fibrous vascular remnants causing contraction of the
peripapillary retina and macula (see also Chapter 65).
FIGURE H-8 Optic disc pit. A:Fundus image of an 8-
year-old boy presenting with a chronic serous retinal
detachment due to an optic disc pit. Visual acuity has been
20/40 for 2 years. B:Red-free image of the same patient.
C:Optical coherence tomography (OCT) vertical scan
shows schisis-like appearance in the outer plexiform layer
and chronic macular serous detachment. D:Horizontal
OCT scan shows communication between outer plexiform
layer and defect in optic nerve. Vitreoretinal adhesion is
visible (see also Chapter 22). Green line through macular
FIGURE H-9 Optic nerve hypoplasia. A, B:Wide-field
fundus images of a 6-month-old infant with severe
bilateral optic disc hypoplasia. A very small optic nerve is
visible in the center surrounded by peripapillary pigment
epithelial changes. The patient does not have the ability to
fix and follow (see also Chapter 22).
Index
A
AAV. See Adeno-associated virus (AAV) vector
AAV serotype 2 vector (AAV2)
AAV (adeno-associated virus) vector
ABCA4 gene, cone-rod dystrophy
ABD. See Adamantiades-Behçet disease
Abetalipoproteinemia
Ablation, peripheral retinal, for retinopathy of prematurity complications of
Ablative therapy, Coats disease
complications
postoperative care
supporting evidence and procedural technique
Absent septum pellucidum
Achromatopsia
ACHM gene therapy
using recombinant viruses, for gene therapy
CNGB3 mutations
complete
ERG
gene associated with
incomplete
Acid ceramidase (AC)
Acidosis, lactic, with mitochondrial encephalomyopathy and stroke-like
episode
Acidosis-induced retinopathy (AIR)
Acute lymphoblastic leukemia (ALL)
Acute posterior multifocal placoid pigment epitheliopathy
N-Acylsphingosine amidohydrolase (ASAH)
Adamantiades-Behçet disease
Adeno-associated virus (AAV) vector
Adenosine deaminase (ADA) deficiency
ADOA. See Autosomal dominant optic atrophy (ADOA)
Adult NCL (ANCL)
AGA (appropriate for gestational age)
Age-related macular degeneration (AMD)
AIR (acidosis-induced retinopathy)
Airway, laryngeal mask, for premature infant
Albendazole, for ocular toxocariasis
Albinism
clinical symptoms and signs
diagnostic studies
differential diagnosis
environmental factors
ethical considerations
foveal hypoplasia
future treatments
genetics
management and visual rehabilitation
dermatologic issues
ophthalmic issues
systemic issues
nonsyndromic albinism
ocular albinism
oculocutaneous albinism, type 1
oculocutaneous
pathophysiology
prevalence
syndromic albinism
Chediak-Higashi syndrome
hermansky-pudlak syndrome
worldwide impact
Alström syndrome
Amaurosis Leber congenital
CRX gene
misdiagnosis of
Amblyopia
diagnostic studies
anisometropia
in pediatric retina practice
refractive error
strabismus
visual deprivation
differential diagnosis of, retinoschisis
future treatments
genetics
management of
pathophysiology
physician and health care providers
prevalence
refractive error
anisometropia
lensectomy
organic amblyopia
posterior uveitis
retinal detachment
retinopathy of prematurity
vitreous and retinal hemorrhages
treatment and vision rehabilitation
worldwide impact
AMD (age-related macular degeneration)
Amino acid disorders
Anesthesia
examination under
general, in premature infants
nitrous oxide
for surgery, inhalation, intraocular gas and
Angiogenesis
Angiography
fluorescein (see Fluorescein angiography (FA))
indocyanine green, in choroidal hemangioma diagnosis
Angioma
Anisometropia
Anomalous posterior vitreous detachment
Anterior chamber, in acute pediatric eye trauma, ocular examination
Anterior chamber paracentesis, for uveitis
Antioxidant therapy
Antioxidants, on retinopathy of prematurity
Antivascular endothelial growth factor
Anti-VEGF treatment. See also Retinopathy of prematurity
aflibercept
bevacizumab
cryotherapy
intravitreal injections for
laser photocoagulation
long-term safety and neurodevelopmental outcomes
pegaptanib
pharmacodynamics and pharmacokinetics
ranibizumab
rationale for drug
recurrence and retreatment of
success rates
trials
safety
VEGF binding receptors
Apoptosis, in retinoblastoma
Appropriate for gestational age (AGA)
Arthritis
inflammatory bowel disease
rheumatoid, juvenile, uveitis in, surgical approaches to
Artificial intelligence (AI) algorithm
Astigmatism
Astrocytes, retinal, development of
Astrocytic hamartoma
Atrophy, autosomal dominant optic
Autosomal dominant disorders
Autosomal dominant optic atrophy (ADOA)
clinical signs and symptoms
diagnostic studies
genetics
management of
pathophysiology
prevalence of
systemic manifestations
worldwide impact
Autosomal dominant retinitis pigmentosa (PRPF8 mutation)
Autosomal dominant Stargardt-like macular dystrophy
clinical characteristics and disease course
differential diagnoses
molecular genetics of
Autosomal dominant vitreoretinochoroidopathy
Autosomal recessive bestrophinopathy (ARB)
Autosomal recessive disorders
Azathioprine
intermediate uveitis
JIA-associated uveitis
for sarcoidosis
B
Band keratopathy, for uveitis
indications
in juvenile rheumatoid arthritis-associated uveitis, surgery
procedure
Bardet-Biedl syndrome (BBS)
BBS1 mutations
anosmia
developmental disability
diabetes mellitus
hearing loss and glue ear
obesity
polydactyly
renal and genital anomalies
retinal degeneration
diagnostic studies in
gene therapy
genetics
health care providers
management of
pathophysiology
physicians role
prevalence of
vision rehabilitation
worldwide impact of
Bassen-Kornzweig syndrome. See Abetalipoproteinemia
Batten disease
BBS. See Bardet-Biedl syndrome (BBS)
Benzodiazepines
Bergmeister’s papilla
Berlin edema. See Commotio retinae
Best corrected visual acuity (BCVA)
Best disease
choroidal neovascularization
clinical findings
genetics
pathophysiology
Best dystrophy
Best vitelliform macular dystrophy (BVMD)
BETTS (Birmingham Eye Trauma Terminology System)
Bevacizumab Eliminates the Angiogenic Threat of Retinopathy of
Prematurity (BEAT-ROP) study
β-galactosidase deficiency
β-glucocerebrosidase deficiency
β-hexosaminidase α-subunits, deficiency in
Bilateral microphthalmia
Bilateral nasal tilted disc syndrome
Bilateral optic disc drusen
Binocular indirect ophthalmoscopy (BIO)
Biopsy
chorioretinal, in uveitis diagnosis
conjunctival, in sarcoidosis diagnosis
retinal, in uveitis diagnosis
Birmingham Eye Trauma Terminology (BETT) System
Birth weight standard deviation score (BWSDS)
Bitemporal hemianopia
Blindness
childhood (see Childhood blindness)
measles-related
cases data
corneal scarring
immunization
keratomalacia
malabsorption
malnutrition and overcrowding
primary prevention
protein-losing enteropathy
secondary prevention
Bloch-Sulzberger syndrome. See also Incontinentia pigmenti (IP)
Blood–retinal barrier (BBB)
development and maintenance
molecular mediators
Hedgehog signaling pathway
Mfsd2a
PDGF-B
retinoic acid
Wnt/wingless pathway
Blue cone monochromatism (OPN1LW mutation)
Body awareness
Bradyopsia
clinical characteristics
genes associated with
Bronchopulmonary dysplasia (BPD ), in VLBW infant, general anesthesia
and
Bronchospasm
complicating general anesthesia in premature infants
intraoperative, acute, pharmacologic therapy for
BWSDS (birth weight standard deviation score)
C
Candida chorioretinitis, flecks and spots
CAPS. See Cryopyrin-associated periodic syndromes (CAPS)
Capsulotomy laser for uveitis
Carbon dioxide, retinopathy of prematurity and
Cardiovascular drips
Carriers, inheritance patterns
Cataract(s)
in uveitis
indications
inflammation management
for intermediate uveitits
in sarcoidosis, extraction of
surgical technique
visual rehabilitation
Catscratch disease
cCSNB. See Complete form of CSNB (cCSNB)
Central nervous system (CNS)
incontinentia pigmenti
lesions of, childhood blindness from
CEP290 mutations (ProQR)
Cerebral visual impairment
Cerebrohepatorenal syndrome
Cerliponase alfa (recombinant TPP1)
Chédiak-Higashi syndrome (CHS)
Chemoreduction, for retinoblastoma
Chemotherapy
immunosuppressive, for intermediate uveitis
systemic, for retinoblastoma
Cherry-red spot myoclonus syndrome
Chikungunya
Childhood blindness
causes of
avoidable
change in, over time
classification
congenital rubella syndrome
data on
fetal alcohol syndrome
harmful traditional eye remedies
lesions of central nervous system
malaria as
measles as
myopia
ophthalmia neonatorum
regional variation in
vitamin A deficiency disorders
definition
epidemiology of
incidence of
magnitude of
mortality
prevalence of
universal health coverage
World Bank economic region
worldwide causes of
Childhood retinal diseases
intravitreal surgery
child phakic or pseudophakic or aphakic
intraoperative considerations
port placement
postoperative evaluation
preoperative assessment
wide-angle viewing system
scleral buckle (see Scleral buckle)
Children
blindness (see Childhood blindness)
endoscopic vitrectomy
external and minor surgical procedures (see Noninvasive/minimally
invasive interventions)
ultrasonography (see Ultrasonography)
Children’s Hospital in Philadelphia Retinopathy Prematurity model (CHOP-
ROP)
Chondroitin sulfate, in vitreous
Choriocapillaris (CC)
embryonic and fetal human development
14 to 16 weeks of gestation
hemovasculogenesis
alkaline phosphatase (APase)
carbonic anhydrase IV (CA IV)
CD31 immunolabeling
CD39-immunolabeled
embryonic hemoglobin (Hb-e) colocalization
endothelial nitric oxide synthase (eNOS)
6.5 WG fetal choroid
time line of
vascular development
Chorionic villus sampling (CVS), genetic testing
Chorioretinal biopsy, in uveitis diagnosis
Chorioretinitis, differential diagnosis of, retinoschisis
Chorioretinitis sclopetaria. See Traumatic chorioretinal rupture
Choroid, coloboma of
Choroidal effusion
Choroidal hemangioma
circumscribed
Choroidal neovascularization
Choroidal osteoma
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
worldwide impact
Choroidal rupture
Choroideremia
clinical presentation and disease course
molecular findings
natural history studies, gene therapy
Chronic progressive external ophthalmoplegia (CPEO)
Ciliary body medulloepithelioma, masquerade uveitis syndromes
Circumscribed choroidal hemangioma
diagnostic studies
genetics
intravitreal bevacizumab
management
pathology and pathophysiology
physicians role
prevalence
worldwide impact
Classification of eye movement abnormalities and strabismus (CEMAS)
Clinical heterogeneity
CLN3 disease
CLN3 gene, neuronal ceroid lipofuscinosis
CLN4 disease
CLN5 disease
CLN6 disease
CLN7 disease
CLN8 disease
CLN9 disease
CLN12 disease
CLN13 disease
Closed globe injury
definition
ocular sequelae management
choroidal rupture
commotio retinae
giant retinal tear
retinal break
retinal detachment
retinal dialysis
traumatic chorioretinal rupture
traumatic macular hole (see Traumatic macular hole)
vitreous hemorrhage
open globe injury zone
ultrasound imaging
CNS (central nervous system)
lesions of, childhood blindness from
Coats disease
antivascular endothelial growth factor therapy
classification of
clinical features of
cryopexy
definition
diagnosis
differential diagnosis of
epidemiology of
genetic associations of
laser ablation
optical coherence tomography
outcomes
pathology/pathophysiology
prognosis
surgery
ablative therapies
equipment
indications
intraocular pharmacotherapies
intraocular surgery
symptoms and signs
systemic associations
Wnt signaling pathway and
worldwide impact of
Cobalamin (vitamin B12), disorder of
Cognitive development
Collagens, in vitreous
disorders of
Coloboma(s)
chorioretinal
choroidal
lens, management of
optic nerve
bilateral
complications
macular pigment
PAX2 transcription
retinal detachment
size of
treatment of
Combined anterior and posterior segment surgery
corneal and retinal surgery
glaucoma drainage device implantation and vitrectomy
keratoprostheses
lenticular and retinal surgery
sutureless intrascleral intraocular lens fixation and pars plana vitrectomy
anterior chamber IOL placement
exchange/secondary IOL placement
flanged sutureless intrascleral IOL fixation
forming haptic bulbs/flanges
in Marfan syndrome
in one-piece PMMA IOL
side effects
surgical variations
Combined hamartoma of pigment epithelium and retina. See Combined
hamartoma of the retina and retinal pigment epithelium
Combined hamartoma of the retina and retinal pigment epithelium (RPE)
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
worldwide impact
Combined hamartoma of the retina and RPE surgical considerations
cases
classification
complications
definition
diagnostic studies
equipment
location
postoperative care
procedural techniques
vs. retinoblastoma
unilateral
Commotio retinae
Comparing Alternative Ranibizumab Dosages for Safety and Efficacy in
ROP (CARE-ROP) study
Complete form of CSNB (cCSNB)
genetics and pathophysiology
Computed tomography (CT)
choroidal osteoma
Coats disease
nonaccidental head injury (NAHI)
in ocular trauma evaluation
Computer-Aided Image Analysis of the Retina (CAIAR)
Computer-based imaging analysis (CBIA)
Cone-rod dystrophy
autosomal dominant
autosomal recessive
clinical presentation and disease course
electrophysiologic characteristics
fundus albipunctatus
genes associated with
macular changes
molecular genetics
Stargardt disease
symptoms
systemic diseases
Cone–rod dystrophy (Bornholm eye disease, OPN1LW mutation)
Cone–rod dystrophy and hearing loss (CRDHL)
Cones
differentiation of
20 WG
40 WG
cone density
cone packing
increased oxygen supply
inner nuclear layers
outer nuclear layer
perifoveal anastomosis
photoreceptor elongation wave
postnatal development
S-cones
dystrophy
Congenital disorder of glycosylation (CDG)
Congenital hypertrophy of the retinal pigment epithelium (CHRPE)
associated with Gardner syndrome and familial adenomatous polyposis
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
worldwide impact
Congenital motor nystagmus (CMN)
Congenital pigment epithelial hypertrophy
Congenital rubella retinopathy
Congenital rubella syndrome, childhood blindness
Congenital stationary night blindness (CSNB)
complete form of CSNB
GNB3-CSNB
incomplete form of CSNB
Oguchi disease
Riggs type of
Schubert-Bornschein type
diagnostic studies
ERG phenotype
future treatments
general clinical characteristics
complete form of CSNB
gene defects in
GNB3 gene defect
incomplete form of CSNB
Oguchi disease
Riggs type
mode of inheritance of
prevalence
referral to low vision specialists
Conjunctiva
in acute pediatric eye trauma, ocular examination
biopsy of, in sarcoidosis diagnosis
Consent, informed, for surgery
Contact camera retinopathy of prematurity
e-ROP
International Classification of ROP
Cornea, in acute pediatric eye trauma, ocular examination
Cortex, vitreous
Corticosteroids
for inflammation control in cataract surgery in uveitic eye
for intermediate uveitis
for juvenile rheumatoid arthritis-associated uveitis
for ocular toxocariasis
for sarcoidosis
for uveitis
CPEO (chronic progressive external ophthalmoplegia)
CPK80 deficiency
CRDHL (cone-rod dystrophy and hearing loss)
CRISPR-Cas systems
Cryopexy
for lattice degeneration
for peripheral retinal ablation
retinal, for intermediate uveitis
Cryopyrin-associated periodic syndromes (CAPS)
familial mediterranean fever
periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis
(PFAPA) syndrome
tumor necrosis factor receptor–associated periodic syndrome
CRYO-ROP trial. See Cryotherapy for Retinopathy of Prematurity (CRYO-
ROP) study
Cryosurgery, for retinopathy of prematurity, anesthesia for
Cryotherapy
for Coats disease
for peripheral neovascularization in uveitis
for retinoblastoma treatment
for retinopathy of prematurity
Cryotherapy for Retinopathy of Prematurity (CRYO-ROP) study
CSNB. See Congenital stationary night blindness (CSNB)
Cyclophosphamide
for sarcoidosis
Cycloplegics, for sarcoidosis
Cyclosporin
for sarcoidosis
Cystine-depleting therapy
Cystinosis
D
Data Safety Monitoring Board (DSMB)
de Morsier syndrome
Deep learning (DL)
Deep neural networks (DNNs)
Degenerative retinoschisis, differential diagnosis of, retinoschisis
4-Degree grating test
Dengue fever (DF)
Depression
Dexamethasone implantation procedure
Diabetic retinopathy, fluorescein angiography
Digital fundus imaging, neonatal intensive care units
automated algorithms
binocular indirect ophthalmoscopy
digital photography
Icon camera
3nethra Neo
Panocam
repeated examination
RetCam
Retcam3 camera
risk for ROP
software algorithms
standard images
topical anesthesia
Digital wide-field imaging systems. See RetCam
Diligent cysteamine therapy
DNA testing, in retinoblastoma
Dominant exudative vitreoretinopathy
Dragged optic disc
Drugs
pediatric anesthesia
MAC concentrations of
pediatric resuscitation
Drusen, optic head
Dysostosis multiplex
E
Early childhood–onset retinal dystrophies (ECORD)
Early intervention and rehabilitation
cerebral visual impairment
challenging test situations
clinical examination and assessment
communication
4-degree grating test
high-risk groups of infants and children with vision loss
at home and school
initial information
passive “vision stimulation,” 149–150
problematic behaviors and situations
school age
tectopulvinar pathway
during treatment
visual milestones
Early Treatment for Retinopathy of Prematurity (ET-ROP) study
Early-onset inherited retinal degeneration (IRD), animal models of
Early-onset retinal degenerations (EORDs)
Ebola viruses
EBRT (external beam radiation therapy), for retinoblastoma
ECORD (Early childhood-onset retinal dystrophies)
EFTFs (eye-field transcription factors)
Ehlers-Danlos syndrome (EDS)
Electronegative waveform, in X-linked retinoschisis
Electroretinogram (ERG)
Usher syndrome
X-linked retinoschisis (XLRS)
Electroretinographic (ERG) waveforms
achromatopsia (CNGB3 mutations)
Alström syndrome (ALMS1 mutations)
Autosomal dominant retinitis pigmentosa (PRPF8 mutation)
Bardet-Biedl syndrome (BBS1 mutations)
Batten disease (CLN3 mutations)
Blue cone monochromatism (OPN1LW mutation)
b-wave development
cone–rod dystrophy (Bornholm eye disease, OPN1LW mutation).
Enhanced S-cone syndrome (NR2E3 mutations)
in healthy adult and 10-week-old infant’s
ISCEV standard protocol
Joubert (INPP5E mutations)
Leber congenital amaurosis, LCA (CEP290, RPE65, CRB1 mutations)
Night blindness
posterior microphthalmos with retinal folds (MFRP mutations)
Stargardt disease (ABCA4 mutations)
Stickler syndrome (COL11A1 mutation)
Usher syndrome (MYO7A, USH2A mutations
X-linked congenital stationary night blindness, CSNB1 (NYX mutation)
and CSNB2 (CACNAF1 mutation)
X-linked juvenile retinoschisis (RS1 mutation)
X-linked retinitis pigmentosa (RPGR mutation)
Embryology, of retina
Embryonic hyaloid vasculature, vitreous in
Encephalomyopathy, mitochondrial, with lactic acidosis and stroke-like
episodes
Endoscopic vitrectomy
advantage of
clinical application
20-gauge endoscope
hand-to-eye coordination
instrument development
learning curve of
persistent fetal vasculature syndrome
sclerotomy placement
surgically relevant properties
anatomical perspective
optical property
pars plana vitrectomy
Endothelial growth factor, vascular, intravitreous neovascularization
Endotracheal tube size and length by weight and age
End-stage disease and developmental disorders, gene therapy
Enhanced S-cone (Goldmann-Favre) syndrome
Enhanced S-cone syndrome (ESCS)
Environmental factors
Bardet-Biedl syndrome
choroidal osteoma
circumscribed choroidal hemangioma
congenital hypertrophy of the retinal pigment epithelium
Kearns-Sayre syndrome
in Leber’s hereditary optic neuropathy
medulloepithelioma
neuropathy, ataxia, and retinitis pigmentosa
in retinoblastoma
Stargardt macular dystrophy/fundus flavimaculatus
Enzymatic vitreolysis, in retinopathy of prematurity
Enzyme replacement therapy (ERT)
Epilepsy
Epiretinal membrane (ERM)
cellophane type
classification
clinical appearance
Coats disease
complications
diagnostic studies
equipment
indications
macular
postoperative care
recurrence
vasocentric
Epstein-Barr virus
ERG. See Electroretinogram (ERG)
e-ROP (multicenter studies)
ERT (enzyme replacement therapy)
Erythropoietin
in ROP and cognitive development
Erythropoietin derivatives (EPO)
associated with severe ROP
ESCS (enhanced S-cone syndrome)
Essential polyunsaturated fatty acids, lack of
Ethical considerations
albinism
amblyopia
choroidal osteoma
combined hamartoma of the retina and retinal pigment epithelium
future treatments
infantile nystagmus syndrome
Leber congenital amaurosis
medulloepithelioma
myopia
neurofibromatosis
neuronal ceroid lipofuscinoses
on retinopathy of prematurity
Sturge-Weber syndrome
tuberous sclerosis
Usher syndrome
Von Hippel-Lindau syndrome
Wyburn-Mason syndrome
X-linked retinoschisis
ET-ROP trial. See Early Treatment for Retinopathy of Prematurity (ET-ROP)
study
Evaluating Acute-Phase ROP (e-ROP) study
Examination under anesthesia (EUA)
infants and children retinal evaluation
ocular examination in acute pediatric eye trauma
scleral buckles
Exogenous erythropoietin
External beam radiation therapy (EBRT), for retinoblastoma
Eye(s)
embryology of
examinations, digital fundus imaging
equipment
potential systemic complications
pupils dilation
schedule
terminated
hyaloidal vasculature (see Hyaloidal vasculature)
remedies for, traditional, harmful blindness from
trauma to
wall of, definition of
Eye-field transcription factors (EFTFs)
F
FA. See Fluorescein angiography (FA); Fundus albipunctatus (FA)
Fabry disease
diagnostic studies
genetics
management
pathophysiology
Familial exudative vitreoretinopathy (FEVR)
basic considerations for technique
classification of
complications after
laser treatment
vitreoretinal surgery
diagnostic studies
endoscopic vitrectomy (see Endoscopic vitrectomy)
equipment
genetics of
outcomes
postoperative care
retinopexy and therapeutic injections
vitrectomy and/or scleral buckle
serial laser treatments
surgical management
treatment indication
for laser and anti-VEGF
for surgery
vitrectomy (see Vitrectomy)
classification system
clinical course
diagnosis
genetics
history
presentations
treatment
Familial mediterranean fever
Farber bodies
Farber disease
diagnostic studies
genetics
pathophysiology
FAZ. See Foveal avascular zone (FAZ)
Fetal alcohol syndrome, childhood blindness from
FEVR. See Familial exudative vitreoretinopathy (FEVR)
ffERG (full-field electroretinogram)
Fibrillins, in vitreous
Fluocinolone implantation procedure, for uveitis
Fluorescein angiography (FA)
CHRRPE
in Coats disease
contact imaging modalities
epiretinal membrane
field of view
fluorescein dye
indocyanine green angiography
noncontact imaging modalities
Coats disease
diabetic retinopathy
familial exudative vitreoretinopathy
sickle cell retinopathy
uveitis
Norrie disease
oral fluorescein dye
retinal nonperfusion
Focal retinopexy
Focal therapies, for retinoblastoma
adjuvant treatment
complications
cryotherapy
external beam radiotherapy
transpupillary thermotherapy
Folds
Foreign body, intraocular
B-scan ultrasonography studies
computed tomography imaging
definition of
indirect ophthalmoscopy
open globe injury complication
removal of
Fovea
cones differentiation
formation of
postnatal development of
prematurity impacts, development of retinal vessels
retinal, development of
Foveal avascular zone (FAZ)
avascular
EphA
in ganglion cell layer
Foveal hypoplasia, albinism
Foveal schisis, X-linked retinoschisis
FST (full-field stimulus or sensitivity testing) techniques
Full-field electroretinogram (ffERG)
Full-field stimulus or sensitivity testing (FST) techniques
Fundus albipunctatus (FA)
autofluorescence of
flecks and spots in
fundus color photographs
optical coherence tomography (OCT)
RDH5 gene mutation
Fundus autofluorescence (FAF) imaging
Fundus flavimaculatus. See Stargardt macular dystrophy/fundus
flavimaculatus
G
Galactosialidosis
diagnostic studies
genetics
pathophysiology
α-Galactosidase A (GLA) deficiency
Galsulfase, ERT with
Gaucher cells
Gaucher disease
diagnostic studies
genetics
pathophysiology
phenotypes
prevalence
Gene(s), retinoblastoma
Gene augmentation therapy
retinal genes targeted in
Gene therapy
animal models of
early-onset inherited retinal degeneration
application of, to pediatric retinal disease
in clinical trials, current path for
current challenges
definition
end-stage disease and developmental disorders
future prospective
gene transfer agents
inherited pediatric-onset retinal diseases
inherited retinal diseases
intraocular gene delivery approaches
intravitreal delivery
agents dispersed into vitreous chamber
clinical trials
complications
disadvantages
equipment for
for gene- or cell-based therapies
medications
postoperative care
procedural technique for
for rhegmatogenous retinal detachment
vitreous gel
for leber congenital amaurosis (LCA)
natural history studies
new disease targets
nucleic acids, clinical trials for retinal diseases
outcome measures and therapeutic windows
posterior segment
principles of
recombinant viruses, clinical trials for retinal diseases
for retinoblastoma
RPE65 mutant animals
status, for genetic disease
transchoroidal subretinal delivery
equipment for
transvitreal subretinal delivery
basic considerations
benefits, risks, and potential complications
biallelic heterozygous mutations in RPE65
complications
equipment for
outcomes
postoperative care
in utero
viral vectors in
Gene transfer agents in gene therapy
Gene-based therapies, genetic counseling for pediatric retinal diseases
Genetic counseling
clinical diagnosis
counselor role, in ophthalmology
goal of
patient education
providing support resources
timing
family history in
gene-based therapies
genetic complexity of, inherited
genetic testing
appropriate medical management
changing diagnosis
chorionic villus sampling (CVS)
multi-gene testing
and privacy
results
selection of
in vitro fertilization (IVF)
inheritance and risk
inheritance patterns
autosomal dominant disorder
autosomal recessive disorders
Mendelian inheritance, genetic conditions with
mitochondrial DNA mutations
multifactorial inheritance
pedigree construction, symbols used in
penetrance
X-linked disorder
on retinoblastoma
risk assessment
Genetic heterogeneity
Genetic Information Nondiscrimination Act (GINA)
Genetic testing
retinoblastoma
Stargardt macular dystrophy
Genetics
in autosomal dominant optic atrophy
choroidal osteoma
in chronic progressive external ophthalmoplegia
of Coats disease
of familial exudative vitreoretinopathy (FEVR)
glycosylation disorders
in Kearns-Sayre syndrome
in Leber hereditary optic neuropathy
in Leigh syndromes
masquerade uveitis syndromes
medulloepithelioma
myopia
of neuropathy, ataxia, and retinitis pigmentosa
Norrie disease
of papillorenal syndrome
of Refsum disease
of retinoblastoma
clinical
retinoblastoma gene and protein product
Stargardt macular dystrophy
Usher syndrome
Germline mutation, in heritable retinoblastoma
Giant retinal tears (GRTs)
Glaucoma
in juvenile rheumatoid arthritis-associated uveitis, surgery for
in sarcoidosis, surgery for
in uveitis
angle-closure
mechanisms of
Glioma, retinal
Global Education Network for Retinopathy of Prematurity (GEN-ROP)
Global Universal Eye Screen Testing (GUEST)
GLP (Good Laboratory Practices)
Glybera
Glycosaminoglycans, in vitreous
Glycosylation, congenital disorders of
GM1 gangliosidosis
diagnostic studies
genetics
pathophysiology
GM2 gangliosidosis (Sandhoff disease)
diagnostic studies
genetics
pathophysiology
GM2 gangliosidosis (Tay-Sachs disease)
cherry-red spot
diagnostic studies
genetics
management
pathophysiology
prevalence
GNB3 gene, congenital stationary night blindness
Goldmann-Favre syndrome
Goldmann-Witmer (GW) coefficient, in uveitis evaluation
Gonococcal keratoconjunctivitis
Good Laboratory Practices (GLP)
Granular osmiophilic deposits (GRODs)
Granuloma, posterior pole, in ocular toxocariasis
Gyrate atrophy
H
HA. See Hyaluronan
Haltia-Santavuori
Hamartoma
astrocytic
combined
Health Insurance Portability and Accountability Act of 1996
Hemangioma, circumscribed choroidal
Hematopoietic stem cell transplant
Hemianopic field
Hemorrhage(s)
intravitreal
preretinal
retinal
retrohyaloidal
subhyaloidal subfoveal
Heparan sulfate, in vitreous
Heritable retinoblastoma
Hermansky-Pudlak syndrome (HPS)
Herpes simplex viruses
Hexosaminidase A deficiency
cherry-red spot
diagnostic studies
genetics
management
pathophysiology
prevalence
High-risk retinoblastoma
Human patient-derived induced pluripotent cell (iPSC) models
Hunter syndrome
diagnostic studies
genetics
management
pathophysiology
Hurler-Scheie syndrome
Hyalocytes, in vitreous
Hyaloidal vasculature
hyaloid artery
in 40-mm human embryo
during atrophy
prepapillary/vitreous hemorrhage
structure of
hyaloidal vascular regression
angiopoietin receptor
macrophages
vascular endothelial growth factor
Wnt7b deficiency
pupillary membrane
tunica vasculosa
Hyaluronan (HA), in vitreous
Hydrocephalus
Hypermetropia
Hyperoxaluria type 1, primary
Hyperoxia
Hyperoxia-induced vaso-obliteration
Hypoxia
hypoxia-inducible factor-1
physiologic
VEGF accessibility
VEGF mRNA stability
Hypoxia-induced VEGF expression
Hypoxia-inducible factor (HIF)
heterodimeric transcription factors
prolylhydroxylases
regulated genes and general pathway analysis
safety and efficacy
stabilization
structure
synergy between liver and retina
Hypoxic retina
I
icCSNB. See Incomplete form of CSNB (icCSNB)
ICROP (International Classification of Retinopathy of Prematurity)
Idiopathic nonspecific orbital inflammatory syndrome
Iduronate-2-sulfatase (IDS), deficiency of
α-L-Iduronidase (IDUA), 311
Idursulfase, ERT with
IGF-1 (insulin-like growth factor)
IGF-1R (insulin-like growth factor receptor)
IGF-I (insulin-like growth factor I)
Image storage and retrieval and telemedicine
definition
remote digital fundus imaging
experimental analysis of
implementation of
KIDROP
limitations
ophthalmic technology assessment
real-world application
safety
SUNDROP
screening guidelines
Imaging and Informatics in Retinopathy of Prematurity (I-ROP)
Immunosuppressive therapy
for sarcoidosis
Impaired vision
In silico approaches
In utero ocular gene therapy
In vitro fertilization (IVF), genetic testing
Incomplete form of CSNB (icCSNB)
Incomplete penetrance
Incontinentia pigmenti (IP)
adult vitreoretinal sequelae
central nervous system involvement
complications
diagnostic studies
diagnostic testing
disease pathogenesis of
enucleation
equipment
future treatments
genetics
indications
intravitreal antivascular endothelial growth factor therapy
macular pathology
media opacity
optic nerve atrophy
management
outcomes
peripheral ablation
postoperative care
prevalence and environmental factors
retinal pigment epithelium abnormality
roles of other providers
scleral buckle
screening for
strabismus
worldwide impact
Indirect ophthalmoscopy, retinopathy of prematurity
Individual educational plan (IEP)
Individual learning plan (ILP)
Indocyanine green angiography, in choroidal hemangioma diagnosis
Infant(s)
premature, general anesthesia in
bronchospasm complicating
drugs for
extubation after
retinopathy of prematurity (ROP)
oxidative stress in preterm infant
postnatal oxygen
postnatal stresses
in utero and perinatal setting
surgical approaches (see Intravitreal surgery; Scleral buckle)
ultrasonography (See Ultrasonography)
Infantile CLN1 disease
Infantile NCL (INCL) disease
Infantile nystagmus syndrome (INS)
diagnostic approach
brain imaging
clinical examination
clinical history
electroretinography
eye movement testing
molecular genetic testing
testing
future treatments
genetics
management
extraocular muscle surgery
optical treatments
pharmacologic therapies
subretinal gene therapy
pathophysiology
physicians and health care providers
prevalence
worldwide impact
Infantile Refsum disease
Inflammatory bowel disease (IBD) arthritis
Informed consent, surgical intervention for infants and children
Inherited pediatric-onset retinal diseases, gene therapy
nonviral vectors, studies using
CEP290 mutations (ProQR)
viral vectors, studies using
achromatopsia (ACHM) gene therapy
choroideremia
leber hereditary optic neuropathy
retinitis pigmentosa
RPE65 deficiency
Stargardt disease
XL retinoschisis
Inherited retinal disease (IRD)
gene therapy in (see Gene therapy)
Insulin-like growth factor (IGF-1)
Insulin-like growth factor 1 (IGF-1)
Insulin-like growth factor I (IGF-I)
Insulin-like growth factor receptor (IGF-1R)
Interfactants, in pharmacologic vitreolysis
Intermediate uveitis
age of onset
bilateral disease
diagnosis
management
antimetabolites
cataract surgery
corticosteroids
immunosuppressive drugs
oral NSAIDs
peripheral laser/cryotherapy
peripheral retinal cryopexy
vitrectomy
symptoms and signs
International classification of retinoblastoma (ICRB)
International Classification of Retinopathy of Prematurity (ICROP)
International Pediatric Ophthalmology and Strabismus Council (IPOSC)
International Society for Clinical Electrophysiology of Vision (ISCEV)
Intra-arterial chemotherapy (IAC), retinoblastoma
Intracameral chemotherapy (ICC), retinoblastoma
Intraocular foreign body (IOFB)
B-scan ultrasonography studies
computed tomography imaging
definition of
open globe injury complication
removal of
Intraocular gene delivery approaches
Intraocular hemorrhage
Intraocular pharmacotherapy, in Coats disease
complications
equipment
Intraocular pressure, in acute pediatric eye trauma, ocular examination
Intravenous (IV) access, in premature infant
Intravenous chemotherapy (IVC), retinoblastoma
Intravitreal chemotherapy, retinoblastoma
Intravitreal injection, of pharmacologic agents
Intravitreal lipid hydroperoxides (LHP)
Intravitreal neovascularization
Intravitreal surgery
child phakic or pseudophakic or aphakic
intraoperative considerations
adjuvant scleral buckle
choice of tamponade
drainage of subretinal fluid
membrane stripping
posterior hyaloid separation and vitrectomy
port placement
postoperative evaluation
preoperative assessment
vitrectomy
wide-angle viewing system
Intravitreal triamcinolone (IVT), in Coats disease
IOFB. See Intraocular foreign body (IOFB)
IP. See Incontinentia pigmenti (IP)
iPSC (human patient-derived induced pluripotent cell) models
IRD (inherited retinal disorders)
Iridectomy, laser, for uveitis
Iridocyclitis. See Juvenile idiopathic arthritis–associated (JIA) uveitis
Iris
nodules, in sarcoidosis, surgical excision of
stromal cysts
IRVAN syndrome
ISCEV (International Society for Clinical Electrophysiology of Vision)
ISCEV standard protocol
Isoflurane, for anesthesia in premature infants
J
Janus kinase/signal transducer and activator of transcription (JAK/STAT)
signaling
Joubert (INPP5E mutations)
Joubert syndrome
Juvenile CLN3 disease
Juvenile idiopathic arthritis–associated (JIA) uveitis
band keratopathy
cataract surgery
combined phacoemulsification and pars plana vitrectomy
daunting
indications
IOL implantation
outcomes
pars plana lensectomy
peripheral iridectomy
risk of
glaucoma surgery
incidence
medical therapy
outcomes
medical management
prevalence
surgical indications
symptoms and signs
Juvenile NCL (JNCL)
Juvenile seronegative spondyloarthropathies (JSpA)
ankylosing spondylitis
inflammatory bowel disease (IBD) arthritis
psoriasis
reactive arthritis
Juvenile xanthogranuloma (JXG)
K
Karnataka Internet Assisted Diagnosis of Retinopathy of Prematurity
(KIDROP) program
Kawasaki disease (KD)
diagnostic studies
genetics
management
pathophysiology
prevalence of
worldwide impact
Keratomalacia, vitamin A deficiency disorders
corneal ulceration and necrosis
mortality rate
Keratopathy, band, for uveitis
indications
procedure
Keratoprostheses
Knobloch syndrome
clinical features
genetics
Kufs disease
Kufs type B
L
Laceration, definition of
Lacrimal system, associated injury, evaluation for
Lactic acidosis, with mitochondrial encephalomyopathy and stroke-like
episodes
Lamellar schisis
Langerhans cell histiocytosis (LCH)
Laryngeal mask airway (LMA)
patient selection guidelines
for premature infant
Laser
for retinopathy of prematurity
ablative therapy
anti-VEGF studies
ETROP study
surgical iridectomy
panretinal/segmental photocoagulation
peripheral neovascularization
posterior capsulotomy vs. surgical posterior capsulotomy
Laser capsulotomy, for uveitis
Laser iridectomy, for uveitis
Laser peripheral retinal ablation
Laser photocoagulation
choroidal hemangioma
choroidal osteoma
in Coats disease
for retinoblastoma treatment
for uveitis
Laser retinopexy
for abnormal vitreoretinal interface
Laser-induced hyperthermia
Late infantile CLN2 disease
Late infantile NCL (LINCL)
Lattice degeneration
cryopexy for
laser retinopexy for
LCA. See Leber congenital amaurosis (LCA)
LCHAD deficiency
LCPUFAs (long chain polyunsaturated fatty acids)
Least restrictive educational environment
Leber congenital amaurosis (LCA)
CEP290, mutation in
CEP290, RPE65, CRB1 mutations
classification
CRB1, mutation in
CRX, mutation in
CRX gene
diagnostic studies
future treatments
genetic heterogeneity of
genetics
GUCY2D, mutation in
Luxturna, subretinal injection of
management
misdiagnosis of
molecular pathways causing
pathophysiology
physicians and health care providers
prevalence
RDH12, mutation in
RPE65, mutation in
TULP1, mutation in
types of
using nucleic acids, for gene therapy
worldwide impact
Leber congenital amaurosis, retinal dystrophy
Leber hereditary optic neuropathy (LHON)
diagnostic studies
genetics
management
pathophysiology
prevalence of
worldwide impact
Leber neuroretinitis
Leigh syndromes
Lens
coloboma of, management of
Lensectomy
Norrie disease
for retinopathy of prematurity-related retinal detachment
Lens-sparing vitrectomy, vision rehabilitation
Lentiviral vectors, in gene therapy
Leukemia, masquerade uveitis syndromes
Leukocoria
LHP (intravitreal lipid hydroperoxides)
Lids, evaluation for lacerations evidence
Light, retinopathy of prematurity and
animal studies
clinical trials
Light Reduction in Retinopathy of Prematurity (Light-ROP) study
Lipid metabolism
LCHAD deficiency
Sjögren-Larsson syndrome
Lipid-laden macrophages
Lipoprotein metabolism, disorder of
Liquefaction, in pharmacologic vitreolysis
Literacy, low vision
braille
definition
historical perspective
least restrictive educational environment
optical and electronic magnification devices
LMA (laryngeal mask airway)
patient selection guidelines for
for premature infant
LN11 disease
Long chain polyunsaturated fatty acids (LCPUFAs)
Low vision
barriers to care
clinical low vision rehabilitation evaluation
definition
driving with a visual impairment
functional and educational outcomes
literacy
braille
definition
historical perspective
least restrictive educational environment
optical and electronic magnification devices
optical devices
pediatric visual acuity testing
rehabilitation team
statistics
technology
vision enhancement options
Luxturna
Lysosomal storage disease (LSD)
cystinosis
diagnosis of
Fabry disease
diagnostic studies
genetics
management
pathophysiology
Farber disease
diagnostic studies
genetics
pathophysiology
galactosialidosis
diagnostic studies
genetics
pathophysiology
Gaucher disease
diagnostic studies
genetics
pathophysiology
phenotypes
prevalence
GM1 gangliosidosis
diagnostic studies
genetics
pathophysiology
Hurler-Scheie syndrome
management
mucolipidoses
mucopolysaccharidosis
type I
type II
type III
type IV
type VI
type VII
neuronal ceroid lipofuscinoses
diagnostic studies
future treatments
genetics
incidence
management
pathophysiology
physicians and health care providers, role of
Niemann-Pick disease (NPD)
diagnostic studies
genetics
management
pathophysiology
Sandhoff disease
diagnostic studies
genetics
pathophysiology
Scheie syndrome
sialidosis
diagnostic studies
genetics
pathophysiology
sphingolipidoses
Tay-Sachs disease
cherry-red spot
diagnostic studies
genetics
management
pathophysiology
prevalence
M
Machine learning
Macula
detachments
dystrophies
autosomal dominant Stargardt-like macular dystrophy
clinical characteristics and disease course
differential diagnoses
molecular genetics of
Best disease
clinical findings
genetics
pathophysiology
North Carolina macular dystrophy
clinical findings
molecular genetics
pattern dystrophy
Sorsby fundus dystrophy
Macular halo syndrome
Macular hole
Coats disease
Macular pseudocolobomas
Maculopathy
optic disc pit
cerebrospinal fluid (CSF) pressure
intraocular pressure
neuroectodermal/astroglial tissue
posterior vitreous detachment (PVD)
prepapillary glial material
treatment of
Magnetic resonance imaging (MRI)
medulloepithelioma
nonaccidental head injury (NAHI)
retinoblastoma
Malaria, childhood blindness from
Marfan syndrome
clinical features
genetics
pediatric rhegmatogenous retinal detachment
vitreous in
Maroteaux-Lamy syndrome
child with
diagnostic studies
genetics
management
pathophysiology
Masquerade uveitis syndromes
cause of
definition
genetics
health care providers role
lymphoma
primary CNS
systemic non-Hodgkin
uveal
metastatic tumors
neoplastic disease entities
ciliary body medulloepithelioma
juvenile xanthogranuloma
langerhans cell histiocytosis
leukemia
retinoblastoma
nonneoplastic entities
idiopathic nonspecific orbital inflammatory syndrome
intraocular foreign body
iris stromal cyst
retinal degenerations
retinal detachment
vitreous hemorrhage
physicians role
posttransplant lymphoproliferative disorder
prevalence
uveal melanoma
Measles virus
Measles-related blindness
cases data
corneal scarring
immunization
keratomalacia
malabsorption
malnutrition and overcrowding
primary prevention
protein-losing enteropathy
Mebendazole, for ocular toxocariasis
Mechanical ventilation, for VLBW infants, general anesthesia
Medulloepithelioma
diagnostic studies
genetics
management
pathophysiology
physicians role
prevalence
worldwide impact
MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like
episode syndrome)
Melatonin
Mendelian genetic diseases
Metabolic disorder
abetalipoproteinemia
cobalamin-C–type methylmalonic aciduria with homocystinuria
congenital disorder of glycosylation
Gyrate atrophy
lipid metabolism
LCHAD deficiency
lysosomal storage disease
cystinosis
mucolipidoses
mucopolysaccharidoses
neuronal ceroid lipofuscinosis
sphingolipidoses
mitochondrial disorders
mitochondrial variants A3243G and T8993G
peroxisomal disorders
primary hyperoxaluria
Zellweger spectrum disorders
prevalence
Metastasis, of retinoblastoma, risk of
Metastatic tumors, masquerade uveitis syndromes
Methotrexate
for sarcoidosis
Microcornea, persistent fetal vasculature syndrome
Microglia, retinal, development of
Migalastat, Fabry disease
Mild optic nerve hypoplasia
Mitochondria
disorders of
autosomal dominant optic atrophy
chronic progressive external ophthalmoplegia
future treatments
Kearns-Sayre syndrome as
Leber hereditary optic neuropathy
Leigh syndromes
mitochondrial encephalomyopathy, lactic acidosis, and stroke-like
episode syndrome
mitochondrial variants A3243G and T8993G
neuropathy, ataxia, and retinitis pigmentosa
physician role
feature of
function
Mittendorf dot
Mizuo phenomenon, X-linked retinoschisis
Mizuo-Nakamura phenomenon
Morning glory disc anomaly
clinical presentation
complications
etiology of
Morquio syndrome
Morquio A syndrome
diagnostic studies
genetics
management
pathophysiology
Morquio B syndrome
diagnostic studies
genetics
management
pathophysiology
Mortality, in blind children
Mosaicism, genetic in retinoblastoma
Motor development
Mucolipidoses
Mucopolysaccharidosis
type I
type II
type III
type IV
type VI
type VII
Multifactorial inheritance
Multi-gene testing, genetic counseling for pediatric retinal diseases
Multi-luminance mobility test (MLMT)
Mutation
germline mutation, in heritable retinoblastoma
X-linked juvenile retinoschisis (see X-linked juvenile retinoschisis)
Mycophenolate
Myopia
childhood blindness from
clinical features
diagnostic studies
genetics
management
atropine
orthokeratology
outdoor time and myopia
prophylactic retinopexy in stickler syndrome
refractive surgery
pathophysiology and environmental factors
prevalence
scleral collagen cross-linking
tilted disc syndrome
vitreous in
worldwide impact
N
NAHT. See Nonaccidental head trauma (NAHT)
NARP. See Neuropathy, ataxia, and retinitis pigmentosa (NARP)
Nasal tilted disc syndrome, fundus appearance
National Reading Media Assessment (NRMA)
NCL. See Neuronal ceroid lipofuscinosis (NCL)
NCMD. See North Carolina macular dystrophy
Neonatal intensive care units (NICUs)
digital fundus imaging
automated algorithms
binocular indirect ophthalmoscopy
digital photography
Icon camera
3nethra Neo
Panocam
repeated examination
RetCam
Retcam3 camera
risk for ROP
software algorithms
standard images
topical anesthesia
eye examinations
equipment
potential systemic complications
pupils dilation
schedule
terminated
Neural and vascular interaction
Neurofibromatosis
clinical features
diagnostic studies
future treatments
genetics
management
neurofibromatosis 1 associated with glaucoma
pathophysiology
physicians role
prevalence
Neuronal ceroid lipofuscinosis (NCL)
CLN3 gene
diagnostic studies
future treatments
genetics
incidence
management
pathophysiology
physicians and health care providers, role of
Neuronal migrational anomalies
Neuropathy, ataxia, and retinitis pigmentosa (NARP)
diagnostic studies
genetics
management of
pathophysiology
prevalence of
worldwide impact
Neuropathy, optic, Leber hereditary
Neuroretinitis
Newborn Eye Screen Testing (NEST)
Next generation sequencing (NGS)
NICUs. See Neonatal intensive care units (NICUs)
Niemann-Pick disease (NPD)
diagnostic studies
genetics
management
pathophysiology
Night blindness
Nitric oxide (NO)
Nonaccidental head trauma (NAHT)
brain lesions
case studies in
cause
definition
incidence
with localized macular retrohyaloidal hematoma
ocular lesions
diagnostic studies
macular changes
neovascularization and retinal ischemia
neural damage
pathophysiology and specificity
retinal detachment
retinal folds
retinal hemorrhages (see Retinal hemorrhages)
schisis
prevalence
vision rehabilitation (see Vision rehabilitation)
Nonarteritic anterior ischemic optic neuropathy (NAION)
Noninfectious uveitides
Adamantiades-Behçet disease
cryopyrin-associated periodic syndromes (CAPS)
familial juvenile systemic granulomatosis
granulomatosis with polyangiitis
intermediate uveitis
juvenile dermatomyositis
juvenile idiopathic arthritis–associated uveitis
juvenile seronegative spondyloarthropathies (JSpA)
ankylosing spondylitis
inflammatory bowel disease (IBD) arthritis
psoriasis
reactive arthritis
Kawasaki disease (KD)
masquerade syndromes
polyarteritis nodosa
sarcoidosis
sympathetic ophthalmia (SO)
systemic lupus erythematosus
tubulointerstitial nephritis and uveitis
Vogt-Koyanagi-Harada disease
Noninvasive/minimally invasive interventions
cryopexy
for peripheral retinal ablation
focal retinopexy
intravitreal injection, of pharmacologic agents
laser retinopexy
for abnormal vitreoretinal interface
neonatal intensive care units (see Neonatal intensive care units (NICUs))
peripheral retinal ablation
cryopexy for
for vascular/VEGF mediated retinopathies
Nonsteroidal antiinflammatory drugs (NSAIDs)
Nonsyndromic albinism
ocular albinism
oculocutaneous albinism
type 1
type 2
type 3
type 4
type 5
type 6
type 7
Norrie disease
clinical diagnosis of
complications
equipment
genetics
indications
management
missense mutation
outcomes
pathophysiology
postoperative care
prevalence
clinical course
diagnosis
genetics
history
presentations
treatment
worldwide impact
North Carolina macular dystrophy
clinical findings
molecular genetics
Northern epilepsy (NE)
NPD. See Niemann-Pick disease (NPD)
Nutrition, prenatal weight gain
Nystagmus
O
Obesity, Bardet-Biedl syndrome
Object permanence
OCT. See Optical coherence tomography
Ocular albinism
Ocular inflammatory disease
Ocular lesions, in nonaccidental head injury
diagnostic studies
macular changes
neovascularization and retinal ischemia
neural damage
pathophysiology and specificity
retinal detachment
retinal folds
retinal hemorrhages (see Retinal hemorrhages)
schisis
Ocular toxocariasis
clinical signs of
diagnosis of
diagnostic features of
management
medical
surgical
outcomes
pathogenesis of
postoperative care in
surgical indications
Ocular trauma
acute open globe injury management
complications
equipment
intraocular foreign body removal
outcomes
postoperative care
prophylactic scleral buckle surgery
systemic antibiotic therapy
tetanus shot/booster administration
vitreoretinal surgery complications (see also Vitreoretinal surgery)
Birmingham Eye Trauma Terminology for
definitions
closed globe injury (see also Closed globe injury)
eye wall
laceration
open globe injury (see also Open globe injury)
penetrating injury
perforating
superficial foreign body
diagnostic studies
ocular examination in acute pediatric eye trauma
anterior chamber depth assessment
conjunctiva
cornea
examination under anesthesia
external evaluation
iris
lens
physical examination
pupillary reflex and intraocular pressure
sclera
visual acuity measurement
patient history
posterior segment manifestations
prevalence
Oculocutaneous albinism (OCA). See also Albinism
type 1 (OCA1)
type 2 (OCA2)
type 3 (OCA3)
type 4 (OCA4)
type 5 (OCA5)
type 6 (OCA6)
type 7 (OCA7)
OD. See Oguchi disease (OD)
OFF-bipolar cell
Oguchi disease (OD)
OIR. See Oxygen-induced retinopathy; Oxygen-induced retinopathy (OIR)
Oligocone trichromacy
ONA (optic nerve aplasia)
ON-bipolar cells
ONH (optic nerve hypoplasia)
ONHD (optic nerve head drusen)
Open globe injury
management
complications
equipment
intraocular foreign body removal
outcomes
postoperative care
prophylactic scleral buckle surgery
systemic antibiotic therapy
tetanus shot/booster administration
vitreoretinal surgery complications (see also Vitreoretinal surgery)
occurrence
as rupture
zones of
Ophthalmia neonatorum, childhood blindness from
Optic atrophy. See also Autosomal dominant optic atrophy
Optic disc edema (ODE)
Optic disc pit
maculopathy
cerebrospinal fluid (CSF) pressure
intraocular pressure
neuroectodermal/astroglial tissue
posterior vitreous detachment (PVD)
prepapillary glial material
treatment of
Optic nerve
abnormalities of
coloboma
optic disc pit
optic nerve aplasia
optic nerve head drusen (ONHD)
optic nerve hypoplasia
superior segmental optic hypoplasia
tilted disc syndrome
coloboma
development of
Optic nerve aplasia (ONA)
diagnostic criteria
embryology
Optic nerve coloboma
bilateral
complications
macular pigment
PAX2 transcription
retinal detachment
size of
treatment of
Optic nerve head drusen (ONHD)
clinical evaluation of
calcified drusen
color fundus and autofluorescence
hyperreflective
subretinal hyporeflective space (SHYPS)
complications
NAION
Optic nerve hypoplasia (ONH)
bilateral
double ring sign
nystagmus
ophthalmoscopy
poor visual behavior
endocrine anomaly
etiology of
mild
prenatal and perinatal risk factors
prevalence of
severe
Optic neuropathy, Leber hereditary
Optical coherence tomography (OCT)
choroidal osteoma
choroidal rupture
CHRRPE
epiretinal membrane
intraretinal edema
maculae
medulloepithelioma
normal full-term retina
pediatric imaging
Coats disease
foveal avascular zone morphology
infant foveal development considerations
methods and limitations
pediatric choroidal neovascular membranes
pediatric epiretinal membranes
retinoblastoma
trisomy 21
retinal layers
Stargardt disease
traumatic macular hole
Optical coherence tomography angiography (OCT-A)
abnormalities
application
Opticin, in vitreous
Optogenetic gene therapy
Oral cysteamine therapy
Orbital pseudotumor. See Idiopathic nonspecific orbital inflammatory
syndrome
Orbital trauma
Organic amblyopia
Ornithine metabolism, disorder of
Osteoporosis pseudoglioma syndrome (OPPG)
Oxidative signaling pathways
Oxidative stress, retinopathy of prematurity and
lutein
vitamin E
Oxidative stress in preterm infant
Oxidative stress–related therapy
Oxygen, retinopathy of prematurity and
clinical trials
STOP-ROP study
SUPPORT study
high oxygen
monitoring and management
Benefit of Oxygen Saturation Targeting (BOOST) study
effect of O2 saturation
HOPE-ROP infants
length of time
STOP-ROP trial
TcPO2 levels
oxygen fluctuations
oxygen supplementation
saturation of fetus in utero, 563
Oxygen-induced retinopathy (OIR)
animal models of
clinical studies
anti-VEGF
erythropoietin in ROP and cognitive development
insulin-like growth factor (IGF-1)
hyperoxia-induced vaso-obliteration
with isolectin staining
neural and vascular interaction
oxidative stress–related therapy
oxygen fluctuations and OIR severity
physiologic retinal vascular development and pathogenic
vasoproliferation
potential antioxidant therapies
signaling pathways, postnatal stresses
antioxidants
erythropoietin
oxidative signaling pathways
VEGF signaling
VEGFA
treatment strategies
variable oxygen
P
Pain, in postanesthesia care unit, treatment of
Panda sign, in sarcoidosis
Papilla, Bergmeister
Papillorenal syndrome
Paracentesis, anterior chamber, for uveitis
Parenchymal anoxic ischemic injuries
Pars plana vitrectomy (PPV)
cataract
juvenile idiopathic arthritis
ocular toxocariasis
sarcoidosis
sutureless intrascleral intraocular lens fixation and
anterior chamber IOL placement
exchange/secondary IOL placement
flanged sutureless intrascleral IOL fixation
forming haptic bulbs/flanges
in Marfan syndrome
in one-piece PMMA IOL
side effects
surgical variations
for uveitis
vitreoretinal complications
Pattern dystrophy
Pax6 mutations
Pediatric choroidal neovascular membranes, optical coherence tomography
Pediatric epiretinal membranes, optical coherence tomography
Pediatric Eye Disease Investigator Group (PEDIG) study
Pediatric retinal diseases, genetic counseling for
clinical diagnosis
counselor role, in ophthalmology
goal of
patient education
providing support resources
timing
family history
gene-based therapies
genetic complexity of, inherited
genetic testing
appropriate medical management
changing diagnosis
chorionic villus sampling (CVS)
multi-gene testing
and privacy
results
selection of
in vitro fertilization (IVF)
inheritance and risk
inheritance patterns
autosomal recessive disorders
Mendelian inheritance, genetic conditions with
mitochondrial DNA mutations
multifactorial inheritance
pedigree construction, symbols used in
penetrance
X-linked disorder
risk assessment
Pediatric rhegmatogenous retinal detachment (PRRD)
after ocular surgery
associated inherited disorders
Ehlers-Danlos syndrome (EDS)
Marfan syndrome
Stickler syndrome
atopic dermatitis
causes of
clinical examination
diagnostic studies
epidemiology
etiology of
incidence
indications
nontraumatic inferotemporal retinal dialysis
occurrences
outcomes
postoperative care
surgerical approach
general considerations
inherited disorders
scleral buckling
traumatic/nontraumatic retinal dialysis
vitrectomy
trauma
Pediatric uveitis
diagnostic approach
age of presentation
anatomic regions
categories
juvenile idiopathic arthritis
epidemiology
history
incidence and prevalence
infectious uveitides
catscratch disease
chikungunya virus
cytomegalovirus
dengue virus
diffuse unilateral subacute neuroretinitis
Ebola viruses
Epstein-Barr virus
herpes simplex viruses
Lyme disease
lymphocytic choriomeningitis virus
measles virus
postinfectious and vaccination-associated uveitis
rubella
subacute sclerosing panencephalitis (SSPE)
syphilis
toxocariasis
toxoplasmosis
tuberculosis
varicella–zoster virus
West Nile virus
Zika virus
medical management
noninfectious uveitides (see Noninfectious uveitides)
types of
visual outcomes
Pericytes, retinal, development of
Periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis
(PFAPA) syndrome
Peripapillary choroidal neovascularization
Peripheral retinal ablation
cryopexy for
for vascular/VEGF mediated retinopathies
Peroxisomal disorders
primary hyperoxaluria
Peroxisomes
biosynthetic functions
catabolic and anabolic processes
disorders of
biogenetic, affecting retina
classification
clinical manifestations
measuring saturated very longchain fatty acids
prevalence
Refsum disease as
worldwide impact
primary hyperoxaluria type 1
proteins required for
Persistent fetal vasculature syndrome (PFVS)
anterior
bilateral
characteristic findings of
combined
complication
diagnostic studies
color Doppler imaging
fundus photography
magnetic resonance imaging
ultrasonography
visual evoked potential
genetics
glucose 6-phosphate dehydrogenase (G6PD) deficiency
long-term management
management
nomenclature and types
nonsurgical treatments
outcomes
pathophysiology
posterior
postoperative care
surgery
incisions
indications
intraoperative optical coherence tomography
limbal approach
pars plicata approach
small-gauge surgery
timing of
vitrectomy (see Vitrectomy)
Persistent hyperplastic primary vitreous (PHPV)
Persistent maculopathy
PFVS. See Persistent fetal vasculature syndrome (PFVS)
Pharmacologic vitreolysis
adverse events
indications and contraindications
interfactants
liquefactants
pediatric development
Phenotypic heterogeneity
Photocoagulation, laser. See Laser photocoagulation
Photodynamic therapy (PDT)
with verteporfin
Photophobia
Photopic ERGs
Photoreceptors
development
differentiation of
retinoschisin synthesis
PHPV (persistent hyperplastic primary vitreous)
Pigmentary changes
Platelet-derived growth factor (PGDF) signaling
angiopoietin-1/tie-2 signaling
transforming growth factor beta1
Plus disease, diagnosis of
Polyarteritis nodosa (PAN)
Polydactyly, Bardet-Biedl syndrome
Postanesthesia care unit, pain in, treatment of
posterior microphthalmos with retinal folds (MFRP mutations)
Posterior pituitary hypoplasia
Posterior segment
hemorrhages in
masses
Posterior uveitis
Postnatal growth, timing of
Postnatal Growth and Retinopathy of Prematurity study (G-ROP)
Postnatal oxygen
Postnatal stresses, increase risk of ROP
Preimplantation genetic diagnosis (PGD). See In vitro fertilization
Premature infants, general anesthesia in
bronchospasm complicating
drugs for
endotracheal tube size and length for
extubation after
intravenous access
Premature Infants in Need of Transfusion ROP (PINT-ROP) study
Prematurity
fovea
at 24 WG
at 28 WG
by 32 WG
anatomy
Eph receptor signaling pathways
FAZ diameter
hypoxia
increased retinal thickness
oxygen-enriched environment exposure
retinopathy of (see Retinopathy of prematurity (ROP))
Prenatal weight gain
in animal studies
in clinical studies
alarm, time after birth
hydrocephalus
hyperoxia of
insulin-like growth factor I (IGF-I)
mild versus severe ROP
ROP Score, 521
at 6 weeks
WINROP
factors contributing to
inappropriate nutrition
increased metabolic rate
lack of essential polyunsaturated fatty acids
lack of growth factors and nutrients
oxygen supplementation
future interventions
nutrition
timing of postnatal growth
as risk factor for ROP
small for gestational age (SGA)
versus AGA
definition of
in mature infants
retrolental fibroplasia
standard deviation scores
Preserved para-arteriolar RPE (PPRPE)
Primary hyperoxaluria
Primary hyperoxaluria type 1
Print Media Assessment Process (PMAP)
Progressive macular atrophy
Progressive rod–cone dystrophy
Protein(s)
retinoblastoma
structural, noncollagenous, in vitreous
Pupillary reflex, in acute pediatric eye trauma, ocular examination
R
Radiation therapy
for retinoblastoma
Ranibizumab Compared With Laser Therapy for the Treatment of Infants
Born Prematurely with ROP (RAINBOW) study
RDFI. see Remote digital fundus imaging
RDH5 gene, congenital stationary night blindness
RDH12-LCA, full-field sensitivity (FST) testing in
Reactive oxygen species (ROS)
Recombinant adeno-associated virus vectors (rAAV)
Recombinant adenoviral vectors, in gene therapy
Recombinant α-L-iduronidase (laronidase)
Recombinant DNA engineering techniques
Refractive error
anisometropia
lensectomy
organic amblyopia
posterior uveitis
retinal detachment
vitreous and retinal hemorrhages
Refsum disease
Rehabilitation
early intervention and
cerebral visual impairment
challenging test situations
clinical examination and assessment
communication
4-degree grating test
high-risk groups of infants and children with vision loss
at home and school
initial information
passive “vision stimulation,” 149–150
problematic behaviors and situations
school age
tectopulvinar pathway
during treatment
visual milestones
retinoblastoma
visual (see also Vision rehabilitation)
after cataract surgery in uveitic eye
choroidal osteoma
Relative afferent pupillary defect (RAPD)
Remote digital fundus imaging (RDFI)
experimental analysis of
implementation of
binocular indirect ophthalmoscopy (BIO) evaluation
discharge
graders
image transfer and storage
personnel
PHOTO-ROP study
RDFI team
reports
KIDROP
limitations
ophthalmic technology assessment
real-world application
safety
SUNDROP
Respiratory compromise, in VLBW infant, general anesthesia and
Resuscitation medications
RetCam
Retina
4B
predominantly effusive, mechanism of
predominantly tractional, mechanism of
abnormal vasculature
biopsy of, in uveitis diagnosis
complications of sarcoidosis involving
degeneration of
degenerations of
animal models of
en-face imaging in
detachments of in retinopathy of prematurity, clinical features of
development of
embryology of
ERG waveforms, inherited pediatric diseases
eye field
foveal development
ganglion cell death
histogenesis, cell types
astrocytes
Dicer gene
FoxN4, 81
H-thymidine “birthdating” technique
lineages tracking
microglia
miRNA production
Müller glial cells
Pax6
progenitor cells
inner
amacrine cells
bipolar cells
cell adhesion molecules
contact inhibition
dendrites
ganglion cells
ganglion cells migration
inner plexiform layer
tiling
laser peripheral retinal ablation
optic cup
peroxisomal biogenesis disorders affecting
diagnostic studies
genetics
management of
systemic manifestation
photoreceptor development
retinopathy of prematurity-related, surgical anatomy of
rhegmatogenous (see Rhegmatogenous retinal detachment)
vascular development
Retinal and choroidal dystrophies
achromatopsia
complete
ERG
gene associated with
incomplete
choroideremia
cone-rod dystrophy (see Cone-rod dystrophy)
enhanced S-cone syndrome
CRX gene
misdiagnosis of
Retinal cavernous hemangioma
Retinal degenerations
Retinal detachment
Retinal dialysis, pediatric rhegmatogenous retinal detachment
Retinal dysplasia
Retinal glioma
Retinal hemorrhages
causes of
circular hypopigmented retinal fold
hemorrhages resolution
histopathologic studies
incidence
vs. intraocular hemorrhage
with localized macular retrohyaloidal hematoma
Roth spots with fibrin
with traumatic macular hole
Retinal hypoxia
Retinal image analysis
challenges
deep learning
future directions
initial computer based imaging analysis approaches
machine learning
plus disease, diagnosis of
Retinal Image multiScale Analysis (RISA)
Retinal imaging
benefits and drawbacks
central, external and internal angles
classification
clinical utility
evolution
imaging consideration
advantages
choroidal circulation
contact cameras
flying baby position
limitations
noncontact U-WF imaging module
Optos device
RetCam-III
stage 4 ROP
vertical red/black line artifacts
limitations
Phantom eye
universal retinal screening of newborn
Retinal pigment epithelium (RPE)
atrophy
development of
Retinitis pigmentosa. See also Progressive rod–cone dystrophy
Retinitis pigmentosa (RP)
diagnostic studies in
gene therapy
genetics
management of
pathophysiology
physicians role
prevalence of
worldwide impact of
natural history studies, gene therapy
neuropathy, ataxia
X-linked, using recombinant viruses, for gene therapy
Retinitis punctata albescens
Retinoblastoma (RB)
classification
clinical presentation of
diagnostic studies
diffuse
epidemiology and prevalence
ethical consideration in
follow-up of
future treatments
gene therapy for, animal models of
genetic counseling and DNA testing
genetic mosaicism
genetic testing
genetics of
clinical
retinoblastoma gene and protein product
group classification
heritable
historical context of
low-expressivity
low-penetrance
management
decision tree by patient presentation
enucleation
factors for
focal therapies (see Focal therapies)
high-risk retinoblastoma
intra-arterial chemotherapy
intracameral chemotherapy
intravenous chemotherapy
plaque radiotherapy
precision intravitreal chemotherapy
metastasis, risk of
metastatic disease
monitoring children with
nonheritable
orbital disease
pathophysiology of
physicians role
prognosis of
screening for
second tumor predisposition
staging examination
trilateral
worldwide impact of
Retinocalcarine pathway
Retinocytoma
Retinoma
Retinopathy of prematurity (ROP)
advanced stages of
age of onset
aggressive posterior
anti-VEGF
safety
VEGF binding receptors
anti-VEGF treatment
aflibercept
bevacizumab
cryotherapy
intravitreal injections for
long-term safety and neurodevelopmental outcomes
pegaptanib
pharmacodynamics and pharmacokinetics
ranibizumab
rationale for drug
recurrence and retreatment of
success rates
trials
BEAT-ROP study
blindness (see Blindness)
childhood blindness
epidemics due to
in high-income countries
primary prevention
risk factors for
screening and treatment
supplemental oxygen
tertiary prevention
visual impairment and blindness
clinical trials in
complications
course of
cryotherapy for
development of
diagnosis of
contact camera
diagnostic features of
education and management
in low-income countries
in middle-income countries
preventative measures
telemedicine and tele-education
in United States (see United States)
worldwide
effect of oxygen, historical perspectives on
endoscopic vitrectomy (see Endoscopic vitrectomy)
environmental factors in
genetics
light
neurovascular effects
oxidative stress
oxygen
pathophysiology
worldwide impact
enzymatic vitreolysis
erythropoietin derivatives
ethical considerations on
ET-ROP trial
evolution and differences worldwide
first and second epidemic of
general anesthesia for
global workforce shortage
historical context
and hypoxia-inducible factors (see Hypoxia-inducible factor (HIF))
Imaging and Informatics in Retinopathy of Prematurity
incidence
infant(s)
oxidative stress in preterm infant
postnatal oxygen
postnatal stresses
in utero and perinatal setting
International Classification of
intravitreal anti-VEGF therapy
late stages of pathogenesis
low weight at birth
management of
neonatal care and
Postnatal Growth and ROP (G-ROP) study
screening
WIN-ROP algorithm
nonocular treatments on
carbon dioxide
oxygen
steroids administration
surfactant therapy
outcomes
oxygen, primary determinant of
oxygen and angiogenesis
oxygen-induced retinopathy (see also Oxygen-induced retinopathy
(OIR))
animal models of
anti-VEGF
erythropoietin in ROP and cognitive development
hyperoxia-induced vaso-obliteration
insulin-like growth factor (IGF-1)
with isolectin staining
neural and vascular interaction
signaling pathways, postnatal stresses
variable oxygen
pathogenesis of
pathophysiology
broad anti-VEGF effects
optimal anti-VEGF dose inhibits IVNV
targeted VEGFR2 inhibition
persistent retinal avascularity, premature birth
phenotype and treatment
physicians role
postnatal weight gain
in animal studies
prematurity in clinical studies
severe ROP prediction
postoperative care
prenatal weight gain as risk factor for (see also Prenatal weight gain)
prethreshold
prevalence
prevention of
lensectomy and vitrectomy
lens-sparing vitrectomy
scleral buckle
rat OIR model
refractive error
retinal detachment in
equipment
4B, mechanisms of
surgical anatomy of
timing of surgery
retinal hypoxia
retinal image analysis
risk factors for
severe
studies on
tele-education
telemedicine (see Telemedicine)
third epidemic of
threshold
treatment
anti-VEGF treatment
bilirubin
D-penicillamine
laser
predicting progressive stage 4
propranolol
unfavorable macular outcome
unfavorable retinal outcomes
vasopermeability factor
vision rehabilitation for
Retinoschisin synthesis, photoreceptors
Retrolental fibroplasia
Retroviral vectors, in gene therapy
Rhegmatogenous retinal detachment (RRD)
occurrence
scleral buckling
Rheumatoid arthritis, juvenile, uveitis in, surgical approaches to
Riggs type of congenital stationary night blindness
RLBP1 deficiency
Rod–cone dystrophy. See also Leber congenital amaurosis (LCA)
differential diagnosis of, retinoschisis
Rod-derived cone viability factor (RdCVF)
Rod-photoreceptor dysfunction. See Fundus albipunctatus (FA)
Rods
ROP. See Retinopathy of prematurity (ROP)
ROP Score, 521
ROS (reactive oxygen species)
RPE (retinal pigment epithelium)
RPE65 deficiency
RPE65 mutant, in gene therapy
RPE65-associated retinal dystrophy
Rubella
Rubella syndrome, congenital, childhood blindness from
S
Sandhoff disease
diagnostic studies
genetics
pathophysiology
Sanfilippo syndrome
diagnostic studies
genetics
pathophysiology
Sarcoid choroiditis, flecks and spots in
Sarcoidosis
clinical signs
conjunctival biopsy
diagnosis of
medical management
phorocoagulation for
surgerical indications
surgical management
cataracts
glaucoma
iris nodules
vitreoretinal complications
Scheie syndrome
Schisis
Schubert-Bornschein type of congenital stationary night blindness
Schwartz-Matsuo phenomenon
Sclera, in acute pediatric eye trauma, ocular examination
Scleral buckle
childhood retinal diseases
combined with pars plana vitrectomy
complications
equipment
general considerations
axial length
familial exudative vitreoretinopathy (see also familial exudative
vitreoretinopathy (FEVR))
imaging technology
partial-thickness scleral sutures
persistent fetal vasculature (PFV) (see also Persistent fetal
vasculature syndrome (PFVS))
placement
PVR
retinal breaks
retinal dialysis
scleral thickness
Stickler syndrome
trauma
indications
outcomes
pediatric rhegmatogenous retinal detachment (see also Pediatric
rhegmatogenous retinal detachment)
complications
fishmouth effect
laser/cryotherapy
postoperative care
chandelier-assisted scleral buckling
clean tenon capsule
encircling band
external drainage
general principles
retinal break localization
360-degree conjunctival peritomy
vitreoretinal traction
for retinopathy of prematurity-related retinal detachment
vitrectomy and
Sclopetaria. See Traumatic chorioretinal rupture
Scotopic ERGs
SDS (standard deviation scores)
Sea-blue histiocytes
SECORD (severe early childhood-onset retinal dystrophy)
Senior-Löken syndrome (SLSN)
Septo-optic dysplasia (SOD)
Severe combined immune deficiency (SCID)
Severe early childhood–onset retinal dystrophy (SECORD)
Sevoflurane, for anesthesia in premature infants
SGA. See Small for gestational age (SGA)
Shaken baby syndrome (SBS). See Nonaccidental head trauma (NAHT)
Sialidosis
diagnostic studies
genetics
pathophysiology
Sickle cell disease
Sickle cell retinopathy, fluorescein angiography
Signaling pathways, postnatal stresses
antioxidants
erythropoietin
oxidative signaling pathways
VEGF signaling
VEGFA
Slow-release steroid treatment, for uveitis
dexamethasone implantation procedure
FA reservoir device
fluocinolone implantation procedure
fluorescein angiogram
optic nerve swelling
peripapillary necrotizing retinitis
SLSN (Senior-Löken syndrome)
Sly syndrome
diagnostic studies
genetics
pathophysiology
Small for gestational age (SGA)
vs. AGA
definition of
in mature infants
retrolental fibroplasia
SMDCRD (spondylometaphyseal dysplasia with cone-rod dystrophy)
Solar retinopathy
Sonic hedgehog (Shh) gene
Sorsby fundus dystrophy
Sorsby pseudoinflammatory macular dystrophy. See Sorsby fundus dystrophy
Spatial concepts
Sphingolipidoses
Spondylometaphyseal dysplasia with cone–rod dystrophy (SMDCRD)
Standard deviation scores (SDS)
Stanford University Network for Diagnosis of Retinopathy of Prematurity
(SUNDROP)
Stargardt disease. See also Stargardt macular dystrophy/fundus
flavimaculatus
ABCA4 mutations
pisciform flecks in
animal therapeutic studies
bivariate contour ellipse area
clinical classification
autofluorescence classification
ERG classification
ophthalmoscopic/electrophysiologic/psychophysical classification
eccentric fixation and preferred retinal locus in
future treatments
gene discovery
genetic testing for
genetics
historical perspectives
management of
pathologic studies
pathophysiology
prevalence
retinol-binding protein antagonists
BPN-14136
fenretinide
RPE65 (emixustat), inhibition of
school-age issues
subtypes of
worldwide impact
Steroids
retinopathy of prematurity and
for uveitis
glaucoma
slow-release (see Slow-release steroid treatment, for uveitis)
Stickler syndrome
bilateral spontaneous retinal detachments
clinical features
COL11A1 mutation
genetics
inherited and childhood retinal detachment
ocular features of
orofacial findings
buckle/vitrectomy
giant retinal tears associated with
inherited connective tissue disorders
surgical treatment
posterior retinal break
prophylactic retinopexy
complications
equipment
outcome expectations
supporting evidence and cryopexy and laser techniques
right eye, with prophylactic laser
risk of vision loss
type 1, pediatric rhegmatogenous retinal detachment
Col2A1 mutation
cryoprophylaxis
Stiles-Crawford effect
STOP-ROP (Supplemental Therapeutic Oxygen for Pre-threshold
Retinopathy of Prematurity)
Store-and-forward programs
Strabismus surgery
open globe injury repair
Stroke-like episodes, mitochondrial encephalomyopathy, and lactic acidosis
with
Student, Environment, Tasks, and Tools (SETT) framework
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
Subacute sclerosing panencephalitis (SSPE)
Subdural hemorrhage (SDH)
Superior segmental optic hypoplasia
Supplemental oxygen saturation (STOP-ROP)
SUPPORT study (Surfactant, Positive Pressure, and Pulse Oximetry
Randomized Trial)
Surfactant therapy, retinopathy of prematurity and
Surgery
anatomy for
anesthesia for, inhalation, intraocular gas and
cataract (see Cataract(s))
Coats disease, intraocular
complications
equipment
for gene therapy (see Gene therapy)
informed consent for
intravitreal (see Intravitreal surgery)
patient position for
postoperative care
scleral buckle (see Scleral buckle)
team for
for uveitis (see Uveitis)
vitreoretinal interface in
warning bracelet
Sympathetic ophthalmia (SO)
Syndromic albinism
Chediak-Higashi syndrome
hermansky-pudlak syndrome
Systemic lupus erythematosus
T
Tadoma
Tay-Sachs disease
cherry-red spot
diagnostic studies
genetics
management
pathophysiology
prevalence
Teachers of the visually impaired (TVI)
Tectopulvinar pathway
Tele-education
Telemedicine
Teller Preferential test
Therapeutic surgery, for uveitis
angle-closure glaucoma
band keratopathy
indications
procedure
cataract
indications
surgical technique
glaucoma
lasers (see Laser)
uveitic glaucoma
vitreoretinal surgery (see Vitreoretinal surgery)
Thiabendazole, for ocular toxocariasis
Tilted disc syndrome
bilateral nasal
clinical appearance
astigmatism
bitemporal hemianopia
inferonasal optic nerve fiber loss
macular detachments
marked central excavation
myopia
pigmentary accumulation
etiology
prevalence of
Title VII of the Civil Rights Act of 1964
Total tractional retinal detachment (TRD)
Toxocariasis
ocular (see Ocular toxocariasis)
Toxoplasmosis
Traditional eye remedies, harmful blindness from
Transient PRL (TPLR)
Transpupillary thermotherapy (TTT), for retinoblastoma
Transvitreal vitrectomy, vision rehabilitation
Traumatic chorioretinal rupture
clinical features
globe deformation
long-term follow-up
occurrence
retinal and choroidal defect
ultrasound imaging
Traumatic macular hole
equipment
indications and basic considerations
management
OCT
origin
outcomes
pathophysiology
postoperative care
ILM peeling
intraocular gas tamponade
ocriplasmin
plasmin-assisted pars plana vitrectomy with C3F8 gas
platelet rich plasma
silicone oil tamponade
wound closure
subsequent vitreoretinal surgery
visual outcomes
vitreous traction
Trihexyphenidyl
Triple freeze–thaw technique
Trisomy 21, optical coherence tomography
Trophic feeds
Tuberous sclerosis
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
Tumor(s)
choroidal osteoma
combined hamartoma of retina and retinal pigment epithelium
medulloepithelioma
phakomatoses
neurofibromatosis
tuberous sclerosis
retinoblastoma as
Tumor angiogenesis, in retinoblastoma
Tumor necrosis factor receptor–associated periodic syndrome
Type 1 sialidosis
Type 2 sialidosis
U
Ultrasonography
choroidal osteoma
Coats disease
Norrie disease
retinoblastoma
in trauma evaluation
Ultrasound
A-scan probe
benefits of
B-scan probe
choroidal effusion
Coats disease
definition
examination
in infants and children
in leukocoria
limitations
ocular toxocariasis
persistent fetal vasculature
piezoelectric effect
retinal detachment
scleral rupture with vitreous incarceration
Ultra–wide-field fluorescein angiography (UWFFA)
Unilateral amblyopia
United States, education and management of ROP
digital retinal image
international ophthalmology trainees
laser photocoagulation procedures
mean distribution
misdiagnosis
ophthalmology training in
overdiagnosis
potential solutions
residents and residency program
type 2 prethreshold ROP
web-based surveys of training programs
United States Eye Injury Registry
Universal health coverage (UHC)
UPI (uteroplacental insufficiency)
Usher syndrome (MYO7A, USH2A mutations
Usher syndrome (USH)
among deaf/hard-of-hearing population
cause of
classification
clinical trials
diagnostic studies
electroretinogram
examination of retinitis pigmentosa
genetic testing
lip reading
newborn hearing screening
future treatments
genetics
management
pathophysiology
physician role
type 2
Usher proteins
in human hair cell
in photoreceptor
Uteroplacental insufficiency (UPI)
Uveal melanoma, masquerade uveitis syndromes
Uveitis
masquerade syndromes (see Masquerade uveitis syndromes)
pediatric (see Pediatric uveitis)
steroids for
slow-release
surgical approaches to
anterior chamber paracentesis
by disease (see also Intermediate uveitis; Juvenile idiopathic
arthritis–associated (JIA) uveitis; Ocular toxocariasis;
Sarcoidosis)
considerations for
diagnostic
retinal or chorioretinal biopsy
therapeutic (see Therapeutic surgery, for uveitis)
vitrectomy
V
VADD. See Vitamin A deficiency disorders, (VADD), childhood blindness
Variable oxygen
Variant LINCL
Variants of uncertain significance (VUS)
Varicella–zoster virus
Vascular endothelial growth factor (VEGF)
accessibility
anti-VEGF treatment ( see Anti-VEGF treatment)
experimental studies on pathophysiology of ROP
expression of
gradient
mRNA stabilization
in normal vascular development
notch signaling
role
Vascular remodeling, retinal, development of
Vascular system
choroid
choriocapillaris (CC) (see Choriocapillaris (CC))
Haller layer
regulation of
Sattler layer
embryonic, vitreous disorders from
notch signaling
retinal, development of
astrocytes
blood-retinal barrier development and maintenance
fovea
in human
microglia
in mouse
mRNA stability
notch signaling
physiologic hypoxia
platelet-derived growth factor signaling
vascular endothelial growth factor (see Vascular endothelial growth
factor)
vascular remodeling
vasculogenesis and angiogenesis
Vasculature
hyaloid artery
during atrophy
prepapillary/vitreous hemorrhage
structure of
hyaloidal vascular regression
angiopoietin receptor
macrophages
Wnt7b deficiency
pupillary membrane
tunica vasculosa
Vasculogenesis
VEGF. See Vascular endothelial growth factor (VEGF)
VEGF signaling
anti-VEGF
evolution and differences worldwide
oxygen and angiogenesis
pathophysiology
phenotype and treatment
rat OIR model
vasopermeability factor
VEGFA
VEGFRs
Ventilation, mechanical, for VLBW infants, general anesthesia
Very low birth weight (VLBW) infant, general anesthesia in
Vessel tortuosity. See also Plus disease
Viral vectors, gene therapy for inherited pediatric-onset retinal diseases
achromatopsia (ACHM) gene therapy
choroideremia
leber hereditary optic neuropathy
RPE65 deficiency
Stargardt disease
XL retinoschisis
Viral vectors, in gene therapy
Vision rehabilitation
amblyopia
asymmetric vitreous hemorrhage
choroidal osteoma
combined hamartoma of the retina and retinal pigment epithelium
equipment
indications
infantile nystagmus syndrome
medulloepithelioma
mitochondria disorders
myopia
neurofibromatosis
neuronal ceroid lipofuscinosis
Norrie disease
outcomes
retinoblastoma
surgical technique
tuberous sclerosis
Visual acuity, in acute pediatric eye trauma, ocular examination
Visual and retinal function
achromatopsia
congenital stationary night blindness
electroretinography
full-field
multifocal
leber congenital amaurosis
normal development
normal scotopic retinal sensitivity
background adaptation
dark adaptation after exposure to bright light
dark-adapted visual threshold
rod-mediated visual thresholds
spatial summation
temporal summation
normal visual acuity
Visual function testing
Vitamin A deficiency disorders, (VADD), childhood blindness
in animal food sources
Bitot spot in
causes of
communities risk
conjunctival xerosis
definition of
keratomalacia
low dietary intake
malabsorption
in preschool-aged children
primary prevention for
supplementation coverage
tertiary prevention
xerophthalmia
Vitamin E, retinopathy of prematurity and
Vitrectomy
enzymatic vitreolysis
membrane dissection
primary RRD
rhegmatogenous detachments
silicone oil tamponade
with traction retinal detachment
with/without lensectomy
glaucoma drainage device implantation and
intravitreal surgery (see Intravitreal surgery)
lensectomy and, for retinopathy of prematurity-related retinal
detachment
lens-sparing, for retinopathy of prematurity-related retinal detachment
Norrie disease
pars plana
for traction macular detachment in ocular toxocariasis
for uveitis
bimanual approach
complications
intraocular gas C3F8
perfluorocarbon liquid
PVD induction
retinal detachments
anterior
posterior and combined
for uveitis
indications
intermediate uveitits
procedure
Vitreolysis
enzymatic, in retinopathy of prematurity
pharmacologic
adverse events
indications and contraindications
interfactants
liquefactants
pediatric development
Vitreopathy, myopic
Vitreoretinal interface
surgery and
Vitreoretinal procedures
Vitreoretinal surgery
complications for open globe injuries repair
equipment
indications
outcomes
postoperative care
techniques
Vitreoretinal surgery for uveitis
indications
course of inflammation
cyclitic membranes, removal of
opacities removal
vitreomacular membranes removal
procedure
scleral buckle procedures
Vitreoretinopathies. See also Familial exudative vitreoretinopathy (FEVR)
atropine
familial exudative (see also Familial exudative vitreoretinopathy
(FEVR))
orthokeratology
outdoor time and myopia
prophylactic retinopexy in stickler syndrome
refractive surgery
Vitreous
abnormal development of
biochemistry of
collagen disorders
cyst(s)
development and aging
persistent hyperplastic primary (See also Persistent fetal vasculature
syndrome)
persistent hyperplastic primary vitreous
primary
secondary
structure of
supramolecular organization
Vitreous and retinal hemorrhages
Vitreous base, structure of
Vitreous body, biochemistry of
Vitreous cortex
Vitreous hemorrhage, masquerade uveitis syndromes
Vitreous seeds, in retinoblastoma
Vitreous surgery, for retinopathy of prematurity goals of
Vogt-Koyanagi-Harada disease
clinical features
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
Vulnerable subjects
W
Wagner syndrome
clinical features
genetics
Wagner vitreoretinal dystrophy
Weight IGF-I Neonatal ROP (WINROP)
West Nile virus (WNV)
Wide-field fluorescein angiography, incontinentia pigmenti
WINROP (Weight IGF-I Neonatal ROP)
Wnt signaling pathway
and Coats disease
and coloboma
discovery
microcornea and
molecular pathways in
Norrie disease and FEVR
clinical course
diagnosis
genetics
history
presentations
treatment
and osteoporosis pseudoglioma syndrome
and persistent fetal vasculature syndrome
posterior megalolenticonus and
retinal vasculature development
and retinopathy of prematurity
Wyburn-Mason Syndrome
clinical features
diagnostic studies
future treatments
genetics
management
pathophysiology
physicians role
prevalence
X
XL retinoschisis
X-linked cone-rod dystrophy
X-linked congenital stationary night blindness, CSNB1 (NYX mutation) and
CSNB2 (CACNAF1 mutation)
X-linked disorders
X-linked dominant inheritance
X-linked juvenile retinoschisis (RS1 mutation)
X-linked retinitis pigmentosa (RPGR mutation)
classification system for
diagnostic studies of
ethical considerations and genetic counseling
future treatments
genetics
human XLRS gene therapy
low-vision aids
medical management
pathophysiology
physicians role
prevalence of
surgical treatment of
Z
Zellweger syndrome
Zika virus
APPENDIX OF PROCEDURAL
VIDEOS
Georges Caputo
A. COATS DISEASE
Video 1 Vitrectomy and Photocoagulation in

