The Washington Manual of Surgical Pathology

The Lauren V.
Ackerman Laboratory of Surgical Pathology

Barnes-Jewish and St. Louis Children's Hospitals
Washington University Medical Center
Department of Pathology and Immunology
Washington University School of Medicine
St. Louis, Missouri
Editors
Peter A. Humphrey, MD, PhD
Lou is P. Dehner, MD
John D. Pfeifer, MD, PhD
I
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© 2012 by Department o£ Pathology and Immunology, Washington University School of Medicine

First Edition © 2008 by Department of Pathology and Immunology, Washington University School
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Library o£ Congress Cataloging-in-Publication Data

The Washington manual of surgical pathology I editors, Peter A. Humphrey, Louis P. Dehner,
John D. Pfeifer. -2nd ed.
p.;cm.
Surgical pathology
Includes bibliographical references and index.
Summary: "The aim of this book is to provide a practical manual that is helpful to pathologists and
residents in the daily practice of surgical pathology. The text encompasses all anatomic sites in the
human body. Each chapter covers tissue handling, gross examination, gross dissection and diagnosis,
processing, slide preparation, microscopic diagnosis, special studies, and diagnostic reporting. The
format of the book would be modded after the Washington Manuals for other specialties (such as
Medical Therapeutics, Oncology, and Surgery)"-Provided by publisher.
ISBN 978-1-4511-1436-2
I. Humphrey, Peter A. II. Dehner, Louis P., 1940- III. Pfeifer, John D. IY. Title: Surgical pathology.
[DNLM: 1. Pathology, Surgical-Handbooks. WO 39]
LC classification not assigned
617'.07---dc23 2012000549
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10 9 8 7 6 5 4 3 2 1
My wife Kay, and children Tom and Jennifer-P.A.H.
For the continued opportunity, joy, and satisfaction of working with great
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P.J., S.M., and L.P.D., for their friendship and support-J.D.P.
Contributors
Ashima Agarwal, MD Danielle H. Carpenter, MD
Instructor Surgical Pathology Fellow
Department of Pathology and Immunology Department of Pathology and Immunology
Washington University School of Medicine Washington University School of Medicine
St. Louis, Missouri St. Louis, Missouri
Hussam Al-Kateb, MSc, PhD Yumei Chen, MD, PhD

Assistant Professor Molecular Genetic Pathology Fellow
Associate Director of Cytogenomics and Department of Pathology and Immunology
Molecular Pathology Washington University School of Medicine
Department of Pathology and Immunology St. Louis, Missouri
St. Louis, Missouri
Rebecca D. Chernock, MD
Assistant Professor
Craig Allred, MD
Professor
Section Head, Breast Pathology
St. Louis, Missouri
St. Louis, Missouri Brian Collins, MD
Associate Professor
Catalina Amador-Ortiz, MD Section Head, Cytopathology
Resident Department of Pathology and Immunology
Department of Pathology and Immunology Washington University School of Medicine
Washington University School of Medicine St. Louis, Missouri
St. Louis, Missouri
Catherine E. Cottrell, PhD
Emily A. Bantle, MD Assistant Professor
Chief, Dermatopathology Associate Director of Cytogenomics and
Department of Pathology Molecular Pathology
David Brant Medical Center Department of Pathology and Immunology
Travis Air Force Base, California Washington University School of Medicine
St. Louis, Missouri
Nils Becker, MD
Resident Kimberly G. Crone, MD
Department of Pathology and Immunology Dermatopathologist
Washington University School of Medicine Associated Dermatologists of West
St. Louis, Missouri Bloomfield and Commerce
Commerce Township, Michigan
Elizabeth M. Brunt, MD
Professor
Section Head, Liver!GI Pathology Erika Crouch, MD, PhD
Department of Pathology and Immunology Professor
Washington University School of Medicine Department of Pathology and Immunology
St. Louis, Missouri Washington University School of Medicine
St. Louis, Missouri
Dengfeng Cao, MD, PhD
Assistant Professor Rosa M. Davila, MD
Department of Pathology and Immunology Medical Director, Clinical Laboratory
Washington University School of Medicine Barnes jewish West County Hospital
y
vi I CONTRIBUTORS
Louis P. Dehner, MD Johann D. Hertel, MD, MA

Professor of Pathology and Immunology Resident
Professor of Pathology in Pediatrics Department of Pathology and Immunology
Department of Pathology and Immunology Washington University School of Medicine
Washington University School of Medicine St. Louis, Missouri
St. Louis, Missouri
Jena Beth Hudson, MD
Samir K. El-Mofty, DMD, PhD Resident
Professor of Oral and Maxillofacial Department of Pathology and Immunology
Pathology Washington University School of Medicine
Associate Professor of Otolaryngology St. Louis, Missouri
and Head and Neck Surgery
Associate Professor of Pathology Phyllis C. Huettner, MD
Section Head, Head and Neck Pathology Associate Professor of Pathology and
Department of Pathology and Immunology Immunology
Washington University School of Medicine Associate Professor of Obstetrics and
St. Louis, Missouri Gynecology
Section Head, Gyn Pathology
John L. Frater, MD Department of Pathology and Immunology
Assistant Professor Washington University School of Medicine
St. Louis, Missouri
Michael E. Hull, MD
Assistant Professor
Joseph P. Gaut, MD, PhD Department of Pathology and Immunology
Instructor Washington University School of Medicine
Washington University School of Medicine Peter A. Humphrey, MD, PhD
St. Louis, Missouri Ladenson Professor of Pathology and
Omar Hameed, MD Immunology
Associate Professor Chief of Anatomic and Molecular
Department of Pathology, Microbiology Pathology
and Immunology; Department of Surgery
Vanderbilt University
St. Louis, Missouri
Medical Director, Surgical Pathology;
Associate Medical Director, Anatomic Mohammad 0. Hussaini, MD
Pathology Resident
Vanderbilt University Medical Center Department of Pathology and Immunology
Nashville, Tennessee Washington University School of Medicine
St. Louis, Missouri
George J. Harocopos, MD
Assistant Professor of Ophthalmology and Michael Isaacs, BS
Visual Sciences Director of Information Systems
Assistant Professor of Pathology and Department of Pathology and Immunology
Immunology Washington University School of Medicine
Department of Ophthalmology and Visual St. Louis, Missouri
Sciences
Washington University School of Medicine Jason A. Jarzembowski, MD, PhD
St. Louis, Missouri Assistant Professor
Department of Pathology
Anjum Hassan, MD Medical College of Wisconsin
Assistant Professor Program Director, Perinatal Pathology
Department of Pathology and Immunology Department of Pathology
Washington University School of Medicine Children's Hospital of Wisconsin
St. Louis, Missouri Milwaukee, Wisconsin
CONTRIBUTORS vii
Jeffery M. Klco, MD, PhD Jochen K. Lennerz, MD, PhD

Research Instructor Pathologist
Department of Pathology and Immunology Institute of Pathology
Washington University School of Medicine University Ulm
St. Louis, Missouri Ulm, Germany
Michael J. Klein, MD James S. Lewis, Jr., MD

Professor of Pathology and Laboratory Assistant Professor
Medicine Department of Pathology and Immunology
Department of Pathology and Laboratory Washington University School of Medicine
Medicine St. Louis, Missouri
Weill Cornell School of Medicine
Pathologist-in-Chief and Director Helen Liapis, MD
Department of Pathology and Laboratory Professor
Medicine Section Head, Renal Pathology
Hospital for Special Surgery Department of Pathology and Immunology
New York, New York Washington University School of Medicine
St. Louis, Missouri
Friederike Kreisel, MD
Assistant Professor Anne C. Lind, MD
Department of Pathology and Immunology Associate Professor
Washington University School of Medicine Section Head, Dermatopathology
St. Louis, Missouri Department of Pathology and Immunology
Elise L. Krejci, MD St. Louis, Missouri
Staff Pathologist
Sherman Hospital Ta-Chiang Liu, MD, PhD
Elgin, Illinois Resident
Hannah R. Krigman, MD Washington University School of Medicine
Associate Professor St. Louis, Missouri
Department of Laboratory Medicine and
Pathology Dongsi Lu, MD, PhD
University of Minnesota Assistant Professor
Minneapolis, Minnesota Department of Pathology and Immunology
Shashikant Kulkarni, PhD St. Louis, Missouri
Associate Professor
Medical Director, Cytogenomics and Changqing Ma, MD, PhD
Molecular Pathology Resident
Julie Elizabeth Kunkel, MD Mitra Mehrad, MD

Staff Pathologist Resident
Department of Pathology Department of Pathology and Immunology
San Antonio Military Medical Center Washington University School of Medicine
San Antonio, Texas St. Louis, Missouri
Kathryn M. Law, MD ILKe Nalbantoglu, MD

Instructor Assistant Professor
viii I CONTRIBUTORS
TuDong Nguyen, MD, PhD Jon H. Ritter, MD

Assistant Professor Professor
Department of Pathology and Immunology Director of Anatomic and Surgical
Washington University School of Medicine Pathology
St. Louis, Missouri Department of Laboratory Medicine and
Pathology
Deborah Novack, MD, PhD University of Minnesota
Associate Professor of Internal Medicine Minneapolis, Minnesota
Associate Professor of Pathology and
Immunology Souzan Sanati, MD
Washington University School of Medicine Assistant Professor
St. Louis, Missouri Department of Pathology and Immunology
Sushama Patil, MD St. Louis, Missouri
Washington University School of Medicine Robert E. Schmidt, MD, PhD
St. Louis, Missouri Professor
Chief of Neuropathology
Richard J. Perrin, MD, PhD Department of Pathology and Immunology
Assistant Professor Washington University School of Medicine
St. Louis, Missouri Kevin Selle, MT, HTL (ASCP)
Lab Superoisor
Arie Perry, MD Department of Surgical Pathology
Professor of Pathology and Neurological Barnes-]ewish Hospital
Surgery St. Louis, Missouri
Director of Neuropathology
Department of Pathology, Division of Maria F. Serrano, MD
Neuropathology Physician
University of California San Francisco Department of Pathology
School of Medicine Kaiser Permanente San Rafael Medical
San Francisco, California Center
San Rafael, California
John D. Pfeifer, MD, PhD David E. Spence, MD
Professor of Pathology and Immunology
Pathologist
Professor of 0 bstetrics and Gynecology
Department of Pathology
Vice Chairman for Clinical Affairs
Associated Pathologists at Erlanger
Hospital
Chattanooga, Tennessee
St. Louis, Missouri
Kiran R. Vij, MD
Meredith E. Pittman, MD, MSCI Fellow in Clinical Cytogenomics
Resident Department of Pathology and
Department of Pathology and Immunology Immunology
Samuel J. Pruden, II, MD, FACS Nathan C. Walk, MD

Dermatopathology Fellow Pathologist/Dermatopathologist
Washington University School of Medicine Middlesex Hospital
St. Louis, Missouri Middletown, Connecticut
CONTRIBUTORS ix
Hanlin L. Wang, MD, PhD Frances V. White, MD

Professor Associate Professor
Department of Pathology and Laboratory Department of Pathology and Immunology
Medicine Washington University School of Medicine
David Geffen School of Medicine Interim Chief of Pathology, St. Louis
University of California Los Angeles Children's Hospital
Los Angeles, California St. Louis, Missouri
Joshua I. Warrick, MD Heather N. Wright, MD

Resident Surgical Pathology Fellow
Lourdes R. Ylagan, MD, FIAC
Mark Watson, MD, PhD Associate Professor
Associate Professor Department of Pathology and Laboratory
Department of Pathology and Immunology Medicine
Washington University School of Medicine Roswell Park Cancer Institute
St. Louis, Missouri Buffalo, New York
Rao Watson, MD Barbara Zehnbauer, PhD, FACMG
Resident Adjunct Professor of Pathology
Washington University School of Medicine Emory University School of Medicine
St. Louis, Missouri Atlanta, Georgia
Preface
Welcome to the second edition of the The Washington Manual ofSurgical Pathology. As
with the first edition, the book draws on the rich heritage of surgical pathology at Barnes
Hospital (now Barnes-Jewish Hospital), which began with Lauren V. Ackerman, MD.
When Dr. Ackerman joined the institution in 1948 as director of surgical pathology,
he was the first trained and board-certified pathologist to occupy a position previously
held by surgeons. Dr. Ackerman initiated the paradigm of diagnostic excellence with
a central focus on the patient, and advanced the vital role of the surgical pathologist
as a consultant to clinicians. He was keenly aware of the role of surgical pathologists
as educators. Dr. Ackerman also emphasized that, as investigators of illness from an
observational perspective, surgical pathologists were uniquely qualified to correlate
morphologic findings with the clinical behavior of disease. If he were alive today,
Dr. Ackerman would likely enthusiastically add that surgical pathologists are also
uniquely qualified to correlate morphologic findings with the molecular features of
disease.
Although multivolume textbooks as well as subspecialty texts continue to have
a central role in education and everyday clinical practice, this book is an attempt
to respond to the immediate needs of an ever-accelerating world in which pathology
residents and fellows never seem to have enough time to "get it all done." Likewise,
pathologists in practice never seem to be able to sign out cases quickly enough to satisfy
their clinical colleagues, and clinicians themselves need a ready and available reference
in surgical pathology. As with the first edition, it is our goal that this book, continuing
in the fast-access Washington Manual outline format, will fill a niche in our hectic
world.
This edition of the The Washington Manual of Surgical Pathology includes several
significant revisions and additions. In addition to updates to every chaptet; several
new sections have been added to the manual, including new subchapters on renal
transplant pathology and cystic diseases of the kidney, a new chapter on diseases of the
retroperitoneum, and a reorganized chapter on molecular genetic testing. Over 1000
new images have been added, and the text includes the updated AJCC cancer staging
schemes released in 2010. As with the first edition, a companion website offers the text
and image bank (which now contains over 2850 full-color images) in an electronic and
fully searchable format (see the inside front cover for website access information).
As with the first edition, most of the individuals who have contributed to this work
had their surgical pathology training in the Lauren V. Ackerman Laboratory of Surgical
Pathology at Barnes-Jewish Hospital or are current members of the faculty. As is the
tradition of the Washington Manual series, residents and fellows have contributed to
many of the chapters.
Peter A. Humphrey, MD, PhD