Coats Disease
In patients with Coats disease, continuous low power laser can be delivered
directly to telangiectatic areas and scatter treatment in the regions of
avascular retina. However, in this case of a 9-year-old boy, previous
treatment was insufficient. The boy presented with stage 3 Coats disease and
intravitreal hemorrhage; after laser, exudation did not resolve. A 25-G
vitrectomy was performed to release the posterior hyaloid and apply direct
endolaser to telangiectasia followed by silicone oil tamponade. Several
months after silicone oil removal, there was resolution of exudation, and
vision improved from 20/200 to 20/80.
Video 2 Laser Photocoagulation in Coats

Disease
Even in a case of total retinal detachment, if telangiectasia are clearly visible,
laser (yellow or green) can be applied to the telangiectatic vessels shown here
with a contact lens or by indirect delivery (see also chapter 64) and result in
resolution of the retinal detachment over a month or more. Additional
sessions may be needed.
Video 3 Technical Issues in Vitrectomy for
Coats Disease
In case of total retinal detachment with insufficient visualization to treat
retinal telangiectasia, surgery is performed. An external drainage sclerotomy
is performed to release subretinal fluid prior to cautious introduction of an
infusion line and active ports into the posterior vitreous cavity. This is
important to avoid internal drainage and the risk of proliferative
vitreoretinopathy. Perfluorocarbon liquid can be used to push thick subretinal
fluid toward the area of the external drainage location. Direct endolaser is
applied to telangiectatic vessels prior to the infusion of silicone oil.
B. FAMILIAL EXUDATIVE
VITREORETINOPATHY (FEVR)
Video 4 Managing Epiretinal Membrane in

FEVR
Epiretinal membranes can form in FEVR as shown in this case in which a
thick vascularized membrane was present. The patient had undergone
previous peripheral laser to avascular retinal areas. Careful dissection of the
membrane was performed to avoid causing retinal breaks. Additional
peripheral laser was added to the regions of avascular retina at the end of the
procedure with a favorable postoperative outcome.
Video 5 Active FEVR in Young Infants

This young girl was early diagnosed with FEVR, and peripheral laser was
applied in her first year of life. She was lost to follow-up for 3 years. An
intravitreal hemorrhage was present necessitating a 25-gauge vitrectomy and
further laser treatment.
Video 6 Managing Exudative FEVR by

Vitrectomy
In 2012, 4-year-old boy presented with FEVR and vision of 20/60 in the right
eye after one cryotherapy and laser session. Over the next 2 years, four
additional laser sessions were performed and vision was 20/30. Over the next
2 years, three additional sessions of laser and cryotherapy were performed
with an injection of bevacizumab and vision was preserved at 20/40. One
year later, despite additional sessions laser and cryotherapy and bevacizumab,
severe exudation occurred and a shallow retinal detachment causing VA to
worsen to 20/400.
A 25-G vitrectomy was performed to release the posterior hyaloid and
peel preretinal membranes. The peripheral vitrectomy and dissection was
added with internal slit lamp illumination and careful light scleral depression
indentation.
Mild peripheral cryotherapy was performed. The procedure led to
resorption of exudates, flattening of the posterior pole, and vision of 20/100.
A mild ERM recurred.
NOTE WELL: USE OF ANTI-VEGF IS NOT ROUTINELY USED
IN FEVR
C. MACULA
Video 7 Management of Epiretinal Membrane in

Children
Initially treated by occlusion therapy, this 10-year-old boy with 20/200 vision
was operated on by 25-G vitrectomy to remove a diffuse epiretinal
membrane. VA was 20/50. Neurofibromatosis is considered, and in this case,
a brain MRI was performed.
Video 8 Vitrectomy for Traumatic Macular Hole

With Flap
A 11-year-old boy presented with a traumatic macular hole of 2 years
duration and VA 20/200. A 25-G vitrectomy is performed with posterior
vitreous separation and injection of dual blue to aid in peeling the internal
limiting membrane (ILM). The ILM membrane flaps are positioned in the
hole, and a fluid–air exchange is performed. Long-acting gas was exchanged
for air. At 3 months, the hole was closed and VA was 20/40.
PLEASE NOTE: DUAL BLUE IS AVAILABLE IN EUROPE BUT
MAY NOT BE AVAILABLE EVERYWHERE. INDOCYANINE
GREEN ALSO STAINS THE INTERNAL LIMITING MEMBRANE.
Video 9 Optic Disc Pit Management

A persistent serous retinal detachment and decreased VA from optic pit
maculopathy in this 5-year-old boy with 20/400 vision was treated by 25-G
vitrectomy and gas tamponade. Although the ILM is not always peeled, here
the ILM was peeled with a favorable outcome over 6 months later.
Resolution of serous detachments with optic nerve pits can take months.
NOTE WELL: THE LITERATURE DOES NOT PROVIDE HIGH-
GRADE EVIDENCE THAT ILM PEELING IS NECESSARY.
PATIENTS ARE APPROACHED CASES BY CASE AND
RESOLUTION OF DETACHMENTS CAN BE SUCCESSFUL WITH
POSTERIOR HYALOID REMOVAL WITHOUT ILM PEELING OR
PERIPAPILLARY LASER.
D. RETINAL DETACHMENT
Video 10 Management of Retinal Detachment in
Stickler Syndrome
This one eye boy presented a recent retinal detachment due to a giant superior
retinal tear although a few months before, a thorough retinal examination did
not find any predisposing lesions. After placing a 3-mm scleral buckle, a 25-
G vitrectomy is performed. A dye, such as sodium fluorescein, allows better
visualization of the posterior vitreous, and the retinal tear appears to be at the
ora serrata. Perfluorocarbon liquid (PFCL) is used to reposition and stabilize
the retina to apply peripheral laser. A PFCL/silicone oil exchange was then
performed. Long-term results are presented.
PLEASE NOTE: DYE MUST BE APPROVED FOR INTERNAL
USE IN THE VITREOUS AND MAY NOT BE AVAILABLE
EVERYWHERE.
E. PERSISTENT FETAL VASCULATURE

(PFV) AND RETINAL DYSPLASIA
Video 11 Complete Anterior PFV

A complete anterior form of persistent fetal vasculature in a 3-month-old
infant with a vascularized retrolental membrane and immature iris. Surgery
consists of an anterior approach to remove the lens, diathermized vessels on
the retrolental membrane, and carefully dissect bimanually up to the ciliary
processes. In this case, prophylactic peripheral endolaser was performed to
avoid a secondary retinal detachment. Prophylactic laser may not be used in
all cases.
Video 12 Incomplete Anterior PFV

Partial anterior persistent fetal vasculature syndrome with tractional retinal
detachment showing anterior approach to avoid injury to the retina,
lensectomy, anterior vitrectomy, and removal of capsule and fibrotic
membranes, followed by prophylactic peripheral endolaser assisted with
scleral depression. There was no retinal break noted in this case, and the
retinal detachment was believed to be all tractional.
F. POSTERIOR PFV
Video 13 Posterior Retinal Fold With Marked

Cataract
A 18-month-old infant with milky cataract and retinal fold. The surgical
approach is initially anterior to allow for intraocular lens implantation. The
periphery of the retina is not involved, which allows a 25-G vitrectomy to
remove the fibrotic medallion adherent to the posterior capsule, thereby
removing the tractional component of the retinal fold. A posterior vitrectomy
frees the retinal fold.
Video 14 Posterior Retinal Fold With Lens

Attachment
This form of posterior PFV presents with a tractional retinal fold centrally
attached to the lens. After assuring no abnormalities of the peripheral retina,
which are common in PFV, a posterior approach to vitrectomy was taken.
After cutting the fibrous stalk to avoid retinal traction, the posterior medallion
is gently peeled off the posterior capsule of the lens, avoiding the need to
remove the lens.
Video 15 Posterior Retinal Fold With Lens
Attachment
The clinical presentation is the same as the latter case; the bimanual
dissection of the medallion leads to a break in the posterior capsule, which
necessitates the extraction of the lens. Care is exercised to only cut the
posterior stalk to release traction so as not to create an inadvertent break in
the retina (retina is often pulled up into the talk and not appreciated easily).
An intraocular lens was implanted and the operation was terminated with
success.
G. ROP
Video 16 Laser Treatment Procedure in Stage 3

ROP
After screening through a telemedicine solution a premature baby with stage
3 plus disease, the infant is transferred from the NICU to the operating room.
Monitoring and intubation allow pretreatment photographs; green laser is
applied through an external adapter for the microscope, which allows laser
delivery through a pediatric quadraspheric lens. After post-treatment
photographs, the baby is sent back to the NICU with the incubator.
PLEASE NOTE: THERE IS VARIABILITY ACROSS PRACTICES
WITH SOME SURGEONS PERFORMING LASER IN THE OR AND
SOME UNDER CONSCIOUS SEDATION, SUCH AS IN THE
NEONATAL NURSERIES.
Video 17 Stage 5 ROP Management by Open

Sky Vitrectomy in Case of Corneal Opacity
Open sky vitrectomy may be used in the case of severe corneal opacification
due to lens contact from stage 5 ROP. The surgical technique is presented in
detail. After positioning a Flieringa ring, and the infusion line at the limbus,
iris hooks incisions are made. Seven mm corneal trephination is done, the
adhesions of the iris to the cornea are carefully dissected, and the corneal
button is removed and preserved in media for corneal transplantation. The
preretinal fibrosis is dissected from the periphery toward the center 360
degrees. The anterior fibrosis is removed to allow dissection of the retinal
funnel. Bleeding is controlled by active aspiration and cautious diathermy.
The corneal button is replaced with 16 separated nylon sutures. The cornea
will clear some if Descemet membrane is not severely damaged.
Video 18 Management of Stage 5 ROP by

Closed Vitrectomy
Surgical approach of stage 5 ROP with clear media consists in closed
vitrectomy through limbal sclerotomies. Iris hooks are positioned to allow
good peripheral visualization, and iridectomies are done through the
sclerotomies to maintain longer corneal clearness. The lens is aspirated with
the vitreous probe and the capsule removed cautiously. Bimanual dissection
is conducted from the periphery toward the center, and peeling with two-
handed technique using forceps is useful. No tamponade is used at the end of
the procedure.
PLEASE NOTE: CLOSED TECHNIQUES CAN ALSO BE DONE
FROM THE CENTER OF THE MEMBRANE AND EXTENDED
PERIPHERALLY AS SHOWN IN CHAPTER 55.
All videos Courtesy Dr. Caputo.
1
Basics of Genetic Testing and
Methodology
Virginia Miraldi Utz and Diana Schorry Brightman
BACKGROUND
AAO Guidelines for Ordering Genetic Testing for
Inherited Eye Diseases (1)
Genetic t1esting should be performed by a clinician or genetic counselor
familiar with such testing.
Avoid direct-to-consumer testing.
When possible, use a Clinical Laboratory Improvements Amendments
(CLIA)-certified laboratory to ensure quality testing that meets federal
standards for dealing with human clinical specimens and providing a
formal report. (For example, a patient with genetic testing completed in
a non–CLIA-certified lab may be ineligible for a clinical trial until
confirmed by a CLIA-certified lab.)
Key Resource: http://www.ncbi.nlm.nih.gov/gtr/ (You can filter based on

CLIA certification, among other filters)
Use the patient's personal and family history when ordering tests to
identify the most specific test for the patient.
Consider genotype–phenotype correlations when available.
Avoid genetic testing for conditions that do not have a well-defined
molecular association, such as multifactorial conditions, like
nonsyndromic myopia.
The Essential Role of Genetic Counseling
Genetic counseling is recommended for individuals undergoing genetic
testing. This process includes the following goals:
Interpretation of family and medical history to assess the chance of
disease recurrence in future offspring.
Education about inheritance, testing, management, prevention,
resources, and research.
Counseling to promote informed choices and adaptation to the risk
or condition.
Medical geneticists are MDs who specialize in diagnosing, treating, and
counseling for genetic disorders.
Genetic counselors are allied health care professionals who have
specialized education in medical genetics and counseling to interpret
genetic test results and to guide patients through the genetic testing
process.
For more information on genetic counseling, please refer National
Society of Genetic Counselors webpage: https://www.nsgc.org
Ophthalmic geneticists are ophthalmologists who had done special
training and/or have a special interest in genetic eye disorders.
Ophthalmic geneticists diagnose, guide treatment and rehabilitation, and
counsel patients and families about genetic eye disorders. Ophthalmic
geneticists have the expertise to discuss prognosis as well as current
treatment and clinical trial options tailored to each disorder and to each
individual.
Medical geneticists, genetic counselors, and ophthalmic geneticists
working together provide the most comprehensive care to patients with
genetic eye disorders (2).
Key Concepts in Determining Pathogenicity of a

Variation
Definitions
1. Exome—refers to all DNA sequences encoding proteins
2. Genome—refers to all coding and noncoding regions of the DNA
Key Point: A typical patient's exome has 40,000 variants that will differ from
the reference sequence. A typical patient's genome has 3,000,000 variants
that differ from the reference sequence.
Mutation versus Variant