Louis P. Dehner, MD
John D. Pfeifer, MD, PhD
X
Acknowledgments
The second edition of The Washington Manual of Surgical Pathology would not have
been possible without the participation of our colleagues, who so willingly and gener-
ously provided their time and expertise to the project. All but one of the authors (again,
special thanks to Dr. Michael Klein) are, or were, faculty or house staff in the Depart-
ment of Pathology and Immunology at Washington University School of Medicine,
which emphasizes that surgical pathology at Washington University has always been a
collaborative venture between the faculty and trainees. The Department's Chairman,
Dr. Herbert (Skip) Virgin, has not hesitated to continue to provide the support nec-
essary to maintain the tradition of diagnostic excellence, academic productivity, and
education that forms the foundation of academic surgical pathology at our institution.
We extend special thanks to our administrative assistants, Elease Barnes, Jeannie
Doerr, and Shari Jackson, who handled most of the logistical and secretarial work
required to make the second edition a reality. We also acknowledge the artistic expertise
of the staff at MedPIC (the medical illustration service of Washington University School
of Medicine), especially Marcy Hartstein and Vicki Friedman, who together revised the
figures for the book. Walter Clermont expertly and patiently edited all the electronic
figures found at the associated website. We are lucky to continue to have had the
opportunity to work with several wonderful people at Lippincott William & Wilkins,
including Jonathan W. Pine, Jr. (who enthusiastically suggested at an early stage that
we produce a second edition of the manual), Emilie Moyer, Marian Bellus, and Martha
Cushman.
And our families. Their love, patience, and support made the book possible, and
make it all worthwhile.
Peter A. Humphrey
Louis P. Dehner
John D. Pfeifer
xi
Contents
Contributors v
Preface x
Acknowledgments xt
I HEAD AND NECK 1

Oral Cavity and Oropharynx 1
• Rebecca D. Chernock and James S. Lewis Jr.
- . Larynx 19
Iii James S. Lewis Jr.
Ell Nasal Cavity, Paranasal Sinuses, and
. . Nasopharynx 34
Heather N. Wright and James S. Lewis Jr.
Tumors and Cysts of the Jaws

II David E. Spence and Samir K. EI-Mofty
51
Ell The Eye 63

. . George J. Harocopos
El Salivary Glands 81
. . Joshua I. Warrick, Elise L. Krejci, and James S. Lewis Jr.
Cytopathology of the Salivary Glands 95
Brian Collins
The Ear 97
Ill Peter A. Humphrey and Rebecca D. Chernock
II THORAX 104
Ell Lung 104
. . Jon H. Ritter and Hannah R. Krigman
Cytopathology of the Lung 145
Hannah R. Krigman
Ell Cardiovascular System 149

. . Jochen K. Lennerz and John D. Pfeifer
lli1 Mediastinum 165

. . Louis P. Dehner
xii i
xiv I CONTENTS
Serosal Membranes 177

Jon H. Ritter and John D. Pfeifer
Ill Gl TRACT 190

n;J The Esophagus 190
Ill Danielle H. Carpenter and Elizabeth M. Brunt
Cytopathology of the Esophagus 199
Julie Elizabeth Kunkel
IE1 The Stomach 201

. . Kathryn M. Law and Elizabeth M. Brunt
Cytopathology of the Stomach 212

The Intestines, Appendix, and Anus 214

II ILKe Nalbantoglu and Elizabeth M. Brunt
1m The Liver 248

. . Ta-Chiang Liu and Elizabeth M. Brunt
Cytopathology of the Liver 271

Julie Elizabeth Kunkel and Hannah R. Krigman
IT:I The Gallbladder and Extrahepatic

. . Biliary Tree 274
Ta-Chiang Liu and Elizabeth M. Brunt
Cytopathology of the Gallbladder and Extrahepatic Biliary Tree 279

Brian Collins and Julie Elizabeth Kunkel
IR The Pancreas 282

. . Dengfeng Gao and Hanlin L. Wang
Cytopathology of the Pancreas 294

IV BREAST 298
11:1 Breast Pathology 298
. . Souzan Sanati, Omar Hameed, Joshua I. Warrick,
and Craig Allred
Cytopathology of the Breast 326

Lourdes R. Ylagan
CONTENTS I XY
V URl NARY TRACT 329

11!1 Medical Diseases of the Kidney 329
. . Joseph P. Gaut and Helen Liapis
~~~ Surgical Diseases of the Kidney 344
Maldevelopment and Nonneoplastic

Cystic Diseases 344
Johann D. Hertel, Peter A. Humphrey, and Helen Liapis
Pediatric Renal Neoplasms 347
Jason A. Jarzembowski and Frances V. White
Adult Renal Neoplasms 357
Maria F. Serrano and Peter A. Humphrey
Cytopathology of Renal Neoplasms 371
Souzan Sanati
OJ Renal Pelvis and Ureter 373

Ill Souzan Sanati, Frances V. White,
and Peter A. Humphrey
r;J;J The Urinary Bladder 379

IIIII Omar Hameed and Peter A. Humphrey
Cytopathology of the Urinary Bladder 396
Rosa M. Davila
&1 Urethra 399

IIIII Souzan Sanati and Peter A. Humphrey
VI ENDOCRINE SYSTEM 404

tm Thyroid 404
. . Changqing Ma, James S. Lewis Jr., and Rebecca D. Chernock
Cytopathology of the Thyroid 418
Michael E. Hull and Julie Elizabeth Kunkel
rwt Parathyroid Glands 422

IIIII James S. Lewis Jr.
Cytopathology of the Parathyroid Glands 427
Hannah R. Krigman
r;r:t The Adrenal Gland and Paraganglia 428

IIIII Louis P. Dehner
I CONTENTS
xvi
DJ Pituitary Gland 446

. . Richard J. Perrin, Sushama Patil, and Arie Perry
VII REPRODUCTIVE TRACT 451

Jm1 Testis and Paratestis 451
liilill Kiran R. Vij and Peter A. Humphrey
1WJ Prostate 467
1111 Peter A. Humphrey
ETi1 Penis and Scrotum 483
. . Peter A. Humphrey
Ell The Ovary 493

. . Meredith E. Pittman, John D. Pfeifer, and Phyllis C. Huettner
EJ;J Fallopian Tube 511

IIIII Mitra Mehrad, John D. Pfeifer, and Phyllis C. Huettner
Ea Uterus (Corpus) 517
. . Jena Beth Hudson, John D. Pfeifer, and Phyllis C. Huettner
Ell Uterine Cervix 535

. . Michael E. Hull
Cytopathology of the Uterine Cervix 547
Michael E. Hull
Em Vagina 551
. . Rao Watson, John D. Pfeifer, and Phyllis C. Huettner
ET:I Vu Iva 558

. . Danielle H. Carpenter, John D. Pfeifer, and Phyllis C. Huettner
E6 Placenta 566
Iiiii Phyllis C. Huettner
VIII SKIN 579
Ern Skin: Nonneoplastic Dermatopathology 579
. . Samuel J. Pruden II, Kimberley G. Crone, and Anne C. Lind
Em Nonmelanocytic Tumors of the Skin 595

llflll Nathan C. Walk, Yumei Chen, Anne C. Lind,
Friederike Kreisel, and Dongsi Lu
CONTENTS xvii
twi1 Skin: Melanocytic Lesions 613

1M Anne C. Lind, Nils Becker, Emily A. Bantle, and Louis P. Dehner
~ NERVOUSSYSTEM 625
HI Central Nervous System: Brain, Spinal Cord,
1M and Meninges 625
Richard J. Perrin, Sushama Patil, and Arie Perry
Cytopathology of the Central Nervous System 671
Souzan Sanati and Lourdes R. Ylagan
R;J Nerve Biopsies 673

Ill Robert E. Schmidt
X HEMATOPOIETIC SYSTEM 683

lEI Lymph Nodes 683
1M Anjum Hassan and Friederike Kreisel
Lymph Node Cytopathology 706
Ill Bone Marrow Pathology 709

. . Jeffery M. Klco and John L. Frater
~ Spleen 729
1M Mohammad 0. Hussaini and Anjum Hassan
XI SOFT TISSUE AND BONE 739

~"fit Soft Tissue 739
1M John D. Pfeifer and Louis P. Dehner
1"6 Retroperitoneum 781
. . Catalina Amador-Ortiz, Louis P. Dehner,
and John D. Pfeifer
1m Bone Neoplasms and Other Nonmetabolic

1M Disorders 790
Omar Hameed and Michael J. Klein
1m Metabolic Diseases of Bone 816

1M Deborah Novack
xviii I CONTENTS
er.1 Joints and Synovium 822

XII CYTOPATHOLOGY 828

ell General Principles of Cytopathology 828
1M Brian Collins
XIII ANCILLARY METHODS 832
1WJ Frozen Sections and Other Intraoperative
Ill Consultations 832
Michael E. Hull, Peter A. Humphrey, and John D. Pfeifer
ft1 Electron Microscopy 838

. . Ashima Agarwal and Frances V. White
ell Histology and Histochemical Stains 842

. . Kevin Selle
6Et Immunohistochemistry 851

er:t Immunofluorescence 856

. . Anne C. Lind, Joseph P. Gaut, and Rosa M. Davila
13J Flow Cytometry 859

. . Friederike Kreisel
eJ!1 Cytogenetics 863

lilill Shashikant Kulkarni, Hussam AI-Kateb, and Catherine Cottrell
1K!1 Fluorescence in Situ Hybridization 876
1M TuDong Nguyen, Arie Perry, and Anjum Hassan
r:ti1 Direct and Indirect Methods for DNA
. . Sequence Analysis 887
TuDong Nguyen, Barbara Zehnbauer, and John D. Pfeifer
r:n Microarrays 903

1M Mark Watson
CONTENTS I xix
rwJ Biospecimen Banking 914

lilliiil Mark Watson
r:ra Imaging Technologies in Surgical Pathology:
Ill Virtual Microscopy and Telepathology 926
Jochen K. Lennerz, Michael Isaacs, Erika Crouch, and John D. Pfeifer
Index 931
SECTION I
Head and Neck
Oral Cavity and

Oropharynx
Rebecca D. Chernock and James S. Lewis Jr.
I. NORMAL ANATOMY
A. Oral cavity. The anterior aspect of the oral cavity extends from the mucocu-
taneous junction (vermilion border) of the lips to include the buccal mucosa
(inside of cheek), maxillary and mandibular arches (teeth), retromolar trigone,
anterior two-thirds of the tongue (oral tongue), floor of mouth, and hard palate.
Posteriorly, the oral cavity freely communicates with the oropharynx; the border
between the two is marked by the junction of the hard and the soft palate superi-
orly and the line of circumvallate papillae on the dorsal tongue (border between
the anterior two-thirds and posterior one-third of the tongue) inferiorly.
1. The oral tongue is freely mobile and composed mainly of skeletal muscle. It
has a dorsal (exposed) surface, ventral surface, and tip. The dorsal surface
contains numerous papillae that have specialized taste receptors. The floor
of the mouth lies beneath the tongue and is divided into sides by the midline
frenulum (mucosal fold) of the tongue. It contains ostia of the submandibu-
lar and sublingual salivary glands, whereas the main duct of the parotid
gland (Stensen's duct) enters the oral cavity through the buccal mucosa. The
hard palate forms the roof of the oral cavity and consists of portions of the
maxillary and palatine bones.
2. The oral cavity is lined by stratified squamous mucosa with prominent
mucoserous glands in the submucosa. Most of the mucosa is nonkeratiniz-
ing with the exception of the hard palate, gingiva, and dorsal tongue, which
become keratinized due to the friction of mastication.
B. Oropharynx. The oropharynx is the space posterior to the oral cavity that com-
municates with the nasopharynx superiorly and the larynx and hypopharynx
inferiorly. The soft palate (posterior one-third of the palate) marks the superior
aspect of the oropharynx and is suspended from the posterior aspect of the hard
palate. In contrast to the hard palate, the soft palate is fibromuscular without a
bony skeleton. Whereas the oral aspect is covered with nonkeratinizing stratified
squamous epithelium, the nasal surface is covered by pseudostratified ciliated
columnar (respiratory-type) epithelium. Numerous mucoserous glands lie in its
submucosa. The uvula extends down from the posterior aspect of the soft palate
and has an identical histology. The posterior one-third of the tongue is rich in
lymphoid tissue (known as the lingual tonsil). The palatine and lingual tonsils,
together with the pharyngeal tonsil in the nasopharynx, are collectively known
as Waldeyer's ring.
2 I SECTION 1: HEAD AND NECK
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION

A. Open and endoscopic biopsies. The majority of specimens from the oral cav-
ity and oropharynx consist of open biopsies of lesions that can be visualized
by the naked eye. The small biopsy pieces should be placed immediately into
10% buffered formalin or other appropriate fixative. Processing of the biopsies
should include gross description of the tissue fragments with documentation of
the number of pieces present; the biopsies should be entirely submitted, with
three levels cut from each paraffin block for hematoxylin and eosin (H&E)
examination.
B. Resections
1. Open procedure specimens are widely variable depending on the location