Variant: a change in the DNA sequence that differs from the reference
sequence (i.e., the DNA sequence in normal controls)
DNA directs the production of protein.
A DNA variant may change or abolish protein function, or it may
have little or no effect on protein.
If there is little or no effect on protein, it is termed a benign variant
or polymorphism.
If the effect on protein is unknown, it is termed a variant of
unknown significance (VUS).
If the protein is abolished or severely abnormal, the variant is likely
to cause disease and is called a disease-causing mutation.
Mutation: a genetic variation known to cause disease (i.e., a pathogenic
variant).
Copy number variants are areas of the genome that are either duplicated
or deleted. They can be classified based on size of the deletion or
duplication into single nucleotide variants (SNV), large and small
copy number variants (CNV), structural rearrangements, and
numerical changes. Deletions are only disease causing if they are large
enough and in the right location to delete part of all of a gene or genes,
or regulatory elements. The larger the deletion, the higher the chance of
having a pathogenic effect. Depending on the inheritance (autosomal
recessive vs. autosomal dominant vs. X linked) and the status of the
other allele will determine whether loss of a specific gene is
deleterious. For example, a deletion in an autosomal dominant gene
is likely to lead to disease through haploinsufficiency. Many normal
people have deletions that have no consequence to health. Small
duplications are relatively more benign (unless the gene is subject to
anticipation and trinucleotide expansion) but sometimes cause a
milder phenotype of the same kind that a deletion of that gene causes.
Large duplications (e.g., trisomy 21, 18, 13), however, can lead to
severe disease manifestations.
How does one determine if a variation is pathogenic? (3)
The probability of pathogenicity is determined in most labs by 28
criteria (see Richards et al., 2015). Some of the features of the criteria
include the following:
The segregation of the variant within families
The frequency of the variant in affected and unaffected individuals
The specificity of the phenotype for the gene
Bioinformatics to predict the effect on protein function
Conservation of the residue in other species through evolution
Allelism
Classification of a New Variant

Pathogenic variation—probability of pathogenicity is >99%
Likely pathogenic—probability of pathogenicity is between 90% and
99%
Variation of uncertain significance (VUS)—the probability of
pathogenicity is <90%
What Type of Variations Are Present in the Genome

(Figure 1-1)?
FIGURE 1-1
Genetic Testing Modalities (Table 1-1)
TABLE 1-1 Targeted (genotyping) vs. nontargeted

methods
BRIEF EXPLANATIONS OF
CYTOGENIC, ARRAYS, AND
SEQUENCING TECHNIQUES WITH
CLINICAL EXAMPLES
Cytogenic Techniques (Table 1-2)
TABLE 1-2
Array-Based Techniquesa (Table 1-3)
TABLE 1-3
aChromosome microarray (CMA) is also an array-based technique.
Sequencing Tests (Table 1-4, Figure 1-2)
TABLE 1-4
FIGURE 1-2 Algorithm for testing strategy of a patient
with a suspected genetic disorder. Pre- and posttest genetic
counseling is strongly recommended.
REFERENCES AND ADDITIONAL READINGS

1. Drack AV, Miraldi Utz V, Wang K, et al. Survey of practice patterns for the management of
ophthalmic genetic disorders among AAPOS members: report by the AAPOS Genetic Eye
Disease Task Force. J AAPOS. 2019;23(4):226-228.e1.
2. Stone E, Aldave A, Drack A, et al. Recommendations for genetic testing of inherited eye
diseases: report of the American Academy of Ophthalmology task force on genetic testing.
Ophthalmology. 2012;119(11):2408-10.
3. Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence
variants: a joint consensus recommendation of the American College of Medical Genetics and
Genomics and the Association for Molecular Pathology. Genet Med. 2015;17(5):405-424. doi:
10.1038/gim.2015.30.
2
Selected Pediatric Retinal Differential
Diagnosis Based on Clinical
Symptoms and Signs
Virginia Miraldi Utz and Elias I. Traboulsi
TABLE 2-1
aBoth usually have retinal manifestations (foveal hypoplasia), but the etiology of photodysphoria arises
largely from iris transillumination and hypoplasia, respectively.
bPatients with these diagnoses are not uniformly photodysphoric, initial symptoms depend largely on
whether cone dysfunction predominates in early phases of disease.
AD, autosomal dominant; ANCA, antineutrophil cytoplastic antibody; APS, antiphospholipid
syndrome; AR, autosomal recessive; BCM, blue cone monochromacy; BBS, Bardet Biedl syndrome;
BED, Bornholm eye disease (BED); BEM, bull's eye maculopathy; CHRPE, congenital hypertrophy of
the retinal pigment epithelium; cCSNB, complete congenital stationary night blindness; CNS, central
nervous syndrome; CPEO, chronic progressive external ophthalmoplegia; CRAO, central retinal artery
occlusion; CRD, cone–rod dystrophy; D-15, Farnsworth D-15 dichotomous color blindness test;
ffERG, full-field ERG; ED-OCT, enhanced-depth OCT; EEG, electroencephalogram; GVF, Goldmann
VF; HIV, human immunodeficiency virus; iCSNB, incomplete congenital stationary night blindness;
IOP, intraocular pressure; IP, incontinentia pigmenti; IVFA, intravenous fluorescein angiography;
LCA, Leber congenital amaurosis; LP, lumbar puncture; mfERG, multifocal ERG; MRI, magnetic
resonance imaging; MRV, magnetic resonance venography; NARP, neuropathy, ataxia and retinitis
pigmentosa; POFLs, pigmented ocular fundus lesions; PFV, persistent fetal vasculature (also known as
PHPV, persistent hyperplastic primary vitreous); PVL, periventricular leukomalacia; SD-OCT, spectral
domain optical coherence tomography; SECORD, severe early childhood onset retinal dystrophy;
SNHL, sensorineural hearing loss; SLE, systemic lupus erythematosus; RP, retinitis pigmentosa; RPE,
retinal pigment epithelium; TB, tuberculosis; TORCH, Toxoplasmosis, Other (Syphilis, Zika,
Parvovirus B19), Rubella, Cytomegalovirus, Herpes (simplex, zoster); VF, visual field; VKH, Vogt-
Koyanagi-Harada disease or Harada disease (just ocular involvement); XL, X-linked.
3
Online Resources for Pediatric
Genetic Retinal Disorders
Arlene Drack
INTRODUCTION
Online resources can be and are updated on a regular basis for sites related to
pediatric genetic eye disorders. Because of the rapid expansion of knowledge
in genetics, online resources listing genes that have been identified, which
mutations are pathologic versus normal variants, and which clinical trials are
recruiting are preferable to print, which will be out of date almost as soon as
they are published.
Genetic variation is responsible for the breadth and richness of human
characteristics. Because our knowledge of the human genome is young, our
understanding of the “normal” genetic sequences of genes is incomplete.
Normal human DNA sequences have been culled from a relatively small
number of people who were described as normal, but who may have harbored
genetic abnormalities of which they were unaware. In addition, they did not
represent the full panoply of normal human genetics across the globe.
Subsequently, the range of normal has been expanded and will continue to
be. Novel mutations in known disease-causing genes, new mutations, and
types of mutations in genes previously not connected to a specific phenotype
are also constantly being found. In addition, some variations initially thought
to be benign may actually contribute to polygenic or very common disorders.
In order for a variant in DNA to be classified as disease causing, it must be
proven that it changes the protein product of the gene in a way likely to
disrupt function, and it must not be more common in the general, healthy
population than the disorder is. Because certain variants, either benign or
deleterious, are more common in populations from certain ancestral groups
than others, unless the normative database includes a diverse population,
there will be errors in designation of variants (1). In addition, because some
disorders are not fully penetrant, that is, not everyone who has the
mutation(s) gets the disease, and some genetic conditions are phenotypically
heterogeneous with some affected people having all the features of a
syndrome and others only one or a few, our estimates of disease frequency
may be in error. In this case, mutations found in the reference population may
be deemed too common to cause the disease because the frequency of the
disorder is underestimated (2).
The sites below are a few examples of sites found to be especially useful
to clinicians caring for children with genetic retinal disorders. This list is not
exhaustive; many other sites exist and many more than are available at this
writing are likely to be developed in the years to come. The examples below
provide a starting point to using resources online to aid in the understanding
and treatment of our patients with genetic retinal disorders.
RETINAL INFORMATION NETWORK

(RETNET)
1. https:sph.uth.edu/retnet/
a. This site is a service of the Laboratory for the Molecular
Diagnosis of Inherited Eye Diseases, the Hermann Eye Fund,
Ruis Department of Ophthalmology and Visual Science, and
The Human Genetics Center, School of Public Health at The
University of Texas-Houston Health Science Center.
b. The site has tables of genes and loci that have been reported to
cause inherited retinal diseases.
i. Use this database:
1. To find out how many/which genes have been reported to cause a
specific genetic retinal disease
2. To find out which genetic retinal conditions have been reported to be
caused by a gene of interest
3. To search for genes that cause genetic retinal disorders by chromosome
ii. Case example
1. A 6-year-old child presents with poor night vision, decreased central
vision, and a history of polydactyly at birth, s/p surgical removal of
extra axial digits. You suspect Bardet-Biedl syndrome and wonder how
many genes can cause this disorder.
a. On the RetNet site, click on Home Page, then Diseases, then
Summaries. Table A lists number of genes and loci by disease.
Under Bardet-Biedl syndrome, the total number of genes and
loci known is 18. This was last updated July 1, 2019. The last
update date is always listed at the top of the table.
2. An adult patient presents with a lifelong diagnosis of Leber congenital
amaurosis (LCA). The patient is ready to start a family. She has no
family history of LCA. She wonders if there is any chance she can pass
this condition directly to her child, in an autosomal dominant fashion.
a. On RetNet, click on Home Page, then Disease Genes and Loci,
then Summaries. Go to Table B. Gene and Locus Symbols by
Disease Category. Under Leber congenital amaurosis, there are
two types: autosomal dominant and autosomal recessive. There
are 3 autosomal dominant genes that have been reported while
there are 23 autosomal recessive. You can tell your patient that
autosomal dominant transmission is possible, though not
common. You can choose a genetic testing panel that includes
all of these LCA genes, autosomal dominant and recessive. This
information was last updated July 1, 2019.
GENEREVIEWS
1. https://www.ncbi.nlm.nih.gov>books>NBK1116
a. GeneReviews is published by the University of Washington,
Seattle.
b. This resource is geared toward clinicians and provides
information on diagnosis, genetics, management, and genetic
counseling.
c. Each chapter focuses on a single phenotype/disease or a single
gene or summarizes the genetics of a common condition.
d. There are currently 759 chapters.
e. Each chapter is written by experts in the field, is peer reviewed,
and is updated at least every 2 to 4 years.
f. GeneReviews chapters are accessible through PubMed.
i. Use this resource
1. To quickly learn about the genes causing a specific condition
2. To quickly learn about the clinical manifestations of a specific condition
3. To review what clinical findings might be present in a patient with
mutation or deletion of a specific gene
ii. Case example
1. A 10-year-old boy is being referred for night blindness. Visual acuity is
20/30, and there is no bone spicule like pigmentation. You suspect X-
linked congenital stationary night blindness. What is the appropriate
workup?
a. Access GeneReviews. Search X-linked congenital stationary
night blindness in the search box.
i. Under Diagnosis/testing, an ERG and genetic testing of the genes
CACNA1F and NYX are suggested. Description of the
characteristic ERG waveform is provided. Different approaches to
genetic testing are outlined. Under Suggestive Findings, myopia is
listed, which can be ascertained on clinical exam.
PUBMED
1. https://www.ncbi.nlm.nih.gov>pubmed
a. This is a free search engine of references and abstracts primarily
in the life sciences and biomedical research. It is maintained by
the United States National Library of Medicine at the National
Institutes of Health.
i. Use this database
1. To find the latest research publications on a specific disease or gene
2. To find high-quality scientific summary and survey articles on a topic
ii. Case example
1. An infant presents with nystagmus and very poor vision. You suspect
LCA. Parents would like to know whether research is being done on
LCA and if so what have the results been.
a. Access PubMed. Enter Leber congenital amaurosis in the search
space.
i. 996 articles are listed ranging from treatment with topical
brinzolamide for associated cystoid macular edema to numerous
gene-specific subtypes such as RDH12 and GUCY2D, to animal
model studies in zebrafish and mice. This gives an overview of
the many facets of research into this condition; you can tell the
parents that this is a very active field of research and there are
many different genetic subtypes—genetic testing or referral to
medical genetics for testing is indicated.
ii. Now, narrow your search to Leber congenital amaurosis and
clinical trial. Results from clinical trials for RPE65 and CEP290
LCA as well as overview articles on retinal gene therapy come up.
These can be summarized or printed for parents.
CLINICALTRIALS.GOV
1. www.clinicaltrials.gov
a. A database of clinical trials provided by the U.S. National
Library of Medicine.
b. As of 9/28/19, there were listings for 317,735 research studies
being conducted in 50 states and 209 countries, including both
privately and publicly funded studies.
c. The U.S. government does NOT evaluate vet or endorse studies
listed on this site, it simply maintains the list.
1. To learn about which clinical trials are being conducted for specific
diseases and molecular genetic diagnoses
2. To learn which clinical trials are recruiting and what the inclusion and
exclusion criteria are
3. To find contact information for researchers conducting clinical trials
ii. Case example
1. An 18-year-old male patient presents with decreased vision and a strong
family history of choroideremia. He is interested in learning about any
clinical trial that he may be able to enroll in.
a. Go to clinicaltrials.gov. In the search box, type choroideremia.
Twenty studies come up; 6 have been completed; 1 of these has
results available, which can be reviewed and shared with the
patient. Five are recruiting, but only 2 of these are
interventional studies, the others are observational. One of the
interventional studies is a phase 2 safety trial open to males 18
years of age or older with molecular genetically confirmed
choroideremia. The other is a phase 3 efficacy study with the
same inclusion criteria. Some patients will be randomized to a
nontreatment control group for a year in the phase 3 study.
Contact information is given for both studies. Thus, there are
several options with different pros and cons for this patient, and
the first step is molecular genetic testing. A clinical diagnosis,
even with family history, is not adequate to be considered for
the studies.
EXOME AGGREGATION
CONSORTIUM (EXAC)
1. exac.broadinstitute.org
a. This is a collaborative database produced by a coalition of
investigators.
b. As of 9/28/19, it included data on 60,706 individuals who had
their DNA sequenced as part of disease-specific and population
genetic studies.
c. It gives information on how frequent individual variants and
disease-causing mutations are in a wide array of genes.
1. To study a gene of interest
2. To look into whether a variant found in a patient is likely to be disease
causing
ii. Case example
1. A 10-year-old patient presents with a year long history of decreased
central vision. Fundus appears normal. OCT shows disruption of the
outer nuclear layer in the fovea. Genetic testing for juvenile
maculopathy reveals 2 variants in the ABCA4 gene, one a known
disease-causing mutation, the other a variant thought to be benign.
ExAC can be consulted to further understand the unknown variant.
a. In ExAC, ABCA4 is noted to have 2,342 different variants
reported.
b. The patient's second variant, c.6764 G>T (p.Ser2255Ile),
1:94461717 C/A, has been found as homozygous variants in
1,273 unaffected people, making it very unlikely to be disease
causing.
c. The frequency of this allele across populations is 0.08, and in
people of African descent, it is 0.49; these frequencies are too
high for a disease as rare as autosomal recessive Stargardt
disease.
d. Conclusion: The second variant found is not likely to be disease
causing. Either the patient has a second, occult Stargardt
disease causing mutation, or the patient has a phenocopy of
Stargardt and another etiology of their maculopathy must be
suspected. One caveat: Some variants that are benign in the
homozygous state can be disease causing if they occur with a
much more damaging allele.
REFERENCES
1. Whiffin N, Ware James S, O'Donnell-Luria A. Improving the understanding of genetic variants
in rare disease with large-scale reference populations. JAMA 2019;322(13): 1305–1306.
2. Lek M, Karczewski KJ, Minikel EV, et al. Analysis of protein-coding genetic variation in
60,706 humans. Nature 2016;5369(7616):285–291.
4
Genetic Mutations and Related
Proteins Associated with Inherited
Retinal Diseases
Robert S. Molday
GENERAL CONSIDERATIONS
Inherited retinal diseases are a heterogeneous group of disorders that
represent a significant cause of blindness in the world. They are typically
characterized by the loss in vision resulting from mutations in genes that
encode proteins essential for photoreceptor cell function and/or survival.
Over 271 genes are now known to cause various retinopathies, and the
chromosomal loci of another 36 disease-causing genes have been mapped
using a variety of genetic strategies including candidate gene approaches,
linkage analyses, and next-generation sequencing (NGS)
(http://www.sph.uth.tmc.edu/RetNet/). This accounts for over 85% of the
inherited retinal diseases and includes the most prevalent retinopathies (1,2).
With continued genetic screening of affected patients, the remaining genes
associated with loss in vision will be identified in the near future. These
studies will also provide additional information on the number and types of
disease-causing mutations thereby expanding our understanding of genetic
and molecular mechanisms responsible for retinal diseases, advancing our
knowledge on phenotype–genotype correlations, and importantly serving as a
basis for developing novel rationale treatments that will slow or prevent
vision loss in affected patients.
A significant fraction of genes linked to inherited retinal diseases are
solely or predominantly expressed in developing or adult photoreceptor cells
or retinal pigment epithelial (RPE) cells. These genes encode proteins that
play crucial roles in any of a wide variety of biochemical pathways essential
for the development, structure, function, or survival of these cells. They
include the visual cycle, phototransduction, photoreceptor outer segment
structure and renewal, cilium structure and transport, protein trafficking,
RNA splicing, lipid and nucleotide metabolism, translation and transcription,
posttranslational protein modifications, extracellular matrix, cell–cell and
cell–matrix adhesion, and synaptic structure and function among others
(Figure 4-1). Some genes associated with inherited retinal diseases are
expressed in other tissues as well as the retina. Mutations in these genes may
trigger phenotypes associated with these tissues. Such diseases are referred to
as syndromic diseases. In other cases, however, defective genes are expressed
in a wide variety of tissues as well as the retina, but the phenotype associated
with the genetic defect is only apparent in the retina. Such genes may have
evolved to encode proteins that play essential roles in retinal cell biology but
only redundant functions in other tissues.
FIGURE 4-1 Schematic diagram of a cone and rod

photoreceptor cell and RPE cells showing subcellular
organization and many of the proteins and cellular
processes/pathways associated with inherited retinal
diseases in these cells. (STGD1, autosomal recessive
Stargardt macular degeneration; LRAT, lecithin retinol
acyltransferase; LCA, Leber congenital amaurosis; RDH,
retinol dehydrogenase; IRBP, interphotoreceptor retinoid-
binding protein; RP, retinitis pigmentosa; RGR, RPE-
retinal G-protein-coupled receptor; CSNB, congenital
stationary night blindness; GNAT1, rod transducin α-
subunit; PDEA/B/G, phosphodiesterase ◻◻
◻◻◻◻subunits; arrestin; GRK1, G-protein coupled
receptor kinase 1; CNGA1/B1, rod cyclic nucleotide-gated
channel ◻◻and ◻; NCKX1, rod Na/Ca-K exchanger;
GC1, guanylate cyclase 1; CRD, cone–rod dystrophy;
GCAP1/2, guanylate cyclase activating protein 1 and 2;
CD, cone dystrophy; CNGA3/B3, cone cyclic nucleotide-
gated channel subunits α and β; GNAT2 cone transducin
α-subunit; ACHM, achromatopsia; PDE6C/H, cone
phosphodiesterase α and γ; MD, macular dystrophy;
digRP, digenic RP; RCD, rod–cone dystrophy; USH,
Usher syndrome; BBS, Bardet-Biedl syndrome; CHM,
choroideremia; ESC, enhanced S-cone syndrome; SFD,
Sorsby fundus dystrophy; ML, Malattia Leventinese.)
In addition to the clinical and genetic analysis of retinal diseases, the

molecular and cellular characterization of the proteins encoded by disease-
causing genes is an essential step in understanding pathogenic mechanisms.
Determining how disease-causing mutations affect the expression,
localization, structure, and function of a protein is crucial for a complete
understanding of disease mechanisms. Importantly, knowledge of disease
mechanisms is critical for designing and developing promising therapeutic
treatments for these diseases (3).
A large repertoire of established and newly emerging biochemical,
molecular, cellular, physiologic, optical imaging techniques, and diagnostic
procedures together with valuable animal models has greatly enhanced our
knowledge of retinal diseases. This has led to a significant increase in
preclinical and clinical trials for a number of pediatric retina diseases (3, 4, 5)
and is highlighted by the successful gene therapy trials for Leber congenital
amaurosis type 2 (LCA2) associated with mutations in the retinoid
isomerohydrolase RPE65 (6, 7, 8, 9) (see also Chapter 28). Recent advances
have also been made in the areas of cell and drug therapies, some of which
are in clinical trials. This chapter provides an overview of genetic defects that
underlie inherited retinal diseases and key biochemical pathways and cellular
processes that are compromised.
GENES
Genetic studies have highlighted the extreme heterogeneity of retinal diseases
(2,10). Clinically defined disease phenotypes often arise from mutations in
any of a large number of different genes. To date, over 80 genes are currently
known to cause retinitis pigmentosa (RP); 27 genes are associated with cone
dystrophy (CD) and cone–rod dystrophies (CRDs); 14 genes have been
linked to Leber congenital amaurosis (LCA1–14); 15 genes are associated
with Usher syndrome; 18 genes are responsible for Bardet-Biedl syndrome
(BBS); and 13 genes are responsible for congenital stationary night blindness
(CSNB) (for an updated listing, see the RetNet Web site:
http://www.sph.uth.tmc.edu/RetNet/) (see also Section on Genetics in
Pediatric Retina and Resources in e-book). Additional genetic screening and
NGS methods including whole-exome and whole-genome NGS will add new
genes to the list of inherited retinal diseases (11, 12, 13).
The inheritance patterns can vary for many retinal diseases. This is best
exemplified for RP, a retinal degenerative disorder typically involving the
degeneration of rod photoreceptor cells and progressive loss in cone cells
(14). The pattern of inheritance can be autosomal dominant, autosomal
recessive, X linked, or in rare cases maternal mitochondrial and digenic. In
the case of autosomal dominant RP (ADRP), a disease-causing mutation in
only one allele is sufficient to cause the progressive loss in vision associated
with the degeneration of photoreceptor cells; for autosomal recessive RP
(ARRP), a disease-causing mutation in each allele has to be present; and for
X-linked RP (XLRP), the defective gene is mapped to the X chromosome.
Currently, 22 genes are known to cause ADRP accounting for about 30% to
40% of RP cases, 41 genes have been implicated in ARRP representing 50%
to 60% of RP cases, and 2 known genes are identified for XLRP accounting
for 5% to 15% of the RP cases of known inheritance pattern. The mode of
inheritance of a large number of RP cases is unknown. These are commonly
referred to as simplex cases. Mutations in the RHO gene encoding rhodopsin
account for approximately 30% of the ADRP cases, mutations in the USH2A
gene encoding usherin, a large membrane protein containing numerous
extracellular domains, account for up to 17% of ARRP cases, and mutations
in RPGR encoding the X-linked RP GTPase regulator, RPGR, that functions
in cilium structure and function account for about 80% of the genetically
identified forms of XLRP (14). In rare cases, RP can arise through digenic
inheritance involving mutations in two different genes. Individuals with
either a L185P mutation in peripherin-2 (PRPH2) or a null mutation in
ROM1 are unaffected, whereas individuals who inherit both mutations
display a RP phenotype (15).
In a number of cases, a given disease phenotype arises from mutations in
a single gene. For example, autosomal recessive Stargardt macular
degeneration (STGD1) is caused by mutations in the ABCA4 gene encoding
the photoreceptor ATP binding cassette transporter ABCA4 (16);
choroideremia is associated with mutations in the CHM gene encoding the
rab escort protein 1, REP1 (17); X-linked juvenile retinoschisis (XLRS1) is
caused by mutations in the RS1 gene encoding the extracellular protein
retinoschisin (also known as RS1) implicated in cell–cell adhesion (18);
Sorsby fundus dystrophy is linked to mutations in TIMP3 gene encoding the
metalloproteinase inhibitor 3 protein, TIMP3 (19); and Best vitelliform
macular dystrophy (BEST1) is caused by mutations in the VMD2 gene
encoding the chloride channel bestrophin-1 in RPE cells (20).
In most cases, any one of a large number of different mutations within the
gene can cause a given clinically defined disease. Over 170 mutations in
RHO are known to cause ADRP, 150 mutations in RPGR cause XLRP, 130
mutations in RS1 cause X-linked retinoschisis, and over 1,000 mutations in
ABCA4 cause autosomal recessive Stargardt macular degeneration and related
retinopathies. The specific mutation within a gene can have a major influence
on the phenotypic features of the disease. Some mutations can result in a
significant reduction in protein expression and accordingly cause a severe,
early-onset disease, whereas others may only partially affect protein
expression leading to a milder, later-onset disease. In some cases, clinical
diagnosis of individuals with early- and late-onset diseases results in distinct
clinical disease classifications. For example, early diagnosis of disease-
causing mutations in ABCA4 reveals characteristics of Stargardt disease,
whereas later stages of the disease may be more characteristic of CRD (21,
22, 23, 24).
In many cases, however, a clinical phenotype can arise from any of a
number of different genes encoding proteins that function in distinct cellular
processes. Genes associated with RP encode proteins that function in
phototransduction, the visual cycle, outer segment disk structure and
morphogenesis, cilium structure and protein trafficking, pre-RNA splicing,
and transcription among others. Defects in genes associated with LCA, a
severe early-onset retinopathy in which children are born with little or no
vision, typically encode key proteins required for photoreceptor or RPE cell
survival and in some cases development. These genes are often
predominantly expressed in both rod and cone photoreceptor cells or RPE
cells required for photoreceptor survival.
Heterogeneity of retinal degenerative diseases is further evident in the
finding that different mutations in a given gene can cause clinically distinct
diseases. Some missense mutations in the PRPH2 gene encoding the
photoreceptor structural protein peripherin-2 (also known as peripherin/rds or
PRPH2) are associated with ADRP, while other mutations in PRPH2 cause
autosomal dominant pattern macular dystrophy (25,26). Genetic defects in
the GUCY2D gene encoding guanylate cyclase 1 (GC1 or RetGC1) can cause
autosomal recessive LCA type 1 (LCA1) or autosomal dominant CRD
(27,28). Mutations in RPE65 localized in RPE cells have been reported to
cause autosomal recessive LCA2, ARRP, or ADRP depending on the
mutation and diagnosis (29, 30, 31). Furthermore, different family members
with the same genetic defect may exhibit different clinical phenotypes as in
the case of different retinopathies associated with a given mutation in the
PRPH2 gene (32). This highlights the difficulty in assigning a disease
phenotype to a genetic mutation. Although the genotype plays a central role
in determining the disease phenotype, other factors including modifier genes,
age of diagnosis of the affected individual, and socio-environmental factors
can alter the clinical description of the disease.
The candidate gene approach was first used to identify genes associated
with retinal degenerative diseases. In this approach, patients with RP or other
diseases were screened for mutations in genes encoding well-characterized
photoreceptor-specific proteins. A mutation in the RHO gene was first
identified by this approach 30 years ago and shown to cause ADRP (33).
Subsequently, individuals with RP were screened for mutations in the PRPH2
and PDE6B, genes which had previously been reported to cause retinal
degeneration in the rds and rd1 mice, respectively (34,35). Genetic screens
have confirmed that mutations in most genes encoding key proteins involved
in phototransduction, the visual cycle, and photoreceptor structure are
responsible for various types of retinopathies (Figure 4-2).
FIGURE 4-2 Diagrams showing the principal proteins

involved in phototransduction in rod outer segments and
their association with various inherited retinal diseases.
Light isomerizes 11-cis retinal to all-trans retinal (not
shown) within rhodopsin. Activated rhodopsin catalyzes
the exchange of GDP for GTP on transducin α-subunit.
The transducin α-subunit dissociates from the β, γ-subunits
and binds to the inhibitory subunits of phosphodiesterase
(PDE). This activates the catalytic activity of PDE
resulting in the hydrolysis of cGMP to 5'GMP. As the
cGMP level falls, cGMP dissociates from the cyclic
nucleotide–gated channel causing the channel to close.
This prevents the influx of Na+ and Ca2+ into the outer
segment and leads to a hyperpolarization of the cell. The
cell returns to its dark state through (a) the inactivation of
rhodopsin via phosphorylation by rhodopsin kinase
(GRK1) and the binding of arrestin; (b) inactivation of
PDE when GTP bound to the transducin α-subunit is
hydrolyzed to GDP, a reaction catalyzed by RGS9/R9AP
complex; (c) restoration of cGMP levels through the
activation of guanylate cyclase (GC1) via the effect of low
Ca2+ levels on the guanylate cyclase activating protein
GCAP. The latter is caused by the closure of the cGMP-
gated channel (CNGA1/B1) and continued extrusion of
Ca2+ by the Na/Ca, K exchanger (NCKX1); and (d)
reopening of the cyclic nucleotide-gated channel as cGMP
binds to the channel resulting in the return of the cell to its
depolarized state. Diseases associated with various
proteins: RP, retinitis pigmentosa; CSNB, congenital
stationary night blindness; CD, cone dystrophy; CRD,
cone–rod dystrophy; ACHM, achromatopsia (cones);
LCA, Leber congenital amaurosis; bradyopsia.
In some instances, genes associated with retinal diseases are expressed in

other tissues as well as the retina and cause syndromic diseases. Usher
syndromes characterized by RP and deafness are typically caused by
mutations in genes encoding proteins, which are crucial for the development,
function, and survival of photoreceptors and hair cells or related structures of
the inner ear (36). Defects in more widely expressed genes can result in
pleiotropic disorders. BBS is an example in which mutations in a number of
genes crucial for basal body and ciliary biogenesis, structure, and function are
responsible not only for RP but also obesity, polydactyly, diabetes, mental
and growth retardation, auditory deficiencies, and developmental delays (37).
Finally, some genes are expressed in virtually all cells, but loss in function
mutations only cause retinal degenerative diseases. This is exemplified for
mutations in the ubiquitously expressed pre-RNA splicing genes PRPF3,
PRP31, and PRPC8, which cause ADRP. Likewise, RPGR and RP2
associated with XLRP are ubiquitously expressed, but most disease-causing
mutations only cause loss in vision due to the degeneration of photoreceptor
cells.
GENETIC MUTATIONS
A wide variety of genetic mutations are known to cause retinal degenerative
diseases. These include nonsense mutations, indels (insertions or deletions),
splice site, and missense mutations. Nonsense mutations produce premature
stop codons typically resulting in a shortened, nonfunctional protein.
Insertions and deletions that are not divisible by 3 typically cause a
downstream frameshift that results in an altered, often truncated protein.
Together with nonsense mutations, indels are often considered as null alleles
since the altered protein lacks function and is typically unstable and rapidly
degraded by the cell. Splice site mutations often result in defective translation
leading to a null allele, although in some cases defective splicing may be
incomplete resulting in reduced levels of the transcripts and encoded protein
(38).
Disease-causing missense mutations result in the substitution of a key
amino acid for another amino acid. Such substitutions can produce a highly
misfolded, nonfunctional protein that is retained in the endoplasmic reticulum
(ER) by the quality control system of the cell and rapidly degraded by the
proteasome (39). Alternatively, a missense mutation may result in a stable
protein, but its function and/or its cellular localization is drastically impaired
(40, 41, 42, 43, 44). Such mutations can result in either a gain or loss in
function. Most missense mutations in the RHO gene that cause ADRP result
in altered expression, localization, and functional properties of rhodopsin
leading to progressive rod photoreceptor cell death (45,46). Autosomal
dominant diseases can also be caused by haploinsufficiency. In this instance,
the protein encoded by the nonmutated (wild-type) allele is produced in
quantities below that required for normal cellular function and/or survival.
This is evident for some mutations in PRPH2 encoding the outer segment
structural protein peripherin-2 and linked to autosomal dominant
retinopathies (47). Most missense mutations linked to recessive retinal
diseases cause protein misfolding and/or a loss in function due to the removal
of a key amino acid (40,43,48,49).
Although most disease-associated mutations are found within the coding
regions of the genes, in some instances, disease causing mutations have been
found in noncoding regions. NGS has been particularly useful in identifying
and locating disease-causing mutations within introns of genes as in the case
of mutations in ABCA4 associated with STGD1 (50,51).
Many studies have been directed toward correlating genotypes with
phenotypes for specific retinal degenerative diseases (32,40,52, 53, 54). For
the most part, this has been a daunting task as the phenotype for many
diseases not only results from the specific mutation but also from other socio-
environmental and genetic factors. In a number of cases, the disease severity
has been shown to be dependent on the residual expression and functional
activity of the mutated protein (40). Nonsense, frameshift mutations, and
splice site mutations that effectively result in null alleles often cause an
earlier onset and more severe phenotype, whereas missense mutations that
produce proteins that retain residual functional activity may result in less
severe, late onset, phenotype (10). In some cases, however, the expression of
a highly misfolded protein can result in a more severe phenotype due to
cellular stress related to the expression and removal of the misfolded protein.
PROTEINS AND MODEL SYSTEMS

The molecular and cellular characterization of the wild- type and mutant
proteins is an essential step in understanding pathogenic mechanisms
underlying inherited retinal diseases. This includes identifying cells
expressing the protein of interest; determining protein expression profiles
during retinal development and cell differentiation; localizing the protein to
subcellular compartments in cells; evaluating splice variants if present;
analyzing the structural, functional, and regulatory properties of the proteins;
identifying interacting proteins; defining their roles in cellular pathways and
related processes; determining how disease-causing mutations affect protein
localization, structure, function, and stability; and elucidating the biochemical
pathways that lead to cell death. Proteins encoded by disease-associated
genes have been found to participate in a wide variety of cellular processes
(Figure 4-1). The roles of a significant number of disease-associated proteins
in retinal cell biology and pathology, however, remain to be elucidated at a
molecular level.
An understanding of the role of proteins encoded by specific disease-
associated genes has been facilitated through analysis of model systems
(42,45,46,55,56). Rodents (mice and rats), zebrafish, and Xenopus laevis
have been most widely used to investigate the effect of genetic mutations on
vertebrate retinal cell structure and function and as models for human
retinopathies. Mice harboring naturally occurring genetic mutations
responsible for photoreceptor degeneration have been effectively used to
identify disease-causing genes and define the role of the encoded proteins in
photoreceptor cell structure and function (57). Early studies focused on the
rd1 mouse displaying a rapid degeneration of rod photoreceptors and the rds
mouse exhibiting a slow degeneration of rod and cones. Genetic and
biochemical studies showed that the rd1 mouse harbored a mutation in the
Pde6B gene encoding the beta subunit of phosphodiesterase (PD6B), a key
enzyme involved in the hydrolysis of cGMP (58,59). A nonfunctional
PDE6B subunit resulted in the accumulation of toxic levels of cGMP and a
rapid loss in rod photoreceptor cells. The rds mouse (also known as rd2) was
also a subject of intense investigation. Homozygous rds mice fail to produce
outer segments and heterozygous mice develop highly disorganized outer
segments (60). Genetic studies showed that the underlying defect was a viral
insertion in Prph2 gene encoding peripherin-2, a tetraspanin membrane
protein localized along the rim region of photoreceptor outer segment disk
membranes (61, 62, 63). These studies subsequently led to the identification
of mutations in the human PDE6B gene that cause a subset of autosomal
recessive RP and mutations in the human PRPH2 gene that cause autosomal
dominant RP and autosomal dominant pattern macular degeneration
(26,34,35).
The RCS rat has also been widely used to identify the biochemical and
genetic defects responsible for retinal degeneration in this rodent. Wide
ranging studies showed that the RCS rat is deficient in phagocytosis of aged
outer segments due to a genetic defect in the Mertk gene (64). Subsequent
genetic screens showed that mutations in the human MERTK gene are
responsible for a subset of autosomal recessive RP (65). Although naturally
occurring mutant mice continue to be useful for the analysis of retinal
diseases (66), most studies now use genetic techniques to produce mice in
which the gene of interest is knocked out or replaced with a disease-causing
mutant gene (44,67, 68, 69). These studies are facilitated through the recent
development of Crispr/Cas9 gene editing technology.
The effect of the mutated gene on retinal structure and function can be
analyzed in knockout and transgenic mice using a wide variety of
noninvasive methods. Optical coherence tomography (OCT) is a valuable
technique to assess retinal structure, whereas electroretinography (ERG) and
optokinetic analysis can be used to assess visual function. Scanning laser
ophthalmoscopy is also useful for measuring fluorescent lipofuscin deposits,
which accumulate in photoreceptors and RPE cells in animal models and
patients with certain retinal diseases (44,70). These techniques complement
the widely used light and electron microscopic analysis of retinal sections and
cellular and biochemical studies including proteomics designed to measure
the functional activities of the proteins and their interaction with other
proteins (56,66,71).
Although rodents have been widely used to study retinal diseases,
zebrafish and Xenopus laevis have also served as important animal models.
These animals have a number of advantages. They are less expensive to
generate and maintain than rodents, easier and faster to produce in large
numbers, and typically have large rod and cone photoreceptors for structural
and functional analysis. In addition, the phenotype of the variants can be
readily analyzed using imaging, physiologic, genetic, and cell biology
techniques to elucidate molecular and cellular mechanism underlying retinal
diseases and determine the disease progression (72,73). However, as they are
evolutionarily more removed from humans than rodents, the modified gene
may not always replicate the phenotype observed for the human disease.
An in depth understanding of the cellular and molecular basis for retinal
degenerative diseases in animal models can be obtained using
immunocytochemical and biochemical analysis. For analysis by
immunocytochemistry, the retina tissue from sacrificed mice can be fixed and
labeled with specific antibodies to analyze the effect of gene mutation on the
localization of proteins in the retina of animal models using fluorescence
and/or electron microscopy (71). At a molecular level, biochemical
techniques including immunoprecipitation methods and protein analysis can
be used to gain insight into how mutations in proteins affect their structure,
function, and association with interacting proteins (42). In one example, the
function of RD3, a small protein encoded by a gene linked to LCA12, was
elucidated by comparing the localization and interaction of the RD3 protein
with other photoreceptor proteins (66). These studies revealed that in the
absence of RD3 expression in the rd3 mouse, guanylate cyclases (GC1 and
GC2) failed to traffic to photoreceptor outer segment, but instead were
retained in the ER of photoreceptor cells. Protein interaction studies
confirmed the direct interaction of RD3 with GC1 and GC2 in the retina of
wild-type mice. Accordingly, one function of RD3 is to serve as a chaperone
protein promoting the folding and trafficking of guanylate cyclase to outer
segments (48). RD3 has also been shown to inhibit guanylate cyclase activity,
a property which is important in preventing the production and accumulation
of cGMP in the inner segment of photoreceptor cells where it may activate
proteins that cause photoreceptor degeneration (74). Interestingly, mutations
in RD3 and GC1 cause LCA1 and LCA12, respectively, consistent with their
importance in a common key photoreceptor cellular pathway (75,76).
In many cases, the characteristic features observed in mice and other
animal models compare favorably to those of the human disease and,
therefore, they serve as a valuable system for detailed analysis of the role of
the proteins in retinal cell biology and physiology and excellent models for
the development and assessment of therapeutic treatments for specific
diseases (46,55,56,77). However, in some instances, the phenotype of the
knockout mice does not correlate well with the phenotype observed in
humans. For example, a number of mouse models for Usher syndrome show
little if any retinal degeneration that are observed in humans with Usher
syndrome although they typically exhibit a deficiency in hearing (78).
Likewise, a knockout mouse may not represent a useful model for some
diseases including many autosomal dominant inherited diseases. In these
cases, it may be necessary to develop transgenic mouse models in which the
mutated gene is expressed along with the normal gene or in which the
mutated gene replaces the normal gene to more fully evaluate the effect of the
mutation on retinal structure and function (77,79).
In addition to animal models, cell culture systems have served as valuable
tools to understand the effects that disease-associated mutations have on
protein expression, localization, structure, and function. Typically, plasmids
containing the wild-type and mutant cDNAs are used to transfect HEK293,
COS7, CHO, or other cells for the transient or stable expression of proteins.
Mutant protein expression and localization can be determined by western
blotting and immunofluorescence microscopy for comparison with the wild-
type protein (40). The extent of expression provides a reliable indicator of the
stability of a mutant protein relative to wild-type counterpart. Low expression
of mutant proteins generally reflects severe protein misfolding,
mislocalization, and rapid degradation by the cell (80). Mutants that express
at sufficient levels (>25%) are suitable for further functional analysis.
Immunofluorescence microscopy using cell-specific markers provides a
means for comparing the localization of expressed mutant protein with the
wild-type protein (43,44,48). If a high-quality antibody is not available, then
the protein of interest can be engineered with an epitope tag or green
fluorescence protein (GFP) at one end of the protein. In most cases,
membrane proteins are synthesized in the ER of the cell and transported to
their cellular location via vesicle trafficking mechanisms. Mutations can
affect the folding of the protein or the targeting sequence resulting in
mislocalization. Indeed, many mutated membrane proteins are misfolded and
retained in the ER by the quality control system of the cell and hence exhibit
a markedly different distribution than the wild-type protein (43,45,66,81). ER
localization can be confirmed using a specific ER marker such as calnexin
(71). In some cases, only a fraction of the mutated variant may be misfolded
and hence the cellular distribution may be characteristic of both the wild-type
and mutant protein (82). Some proteins containing ER retention sequences
are naturally retained in the ER such as ELOVL4, an enzyme associated with
very long chain fatty acid elongation (83). It should be noted, however, that
the localization of proteins overexpressed in culture cells may not necessarily
reflect their localization in situ. Hence, it is important to combine these
heterologous expression studies with immunocytochemical labeling in retina
tissue or other suitable natural cell sources to more accurately define their
true cellular localization (81).
Culture cell expression is also useful for analyzing the functional
properties of wild-type and mutant proteins. In some cases, the function of a
membrane protein can be analyzed in extracts of cells expressing the protein
of interest. Alternatively, it may be necessary to purify the protein for direct
functional analysis (84,85). The purification of proteins can be achieved
using either conventional chromatographic methods or affinity techniques
(41,82). Comparison of the functional properties of the mutant proteins with
the wild-type proteins can provide important insight into the effect of
mutations of the structure and function of the proteins.
BIOCHEMICAL PATHWAYS AND

CELLULAR PROCESSES ASSOCIATED
WITH RETINAL DISEASES
Phototransduction
The principal function of rod and cone photoreceptor cells is to carry out
phototransduction, the process by which light entering the eye is captured and
converted into an electrical signal (86, 87, 88, 89). The signal is then
transmitted to secondary neurons in the retina where it is further processed
and finally sent to the brain via the optic nerve for visual perception.
Phototransduction in rod cells is initiated when a photon of light converts
11-cis retinal (11-cis retinaldehyde) to its all-trans isomer within the G-
protein–coupled receptor rhodopsin (Figure 4-2). This transforms rhodopsin
from its resting state to its activated R* or Met II state. Activated rhodopsin
catalyzes the exchange of GDP for GTP on the α-subunit of the trimeric G-
protein transducin. The GTP-α-transducin subunit (GNAT1) dissociates from
the β,γ subunits of transducin and binds to the inhibitor subunits of
phosphodiesterase, a protein complex consisting of two catalytic subunits
(PDE6A and PDE6B) and two identical inhibitor subunits (PDE6G). The
binding of GTP-bound transducin releases the inhibitory restraint on the
enzyme allowing PDE6A:PDE6B complex to efficiently catalyze the
hydrolysis of cGMP to 5'GMP. In the dark, a significant number of cyclic
nucleotide–gated (CNG) channels in the plasma membrane are maintained in
their open state through the binding of cGMP to the channels. The reduction
in intracellular cGMP following photoexcitation causes cGMP to dissociate
from the CNG channel, a heterotetrameric complex of 3 CNGA1 subunits
and 1 CNGB1 subunit (90). As a result, the CNG channels close leading to a
reduction in the influx of Na+ and Ca2+ into the outer segment, a
hyperpolarization of the photoreceptor cell, and an inhibition of glutamate
release at the rod cell synapse. Closure of the CNG channels also causes a
decrease in intracellular Ca2+ levels as Ca2+ continues to be extruded from
the rod outer segment through the sodium–calcium–potassium exchanger
(NCKX1) in the plasma membrane of rod cells.
Following photoactivation, the rod cells recover to its dark state through a
series of biochemical reactions that 1) turn off reactions that were initiated
through photoactivation and 2) restore intracellular cGMP to its dark state
level (86,89). Rhodopsin is deactivated by phosphorylation of serine and
threonine residue at its C-terminus, a reaction catalyzed by rhodopsin kinase
(GRK1). Arrestin binds to phosphorylated rhodopsin further inactivating
rhodopsin. Transducin is returned to its dark state through the hydrolysis of
bound GTP to GDP on the GNAT1 subunit, a reaction that is catalyzed by the
GAP protein RGS9 (91). This also returns phosphodiesterase to its basal level
of activity as the GDP-α-transducin dissociates from the inhibitory subunit
PDE6G. GDP-α-transducin recombines with the β,γ subunits of transducin to
form the inactive transducin trimeric complex. Finally, guanylate cyclase
isoforms (GC1 found in both rods and cones, and GC2 expressed only in
rods) are activated by low Ca2+ levels via their regulation by guanylate
cyclase activating proteins (GCAP1 and GCAP2) (92). The activated
guanylate cyclases catalyze the conversion of GTP to cGMP leading to a rise
in intracellular cGMP levels and restoration of the CNG channels to their
open state. As a result, the Ca2+ concentration also increases to its dark state
level as Ca2+ enters the rod outer segment through the CNG channels. This
returns activated GC1 and GC2 to their basal “dark state” level of activity.
Cone photoreceptors use a similar phototransduction mechanism,
although several of the key proteins are encoded by different genes (93). In
particular, cone photoreceptors express distinct cone opsins (blue, green, and
red opsin), α-transducin subunit (GNAT2), PDE subunits (two catalytic
PDE6C subunits and two inhibitory PDE6H subunits), and CNG channel
subunits (CNGA3, CNGB3). Differences in the functional and regulatory
activities of the proteins encoded by cone- and rod-specific genes are
responsible, in part, for the differences in color vision, sensitivity to light, and
response kinetics between cone and rod cells.
The importance of phototransduction signalling in rod and cone function
and survival is evident in the finding that genetic defects in most genes
encoding proteins involved in rod and cone phototransduction are responsible
for any of a variety of retinal diseases (Figure 4-2). Mutations in rhodopsin
and GCAP2 are known to cause ADRP; mutations in PDE6A, PDE6B,
PDE6G, CNGA1, and CNGB1 are responsible for ARRP; and mutations in
the α-subunit of transducin (GNAT1), arrestin-1 (SAG), GRK1, and NCKX1
are associated with various forms of CSNB. GC1 present in both rod and
cone outer segments are associated with LCA1 and CRD, and mutations in
GCAP1 are known to cause CRD and CD. Mutations in green cone opsin
(OPN1LW) and red cone opsin (OPN1MW) cause macular dystrophy and
colorblindness, whereas mutations in the cone-specific genes, CNGA3,
CNGB3, PDE6C, PDE6H, and GNAT2 are known to cause achromatopsia
and CD. Finally, mutations in RGS9 are associated with bradyopsia, a visual
disorder characterized by difficulties in adjusting to changes in light
intensities.
Visual Cycle
The visual or retinoid cycle is another key pathway in the visual response
(94,95). This process incorporates a number of retinoid binding proteins and
enzymes, which function in the conversion of all-trans retinal generated from
photoactivation of rhodopsin and cone opsin to 11-cis retinal for the
regeneration of photopigment (Figure 4-3). In the conventional visual cycle,
a significant fraction of the all-trans retinal derived from the photoactivation
of rhodopsin or cone opsin is first reduced to all-trans retinol by retinal
dehydrogenase 8 (RDH8) present on the cytoplasmic side of photoreceptor
disk membranes. All-trans retinol is transported to RPE cells by the
interphotoreceptor retinoid binding protein (IRBP), and with the aid of
cellular retinol binding protein 1 (RBP1 also known as CRBP1), it is
delivered to lecithin retinol acyltransferase (LRAT) for esterification and
hydrolysis prior to isomerization to 11-cis retinol by retinoid
isomerohydrolase RPE65. Retinal dehydrogenase (RDH5) subsequently
oxidizes 11-cis retinol to 11-cis retinal. With the aid of retinaldehyde binding
protein (RLBP1 also known as CRALBP) and IRBP, 11-cis retinal is
returned to the photoreceptor outer segment disk membranes where it
combines with opsin to regenerate rhodopsin.
FIGURE 4-3 Visual cycle showing the key retinoids and

enzymes involved in the resynthesis of 11-cis-retinal from
all-trans-retinal following photobleaching of rhodopsin.
All-trans-retinal is reduced to all-trans-retinol by RDH8
and transferred to RPE cells by IRBP. All-trans retinol
binds to retinal binding protein RBP1 (retinol binding
protein) and converted to retinylesters by lecithin retinol
acyltransferase LRAT, isomerized to 11-cis-retinol by
retinoid isomerohydrolase RPE65, and oxidized to 11-cis-
retinal by 11-cis retinol dehydrogenase RDH5. The 11-cis-
retinal is transferred back to outer segments with IRBP
where it recombines with opsin to regenerate rhodopsin.
ABCA4 in disk membranes transports N-retinylidene-
phosphatidylethanolamine from the lumen to the
cytoplasmic side of disk membranes insuring that all
excess 11-cis-retinal and all-trans-retinal are removed
from outer segments via the visual cycle. Retinol
dehydrogenase 12 localized in the inner segment reduces
retinal to retinol that has diffused from the outer segment
to the inner segment. Diseases associated with various
proteins: RP, retinitis pigmentosa; CSNB, congenital
stationary night blindness; CRD, cone–rod dystrophy;
fundus albipunctatus; LCA, Leber congenital amaurosis.
Some of the all-trans retinal (~30%) released from rhodopsin or cone opsin
after photoexcitation reversibly react with the phospholipid,
phosphatidylethanolamine (PE) to form the Schiff base adduct, N-
retinylidene-phosphatidylethanolamine (N-Ret-PE) (55,96,97). A significant
fraction (~20% to 30% of N-Ret-PE) is trapped on the lumen side of the disk
membrane. The function of the ATP-binding cassette transporter ABCA4 is
to flip N-Ret-PE from the luminal to the cytoplasmic side of the disk
membrane. There, N-Ret-PE dissociates into all-trans retinal and PE enabling
all-trans retinal to be reduced to all-trans retinol by RDH8 for re-entry into
the visual cycle. This transport function of ABCA4 insures that all of the all-
trans retinal generated from photoactivated rhodopsin and cone opsin is
effectively cleared from disk membranes preventing the buildup of toxic bis-
retinoid compounds. ABCA4 also insures that any excess 11-cis retinal that is
not required for the regeneration of rhodopsin or cone opsin is also removed
via the visual cycle (97).
Recent studies indicate that cone opsin can be regenerated using another
series of reactions that involve cone photoreceptors and Müller cells (98). In
this retinoid cycle, all-trans retinol obtained from the reduction of all-trans
retinal by RDH8 in disk membranes is delivered to Müller cells where it is
isomerized to 11-cis retinol and returned to cone photoreceptors. There it is
oxidized to 11-cis retinal by a dehydrogenase for regeneration of cone opsins.
Several other proteins have also been implicated in the retinoid cycle and
in the detoxification of retinal. Retinal G-protein–coupled receptor (RGR)
encoded by the RGR gene binds and photoisomerizes all-trans retinal to 11-
cis retinal in RPE and Müller cells and may contribute to the regeneration of
photopigments (99). Retinol dehydrogenase 12 (RDH12) encoded by the
RDH12 gene and localized to the inner segments of photoreceptors catalyzes
the reduction of both all-trans and 11-cis retinal to their corresponding retinol
derivative (100). RDH12 is generally thought to play a crucial role in
protecting photoreceptor inner segments from the toxic effect of retinal that
may escape from outer segments. RDH12 may also protect photoreceptors
against toxic aldehyde products resulting from lipid peroxidation.
Mutations in genes encoding proteins that function in the visual cycle and
retinoid processing are known to cause a variety of retinal diseases.
Autosomal recessive Stargardt macular degeneration (STGD1), CRD, and a
subset of ARRP are caused by mutations in ABCA4. LCA14 is caused by
genetic defects in LRAT. Mutations in RPE65 have been reported to cause
LCA2, ADRP, and ARRP, whereas mutations in RDH5 cause fundus
albipunctatus, a form of CSNB display whitish-yellow flecks upon fundus
examination, and mutations in IRBP, RGR, and RLBP1 have been reported to
cause ARRP. Finally, mutations in RDH12 have been linked to LCA13 and
ARRP. Interestingly, to date, disease linked to mutations in RDH8 have not
been reported despite its prominent role in the visual cycle.
Photoreceptor Outer Segment Structure and Renewal

The nonmotile cilium of retinal photoreceptor cells consists of the light-
sensitive outer segment compartment and a thin connecting cilium that joins
outer segment to the inner segment (101, 102, 103). In rod photoreceptors,
the outer segment contains a stack of over 800 closed disks surrounded by a
separate plasma membrane (Figure 4-4). In cones, the outer segment, often
displaying a tapered “cone” shape, consists of disk-like structures that are
contiguous with the plasma membrane. The flattened region of each disk is
packed with the photopigment rhodopsin in rod outer segments and cone
opsins in cone outer segments. This organization allows the photopigments to
be densely packed and oriented toward incoming light for optimal photon
capture. The planar disk surface also serves as a platform for many of the
biochemical reactions involved in photoexcitation and photorecovery. In
addition to rhodopsin, rod outer segment disks contain GRK1, arrestin-1,
transducin, PDE6, and the RGS9-Gβ5-R9AP complex directly involved in
the visual response and several other proteins including the progressive rod–
cone degeneration (PRCD) protein important in stabilizing the outer segment
disks and the ATP8A2-CDC50A, a complex that actively flips
phosphatidylserine and PE across membranes (104, 105, 106). The two
planar regions are circumscribed by a highly curved rim region to enclose an
intradiscal compartment within each disk. One or more incisures penetrate
from the rims toward the center of the rod outer segment disk. Such incisures
are not found in cone outer segment disk-like structures. The highly curved
rim region of disk membranes including the incisures contains a number of
transmembrane proteins important in outer segment structure and function
(Figure 4-4). These include GC1 and GC2, ABCA4, and the peripherin-
2:rom1 complex (86).
FIGURE 4-4 Schematic of the rod outer segment showing
key membrane proteins localized in the plasma membrane
and disk membranes of rod outer segment structure and
their association with inherited retinal diseases. Proteins
include rhodopsin, progressive rod–cone dystrophy
(PRCD) protein, cyclic nucleotide–gated channel subunits
CNGA1 and CNGB1, sodium–calcium–potassium
exchanger NCKX1, guanylate cyclase 1 (GC1); ATP-
binding cassette transporter ABCA4, peripherin-2; rom-1
PRPH2:ROM1 complex, prominin1; protocadherin 21
PCDH21 and include the key complexes BBSome and ITF
complex. Numerous other proteins involved in connecting
cilium and basal body and linked to nonsyndromic and
syndromic diseases (ciliopathies) are not shown.
The plasma membrane of rod outer segments has a different protein

composition than disk membranes (107). The CNG channel and the NCKX1
exchanger are exclusively localized to the plasma membrane where they form
a complex (108,109). Embigen, a highly glycosylated membrane protein, and
a number of less well characterized membrane proteins are also exclusively
localized to the plasma membrane. Rhodopsin is found in the plasma
membrane but at a lower concentration than in disk membranes (107,110).
Disks in rod outer segments are not free floating as originally thought,
but instead are tethered to each other and to the plasma membrane by thin
filamentous structures as visualized by electron microscopy (111). The
molecular components that comprise these filaments have not been well-
characterized although interaction between peripherin-2 along the rim region
of disks and the glutamic acid-rich protein (GARP) region of the CNGB1
channel subunit in the plasma membrane helps to stabilize the rod outer
segment structure (112). Free GARPs also have been proposed to mediate
disk–disk interactions, although evidence for this is lacking (102). Peripherin-
2 and its homologue rom1 are tetraspanin protein that associate to form
peripherin-2 homotetramers and peripherin-2:rom-1 heterotetramers (42).
These proteins further assemble into higher order oligomeric complexes that
appear to play an essential role in generating and modeling the highly curved
rims of rod and cone outer segments disks (113,114).
Photoreceptor outer segments are continually renewed every 10 days in
most vertebrate species (102). One-tenth of the outer segment is removed
each day from the distal end of the outer segment through a phagocytic
process orchestrated by the apical structures of the RPE cells. New disks are
formed at the proximal end of the outer segment. The process of disk
morphogenesis at the base of the rod outer segment is not well understood at
the molecular level, but at a cellular level, the classic model of Steinberg et
al. is supported by more recent electron microscopic imaging studies
(115,116). In this model, new disks form through evagination of the ciliary
plasma membrane followed by an extension of the rim region from the
axoneme to surround disk evaginations. Subsequently, the disks separate
from the plasma membrane through an undefined fission process. A similar
process is suggested for the generation of disk-like structures of cone
photoreceptors although the separation of disks from the plasma membrane is
incomplete (117).
Disk morphogenesis is critically dependent on peripherin-2, a tetraspanin
protein that assembles into tetramers and higher order oligomers with itself
and with it homologous tetraspanin protein rom1 (63,102). Mice deficient in
peripherin-2 (homozygous rds mice) develop connecting cilia but fail to
elaborate these structures into rod outer segments (60). Even heterozygous
rds mice with reduced levels of peripherin-2 display highly disorganized
outer segments. Recent studies have shown that peripherin-2 plays a role in
the initial stage of disk morphogenesis by preventing the release of ectosomes
from the photoreceptor connecting cilium and redirecting proteins to
elaboration of outer segment structures (118). Rom1 is not required for disk
morphogenesis since rom1 knockout mice form outer segment disks that are
only moderately disorganized and smaller in size (119). Instead, rom-1
appears to regulate the size and organization of disks through its association
with peripherin-2. Rom1 may play a more important role in cone outer
segment morphogenesis, although further studies are needed (120).
Rhodopsin, the major protein in rod disk membranes, also plays a crucial role
in disk morphogenesis since homozygous rhodopsin knockout mice fail to
generate disk membranes (67).
Prominin1, a membrane glycoprotein localized to protrusions in many
cells, appears to also contribute to disk morphogenesis (121).
Immunolabeling studies indicate that prominin1 in rod and cone outer
segments is localized at the outer edge of disk evaginations present at the
base of the outer segments where it interacts with protocadherin 21
(PCDH21) (122, 123, 124). Mice deficient in prominin-1 undergo
progressive photoreceptor degeneration, although they do elaborate outer
segments that are highly disorganized. Other outer segment proteins
examined to date are not essential for disk morphogenesis but likely play a
direct or indirect role in generating highly organized, stable outer segment
disks.
The detailed molecular mechanism underlying phagocytosis of aged
photoreceptor outer segments by RPE cells is not well understood. However,
studies of the RCS rat deficient in outer segment phagocytosis have shown
that proto-oncogene tyrosine-protein kinase MER encoded by the MERTK
gene plays an essential role in this process (64). Exposure of
phosphatidylserine at the tips of rod outer segments may also serve as a
signal for initiation of phagocytosis (125).
Various retinal degenerative diseases have been associated with
mutations in proteins implicated in photoreceptor outer segment
morphogenesis, structure, and renewal. Mutations in PRPH2 encoding
peripherin-2 are responsible for about 5% of cases of ADRP (34).
Interestingly, some mutations in PRPH2 have been reported to cause pattern
macular dystrophy (26). Disease-associated mutations in the ROM1 gene are
rare. However, a mutation in the ROM1 gene has been reported recently to
cause a late-onset pattern macular dystrophy (126). A digenic form of RP has
also been reported to involve mutations in ROM1 coupled to a mutation in
PRPH2 (15). In this case, a base-pair insertion that creates a frameshift early
in the ROM1 gene causes RP when coupled to a Leu185Pro mutation in
PRPH2. Mutations in PROM1 gene encoding prominin-1 have been
associated with autosomal dominant macular dystrophy (STGD4) and in
some cases recessive RP (124). Mutations in PCDH21 gene have also been
reported to cause progressive retinal degeneration (127).
Connecting Cilium
The connecting cilium plays a crucial role in transporting proteins and
metabolites between the inner segment and outer segment of photoreceptors.
This structure, equivalent to the transition zone in a nonphotoreceptor cilium,
contains a microtubule-based axoneme that extends from the basal body in
the inner segment far into the outer segment (Figure 4-4). The axoneme in
the photoreceptor is organized with nine microtubules in a circle with no
microtubules in the center known as a 9 + 0 arrangement. Within the
proximal inner segment and connecting cilium, the microtubules exist as nine
doublets. Further into the outer segment, however, the nine microtubules
appear as nine singlets. An intraflagellar transport complex IFT and a protein
complex known as the BBSome together with the motor protein kinesin (128)
transport proteins into and out of the outer segment (39,129, 130, 131).
Biochemical studies indicate that a wide range of membrane and soluble
proteins coordinate the trafficking of proteins through the connecting cilium
(128). Many of these proteins are also found in the cilium of
nonphotoreceptor cells, whereas others appear to be selectively expressed in
photoreceptors.
Mutations in genes encoding a large variety of proteins that play a role in
basal body and ciliary structure and transport have been linked to
nonsyndromic and syndromic diseases. These diseases are collectively known
as ciliopathies and in some classifications include retinal diseases caused by
mutations in outer segment proteins (2). Nonsyndromic diseases associated
with proteins involved in connecting cilium structure and function including
RP, CRDs, and LCA. For example, RP1, a protein associated with
microtubules and important in maintaining the stability of rod outer segment
disks, has been linked to autosomal recessive RP (132), whereas mutations in
CEP290, a large protein that links the ciliary membrane to microtubules of
the axoneme, are known to cause LCA10. Syndromic diseases include Usher
syndrome (see also Chapter 33) involving photoreceptor degeneration and a
loss in hearing; BBS (see also Chapter 32) associated with a loss in vision
due to photoreceptor degeneration and postaxial polydactyly, central obesity,
mental retardation, hypogonadism, and renal dysfunction; Senior-Løken
syndrome (SLS) resulting in early-onset photoreceptor degeneration and
juvenile nephronophthisis; and Joubert syndrome (JBS), a neurologic
disorder characterized by hypotonia, ataxia, abnormal breathing,
developmental delay, and abnormal ocular movements together with
photoreceptor degeneration, among others (see also e-Chapter 2). A partial
list of proteins implicated in basal body and connecting cilium structure and
transport function and associated with nonsyndromic and syndromic diseases
include CEP290, lebercilin, RPGRIP1, RPGR, RP1, USH2A, USH2C,
USH2D, ITF27, ITF38, ITF81, ITF140, ITF144, ITF172, and BBS1-7.
Although considerable progress has been made in identifying proteins
encoded by genes associated with connecting cilium diseases, the
biochemical properties of these proteins have not been thoroughly
investigated and their role in cilium structure and transport is not well
understood.
Metabolism and Protein Trafficking

Genetic screens have identified a number of defective genes encoding
proteins that play crucial roles in protein modifications, lipid and nucleotide
biosynthesis and metabolism, chaperones involved in protein trafficking,
transcription and pre-RNA splicing, molecular scaffolds, and retinal
development. In the case of lipid biosynthesis, mutations in ELOV4 gene
encoding an enzyme that functions in the elongation of very long chain fatty
acids ELOV4 cause an autosomal dominant Stargardt-like macular
degeneration (STGD3) (133). This supports the view that very long chain
fatty acids found in photoreceptor outer segments are important in
maintaining the survival of photoreceptor cells (134). Mutations in
dehydrodolichyl diphosphate synthetase (DHDDS), an enzyme involved in
the synthesis of dolichol monophosphate, are responsible for RP59 (135).
Mutations in the inosine monophosphate dehydrogenase 1 gene IMPDH1
encoding an enzyme, which catalyzes the rate-limiting step in de novo
guanine synthesis, cause autosomal dominant LCA11 and RP10 (136).
Transcription factors important in retinal development and differentiation
also have been implicated in retinal degenerative diseases. Notably,
mutations in cone–rod homeobox protein CRX are responsible for LCA7 and
CRD (137); mutations in the neural retina-specific leucine zipper protein
NRL important in rod cell differentiation cause autosomal dominant and
recessive RP (138); and mutations in NR2E3, another member of the nuclear
receptor transcription family, cause a loss in rod photoreceptors and an
increased number of S-cone photoreceptors associated with recessive-
enhanced S-cone syndrome (139) and RP (140). In addition to these and other
transcription factors, mutations in genes involved in pre-RNA splicing cause
ADRP. These include ubiquitous core snRNP proteins (Prpf3, Prpf4, Prpf6,
Prpf8, and Prpf31) and two additional splicing factors RP9 and DHX38.
PRPF31 mutations are the most prevalent of the splicing factor mutations and
account for about 2.5% of ADRP cases (141).
Genetic defects in proteins involved in vesicle trafficking also contribute
to a variety of retinal degenerative diseases. Truncation mutations in RD3, a
small 24 kDa protein, cause severe retinal degeneration in the rd3 mouse,
CRD in the rcd2 collie, and patients with LCA12 (75,142).
Immunocytochemical and biochemical studies have shown that RD3 plays a
crucial role in the stable expression and trafficking of guanylate cyclases to
rod and cone photoreceptor outer segment (66). Genetic defects in
arylhydrocarbon-interacting receptor protein-like 1 (AIPL1) are responsible
for LCA4 (143). Biochemical studies indicate that AIPL1 functions as a
chaperone protein for the PDE complex via binding to farnesyl lipid moiety
of PDE (144). Other proteins that are involved in protein trafficking and have
been linked to retinal degenerative diseases include TULP1, RP2, and ARL6.
Synaptic Region and Extracellular Matrix

In addition to carrying out phototransduction, rod and cone cells are directly
involved in the transmission of electrical signals to secondary neurons of the
retina and in particular bipolar and horizontal cells. This takes place at the
ribbon synapses of rod and cone photoreceptors and involves the release of
the chemical transmitter glutamate stored in synaptic vesicles. Although the
structure and function of the ribbon synapses are not completely understood
at a molecular level, a number of proteins that localize within this region of
rod and/or cone photoreceptors have been identified. Their importance in rod
and cone function and survival is highlighted by the finding that mutations in
a number of proteins have been associated with retinal diseases. For example,
mutations in UNC119, a protein homologous to C. elegans neuroprotein
unc119 and localized to the synaptic ribbons of rod and cone photoreceptor,
have been linked to CRD (145). Mutations in the CACNA1F gene encoding
the L-type voltage-gated calcium channel alpha-1 subunit have been shown
to cause CSNB and X-linked progressive CRD (146, 147, 148), and
mutations in RIMS1 encoding RIMS1 (also known as RIM), a protein
implicated in regulating synaptic membrane exocytosis and voltage-gated
calcium channels, are responsible for a dominant form of CRD (149,150).
Although most disease-associated proteins play key roles in biochemical
processes that occur within photoreceptors and RPE cells, a number of
proteins synthesized by these cells are secreted into the extracellular
environment. Retinoschisin (also known as RS1), a multi-subunit protein
predominantly expressed and secreted from photoreceptor cells, has been
implicated in maintaining the structural organization and synaptic structure of
photoreceptors (18,86,151,152). Over 130 mutations in the XLRS1 gene
encoding retinoschisin cause X-linked retinoschisis, an early-onset retinal
degenerative disease associated with a splitting of the retina, a marked
reduction in the b-wave of ERGs, and progressive loss of rod and cone
photoreceptors (153,154) (see also Chapter 34). Another prominent disease
associated with an extracellular protein is Sorsby dystrophy, an autosomal
dominant macular dystrophy caused by mutations in the TIMP3 gene
encoding the tissue inhibitor of metalloproteinase 3 TIMP3 that is involved in
retinal extracellular matrix remodelling of the Bruch membrane (19).
CONCLUSIONS AND PERSPECTIVE

Extensive clinical and genetic studies have confirmed the extreme
heterogeneous nature of inherited retinal diseases. These disorders range from
extensive or complete loss in vision at or just after birth as in the case of LCA
to more limited and in some cases stationary loss in vision as found for
various forms of CSNB and late-onset diseases. Approximately, 300 different
genes have been implicated in various inherited retinal diseases with most
being expressed in photoreceptors or RPE cells and encoding proteins
essential for the function and/or survival of these cells. The heterogeneity of
inherited retinal diseases further extends to an extremely diverse set of
biochemical pathways and cellular processes, which are compromised in
these disorders. Molecular studies have shown that mutations in most
proteins involved in phototransduction, the visual cycle, and outer segment
structure and renewal cause nonsyndromic retinal diseases. Furthermore, the
underlying cause of many other nonsyndromic and syndromic diseases
involves mutations in a diverse set of proteins required for connecting cilium
and basal body biogenesis, structure, and function. Additionally, mutations in
proteins that function in other key biochemical pathways including lipid and
nucleotide metabolism, transcription, vesicle trafficking, ion transport, and
synaptic structure and function are responsible for various retinal diseases.
Although significant progress has been made in understanding how
disease-causing mutations affect phototransduction and the visual cycle
through the use of animal and cellular models together with an array of
physiologic, biochemical, and imaging tools, we still know relatively little
about how specific proteins function in other key photoreceptor processes
including photoreceptor disk morphogenesis, outer segment phagocytosis,
connecting cilium structure and transport function, vesicle transport,
photoreceptor development and differentiation, and synaptic structure,
function, and regulation. These and other areas require more detailed studies
to elucidate pathogenic mechanisms that contribute to various types of
inherited retinal diseases and to design and develop novel therapeutic
treatments for these diseases.
ACKNOWLEDGMENTS
This work was supported in part by grants from the National Institutes of
Health (EY002422) and the Canadian Institutes of Health Research (PJT
148649).
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5
Extracellular Matrix Regulation of
Vascular Development in the Retina
Saptarshi Biswas, William J. Brunken and Dale D. Hunter
INTRODUCTION
The retina, as the rest of the central nervous system (CNS), vascularizes
primarily by angiogenesis (reviewed in (1,2)). Mechanisms that regulate
angiogenesis remain active throughout life. Disruptions in mechanisms
regulating retinal angiogenesis are the leading causes of adult blindness in the
developed world (reviewed in (1)). For example, diabetic retinopathy is the
main cause of blindness within the working-age population in industrialized
nations (reviewed in (3)). Also, vessel tortuosity and abnormal arterial
collateral formation have been reported in retinal vessel occlusion (4).
Moreover, abnormal arterial and venous patterning, characterized primarily
by direct arteriovenous shunt formation, has been reported in several diseases
with ocular manifestation (4, 5, 6). In premature infants, retinopathy of
prematurity (ROP) is a major cause for blindness (reviewed in (7)).
Therefore, a detailed understanding of the mechanisms that govern retinal
angiogenesis is important for the development of potential treatments.
CNS blood vessels, including those in the retina, acquire a unique
structure called the neurovascular unit (reviewed in (8,9)). The neurovascular
unit, a shared structural component among all mammals, is comprised of
different cell types including endothelial cells, mural cells (pericytes and
vascular smooth muscle cells), astrocytes, neurons and microglia, as well as
the vascular basement membrane (BM). Retinal vasculature sprouts from an
existing artery in the optic nerve head in response to the metabolic demand of
newly active retinal neurons (10, 11, 12), which active glial cells that act to
promote vascular growth and development. Initial events in vascular
development are initiated by retinal ganglion cells, which induce astrocyte
migration over the retinal surface forming a template for the expansion of the
retinal vascular tree (13, 14, 15). Glial cells, astrocytes, and Müller cells
express and secrete vascular endothelial growth factor (VEGF), which drives
endothelial proliferation (15).
Microglia, the resident macrophages of the CNS, regulate branching of
the newly formed vascular network by facilitating anastomosis of emerging
vascular sprouts and mediating turning of vascular sprouts in the deeper
layers of the retina (16,17). Moreover, depending on their activation state,
microglia regulate endothelial cell proliferation. While resting microglia
produce antiangiogenic factors that inhibit endothelial cell proliferation,
activated microglia secrete proangiogenic factors that induce endothelial cell
proliferation (18). It was also known that microglia interact with the
astrocyte-derived components of the vascular BM (19). However, whether
BM components regulate microglial recruitment and activation around the
growing vasculature, the physiologic consequence of that regulation has not
been extensively studied.
Our work focused on the mouse model has shown that γ3-containing
laminins are negative regulators of vascular branching in the mouse. Vascular
branch point-associated microglial density is specifically increased at the
nascent plexus of the Lamc3−/− retina. Consequently, vascular branching
density is also increased in the Lamc3−/− retina, suggesting an increased
occurrence of microglia-mediated vascular anastomotic events. We further
showed that microglia interact with astrocyte-derived laminins via integrin
β1-mediated signaling. Laminins containing γ3-chain also negatively regulate
microglial activation. Microglia become hyperactivated in the Lamc3−/−
retina or when tested in vitro with Lamc3−/− astrocyte-derived matrix. In
addition, mural cells, such as vascular smooth muscle cells and pericytes,
help stabilizing maturing blood vessels (reviewed in (20)).
Several studies demonstrated that BM molecules, especially laminins,
influence several aspects of retinal angiogenesis either by direct endothelial
cell–BM interactions (21) or by mediating cross-talk between different cell
types (22, 23, 24). Moreover, all these cell types as well as the vascular BM
play critical roles in maintaining the barrier property of CNS vessels
(reviewed in (25)). BM disorganization and remodeling occur in several
pathologic conditions with an ocular manifestation such as diabetes (26,27)
and ischemia (reviewed in (25)). Indeed, the inhibition of matrix
metalloproteinase-mediated degradation of the extracellular matrix (ECM)
has been reported to reduce infarct size in the CNS following ischemia (28).
Here, we summarize the role of the BM during retinal angiogenesis, that is,
retinal vascular development, in the context of cell–cell interactions, as well
as direct signaling between the BM and endothelial cells. We focus mainly on
laminins, a major component of the BM (reviewed in (29)).
ANATOMY OF THE NEUROVASCULAR

UNIT
In the CNS, the vascular compartment (endothelial cells and mural cells) is
tightly coupled with the neural compartment (glial cells, microglia, and
neurons) in a neurovascular unit (Figure 5-1; reviewed in (8,9)). CNS
vasculature, including those in the retina, grows in response to the metabolic
demand of neurons; the neuronal metabolic demand is transduced into
proangiogenic factors, largely VEGF, produced by retinal glial. For example,
the superficial vascular tree in the retina forms in response to the metabolic
demand of ganglion cells (11) and the VEGF production by astrocytes, the
deep vascular plexus forms in response to the metabolic demand of
photoreceptors (10), and the intermediate vascular plexus forms in response
to the metabolic demands of interneurons, such as amacrine cells (12) with
the Müller cell as likely producer of VEGF.
FIGURE 5-1 Schematic of the anatomy of the
neurovascular unit. Endothelial cells surround the vascular
lumen. The vascular basement membrane (BM) surrounds
endothelial cells. The vascular BM is a composite
structure with components derived from both endothelial
cells and astrocytes. Astrocytes contact the vascular BM
with their endfeet. Pericytes are embedded in the vascular
BM and make direct contact with endothelial cells.
Activated microglia are in contact with the astrocyte-
derived components of the vascular BM. Astrocytes and
microglia also make contact with neurons.
Anatomically, endothelial cells are surrounded by the vascular basement

membrane (BM) in a CNS vessel. Pericytes are embedded in the vascular BM
and make direct contact with endothelial cells. Large arteries and arterioles
also have vascular smooth muscle cells surrounding them. On the neural side,
astrocytes surround the vascular BM with their endfeet (reviewed in (8,9)).
The vascular BM is a composite structure consisting of components derived
from both endothelial cells and astrocytes (30). Microglia, resident
macrophages of the CNS, interact with the astrocyte-derived layer of the
vascular BM (19,22). Astrocytes and microglia also maintain contact with
CNS neurons (reviewed in (9,25)). Integrity of this neurovascular unit is
critical for the proper maintenance of the barrier property, which is disrupted
in pathologic conditions such as ischemia (reviewed in (25,31)). Indeed, the
expression of the matrix degrading enzymes, matrix metalloproteinase-2 and
-9 (MMP-2 and MMP-9), are increased in the brain following ischemia (28),
and inhibiting MMP-9–mediated degradation of vascular BM reduces
cerebral infarct size and neuronal death (28,32).
MOUSE RETINA AS A MODEL TO

STUDY ANGIOGENESIS
The mouse retina is a useful model to study mechanisms of angiogenesis in
the CNS, because vascularization of the mouse retina is entirely postnatal,
and there are a large number of genetic variants available. After birth,
embryonic hyaloid vessels gradually regress as retinal vessels form
(17,33,34). There are three distinct, interconnected vascular beds in the
mouse retina: (1) the superficial vascular tree; (2) the deep vascular plexus
(DVP); and (3) the intermediate vascular plexus (IVP). The superficial
vasculature radially expands from the optic nerve head toward the retinal
periphery over the ganglion cell layer (GCL) and nerve fiber layer (NFL) and
follows the astrocyte template that migrates through the inner limiting
membrane (ILM) (Figure 5-2). Eventually, vascular sprouts from the
superficial vessels dive deep into the retina guided by Müller cell processes
(reviewed in (35)) and form the DVP in the outer plexiform layer (OPL)
between postnatal day (P)8-P12. Finally, the IVP forms in the inner
plexiform layer (IPL), completing vascularization of the mouse retina by P21
(36).
FIGURE 5-2 Time course of expansion of the retinal
vascular plexus. Drawings are representations of sections
through a representative vascularized mammalian retina.
Five stages of vascularization are depicted, with stages of
development for rat, cat, and mouse stated at the left. The
developing vascular tree is depicted in red. In these
animals, astrocytes migrate out of the optic nerve over the
inner limiting membrane (ILM) responding to cues
derived from ganglion cells. Angiogenesis proceeds are
endothelial cells sprout off the existing hyaloid artery. The
endothelial cells follow over the astrocytic template along
the ILM coming to lie superficial to the ganglion cell layer
(GCL). Later in development, in a central to peripheral
pattern, the superficial vessels branch driving through the
GCL and inner plexiform layer (IPL) ultimately
terminating in the outer plexiform layer (OPL). The deep
vascular bed is completely formed by postnatal day
(P)12.5; branches from the deep vascular bed arise and
give rise to the intermediate vascular bed that forms in the
IPL (not depicted here). The mouse retina becomes
completely vascularized by P21. In human and dog, the
cellular events may be different than these model systems,
but many of the same molecular regulatory pathways are
involved. Figure modified from Stone et al. (15).
The growing retinal superficial vascular tree may be subdivided into three
developmental zones: (1) the vascular front, where tip cell/stalk cell
specification takes place; (2) the nascent plexus, where newly formed vessels
branch and proliferate; and (3) the remodeling zone, where morphologically
distinct arterial and veins arise from the undifferentiated nascent plexus
(Figure 5-3; (37)). As the temporal sequence of angiogenesis is spatially laid
out over the retinal surface, the retinal superficial vascular tree proves to be
an ideal model to study all three aspects of developmental angiogenesis in the
CNS.
FIGURE 5-3 Distinct developmental regions in a growing
retinal superficial vascular tree. The lectin, IB4, binds to
sugar moieties that are expressed on endothelial cells. In
the mouse at P5, three different developmental regions
(vascular front, nascent plexus, and remodeling zone) are
present. Vascular sprouts align themselves toward the
angiogenic cue at the vascular front, where tip cell/stalk
cell specification takes place. Behind the vascular front,
newly formed vessels form an immature nascent plexus,
where most endothelial cell proliferation occurs. Further
behind, this nascent plexus undergoes extensive
remodeling to form morphologically distinct arteries,
veins, and the capillary network.
LAMININ COMPOSITION OF THE CNS

VASCULAR BASEMENT MEMBRANE
The basement membrane (BM) is a three-dimensional structure composed of
various extracellular matrix (ECM) molecules such as laminins, collagen IV,
perlecan, agrin, and nidogen (reviewed in (29)). BM molecules have been
reported to be critical regulators of cell adhesion, migration, proliferation,
and differentiation (reviewed in (29)). Of these molecules, laminins have
been shown to be especially important in regulating several aspects of retinal
vascular development (21, 22, 23, 24).
Laminins are heterotrimeric glycoproteins made of ◻-, ◻-, and ◻-chains
(Figure 5-4A). Five ◻-, three ◻-, and three ◻-chains have so far been
identified that combine into 16 known isoforms (38,39); a simplified numeric
naming system is used and designates the isoforms as laminin 111 or 521, for
the component chains. The coiled-coil domains of laminin chains are
essential to form the functional heterotrimer. The LN domains of laminin
chains are important for the self-polymerization of laminin molecules.
Finally, the LG domains of the laminin ◻-chain are critical for the receptor
binding (reviewed in (29)).
FIGURE 5-4 Structure of a laminin heterotrimer and

schematic of laminin-receptor interaction. A.Laminin
heterotrimers are composed of 1 ◻, 1 ◻, and 1 ◻◻chain.
There are 5 different ◻ chains, 3 ◻ chains, and 3 ◻ chains
that combine into 16 different heterotrimers (isoforms).
The coiled-coil domains are critical for the laminin
heterotrimer formation. The LN domains are critical for
the self-polymerization of laminin heterotrimers. The LG
domains are critical for laminin receptor binding. The LG
domain cluster 1 to 3 is separated from 4 to 5 by a short
hinge. B.Spatial expression patterns of different laminin
chains in the mature retinal superficial vascular tree. The
laminin ◻2, ◻4, ◻1, and ◻1 chains are uniformly present
throughout the mature retinal superficial vascular tree. In
contrast, the laminin ◻5 and ◻2 expression is higher in
the arterial BM than either venous or capillary BM.
Laminin ◻3 chain is absent in the mature arterial BM but
present in venous and capillary BMs. C.Laminins bind to
various cell membrane receptors, including integrins and
dystroglycan. Both integrins and dystroglycan are
heterodimers, containing ◻ and ◻ subunits. Both ◻ and ◻
subunits of integrins are transmembrane. In contrast,
heavily glycosylated ◻ subunit of dystroglycan is
extracellular and only the ◻◻subunit is transmembrane.
Laminin LG domain cluster 1-3 is thought to responsible
for integrin binding, whereas LG domains 4-5 are thought
to bind ◻-dystroglycan.
Using experimentally induced encephalitis models, Lydia Sorkin and her

colleagues showed that the vascular BM is composed of components secreted
by both endothelial cells and glial cells (Figure 1, (30)). Indeed they
demonstrated that endothelial cell-derived laminins and astrocyte-derived
laminins are distinct isoforms and have specific contributions to the blood–
brain barrier. The main laminin ◻-chains made by endothelial cells are ◻4
and ◻5, whereas the main laminin ◻-chains made by astrocytes are ◻2 (30)
and ◻5 (21). Additionally, vascular endothelial cells make laminin ◻◻-◻
and ◻◻-chains to form laminin-411 and 511, whereas astrocytes make
laminin ◻◻-◻ and ◻◻-chains to form laminin-211 (30) and possibly
laminin-511. Astrocytes and possibly endothelial cells also make laminin ◻2-
◻ (24,40,41) and ◻3-chains (22,24,40). However, the specific laminin
trimers containing ◻2-◻ and ◻3-chains that are present in the vascular BM
are yet to be identified; this is ongoing work in our laboratory. Unfortunately,
most current literature focuses on the role of ◻1-◻ and ◻1-containing
isoforms (laminin-211, -411, and -511) in CNS vascular development.
Although studies from our laboratory provided strong evidence of the critical
role of ◻2-◻ and ◻3-containing laminins in CNS angiogenesis, other groups
have largely ignored these isoforms so far. Thus, it is of critical importance to
study these isoform-mediated signaling in various aspects of CNS
angiogenesis in great detail.
One of the questions that has not been completely answered yet is how
the expressions of different laminins in the vascular BM are regulated.
Previous studies suggested that the expression of different laminin chains in
the vascular BM might be coordinately regulated. For instance, genetic
deletion of the laminin ◻4-chain led to decreased expression of laminin ◻1-
chain in the vascular BM of the skeletal muscle (42). Genetic deletion of the
astrocytic laminin ◻1-chain led to decreased expressions of laminin ◻1- and
◻2-chains in the vascular BM of the brain (43). Finally, genetic deletion of
the laminin ◻2-chain led to decreased expressions of laminin ◻5- and ◻1-
chains in the vascular BM of the brain (44). However, most of these studies
examined the expression laminin ◻1- and ◻1-chains in their knock-out
mouse models, but very little is known about the coordinated regulation of
laminin ◻2- and ◻3-chain expressions in the vascular BM. Further studies
are required to thoroughly examine the coordinated expression of different
laminin chains in the CNS vascular BM, including those in the retina.
DEVELOPMENTAL REGULATION OF
LAMININ EXPRESSION PATTERNS IN
THE RETINAL VASCULAR BM
Very little is known about the expression patterns of different laminin chains
in the retinal vascular BM and how their expressions are regulated
developmentally. Previous studies suggested that laminin expressions in
retinal vessels are spatially regulated (Figure 5-4B). For example, laminin
◻◻-chain is present throughout the retinal vasculature with higher
expression at the growing vascular front. In contrast, laminin ◻◻-chain is
most prominently present in the nascent plexus immediately behind the
vascular front (21). It has also been reported that there is a patchy distribution
of the laminin ◻◻-chain in the venules in the brain ((45); reviewed in (46)).
In our preliminary results, we have found a more prominent expression of the
laminin ◻◻-chain in arteries than veins in the P10 retina (unpublished
results). The developmental expression pattern of laminin ◻1-chain in the
retinal vasculature has not been extensively studied. In contrast, laminin ◻2-
chain has been shown to be present throughout the retinal vascular tree with
higher expression in the arterial BM than the venous counterpart (24). The
laminin ◻◻-chain is expressed in the basement membrane throughout the
vasculature in the adult retina (40), whereas the laminin ◻3-chain expression
is restricted to the venous and capillary BMs of the mature retinal vasculature
(24,40).
Along with spatial expression differences, there seems to be also a
temporal regulation of laminin expression in the CNS vascular BM. For
example, whereas the laminin ◻◻-chain is expressed in the CNS vascular
BM in all stages of development, the laminin ◻◻-chain expression is
postnatal (reviewed in (46)). However, whether there is a specific temporal
regulation of laminin ◻4 and ◻5 expression patterns in the retina has not
been studied. In a growing retinal vascular tree, laminin ◻3-chain expression
is excluded from tip cells at the vascular front but present around stalk cells.
At the nascent plexus, ◻3-containing laminins are present at vascular branch
points (24). In the mature retinal vascular tree, laminin ◻3-chain expression
is virtually absent in the arterial BM but present in venous and capillary BMs
of a mature retinal vascular tree (24,40). These complex spatial and temporal
expression patterns of various laminins in the vascular BM suggest that
specific laminin isoforms play critical role in different aspects of vascular
development and stabilization in specific regions of the vascular tree.
LAMININ RECEPTORS IN THE CNS

VASCULATURE
Laminin binding and signaling is mediated by various transmembrane
receptors, including sulfated glycolipids, Lutheran glycoproteins,
dystroglycan, and integrins (reviewed in (29)). Among these receptors, the
role of dystroglycan and integrins has been studied most extensively in the
context of vascular development in the CNS.
Both dystroglycan and integrins are heterodimers containing ◻- and ◻-
subunits. The heavily glycosylated ◻-subunit of dystroglycan, the ligand
binding subunit, is extracellular and the ◻-subunit is transmembrane,
whereas both ◻- and ◻-subunits of integrins are transmembrane (Figure 5-
4C). A single gene encodes both ◻- and ◻-subunits of dystroglycan
(reviewed in (47)), whereas different genes encode the ◻- and ◻-subunits of
integrins (48). Moreover, dystroglycan and integrins are thought to bind
different regions of laminin LG domains. Integrins are thought to bind the
laminin LG domain cluster 1 to 3, whereas ◻-dystroglycan binds laminin LG
domains 4 to 5 (reviewed in (29)). The extensive glycosylation of ◻-
dystroglycan is critical for laminin binding (reviewed in (47)). Indeed,
genetic deletion of the POMGnT1 gene (encodes for protein O-linked-
mannose beta-1,2-N-acetylglucosaminyltransferase 1) results in defective ◻-
dystroglycan glycosylation and perturbed laminin-binding ability (49).
In the neurovascular unit, both endothelial cells and astrocytes express
dystroglycan (reviewed in (25)). In vertebrates, there are 18 ◻- and 8 ◻-
integrin subunits that can form 24 different heterodimers that vary in their
ligand-binding properties and tissue distributions. Of these, integrin ◻3◻1,
◻6◻1, ◻7◻1, and ◻6◻4 are known to bind laminins (reviewed in (50)). Of
these integrins, endothelial cells and astrocytes express integrins ◻3◻1,
◻6◻1, and ◻◻6◻4 (reviewed in (25)), whereas microglia express integrin
◻6◻1 (19). On the other hand, neurons have been reported to express all the
laminin-binding integrins (51). Interestingly, not all laminins bind integrins.
For example, laminins containing the ◻3-chain do not bind integrins due to a
conformational change in the C-terminal LG domain of laminin (52).
LAMININS REGULATE VASCULAR

EXPANSION
The retinal superficial vasculature radially expands from the optic nerve head
(ONH) following the astrocyte template. Astrocytes are one of the major
sources of vascular endothelial growth factor A (VEGFA), which guides
vascular development in the retina (14,53). Astrocytes enter the retina
through the ONH around embryonic day 17 to 18 (E17 to 18) (54,55) and
centrifugally migrate across the retinal surface (13,56). Studies from our
laboratory have demonstrated that astrocytes migrate through the laminin-rich
ILM in the retina (24).
Most studies that examined the role of laminins on retinal vascular
development focused on the vascular expansion. A recessive point mutation
of the laminin ◻1-chain (Lama1nmf223) disrupted astrocyte patterning in the
retina with some astrocytes growing into the vitreous. Consequently, vascular
growth was also disrupted in Lama1nmf223 retinae with the persistence of
embryonic hyaloid vessels and abnormal retinal vessel growth into the
vitreous (23). Genetic deletion of the laminin ◻2-chain (Lamb2−/−) also
severely delayed astrocyte migration and patterning in the retina, thereby
resulting in retarded vascular growth and persistent hyaloid vessels (Figure
5-5; (24)). On the other hand, genetic deletion of the laminin ◻3-chain only
moderately delayed astrocyte migration into the retina up to P3 and resulted
in delayed vascular growth up to P5 (24). Further supporting the concept that
laminins regulate astrocyte migration and retinal vessel growth, recent studies
have shown that disrupting cellular receptors for laminins produces similar
defects. In the POMGnT1−/− retina where dystroglycan's ligand-binding
ability is genetically disrupted, astrocytes and blood vessels showed
abnormal patterning (57). Moreover, astrocyte migration into the retina is
dependent on integrin ◻1-mediated signaling (24). These observations
suggest that the interaction between laminins in the inner limiting membrane
(ILM) and laminin receptors in the astrocyte is a critical regulatory element
for proper astrocyte migration and patterning in the retina, which in turn
influence retinal vascular expansion.
FIGURE 5-5 Deletion of laminins retards vascular
growth. Vascular growth was assayed by CD31 expression
at P5 and P15 in the indicated genotypes. Growth is
markedly reduced in all laminin nulls at P5 and recovers in
the Lamc3−/− animals, but not in other genotypes. In the
Lamb2−/− animals, persistent hyaloid vessels are seen at
P5 by P15 abnormally patterned vascular tufts have
emerged from the vessels. The phenotype of this animal
resembles ROP in many respects. Abnormal sprouting and
fusion along the vascular front in the Lamc3−/− animals is
also seen, this is the result of abnormal microglial
activation by mutant astrocyte-derived microglial (see
text). Images from Gnanaguru et al. (24).
LAMININS REGULATE TIP

CELL/STALK CELL SPECIFICATION
At the growing vascular front, blood vessels respond to the glia-derived
VEGFA by directed filopodia extension by leading endothelial tip cells. The
following endothelial stalk cells proliferate and stabilize the newly formed
vascular network (14). The Dll4/Notch signaling has been shown to play a
critical role in tip cell/stalk cell specification (58,59). The canonical model
postulates that VEGFA-mediated signaling induces Dll4 expression in tip
cells, which subsequently activates the Notch pathway in stalk cells (58,59).
However, in vitro studies have demonstrated that laminin-111 and
laminin-411 could also directly induce endothelial Dll4 expression via
integrin-mediated signaling (21,60). Genetic deletion of the laminin ◻4-chain
(Lama4−/−) down-regulated Dll4 expression in endothelial tip cells, leading
to an excessive number of tip cells (21) and phenocopying the effect of Dll4
deletion on retinal vasculature (58,59,61). Endothelial cell-specific deletion
of integrin ◻1 also led to a similar phenotype (21). These studies suggest that
the direct signaling between basement membrane laminins and endothelial tip
cells is critical for tip cell/stalk cell specification at the vascular front in an
isoform-specific manner. In contrast, genetic deletion of the laminin ◻3-
chain (Lamc3−/−) led to a decrease in tip cell number (24). The exact
mechanism by which ◻3-containing laminins regulate tip cell number is not
understood, and further studies are required to elucidate laminin ◻3-mediated
signaling in the context of tip cell/stalk cell specification.

BRANCHING AND ENDOTHELIAL
CELL PROLIFERATION
Immediately behind the growing vascular front, newly formed vessels form
an undifferentiated nascent plexus. The nascent plexus forms by vascular
sprouts anastomosing with each other (16). Most of the endothelial cell
proliferation takes place in the nascent plexus (37). It has been reported that
genetic deletion of the laminin ◻4- or ◻3-chain led to increased vascular
branching density in the nascent plexus (21,22,24). We have shown that
laminin isoforms containing the ◻3 and ◻2 chains modulate nascent plexus
patterning (22).
Microglia in the CNS have been shown to facilitate vascular anastomosis
and thereby positively modulate vascular branching density (16). Microglia
are recruited and localized at the sites of vascular anastomosis in the retina
(our unpublished data). Microglial density at vascular branch points is
increased in the Lamc3−/− nascent plexus, which suggests an increased
occurrence of vascular anastomotic events in these retinae, leading to the
formation of a hyperbranched nascent plexus (22,24).
Recent studies have also shown that BM laminins regulate endothelial
cell proliferation. Genetic deletion of the laminin ◻4- or ◻3-chain led to
increased endothelial cell proliferation in the nascent plexus (21,22). On the
other hand, genetic deletion of the laminin ◻2-chain led to decreased
endothelial cell proliferation in the nascent plexus (22). However, how BM
laminins regulate endothelial cell proliferation is not completely understood.
Microglia have been implicated in regulating endothelial cell
proliferation. Depending on whether they are in their resting or activated
state, microglia secrete anti- or proangiogenic factors, respectively (18). Our
results demonstrate that activated microglia are in close association with
mature astrocytes in the retina (22). Moreover, microglial interaction with
astrocyte-derived laminins in the vascular BM regulates their activation via
integrin ◻1-mediated signaling (22). Activated microglial density is
significantly increased and microglial TGF-◻1 expression is down-regulated
around the Lamc3−/− nascent plexus, consistent with increased endothelial
cell proliferation in these retinae (22). In contrast, microglial TGF-◻1
expression is up-regulated in the Lamb2−/− nascent plexus, consistent with
decreased endothelial cell proliferation in these retinae (22). These results
suggest that BM laminins regulate vascular branching and endothelial cell
proliferation in an isoform-specific fashion.

REMODELING AND ARTERIOVENOUS
MORPHOGENESIS
As the retinal superficial vasculature develops, the newly formed vessels
undergo extensive remodeling to form arteries, veins, and the capillary
network. Arteriogenesis is comprised of a series of developmental steps that
include adoption of arterial fate by endothelial cells, mural cell recruitment to
the arterial wall, arterial elongation, and the establishment of arterial
branching pattern (reviewed in (62)). Abnormal arterial morphogenesis leads
to altered blood flow and defective tissue perfusion (63). In the retina, all
endothelial cells in the superficial vasculature initially express venous
markers. Distinct arterial morphology and molecular signature is not apparent
until P3-4 in the mouse (55). However, how this shift from venous to arterial
signature occurs during retinal vascular development is poorly understood.
Dll4/Notch signaling has been reported to be a critical regulator of arterial
specification and morphogenesis. Conventional model postulates that VEGF-
mediated signaling induces Dll4/Notch signaling pathway in arterial
endothelial cells (reviewed in (64,65)). However, retinal arteries still form
despite pharmacologic or genetic disruption of VEGF/VEGFR2 pathway
(61,66). These observations suggest that VEGF-mediated signaling is not the
only regulator of arterial morphogenesis in the retina.
The specific spatial and temporal regulation of the expressions of various
laminin chains in the arterial compartment suggests that laminin-mediated
signaling may play a role in arterial development in the retina. Until recently,
very little was known about the role of specific laminin-mediated signaling in
arterial specification and morphogenesis. We have shown that laminins,
especially ◻3-containing isoforms, regulate arteriogenesis in the retina by
modulating arterial Dll4/Notch signaling pathway (67).