of the tumor. Because many lesions encroach upon or invade the bone of
the mandible or maxilla, composite resections with bone and soft tissue are
common. As a generalization, all specimens need to be oriented appropriately,
the soft tissue margins inked, and mucosal and soft tissue margins evaluated
followed by sectioning of the tumor relative to cartilage/bone and tissue
margins. Margins should be evaluated by either shave or radial sections,
depending on the nature of the specimen. If the tumor is relatively distant
from a margin, 1- to 2-mm shave sections are preferred. If the tumor grossly
approximates a margin to <1 to 2 mm, radial sections should be taken.
2. Partial resections with a C02 laser under an operating microscope are becom-
ing more common. Because the inherent approach of this procedure is to
excise the tumor piece by piece, the surgeon inks the individual pieces as
he/she alone knows what constitutes the true margin. In the gross room all
that is therefore required is to measure the pieces provided, describe them,
and submit them entirely in sections perpendicular to the ink. If there is an
additional orienting marker such as a suture, then one should submit the
sections sequentially from that end to the opposite end.
C. Frozen sections are a critical element of surgical therapy for tumors of the head
and neck region. Although practices vary, at most institutions shave margins
are taken by the surgeon from the periphery of the surgical defect after the
tumor has been removed, and separately submitted. In other cases, the surgeon
may sample the tumor or suspicious sites to confirm and/or map the lesion,
with additional samples from areas where the tumor is felt to be closest to the
surgical margin.
For frozen sections, the tissue is submitted in saline to the pathology lab
and then frozen in its entirety. The tissue pieces should be evaluated grossly for
mucosa (typically the shiny and pink-tan surface of the tissue) and, if mucosa is
present, the specimen should be oriented in the frozen section block to demon-
strate this surface on edge. It is critical to cut deeply into the block to obtain
sections that represent the entire tissue submitted so that small foci of tumor
are not missed by inadequate sampling. At our institution, we cut three rather
than two H&E frozen section slides to reduce these sampling errors.
The tissue that remains after frozen section is submitted for evaluation by
permanent sections, which helps assure adequate sampling. Permanent sections
can help resolve a number of issues from frozen section including freezing and
cautery artifact, volume of tumor, and orientation (note that the margins of
the main resection specimen should also be evaluated throughout their entirety
because the separate frozen section specimens almost never cover the entire mar-
gins of a resection specimen). The final margin status is then a conglomerate of
three sources: frozen section slides, permanent slides of the frozen tissue, and the
tissues not submitted for frozen section, and the margins of the main specimen.
Ill. DIAGNOSTIC FEATURES OF COMMON BENIGN DISEASES
A. Inflammation. Lichen planus, pemphigus vulgaris, and cicatricial pemphigoid are
autoimmune disorders that predominately affect middle-aged adults and occur
Chapter 1 • Oral Cavity and Oropharynx I 3
more frequently in women than men. They are diseases that affect mucosal sites
as well as skin, so the oral cavity is sometimes involved as well.
1. Lichen planus. This disorder commonly affects the oral mucosa. Whereas
skin involvement is usually self-limited, oral lichen planus follows a more
protracted waxing and waning course. The oral lesions are typically asymp-
tomatic unless ulceration occurs.
Any oral mucosal surface may be involved, and several patterns can be
seen. The classic pattern is reticular with intersecting white keratotic streaks
(Wickham's striae); the lesions are ill-defined, and the background may be
erythematous due to mucosal atrophy. Some lesions may be mostly erythema-
tous with minimal keratotic streaks, whereas others may show extensive ker-
atinization and/or ulceration or form bullae. Microscopically, a dense submu-
cosal band of lymphocytes is present, which may be less distinct in ulcerated
lesions (e-Fig. 1.1)."' The rete ridges may be hyperplastic (saw-toothed) or
flattened. There is loosening of the basal layer of the epithelium, with degener-
ation of individual keratinocytes that may form eosinophilic colloid (dysker-
atotic, cytoid, or Civatte) bodies (e-Fig. 1.2). The surface may show hyper-
or parakeratosis. Although the histologic findings of lichen planus have been
well defined, they are nonetheless not specific; for example, some oral lesions
with a similar histologic picture may be due to a contact hypersensitivity reac-
tion. In addition, a lichenoid infiltrate may accompany dysplastic lesions.
Asymptomatic patients require no treatment, but steroids (particularly
topical) are often used for erosive or erythematous lesions. Some data sug-
gest that there is an increased risk of malignant transformation in the erythe-
matous, ulcerative, and bullous forms of oral lichen planus. Although this
proposed risk of malignancy is controversial, these lesions at least require
closer clinical follow-up.
2. Pemphigus vulgaris. This is an uncommon disorder that causes superficial
ulceration of the skin and mucous membranes. Involvement of the oral
mucosa may precede the development of skin lesions. The disease is caused
by autoantibodies to desmogleins 1 and 3, cellular transmembrane proteins
involved in the assembly of desmosomes. Cell-to-cell adhesion is impaired
in the suprabasal epithelium, leading to clefting and ulceration. Flaccid bul-
lae that easily rupture to form painful erosions can be seen on any oral
mucosal surface. The lesions heal without scarring. Microscopically, intraep-
ithelial separation with edema and acantholysis, which imparts a "tomb-
stone" appearance to the remaining attached basal cell layer (e-Fig. 1.3),
is seen at the edge of the ulce1:. Acute and chronic inflammation are fre-
quently present in the submucosa. Direct immunofluorescence is positive for
immunoglobulin G (lgG) along cell membranes throughout the epidermis.
Paraneoplastic pemphigus, which is associated with an underlying malig-
nancy, may be distinguished from pemphigus vulgaris by the identification
of a different pattern of antibody staining by direct immunofluorescence.
3. Cicatricial or mucous membrane pemphigoid. This is a rare disease caused by
various antibodies that target the basement membrane of mucous membranes
and occasionally the skin. The oral mucosa is almost always involved, most
commonly the gingiva. In contrast to pemphigus vulgaris, the variably sized
bullae are not flaccid, and ruptured bullae heal with scarring. Microscopi-
cally, there is clefting between the epithelium and the basement membrane,
and the space may be filled with serous fluid containing sparse inflammatory
cells. Direct immunofluorescence shows a linear band of IgG and C3 on the
"'All e-figures are available online via the Solution Site Image Bank.
basement membrane. Treatment is with immunosuppression, but the disease

is often progressive despite therapy.
B. Infections. Only a few of the numerous infections that may involve the oral
cavity are discussed here.
1. Fungal. Candida species cause most of the fungal infections of the oral cavity.
Other fungal infections that occur less frequently in the oral cavity include
histoplasmosis, blastomycosis, and coccidiomycosis. Candida species are a
part of the normal oral flora; candidiasis occurs due to overgrowth, usually
in the setting of a predisposing factor, and Candida albicans is the most
frequently isolated species. Local and systemic factors that favor overgrowth
include immunosuppression, use of steroids or antibiotics, radiation therapy,
xerostomia, use of dentures, and anemia. The extremes of age are more often
affected as well, and infection may be acute or chronic. Symptoms include a
burning sensation or foul odor, although the infection may be asymptomatic.
Several clinical patterns of oral candidiasis are seen. White plaques that
are easily scraped off underlying erythematous mucosa are called pseu-
domembranous candidiasis (oral thrush). This is the most common type of
oral candidiasis. Erythematous candidiasis appears as a red patch due to
atrophy of the mucosa. Median rhomboid glossitis is a type of erythematous
candidiasis that occurs in a specific location, namely a rhomboid-shaped
area on the midline dorsal tongue, which over time may develop a nodular
appearance. Angular cheilitis causes red fissuring and scaling at the labial
commissures; predisposing factors include drooling and ill-fitting dentures.
Chronic hyperplastic candidiasis presents as asymptomatic, white patches
(due to the thickened, hyperplastic mucosa) that cannot be removed by scrap-
ing. This pattern is more common in immunocompetent individuals and may
predispose to the development of carcinoma, although a causal relationship
between the two has not been clearly demonstrated.
Microscopically, an intraepithelial infiltrate of neutrophils is seen in all
types of candida! infection. The epithelium may be ulcerated, although
in chronic hyperplastic candidiasis it is thickened and hyperkeratotic
(e-Fig. 1.4). Fungal pseudohyphae may be difficult to identify on routine
H&E-stained slides. However, methenamine silver or periodic acid-Schiff
(PAS) stains will highlight the fungal elements within the keratin and in the
superficial squamous epithelium (e-Fig. 1.4, inset A).
2. Viral. Oral viral infections are highly prevalent, although frequently asymp-
tomatic.
a. Human papillomavirus {HPV) does not cause specific clinically symptomatic
infection but is associated with several benign and malignant neoplasms
in the oral cavity and oropharynx including squamous papillomas and
squamous cell carcinoma (SCC) (see respective sections to follow).
b. Herpes simplex virus {HSV} causes a common oral viral infection with sero-
prevalence rates of up to 80% of the population. There are two common
serotypes (HSV-1 and HSV-2), and HSV-1 is primarily associated with
oral lesions. Gingivostomatitis occurs in 10% of initial infections, pre-
dominately in children, and is characterized by fever and a vesicular rash.
The virus then latently infects sensory ganglia and may be reactivated
periodically throughout life. Recurrent disease is manifested by clusters
of vesicles that may cause a burning sensation at the mucocutaneous junc-
tion of the lip or the nose. Intraoral lesions can also occur.
Microscopically, the lesional mucosa is often ulcerated and acan-
tholytic, with marked acute and chronic inflammation. There are typically
individual necrotic squamous cells. Identification of the classic intranu-
clear eosinophilic inclusions within squamous epithelial cells is diagnostic
of herpes virus infection; the inclusion-harboring cells are often single,
detached, and multinucleated with molding of the nuclei to each other

(e-Fig. 1.5). Immunohistochemical stains may be useful to confirm the
diagnosis.
c. Epstein-Barr virus (EBV). Acute EBV infection, although frequently asymp-
tomatic, may cause pharyngitis and tonsillitis. The virus enters the host
through oral epithelial cells, where it then gains access to and infects B
lymphocytes. Acute EBV infection may produce reactive changes in the
tonsils and lymph nodes that can mimic a hematopoietic malignancy.
Latent EBV infection is virtually universal in adults and is usually
asymptomatic. However, an EBV-driven proliferation of tongue epithe-
lial cells, known as oral hairy leukoplakia, occurs in immunosuppressed
patients, approximately 80% of whom are HIV positive. The lesions are
asymptomatic unless superinfection with Candida occurs. Grossly, hairy
leukoplakia appears as a flat, white, shaggy plaque on the lateral tongue.
Microscopically, the epithelium is acanthotic with hyper- and parak-
eratosis. Perinuclear clearing forming "balloon cells" is characteristic
(e-Fig. 1.6), and viral replication rna y cause "nuclear beading" (e-Fig. 1. 7).
Inflammation is typically sparse. Definitive diagnosis relies on detection of
EBV within the lesion by immunohistochemistry or in situ hybridization
(e-Fig. 1.8). Oral hairy leukoplakia is self-limited with no propensity for
malignant transformation.
Latent EBV infection has been implicated in the development of a
variety of hematopoietic and nonhematopoietic malignancies as well (see
Chaps. 3 and 43).
3. Bacterial. Cervicofacial actinomycosis ('•lumpy jaw"). Actinomyces are gram
positive, saprophytic anaerobes that are part of the normal oral flora. The
organisms are often incidentally found in sections of the tonsillar crypts.
Occasionally, they are introduced into the soft tissues through trauma, par-
ticularly from dental manipulations, where an acute or chronic infection may
ensue. Actinomyces israeli is the most common pathogenic species.
Acute infections are suppurative, creating a nontender fluctuant mass.
In the chronic phase, infections may form a more extensive firm fibrous
mass mimicking a neoplasm. Sinus tracts may exit either the skin or mucosa
(e-Fig. 1.9) and often discharge yellow clusters of tightly adherent Actino-
myces bacteria that have the appearance of sulfur granules. Osteomyelitis
may develop in adjacent bone. Microscopically, collections of radiating, fila-
mentous organisms are seen in a background of neutrophils with surrounding
granulation tissue and/or fibrosis. Cultures are often negative due to over-
growth of other organisms. Treatment with prolonged antibiotics is usually
successful, although incision and drainage may be necessary.
C. Other non-neoplastic lesions
1. Fibrous lesions. A number of different types of fibrous lesions occur in and
around the oral cavity.
a. Irritation fibroma. These are the most common oral mucosal mass lesions.
They are painless reactive proliferations of fibrous tissue that develop in
response to trauma from teeth or dentures. The lateral tongue and buccal
mucosa along the bite line are the most common sites. Multiple fibro-
mas may be seen in inherited syndromes including Cowden' syndrome
and tuberous sclerosis. Linear, grooved fibromas occurring in the mucosa
opposing the teeth or sulcus of the alveolar ridge are called epulis fissur-
atum and are denture-related.
Grossly, irritation fibroma is usually pink to white, dome-shaped, and
only a few millimeters in maximal diameter. Microscopically, there is a
nodular deposition of dense collagen with associated chronic inflamma-
tion and overlying thinned mucosa (e-Fig. 1.10). Trauma-related changes
such as hyperkeratosis and ulceration may be seen. The fibroblasts are

spindled and indistinct. If larger, stellate fibroblasts are present, the lesion
is called a giant cell fibroma, which, in contrast to irritation fibromas,
is not associated with trauma and occurs at a younger age (e-Fig. 1.11).
Whereas typical irritation fibromas do not recur after simple resection,
giant cell fibromas may recur.
b. Gingival fibromatosis. This is generalized, but not necessarily symmetrical,