INTEGRITY AND BARRIER FUNCTION
Blood vessels in the CNS acquire a unique barrier property called the blood–
brain barrier (BBB) and blood–retinal barrier (BRB), which separate the
systemic circulation from the neural parenchyma. Laminins in the vascular
BM and laminin receptors in the neurovascular cells have been shown to be
critical to maintain BBB integrity (reviewed in (25)). One group has recently
demonstrated that the genetic deletion of the laminin ◻2-chain led to
disrupted vascular mural cell coverage, astrocyte polarization as well as
perturbed endothelial junctional protein expression, leading to vascular
leakage (44). Another group has demonstrated that in a model of
experimental autoimmune encephalomyelitis (EAE), T-cell infiltration occurs
around the vascular BM that lacked laminin ◻5-chain (30). The same group
further demonstrated that ◻5-containing laminins directly inhibit integrin
◻6◻◻-mediated extravasation of T cells (45). Another study demonstrated
that the genetic deletion of the laminin ◻1-chain in astrocytes led to vascular
smooth muscle cell fragmentation, disassembly of the vascular wall, and
hemorrhagic stroke (43). The same group further demonstrated that the loss
of astrocytic laminin ◻1 led to pericyte differentiation from a resting stage to
a contractile stage, disruption of astrocyte endfeet polarity, and perturbed
endothelial tight junction protein expression (68). Although most of these
studies focused on the role of laminins in BBB rather than BRB, a similar
relationship is likely to exist in the retina as well.
Interestingly, all these studies attributed these phenotypes to the absence
of either laminin-211 or -511. However, our laboratory has recently
demonstrated that the genetic deletion of laminin ◻2-chain also led to
vascular leakage in the retina (24). These observations suggest that laminin
◻2-containing trimers (perhaps laminin-221 and -521) are also important for
maintaining the barrier property in the retinal vasculature. It will be of great
importance to identify specific laminin trimers in different regions of the
vascular tree, such as artery, vein, and capillary, and their specific roles in
maintaining the barrier property of CNS vessels.
SUMMARY
Angiogenesis in the retina is a complex developmental process, the
mechanisms that regulate sprouting, proliferation, remodeling, and mural
stabilization remain active throughout life to respond to metabolic needs.
These processes can lead to neovascular diseases as the result of
dysregulation at any step in the process. Our laboratory, together with several
others, has shown that particular isoforms of laminins play critical roles in all
the processes of retinal angiogenesis (Figure 5-6). Laminins in the inner
limiting membrane guide astrocyte migration over the retinal surface
providing a template for endothelial migration and tube formation. Laminins
play key roles in the interactions between endothelial cells and microglial
cells regulating branching, endothelial proliferation, and turning into the
deeper layers of the retina. Laminins also play key roles in the remodeling
events as the nascent plexus is specified into arterial and venous sides.
Laminins also play roles in recruiting and stabilizing pericytes to the vascular
wall. Finally, laminins play critical roles in the blood–retinal and blood–brain
barrier.
FIGURE 5-6 Laminins play central role in retinal

angiogenesis. Retinal angiogenesis proceeds in a stepwise
fashion from the migration of astrocytes over the retinal
surface to the stabilization of the vascular wall and
formation of a stable neovascular unit (outer circle).
Laminins impact each of these steps, providing cues for
migration of astrocytes; tip cell mediated branching and
stalk cell proliferation; vascular remodeling; pericyte
recruitment; and finally vascular stability.
Future work is needed to further understand how neuronal activity and

vascular development are coupled. In particular, what are the mechanisms
regulating the spatial and temporal expression of laminins in various vascular
compartments, that is, venous, arterial, and capillary? Also of interest is
which of the mural or glial cells contribute to the vascular basement
membrane and whether glial-derived and endothelial-derived basement
membrane compartments are isolated from each other or they miscible.
Further work also needs to refine the signaling pathways involved in laminin
regulation of angiogenesis, but to date these pathways appear to involve
integrins, dystroglycan, Notch, and Wnt pathways. Finally, therapeutics
targeting laminin signaling pathways are likely to be important in for
controlling abnormal neovascularization that is not responsive to anti-VEGF
therapies.
ACKNOWLEDGMENTS
The work presented here was funded in part by the NEI R01EY12676 (WJB);
Research to Prevent Blindness, Inc. Unrestricted Grant to the Department of
Ophthalmology and Visual Sciences; Research Development funds from
Upstate Medical University (WJB). The authors also acknowledge the input
and support of fellow members of the Vision Sciences community at Upstate
Medical University, including Francesca Pignoni, Andrea Viczian, and
Huaiyu Hu among others.
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s
6
Visual Anomalies Associated With
Reduced Retinal Pigmentation
Donnell J. Creel
BACKGROUND
From primitive nervous systems, such as the fruit fly Drosophila, through
vertebrates to primates, each side of the nervous system often receives both
ipsilateral and contralateral visual fibers. During stages of development, most
vertebrates have some ipsilaterally projecting optic ganglion fibers. In birds
and fish, these are often transient pathways. Some species of birds have
ipsilateral optic fibers during embryonic periods that do not survive into
adulthood. Most adult birds have few to no ipsilateral projections. The
organization of ipsilateral retinal ganglion cell (RGC) projections in most
vertebrates other than mammals can be described as haphazard at best. In
some vertebrates, ipsilateral fibers re-cross back to the contralateral fiber
tract.
Even closely related vertebrate species often differ dramatically in their
ipsilateral optic projections. Example, most fish have some ipsilateral optic
fibers, whereas the popular genetic model zebrafish have none. In birds, fish,
amphibians, and reptiles, the ipsilateral ganglion fibers originate from
throughout the retina compared to the temporal retinal origin in mammals (1).
Amphibians vary considerably between species with most having some
ipsilateral projections to pretectal, hypothalamic, and primitive lateral
geniculate nucleus (LGN).
Many fish display ipsilateral connections mostly to hypothalamic, ventral
and lateral LGN, and pretectal areas. In nonmammals, some birds and some
fish, the nerves simply overlap at the chiasm with little contact (2). Optic
nerves in fish generally overlap each other rather than interdigitate. Optic
nerves of birds usually interdigitate. Only in the chiasm of primates and
marsupials does one see spatial organization with the ipsilateral fibers of
temporal origin coursing through the chiasm as an organized lateral cluster
(3). In most vertebrates, including cats, the passage of ipsilateral ganglion
cells through the optic chiasm is haphazard.
A feature of the mammalian visual system is the progressive increase of
the temporal retina as the eyes progress frontally from lateral locations in
guinea pig, rabbit, mouse, rat to the frontal position in primates,
commensurate with increase of noncrossing temporal retina ganglion cells at
the optic chiasm. Ipsilateral input progresses from little in rodents to succinct
point to point mapping of the visual world from LGN to cortical binocular
cells in cats, primates, and some marsupials (4). Complete separation of
ipsilateral and contralateral RGCs at the optic chiasm is limited to primates
and some marsupials (5). Figure 6-1 is a schematic of the distribution of
temporal and nasal RGCs at the chiasm in normally pigmented humans.
Primate vision is the most complicated of mammals. The principal hypothesis
of why there is nearly equal representation in each hemisphere of nasal and
temporal retina in primates is that equal representation developed in concert
with frontal vision and stereopsis to aid visually guided hands (6).
FIGURE 6-1 Drawing of approximate distribution of
crossed and uncrossed retina ganglion cells at the optic
chiasm of human beings.
Within a species, visual system organization is usually consistent. Rodents

and rabbits with laterally placed eyes and little temporal retina have few
ipsilateral optic ganglion fibers, numbering <10% (Figure 6-2). In 1965,
Sheridan (7) compared the interocular transfer of brightness and pattern
discriminations of corpus callosum–sectioned albino and pigmented rats.
Sheridan found dramatic differences between the performance of ipsilateral
optic fibers comparing albino and pigmented rats. He hypothesized, “Perhaps
the paucity of uncrossed fibers that characterizes rodents in general is even
further reduced in the albino.” Lund (8) anatomically verified Sheridan's
hypothesis stating albino rats have no organized uncrossed optic fibers.
FIGURE 6-2 Drawing of range of crossed and uncrossed
retinal ganglion fibers at optic chiasm of rodents and
rabbits.
Rodents possess a simple LGN with the 90%+ of crossed RGC terminations
filling the nucleus except for a pocket that contains <10% of the total
terminations located on the ipsilateral side. The LGN of albino rodents lacks
a defined area or pocket of ipsilateral RGC terminations. The few ipsilateral
fibers terminating in the LGN are fragmented with poor organization (Figure
6-3).
FIGURE 6-3 The distribution of optic fiber terminations
in dorsal lateral geniculate nucleus (dLGN) of BLACK
and ALBINO rats shown in serial sections at three levels
of dLGN.
A review of behavioral studies designed to assess visual learning capability

of the limited uncrossed optic fibers in rats reveals that outcomes can be
predicted based on whether albino or pigmented rats were used. Pigmented
rats learn, whereas albinos do not if visual information is limited to uncrossed
optic fibers (7,9, 10, 11, 12, 13).
Normally, pigmented cats, primates, and some marsupials have clearly
laminated LGN nuclei organized as alternating crossed and uncrossed
laminations with point to point representation of visual space maintained in
columns through the laminations of the LGN (Figure 6-4). Some cells at the
cortical level receive connections from both retinae via adjacent laminations.
This is the underlying anatomical basis of binocular stereovision. The
retinotopic organization and integration of binocular input progresses in an
orderly fashion from retina to LGN laminations and then on to cortical cells.
FIGURE 6-4 Organization of central visual pathways in
primates and cats. Schematic of the lamination of LGNd in
cats and primates. The visual field projected on to the
retina is maintained though LGN to cortex producing
anatomical substrate enabling stereovision.
VISUAL SYSTEM ORGANIZATION IN
CATS AND PRIMATES
For several years, the anatomical anomaly of few uncrossed RGCs at the
optic chiasm in albino mammals appeared to be limited to rats, guinea pigs,
and rabbits (14). Guillery (15) described aberrant retinogeniculate
organization in Siamese cats, but the connection to albinism was not
recognized.
The first connection of Siamese cats with albinism and suggestion that
reduced uncrossed optic fibers likely is a “highly general transspecies
phenomenon in albinic mammals” was in 1971 (16,17). Siamese cats possess
a tyrosinase-locus mutation that is a temperature-sensitive pigmentation
defect, that is, pigment forms only on the cold parts of the body similar to
Himalayan mice, rats, and rabbits. Siamese cats demonstrate significantly
reduced uncrossed optic pathways. Siamese cat's cortical visual area shows a
preponderance of contralateral innervation with few binocular cells.
Phylogenetically older connections are not as affected in albino
mammals. Retinohypothalamic projections predate vertebrates found in early
chordates possessing retinal pigment matching vertebrates (18). Melanopsin-
sensitive RGCs are completely crossed or bilaterally projecting to
suprachiasmatic nuclei and are not affected in albinos (19,20). The
phylogenetically older projections to suprachiasmatic nuclei in albino cat
appear to be unaffected (21). The crossed/uncrossed ratio of approximately
1.4:1 of retinal ganglion fibers to the suprachiasmatic nuclei in albino cats
corresponds to the ratio reported for normally pigmented cats (22). The later-
evolving RGC system through the optic chiasm is vulnerable to alteration in
albino mammals, whereas the phylogenetically older melanopsin pathway is
not.
Anatomical studies in the 1970s reported that all albino mammals with
oculocutaneous, or only ocular albinism, demonstrate reduced uncrossed
optic projections (23, 24, 25). Figures characterizing variation of visual
organization in the Siamese cat are in R.W. Guillery's 1974 publication in
Scientific American (26). The consequence of excessive retinal fibers that
cross at the chiasm and terminate in the LGN is fragmentation of the laminae
leading to disruption of the point to point mapping of visual space within the
LGN laminae. Disruption continues onto cortical projections and greatly
reduces the number of binocular cells. The consequence to visual perception
is that patients with albinism have poor stereovision.
Excessive temporal RGCs crossing at the chiasm arrive in the LGN and
overcrowd the crossed lamina. The misrouted RGC terminations spill over,
invade the adjacent uncrossed layer, and intrude into adjacent ipsilateral
laminations, thereby fragmenting the LGN laminae and disrupting point to
point mapping in the LGN that is passed on to the visual cortex. Figure 6-5
depicts the concept of how excessive crossed RGCs fragment projections
from retina to LGN. Temporal retina fibers projecting to the contralateral
LGN interrupt ocular dominance columns through the LGN (21,27).
Fragmentation of a layer devoted to uncrossed RGC terminations pictured in
Figure 6-5 would be a minor expression in Siamese cats. The majority of
Siamese and type 1A albino cats show greater fragmentation of uncrossed
laminae. A severe expression of an abnormal LGN is that of type 1A albino
cats. Most retinal fibers cross at the chiasm depicted in Figure 6-6.
FIGURE 6-5 Schematic example of disruption and
fragmentation of the lamination of LGNd in an albino
producing serious misrepresentation of the visual field to
cortex reducing the number of cortical binocular cells.
FIGURE 6-6 Dark-field autoradiograph of coronal section
through optic chiasm of a type 1A albino cat after injection
of tritiated leucine in right eye showing difference between
contralateral and ipsilateral RGC projections. Arrows point
to projection into suprachiasmatic nuclei. Scale bar 1 mm.
From Creel D, Hendrickson AE, Leventhal AG. Retinal projections in tyrosinase-negative albino cats. J
Neurosci 1982;2(7):907–911.
Figure 6-7 depicts the geniculate nuclei of a type 1A albino cat compared to
a normally pigmented cat. The dorsal lateral geniculate nuclei (dLGN) of
albino cats, which includes only one small pocket of ipsilateral RGC
terminations, is as primitive as a rodent dLGN. (The dLGN is the relay
station in the thalamus where RGCs terminate. The next cells send the retinal
information to the cortex.) Four patterns of geniculocortical organization are
described in albino mammals (26, 27, 28, 29). All forms likely exist in
patients with albinism.
FIGURE 6-7 Dark-field autoradiograph of coronal
sections through the ipsilateral (A) and contralateral (B)
dLGN of a pigmented cat, and ipsilateral (C) and
contralateral (D) dLGN of a type 1A albino cat. The one
small pocket of ipsilateral RGC terminations in albino cat
is indicated by arrow in panel D.
The brain of an albino patient (30,31) and an albino monkey brain were
studied and both had anomalous LGNs (32). Defects identified in albino
mammals encompass a decreased number of photoreceptors, foveal/area
centralis hypoplasia, misrouting of the temporal RGC axons across the
chiasm, variation of central RGC terminations, vascular intrusion into area
centralis, anomalous cortical organization, fewer cones and RGCs in the
macular area, and panretinal reduction in the number of rods (33, 34, 35, 36,
37, 38).
The first evidence of optic misrouting at the chiasm in humans was
published in 1974. Scalp-recorded visually evoked potentials (VEPs)
recorded from 20 patients including four genetic forms of oculocutaneous
albinism exhibited electrophysiologic evidence of reduced uncrossed optic
fibers (39). Similar results were published for patients with ocular albinism
(40) and replicated in early studies by Taylor (41) and Coleman et al. (42).
Reviews of research are found in the references (3,35,43,44).
In the normally pigmented primate, there is a distinct dividing line, the
vertical meridian, at the fovea. Retinal ganglion fibers temporal to the fovea
remain ipsilateral in an organized bundle at the chiasm, whereas retinal
ganglion fibers nasal to the fovea cross into the contralateral hemisphere at
the optic chiasm. Figure 6-8 shows the approximate difference in distribution
of retinal ganglion fibers at the chiasm comparing pigmented individuals to
patients with albinism. Ganglion fibers from up to 15 degrees of temporal
retina also cross at the chiasm in patients with albinism. The proportion of
misrouted ganglion fibers and associated anomalies is quite variable,
including in patients with albinism (44), probably due to a combination of
variation in degree of pigment and individual genetic factors. The level of
retinal pigmentation correlates negatively with the severity of expression of
visual anomalies (45,46). It is likely that the more pigment present in the
retina, the more normally routed RGC fibers will be present. Drack (47)
reported that more albinism-related mutations in an individual are associated
with greater expression of albinism.
FIGURE 6-8 Diagram of approximate distribution of the
optic nerve ganglion cells at the optic chiasm in a
pigmented individual and a patient with albinism.
All type 1A albino cats and most patients with albinism have eye movement
disorders. Most patients with albinism have searching nystagmus due to poor
foveal development. Additionally, many Siamese cats, most type 1A albino
cats, and many patients with albinism have strabismus and reversed
optokinetic nystagmus (OKN) due to miswiring of optic projections in the
brainstem (48).
Located in the brainstem are the nuclei integrating visual sensory input
including the superior colliculus (SC) and other nuclei involved in eye
alignment and fixation. Reduced ipsilateral RGCs in albino mammals include
reduced ipsilateral RGCs to mid-brain nuclei that affect strabismus,
nystagmus, and atypical OKN in albino mammals further complicating the
elaborate connections between these nuclei. In albino guinea pigs, rabbits,
and ferrets, there are no ipsilateral optic projections to the ventral lateral
geniculate nucleus (LGNv), SC, pretectum, nucleus of the optic tract (NOT),
or accessory optic system (AOS) (44,49, 50, 51).
In the normally pigmented cat, about 50% of RGCs project via the LGN
to the ipsilateral cortex. Twenty-four percentage of RGCs originating in
temporal retina project to the ipsilateral SC (52), and about 15% of RGCs
project to the ipsilateral pretectum (53). The type 1A albino cat has very few
ipsilateral projections to the pretectum and almost no ipsilateral projections to
SC (21). All type 1A albino cats have nystagmus and many have strabismus.
Aberrant OKN is described in patients with albinism (54, 55, 56). The
nuclei responsible for the OKN are the terminal nuclei of the AOS: the dorsal
terminal nucleus (DTN), lateral terminal nucleus (LTN), medial terminal
nucleus (MTN), and the NOT. For detailed account of brainstem nuclei
controlling eye movement and dissociated vertical deviation (DVD), see
Creel (57).
Readily observable symptoms of albinism, many of which are not limited
to patients with albinism, include iris transillumination, poor foveal
development, and intrusion of capillaries into the foveal avascular zone that is
apparent during ocular fundus examination and fluorescein angiography.
Also, poor foveal development can be apparent in ocular coherence
tomography (OCT), and reduced visual acuity can be seen. The consequences
of having few binocular cells in cats and patients with albinism are poor
binocular stereovision.
Figure 6-9 compares a normal fovea to that of a patient with reduced
retinal pigment. The OCT is in color to emphasize that the nerve fiber layer
in patients with albinism usually continues over the fovea similar to a mouse
that lacks a fovea. Expression of degree of foveal development varies
considerably as does visual acuity.
FIGURE 6-9 Comparison of optical coherence
tomography (OCT) through the foveal area of an ocularly
pigmented human being (A) and a patient with albinism
(B) foveal areas. In (A) note, the foveal depression and
that the nerve fiber layer, that is, red layer on top, is not
present across the foveal area. In most patients with
albinism, the nerve fiber layer (B) continues over the
potential foveal area.
Most patients with albinism have poor visual acuity, but some have good
acuity in spite of no foveal pit. Figure 6-10 shows OCT of a patient with
albinism with 20/30 visual acuity. Although seemingly surprising, there are
normally pigmented individuals with good acuity despite no foveal pit (58).
FIGURE 6-10 Optical coherence tomography (OCT)
through the foveal area of a patient with albinism with
20/30 visual acuity.
A new method for quantifying pigment in the retina is fluorescence lifetime

imaging ophthalmoscopy (FLIO). FLIO is a novel method quantifying
pigments in the retina (59) that measures the fluorescence lifetimes from
fluorophores within the retina other than lipofuscin that dominates fundus
autofluorescence (FAF). FLIO has been used to characterize albinism
(Figure 6-11). Due to lack of macular pigment, patients with albinism are
missing short FLIO lifetimes indicated by red color in the top FLIO images
of a healthy control, but the technology is new and still being investigated.
FIGURE 6-11 Fluorescence lifetime imaging
ophthalmoscopy (FLIO) images from both spectral
wavelength channels (SSC, short spectral channel, 498 to
560 nm; LSC, long spectral channel, 560 to 720 nm),
fundus autofluorescence intensity, and OCT images from a
healthy 29-year-old as well as a 44-year-old patient with
albinism. The FLIO image shows the absence of macular
pigment (missing short central lifetimes depicted in red
color) in the patient with albinism. The autofluorescence
image shows vascular intrusion into the albino's foveal
area, and the albino OCT lack of fovea.
There are at least 21 types of human albinism. Many are rare limited to small
groups or associated with syndromes. More will be found. More exist in
animal models. Patients with oculocutaneous and ocular albinism with known
loci include those below (Table 6-1; see also (60)):
TABLE 6-1 Human Albinism
Evidence of optic misrouting similar to that seen in patients with albinism is

found in other conditions, possibly linked to albinism. Many are located on
chromosome 15, similar to OCA 2. These include the Prader-Willi syndrome
(15q11-13), Angelman syndrome (15q11-13), Kartagener syndrome (15q24-
25), and some with Asperger syndrome candidate gene location 15q11-13
(60, 61, 62, 63, 64, 65, 66). Chiasmic misrouting is not found in cases of
reduced hypopigmentation due to embryonic migration defects of neurons
and pigment cells from the neural crest such as in Waardenburg syndrome
and deaf white cat (W gene), vitiligo, or piebald spotting, nor other forms of
hypopigmentation such as phenylketonuria. These disorders do not exhibit
visual anomalies associated with albinism.
A consistent feature across all forms of mammalian albinism is
misrouting of temporal retinal fibers to the contralateral hemisphere at the
optic chiasm. Misrouting can be detected recording with only two scalp
electrodes across occipital scalp (Figure 6-12).
FIGURE 6-12 Drawing of International 10-20 System

scalp electrode locations highlighting locations H3 and H4
that often better demonstrate misrouting.
TECHNIQUE TO ASSESS OPTIC

MISROUTING
The best scalp locations are more lateral than the 10-20 International System
O1 to O2, near H3 to H4 (67). Place one EEG electrode at H3 and one at H4.
Plug one into positive and one into negative. A ground lead can be anywhere
on head. Because most patients with albinism have poor acuity and
nystagmus do not use pattern reversal stimuli. Pattern reversal stimuli
exacerbates nystagmus producing little or no recordable VEPs. Once one eye
is patched, record the flash or pattern-onset VEPs (68) and then switch the
patch and record the VEPs stimulating the other eye. In subjects with no optic
misrouting, there will be some asymmetry but little difference in the VEPs
recorded across occipital scalp (Figure 6-13).
FIGURE 6-13 Sample monocular VEP waveforms

recorded across occipital scalp of a normally pigmented
individual showing little change in waveform when each
eye is stimulated monocularly. Time epoch 250 ms.
Nearly all retinal projections in patients with albinism terminate in the

contralateral occipital hemisphere. Monocular VEPs send most information
to the contralateral occipital pole and produce a reversal in polarity of most
VEP components when VEPs are compared between eyes (Figure 6-14).
FIGURE 6-14 Sample monocular VEP waveforms
recorded across occipital scalp of an albino showing
reversal of polarity when each eye is stimulated. This
reflects that almost all optic nerve fibers cross at the optic
chiasm to opposite hemisphere in albino mammals. Time
epoch for VEP 250 ms.
The etiology of abnormalities in the visual system of mammals has been

discussed since the 1970s. The mechanisms controlling visual system
development including the optic chiasm are pretty well known. See Rasband
et al. (69), Jeffery and Erskine (70), Prieur and Rasband (71), and Herrera et
al. (72) for theories of molecular mechanisms directing ganglion cell routing
at chiasm. No solution for initiation of anomalies associated with reduced
retinal pigmentation within mammals has general agreement.
Originally, studies for the cause of misrouting in mammals associated
with reduced retinal pigmentation looked for a role during embryologic
development for melanin pigmentation in the optic chiasm or retina. In
mammals with reduced retinal pigmentation, many anatomical features are
affected besides chiasmic misrouting. Since multiple features are affected, the
abnormalities likely originate in an early event that impacts a wide population
and number of cell types over several time periods. Normal optic pathway
development appears to depend on interactions between RGCs from each eye
during specific temporal periods.
Finding misdirected temporal projections in heterozygotes carrying the
type 1A albino cat gene (73) and an analogous one in an ephrin-B2 knock-in
mice (74) provides evidence that chiasmic misrouting and the cascade of
related events are likely not directly related to lack of pigment but to a gene
effect of albinism that is initiated early in embryonic development.
THEORY
What is considered abnormal visual system organization in mammals with
reduced retinal pigmentation is normal visual system organization in retinally
pigmented vertebrates phylogenetically preceding these species. Cat visual
system disorganization can approach that of normally pigmented rodents. The
organization of the visual system in rodents resembles premammalian
vertebrates. This theory suggests that an early event in genetic coding for
melanin pigment in the nascent retina is the positive inductive signal that
launches normal retinal development and RGC targeting (57). Genetic
instruction for mammals with insufficient retinal pigmentation defaults to a
simpler, more entrenched genetic platform skewed toward complete crossing
of RGCs at the chiasm. In mammals' retinal pigmentation, retinal
embryogenesis defaults to the conserved genetic package that existed when
the species first evolved and has a cascade of consequences.
Ipsilateral fibers are not consistently present even among closely related
nonmammalian vertebrate species suggesting a vulnerability of ipsilateral
fibers being expressed. Within all vertebrate groups, the contralateral visual
projections are more stable across species than are ipsilateral projections.
Contralateral projection of RGCs at the optic chiasm is the conventional
ancestral state in nonmammalian vertebrates, and reduced nondecussated
optic fibers are the norm in ancestral mammals.
The essence of this theory is that retinal melanin pigment is so
fundamental to retinal, vascular, and early RGC targeting that coding for
insufficient retinal melanin triggers a switch, which thwarts the completion of
genetic instructions. Instead an atavistic expression of the conserved, more
stable, genetic platform skewed toward complete crossing of RGCs at the
chiasm develops consistent with genetics of each species' evolutionary origin.
Phylogenetically, late genetic addenda to conserved instructions are
vulnerable to not being expressed if cues, such as sufficient retinal melanin
pigmentation, are not present to support variation from highly conserved
coding.
ACKNOWLEDGMENTS
Supported in part by an Unrestricted Grant from Research to Prevent
Blindness, Inc., New York, NY, to the Department of Ophthalmology &
Visual Sciences, University of Utah. Images provided courtesy of Moran
Ophthalmic Imaging.
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7
Applications for Human Pluripotent
Stem Cell–Derived Retinal Cells in
Development, Disease, and Cellular
Replacement
Kirstin B. VanderWall, Clarisse M. Fligor, Sailee S. Lavekar,
Kang-Chieh Huang and Jason S. Meyer
INTRODUCTION
Human pluripotent stem cells (hPSCs), including both human embryonic
stem (hES) cells and human-induced pluripotent stem (hiPS) cells, provide an
advantageous model for studying cellular development and disease in vitro,
as they can be cultured indefinitely and have the ability to differentiate into
all cell types of the body. Embryonic stem (ES) cells are cultured from the
inner cell mass of the blastocyst in their pluripotent state and were among the
first types of stem cell to be used for studies of human development in vitro
(1,2). Unlike ES cells, iPS cells can be generated from patient sources
through the genetic reprogramming of cells back into a pluripotent state using
the Yamanaka factors (Oct4, Sox2, Klf4, and c-Myc) (3,4). This technology
has transformed the field of cellular biology with the ability to collect
samples from patients harboring specific genetic determinants for various
diseases and using these cells to study mechanisms and phenotypes in some
of the earliest stages of disease pathogenesis (5,6). More so, the development
of iPS cells has allowed for the new opportunity to screen therapeutic
compounds in vitro, as well as develop novel cellular replacement therapies
(7).
Research in the retina has taken full advantage of hPSCs for studies of
development and disease. Over the past decade, researchers have
demonstrated the differentiation of hPSCs into all major retinal cell types
including retinal pigment epithelium (RPE), photoreceptors, and retinal
ganglion cells (RGCs) (Figure 7-1) (8, 9, 10). Numerous differentiation
protocols have been established that utilize either direct differentiation of
retinal cells using small molecules or stepwise protocols through default
neural specification (Table 7-1). More recent research has largely focused on
the development of retinal organoids from hPSCs, which are three-
dimensional tissue-like structures that mimic the spatiotemporal specification
of retinal neurons during development (18,33). Retinal organoids are of great
interest as they recapitulate the cellular interactions of retinal neurons in vivo
and can be a resourceful tool for reproducibly differentiating large
populations of retinal neurons for isolation and cell replacement strategies.
FIGURE 7-1 Derivation of retinal cell types. hESCs and

hiPSCs can be differentiated into three major cell types of
the retina: RPE, photoreceptors, and retinal ganglion cells.
TABLE 7-1 Summary of features used to
characterize hPSC-derived retinal cells
The ability to derive all major cell types of the retina from hPSCs allows for
in-depth studies of retinal fate specification with the ability to observe some
of some the earliest events of human retinogenesis. Patient-specific iPS cells
provide an advantageous tool for studying retinal diseases and the
mechanisms that result in degeneration of specific cell types. The recent
advent of gene editing approaches, like CRISPR/Cas9, allow for the
development of new disease models through the insertion of a mutation into
isogenic control parent lines as well as the correction of specific mutations in
existing patient iPSCs. Isogenic controls are highly important in the stem cell
field due to the variability known to exist between different cell lines. More
so, these cells can also be utilized for large-scale pharmaceutical screenings
for targeted therapeutics in retinal degeneration. Lastly, hPSCs may serve as
a valuable resource for cell replacement strategies with the ability to derive
unlimited quantities of cells in a reproducible and efficient manner. Current
efforts in the field have demonstrated some degree of success for the
replacement of stem cell–derived RPE and photoreceptors and limited studies
to date demonstrating the ability to replace inner retinal neurons such as
RGCs that degenerate in glaucoma. Ongoing studies of hPSC-derived retinal
cells will assuredly continue to focus on the efficacy of outer cell replacement
strategies and the future development of therapies for inner retinal diseases
causing death and degeneration of RGCs.
RETINAL PIGMENTED EPITHELIUM
Functions and Diseases of the RPE in the Retina
The retina is a highly organized structure located within the back of the eye
that consists of numerous interconnecting cell types working together to
allow for proper vision (Figure 7-2) (34). Each of the cell types that make up
the retina performs specific functions in the visual transduction pathway, and
these functions are perturbed in diseases that cause degeneration of the retina.
The outermost cell type of the retina is the retinal pigmented epithelium
(RPE), which forms a monolayer of pigmented cells between the neuroretina
and the retinal vasculature of the choroid. The apical face of the RPE
interacts closely with photoreceptor outer segments, whereas the basal edge
of the RPE faces the Bruch membrane, which is the innermost layer of the
choroid (35). The close association of the RPE to these two structures heavily
relates to their physiologic functions within the retina.
FIGURE 7-2 Cellular architecture of the retina. A

schematic of the retina representing a highly organized,
multilayered structure consisting of numerous neuronal
and supporting cell types.
The RPE serves a variety of important functions within the retina, including
the phagocytosis of photoreceptor outer segments, absorption of light,
transport of nutrients and ions, release of growth factors, and recycling of
important visual cycle compounds (35). Additionally, the RPE contribute to
the outer blood–retinal barrier, which regulates the contents that enter and
exit the retina and allows for its prolonged stability and integrity. As a result
of the continuous visual cycle, photoreceptors are subject to damage caused
by the accumulation of light-induced toxic substances, such as photooxidated
lipids and lipoproteins. Therefore, photoreceptor outer segments are rapidly
shed and replaced with the ingestion of old outer segments by RPE through
phagocytosis. Once inside the RPE, these segments are broken down into
essential molecules and transported back to the photoreceptors. The cycle of
outer segment renewal and phagocytosis is critical for proper
phototransduction, and disruptions to this cycle constitute a main phenotype
in blinding disorders.
When considering the anatomy of the retina, RPE plays the middleman in
the transport of ions and water out of the retina into the choroid through the
Bruch membrane as well as the transport of nutrients and glucose from the
blood into the photoreceptors through transepithelial transport (36). RPE
utilize a number of transporters and channels to serve in this capacity,
including glucose transporters, Na+/K+ ATPase, and Cl−/K+ transporters.
The high metabolic rate of retinal neurons creates a need for the constant
movement of glucose into the retinal layers for energy and the subsequent
removal of water from the inner retina into the choriocapillaris and subretinal
layers through aquaporin-1 channels in the RPE (35). The RPE also delivers
other nutrients into the photoreceptors from the blood such as retinol,
ascorbic acid, and fatty acids, and each of these components plays an
important role in the visual cycle.
With these many functions in mind, the overall health and maintenance of
RPE is crucial to proper visual transduction and longevity of the retina. As
such, many retinal diseases evolve because of dysfunction of the RPE,
including age-related macular degeneration (AMD), Bestrophinopathies (e.g.,
Best disease), Stargardt disease (see Chapter 30), Leber congenital amaurosis
(LCA) (see Chapter 27), and retinitis pigmentosa (RP) (please see e-Chapter
4), and each of these diseases is associated with RPE dysfunction leading to
visual decline. The pathogenesis of each of these diseases can be related to
important RPE pathways that result in a lack of support to the photoreceptors
and lead to loss of vision. Many of these diseases are known to be caused by
hereditary mutations in visual proteins.
RPE dysfunction and vision loss stem from hereditary genetic mutations
in proteins known to play a major role in the visual pathway. Best disease is
an autosomal dominant disease caused by mutations in the BEST1 protein,
which functions as calcium-activated chloride channel; Best disease
manifests in early childhood by the appearance of a “yolk-like lesion” in the
macula (37). A variety of pathophysiologic mechanisms have been associated
with the onset of Best disease, including abnormal fluid flux associated with
the inability to properly move chloride ions out of the retina, accumulation of
proteins and debris, impaired outer segment degradation, and oxidative stress
(38). Dysfunction of these pathways in Best disease leads to progressive
vision loss with increasingly reduced ability to perform tasks, such as reading
and driving.
Stargardt disease also affects the central retina and is the most common
form of hereditary dystrophy. This disease is autosomal recessive and caused
by mutations in the ABCA4 gene or (less commonly the ELOVL4 gene as an
autosomal dominant mutation; see also Chapter 30) and appears in childhood
and early adulthood (39). Lack of ABCA4 results in the impaired transport of
visual cycle by-products and accumulation of bisretinoid A2PE in outer
segment discs. A2PE can be hydrolyzed to form A2E, a highly toxic
metabolite, and accumulates with lipofuscin in the RPE leading to RPE
dysfunction and death with secondary photoreceptor loss. Clinically, patients
demonstrate macular atrophy and display the appearance of yellow-white
flecks in RPE ultimately resulting in central vision loss. Additionally, another
hereditary disease, which affects proteins in the visual cycle is LCA. In a
subset of cases, LCA can be caused by mutations in RPE65 (40). RPE65 is a
protein produced and expressed by the RPE and functions as an isomerase
that converts all-trans-retinyl ester to 11-cis-retinol during the visual cycle.
Mutations in RPE65 result in reduced amounts of 11-cis retinol and esters
accumulation within the RPE leading to abnormal RPE function and early-
onset blindness.
Lastly, forms of hereditary retinitis pigmentosa result in primary RPE
pathology and dysfunction (41). Similar to other proteins mentioned above,
CRALBP is expressed within the RPE and is implicated in the visual cycle. It
has been shown that CRALBP functions during the isomerization step and is
a major acceptor of 11-cis-retinol, which stimulates its conversion from all-
trans retinol during the rod cycle. Clinical manifestations of the disease
include early childhood night blindness and the presence of white dot-like
deposits that remain visible into adulthood. Accumulation of 11-cis retinol in
the RPE alters the normal function of RPE and the visual cycle. Taken
together, numerous hereditary and idiopathic diseases result in the
dysfunction of the RPE and lead to vision loss. These diseases can manifest
from time points of early childhood through elderly adulthood. Major efforts
in recent years have focused on a greater understanding of the underlying
disease mechanisms of these disorders with applications to develop
treatments and therapeutics targeted at improving and restoring vision,
including the use of hPSCs for each of these purposes.
Utilization of hPSCs for Differentiation and Disease

Modeling of RPE
The ability to derive RPE from hPSCs has been widely researched over the
past decade with various protocols describing the derivation of such cells
(Table 7-1). Previous studies have utilized either spontaneous differentiation
of hPSCs into RPE through the removal of basic fibroblast growth factor
(bFGF) (11,12,42, 43, 44, 45), or directed differentiation of hPSCs into a
retinal lineage largely following the major steps of retinogenesis and yielding
all cell types of the retina (8,10,13, 14, 15, 16,46, 47, 48). Although either
method of differentiation yields viable sources of RPE from hPSCs, key
differences may affect the quality and efficiency of yielding this cell type.
There is a pressing need to increase the yield of RPE from hPSCs as well as
decrease the time of differentiation for their use in therapeutics and cell
replacement strategies to treat and study diseases affecting the RPE.
Using spontaneous differentiation methods, initial studies describing the
differentiation of RPE from hPSCs resulted in areas of pigmentation that
could be identified and manually dissected from the dish to be expanded and
passaged. Once pigmented areas are removed from adherent culture through
these protocols, these populations produce an extremely pure population of
RPE that possess a hexagonal and pigmented morphology and express
characteristic RPE markers such as MITF and ZO-1 (Figure 7-3).
Alternatively, approaches for generating hPSC-RPE through a directed
differentiation have been established. In some cases, this differentiation
begins with neural specification giving rise to cells of the eye field and
further to retinal progenitors. Retinal progenitors then have the capacity to
differentiate into all cell types of the retina, which can be modified by the
addition of various growth factors and inhibitors (8,10,13,15,47,48).
Numerous studies have demonstrated the importance of the WNT and
transforming growth factor beta (TGF-β) pathways for RPE specification
with the addition of small molecules at specific time points of differentiation
that increase the efficiency of RPE differentiation (13,15,47, 48, 49, 50, 51,
52, 53). In a more recent study, differentiation of RPE was described by
blocking the FGF/MAPK pathway resulting in RPE differentiation without
the inhibition of WNT/nodal signals with this differentiation significantly
improved with the secondary inhibition of the PKC/BMP pathway (17).
Furthermore, studies have now switched to optimizing xeno-free and cGMP
environments for the differentiation of RPE from hPSCs in order to make
these cells more suitable for cell transplantation in future studies (54, 55, 56).
Utilization of directed differentiation protocols increases the efficiency of
hPSC-RPE generation and can reproducibly yield large quantities of RPE
expressing characteristic genes and proteins for disease modeling, cell
replacement therapies, and large-scale drug screening.
FIGURE 7-3 hPSC-derived RPE characteristics. A:)

Bright fight microscopy demonstrates the hexagonal
monolayer of pigmented epithelial cells. B, C:hPSC-
derived RPE expresses characteristic markers of RPE
including ZO-1 and Ezrin. Scale bar equals 10 μm.
In order for hPSC-derived RPE to serve as a reliable source for cell

replacement and a model system for RPE diseases, these cells must exhibit
functions and characteristics of the in vivo cell type. Previous studies have
shown the ability of hPSC-derived RPE to perform phagocytosis of
photoreceptor outer segments (11,13,45,50,52,53,56, 57, 58), polarization of
cell membranes (13,53,57,58), calcium responses (47,57), as well as exhibit
electrical properties including the maintenance of a trans-epithelial resistance
through formation of tight junctions (56,57). The demonstration of these
various characteristics is even more important when deriving RPE from
patient-specific sources to model various inherited diseases, with many of
these functional properties compromised in diseases affecting the RPE.
Several studies have been reported using RPE derived from patient-
specific sources to model various diseases including gyrate atrophy (59), Best
disease (60, 61, 62, 63), LCA (64, 65, 66), AMD (67, 68, 69), and retinitis
pigmentosa (70, 71, 72, 73). Access to patient-specific sources allows for the
generation of reproducible RPE from hiPSCs that harbor specific genetic
mutations linked to RPE-related diseases and provide a reliable model system
for discovering phenotypic attributes to clinical presentation in order to
understand molecular mechanisms that result in the degeneration of RPE. In
some of the earliest hiPSC modeling studies, RPE exhibited phagocytotic
deficits, oxidative stress, altered calcium homeostasis, and
disorganization/loss of cellular characteristics in relation to various diseases
targeting RPE.
hiPSC-derived RPE can be produced in large and reproducible quantities,
which provides an ideal tool for drug screening applications targeting specific
disease deficits and allow for the future development of therapeutics to
protect RPE from further damage. In 2015, the rescue of photoreceptor outer
segment degradation was demonstrated by the addition of valproic acid in a
hiPSC model of Best disease (62). Additionally, the ability for the drug
PTC124 to significantly improve RPE phagocytosis of photoreceptor outer
segments in RPE grown from a patient with retinitis pigmentosa (72) has
been recently demonstrated. Thus, the ability to generate RPE from patient-
specific sources allows for the modeling of disease mechanisms targeted at
RPE in various diseases and provides a reliable platform for the discovery of
new therapeutic drug compounds that can ameliorate degeneration of these
cells with the hope of validating future compounds for clinical treatments.
Alternatively, gene therapy and gene augmentation to restore proper
protein function is also a large area of study for therapeutic intervention of
RPE diseases. This type of therapy usually involves the delivery of an
unaffected protein via adeno-associated viral (AAV) or lentiviral vectors.
This type of therapy has been used in a variety of hiPSC studies including
retinitis pigmentosa through the delivery of membrane frizzled-related
protein (70), for LCA through the lentiviral delivery of CEP290 (74), and
most recently the lentiviral gene augmentation to rescue Kir7.1 channel
function in an hiPSC model of LCA (66). CRISPR/Cas9 gene editing is
another attractive option for gene therapy that can be used to correct gene
mutations in hiPSCs and generate healthy, functioning RPE from patient-
specific sources (58,75). This technology holds great promise for the ability
to transplant autologous and gene-corrected RPE for treatments of inherited
eye disorders.
hPSCs as a Platform for RPE Replacement for Treating

Blinding Disorders
One of the most promising aspects of hPSC technology is their potential for
cellular transplantation to treat retinal diseases. hPSC-derived RPE represent
a highly attractive source for cellular replacement as they can be generated in
reproducibly large quantities and can be patient specific to reduce immune
reactions. In the past decade, numerous studies and clinical trials have been
conducted in order to understand the safety, efficacy, and efficiency of using
hPSC-derived RPE for cell replacement strategies to treat outer retinal
disorders. Research regarding the ability to use hPSCs for cell replacement
made headway back in the 2000s (44,76) utilizing a rat model of AMD.
These studies provided the first evidence for the safety and utilization of
hPSC-derived RPE to treat an AMD model as their transplantation resulted in
cell survival up to 200 days and indicated a lack of tumor formation. In 2012,
the results of the first human phase I/II clinical trial sponsored by Astellas for
hPSC-RPE cell replacement for AMD and Stargardt was published with their
follow-up results published in 2015 (73,77,78). Patients were administered
transient dosages of different concentrations of hPSC-derived RPE into the
subretinal space and demonstrated pigmentation, increased visual acuity, and
no evidence of proliferation, rejection, or systemic safety issues. It is
currently expected that the first long-term follow-up results will be available
in December of 2019. In November of 2018, an additional phase I/II clinical
trial was published and focused on the preservation of retinal structure
following subretinal injection of hESC-derived RPE in patients with
advanced Stargardt disease (79). Patients in the study demonstrated subretinal
hyperpigmentation and no sign of uncontrolled proliferation or inflammatory
responses, although at the highest dose some retinal thinning and decreased
sensitivity was apparent. These two studies utilized the delivery of hPSC-
derived RPE as a single cell suspension into the subretinal space, although
new methods of hESC-derived RPE growth and delivery are being currently
researched.
A large majority of this research has been placed on the development of a
more permanent implantation of hPSC-derived RPE in which cells are
transplanted in an intact monolayer and grown on various substrates and
artificial scaffolds before transplantation (80, 81, 82). Growth of RPE in such
a way yields functional and polarized RPE with the hope of this functionality
translated once implanted in the host retina. Regenerative Patch Technologies
has sponsored a phase I/IIa clinical trial for the transplantation of hESC-
derived RPE grown on a parylene membrane to patients with advanced dry
AMD, and primary results are expected in 2019 with completion in 2023. In
Brazil, a comparative phase I/II clinical trial is currently under way
examining the injection of suspended hESC-derived RPE into the subretinal
space versus the surgical implantation of hESC-derived RPE seeded on
polymeric substrate with results of this study expected within the year.
Lastly, Pfizer is currently sponsoring a phase I clinical trial for the
transplantation of hESC-derived RPE grown in monolayer on a polyester
membrane in patients with wet AMD, with results from this study also
expected in 2019.
Thus, by the conclusion of 2019, the results from multiple clinical trials
are expected to become available from both cell suspensions of RPE as well
as transplanted sheets of RPE derived from hPSCs. These results will provide
essential information and allow for the development of a more standardized
approach for replacing RPE in diseases like Stargardt disease and AMD.
Aside from suspension versus sheet transplantation, a variety of other
challenges should be addressed in future studies including immunogenicity,
functional integration, and cell survival. The advancement of CRISPR/Cas9
technology to correct gene mutations allows for unique patient-specific
treatment with a lower chance of immune rejection, although studies have
been published regarding the differential efficiency of RPE differentiation
between cell lines. To this extent, allogenic transplantation using a
standardized cell line may provide a more scalable and consistent approach
for cellular replacement. Additionally, once transplanted, RPE need to
survive and functionally integrate into the retina in order to efficiently restore
vision, which may be better achieved with supplementation of growth factors.
More studies will need to be conducted in the future to optimize these
challenges, although the positive results from current clinical trials for hPSC-
derived RPE transplantation demonstrate the potential to eventually improve
and even restore vision for patients.
PHOTORECEPTORS
Functions and Diseases of Photoreceptors in the Retina
Photoreceptors are the main light-sensing neurons of the retina and are
responsible for the conversion of light signals into an electrical impulse.
These cells are found in the back of the retina adjacent to the RPE and
constitute the outer nuclear layer (Figure 7-2). The anatomy of
photoreceptors is broken down into five main parts: the outer segments (OS),
the connecting cilium (CC), the inner segment (IS), the nuclear region, and
the synaptic region (Figure 7-4). The OS are found apically and interact
closely with neighboring RPE. Membranous discs are found throughout the
outer segment and contain photopigments like opsins. When light is detected
by the photoreceptors, 11-cis retinal is isomerized to all-trans retinal and
activates opsins. A cascade of events leads to the closing of ion-gated cyclic
nucleotide channels that results in hyperpolarization of the cell. The
hyperpolarization travels through the CC and IS and then finally through the
nuclear region to a specialized ribbon synapse to release glutamate in a
graded fashion to connecting horizontal and bipolar cells that eventually
reach the RGCs, which send visual information to the brain (83).
FIGURE 7-4 Major structural components of the

photoreceptor. A schematic depicting the major structures
of rods and cones including the outer segments (OS),
connecting cilium (CC), inner segment (IS), nuclear
region, and synaptic region.
Photoreceptors come in two variations, rods and cones. Cone photoreceptors

contain three subtypes, S-, M-, and L-cones, which are responsible for color
perception through specialized opsins. Cones are found in high density in the
central fovea and allow for high spatial acuity vision. There is only one type
of rod photoreceptor responsible for dim light detection with low spatial
acuity. Compared to cones, rods make up about 95% of the photoreceptor
population and can be found in peripheral regions of the retina, whereas
cones make up 5% and can be found mainly in the central retina. Both rods
and cones rely heavily on adjacent RPE for routine maintenance in degrading
OS, recycling of visual molecules, and transporting of nutrients in order to
function properly and allow for visual transduction (84).
The visual transduction pathway is affected in numerous outer retinal
disorders, including AMD and inherited retinal disorders (IRDs). In contrast
to AMD, which is a cone-targeted disease that results in distinct central
vision loss and can be caused by numerous mutations and environmental
factors, IRDs are a group of diseases caused by inherited monogenic
mutations including retinitis pigmentosa, LCA, Stargardt disease, Usher
syndrome, and Bardet-Biedl syndrome (BBS) (see Chapter 32) (85,86).
These diseases affect the rod and cone photoreceptors selectively and are a
major cause for visual impairment of the outer retina.
Retinitis pigmentosa (RP), also known as rod–cone dystrophy, is one of
the most common forms of inherited rod-specific degeneration. As this
disease targets the rod photoreceptors initially, typical symptoms of the
disease include night blindness and tunnel vision. Following the initial
degeneration of rods, the cone photoreceptors degenerate secondarily in later
stages of disease progression and result in complete blindness. In the clinic,
RP can be identified by the abnormal fundus, abnormal a- and b-waves
during ERG, and reduced visual field acuity (87). Numerous mutations have
been implicated with the development of RP, including autosomal dominant,
autosomal recessive, and X-linked recessive inheritance patterns. Genes that
are found to be photoreceptor-specific are most commonly associated with
RP progression, including mutations in rhodopsin (present in 25% of
autosomal dominant cases) with rhodopsin serving in one of the first steps in
the visual transduction pathway. Additional genes found to harbor gene
mutations in RP include PRPF31, PRPH2, and RP1 with each of these genes
playing a major role in photoreceptor characteristics and functionality
(88,89).
Syndromic forms of RP have also been highly implicated in retinal
degeneration including Usher syndrome and Bardet-Biedl syndrome. Usher
syndrome (see Chapter 33) is autosomal recessive inheritance and results in
hearing loss, vestibular dysfunction, and RP-like visual deficits with these
deficits identifiable in the clinic from early adolescence to adulthood
depending on disease type and severity (90,91). Thirteen genes have been
identified to cause this disease including USH2A, DFNB31, and GPR98 with
their respective proteins known to be expressed in photoreceptor IS, CC, and
the synapse, all of which can play a role in disease mechanisms leading to
visual decline. BBS is also characterized by RP-like retinal degeneration
along with symptoms such as polydactyly, cognitive impairment, and obesity.
This syndrome is also inherited in an autosomal recessive fashion, with more
than 14 genes identified for BBS. The five major genes implicated in BBS
cases are BBS1, BBS10, BBS2, BBS9, and BBS6 (92). In vivo models of the
disease have indicated mislocalization of rhodopsin from the OS to the IS
leading to rod degeneration and apoptosis, and may play a major role in
clinical symptoms such as night blindness (93).
Mutations in CEP290 (BBS14) have been associated with the
development of BBS as well as severe LCA. CEP290 is expressed within
photoreceptor cells and has plays an important role in formation of cilia and
OS protein trafficking (40,94). As such mutations in CEP290 lead to
abnormal photoreceptor development and function in early childhood
resulting in significant vision loss. Each of the described diseases severely
affects either cone or rod photoreceptor development and function and lead to
permanent vision loss with the specific mechanisms known to cause this
degeneration in human cells. Although numerous in vivo models of these
outer retinal diseases have been developed, models are not accurate in
directly predicting how human cells will degenerate in response to specific
mutations. Therefore, the use of hPSCs to model photoreceptor development
and disease would provide a more translatable model system with the ability
to understand the molecular mechanisms of photoreceptor specification as
well as identify novel pathways that may be important in outer retinal disease
pathogenesis in humans.
Utilization of hPSCs for the Differentiation and

Disease Modeling of Photoreceptors
Within the past decade, numerous differentiation protocols have been
established and optimized to study the development of photoreceptors,
including their use in disease models and preclinical transplantation studies
(Table 7-1). In 2006, the first study was published describing the
differentiation of retinal cell types, including photoreceptors, from hPSCs (9).
This differentiation protocol utilized a suspension embryoid body (EB)
system with the addition of Dkk-1, BMP agonist, and IGF-1. Results of this
protocol resulted in the production of CRX-expressing photoreceptors
consisting of 10% of the cell population, although the number of these
photoreceptors that expressed mature markers of cones and rods were
minimal. A second protocol was published in 2008, and utilized the same
inhibition of the (bone morphogenic protein) BMP and WNT pathways
during early differentiation (95). In a follow-up study, the addition of retinoic
acid and taurine was added at later time points to enhance photoreceptor
maturation. Within 200 days of differentiation, photoreceptors were found to
express more mature markers of rods and cones, including rhodopsin,
red/green opsin, and blue opsin, and the addition of taurine and retinoic acid
resulting in 10% to 20% of cells expressing these markers (96). Additional
methods have described the generation of photoreceptor precursors through a
more direct, stepwise differentiation, with that caused these cells to express
CRX within 80 days of differentiation and co-express more mature markers
like recoverin and opsin in 46% of that population (10). These protocols
represented the foundational studies that resulted in the differentiation of
photoreceptors from hPSCs expressing a variety of associated markers
(Figure 7-5) and enabled the optimization of photoreceptor differentiation in
future studies.
FIGURE 7-5 Photoreceptors derived from hPSCs in 2D

culture. Following differentiation, hPSC-derived
photoreceptors express numerous associated markers,
including CRX, NeuroD1, OTX2, and recoverin. Scale
bars equal 25 μm for (A) and (B) and 15 μm for (C).
More recently, numerous differentiation protocols have continued to utilize

the initial inhibition of the BMP/WNT pathways with various modifications
to enhance the differentiation of rods and cones (19,97, 98, 99). The addition
of factors, including T3, FGFs, retinoic acid, and taurine were added to
culture media and found to enhance the differentiation of photoreceptors from
retinal progenitor cells (19,98,100,101). Furthermore, the addition of Coco
demonstrated a preferential differentiation of S-cone photoreceptors with
these cells found in 60% to 80% of the population; the addition of T3 led to
M/S-cone photoreceptors (99).
In recent years, many of these protocols have been modified to yield
three-dimensional retinal tissues (Figure 7-6). This type of culturing method
was first described by Meyer et al. in 2011, where neurospheres were
suspended in floating culture and allowed to differentiate from an optic
vesicle stage, expressing retinal progenitor markers up to 100 days where
these structures expressed rhodopsin and S-opsin (59,102). In 2012, a
different study described the formation of bilayered optic cup structures,
which demonstrated an invagination similar to what is seen during in vivo
retinogenesis with these structures later named retinal organoids (33). These
retinal organoids first expressed CRX, then recoverin, and then rhodopsin by
120 days of differentiation with this expression found exclusively on outer
layers similar to the layering of the in vivo retina. In the coming years,
numerous other protocols have adopted this three-dimensional retinal
organoid differentiation protocol to yield more mature photoreceptor cells
(18,20,21,62,103, 104, 105, 106). Using this type of differentiation, hPSC-
derived photoreceptors were able to demonstrate various morphologic and
function characteristics including responses to light (18,21,59,62,106,107),
the formation of cilia (21,65,106,108), and the formation of rudimentary
outer segment discs (18,21,22,106). One of the most appealing aspects of
retinal organoid differentiation is the recapitulation of the laminated retina
and the interactions of retinal cell types within this tissue resulting in more
mature photoreceptors.
FIGURE 7-6 hPSC-derived retinal organoids recapitulate
the layering of the retina. A:Retinal organoids
demonstrated morphologic characteristics indicative of
retinal-like organization. B:Retinal organoids recapitulate
the organization of the retina with RGCs expressing BRN3
within inner layers and photoreceptors expressing
recoverin in outer layers. C, D:Retinal organoids also
contain interneurons including PROX1-positive horizontal
cells and AP2◻-expressing amacrine cells separate from
photoreceptors expressing NeuroD1 and OTX2. Scale bars
equal 100 μm for (A) and 50 μm for (B–D).
The development and optimization of protocols that reproducibly generate

photoreceptors allows for the ability to obtain patient-specific sources as well
as CRISPR/Cas9 engineered hPSC disease models for outer retinal disease.
This type of modeling for photoreceptor degeneration was first demonstrated
by Jin et al., who examined five mutations causative for RP from patient-
specific sources (109). Results of these studies demonstrated the appearance
of oxidative and (endoplasmic reticulum) ER stress as well as varying
responses to vitamin E and recapitulated phenotypes observed in the clinic.
Additionally, this study utilized drug screening for RP mutations and
identified the therapeutic rescue of Rhodopsin-expressing rods from patients
with the RP9 mutation by administration of alpha-tocopherol. This study was
followed up a year later by a further examination of the RHO mutation from
iPSCs and displayed significant ER stress in rod cells and progressive
degeneration of rod cells in culture over time (110). In 2013, Tucker et al.
conducted a study of recessive RP through the USH2A mutation in patient-
specific iPSCs and demonstrated protein misfolding and ER stress associated
with photoreceptor precursor cells (98). In 2014, an iPSC line was created
from patient-specific cells bearing the E181K mutation in the RHO gene
known to cause RP (111). Results of this study were one of the first to use
adenoviral vector gene transfer to amend the mutation and study its
mechanisms in photoreceptor degeneration and identified decreased survival,
increased apoptosis, and higher ER stress in photoreceptor cells bearing the
mutation. The addition of selected compounds was also shown to increase
survival and decrease ER stress, thereby further validating the use of iPSC-
derived photoreceptors for the screening of therapeutic compounds.
In more recent studies, retinal organoids have been used as a platform for
RP disease modeling. Disease phenotypes associated with mutations in
RPGR, TRNT1, and PRPF have been identified in patient-specific sources
including deficits in photoreceptor structure and electrophysiologic responses
(112), altered actin polymerization (113), autophagy disruption and oxidative
stress (114), and cilia formation disruption and cellular stress (115). Deng et
al. also demonstrated the correction of the RPGR mutation using
CRISPR/Cas9 gene editing technology and rescued photoreceptor phenotypes
in retinal organoids, further facilitating the use of these structures for gene
editing and gene therapy as a potential therapeutic strategy (112).
LCA has also been modeled using hPSCs to reveal phenotypes and
photoreceptor degeneration. In 2014, two studies examined alternative
splicing of CEP290 known to be the most causative form of LCA. Results
from these studies demonstrated cilia defects in CEP290-associated LCA
(65,74) with the blockage of aberrant splicing, thus rescuing cilia function
(65). Furthermore, in 2017 an additional study performed by Shimada et al.
demonstrated a specificity of cilia defects only in photoreceptors of CEP290-
LCA-derived retinal organoids compared to the fibroblasts further
implicating the importance of cilia dysfunction in LCA disease progression
and allowing for a potential therapeutic target (116). hPSCs have become an
invaluable tool for modeling diseases affecting photoreceptors with the
ability to identify degenerative mechanisms, correct gene mutations, and
consider these cells as a platform for drug screening and therapeutic
intervention.
hPSC-Derived Photoreceptor Transplantation and

Future Considerations
The ability to derive photoreceptors from hPSCs in large and reproducible
quantities allows for the utilization of these cells for cellular replacement to
treat outer retinal diseases and restore vision. Although a variety of human
studies are ongoing for hPSC-derived RPE cellular replacement, limited
human studies exist for replacement of photoreceptors with numerous
challenges to be overcome before these types of studies can be conducted. To
date, the replacement of photoreceptors has primarily focused on
transplantation into animal models of outer retinal diseases with the first
demonstration of photoreceptor replacement into Crx−/− mice in 2009
(97,117). Since then, a multitude of other studies have published the ability to
differentiate and transplant populations of photoreceptors into rodent and
primate model systems, including the transplantation of purified
photoreceptor precursor cells or intact retinal sheets derived from retinal
organoids (118, 119, 120, 121, 122, 123). In 2018, Zhu et al. demonstrated
the ability to derive transplantable photoreceptors from a current good
manufacturing practice (GMP) human iPSC line with transplanted cells able
to integrate and mature in the host photoreceptor layer (122). This study
provided substantial evidence for the utilization of these types of cell lines for
future clinical applications and human cell replacement.
Many obstacles and challenges still remain for photoreceptor cell
replacement, including understanding the correct developmental time point
for transplantation, the purification of transplantable populations, mode of
delivery, and lastly host–donor material transfer. Previous studies have found
better outcomes for photoreceptor transplant when using postmitotic
photoreceptor precursors (124,125) that yield a challenge for identifying the
developmental counterpart when derived from hPSCs. Therefore, the
majority of studies have relied on the transplantation of late-stage retinal
progenitors derived from hPSCs as a useful source for photoreceptor
replacement. A further characterization of this important developmental
window in hPSC photoreceptor precursors is needed in order to optimize
purification strategies and to further enable success following transplantation.
Thus, the ability to purify populations of photoreceptors and their precursors
is essential for clinical application in the future.
One strategy for photoreceptor enrichment is the use of fluorescence
activated cell sorting (FACS)-sorting based on reporter lines. A variety of
reporter lines have been developed for both photoreceptor precursors and
differentiated photoreceptors that allow for the ability to use FACS to purify
populations within early stages of development (105) or after photoreceptor
commitment has been made (23,108,126, 127, 128). Although reporter lines
allow for FACs purification, this strategy can often be damaging to cells
especially when they are purified in a more differentiated state. Other gentler
approaches have been developed that include magnetic-activated cell sorting,
which utilizes microbeads to isolate cell types based on the expression of cell
surface markers. The utilization of identifiable surface markers for
photoreceptors including CD73, SSEA-1, CD26, CD133, and CD147 permits
the use of magnetic sorting to yield enriched populations of cells for
transplantation with minimal damage that occurs from the sorting (23,129).
This type of sorting can be easily adapted in a cGMP environment and scaled
up for clinical applications.
Other considerations for photoreceptor transplantation are the type of cell
delivery and location of delivery. Cell suspensions have been favored in
many previous studies due to the ability to transplant reproducible numbers
of cells, and the procedure to do so is minimally invasive. Retinal sheet
transplants derived from retinal organoids have the possibility for better graft
survival and the delivery of photoreceptors that are already polarized in a
tissue, although this type of transplantation requires a more invasive surgery
for delivery. A last major consideration for hPSC-photoreceptor transplant is
the concept of material transfer, which has been demonstrated by a number of
animal transplant studies (77,130,131). In order to properly determine
successful integration into the host retina, a clear distinction between donor
and host cells must be established in future studies, which could include
independent labeling of donor cytoplasm and genetic material (83).
Over the past two decades, the ability to derive photoreceptors from
hPSCs has quickly evolved with the ability for this technology to serve in a
multitude of purposes, including developmental modeling, disease modeling,
drug screening, and cellular replacement. Although many of these protocols
have been optimized to yield mature populations of photoreceptors, some
pitfalls remain within this model system and include the lack of supporting
factors like the macula and RPE within hPSC-derived retinal organoids and
the extended amount of time (150 to 200 days) and resources it takes to
differentiate photoreceptors with mature properties. In future studies,
emphasis is needed on the optimization of protocols to accelerate the
maturation of photoreceptors and decrease the time of differentiation to
increase the feasibility for using hPSC-derived photoreceptors for large-scale
disease modeling, drug screening, and cell replacement purposes.
Additionally, future studies should utilize GMP protocols and cell lines to
generate hPSC-derived photoreceptors and precursors with the hope of
moving efforts forward to human clinical trials for cell replacement in outer
retinal diseases.
RETINAL GANGLION CELLS

Functions and Diseases of Retinal Ganglion Cells in
the Retina
RGCs act as the output neurons of the retina that connect the eye to the brain
and allow for the transduction of visual signals (132). RGCs reside within the
inner layer of the retina adjacent to interneurons apically and astrocytes
basally, and their long axons extend out toward the optic nerve head. RGCs
transmit visual information through the formation of complex synapses with
interneurons. Light-induced electrical signals propagating down RGC-axons
through the optic nerve form connections in the brain with numerous
postsynaptic targets. The loss of this important connection can lead to a
number of blinding disorders with limited therapeutics available that target
the restoration of vision in these cases.
As the projection neuron of the retina, the RGC possesses many unique
characteristics and functions compared to other retinal neurons. Firstly, RGCs
acquire a unique morphology that is essential for their proper function with
elaborate dendritic arbors extending into the inner plexiform layer and
forming synapses with amacrine and bipolar cells where the electrical signal
from photoreceptors is passed (133,134). A single RGC axon then extends
outward from the retina. In order to reach the proper synaptic target in the
brain, RGC axons must extend very long distances and utilize intrinsic and
extrinsic cues to pathfind in the complex environment. The lateral geniculate
nucleus and the superior colliculus are the two primary targets of RGC axons
and are responsible for translating visual electrical signals into the image we
see (135).
RGCs are also an extremely diverse population of cells with numerous
subtypes identified to date. RGC subtypes are categorized based upon
morphology, molecular signatures, and functional characteristics with these
different cells all working collectively for visual formation (136,137). Many
of these subtypes have been discovered and characterized in rodent models
including direction-selective, alpha, and intrinsically photosensitive RGCs,
although the translatability of these subtypes into human and nonhuman
primate models remains elusive. In primates, the two main subtypes are
parasol and midget RGCs with these two subtypes primarily characterized
based on morphologic properties (138,139). Additionally, these subtypes
have been shown to exhibit differential susceptibility to injury and disease,
making the discovery of effective therapeutics difficult (140, 141, 142, 143).
Therefore, compared to other retinal cell types, the unique morphologic
features, long distance of travel, and diversity among RGCs pose many
challenges for therapeutic interventions for treating optic neuropathies.
Glaucoma is the most prevalent and irreversible blinding disorder, which
causes degeneration and death of RGCs, ultimately resulting in loss of vision
(144,145). Primary open-angle glaucoma (POAG) results in the degeneration
of RGCs in the presence of high intraocular pressure, and normal-tension
glaucoma (NTG) leads to RGC death without elevated pressure, although the
exact mechanisms leading to cell death in both scenarios may be highly
similar. In premature infants, glaucoma can occur in up to 25% to 30% of
patients from narrow angles and potentially developmental issues (146,147).
Congenital glaucoma is also a concern in the pediatric population. A number
of genetic risk factors have been discovered for glaucoma including the
myocilin and optineurin genes. In the clinic, glaucoma is most commonly
detected through elevated intraocular pressure, although this symptom varies
greatly among cases. As such, imaging of the optic nerve head and nerve
fiber layer provide a more reliable marker of disease progression. Early
detection and diagnosis of glaucomatous neurodegeneration are critical for
intervention as the RGCs will continue to degenerate over time and lead to
progressive loss of vision, with few treatment options available for late stages
of glaucoma.
Leber hereditary optic neuropathy (LHON) is an inherited optic
neuropathy, which also leads to the degeneration of RGCs. Contrary to
glaucoma, more insight is known into the pathogenesis of LHON and
includes mitochondrial and metabolic dysfunction (148). In the clinic, LHON
is usually identified by central vision loss in one eye, followed subsequently
by the other eye within months. The disease is caused by mutations in
mitochondrial DNA, which are associated with components of the oxidative
phosphorylation system (OXPHOS) pathway and electron transport chain
including NADH dehydrogenase 1, 4, and 6. LHON is inherited through
mitochondrial DNA, which is passed on to children through the mother's egg.
Males are predominantly affected by LHON with symptoms beginning in
early adulthood. Mutations in these genes lead to mitochondrial morphologic
deficits, abnormal reactive oxygen species production, and energy and
mitochondrial respiratory dysfunction, with RGCs particularly susceptible to
these phenotypes.
Dominant optic atrophy (DOA) is an inherited optic neuropathy primarily
characterized by the degeneration of the RGC axons along the optic nerve
leading to vision loss. DOA is commonly diagnosed with loss of central
vision and optic disk atrophy (149). Mutations in the mitochondrial
membrane proteins OPA1 and OPA3 have been associated with the onset of
DOA with these genes playing a major role in mitochondrial homeostasis,
apoptosis, and energy metabolism. As such, RGCs may be preferentially
susceptible to mitochondrial dysfunction and lead to optic nerve degeneration
and loss of cells in the retina. Each of these diseases affects the RGCs and
leads to irreversible blindness with no treatments to reverse vision loss and
very few therapeutic options to slow the degeneration. hPSCs can be used as
a valuable tool to generate RGCs for developmental studies and future cell
replacement strategies, as well as model disease mechanisms and
pathogenesis when derived from patient-specific sources.
Utilization of hPSCs for RGC Differentiation, Disease

Modeling, and Cell Replacement
In comparison to RPE and photoreceptors, RGCs are the most understudied
when derived from hPSCs with only a handful of studies focused on the
development of these cells in vitro and their degeneration in optic
neuropathies (Table 7-1). Nevertheless, in the past decade, a number of
foundational studies have been published that describe the differentiation of
RGCs from hPSCs and how they can be utilized for disease modeling, drug
screening, and even some preliminary cell replacement strategies. During
retinal specification, RGCs are among the first cells to become specified,
allowing for a shorter timeline of differentiation compared to later-born cell
types, such as photoreceptors and bipolar cells. A number of protocols have
been published that describe the differentiation of RGCs from hPSCs using
both chemically defined media (24, 25, 26, 27,150) and default specification
of a retinal lineage (59,151, 152, 153). When derived from hPSCs, RGCs
express a multitude of RGC-associated markers including BRN3, SNCG, and
RBPMS and extend MAP2-positive neurites (Figure 7-7). In 2015, the
generation of an mCherry fluorescent reporter cell line for BRN3B allowed
for the ability to identify RGCs in live cultures to study their temporal
development (28). This study was followed up in 2017 with the development
of a tdTomato and Thy1.2 dual reporter system to enrich for RGCs. The
ability to identify and enrich for RGCs in live cultures allows for more in-
depth studies of RGC-associated phenotypes and characteristics (29).
FIGURE 7-7 RGCs derived from hPSCs express
characteristic markers and display elaborate neurite
morphologies. A:Bright field imaging revealed the
neuronal morphology of RGCs with large cell somas
interconnected by numerous processes. B. C:hPSC-
derived RGC neurite morphologies were confirmed by the
expression of MAP2 with RGC-specific transcription
factor BRN3 as well as expression of RGC-associated
protein SNCG. Scale bars equal 100 μm for (A) and 25 μm
for (B) and (C).
In order for hPSC-derived RGCs to be used as a reliable model, these cells

should possess some of the unique characteristics of RGCs including
elaborate dendritic and axonal morphologies, functionality, and diversity.
Previous studies have demonstrated the ability for hPSC-derived RGCs to
extend long, elaborate neurites (26,28, 29, 30,150,154) with the ability for
these neurites to identify appropriate brain targets (155). Additionally, similar
to what is seen in the human retina, hPSC-derived RGCs demonstrate
diversity in gene expression that could be contributed to the presence of RGC
subtypes (31,156). As RGCs are the projection neurons of the retina, they
need to be functionally active in order to transmit visual information to the
brain. RGC functionality has been recapitulated in a number of studies with
the ability for hPSC-derived RGCs to conduct inward sodium currents and
fire action potentials demonstrated in a number of studies (25, 26, 27,
28,150,154).
Efforts in more recent years have been aimed at accelerating and
enhancing these characteristics to yield a cell type similar to what is found in
vivo. A study by Yang et al. observed the extension of long and organized
axons when grown on a nano-imprinted scaffold (157). Another study
published in the same year seeded hPSC-derived RGCs on biodegradable
poly scaffold (PLGA) and observed the formation of dendritic arbors and
functional axons (32). A 2019 study by VanderWall et al. described the
temporal maturation of hPSC-derived RGC neurites and electrophysiologic
characteristics (158), particularly when grown in association with astrocytes.
In the retina, astrocytes are found in close association with the RGCs in the
nerve fiber layer and play a major role in neuronal homeostasis and
maturation. This study observed the acceleration and enhancement of
morphologic characteristics and RGC excitability. The derivation hPSC-
derived RGCs that exhibit a full complement of in vivo characteristics creates
a more reliable model, which can further be used to identify disease
phenotypes when derived from patient-specific sources.
hPSC-derived RGCs have been used as a model for forms of glaucoma
over the past decade. In particular, the E50K mutation in the optineurin
(OPTN) protein has been characterized experimentally with hPSCs with this
mutation leading to protein insolubility (159,160), increased apoptosis
(154,159), Golgi deficits (154), and autophagy dysfunction (159). In these
studies, some of the deficits identified in RGCs were rescued by treatment
with growth factors (154), timonol (159), or BX795 (160). The TBK1
mutation has also been linked to NTG, and in a different study duplication of
this gene resulted in increased LC3II protein in the mutant RGCs (161),
indicative of autophagy disruption. SIX6 is an important retinal development
gene and has also been linked as a risk allele for glaucoma. In a 2017 study,
patient-specific hPSCs harboring the risk allele in SIX6 were differentiated to
a retinal fate and exhibited a lower number of RGCs, deficits in neurite
morphology, and decreased functional capabilities suggesting that this risk
allele plays an important role in RGC development and maturation and could
lead to glaucoma phenotypes (162).
Other studies have utilized patient specific-hPSCs to model disease
phenotypes of LHON (163,164) with RGCs exhibiting neurite deficits,
abnormal respiration, increased oxidative stress, and deficits in
electrophysiologic activity. In 2017 Wong et al. described the ability to
rescue cell death in LHON-diseased RGCs by the cytoplasmic hybrid
(cybrid) correction of mitochondrial DNA, providing a novel technology for
examining disease mechanisms in LHON (165). Lastly, DOA was modeled
in 2016 by using hPSCs derived from patients with an OPA1 mutation.
hPSC-derived RGCs harboring this mutation exhibited increase activation of
apoptosis and developmental abnormalities including misshapen EBs and the
inability to form neural rosettes., Following the identification of these
phenotypes, the addition of Noggin or beta-estrogen promoted RGC
differentiation and survival (166). Thus, hPSCs provide an advantageous
model for RGC development and disease with the ability to identify disease
phenotypes in a number of hPSC-modeled optic neuropathies and the use of
these cells for drug screening purposes to find novel therapeutic strategies to
aid in disease progression.
Optic neuropathies ultimately lead to the loss of RGCs, which do not
have the capacity to regenerate in vivo. Therefore, novel treatments and
therapeutics need to be developed to reverse this loss or replace the damaged
cells with the hope of restoring vision. In a recent study, Teotia et al.
examined the role for the mTOR pathway in RGC differentiation and neurite
outgrowth and guidance (167). Using an in vitro axotomy model using
microfluidics, hPSC-RGC axons were able to regenerate with the mTOR
pathway playing a significant role in this process and providing a potential
therapeutic target for degenerating RGCs in optic neuropathies.
Cell replacement therapy is also an attractive option for treating optic
neuropathies by replacing degenerated RGCs with hPSC-derived RGCs.
These studies are in very preliminary stages due to the complexity and unique
characteristics of RGCs, such as the ability to integrate into the RGC layer,
the formation of synaptic connections, and the extension of long axons out of
the eye to synapse with brain targets. In 2017, Li et al. described the
maturation of hPSC-derived RGCs on an FDA-approved biodegradable
scaffold and the transplantation of this scaffold into rabbit and rhesus monkey
eyes (32). The GFP-expressing scaffold and cells were detected on the retinal
surface and remained up to 3 months without adverse effects or rejection,
although the quality and integration of hPSC-RGCs into the retina were not
extensively explored. In a more recent study, Wang et al. transplanted hPSC-
derived retinal progenitors into NDMA-treated mice and demonstrated the
ability for these cells to integrate, extend neurites, and express mature RGC-
associated markers (168). Some considerations for future hPSC-derived RGC
transplant studies are the proper age and differentiation stage to transplant
cells, the purity of transplanted cells, and the development of assays to
understand proper maturation and integration into host retinas. Nevertheless,
significant progress is being made in preliminary studies for the use of hPSC-
derived RGCs for a potential treatment for optic neuropathies like glaucoma,
LHON, and DOA.
CONCLUSIONS
hPSC technology has revolutionized the field of developmental retinal
biology with access to some the earliest embryonic time points and the ability
to differentiate these cells into RPE, photoreceptors, and RGCs. More so,
when derived from patient-specific sources, hPSCs can be used to model
retinal diseases and understand disease mechanisms causing the degeneration
of retinal cell types and to screen drugs and therapeutic compounds targeted
at affected pathways. Lastly, hPSCs provide an attractive source for cell
replacement strategies with the transplant of hPSC-derived RPE already in
human clinical trials and hPSC-derived photoreceptors and RGCs in more
preliminary stages of transplantation. Progress in these areas is continually
being made with the overarching goal of developing a treatment for blindness
caused by retinal degeneration.
ACKNOWLEDGMENTS
Grant support was provided by the National Eye Institute (R01 EY024984
and R21 EY031120 to JSM), the Indiana Department of Health Spinal Cord
and Brain Injury Research Fund (JSM), an IUPUI University fellowship
(KCH), and startup funds from the Indiana University School of Medicine.
This publication was also supported by a fellowship from the Indiana Clinical
and Translational Sciences Institute (KBV) made possible with partial
support from UL1TR002529 (A. Shekhar, PI) from the National Institutes of
Health, National Center for Advancing Translational Sciences, Clinical and
Translational Sciences Award.
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8
Applications for Human Pluripotent
Stem Cell–Derived Retinal Cells in
Development, Disease and Cellular
Replacement
Kirstin B. VanderWall, Clarisse M. Fligor, Sailee S. Lavekar,
Kang-Chieh Huang and Jason S. Meyer
INTRODUCTION
The discovery of human pluripotent stem cells (hPSCs), including both
human embryonic stem cells (hESCs) in 1998 (1) and human-induced
pluripotent stem cells (hiPSCs) in 2007 (2,3), offered the possibility to
overcome the limited amount of donor retinal tissue and provided exciting
new opportunities to study the earliest stages of human retinogenesis. The
ability to obtain cells from patient sources allows for in vitro disease
modeling applications and makes it possible to study how genetic makeup
influences biologic responses. Indeed, even within the realm of human tissue,
a particular individual's genetic background can influence disease progression
or drug response, making hPSCs ideal candidates for modeling inherited
degenerative disorders of known genetic origin.
The emergence of hPSC-based protocols for retinal differentiation that
recapitulate major molecular and cellular events of human retinogenesis in
vitro has opened up immense research possibilities, including developmental
studies and disease modeling as well as generation of clinically relevant cells.
To date, a number of studies have focused upon the ability to direct the
differentiation of hPSCs to retinal cell types (4, 5, 6, 7, 8, 9), although these
efforts have mostly utilized stochastic methods of hPSC differentiation with
retinal cells heterogeneously arranged. These retinal cells lack the
organization and maturation typically observed in the retina, which limits
their ability to model retinal development as well as retinal disease pathology.
To address this lack of organization, recent efforts have focused on the
differentiation of these cells as three-dimensional (3D) retinal organoids
where differentiation progresses in a stepwise manner analogous to early
stages of retinal development (10, 11, 12, 13, 14, 15). A remarkable
advantage of retinal organoids is the ability to study early human retinal cell
fate decisions including how these cells interact to generate an organized
retinal-like tissue (6,16,17). Also relevant is the possibility to use hPSC-
derived organoids as a tool to model retinal disease progression in vitro with
studies to date focusing upon how cells of the outer retina are affected in
disease states (16,18, 19, 20, 21, 22).
2D VERSUS 3D CULTURE
Differentiation of hPSCs provides a renewable and reliable source of cells to
generate any cell type of the body from hESCs (1) and hiPSCs (2). Over the
years, various groups have developed multiple retinal differentiation
protocols with some utilizing a step-wise protocol (5,6,10,11,13,14,23) that
mimic the major stages of human retinogenesis; other groups have directly
reprogrammed pluripotent stem cells to their final fate (9,24). Some of these
standard protocols utilized two-dimensional (2D) cell culture approaches to
produce retinal neurons and have contributed to multiple significant
translational milestones (25,26). hPSC-derived neurons observed in 2D
cultures display increased activity compared to those in 3D cultures (27) in
part due to increased density, axonal outgrowth, and lack of regulating
signals present in 3D cultures. Furthermore, 2D culture methods are often
less expensive, less complicated to use, and allow for easier downstream
processing. However, 2D models are less biologically relevant as they do not
recapitulate the complex 3D microenvironment of living tissue (17).
Moreover, 2D-based methods of retinal differentiation are unable to produce
all of the structural components of retinal cells, such as vital cell–cell and/or
cell–matrix interactions, making it challenging to recapitulate development
and disease in a dish. Organoid technology is a fast-growing platform used to
study various aspects of human biology. With the discovery of organoid
technology, publications using retinal organoids have significantly increased
in just the past few years (Figure 8-1), indicating the rapid adoption of these
differentiation approaches as well as how the scientific community sees these
organoids as valuable tools for research.
FIGURE 8-1 Number of retinal organoid publications

over the years. Since the discovery of retinal organoids in
2011, there has been a large increase in the number of
publications referencing retinal organoids. This trend is a
reflection of the increasing popularity and utility of retinal
organoids for scientific research.
Organoids Mimic the Spatial and Temporal

Development of the Human Retina
Under physiologic conditions, cells reside in a 3D environment and interact
with other cells and the extracellular matrix (28,29) (ECM). These
interactions are necessary for the proper differentiation, spatial orientation,
and functionality of these cells. The ability to recapitulate human
retinogenesis in a dish provides the opportunity to test hypotheses which
were previously limited to animal models, due to restricted availability and
ethical concerns associated with the use of fetal tissue. While animals are
invaluable assets to all forms of research, human cells differ genetically and
biologically compared to common laboratory animals or even other primates
(5,10,30). Due to these variances, diseases may present themselves
differently, and drug treatments may produce variable effects. Retinal
organoids are 3D retinal tissues grown in vitro that mimic the development,
anatomy, and physiology of the retina in vivo (19,30, 31, 32, 33). In recent
years, multiple publications have demonstrated that 3D retinal organoids can
be produced in a stepwise manner using hPSCs (17,20,22,29,31,34, 35, 36,
37, 38, 39, 40, 41, 42). Access to culture systems that generate organoids,
including retinal organoids, has increased the availability of human tissue for
basic and clinical research. Retinal organoids have a variety of uses, serving
as models of human retinogenesis (17,29,36,42,43) and disease mechanisms
(20, 21, 22,30,44) as well as a source of cells for drug screening (19) and cell
replacement therapies (30,32,33,35,38,44, 45, 46).
These floating cultures of retinal structures allow for the initial
differentiation of retinal progenitor cells (RPCs) into all retinal cell types,
following a sequential manner consistent with in vivo vertebrate retinogenesis
(4,6,14,16,17,34). The ensuing retinal cells self-assemble into a tissue-like
structure that recapitulates the architecture and patterning of the retina. As
such, differentiation within these organoids mimics the developmental timing
and organization of the retina, facilitating their use as models of retinogenesis
and disease progression. Furthermore, organoids allow for direct cell–cell and
cell–matrix interactions within a heterogeneous cell population. This
interaction produces cells with physiologic responses reminiscent of human
biology (11,39,40) and that can recapitulate development and disease
pathology in a way that 2D cell cultures and animal models have been
previously unable to achieve.
Similar to the developing human fetal retina, retinal organoids contain
proliferating/migrating RPCs, retinal pigmented epithelium (RPE),
photoreceptors (PRCs), second-order neurons (INL), and retinal ganglion
cells (RGCs). While BRN3+/Tuj1+ RGCs are found in the basal layers of
retinal organoids, photoreceptors begin to form a distinctly separate apical
layer, separated by CHX10/Ki67+ progenitors in the intermediate zone (36).
The order in which retinal cell types emerge from retinal organoids mirrors
the conserved cell birth order found during vertebrate retinogenesis (36).
RGCs can be detected earliest (25,36,47) followed by horizontal and
amacrine cells (19,41), whereas mature photoreceptors, particularly rod
photoreceptors expressing Rhodopsin, are generated at later stages, followed
by bipolar and Müller glial cells (11,35,48). Therefore, hPSCs are capable of
spontaneously generating stratified retinal tissue that is similar to human fetal
tissue containing a multitude of retinal cells in an appropriate spatially
arranged manner.
Organoids Provide Access to the Earliest Events of

Human Retinogenesis
Importantly, the formation of hPSC-derived optic cups is accomplished by
self-organization (10,23). Cells begin as an aggregate of hPSCs and over
several weeks, self-organize into a layered structure that develops in a
spatiotemporal sequence similar to that which occurs in vivo (17). By 3
weeks in culture, the distal portion of the optic vesicle-like structure begins to
invaginate to form a more optic cup-like structure. Furthermore, the
formation of the optic cup in vitro occurs in the absence of a lens or surface
ectodermal tissue, indicating that invagination is not solely a result of
external structures but rather occurs in a self-directed fashion. These 3D self-
organizing structures develop a cellular composition and architecture similar
to in vivo tissues, thereby replicating biologically relevant intercellular
signaling in vitro. Cell fate specification and maturation follows a sequence
and time course highly reminiscent of healthy retinal development (6,40).
The stepwise differentiation of hPSCs through all of the stages of
retinogenesis helps to ensure the proper differentiation and prospective
identification of retinal progenitors (11,13,16). Furthermore, this stepwise
differentiation allows organoids to effectively model human retinogenesis in
vitro, as researchers can be confident that the resulting neurons are of retinal
origin (49). The first phase in the stepwise production of a retinal phenotype
from undifferentiated pluripotent stem cells is the emergence of eye field
cells within the primitive anterior neuroepithelium. Retinal organoids are the
result of a step-wise differentiation of hPSCs to an anterior neuroectoderm
fate, indicated by the expression of eye field transcription factors (6,31).
Human ESC and iPSC-derived neuroepithelial aggregates express all eye
field transcription factors and the gene and protein expression profiles of the
progenitors forming the vesicular structures reflect a differentiation state of
retinal progenitors close to the optic vesicle stage of retinal development (13).
Following specification of the eye field, optic vesicle-like structures are
formed that further mature to an optic cup-like structure. Optic vesicle-like
structures isolated from hPSCs express early retinal markers which
subsequently produce mature retinal cell types, confirming their status as
RPCs (13). Upon reaching the optic cup-like stage of differentiation, many
cells have already acquired their final differentiated state and display
rudimentary lamination within the hPSC-retinal tissue (10,11,14).
Moving Toward 3D-Based Protocols

Key retinal developmental processes such as optic vesicle and optic cup
formation as well as signaling cascades have been reproduced using mouse
(50, 51, 52) and hESCs (10,12,13) (Figure 8-2). Organoid technology would
not be possible without the fundamental discoveries of previous researchers.
Beginning with the discovery of hESCs in 1998, the field has progressed over
the last 20 years (Figure 8-2). In 2009, the Gamm lab published the first
study generating 3D spheres enriched with RPCs (6), which provided a
foundation for the subsequent development of a retinal induction protocol for
hESCs based on a switch from initial 2D adherent to 3D free-floating culture
conditions (13). Initially, the study demonstrated that differentiation of
hPSCs to a retinal fate involved a progression from neuroectoderm, to eye
field, to optic vesicle, and finally to optic cup-like stages as shown by the
expression of key markers of each developmental stage in a time frame
similar to that of normal human development (13). Adaptation of the
stepwise 3D/2D/3D protocol of differentiation demonstrated that human
iPSC lines could efficiently generate retinal structures (11). In 2014, Zhong et
al. demonstrated the ability to generate 3D retinal organoids that contained all
major retinal cell types arranged into proper layers (11). This adaptation of
the Gamm protocol produced retinal organoids with outer segment-like discs,
capable of photosensitivity. Furthermore, initial reports on cell-based disease
modeling approaches revealed early-stage retinal organoids could be induced
from disease harboring-iPSCs just as effectively as from healthy iPSCs and
model disease-related phenotypes (16).
FIGURE 8-2 Timeline of important events in the

development of retinal organoids. Beginning with the
discovery of human embryonic stem cells in 1998 and
ending with specific cell types within retinal organoids in
2018, multiple keystone events have been fundamental in
developing retinal organoid technology.
Subsequently, in 2011, the Sasai lab reported the formation of a 3D retina

from mouse ESC (mESC) aggregates using a versatile floating culture in
serum-free and growth factor–reduced medium (SFEBq culture, or serum-
free culture of embryoid body-like aggregates with quick aggregation)
(51,52). In 2012, the Sasai lab applied this technique to hESCs and reported
the formation of a bilayered optic cup using reaggregated hESCs cultured as
SFEB in the presence of Matrigel and under high oxygen conditions (10).
This protocol for the generation of hPSC-derived retinal organoids allowed
for the formation of layered retinal structures that harbored high numbers of
photoreceptors beside several other retinal neurons and Müller glia. The Sasai
protocol was the first and only study reporting the formation of a double-
layered optic cup structure (10).
3D culture systems allow for the maintenance of retinal tissue in culture
for more extended periods than 2D systems, most likely because of the higher
degree of structural organization achieved, thus increasing the likelihood of
maturation in vitro (11,27,47,50,53). Moreover, 3D culture systems allow for
the generation of high numbers of clinically relevant retinal cell populations
for transplantation or high-throughput screening approaches (12,31). Retinal
organoids are a reliable tool for studying retinal development, disease
pathophysiology, and experimental cellular therapies. However, 3D culture
systems have added expenses and require more complex culture systems.
Therefore, there are still many obstacles that 3D cell culture needs to address,
such as high variability in differentiation protocols (42,54) and longevity of
the differentiation process (39,53) (up to 300 days) that impairs robust data
collection.
DEVELOPMENTAL MODELING
The eye is a complex organ that consists of many different cell types that are
organized in a spatially specific manner that allows for proper cell–cell
interaction and signal transduction (28,55,56). Recent studies have
demonstrated the differentiation of hPSCs into retinal organoids, which
permit the generation of all retinal cell types in a 3D organized structure
(42,54). Retinal organoids have proven useful as platforms to study different
aspects of development, especially the earlier stages of development that are
otherwise inaccessible to investigation due to limited availability of tissue or
difficulties to perform genetic manipulations. hPSC-derived retinal organoids
reproduce many aspects of embryonic retinal development, including the
formation of a bilayered optic cup (10) and light-detecting photoreceptors
(11,53,57). Retinal organoid neurons can migrate into their appropriate
relative positions, produce neurites, and colocalized synaptic proteins with
appropriate cellular targets (34,36,53).
Live Imaging of Retinal Organoids

Due to the translucent nature of retinal organoids, development and cell
migration can be visualized using live imaging techniques in vitro. Live
imaging of hPSC-derived retinal organoids revealed cells in the distal
epithelium that were highly proliferative and underwent interkinetic nuclear
migration (34). Conventional techniques used to monitor spatial and temporal
changes in structure and function have been limited in their utility due to the
necessary destruction of the tissue. Ideally, live imaging techniques would
enable for nondestructive observation of organoids (58). Researchers have
begun to develop these tools to benefit the rapidly growing field of organoids.
Studies have shown that structural and metabolic changes coordinated with
retinal differentiation in hPSC-derived retinal organoids can be revealed with
live imaging modalities (58,59).
Organoids have also been successfully visualized using powerful two-
photon microscopy approaches such as fluorescence lifetime imaging
microscopy (FLIM) and hyperspectral imaging (HSpec) (34). Both of these
techniques are able to identify and quantify endogenous fluorophores,
eliminating the need for genetic modification of cells to express exogenous
labels. New live imaging modalities detected increased glycolytic activity and
detected retinol and retinoic acid accumulation in the organoid outer layer,
coinciding with photoreceptor genesis (34). With these new techniques, it is
possible to observe the organization and development of retinal organoids by
repeated analysis of the same living organoids providing a more direct means
to characterize changes.
Furthermore, organoid development can be explored with high-resolution
optical coherence tomography (OCT), a technique that is commonly used in
clinical settings as a noninvasive means of imaging ophthalmic structures
(60). The ability to noninvasively monitor retinal organoids can be
accompanied by automated techniques to analyze changes within retinal
organoids. The 3D-ARQ system uses a microplate reading system to rapidly
and effectively quantify the relative fluorescence of reporters in organoid
model systems (59). Live hPSC-derived retinal organoids at different
developmental stages can be examined for microanatomic organization and
metabolic function. Beyond live imaging, techniques exist to image whole
fixed organoids allowing for visualization of specific structures via antibody
labeling (58).
Retinal organoids are mostly translucent, but tissue clearing protocols can
be used which enhance the efficiency of antibody labeling and depth of
imaging at high resolution (58). The combination of tissue clearing
technology and advanced optical sectioning methods such as light-sheet
microscopy provides more defined spatial information and allows for the
study of fine morphology such as photoreceptor ribbon synapses (61).
Enhanced visualization of 3D tissues provides a powerful tool for the
organoid research field, allowing for the detection of molecular structures to
the overall development and structure of the entire organoid.
Composition of Retinal Organoids

The ECM is known to play an important supportive role to the cellular
proliferation, differentiation, migration, guidance, and axonal growth of
neurons (28,36,47,62, 63, 64, 65). However, there has been little focus on
ECM distribution during retinogenesis. A recent study utilized developing
and adult retinas as well as hPSC-derived retinal organoids to study the
expression of crucial ECM components within the retina, with results
indicating the expression of basement membrane ECMs such as Fibronectin
and Collagen IV within Bruch's membrane and inner limiting membrane of
the developing human retina (29). The expression of photoreceptor-specific
ECMs interphotoreceptor matrix proteoglycan 1 (IMPG1), Versican, and
Brevican in the interphotoreceptor matrix (IPM) was conserved between
developing human retina and hPSC-derived retinal organoids (29).
Subsequently, hPSC-derived organoids were used to test the role and
importance of ECM components. Blocking of CD44 and IMPG1 affected
photoreceptor connecting cilia and inner/outer segment formation resulting in
disruption of IPM formation and photoreceptor development (29). Results of
the study demonstrated that ECM components display distinct distribution
patterns throughout the retina and play an essential role in the development of
specific cell types.
Recent rapid improvements in single-cell sequencing technology and
analyses create an opportunity for dissecting organoid cultures composed of
multiple cell types. Understanding the genetic composition of organoids
helps us to better understand tissue biology, cellular behavior, and
interactions. Single-cell RNA sequencing (scRNA-Seq) of retinal organoids
revealed the presence of multiple retinal cell types such as RPE, RGCs, rods
and cones, and Müller glia (40). Remaining clusters contained RPCs and cells
expressing genes involved in homeostasis. Sequencing of retinal organoids at
three different stages of development revealed a decrease in mitotic cells as
earlier born cell types such as RGCs and RPE were formed (40).
Subsequently, rod and cone photoreceptors emerged from photoreceptor
precursors with the number of later-born Müller glia increasing over time
(40). Pseudo-time analysis of organoid development resembled the birth
order of cell during in vivo retinal development, beginning with mitotic
progenitor cells and ending with the emergence of Müller glia (40).
Organoids to Study Cell Fate and Role of Key

Transcription Factors
Retinal organoids begin their development, expressing a complement of RPC
markers (13). These retinal progenitors undergo morphogenic changes by
first developing optic vesicle-like structures, followed by optic cups, and
finally, fully laminated retinas that result in the entire repertoire of retinal cell
types present in the normal retina (10,14,54). Initial studies of retinal
organoid formation began with mouse embryonic stem cells (mESCs) (51).
Results of these studies indicated that the formation of optic cups from
mESCs was most successful when starting cultures contained a high
percentage of RPCs (52). Applying this knowledge to the hPSC-derived
retinal organoid formation, Nakano et al. tested the effects of various
signaling modulators on retinal differentiation (10). Results indicated that the
presence of fetal bovine serum (FBS) in culture media was an effective
enhancer of retinal differentiation (10). Subsequently, in addition to FBS, the
use of a Hedgehog smoothened agonist (SAG) increased the proportion of
Rax-positive RPCs to >70% of total cells (10).
Previous reports have determined that self-organized optic-cup formation
occurs most frequently when the culture contains a high percentage of retinal
progenitors (6,10,66). Based on this research, several intrinsic temporal
factors are known to contribute to RPC competent states. For example, hiPSC
lines expressing the highest levels of DKK1 and to a lesser extent, NOGGIN
at day two also expressed the highest relative levels of anterior neural/eye
field genes at day 10 and VSX2+ OV-like structures at day 20 (6). This study
also reported that early endogenous expression of DKK1 and NOGGIN in
some hPSC lines was sufficient to induce the formation of OV-like structures
without the addition of small molecules antagonizing BMP/TGFB and Wnt
pathways (37). Therefore, hPSC-derived retinal organoids serve as platforms
to study intrinsic regulators of retinogenesis, beginning at developmental
time points inaccessible in humans.
Decades of intricate retinal research has led to a growing understanding
of gene regulatory networks and epigenetic factors contributing to retinal
differentiation (31). Developmental studies of hESC-derived retinal
organoids provide a foundation for studying the role of complex transcription
factors and their dynamic role in human development. Microphthalmia-
associated transcription factor (MITF) is expressed in optic vesicles during
development with direct transcriptional links to cell cycle, apoptosis, and
pigmentation (67). Manipulation of MITF RNA and protein levels in retinal
organoids at early stages of development resulted in decreased expression of
eye field transcription factors and reduced early optic vesicle cell
proliferation (67).
Both hESCs and hiPSCs are well suited to study effects of key neural
retina transcription factors on the differentiation of specific cell types.
However, hiPSCs possess the added benefit of harboring developmental gene
defects when differentiated from affected human patients. hiPSCs produce
retinal organoids in a predictable, stepwise fashion, similar to hESCs
(11,16,23,35). In order to study these developmental processes, Phillips et al.
generated patient-derived hiPSCs harboring a mutation in the transcription
factor Visual Systems Homeobox 2 (VSX2). Results of this study indicated a
conserved role of VSX2 in the maintenance and proliferation of neural RPCs
and the subsequent generation and development of bipolar cells and
photoreceptors (16).
Several studies have reported the derivation of similar mini-retina
structures from hPSCs under different culture conditions (10, 11,
12,17,35,48,53). Cumulatively, these previous results provide a new, highly
sophisticated picture of how a human cell acquires and maintains a specific
cell fate. However, most of these studies have focused on outer retinal cells
such as photoreceptors, with a lack of emphasis upon the development or
maintenance of other cell types within retinal organoids. Beyond the study of
human development, the generation of hPSC-derived organoids provides
opportunities to improve methods of specific retinal neuron production, with
important applications for developmental biology as well as in vitro disease
modeling.
RETINAL GANGLION CELLS

RGCs are the primary output neuron of the retina and consist of complex
dendritic arbors that receive input from preceding retinal circuitry and also
extend large-diameter axons out of the eye and into the brain (68, 69, 70, 71).
RGC axons are capable of transmitting signals to recipient areas, many
centimeters away from the eye (70,72). During pathfinding, RGCs respond to
environmental cues that repel axons from the periphery and direct their
growth toward the center of the retina (73,74). RGC axons converge at the
optic nerve head and project their axons out of the eye and into the optic
nerve (75, 76, 77). Subsequently, this bundle of thousands of axonal fibers
passes information to the next relay stations in the brain, mainly the lateral
geniculate nucleus of the thalamus, the suprachiasmatic nuclei, the superior
colliculi, and other pretectal nuclei (74,78). As the axons grow during
development, they form highly complex connections by pathfinding along
precise routes to ultimately recognize and form synapses with appropriate
target cells (76,79,80). Due to their complex nature, there has been a lack of
emphasis upon the development and organization of RGCs in retinal
organoids.
RGCs Are the First Cell Type Specified in Retinal

Organoids
The production of retinal neurons occurs in a highly conserved temporal
sequence in which RPCs differentiate into various cell types of the retina,
beginning with the differentiation of RGCs (26,81). This development is
conserved in vitro with retinogenesis beginning around week 5 with the
generation of Brn3+ RGCs with RBPMS+ cells becoming abundant at later
stages (11), consistent with BRN3 being a transcription factor required early
to specify RGC development and RBPMS functioning in a differentiating cell
(43,82). Over time, the number of RGCs gradually increases, and the
formation of a distinct RGC layer in the basal-most zone of organoids is
apparent by week 6 (26,36) (Figure 8-3). As the projection neurons of the
retina, RGCs express cytoskeletal markers such as TUJ1 and MAP2 and
RGCs can be identified by combinations of RGC associated markers (Figure
8-3). However, multiple studies have shown that the number of RGCs
decreases over time in 3D cultures (26,34,40).
FIGURE 8-3 Retinal ganglion cells within retinal
organoids. By 40 days in culture, retinal organoids have
self-assembled and begun to produce BRN3+ RGCs in the
innermost layers. These RGCs express multiple RGC-
associated markers that colocalize with BRN3 expression.
This decrease in RGCs is somewhat expected, given that during development

of the visual system, RGCs are generated in an overabundance and send their
axons to synaptic targets in order to ensure sufficient connections (83,84). As
these axons become more dependent on trophic support from postsynaptic
targets, they are subjected to two waves of programmed cell death in order to
eliminate extraneous connections, pruning down the number of axons and
refining the connections to only those receiving sufficient support (68,85). In
vitro, the number of RGCs initially increases over the first few months,
followed by a gradual decline in their numbers (43). Single-cell RNA-
sequencing of retinal organoids found that RGCs decreased from 21% at day
60 to 6.6% at 200 days (40). Ultimately, by 151 days in culture, BRN3+
RGCs are no longer visually detectable (34,53).
As the death of all RGCs within a few months does not typically occur in
vivo, questions have been posed to address this shortcoming of retinal
organoids. RGCs within these retinal organoids have nowhere for axons to
extend due to lack of a postsynaptic target, potentially leading to their loss
due to a lack of retrograde trophic support. Furthermore, organoid-derived
RGC axons lack vasculature, glial support cells, a proper inner limiting
membrane, optic nerve head, and optic tract. The lack of other retinal
structures raises the possibility that the observed decrease in RGC number
may be analogous to RGC apoptosis that occurs in vivo when RGCs are
unable to innervate appropriate targets (86,87). Bax-mediated apoptosis is a
phenomenon that has been demonstrated to be less severe in monolayer
cultures because these stem cell–derived RGCs form synaptically coupled
networks in 2D settings and presumably receive some degree of retrograde
signaling to enhance survival (83).
Beyond studies of RGC development, organoids may also prove useful
for cellular replacement therapies in which extensive axonal outgrowth is
necessary to reach postsynaptic targets. A recent study characterized RGC
differentiation throughout the earliest stages of organoid development, with a
clearly defined RGC layer developing in a temporally appropriate manner
and expressing a complement of RGC-associated markers (36). Significant
enhancement of neurite outgrowth was observed through modulation of both
substrate composition and growth factor signaling. Additionally, organoid-
derived RGCs exhibited diverse phenotypes, extended elaborate growth
cones, and expressed numerous guidance receptors (36). The production of
axons from retinal organoids (36,47,88) suggests that the retinal organoid
environment is providing appropriate factors to induce axon outgrowth, a
process which does not occur by default but instead requires trophic signaling
and is enhanced by electrical activity (89). Thus, retinal organoids can be
used as a valuable tool for studies of RGC development and provide an
effective platform to study factors influencing neurite outgrowth from
organoid-derived RGCs (30,47).
Identification of RGCs Within Retinal Organoids

In vivo, newly born RGCs possess intrinsic properties to help guide their
axons to proper brain targets (84,90,91); therefore, RGCs isolated from early
stages of development are more plastic and amenable to transplant (92,93).
Stem cell–derived RGCs develop in a distinct layer separate from the
photoreceptor layer and account for up to 30% of the cells in the culture,
which is similar to the standard amount of RGCs that are found in the retina,
depending on the stage of development (43,83,94,95). As current 3D
protocols result in a heterogeneous cell population with several retinal cell
types present, a purification step before subsequent applications may be
required in order to reduce contaminating cells (43,45). Techniques that have
existed for decades have proven useful for eliminating residual proliferative
cells and generating nearly pure cultures of stem cell–derived RGCs (26,96).
Enrichment of specific cell populations, such as RGCs, is accomplished by
exploiting RGC-specific expression of surface antigens such as CD90
(Thy1), CD184 (CXCR4), and CD171 (L1CAM) (25,43). Recently,
immunopanning protocols were successfully adapted to sequester RGCs from
retinal neurons and other cells present within retinal organoids using
magnetic activated cell sorting (MACS) (97). Before enrichment, RGCs only
accounted for 4.2% of cells within the differentiated culture (26). MACS
enrichment using Thy1.1 magnetic beads yielded about 77% Thy1.1+ cells
(26). The enriched population of retinal organoid-derived RGCs extended
intricate axons, displayed functional electrophysiologic properties, and was
capable of axonal transport of mitochondria, all suggestive indicators of
maturity (26)The emergence of retinal organoids has created a demand for
tools that facilitate the detection and monitoring of cells within organoids.
Generation of reporter cell lines using CRISPR technology can now be
combined with screening platforms that enable automated reporter
quantification within complex hPSC-derived retinal organoids. A study by
Sluch et al. utilized CRISPR engineering to generate an hPSC line with RFP-
labeled RGCs via a BRN3-linked promoter (25). Fluorescent labeling of
RGCs allowed for the identification of RGCs within organoids, and live
visualization of when and where RGCs are born (36). A subsequent study
further improved this cell line by added the surface expression of thy1.2,
allowing the RGCs to be immunopurified from organoids (98). A fluorescent
reporter in combination with a surface antigen allows for RGCs to be
enriched via MACS and/or FACS sorting.
Obstacles for RGC Replacement