enlargement of the gingiva which may be hereditary, drug-induced, related
to poor oral hygiene, or idiopathic. When it is drug-induced, it is called
fibrous gingival hyperplasia and frequently regresses with cessation of the
inciting drug.
Grossly, the gums are enlarged, smooth-surfaced, and firm. Micro-
scopically, the submucosa shows dense eosinophilic to slightly basophilic
fibrous tissue with associated mild chronic inflammation (e-Fig. 1.12). The
surface squamous epithelium may have chronic inflammation or extreme
elongation of the rete, but is otherwise unremarkable.
2. Inflammatory papillary hyperplasia is a denture-associated lesion and is typi-
cally located beneath a denture base in the hard palate and alveolar ridges.
Occasionally, it is seen in patients without dentures and may be associated
with poor oral hygiene.
Clinically, the mucosa looks "pebbly" with numerous, small, papular pro-
jections. Microscopically, the mucosa may be atrophic or demonstrate pseu-
doepitheliomatous hyperplasia. The underlying submucosa may vary from
edematous to fibrotic, with mild chronic inflammation. Individual nodules
may resemble an irritation fibroma or pyogenic granuloma. The condition
is not premalignant, may subside with less denture weat; or may require
surgical excision.
3. Torus palatinus, torus mandibularis, and buccal exostosis are common develop-
mental anomalies that continue to grow throughout life and typically present
in adulthood. They are site-specific. Torus palatinus occurs in the midline
of the hard palate, torus mandibularis occurs on the lingual surface of the
mandible near the bicuspid teeth, and buccal exostosis is found on the facial
surface of the alveolar bone. Any identical appearing bony proliferations at
other oral sites are generically termed "bony exostosis" or "osteoma" and
are not developmental, but rather trauma-related or true neoplasms that can
be associated with Gardner syndrome.
Grossly, these lesions are broad-based, single, or lobulated masses with
smooth surfaces (e-Fig. 1.13). Microscopically, they are composed of dense
lamellar bone with scattered osteocytes and variable amounts of marrow.
Ischemic changes with marrow fibrosis and loss of osteocytes from lacunae
may be seen. Resection is not necessary except for cosmetic reasons or if the
lesions become large. There is little risk of recurrence, and the lesions have
no malignant potential.
4. Fordyce granules. Sebaceous glands are normally found in the skin associated
with hair follicles. When they are ectopically present in the oral mucosa,
they are called Fordyce granules. They are common (present in up to 80%
of adults) and can occur on any oral mucosal surface, although the buccal
mucosa is most common. Most appear as scattered, 1 to 3 mm, white to
yellow papules. Microscopically, normal sebaceous glands are present in the
submucosa without associated hair follicles; occasionally, Fordyce granules
coalesce to form the larger cauliflower-like lesion termed sebaceous hyper-
plasia. No treatment is necessary unless for cosmetic reasons (biopsies are
rarely performed as the diagnosis is usually clinically apparent).
5. Cysts. Included here are several of the more common soft tissue true cysts
that have an epithelial lining. Odontogenic cysts, bone cysts, and salivary
gland-derived pseudocysts (lacking a true epithelial lining) are discussed in

Chapter4.
a. Epidermoid cyst. Intraoral epidermoid cysts are much less common than
their counterparts in the skin and are thought to represent inclusions
of surface epithelium or cystic change in odontogenic rests. They often
present in teenagers and young adults. The most common site is the gin-
giva. Clinically, they are small (< 1 em) superficial nodules. When they
occur in the midline floor of mouth, howevet; they can become much
larger (>5 em) and interfere with swallowing. Microscopically, they are
lined by thin stratified squamous epithelium, with or without a granular
celllayet; and are often filled with keratinous debris. Rupture with spillage
of keratinous debris may elicit a granulomatous inflammatory reaction.
Treatment is by simple surgical excision.
b. Dermoid cysts. These are similar to epidermoid cysts but contain adnexal
structures, such as sebaceous glands or hair follicles, in the cyst wall. If
other tissue types are present, the lesion is termed teratoid cyst.
c. Nasolabial cysts. These rare cysts occur at the base of the nostril or at
the superior aspect of the upper lip. They are thought to be derived from
remnants of the embryonic nasolacrimal duct. Seventy-five percent of cases
occur in women. More than 10% are bilateral. They present as slow
growing masses, usually < 1.5 em, and they may have irregular contours.
Soft tissue swelling with loss of the nasolabial fold or elevation of the nasal
ala or floor may occasionally cause nasal obstruction. Pressure erosion of
underlying bone is possible.
Microscopically, the cysts may be lined by respiratory type, cuboidal,
and/or stratified squamous epithelium with scattered mucus-filled gob-
let cells, with surrounding chronic inflammation. A fibrous or epithelial
connection to the nasal mucosa is almost always present. Simple surgical
excision is curative.
d. Lymphoepithelial cysts. These cysts are thought to develop from invagina-
tions of crypt epithelium within accessory tonsillar tissue. Clinically, they
are painless submucosal nodules that are almost always <6 mm in diam-
eter and typically occur in teenagers or young adults. Half of cases occur
in the floor of the mouth. The lateral and ventral tongue, as well as the
soft palate, are also common sites. They do not occur in the alveolar soft
tissue.
Microscopically, the cyst lining is an attenuated squamous epithelium
with a poorly formed granular layer. The cyst is filled with orthokeratin
and surrounded tightly by lymphoid aggregates with variable numbers of
germinal centers. The cysts may become dissociated from the epithelium
or remain connected, often with keratin plugging. Microscopically, the
prominent lymphoid aggregates distinguish this cyst from an epidermoid
cyst. Similar appearing cysts can be seen within the tonsils themselves from
blockage of the crypt connection with the surface.
6. Pseudoepitheliomatous hyperplasia is a generic term for benign, downward
proliferation of the epithelium that is important to distinguish from invasive,
well-differentiated SCC. Pseudoepitheliomatous hyperplasia is characteristi-
cally seen in association with specific lesions including inflammatory papil-
lary hyperplasia, submucosal granular cell tumors, and fungal infections. It
can also be seen adjacent to ulcers or be seen associated with myriad other
lesions. Microscopically, irregular and pseudoinfiltrative nests of keratinizing
squamous epithelium are sometimes seen beneath a markedly thickened sur-
face epithelium. The papillae of the squamous epithelium may be markedly
elongated and extend deeply into the submucosa (e-Fig. 1.14). Howevet; in
contrast to squamous dysplasia and carcinoma, cytologic atypia is absent.
I I SECTION •• HEAD AHO HECK
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Squamous papillomas are strongly associated with the nononcogenic

HPV types 6 and 11, but have a very low infectivity so do not appear con-
tagious. In the oral cavity and oropharynx, they occur in adults between
30 and 50 years of age, most commonly on the hard and soft palate and
the uvula.
Squamous papillomas have a characteristic morphology. Grossly, they
are soft, exophytic, granular, and pink-red or tan. Microscopically, they