The central nervous system is notorious for its lack of spontaneous
regenerative capabilities, and RGCs lose their intrinsic ability for
regeneration soon after birth (84,87,89,91,99,100). Therefore, once RGCs
begin to degenerate and die off, opportunities to recover vision loss involve
cell replacement therapies via transplantation or endogenous repair (101, 102,
103). Organoids provide a promising solution for lack of donor tissue for cell
replacement therapies. Stem cell–based transplantation has been studied
although the focus is most frequently on the replacement of RPE into humans
and photoreceptors into animal models. Fortunately, the emphasis on outer
retinal cell replacement has provided helpful insight into successful
translational applications that could potentially be used for other retinal
neurons, including RGCs. However, some obstacles will first need to be
addressed as RGCs are not abundant in stem cell–derived cultures, and RGC
axons must be able to travel an extensive distance in order to connect with
proper target nuclei in the brain.
Replacement of RGCs requires complex local wiring into the inner
plexiform layer, as well as the extension of axons into the optic nerve and to
appropriate brain areas, presenting a more formidable challenge compared to
the replacement of other retinal cell types. Although it may not be the most
efficient strategy, previous studies have suggested that RGC transplantation
is possible (92). This low efficiency has redirected the field in recent years
toward identifying significant physical barriers and how to circumvent them.
The delivery of RGCs into the vitreal space of a host is not the difficult part
of the task but, rather, the challenge begins after RGCs are transplanted as
they must survive and integrate into the host retina (104). However, the retina
contains physical barriers such as the nerve fiber layer and inner limiting
membrane. Beyond increasing integration efficiency, a thinning or disrupted
NFL may better model a diseased retina (83). Therefore, optimizing
transplants into an environment that mimics a diseased retina may be more
appropriate since humans who receive RGC transplants in the future will not
have a healthy retinal environment (83). Nevertheless, the composition of a
diseased retinal environment must be considered for successful
transplantation. Factors such as gliosis, remodeling of cytoarchitecture, and
neural retinal thinning at late stages of degeneration may impede the ability
of transplanted cells to restore the precise neural circuitry of the retina (83).
Retinal Organoids to Study Optic Neuropathies

Many diseases lead to the degeneration of the optic nerve and eventually,
RGC death (71,103,105). These optic neuropathies, which result in loss of
vision or blindness, include diseases such as glaucoma, Leber's hereditary
optic neuropathy, ischemic optic neuropathy, and optic neuritis. Glaucoma is
one of the more common optic neuropathies, and although the exact disease
mechanisms leading to glaucomatous neurodegeneration remain elusive, this
disease leads to progressive degeneration of RGCs and subsequent visual
field loss (106). Previous efforts to model glaucoma in a dish have not proven
successful, as high intraocular pressure (IOP) is challenging to mimic in
vitro. However, there are genetic forms of glaucoma that occur in the absence
of high IOP while still resulting in pronounced RGC degeneration (107, 108,
109). Mutations in the Optineurin (OPTN) gene have been extensively
documented to result in severe RGC degeneration associated with primary
open-angle glaucoma and, therefore, should provide an effective in vitro tool
for studies of underlying disease mechanisms (110, 111, 112, 113, 114, 115,
116). Previous studies have utilized fibroblasts from a patient with an E50K
mutation in the OPTN gene to generate a patient-specific hiPSC line (95).
RGCs differentiated from the E50K line exhibited increased apoptosis and
Golgi deficits compared to control cell lines (95).
As retinal organoid technology continues to develop and improve, the
focus can now shift toward more intricate studies of previously neglected
inner layer neurons. Future directions should focus on optimizing protocols to
generate more substantial quantities of RGCs as well as establishing reliable
methods for long-term survival of RGCs. A recent study described an
optimized cryopreservation method enabling vitrification of retinal organoids
(10,35). According to previous studies, the critical period for generating
robust retinal organoids is within the first 30 to 40 days (42). Retinal
organoids that appear healthy at this stage will continue growing into a
healthy-looking stratified organoid. At this time point, RGCs are already
present. Therefore, it would be useful to make stocks of frozen retinal
organoids at slightly earlier stages when RGC progenitors are present. After
thawing, organoids could be cultured short term to generate newly
differentiated RGCs. This method would make it possible to prepare stocks
of organoids on a large scale that can be kept frozen and delivered to multiple
institutions for clinical trials (30,32). However, replacing a patient's entire
retina with an hESC-derived retina is unrealistic at this point, as it is currently
too challenging to rewire optic nerves to the brain in adult mammals.
In order for hPSC-derived RGCs to be relevant in human clinics, it will
be essential to establish quality control standards that help to address cell line
and differentiation protocol variability (42,54). Furthermore, some
parameters will need to be established between the scientific community and
appropriate governmental bodies such as an agreement on what defines an
RGC (83). A fundamental requirement in the development of cell-based
therapies is the establishment of robust protocols that permit the derivation of
large numbers of donor cells from a renewable source that faithfully
recapitulate the characteristics of the endogenous cell types they are designed
to replace (12). Future studies should aim to improve upon yielding a higher
number of specific cell types, which will allow these methods to be scaled up
as a source of donor cells for translational purposes.
PHOTORECEPTORS
Photoreceptors are light sensitive cells that play a crucial role in retinal
functionality (5). The ability to detect and process light information requires
specialized and unique structures such as photoreceptor inner and outer
segments as well as synaptic end feet containing the ribbon synapse (12).
During human fetal development, the process of generating photoreceptors is
relatively slow with morphologically recognizable photoreceptors beginning
to appear around fetal week 10 at the foveal region (10,117). Over the next
15 weeks, differentiation gradually proceeds in the nonfoveal region, which
accounts for the majority of the retina (10). hPSC organoid-derived
photoreceptors can develop these fundamental processes that participate in
receiving and transmitting visual information (11,12,17,35,42,45,57). The
development and function of these structures occurs in a timely manner that
recapitulates what is known about human photoreceptor development.
Photoreceptor Development Within Retinal Organoids

Although we are now capable of generating bona fide retinal neurons from a
wide array of methods, most methods are still unable to generate large
numbers of specific retinal neurons other than rod photoreceptors (83). Rod
photoreceptors are responsible for vision in dim light and account for 95% of
photoreceptors in the human retina, while the remaining 5% of cone
photoreceptors are responsible for daylight vision (5,118). Therefore, the
abundance of rod photoreceptors should not be too surprising as rods greatly
outnumber other retinal neurons in human retinas in vivo, and organoids are
meant to model this human retinogenesis. However, cones are fundamental in
human vision and are concentrated in the macular region of the retina
associated with high visual acuity. Various subtypes of cones have been
identified with blue (S) cones developing first, followed by green (M) and red
(L) cones (118). After 70 days in culture, recoverin positive photoreceptors
can be found in the apical layer of retinal organoids (Figure 8-4). By 200
days in culture, outer segments are visible around the periphery of the
organoids (Figure 8-4). Furthermore, staining for various photopigments
reveals the presence of rhodopsin-positive rods, s-opsin–positive blue cones,
and M/L-positive green and red cones (Figure 8-4).
FIGURE 8-4 Photoreceptors within retinal organoids.
Earlier stages of retinal organoids (125 days) express
robust photoreceptor markers, which can be visualized in
the outer layers of retinal organoids. Subsequent later
stage differentiation (200 days) produces outer segment-
like structures around the periphery of the organoid.
Immunostaining reveals distinct populations of rod
(rhodopsin) and cone (opsin) photoreceptors within the
outer photoreceptor layer of retinal organoids.
Retinal organoid differentiation can be studied to determine the mechanisms

underlying specification of neuronal subtypes within the human nervous
system. Developmental studies revealed that thyroid hormone (TH) signaling
specifies cone subtypes in human retinal organoids (118). Initial low levels of
TH specify S cones, whereas later high TH signal produces L/M cones (118).
The ability to influence the ratio of cone subtypes is a promising step toward
developing retinal therapeutics and vision repair. Furthermore, studies such
as this validates organoids as a model to study the mechanisms of human
development. Moreover, in vitro models of retinal development are beneficial
as the external environment can be easily manipulated.
Multiple studies have reported generation of hPSC-derived retinal
organoids containing outer nuclear layer–like structures, such as rods and
cones bearing well-formed inner segments and connecting cilia, nascent outer
segments, and presynaptic structures (11,12,17,35,42,45,57). Culturing
hPSC-derived retinal organoids in suspension for up to 6 months revealed the
ability of retinal organoids to form cell layers, including photoreceptors with
outer disc-like protrusions and photosensitivity (11). EM analysis at week 27
showed the presence of extracellular membrane discs reminiscent of outer
segments (11). Supplementation of culture media with retinoic acid promoted
photoreceptor development and maturation, generating more rho+, S-opsin+,
and L/M-opsin+ photoreceptors that also expressed other proteins of the
phototransduction cascade (11). Expression of synaptophysin and vGlut1
between photoreceptors and bipolar cells indicated the formation of synaptic
connections within organoids (22). Long-term culture of retinal organoids
produced electrophysiologically active rod photoreceptor cell in iPSC-
derived 3D organoids (57). A proportion of hPSC-derived photoreceptors
display advanced levels of maturation by responding to cGMP stimulation,
corresponding with photosensitivity found in native photoreceptors
(35,57,119).
The generation of photoreceptors is a long and slow process that can take
hundreds of days to reach maturity (11,53,57). Unfortunately, long-term
cultures often lose the characteristic retinal organization (39). Loss of
organization could be related to incorrect proportions of inner nuclear
neurons as well as failure to establish correct synaptic connections, even
though some synaptic proteins have been reported to be expressed (11,53).
RPE cells were rarely shown to be organized as a monolayer apical to
photoreceptors with the exception being a subfraction of retinal organoids
generated with the Sasai protocol (10). Absence of RPE cells at the apical
side of retinal organoids might be in part accountable for the impaired
generation of proper lamination. Robust formation of fundamental structures
such as cilia and outer segments by hPSC-derived photoreceptors is a crucial
requirement for their utility in understanding human retinal development and
for disease modeling. Studies have reported photoreceptors bearing nascent
outer segments; however, the disc membranes within the outer segments were
not tightly stacked, possibly due to lack of close apposition to the RPE. A
recent protocol reported the generation of RPE spheroids and retinal
organoids simultaneously, reducing the cost of retinal cell preparation and
providing a novel research tool (37).
A recent study by Hallam and colleagues investigated the differentiation
capabilities of five different hiPSC lines, which revealed significant cell line
variability in efficient retinal organoid generation (57). Despite variabilities,
long-term culture (150+ days) yielded retinal organoids capable of generating
light responses that although immature, were comparable to the earliest light
responses recorded from neonatal mice, corresponding with the
developmental period of eye-opening (57). At this time in development,
retinal organoids from all cell lines exhibited well-formed outer nuclear
layers containing photoreceptors with inner segments, connecting cilium, and
outer like segments (57). Therefore, light-responsive retinal organoids can be
derived from careful differentiation of specific cell lines and can be generated
at a scale necessary for pharmacology and drug screening purposes.
Retinal Organoids to Model Diseases Affecting

Photoreceptors
A significant cause of blindness is the degeneration of photoreceptors due to
genetic diseases, including retinitis pigmentosa (RP) (21,22,44), Stargardt
disease (120), and Leber's congenital amaurosis (LCA) (18,20). These
inherited retinal dystrophies, in which photoreceptors die, can arise from
mutations in more than 200 identified genes and present high genetic
heterogeneity (121). Fortunately, hiPSC technology can be used to model
retinal diseases with known genetic factors. Skin fibroblasts from a patient
with a confirmed genetic mutation can be reprogrammed to pluripotency,
then differentiated to the desired cell type of interest. Several laboratories
have developed retinal organoids for disease modeling using patient-derived
iPSCs (7,16,18,20,22,30,120,122, 123, 124). As one of the most common
inherited forms of blindness, RP affects more than 1 million people
worldwide (21,22,44). Mutations in the tRNA Nucleotidyl Transferase
(TRNT1) gene have been linked to early-onset RP (124). To model the
disease in vitro, iPSC-derived retinal organoids were generated from patients
with molecularly confirmed TRNT1-associated RP. TRNT1 mutations caused
reduced levels of full-length TRNT1 protein and patient-derived retinal
organoids exhibited autophagy deficits, evidenced by LC3-II accumulation
and elevated levels of oxidative stress (124). Stem cell–based disease
modeling provides a platform for investigating the thousands of mutations
distributed across more than 100 genes that have been reported to cause RP.
Mutations in the RPGR gene can cause cone–rod dystrophy and have
been identified as one of the many genes associated with RP (21). The RPGR
gene is an important component in the centrosome–cilium interface of
photoreceptors (22). Proof-of-concept studies have recapitulated the
pathogenesis of RP using patient-specific organoids and achieved targeted
gene therapy of RPGR mutations in a dish (22). RPE and retinal organoids
were generated from iPSCs derived from four RP patients with variable
clinical severity. This study used CRISPR/cas9 gene editing technology to
correct a PRPF31 mutation in cells derived from a patient with very severe
RP. RP patient-specific organoids demonstrated developmental defects in cell
morphology and function in photoreceptors (22). RPGR correction led to the
repair of photoreceptor development and rescued transcription defects in
patient retinal organoids (22). Results supported the effectiveness of in situ
gene correction with the rescue of molecular and cellular phenotypes.
Photoreceptors use highly specialized cilia known as outer segments,
which are responsible for detecting light via tightly packed discs containing
the photopigment opsin. Recently, LCA models were created using patient
iPSC-derived optic cups originating from fibroblasts (18). LCA is an
inherited retinal dystrophy that causes missplicing and premature termination
of photoreceptors, resulting in childhood blindness (18). The cilia-related
gene found in the connecting cilium CEP290 has been implicated in this
disease, suggesting a role for CEP290 in cilium assembly (18). Although
cells with a CEP290 mutation were able to form retinal organoids, LCA
organoids displayed abnormal CEP290 splicing and cilia defects (20). Recent
advances in iPSC technology have provided a platform for investing the
effects of patient mutations on neural retinal development and maintenance.
Furthermore, patient-derived organoids can be used to probe disease
mechanisms and test therapeutic compounds. Treatment of diseased retinal
organoids with an antisense morpholino restored normal splicing, increased
CEP290 protein, rescued ciliation, and improved the recruitment of essential
photoreceptor proteins, such as RPGR (18). Mutations in CEP290 have been
implicated in other diseases such as Joubert syndrome–related disorders
(JSRD). A similar study by Shimada et al. used patient-derived retinal
organoids with distinct CEP290 mutations to show a concordance between
ciliary defects and clinical severity of LCA and Joubert syndrome (20).
Studies such as this highlight the value of iPSC-derived retinal organoids in
the study of photoreceptor development and function.
Isolation and Transplantation of Organoid-Derived

Photoreceptors
Patient-derived retinal organoids allow for the interrogation of disease
mechanisms and testing of potential therapies for degenerative blinding
disorders. The introduction of 3D organoid technology dramatically
improved the availability of human retinal tissue and human photoreceptors.
Original protocols developed with 2D differentiation systems had difficulty
producing significant quantities of photoreceptors (5,27). However, retinal
organoids are ideal for cell replacement therapy studies as they efficiently
derive sufficient numbers of integration-competent cells (12,35,38,44, 45,
46), which remains a significant limitation for regenerative medicine to date
(32,83). Based on pioneer stem cell–based clinical trials for the treatment of
retinal degenerative diseases (125), there is a significant interest in bringing
retinal organoid-derived photoreceptors toward clinical applications.
Organoid differentiation protocols recapitulate human photoreceptor
development and allow for the isolation and transplantation of a pure
population of stage-matched cones (12,45). The generation of a fluorescent
cone–rod homeobox (Crx) reporter hPSC cell line using CRISPR/Cas9
technology confirmed that the cell surface antigen CD73 is exclusively
expressed in photoreceptors (45). CD73 is a marker of photoreceptor
precursors and continues to be expressed in differentiated photoreceptors
(35). Floating cultures of isolated structures enabled differentiation of RPCs
into all types of retinal cells in sequential overlapping order with the
generation of transplantation compatible CD73+ photoreceptor precursors
that can be safely transplanted into a host retina (35,45).
Long-term culture conditions allow the maintenance of both mature rods
and cones in retinal organoids until 280 days with specific photoreceptor
ultrastructures (35). Moreover, hiPSC-derived retinal organoids and
dissociated retinal cells can be easily cryopreserved while retaining their
phenotypic characteristics and the preservation of CD73+ photoreceptor
precursors (35). Cryopreservation of the whole retinal organoids
demonstrated that in freeze-thawed structures, the CD73+ photoreceptor
population is preserved (35). Therefore, it could be advantageous to create
stocks of retinal organoids at later stages of development in which the
organoids are just mature enough to possess a large number of photoreceptor
precursors. Similar to immunopanning, methods such as MACS utilize
antibodies contained within magnetic beads to identify cell types of interest.
The magnetic antibodies can be added to a dissociated mix of heterogeneous
cells and run through a magnetic column. The cell type of interest will be
captured within the column, while other cell types simply pass through. As
CD73 is present on the cell surface of photoreceptors, techniques such as
MACS can be used to efficiently isolate CD73+ photoreceptors from hPSC-
derived retinal organoids (45). Moreover, MACS can be performed on
cryopreserved organoids and allow organoids to be stored at varying stages of
differentiation for downstream applications. Preservation of retinal organoids
at specific stages should make it possible to prepare large-scale stocks to
deliver pure transplantation competent cells when needed.
Transplanting Retinal Organoid-Derived
Photoreceptors
Replacement of lost photoreceptors by transplantation represents one of the
few options for the reversal of end-stage degeneration (19,30,32,44,120,126).
The primary challenge of cell replacement is to successfully transplant and
functionally integrate cells that have the capacity to develop into mature
photoreceptors within dysfunctional retinal tissue (30,104). Transplants of
purified human long/medium (L/M) cones have shown survival and
incorporation into the adult mouse retina (12). Although studies have yet to
prove functional integration, the potential use of photoreceptor
transplantation still remains as a treatment for retinal degeneration. To
address this, studies have investigated the feasibility of replacing lost
photoreceptors in a mouse model of advanced degeneration.
A purified population of L/M Opsin+ cones was transplanted into the
Aip1−/− mouse model of LCA between 8 and 12 weeks of age when almost
all of the photoreceptors have died (12). Following transplantation, purified
photoreceptors formed a synapse-like structure in close association to host
interneurons and expressed photopigments, despite being transplanted into a
mouse model of end-stage retinal disease (12). Although mouse models are a
necessary and straightforward tool for the generation of cell replacement
therapies, they are much smaller than humans and do not contain the same
structural anatomy of the human retina (127). The use of larger animal
models such as nonhuman primates will be necessary to determine if
transplanted hPSC-derived photoreceptors can transmit visual information.
Although rescue of the retina may be more productive during early retinal
degeneration, there is a lack of reliable early degenerative markers, and many
patients do not seek treatment until irreversible damage has been done.
Currently, at later stages of degeneration, the restoration of photoreceptors is
being pursued via transplantation of dissociated cells (12,45) or
transplantation of hESC-derived retinal “sheets” (38,46). Because almost all
work on cell replacement in the retina has been done using dissociated cells
rather than intact retinal tissue grafts, the primary emphasis of retinal cell
replacement work has been focused on facilitating cell migration into the
recipient retina. There is no need for cell migration when a 3D retinal tissue
graft is deposited into the subretinal space. However, axonal migration is
needed to enable synaptic integration of the donor retinal tissue into the
synaptic circuitry of the recipient retina with retinal degeneration. The
transplantation of retinal sheets may be more appropriate and effective at
later stages of degeneration (128). Transplantation of hESC-derived retinal
sheets into primate models of injury-induced focal retinal degeneration also
led to the survival of donor cells with photoreceptors organized in clusters
expressing rod and cone proteins, as well as some synaptic markers (129).
The lack of host photoreceptors present during advanced stages of retinal
degeneration may improve the incidence of structured outer segment
formation and long-term cell survival.
While recent years have seen many advances in retinal organoid
technology with a significant focus on the differentiation and maturation of
photoreceptor cells, there are still many challenges that remain that will need
to be addressed in future studies. For example, hPSC-derived photoreceptors
do not fully form light-detecting outer segments (31,32,57). Moreover, the
size of a 3D retinal tissue derived from hPSCs is much smaller (~0.5 mm
length or less), and the photoreceptor density is lower than that in the human
fetal retina (31). Improving transport of nutrients and oxygen by growing
retinal organoids in higher oxygen conditions and/or in rotating-wall vessel
bioreactors may improve retinal organoid technology. Previous studies
indicate that use of bioreactors in retinal organoid culture results in improved
lamination as well as an increase in the yield of photoreceptors bearing cilia
and nascent outer segment like structures (39). The adaptation of organoid
production using bioreactors might allow scaling up the generation of stem
cell–derived PRCs, a demand that will have to be met soon given the
requirement of high cell numbers for cell-based therapies or drug screening
(31).
TRANSLATIONAL APPLICATIONS
The retina is an attractive model to study the central nervous system and has
been studied at the cellular level in excruciating detail for over a century (83).
Due to its location, it is easily accessible with minimal surgery and can be
imaged in vivo. However, there are multiple diseases and injuries that affect
the retina, resulting in loss of vision and/or blindness (19,30,32,33,120,125).
Current therapeutic strategies can slow disease progression, but new
strategies which aim to preserve and restore vision are needed to cure these
disorders. However, many drug discovery platforms rely on simplified 2D
culture systems that do not accurately model in vivo cellular architecture and
physiology (27). With the introduction of hPSC-derived retinal organoids, a
vital tool for generating retinal tissue in vitro has been established and is now
widely used in numerous laboratories worldwide (32,33).
The Age of Organoids

Recent years have witnessed a significant milestone as hPSCs were used to
generate self-organizing 3D structures with properties remarkably similar to
the developing human retina (10,13). With these methodologies, cells pass
through the same developmental stages as cells differentiating in vivo, and
cells develop in the context of a tissue that preserves cell–cell contacts and
intercellular signaling of retinal cells in vivo. All of these factors increase the
likelihood that the molecular nature of the cells will appropriately model
cellular behavior, whether used to model diseases, generate regenerative
therapeutics, study human retina development, screen drug compounds, or
assess toxicity (19,33). Thus, the ability to generate 3D retinal tissue is
providing previously unachievable opportunities to model human retinal
development and disease and has enhanced the probability that stem cell–
derived tissues and cells will contribute to effective treatment of eye disease
(Figure 8-5).
FIGURE 8-5 Translational applications of retinal
organoids. Retinal organoids have many uses for
translational applications such as modeling degenerative
diseases, replacing lost or damaged retinal cells, and high
throughput drug screening for the treatment of retinal
disease and injury.
The use of culture systems based on 3D organoids has significantly improved

the availability of in vitro models for studying differentiation and maturation
of specific target tissues and cells. Notably, by taking advantage of hPSCs,
new opportunities have arisen to investigate human-related development and
disease besides providing sufficient amounts of human cells for clinical
applications (32,130,131). hPSC-derived retinal organoids could permit the
diagnosis of early disease phenotypes, predict disease severity, or optimize
treatments for slowly progressing diseases (132,133). Additionally, hPSC
technology allows for the generation of patient-specific cells that would be
ideal to avoid immune rejection of transplanted cells (134). Furthermore,
cells can be genetically modified using gene-editing tools such as
CRISPR/Cas9 to remove disease-causing mutations (22). Compared with
nonhuman animal models, patient-specific organoids may better recapitulate
some aspects of disease phenotypes and, therefore, could be useful for
studying the mechanisms involved in human disease, as well as in drug
discovery (22).
Bringing Organoids Into the Clinic

The advent of hPSC culture systems to generate 3D retinal organoids
provides new opportunities to improve drug development pipelines (30).
Retinal organoids can be used to speed up the process and increase the
efficiency of transitioning potential therapeutic agents from bench to bedside.
Although organoid technology is not developed enough to replace current
translational paradigms, organoids provide a physiologically relevant human-
based model with the flexibility and level of control of an in vitro system
(19,30,33). Therefore, the addition of retinal organoids, rather than
replacement of existing pathway components, may provide a beneficial
augmentation that harnesses the advantages of this improved pathologic
modeling. In this role, organoids can serve as an accessory to improve
primary drug screening, reduce the number of expensive animal studies, and
provide valuable insight prior to human trials (30).
Cell replacement using hPSC-derived cells is one of several promising
approaches being pursued to restore vision and improve quality of life
(32,44,83,126). In order for cell replacement to be a viable therapy to treat
retinal degenerative diseases, cells must come from an efficient source, be
able to survive for long periods and mature following transplantation, connect
with existing retinal circuitry, and ultimately restore visual function (135).
Significant developments within the last decade have brought therapeutic cell
replacement closer to clinical application with retinal organoid technology
now representing the primary tool in these efforts (19,30,32). Whether retinal
organoid-derived cells are favorable to generate transplantable donor cells for
clinical use will depend on several stipulations.
For medical applications, therapeutic retinal cells must be generated
under defined conditions (136). The success of future clinical applications
will depend on the ability to generate transplantable cell types derived from
hPSCs. An essential prerequisite to use hPSC-derived retinal tissue for
therapeutic purposes is to modify and optimize xeno-free and feeder-free
protocols compliant with Good Manufacturing Practice (GMP) guidelines
(35,137). Clinical cells lines will need to be validated, increasing the overall
length and costs and therapeutic procedures. The added necessity of quality
control testing, including identity, potency, sterility, karyotyping, whole-
genome sequencing, and sequence analysis make it unlikely that autologous
hPSC technology will be used for routine cell therapies (31,138). However,
the field has begun to move in this direction with several groups reporting the
generation of hPSCs in xeno-free conditions (35,137), and several attempts
for setting up cell banks containing GMP-grade hPSCs have been started.
hPSCs can be validated, expanded, and differentiated in large amounts;
therefore, one cell line could be used for multiple patients. The establishment
of cell/tissue banks that contain diverse GMP-approved cell lines from
healthy donors with different immunological profiles may be able to service
large amounts of the population.
Future Directions
The significant unmet challenges for retinal organoids include generating
sufficient numbers of specific cell types, achieving functional integration of
transplanted cells, and surgical delivery of retinal cells or tissue without
triggering immune responses, inflammation, and/or remodeling. Currently,
retinal organoid technology seems unable to produce correct proportions of
all retinal cell types, particularly those localized in the INL (32). Distinct
plexiform layers are not routinely formed, causing the retinal organoids to not
be fully morphologically or functionally mature (31). Several processes could
be potential sources of organoid variation, such as progenitor proliferation,
cell differentiation, and ontogenetic cell death. High heterogeneity between
cell lines and experimental outcomes, extended culture times, and manual and
laborious procedures currently limits the use of human retinal organoids for
clinical applications. Furthermore, the increasing number of differentiation
protocols vary in their efficiency that can be further compounded with cell
line variability (42). Studies attempting to standardize the field have found
that cell line–specific variables are responsible for differentiation efficiency
during early stages, while organoid generation and culture methods determine
the efficiency of maturation at later stages (42). A greater understanding of
mechanisms of organoid formation will be essential to improving and
standardizing protocols for the production of physiologically relevant tissue
across multiple cell lines.
The heterogeneity in production, composition, and maturation of
organoids represent essential challenges that are problematic for the entire
stem cell field. Beyond varying efficiencies in the differentiation and
maturation between cell lines and protocols, variation can occur between
organoids differentiated at the same time and even within different regions of
the same organoid. This heterogeneity has sparked recent efforts to better
evaluate current capabilities and limitations of retinal organoid structure. A
recent study used a single differentiation protocol, which incorporated
advances in differentiation protocols from multiple laboratories to generate
retinal organoids from 16 hPSC lines (54). Results of the study verified
organoid technology as a powerful and reliable tool that can be standardized
and optimized for use in multiple cell lines. Nevertheless, in order to
overcome heterogeneity and ensure standards for the manufacturing of cell
therapy products, the automation of differentiation and maintenance is
definitely needed (30,139). There is also room for improvement upon
cryopreservation protocols that allow easy distribution of cell products (10).
Despite reports of successful cryopreservation of retinal organoids in cGMP
approaches, conditions that allow for reliable preservation of final cell
products will be essential to establish (35). However, by tackling these
challenges, 3D retina culture systems will be of utmost importance for
regenerative approaches to treat currently incurable retinal degenerative
diseases.
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9
Regenerative Medicine: Hypothesis in
Support of In Situ Retinal
Regeneration
Michael Trese, Edward Wood, Antonio Capone Jr and Kimberly
Drenser
INTRODUCTION
Regenerative medicine is a term used to encompass many approaches to try
to repair and regenerate dead or diseased tissue/organs. There have been
many attempts to achieve this end, many of which have been misleading and,
in some cases, damaging to patients. In 2017, the FDA Director made the
statement that the FDA would clamp down on the charlatans but that
regenerative medicine was no longer science fiction but within reach of
modern medicine. There are centers around the world, which focus on
regenerative medicine in several forms: gene therapy, introduction of stem
cells from outside the body replanted in the space of damaged cells, or the
use of dormant in situ progenitor cells that reactivates the healing repair and
regeneration of tissues. This is a process that already exists in the human
body; for example, the gut lining is regenerated every 5 days by the activity
of progenitor cells. This is often referred to as in situ tissue regeneration and
is being tested in hearing loss, cartilage regeneration, hair growth, and wound
healing. We will confine our comments to in situ retinal regeneration. The
approach we use is to provide the microenvironment that can stimulate the
dormant progenitor cells to grow a healthy retina, recapitulating how it
occurred originally. This type of tissue regeneration has been possible in
lizards and other animals but only in limited fashion in humans, occurring in
the gut, hair, and nail growth. In lower animals, the regrowth of organs is
Wnt (Wingless, Int-1) signaling mediated. Wnt signaling is highly conserved
from Drosophila to humans suggesting its importance.
WHAT AND WHY WNT SIGNALING

The Wnt signaling pathway is a cellular signaling pathway that guides tissue
differentiation in the developing fetus and plays several roles in adults,
including angiogenesis; maintenance of the blood–brain barrier (BBB) and
blood–retinal barrier (BRB), essential features of neurovascular tissue; and
promotion of tissue regeneration (8). The Wnt gene family consists of 19
genes encoding proteins that act as extracellular signaling factors (2,9)
primarily impacting vascular and neural tissues. There are two Wnt
pathways: (a) the canonical/β-catenin pathway and (b) the noncanonical
pathway with the canonical pathway primarily controlling cell proliferation,
stem cell self-renewal, and tissue regeneration (10,11). Norrin is a growth
factor protein encoded by the NDP gene on the X chromosome and is the
strongest known activator of the canonical Wnt signaling pathway in the
retina and retinal vascular cells. In the retina, Müller glial cells express NDP
and produce norrin (12). Activation of the canonical Wnt pathway ultimately
culminates in the accumulation of β-catenin, a transcription factor that can
guide gene expression. In the case of vascular endothelial cells, norrin binds
to the retinal endothelial cell Frizzled-4 cell surface receptor (FZD4) in
conjunction with low-density lipoprotein receptor–related protein-5 (LRP5)
and tetraspanin family member-12 (TSPAN12), producing a myriad of
proteins that promote healthy cells (10). While essential in fetal and neonatal
tissue development, these key actuators of the Canonical Wnt signaling
pathway (norrin, FZD4, LRP5, and TSPAN12) remain expressible in the
adult, postnatal retina (13), suggesting that driving the canonical Wnt
pathway with exogenous norrin might shift cells into a developing or
embryologic state and aid in vascular and neural tissue regeneration.
In the eye and retina, a functional Wnt signaling system plays a key
fundamental role in the development of a sufficient vascular and neural
network to support vision. This is evidenced by patients with Wnt mutations
who develop many neurovascular diseases, such as familial exudative
vitreoretinopathy (FEVR), retinopathy of prematurity (ROP), Coats disease,
and Norrie disease (14). In Norrie disease, the developing fetus is unable to
produce a functional norrin protein and thereby unable to produce appropriate
vascular or neuronal structures for the brain, ear, and particularly the retina.
Because norrin-driven Wnt signaling is the primary Wnt system for the eye
(12), 100% of Norrie disease patients are bilaterally blind, yet only 40% are
deaf or developmentally delayed, highlighting that other Wnt signaling
pathways are available in the ear and central nervous system (10).
It is also a common observation that young children can recover from
injury faster than older adults. In particular, our clinical practice has observed
that pediatric patients with traumatic macular holes may heal these injuries
with clinically repaired photoreceptors and retinal pigment epithelial (RPE)
cells. Could it be that children still possess the ability to activate a norrin-
driven Wnt signaling wound repair/regenerative system? It was thought that
these conditions could only be seen in fetal and infant cells; however, our
group has shown that these Wnt functions can be activated in adult human
endothelial and neuronal cells (15). It, therefore, may be possible that through
the exogenous application of norrin, norrin-driven WNT pathway activation
may repair and regrow the retinal vascular and neuronal elements. Herein, we
describe cellular mechanisms by which the external application of the norrin
protein promotes the health, maintenance, and regeneration of retinal vascular
and neural cells.
NORRIN-DRIVEN RETINAL
VASCULAR REGENERATION
The fundamental theme in norrin-driven retinal vascular regeneration is that
the exogenous application of norrin repairs and regenerates vascular
endothelial cells by modifying the gene expression of a myriad of proteins
that improve endothelial cell tight junctions, strengthen the BRB, inhibit the
formation of significant tissue edema, and increase angiogenesis. When blood
vessels in the retina are injured in the setting of retinal vascular disease,
including diabetic retinopathy (DR), retinal vein occlusion (RVO), FEVR,
ROP, Coats, Norrie disease, and others, numerous growth factors, including
vascular endothelial growth factor (VEGF), are up-regulated in an attempt to
promote the growth of blood vessels and increase cellular oxygen delivery
(15). However, VEGF also directs the growth of disorganized and pathologic
immature vessels along the retinal surface and into the vitreous cavity (retinal
neovascularization) and increases the permeability of the BRB, which leads
to leakage of plasma and red blood cells through endothelial cells into the
retina, clinically manifesting as retinal edema. Vascular leakage in the retina
is clinically observed as macular edema or late-angiographic posterior and
peripheral endothelial leakage (LAPPEL) (16). A primary mechanism of
vascular leakage and retinal edema is VEGF-induced expression of
plasmalemma vesicle–associated protein (PLVAP). PLVAP is a cell-specific
protein that plays a pivotal role in transendothelial transport and
pathologically increases within retinal vascular endothelial cells in the setting
of VEGF-driven disease (17).
Contemporary medical therapies for treatment requiring retinal vascular
disease involve tissue destruction with ablative retinal laser therapy and/or
intravitreal injection of anti-VEGF agents (bevacizumab, ranibizumab,
aflibercept) (18). Intravitreal injection of anti-VEGF agents is the most
commonly performed procedure in ophthalmology and possibly all of
medicine (19). Through targeting VEGF, these therapies block an upstream
effector of retinal edema and retinal neovascularization. However, VEGF is
up-regulated for an intended purpose in response to vascular injury, and
numerous reports have shown that the presence of VEGF plays a pivotal role
in wound healing, neuroprotection, and vascular protection (20). Therefore,
long-term anti-VEGF therapy and VEGF suppression may impart unintended
consequences on cells (21). Targeting a more specific effector of retinal
vascular leakage, such as PLVAP, may serve as a more precise approach.
This may be accomplished with the activation of norrin, as the norrin protein
has been shown to directly down-regulate endothelial PLVAP (22), thereby
decreasing retinal edema. Our group has shown that the exogenous
application of norrin significantly decreases PLVAP levels alone and in the
presence of VEGF (Figure 9-1). The exogenous application of norrin also
mobilizes claudin-5, translocating it to the cell membrane, where it can help
restore the BRB and facilitate modulated angiogenesis (10).
FIGURE 9-1 Normalized plasmalemma vesicle-
associated protein (PLVAP) levels within cultured human
retinal endothelial cells in the presence of media alone,
norrin protein (Noregen; 200 ng/mL), VEGF165b (V165b)
25 ng/mL), and VEGF165b (25 ng/mL) and norrin. This
experiment shows that the exogenous application of norrin
significantly decreases PLVAP levels alone and in the
presence of VEGF.
Other proteins favorably driven by the exogenous application of norrin

include vascular endothelial-cadherin, BMP2, and SOX proteins (10).
With the application of norrin, the goal is to promote the health of the
retinal vascular network to decrease tissue ischemia. Our group has realized
this goal by showing that Norrin increases vascularization of the retina and
decreases retinal neovascularization in animal models of disease (23). A well-
perfused retina fails to drive the pathologic up-regulation of VEGF, thereby
avoiding VEGF-related consequences, including retinal edema and retinal
neovascularization, and obviating the need for anti-VEGF therapy.
NORRIN-DRIVEN RETINAL NEURAL

CELL REGENERATION
Exogenous application of the norrin protein has also been shown to promote
the health, maintenance, and regeneration of retinal neural cells, including
retinal ganglion cells (RGCs), bipolar cells, and photoreceptors. It is believed
that this process is due to norrin either improving the vascular perfusion to
neural cells, acting directly on retinal neural cells to increase their health and
repair them, or by acting on neuronal support cells, such as Müller glial cells
(15).
As described above, the exogenous application of norrin can improve
retinal vascularization. Improved tissue perfusion results in many positive
effects on cells, including increased oxygen delivery to diseased tissue and
the protection and repair of neural cells. To this end, our group (15,21) and
others (24) have shown that norrin-induced retinal vascularization increases
the survival of RGCs in an animal model of retinal disease (Figure 9-2).
FIGURE 9-2 In an animal model of oxygen-induced
retinopathy (OIR), eyes exposed to exogenous Norrin
(Norrin-OIR) display a greater number of dendrites
compared to control eyes, indicating that Norrin-OIR
facilitates neuronal maturation in the setting of vascular
disease.
Alternatively, norrin may also act directly and indirectly through trophic
factors on retinal neural cells to improve their health. The Wnt receptor
family of leucine-rich repeat-containing, G-protein–coupled receptors
(LGRs), specifically LGR4, is present within RGCs (25). Norrin binding to
LGR4 activates Wnt signaling and may have a positive effect on RGC health.
Norrin has also been shown to mediate neuroprotection to RGCs by
activating Wnt signaling within Müller cells and promoting them to secrete
neuroprotective trophic factors (26). Additionally, in an animal model where
RPE cells were made to continuously produce the norrin protein, retinal
photoreceptors were protected against light-induced cell damage through
protective effects of brain-derived neurotrophic factor (27). These studies
remind us of the complex relationships between retinal cell types and show
that the norrin protein can promote retinal neural and support cells to act
together for cellular health and repair.
In the presence of norrin, retinal neuronal support cells (e.g., Müller glial
cells) may actually turn into functioning retinal neural cells. This is an
evolutionarily conserved pathway still active in fish and birds where Müller
cells promote continuous and ongoing retinal regeneration (28). In mammals,
it is well known that trauma can induce glial to neural differentiation in the
central nervous system to a small degree that is mediated by inflammation
and related pathways, but there is more evidence pointing toward canonical
Wnt signaling being able to promote this even in the absence of injury (29).
Retinal Müller cells are the last cell type to form from retinal progenitor cells,
indicating that they retain some embryologic memory of turning into other
cell types (30). Essentially, activation of the canonical Wnt signaling pathway
may revert Müller cells back to a progenitor retinal cell type, which
subsequently undergoes differentiation to mature retinal cell types
functioning photoreceptors (31). This process seems to be more prominent in
youth and decreases with age (32), indicating that it may play a role in the
striking ability of children to heal as discussed above. In essence, Müller glial
cells are a source of retinal stem cells, and norrin-driven Wnt signaling may
promote them to participate in the process of retinal regeneration.
CONCLUSION
The concept of in situ retinal regeneration is complex and involves a
multitude of proteins and other signaling systems recreating the complex
milieu, which allowed healthy retinal growth. Certainly the pattern of Norrin
being present as a fetus and infant suggest that Norrin-driven Wnt signaling
is involved in organ generation. Future regenerative medicine clinical studies
and trials will hopefully pave the way for the future of vision restoration.
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10
International ROP: Retinopathy of
Prematurity in Chile
B. Andrés Kychenthal and S. Paola Dorta
INTRODUCTION
Shortly after the publication of the multicenter trial of cryotherapy for
retinopathy of prematurity (Cryo-ROP study) (1), the first ROP screening and
treatment program began in Chile. This chapter describes the personal
experience of the authors in the management of ROP cases in Chile from the
1990s to the present. We will describe how we diagnose and treat ROP in our
country, which has served as a model for several ROP programs in other
Latin American countries.
SCREENING STRATEGY
The development of a screening strategy is a key element in any ROP
program. There are certain elements that need to be considered, because they
may be different in each country. Examples include geography and the ability
of patients to travel to access care, phenotype of ROP appearance, which
provides characteristics of infants requiring screening, and level of neonatal
care.
In any ROP screening program, it is necessary to define which preterm
infants are to be examined, at what age, and how screening is to be
performed.
Which Preterm Infants Should Be Screened?

ROP is a public health problem whose magnitude is related to the level of
health care and technology in each country. Countries in which children
develop ROP must have sufficiently advanced neonatal intensive care units
(NICUs) to ensure the survival and adequate treatment of premature children.
At one extreme, are the NICUs of developed countries, which are so
sophisticated that increasingly smaller premature infants are able to survive
and, therefore, are at risk of developing ROP; at the other extreme, are
countries in which NICUs are not sufficiently advanced, and premature
children do not survive long enough to develop ROP. Developing countries,
among which are the majority of countries in Latin America and Eastern
Europe and now in sub-Saharan Africa, have intermediate-level NICUs that
are able to save premature infants but lack technologic advances and/or
resources to provide adequate care in NICUs. As a consequence, the infants
who develop ROP have different characteristics from those in developed
countries. There is insufficient screening and fewer screening strategies.
Patients may lack resources to access skilled ophthalmologists for early
diagnosis and treatment. Altogether these challenges lead to a high
percentage of children with ROP-induced blindness worldwide from
developing countries (2).
Within the group of developing countries is Chile, where ROP is a
leading cause of child blindness. In a study that examined all blind people in
Schools for the Blind in Chile, it was determined that 18% of blind children
were blind due to ROP. Of blind children over 10 years old, only 10% were
blind due to ROP; however, in children under 10 years, the value was 24%
(3). This suggests that as neonatal care in Chile improves with an increase in
the survival of increasingly smaller preterm infants, the incidence of ROP
will increase and contribute to the increasing number of blind children in the
country if advancements in neonatology are not accompanied by improved
screening strategies that diagnose children at risk of ROP and strategies to
assure treatment in a timely manner.
In 1995, approximately 40% of high-risk preterm infants in our country
were examined in neonatal care units. This was gradually improved by the
implementation of screening programs; in 2005, as part of national health
reform, the screening and treatment of ROP became mandatory throughout
the country. This has allowed practically all children in Chile at risk of ROP
to be screened.
When retrolental fibroplasia was described by Terry (4), children who
developed the disease were generally considered “large” premature infants.
Currently, in industrialized countries, preterm infants at risk of developing
ROP and requiring treatment are mainly those with an extremely low birth
weight < 1000 g. This observation indicates that the population of children at
risk of developing blindness due to ROP has varied over time and is also
different according to the degree of development of the neonatology unit
where the child is receiving treatment.
The majority of multicenter randomized studies come from NICUs in
developed countries. Their inclusion criteria usually include a birth weight
(BW) lower than 1250 g and a gestational age (GE) of 31 weeks. These
criteria cover 99% of the population at risk of developing ROP. However,
these data should be interpreted with caution when defining screening
strategies in developing countries, because the population at risk for ROP
includes infants with a greater BW (range = 410 to 2700 g) and whose
gestational ages (GA) range from 22 to 37 weeks (5).
We analyzed data obtained from 138 infants with threshold ROP treated
with laser photocoagulation by our group between 1995 and 2002 and found
that BW ranged between 495 and 1550 g and GA ranged from 23 to 35
weeks; 10.14% of treated infants had a birth weight between 1251 and 1550 g
(6). The number of patients with threshold disease and BW < 1250 g was
even bigger at the beginning of the screening program (Table 10-1).
TABLE 10-1
Percentage of preterms born at Hospital Salvador in Santiago, Chile that developed threshold disease in
the period 1995–1997 and 1995–2002.
As the National Reference Center for Premature Retinopathy during the

aforementioned period, we observed differences among neonatology units.
One neonatology unit in Chile referred 25% of infants with zone I ROP who
had BWs of 720 to 1110 g and GA of 25 to 30 weeks. The BW and GA did
not differ statistically from infants treated for ROP by our group (495 to 1550
g BW and 23 to 35 weeks GE) in the same period; however, infants from the
referral unit had more severe ROP and unstable clinical courses than infants
from our group. This finding may be interpreted that infants with more
serious complications of prematurity may be at a higher risk of developing
ROP especially in posterior zones. Therefore, how sick preterm infants are in
units may be considered when planning a screening strategy for a unit.
To understand ROP, one must be familiar with the natural history of the
disease. The Cryo-ROP study (1) demonstrated a total incidence of ROP for
those born with birth weights lower than 1251 g of 65.8%, and the incidence
was further subdivided for those with a BW < 750 g of 90% and for those
weighing 1000 to 1250 g at birth of 46.9%. That is, the incidence of ROP
appears inversely proportional to the birth weight of the preterm infant.
However, of the group of premature infants with retinopathy, only a small
percentage requires treatment.
Between 1995 and 2002, our group examined all premature babies with a
BW < 1501 g and/or a GA < 32 weeks in the neonatology unit of “El
Salvador Hospital” in the city of Santiago. Of the premature infants
examined, 52% developed ROP. Although there was a high incidence of
ROP, the vast majority of these children had spontaneous regression, and
treatment was not necessary. The need to apply treatment is inversely
proportional to birth weight (1). In our series, which includes premature
infants from various neonatology units in the city of Santiago examined by
members of our group, we determined that only 4.8% of preterm infants with
a BW < 1501 g and/or with a GA < 32 weeks required treatment, based on
the criteria of threshold retinopathy by the Cryo-ROP study (1) (see also
Chapter 38).
Based on the above findings, the screening guidelines during that period
were (a) premature infants with a BW < 1500 g and/or a GA ≤ 32 weeks and
(b) newborns (NBs) with a BW between 1500 and 2000 g with an unstable
clinical course or with associated risk factors, including repeated blood
transfusions, sepsis, and prolonged oxygen therapy.
At What Age Should Preterm Infants Be Examined for

ROP
The largest study performed that aids in determining when to examine
preterm infants is the evidence-based screening criteria for retinopathy of
prematurity (5). In this study, data on more than 4000 premature infants with
a BW < 1251 g and a postmenstrual age (PMA) < 31 weeks were analyzed. It
was determined that signs of unfavorable outcomes were not found in more
than 1% of patients with a PMA < 31 weeks and/or a chronologic age of 4
weeks or beyond a PMA of 46.3 weeks. That is, the increased risk of
developing severe ROP occurs between PMA of 30.9 and 46.3 weeks or
between chronologic ages of 4.7 and 18.7 weeks (5).
When analyzing the data from patients treated by our group between
1995 and 2002 according to the criteria of the Cryo-ROP study (1), we
observed that out of 138 patients treated, all had BWs ≤ 1550 g and GAs
between 24 and 33 weeks. The age to develop threshold ROP for those in
zone I was of 35.2 weeks PMA (range = 33 to 44 weeks) and 8.6 weeks
chronologic age (range = 4 to 14 weeks). For those who with zone II ROP,
the age to develop threshold was a 37.2 weeks PMA (r = 31.5 to 42.0) and
9.5 weeks chronologic age (r = 6 to 14). All these data are in agreement with
the aforementioned study (5,7).
However, when developing screening strategies, one must also consider
the time necessary to assure adequate conditions for the treatment of the
infant. Delays in transfer can be several days either due to several factors,
including the general condition of the infant, the means of transportation, or
the coordination of space in the receiving unit. As more time passes after the
diagnosis of ROP is made, the worse the prognosis will be even after
treatment. When analyzing our data obtained after laser treatment for ROP,
we observed that for those children in which the screening and treatment was
performed by a member of our group, 92% experienced a favorable outcome
(group A, n = 47) based on the criteria used in the Cryo-ROP study.
However, patients who had to be transferred to our unit for treatment
experienced a greater delay between diagnosis and treatment, and the
favorable outcomes decreased to 78% (group B, n = 117) (7) (Figure 10-1).
FIGURE 10-1 Delay in performing treatment effect:
preterms in which screening and treatment was performed
by a member of our group, 92% experienced a favorable
outcome (group A). However, patients who had to be
transferred to our unit for treatment experienced a greater
delay between diagnosis and treatment, and the favorable
outcomes decreased to 78% (group B).
Accounting for all these data, it was determined that screening begin at a
chronologic age of 4 weeks. The frequency of subsequent examinations was
determined according to the classification of ROP at each examination.
How Preterm Infants Are Examined

In the majority on NICUs in Chile, preterm infants are examined by indirect
ophthalmoscopy.
In 1995, we introduced the use of video-ophthalmoscopy. This
technology allowed us to teach and take images that were not possible to
obtain with previous cameras (Figures 10-2 and 10-3). One of these images
was included in the International Classification of ROP revisited in 2005 (8),
to illustrate AP-ROP.
FIGURE 10-2 Video-ophthalmoscopy: introduced in

1995, the use of video-ophthalmoscopy allowed teaching
and taking images that were not possible to obtain with
previous cameras, as (A) and (B) are images of aggressive
posterior ROP obtained using a 28D condensing lens.
FIGURE 10-3 Clinical characteristics of zone II ROP (A)

compared with zone I ROP (B–D). Zone II retinopathy of
prematurity with its most prominent feature: an elevated
ridge with extraretinal neovascular proliferation. Atypical
morphology in zone I retinopathy of prematurity: a
peripheral arteriovenous arcade located exactly in the line
apparently separating the vascular from avascular retina
(B). Neovascularization that forms a syncytial network of
flat preretinal new vessels (C), flat neovascular
proliferation posterior to the arteriovenous arcade (D).
(From Retina 2006;26:S11–S15 (Figures 1, 2, 3, and 4).)
In 2008, the first RetCam arrived in Chile, and we initiated modern screening
techniques and telemedicine. Since then, more of these cameras are in use in
NICUs throughout the country. The geography of the country and the high
concentration of retina specialists in the capital, Santiago, makes the use of
telemedicine a great contribution to ROP management. RetCam images were
taken initially by physicians only, but today they are taken by nurses and
informed by ophthalmologist with ROP training.
ZONE I ROP
Zone I ROP has represented the greatest clinical and therapeutic challenge in
the management of ROP patients.
During the 1990s, there was misunderstanding regarding posterior ROP,
making the diagnosis more difficult and resulting in delays in treatment and
worse outcomes than those of patients with zone II disease. A publication by
Shapiro et al. from 1993 reported findings similar to our experience at that
time (10).
Later, the Early Treatment for Retinopathy of Prematurity Randomized
Trial in 2003 (9) and the International Classification of ROP revisited in 2005
(8) helped to address this problem.
In 2000, we published results on zone I patients treated with threshold
disease as defined by the Cryo-ROP study. Eight out of 20 eyes with zone I
had unfavorable results (40%). Mean BW and GA were similar in zone I and
zone II ROP patients that developed threshold disease, but the mean PMA at
treatment was significantly different, 35.2 weeks for zone I ROP versus 37.2
weeks for zone II ROP, P = 0.006. In this same paper, it was also noted that
special attention should be given to atypical morphology present in zone I
cases: Some zone I cases showed clinical characteristics that were difficult to
analyze according to the 1984 International Classification of Retinopathy of
Prematurity. At times, instead of a demarcatory line between vascular and
avascular retina characteristic of stage 1 ROP, a demarcatory vessel was
observed. This vessel coursed exactly the line of demarcation between
vascular and avascular retina and had a general direction perpendicular to the
retinal vessels. Classic stage 3 disease was not always observed in zone I
despite the presence of plus disease. In many instances, the neovascular
proliferation associated with plus disease took the form of a flat network of
vessels either at the level of the demarcation or slightly more posterior. The
demarcation between vascular and avascular retina was always noticeable;
even if it did not have height as a ridge, there was an abrupt change in color
(11) (Figure 10-3).
In 2006, another paper by our group regarding the clinical characteristics
and treatment outcomes of zone I patients reported the atypical clinical
features observed in patients with zone I ROP including flat preretinal
neovascularization and a demarcation vessel between the vascular and
avascular retina. In this publication, within zone I, 2 anatomic subgroups
were defined (10). Posterior zone I was defined as a circular area limited by
the radius from the disc to the center of the macula. Anterior zone I was
defined as the annular area bounded by posterior zone I and zone II. The
results of laser photocoagulation were that significantly better outcomes were
obtained in the eyes with anterior zone I ROP, with 31 out 48 (65%) eyes
achieving favorable outcomes compared to eyes with posterior zone I ROP
with no eyes achieving a favorable outcome (P < 0.001) (12) (Figure 10-4).
FIGURE 10-4 Schematic representation of zone I ROP.

Two anatomic subgroups were defined. Posterior zone I
was defined as a circular area limited by the radius from
the disc to the center of the macula (A). Anterior zone I
was defined as the annular area bounded by posterior zone
I and zone II. Clinical images of anterior and posterior
zone I (B, C). (From Retina 2006;26:S11–S15 (Figures 5
and 6).)
Changes in neonatal care improved survival rates of very low birth weight
preterm infants and produced an increase in zone I ROP. In 2005, the mean
BW and GA were 945 g and 27.5 weeks, respectively, and in 2008, they were
859 g and 25.5 weeks, respectively. At this time, the percentage of zone I
ROP represented 34% of the treated patients.
The poor outcomes for zone I compared to zone II ROP significantly
changed with a better understanding of the clinical characteristics of posterior
ROP, through the use of digital imaging for screening that led to earlier
treatment. Also, the addition of antiangiogenic agent, anti-VEGF, as
therapeutic option, improved these cases.
NATIONAL GUIDELINES FOR THE

SCREENING AND TREATMENT OF
ROP IN CHILE
In 2005, health reform in Chile was implemented. As part of this reform, the
diagnosis and treatment of a group of diseases was guaranteed for the entire
population. Within these diseases was ROP.
The most important aspect of this reform is that it provided “Clinical
Guidelines” that were mandatory throughout the country. In this way, a
national screening program was started for all infants born with a BW of
<1500 g, and if treatment was necessary for ROP, was guaranteed. For the
treatment, 5 photocoagulation units were put into operation throughout the
country, and a vitreoretinal surgery center was established, where we
operated on infants from all over Chile.
Highlighted in this chapter is one of the major shortcomings of
developing countries and that is not having a national program for the
detection and treatment of ROP.
Establishing national programs that are mandatory and supervised by a
health authority would reduce blindness due to ROP; the goals are for this to
occur in all countries.
ROP TREATMENT
Laser Photocoagulation in ROP
When we started treating ROP in 1994, indirect photocoagulation with a
diode laser was utilized. At that time, cryo-coagulation was commonly used
in other countries; however, there were advantages of using diode laser:
fewer local adverse effects and better outcomes.
The laser procedure was indicated for threshold ROP, as defined initially
by the Cryo-ROP study (1) but then changed to the criteria (type 1 ROP)
established by the Early Treatment for Retinopathy of Prematurity
Randomized Trial (9).
The unfavorable outcomes decreased noticeably over the years. From
1995 to 2004, the percentage of unfavorable outcomes in a group that
included 166 eyes was 18.1%, whereas from 2005 to 2008, the percentage of
unfavorable cases decreased to 2% in an group that included 133 eyes with
similar clinical characteristics.
The change to more favorable outcomes was influenced by several
factors: first, the greater technical experience of the medical team, and
second, the implementation of Chilean health reform in 2005 that included
mandatory clinical guidelines for all neonatal units in the country. The
clinical guidelines include a maximum time until treatment of 72 hours for
type 1 ROP requiring photocoagulation. As we mentioned previously, prior
to the implementation of this reform, referrals for treatment could be delayed
excessively thereby leading to worse outcomes (7).
However, as screening and treatment were improved in Chile, so did
neonatal care leading to increased survival of very low birth weight preterm
infants. The increase in extremely preterm infants led to a rise in posterior
ROP, the condition responsible for most of the unfavorable outcomes after
laser treatment.
Antiangiogenic Drugs (Anti-VEGFs)

Conventional therapy, that is, laser photocoagulation, has excellent results for
type 1 ROP in zone II, but eyes with ROP in zone I have a higher rate
unfavorable results after laser.
Laser ablation of the avascular retina is a destructive procedure. Eyes
with posterior ROP have large avascular retina that needs to be treated to
control the disease. Even in cases where there is a favorable anatomic result,
the functional outcome can be poor particularly in posterior zone I eyes,
where vascular retinal development does not often extend beyond the macula.
The extensive laser treatment increases the chances of local and systemic
laser-associated complications (Figure 10-5).
FIGURE 10-5 Extensive laser photocoagulation for cases

of posterior zone I (A, B). Laser is a destructive procedure
that could result in poor functional outcomes in these
cases.
The search for complementary or alternative treatments was then necessary

for these cases. Based on several facts (see Chapter 38), the use of anti-VEGF
agents was a logical choice. Therefore, in 2008, we clinically assessed the
safety and efficacy of intravitreal bevacizumab (Avastin) to help manage
ROP. We used Avastin in 3 clinical situations before using it as a first-line
therapy. First, we administered Avastin to 1 group of infants after disease
progression despite laser photocoagulation. Second, we treated infants after
treatment with laser who had posterior zone I ROP, mainly aggressive
posterior ROP (AP-ROP) but did not receive laser in the macular region. In
these cases, we did not laser the area where the macula should develop, but
instead injected Avastin days after photocoagulation as long as there was
vascular activity, so as not to risk an unfavorable result due to insufficient
laser treatment. A third group of patients were administered the drug prior to
vitrectomy because of the presence of a vascularly active retinal detachment
when the infant was first seen (13).
After positive clinical results, we used bevacizumab as first-line therapy.
Between August and December 2008, 12 consecutive eyes of 7 premature
infants with type 1 ROP were treated with a single intravitreal injection of
Avastin (0.625 mg). Nine eyes had zone I ROP, and 3 eyes had zone II ROP.
The infants had birth weights between 600 and 1100 g (mean 846.57 g). The
gestational ages ranged from 23 to 28 weeks (mean 25.57 weeks). Avastin
was injected at an average of 34 weeks PMA for zone I cases and at 40 weeks
PMA for zone II cases. Eyes showed regression of the disease with no need
for additional treatment. We observed that the vascularization of the retina
after ROP regression after Avastin injection followed a normal though slower
pattern. Some patients showed a peripheral avascular retina that remained
unchanged. No complications attributable to the bevacizumab injections were
seen in any of our patients (14) (Table 10-2, Figures 10-6, 10-7, 10-8, 10-9,
10-10, 10-11).
TABLE 10-2
AP-ROP, aggressive posterior ROP; RE, right eye; LE, left eye; M, male; F, female.
FIGURE 10-6 Case 1 posterior zone I ROP (AP-ROP).
Avastin injection with type 1 ROP at 32 weeks
postmenstrual age, at 4 weeks of birth. Retinal
vascularization does not even reach the macular area (A).
At 35 and 56 weeks postmenstrual age, retinal vessels
reaches the foveal area (B). At 46 weeks postmenstrual
age (14 weeks after the Avastin injection), retinal
vascularization within zone III (C). (From Retina
2010;30:S24–S31 (Figure 1).)
FIGURE 10-7 Case 2 posterior zone I ROP. Avastin

injection given at 32 weeks postmenstrual age, when the
baby developed type 1 ROP (A). Regression of the
vascular activity at the anterior and posterior segment was
observed 1 week after the procedure (B). At 43
postgestational weeks, the retinal vascularization reaches
zone III (C). (From Retina 2010;30:S24–S31 (Figure 2).)
FIGURE 10-8 Case 3 zone I ROP. Avastin was given at
34 weeks postmenstrual age. When the ridge starts
regressing the limit between the vascular and avascular
retina looks very neat, giving the impression of a
neovascular strip advancing toward the avascular area (A).
Normal looking vascularization reached zone III at 46
weeks (B). (From Retina 2010;30:S24–S31 (Figure 3).)
FIGURE 10-9 Case 4. Initial fibrosis and ridge and at the
time of the Avastin injection, at 38 and 54 weeks
postmenstrual age (A). Adequate regression without any
traction development after the injection at 39 and 84
weeks (B). (From Retina 2010;30:S24–S31 (Figure 4).)
FIGURE 10-10 Case 5. AP-ROP and Avastin was
injected at 32 and 28 weeks postmenstrual age. Images of
the right and left eyes taken 6 weeks after Avastin
injection. There is a marked vascular strip moving toward
the avascular retina, which, previous to this study, would
have prompted us to perform photocoagulation. (From
Retina 2010;30:S24–S31 (Figure 5).)
FIGURE 10-11 Case 6. Evident fibrosis at the time of
treatment at 40 and 28 weeks postmenstrual age (A).
Regression was slower and 3 weeks post-Avastin
injections (43 and 28 weeks) (B) anterior and posterior
segment activity was still present. In spite of a poor
systemic condition, there was no need for further
treatment, and retinal vascularization reached zone III at
48, 28 weeks (C). (From Retina 2010;30:S24–S31 (Figure
6).)
Since that study, anti-VEGF drugs have been used in Chile for the treatment
of ROP, decreasing the number of unfavorable results and blindness due to
this disease. We believe that photocoagulation for type 1 ROP might not be
the appropriate first-line therapy for every case. We currently treat posterior
ROP with anti-VEGF drugs as the first-line therapy. The availability of anti-
VEGF drugs allows establishing the best choice for each patient. We
recognize that more study is needed regarding safety, but for visual outcomes
(see chapters by Ells (53) and Hartnett (38)), anti-VEGF agents have
improved outcomes in Chile.
The reactivation of ROP after the use of anti-VEGF drugs is an important
clinical problem. There are published series with high percentages of
reactivation even a prolonged time after anti-VEGF drug injection. Our
experience has been different with a very low number of cases of
postinjection reactivation.
We believe that there are important differences between preterm infants
from other series and ours, that is, ours may be more representative of
developing countries. Among these are genetic and racial differences and
others such as a higher BW and older GA, on average, for patients from
developing countries. In addition, infants born at larger BW and older GA are
likely more developmentally mature when Avastin is administered than the
extremely preterm infants in developed countries (see Chapter 38).
Considering the possibility of reactivation of ROP, we believe that
photocoagulation of the peripheral retina in patients treated with anti-VEGF
should be contemplated, especially in those cases where prolonged follow-up
is not possible.
We would perform retinal examinations after regression following anti-
VEGF treatment until 60 weeks of PMA, every 2 weeks until week 52 and
then monthly.
Vitreoretinal Surgery for ROP

Vitreoretinal surgery is an essential therapeutic tool for the management of
advanced cases of ROP. We introduced vitreoretinal surgery for ROP in
Chile in 1999.
Until 2004, we operated using 20-gauge instrumentation. Then, we
started using micro incision techniques, and in 2008, we published on the use
of small gauge vitrectomy in ROP in a series of 13 eyes of 10 patients. We
concluded that surgical intervention with a three-port 25-gauge
transconjunctival sutureless vitrectomy was an effective technique to attach
the retina in cases with stage 4A retinal detachment in ROP (15,16) (Figure
10-12). Since that time, we have been using small gauge vitrectomy in all our
surgeries.
FIGURE 10-12 Ports for the infusion, light pipe, and
cutter are placed in the inferotemporal, superotemporal,
and superonasal quadrants, respectively, to perform 25-
gauge transconjunctival lens-sparing vitrectomy for stage
4A ROP. (From Retina 2008;28:S65–S68 (Figure 1).)
In 2010, we published a study regarding a series of eyes in which vitrectomy

was performed after Avastin injection in vascularly active ROP retinal
detachments. The use of an anti-VEGF drug as an adjunct in these cases
proved very effective. A very significant reduction in the vascular activity of
these eyes could be observed after the injection when vitrectomy was
performed (Figures 10-13, 10-14, 10-15). A change from vascular to fibrous
tissue was observed, but there were no rhegmatogenous retinal detachments
following Avastin injections. We observed that as the extent of neovascular
tissue growth into vitreous increased, the more likely it was for fibrosis
development to occur after Avastin injection. A favorable outcome was
observed for all eyes (3).
FIGURE 10-13 Preterm born at 25 weeks with 810-g
BW. At 38-week PMA, a stage 4A ROP with significant
vascular activity was present. (From Retina
2010;30:S32–S36 (Figure 1).)
FIGURE 10-14 One week after Avastin injection, a
marked reduction in the vascular activity can be observed.
(From Retina 2010;30:S32–S36 (Figure 2).)
FIGURE 10-15 Favorable anatomic outcome after
Avastin and a 25-gauge lens-sparing vitrectomy in the
same patient. (From Retina 2010;30:S32–S36 (Figure 3).)
Timing the vitrectomy, 5 to 7 days after Avastin injection is important to

avoid excessive fibrous proliferation that could increase the chances of an
unfavorable result (Figure 10-16).
FIGURE 10-16 Importance of adequately timing

vitreoretinal surgery after injecting an anti-VEGF as an
adjunct prior to vitrectomy. Vascularly active ROP retinal
detachment (A). Five to seven days postinjection, there is
regression of the vascular activity, signaling the best
moment to perform surgery (B). After that time, fibrous
conversion of the neovascular tissue might induce or
promote progression of the detachment (C).
The role of laser, anti-VEGF drugs, and vitreoretinal surgery in the

management of ROP are represented in Figure 10-17.
FIGURE 10-17 Flow chart of the management of ROP

with laser photocoagulation, anti-VEGF drugs, and
vitreoretinal surgery.
SPECTRAL DOMAIN OPTICAL

COHERENCE TOMOGRAPHY (SD-OCT)
IN ROP
Current management of ROP cases that require treatment, including
photocoagulation of the avascular retina, anti-VEGF drugs, and vitreoretinal
surgery, results in healthy-looking and vascularized retinas in many patients.
One current goal is to detect the factors that are responsible for achieving
good vision in these eyes. The identification of fine retinal characteristics
using SD-OCT may play a key role in achieving this objective. The
identification of prognostic factors will also aid in the selection of better
treatment options.
With that purpose, we studied ROP patients with SD-OCT. In one study,
we investigated 20 eyes of 11 patients treated with bevacizumab only or in
addition to photocoagulation and vitreoretinal surgery. One of the most
interesting findings we observed was the correlation between the foveal
pattern and visual acuity. When a foveal contour was present, vision tended
to be better. In patients without an foveal contour, only 2 of the 5 eyes had
better than 20/400 vision; in contrast, 10 of 11 eyes with a foveal contour had
better than 20/400 vision. The absence of development of a foveal contour
was associated with a younger GA and zone I disease.
Patients without development of a foveal contour can have either good or
poor vision. In those with good vision, the macular thickness is maintained,
and the retinal layers are recognizable; in patients with poor vision, the
macular thickness is reduced, and the inner retinal layers are not clearly
recognizable.
We concluded that anti-VEGF treatment for ROP can result in macular
development and good vision. Notwithstanding normal ophthalmoscopy and
RetCam images, some eyes had abnormal macular configurations on OCT.
As previously mentioned, when a normal macular configuration is
present, vision tends to be better. Thus, morphologic characteristics of retinal
anatomy might be predictive of visual function (17) (Figure 10-18).
FIGURE 10-18 SD-OCT patients with type 1 ROP in
zone I treated with Avastin ▫. A:720-g BW and 24 weeks
GA, with foveal contour development imaged at 28
months after Avastin. Persistence of retinal layers (inner
nuclear, inner plexiform) over the fovea (red bracket).
Signs of macular development: outer segment elongation,
inner segment/outer segment and external limiting
membrane presence (blue arrows), and thickening of the
outer nuclear layer at the fovea (orange bracket). B:650-g
BW and 24 weeks GA, with no foveal contour
development imaged at 38 months after Avastin. Not all
retinal layers are present (nerve fiber layer absent). It is
difficult to differentiate between the ganglion cell layer
and inner plexiform layer.
REFERENCES
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Ophthalmol 2002;120:1470–1476.
6. Andrés Kychenthal B, Ximena Katz V, Paola Dorta S. Outcome After Laser and Surgical
Treatment for Retinopathy of Prematurity. ARVO Meeting, Fort Lauderdale, 2000.
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Review. PubMed PMID: 16009843.
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treatment for retinopathy of prematurity randomized trial. Arch Ophthalmol
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10. Shapiro MJ, Gieser JP, Warren KA, et al. Zone I Retinopathy of Prematurity. Proceedings of the
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1993:149–155.
11. Katz X, Kychenthal A, Dorta P. Zone I retinopathy of prematurity. J AAPOS 2000;4:373–376.
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treatment outcomes. Retina 2006;26:S11–S15.
13. Kychenthal A, Dorta P. Vitrectomy after intravitreal bevacizumab (Avastin) for retinal
detachment in retinopathy of prematurity. Retina 2010;30(4 Suppl):S32–S36. doi:
10.1097/IAE.0b013e3181ca146b.
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in: Retina 2009;29(1):127.
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11
International ROP: Tele-ROP in India
—A Real World Experience
Anand Vinekar
THE NEED FOR TELE-ROP

Tele-retinopathy of prematurity (ROP) services started in India in 2007. The
need arose from the inability of providing mandated ROP screening services
to rural and semi-urban neonatal centers, a vast majority of which did not
have qualified ophthalmologists to perform screening. The lack of services
resulted in “delayed or no screening” and hence blind infants.
The demographics in India are similar to those in several middle-income
countries. A large number of live births, a significant proportion of which are
premies, and lack of trained manpower plague the health care delivery
systems. Annually, of the 27 million babies born in India, 3.5 million are
born premature. With <20,000 ophthalmologists, 2,000 retina specialists, and
200 ROP specialists, the need to screen for and treat ROP outweighs the
demand several fold.
KIDROP
The first program and currently the largest tele-ROP service is the Karnataka
Internet Assisted Diagnosis for Retinopathy of Prematurity (KIDROP),
initiated in Bangalore, India, in 2007. KIDROP has completed to date over
150,000 screening sessions and identified over 2,500 infants with Type 1
ROP (1, 2, 3, 4, 5). The program currently screens in over 126 neonatal units
in the south Indian state of Karnataka, which has a population of
approximately 65 million. KIDROP addressed these problems of unscreened
rural and semi-urban premature infants for ROP using a novel platform of
telemedicine and employing for the first time nonphysician graders who
travel to remote neonatal intensive care units (NICUs) (1, 2, 3). On average,
the teams undertake 7,000 km of travel to reach these centers wherein 2,000
to 2,500 imaging sessions are performed every month.
KIDROP's services are provided zonally. Each zone covers
approximately 5 to 6 “districts” of the state, which are defined by
geographical contiguity. Each zone has its own equipment, including a wide-
field ROP camera, a vehicle, information technology infrastructure, as well as
a trained team. Each team has a fixed schedule of 6 working days a week
during which over 20 to 50 neonatal units are visited.
A single camera is used per zone, and the screening occurs in the
neonatal unit where preidentified infants are dilated and kept ready by the
nursing staff for imaging. Imaging is performed inside the incubator, on the
warmer, or in an adjoining room under medical supervision. Images are
captured in the video mode and the required still images are saved. The
imagers use the ROP camera to capture retinal images of in-house and
follow-up infants in these outreach centers. They then grade and interpret the
images and make management decisions within the NICU itself after
reviewing the images at the end of the session. Images are also shown to the
treating neonatologist and parents, when possible, and documented on the
“ROP card” given to the mother, in the hospital records, in the online
database, and in the image database (2,4,5), All images are backed up onto a
secure online database and are available for the remote expert to evaluate.
Reporting by nonphysicians is facilitated through a “decision-aiding
algorithm,” (1,2) which was developed with a three-way triaging code of
(Figure 11-1):
FIGURE 11-1 The decision algorithm followed by trained
technicians in the KIDROP program. (Source: Vinekar A,
et al. The KIDROP model of combining strategies for
providing retinopathy of prematurity screening in
underserved areas in India using wide-field imaging, tele-
medicine, non-physician graders and smart phone
reporting. Indian J Ophthalmol 2014;62(1):41–49.)
red (requiring treatment or urgent review by the MD)

orange (disease or immature retina that can be followed)
green (mature retina in both eyes)
The imagers are trained to image the retina to include the ora serrata (6). This
reduces the chances of missing ROP that may be present in the retinal
periphery and also allows the “discharge” of the baby without the need of an
ophthalmologist performing indirect ophthalmoscopy to confirm retinal
maturity. All infants are imaged at least once between the postmenstrual ages
of 40 to 44 weeks to coincide with the physiological age of retinal maturity.
This applies to infants who do not develop any stage ROP and vascularize
normally. Those infants who develop any disease or require treatment with
laser are followed up for longer periods. Those infants who have received
anti-VEGF treatment currently need the longest follow-up. This acts as a
“safety net” in a program where there is no manual examination by a ROP
specialist. The remote specialist views and reports these images on his or her
smart phone (1,2). Owing to improved Internet and mobile data packages,
relatively rapid upload and review of images in a single session are possible
in a short period of time. The reporting time on an average after upload is 4
minutes. The time taken to report all “severe cases” of any session that need
urgent attention is <30 minutes (3). These images are uploaded manually
(RetCam) or automatically (Neo) for the ROP specialist to review and report.
Training and accreditation of the “imagers” is a critical contributor to the
success of the KIDROP program since the first “triage of diagnosis and
decision” is made by this cadre of nonphysicians (1,2). The KIDROP STAT
(Score for Training and Accreditation of Technicians) was developed to train
and certify imagers. This comprises three levels (I, II, and III) and has a 20-
point score, which tests the knowledge, skill, and practice patterns of the
imager in his or her native setting (Figure 11-2). On an average, training a
new imager can take between 30 and 90 working days.
FIGURE 11-2 The STAT score developed to train
imagers in the KIDROP program. (Source: Vinekar A, et
al. The KIDROP model of combining strategies for
providing retinopathy of prematurity screening in
underserved areas in India using wide-field imaging, tele-
medicine, non-physician graders and smart phone
reporting. Indian J Ophthalmol 2014;62(1):41–49.)
INNOVATIONS
More recently, a low-cost, portable camera was developed in India, 3Nethra
Neo (“Neo”) (Forus Health, India) with the clinical collaboration of
KIDROP. This affordable, smaller, and more portable device has made
transport, imaging and scaling-up of the program easier (7,8).
The Neo was designed specifically for ROP and infant retinal imaging. It
is a contact camera with a single, monolithic, handheld probe and provides a
120-degree field of view (Figure 11-3). It has a patented liquid lens, which is
integrated into the hand-piece that does not need to be removed after each
session (unlike the RetCam ROP lens). The Neo's illumination source is a
patented warm LED light. The image resolution is 2040 × 2040, compared to
1800 × 1600 of the RetCam (Figure 11-4A and B) and results in a square
image, which provides an extra arc of the superior and inferior retina that
would be cropped out in the rectangular image of other cintact cameras
(Figure 11-5). The systemic and ocular safety of the Neo was established (7).
FIGURE 11-3 The “3Nethra Neo” (“Neo”) ROP Camera
(Forus Health, India) is a monolithic, portable, wide-field,
contact infant retinal camera.
FIGURE 11-4 Comparison of RetCam vs. Neo Images.
A:RetCamShuttle (Natus USA) image of Stage 3 ROP in
the right eye. The image is rectangular due to the 1800 ×
1600 sensor of the camera. B:Neo (Forus Health, India)
camera of the same eye showing a 2040 × 2040 square
image with an additional superior and inferior retinal arc.
(Source: Vinekar A, Bhende P. Innovations in technology
and service delivery to improve retinopathy of prematurity
care. Community Eye Health 2018;31(101):S20–S22.)
FIGURE 11-5 RetCam vs. Neo image. The superimposed
images demonstrate the “extra arcs” of retina (dotted
yellow oval) in the superior and inferior retina,
respectively, that are imaged by the Neo and not on the
RetCam.
An innovation in training was necessary to reduce the time needed to train

and certify imagers. This was possible after the introduction of online
training. The online platform “WISE-ROP” (Wide-field Imaging for
Screening and Education for ROP) has training modules based on the onsite
training curriculum. Imagers read and undergo self-assessment quizzes at the
end of each module. Video sessions and viva voce to correct the technique
and address practical difficulties are scheduled through the platform with an
assigned mentor. Skill is graded as per the STAT-Score levels and tested
before a completion certificate is awarded (9). The WISE-ROP platform is
currently being upgraded to provide image reporting services even after
training is completed. This is particularly useful in states or zones where
there are no qualified specialists to review or report the images.
Software-based innovations have also contributed to KIDROP's impact
on expanding services to underserved regions. Disease severity detection,
enhancing clinical features of “poor-quality” images, and image processing
tools to enhance vascular details noninvasively are now getting integrated
into the imaging platform (10, 11, 12). More recently, automated diagnosis
algorithms are being integrated into the image portals to provide a more rapid
triage of the diagnosis that can aid the grader or the clinician (12).
EXPANDING SERVICE DELIVERY

A significant advantage of ROP screening with imaging rather than indirect
ophthalmoscopy lies in the documentation of other non-ROP conditions. In a
study of 1,450 preterm infants screened by the KIDROP program, 7.7% had a
diagnosis other than ROP, which included conditions as severe as
retinoblastoma (13). Furthermore, the program has expanded to provide
“universal screening” in some zones for full-term, healthy infants consistent
with the Government's Universal Health Coverage program. In a study of
1,021 term infants imaged within 3 days of birth, 4.7% had an abnormality of
whom 1.6% required medical or surgical intervention (14).
The program has now initiated training of resident nurses rather than
outsourced “imagers” to screen infants. This allows “round-the-clock”
screening of ROP and universal screening rather than the weekly fixed
schedule that is currently followed. This program is the real-world
implementation of the “paradigm shift” that Gilbert et al. recommended for
middle-income countries (15). A cloud-based image reservoir is created and
can be accessed and reported by ROP experts.
IMPACT
In its 12th year and since inception, the impact of KIDROP in preventing
infant blindness assessed using the “blind-person-years” (BPY) formula (i.e.,
number of infants × per capita income × life expectancy) is over 154 million
US dollars. An impact assessment of scaling up the program in India showed
that in the 10 high-risk ROP states, with a population of roughly 680 million,
over 35,000 infants would be detected with ROP and over 1,200 would need
treatment annually. The financial saving in “BPY” is estimated at USD 108
million annually (4). The United Nations Development Programme (UNDP)
report on KIDROP (16) and the National Health and Medical Research
Council (NHMRC, Australia) report based on the Center for Disease Control
(CDC) guidelines (17) suggested that “wide-field imaging performed by
trained imagers” is more reliable than “indirect ophthalmoscopy performed
by inadequately trained ophthalmologists” and warned against the possible
“medicolegal liability” that “suboptimal management” could expose the
Government to, since “ROP screening and treatment is now a mandate”. This
would be applicable not only to India but to other nations with similar ROP
demographics. The KIDROP program is now being emulated in other states
of India and some other nations in the region.
SURVEILLANCE AND ADVOCACY

After a Supreme Court of India judgment in 2015, ROP screening is now
under the medicolegal scanner and has received legal mandate. A National
Task force of ROP set up by the Central Government has created operational
guidelines for screening and referral. For the first time, these guidelines
recommend image-based screening by nonophthalmologists as a viable
alternative for ROP screening to indirect ophthalmoscopy (18). Furthermore,
the Indian ROP Society was formed in 2016 and comprised ophthalmologists
skilled in screening for ROP who are collaborating to provide ethical and
evidence-based ROP management to address local needs (19).
SUMMARY
Tele-ROP services in the real world pose unique challenges (20). KIDROP
was built on the tenants of wide-field imaging, wide coverage, and a robust
telemedicine platform. The unique aspects of the program are as follows: (1)
nonphysicians are allowed to report and analyze the images as the first point
of contact, thereby providing the diagnosis and decision to the (rural) mother
before she leaves the center; (2) indirect ophthalmoscopy is not mandated
before an infant can be discharged from screening; and (3) the program aims
at detecting any stage of ROP, not just treatment-warranted ROP. KIDROP,
therefore, differs from other programs that used referral-warranted criteria as
their end-points.
REFERENCES
1. Vinekar A, Gilbert C, Dogra M, et al. The KIDROP model of combining strategies for
providing retinopathy of prematurity screening in underserved areas in India using wide-field
imaging, telemedicine, non-physician graders and smart phone reporting. Indian J Ophthalmol
2014;62:41–49.
2. Vinekar A, Jayadev C, Mangalesh S, et al. Role of telemedicine in retinopathy of prematurity
screening in rural outreach centers in India—a report of 20,214 imaging sessions in the
KIDROP program. Semin Fetal Neonatal Med 2015;20:335–345.
3. Vinekar A, Jayadev C, Bauer N. Need for telemedicine in retinopathy of prematurity in middle-
income countries: EROP vs. KIDROP. JAMA Ophthalmol 2015;133:360–361.
4. Vinekar A, Mangalesh S, Jayadev C, et al. Impact of expansion of telemedicine screening for
retinopathy of prematurity in India. Indian J Ophthalmol 2017;65(5):390–395.
5. Vinekar A, Avadhani K, Dogra M, et al. A novel, low-cost method of enrolling infants at risk
for Retinopathy of Prematurity in centers with no screening program: the REDROP study.
Ophthal Epidemiol 2012;19(5):317–321.
6. Vinekar A, Dogra M, Shetty B. Imaging the ora serrata with the 3Nethra Neo camera—
importance in screening and treatment in retinopathy of prematurity. Indian J Ophthalmol
2020;68(1):270–271.
7. Vinekar A, Rao SV, Murthy A, et al. A novel, low-cost, wide-field, infant retinal camera,
“Neo”: technical and safety report for the use on premature infants. Transl Vis Sci Technol
2019;8(2):2. doi: 10.1167/tvst.8.2.2.
8. Vinekar A, Dogra MR, Jayadev C et al. Evaluation of a new, low-cost, portable, wide-field
digital retinal camera, “Neo” for screening infants or retinopathy of prematurity—a prospective,
multi-center, validation report in Asian Indian infants. Invest Ophthalmol Vis Sci 2016;57(12).
Available at http://iovs.arvojournals.
9. Vinekar A, Bhende P. Innovations in technology and service delivery to improve retinopathy of
prematurity care. Community Eye Health 2018;31(101):S20–S22.
10. Jayadev C, Vinekar A, Mohanachandra P, et al. Enhancing image characteristics of retinal
images of aggressive posterior retinopathy of prematurity using a novel software, (RetiView).
Biomed Res Int 2015;2015:898197
11. Rajashekar D, Srinivasa G, Vinekar A. Comprehensive retinal image analysis for aggressive
posterior retinopathy of prematurity. PLoS One 2016;11(10):e0163923.
12. Vijayalakshmi C, Sakthivel P, Vinekar A. Automated detection and classification of telemedical
retinopathy of prematurity images. Telemed J E Health 2019. doi: 10.1089/tmj.2019.0004.
13. Jayadev C, Vinekar A, Bauer N, et al. Look what else we found—clinically significant
abnormalities detected during routine ROP screening. Indian J Ophthalmol
2015;63(5):373–377.
14. Vinekar A, Govindaraj I, Jayadev C, et al. Universal ocular screening of 1021 term infants
using wide-field digital imaging in a single public hospital in India—a pilot study. Acta
Ophthalmol 2015;93:e372–e376.
15. Gilbert C, Wormald R, Fielder A, et al. Potential for a paradigm change in the detection of
retinopathy of prematurity requiring treatment. Arch Dis Child Fetal Neonatal Ed
2016;101(1):F6–F9.
16. UNDP Social Sector Service Delivery: Good Practices Resource Book 2015 | UNDP in India.
Available at:
http://www.in.undp.org/content/india/en/home/library/democratic_governance/SSS-delivery-
good-practices-2015.html. Accessed January 6, 2017.
17. CDC. Developing an effective evaluation report, 2013. Available at:
https://www.cdc.gov/eval/materials DevelopingAnEffectiveEvaluationReport_TAG508.pdf.
Accessed March 9, 2017.
18. Project Operational Guidelines. Prevention of Blindness from Retinopathy of Prematurity in
Neonatal Care Units. Available at: https://phfi.org/wp-content/uploads/2019/05/2018.ROP-
operational-guidelines.pdf. Accessed May 21, 2019.
19. Vinekar A, Azad R, Dogra MR, et al. The Indian retinopathy of prematurity society: a baby step
towards tackling the retinopathy of prematurity epidemic in India. Ann Eye Sci 2017;2:27.
20. Vinekar A, Dogra M, Azad RV, et al. The changing scenario of retinopathy of prematurity in
middle and low income countries: unique solutions for unique problems. Indian J Ophthalmol
2019;67:717–719.
12
International ROP: Retinopathy of
Prematurity in México
María Ana Martínez-Castellanos, Ana González H. León and
Estephania Feria Anzaldo
INTRODUCTION
Current Mexican Guidelines for ROP Screening
The 2013 reformation on article 61 of the Mexican General Health Law (1)
ruled that ophthalmic assessment of all infants at the 4th week of birth be
mandatory in order to detect early vision and functional threats to vision. It
also emphasized the importance of an extensive ophthalmoscopic
examination in all preterm infants. Moreover, in 2015, the Mexican Secretary
of Health (2) updated the retinopathy of prematurity (ROP) screening and
clinical practice recommendations widening the range for screening to
include a requirement that all infants born at gestational age (GA) ≤ 34 weeks
and/or with a birth weight (BW) of ≤1,750 g be screened. This ruling has
prevented missing many preterm infants at risk of ROP. Because it is not
uncommon to diagnose ROP in infants weighing more than 1,500 g at birth
and/or with a GA of 30 weeks or older in Mexico (2,3); as noted in other
articles, the screening criteria used in more developed nations may not apply
to middle-income countries (4, 5, 6). Mexican guidelines aim to prevent
functional decline and/or organ loss; give an early and appropriate treatment
by establishing standardized detection, diagnosis, and follow-up criteria;
educate healthcare professionals to identify at-risk populations; and refer in a
timely fashion to third-level center patients who require specialized care (2).
SCREENING STRATEGY
Screening for ROP plays an important role in preventing childhood blindness
in preterm infants. Because of the vast diversity in neonatal care, the facilities
available, and the number of trained ophthalmologists who can provide an
adequate screening program around the world, adaptation of guidelines to
each population must be considered (2,4).
In Mexico, Grupo ROP Mexico published the most relevant information
on ROP and the impact in our nation (7). The actual preferred practice
guidelines of ROP management in Mexico (2) acknowledge their
recommendations and suggest ROP screening be performed in the following
infants:
1. All preterm infants ≤34 weeks GA and/or <1,750 g birth weight.

2. All preterm infants >34 weeks GA and >1,750 g birth weight who
received supplemental oxygen and who by criteria of his/her treating
physician needs an exam.
To the treating physician criteria, the preterm infants with associated risk
factors (cardiovascular pathology, associated sepsis, unstable clinical course,
among others).
Since there is an indirect relationship between GA and the time when
ROP manifests, it is recommended to use postmenstrual age (PMA) and is
used to determine the time of the first ophthalmologic exam (2). We base the
beginning of revisions on the Pediatric American Association Guidelines (8)
by commencing screening at a chronologic age of 4 or 32 weeks PMA,
whichever occurs first (2). The frequency and the time of the next check-up
will be determined by the stage of the disease at each examination (9).
Ophthalmoscopic exams should continue until the retinal vasculature is
completely developed or when the disease is regressed after giving treatment
(2).
In Mexico, the majority of screenings are performed with the help of
indirect ophthalmoscopy by an experienced retinal specialist,
ophthalmologist, and, in some cases, the pediatrician or neonatologist (3,10).
Currently, the evaluation by an expert using indirect ophthalmoscopy is the
gold standard for diagnosis and follow-up of patients with ROP; however,
telemedicine is becoming an important tool to detect disease with a similar
sensitivity for zone I disease and type 2 ROP (11); this is especially important
because Mexico lacks sufficient specialized doctors to perform indirect
ophthalmoscopy needed to diagnose infants (12).
ROP TREATMENT
Laser Photocoagulation in ROP
The criteria for laser therapy have been revised from threshold ROP to
include the earlier stage of high-risk prethreshold ROP (13).
Adequate and appropriate laser photocoagulation for ROP is different
from the application of lasers in adult retinal vascular diseases, and many
ophthalmologists need additional training to successfully treat ROP (13,14).
The standard of care in ROP was the diode red (810 nm wavelength)
laser indirect ophthalmoscope. Laser has many advantages over cryotherapy.
There is less posttreatment pain, adnexal edema, exudative retinal
detachment, vitreoretinal traction, and vitreous hemorrhage believed due to
reduced breakdown of the blood–retinal barrier (15).
Visual outcomes reported after laser are better than those after
cryotherapy. It is important to keep close observation after treatment to
ensure there is no need of more (as seen on Figure 12-1) (13,14).
FIGURE 12-1 Fundus photos showing (left side)
confluent grayish white laser burns up to the ridge.
Inadequate regression (top right) requiring more laser and
adequate regression (top bottom) needing only close
follow-up 7 days after laser. (From Good WV, Hardy RJ,
et al. Final visual acuity results in the early treatment for
retinopathy of prematurity study. Arch Ophthalmol.
2010;128:663–671.)
The initial settings on the laser console are chosen based on the fundus
pigmentation and area to be treated. Usually, laser power is 250 mW at 150
ms duration on the repeat mode setting at 300-ms interval. The treatment
must not be faster as this can result in inadequate burns. The intensity is a
grayish white rather than white spot, and placement of spots must be nearly
confluent. The laser power must be varied; less energy is used for the anterior
and superior retina compared to the posterior and inferior retina or to the
retina close to the ridge. A pediatric depressor is used to rotate and stabilize
the globe and to indent the anterior retina for laser (14,15).
It is essential to treat the entire avascular retina from the ridge to the ora
for 360 degrees and not leave untreated “skip” areas near the posterior ridge
of the avascular retina (14).
In a single session, one may place 3,000 to 4,000 spots in each eye to
cover the avascular retina in zone I disease and APROP. A smaller number of
spots, that is, 1,000 to 2,000, are needed to manage type 1 ROP outside zone
I (15).
Antiangiogenic Drugs (Anti-VEGFs)

ROP is a biphasic disease consisting of an initial phase of oxygen-induced
vascular obliteration and delayed physiologic retinal vascular development
by a period of hypoxia-induced vessel proliferation (15) (see also Chapter 52
by Shulman and Hartnett).
Beginning in 2007, several retrospective reports presented promising
experience with off-label use of anti-VEGF in infants with progressive ROP
(15,16). These are covered in more detail in Chapters 52 and 53.
Our injection protocol involves the use of the following:
1. Topical 5% povidone–iodine in periocular skin and the conjunctiva.

2. A sterile lid speculum is placed.
3. Amount of 0.01 mL (0.25 mg) of bevacizumab is injected 1 mm
posterior to the corneoscleral limbus in the inferotemporal quadrant
using a 32-gauge needle and 4 mm.
In a study conducted by our group (16), regression of neovascularization was

observed in all eyes in the first 48 hours. Total of 88.8% required only one
injection and had complete resolution. And 83% had favorable anatomical
outcomes without any signs of retinal detachment, macular dragging, or folds
after 60 months of follow-up. Only one patient with bilateral ROP was
diagnosed with recurrent disease after 2 months of follow-up and required
additional treatment (3). (It should be noted that the GA and BW of infants at
risk of ROP in Mexico are older and larger than in developed countries.)
Intravitreal bevacizumab has recently emerged as an important and
promising yet controversial treatment option in ROP, because it is an easier
treatment to administer than ablative therapy with laser. The procedure is
shorter, does not require special equipment, and can be carried out with short-
acting topical anesthesia, thereby avoiding general anesthesia or sedation
(15).
There are few studies that evaluate recurrence of ROP only with anti-
VEGF or in combination with other therapies. In these publications, follow-
up is carried out closely (every 1 to 3 weeks) according to the clinical
evolution. However, patient follow-up is discontinued after 55 weeks in Latin
America (5).
It should be noted that the largest case series of patients treated with
intravitreal bevacizumab found recurrent ROP in 8.3% (20/241) of infants
and 7.2% (34/471) of eyes that had been treated for ROP, stage 3 with plus
disease, and aggressive ROP. The mean PMA at the time of recurrence was
51.2 ± 4.6 weeks at an interval of 16.2 ± 4.4 weeks between treatments (16).
The main risk factors for recurrent ROP were neovascularization in the
subsequent aggressive ROP, longer hospitalization time, and necrotizing
enterocolitis, especially that required surgical intervention (15,16).
Vitreoretinal Surgery for ROP in Mexico

Stage ROP 5 lensectomy and membranectomy for retrolental membrane and
funnel RD.
Surgical technique:
A corneal approach is used. An anterior chamber maintainer is placed
through a corneal incision at the lower temporal quadrant in order to maintain
the pressure at constant gravity infusion and avoid hemorrhage. Another
incision is made at the 9 or 10 o'clock meridian with a 15° or 1-mm knife (see
Figure 12-2A). Viscoelastic is injected to reform the anterior chamber and to
release iris synechiae to the corneal endothelium or anterior lens capsule (as
seen on Figure 12-2B). Then, an incision is made in the anterior lens capsule,
and the irrigation–aspiration system (Simcoe) is introduced into the lens
(Figure 12-2C). The lens is aspirated, and the lens capsules are removed with
Maxgrip forceps.
FIGURE 12-2 Fundus photos showing (left
A:Superotemporal limbal incision. Usually done between
9 and 10 meridian using a 15-degree knife. B:Fine cannula
to inject viscoelastic, anterior chamber is reformed and
posterior iris synechiae are thorn. C:Simcoe cannula is
used to gently aspirate the lens. D:Three ports can be seen
on the image. The irrigation cannula can be seen at M5,
while there are two superior ports where the scissors and
the grasps are entered to perform the iris sphincterotomy.
E:Diathermy-hand piece with plug-on that allows high-
frequency capsulotomy (Oertli). F:Image shows how the
plug-on tip is used to open and dissect a plane through the
retrolental membrane. G:Image shows how the retrolental
membrane is now open with the help of the diathermy-
hand piece, retina can be visualized behind the membrane.
To improve visualization of structures behind the iris, we perform an iris
sphincterotomy using retinal scissors (as shown on Figure 12-2D).
The Oertli surgical platform is used at high-frequency capsulotomy
(CAPS) (HF) to perform a membranectomy in areas that are free of retinal
adhesions as determined by transillumination with an endoilluminator
(Figure 12-2E shows how the capsulotomy device looks, and Figure 12-2F
and G show how we use it to form a plane through the membrane and dissect
it).
Sometimes, stripping of membranes with the retina forceps is performed,
in addition to external drainage, particularly if extensive subretinal
hemorrhage was found on preoperative ultrasound. For tamponade,
viscoelastic is placed into the vitreous cavity and anterior chamber to
maintain the retinal funnel open, but not to fully attach the retina. Retinal
breaks are avoided. All corneal incisions are closed with 10-0 Nylon.
For the other hand, surgical intervention with a three-port 25-gauge
transconjunctival sutureless vitrectomy is an effective technique to attach the
retina in cases with stage 4A to 4B retinal detachments.
IMAGING IN ROP FOR DIAGNOSIS

Fluorescein Angiography
Image analysis showed two distinct types of peripheral vascular changes:
Group 1 or Nonproliferative
Extensive areas of capillary nonperfusion with widespread arteriovenous
shunting between adjacent primary vessels, tortuosity of primary vessels
(“plus-like” disease), abnormal budding of tertiary vessels and capillaries,
abnormal capillary tuffs, and absence of foveal avascular zone were found.
Vascular changes were readily identified in FA images (17).
Group 2 or Proliferative
Some or all the characteristics of group 1 were seen plus leakage of dye at the
boundary between perfused and nonperfused retina and/or optic disc.
A finding was the absence of avascular foveal zone, which has been
reported by Henaine et al. in 50% of patients born at 36 weeks of GA or more
(17,18).
Fundus Wide-Angle Imaging

Screening and management of ROP detected prethreshold or threshold
disease in some eyes in which images were able to be obtained. See how on
Figure 12-3 we can clearly define the avascular area (11).
FIGURE 12-3 Wide-angle imaging fundus angiography.

A clear line of avascular retina can be found at the
temporal periphery.
Ophthalmologic examinations were needed in 20% of cases that did not reach
threshold or prethreshold disease because of poor image quality or
overestimation of ROP (12).
Optomap Imaging
At the present time, ROP screening is generally performed by binocular
indirect ophthalmoscopy and/or wide-field digital imaging systems. In
Mexico, there are two kinds of wide-field viewing systems in use for the
pediatric age group: contact (3nethra Neo, ICON, PanoCam, RetCam) and
noncontact systems (19).
The Optos uses a noncontact ultra-widefield dual wavelength laser
camera that is able to capture high-quality images from infants with ROP.
The Optomap is a panoramic digital image generated by Optos scanning laser
technology, which shows approximately 82% of the retina (19,20). Figure
12-4 displays an example of a ultra-widefield imaging of a retina in a preterm
male.
FIGURE 12-4 Ultra-widefield imaging. Left eye of a
premature male. Plus disease is seen in all four quadrants.
Avascular retina is seen in all the superior areas.
Retinal imaging in patients with ROP using ultra-widefield imaging system

has recently been popularized by Patel et al. (19,20). This team first reported
that Optos was capable of capturing high-quality images in infants with ROP
using the Optos noncontact ultra-widefield fundus imaging system (19) (see
also Chapter 15). One of the important results from their study was that the
Optomap identified “skip areas” missed by initial laser treatment in the
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second level Hospital in México. Rev Mex Ped. 2016;83:3.
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retinopathy of prematurity. JAMA Ophthalmol. 2018;136:498–504. doi:
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13. Dobson V, Quinn GE, Summers CG, Hardy RJ, Tung B. Visual acuity at 10 years in
cryotherapy for retinopathy of prematurity (CRYO-ROP) study eyes: effect of retinal residua of
retinopathy of prematurity. Arch Ophthalmol. 2006;124:199–202.
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PEDIATRIC RETINA

Antonio Capone Jr, MD, FACS

Copyright © 2021 Wolters Kluwer

Cataloging-in-Publication Data available on request from the Publisher

Thomas M. Aaberg Sr, MD, MSPH

James D. Akula, PhD

Isabelle Audo, PU, PH

Yomtov Robert Barishak, MD

Jean Bennett, MD, PhD

Saptarshi Biswas, PhD

Colin A. Bretz, PhD

Diana Schorry Brightman, PhD, MS, CGC

William J. Brunken, PhD, FARVO

John Peter Campbell, MD, MPH

Antonio Capone Jr, MD, FACS

Megan E. Capozzi, PhD

Joseph Carroll, PhD

R. V. Paul Chan, MD, MSc

Emmanuel Y. Chang, MD, PhD

Christopher D. Conrady, MD, PhD

Donnell J. Creel, PhD

Catherine A. Cukras, MD, PhD

Patricia A. D’Amore, PhD, MBA, FARVO

Paola Dorta Salfate, MD, MHA, MBA

Kimberly A. Drenser, MD, PhD

Estephania Feria Anzaldo, MD

C. Stephen Foster, MD, FACS, FACR, FARVO

Clare E. Gilbert, FRCOpth, MD, MSc

Morton F. Goldberg, MD, FACS, FARVO, FAOS

Ronald M. Hansen, PhD

Anna-Lena Hård, MD, PhD

M. Elizabeth Hartnett, MD, FACS, FARVO

Ann Hellström, MD, PhD

George Hoppe, PhD

G. Baker Hubbard III, MD

Dale D. Hunter, MD, JD

Lea V. M. Hyvärinen, MD, PhD

Raymond Iezzi Jr, MD

Samer Khateb, MD, PhD

Sang Jin Kim, MD, PhD

Baruch D. Kuppermann, MD, PhD

Andrés Kychenthal Bab, MD

Anna La Torre, PhD

Michelle E. LeBlanc, PhD

Katherine Ann Lee, MD, PhD

Ana González H. León, MDPhysician

Michele C. Madigan, PhD

Maria Ana Martinez-Castellanos, MDAssistant Professor

Jason S. Meyer, PhD

Robert S. Molday, PhD, FRSC, FARVO

Timothy G. Murray, MD, MBA

Eric Nudleman, MD, PhD

Joan Marie O’Brien, MD

Dolly A. Padovani-Claudio, MD, PhDPediatric Ophthalmologist

C. K. Patel, MRes(Oxon), FRCOphth

John S. Penn, PhD

Wanda Pfeifer, OC(C), COMT, CO

Jan M. Provis, PhD

Thomas A. Reh, PhD

Jonathan F. Russell, MD, PhD