consist of arborizing, papillary fronds of thickened but maturing squa-
mous epithelium with nuclei that are slightly enlarged and irregular but
not overtly dysplastic. Mitotic activity is usually present but is modest.
Sometimes there is slight hyperplasia of the basal layer. Cells in the mid-
layer often have cytoplasmic clearing, but frank koilocytosis is not regu-
larly observed. Although typically absent, surface keratinization may be
seen (e-Fig. 1.17).
Squamous papillomas are cured by simple excision. They have limited
growth potential, rarely recur, and have essentially no risk of malignant
transformation.
b. Condyloma acuminatum and verruca vulgaris. Both of these lesions, although
much more common on the skin, can also occur in the oral cavity. Condy-
lomas are considered a sexually transmitted disease and usually occur in
young adults on the lips and soft palate as clusters of pink nodules that
coalesce into more exophytic masses. They have more blunted, papillary
fronds and more hyperkeratosis than squamous papillomas.
Verrucae also can occur intraorally, particularly in children, commonly
on the lips and anterior tongue. They have a morphology identical to that
of verrucae of the skin with a broad base, marked papillomatosis, and
hyperkeratosis with parakeratosis.
c. Verruciform xanthoma. This is a peculiar lesion of the oral cavity which has
no relation to HPV and may be reactive in nature. It occurs in middle-
aged to older adults and is most common on the alveolar ridges. Clinically
and grossly, it is a well-demarcated, painless, and soft, slightly elevated
mass. It may have a yellow or white color with a roughened surface.
Microscopically, it consists of broad papillae with intervening cleft-like
spaces covered by a slightly thickened, nondysplastic squamous epithelium
with hyperkeratosis and parakeratosis. The diagnostic cells, which lie in
the superficial submucosa, are foamy macrophages with abundant pale,
flocculent cytoplasm and round to oval, bland nuclei (e-Fig. 1.18). These
cells are filled with lipid and should be distinguished from the eosinophilic
granular cells of granular cell tumor (see Section E to follow).
Verruciform xanthoma is treated by conservative excision and has no
risk of malignant transformation. Recurrences are rare.
2. Precursor (premalignant) squamous lesions. Precursor lesions are defined as
altered squamous epithelium with an increased risk of progression to sec,
and are strongly associated with smoking and alcohol use. They may present
as leukoplakia (white, thickened epithelium; see e-Fig. 1.19), erythroplakia
(thin, erythematous, and red epithelium), or speckled erythroplakia (a mix-
ture of both erythroplakia and leukoplakia).
The terms dysplasia or intraepithelial neoplasia should be used for these
lesions. The likelihood of malignant change relates to the severity of dyspla-
sia, although carcinoma can develop from any grade of dysplasia (as well as
from normal epithelium). Atypia, on the other hand, is not considered syn-
onymous with dysplasia and is used in a more general sense because the
term may also describe changes seen in reactive epithelium. There are a
number of changes that occur in dysplasia including nuclear abnormalities,
architecturaVorganizational abnormalities, and abnormal keratinization.
1D I SECTION I. HEAD AHD NECK
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primary surgery or primary chemoradiotherapy. Approximately 25% are

related to high risk HPV, and these have a more favorable prognosis. For
the non-HPV related tumors, the prognosis is slightly worse than for such
tumors in the oral cavity.
b. Nonkeratinizing SCC. This entity is slowly gaining recognition as a spe-
cific subtype of head and neck SCC. Tumors with this morphology are
almost exclusively seen in the palatine tonsils and base of tongue and are
virtually always HPV-associated (high risk HPV in >95% of cases and
specifically type HPV16 in >90% of such cases). Patients with nonkera-
tinizing sec are, on average, 5 years younger than those with conventional
keratinizing-type SCC. A significant minority are non-smokers, and those
that do smoke are less likely to be heavy smokers. The most common
clinical presentation is as an asymptomatic neck mass. Nonkeratinizing
sec arises in the crypts of the tonsillar tissue, so the tumors tend to be
subsurface and thus clinically subtle or totally hidden. This feature, com-
bined with the tumor•s strong propensity for early metastasis to cervical
lymph nodes, explains why the most common presentation is as a painless
neck mass. Better recognition of this tumor type has led to more intensive
scrutiny of the tonsils and base of tongue in this clinical setting.
Grossly, nonkeratinizing SCC is endophytic, firm, and tan in most
cases. However, because the tumors are often small and do not elicit
much desmoplasia in the surrounding stroma, they can be quite difficult
to identify grossly. Microscopically, they consist of ribbons of tumor cells
lining the crypt epithelium and of nests and sheets of cells with smooth
borders in the submucosa. The overlying surface epithelium is typically
intact without dysplasia (e-Fig. 1.23). The cells are basal in appearance,
with round to oval to spindled nuclei, relatively homogeneous chromatin
without prominent nucleoli, minimal cytoplasm, and very brisk mitotic
activity with abundant apoptosis (e-Fig. 1.24). Central (comedo) necrosis
is common in the tumor cell nests.
In situ hybridization and/or polymerase chain reaction (PCR) are pos-
itive for HPV16 or other high-risk HPV types in almost all cases. More
recently, assays specific for high-risk HPV RNA transcripts have been
applied which are almost always positive in this tumor type. Immunohis-
tochemistry is positive for cytokeratins and for p16 (a tumor suppressor
protein that is aberrantly overexpressed in cells infected by HPV) (e-Fig.
1.25). Strong and diffuse p16 immunoreactivity has become increasingly
recognized as a clinically useful surrogate marker for HPV-driven SCCs of
the oropharynx. Many use this as a single risk stratification test in clinical
practice because, when positive, it indicates a tumor with a much more
favorable prognosis than for head and neck sec in general.
Treatment is most often by surgical resection with neck dissection fol-
lowed by postoperative radiation therapy, although primary chemoradia-
tion therapy is a very effective approach as well. Numerous studies have
shown that the prognosis for HPV-related SCC of the oropharynx is bet-
ter than for keratinizing type sec, despite the fact that tumors commonly
present with lymph node metastases and thus are high stage.
c. Verrucous carcinoma (VC) is a specific, well-differentiated and nonmetas-
tasizing variant of sec. It occurs in the larynx and oral cavity and grossly
appears as a well-circumscribed, warty and exophytic, broad-based, white
or tan mass (e-Fig. 1.26). It can become very large and dramatically invade
soft tissues and bone. Microscopically, VC consists of very thick sur-
face squamous epithelium with club-shaped papillae that have a broad
pushing base. These blunt and downward-pushing projections have some-
times been likened to elephant's feet. There is usually prominent surface
hyperkeratosis, and the sheets of tumor are composed of bland cells with
abundant eosinophilic to clear cytoplasm, sometimes described as glassy
in appearance. There is no cytologic atypia. The stroma directly beneath
the tumor typically demonstrates prominent chronic inflammation, often
with abundant plasma cells (e-Fig. 1.27).
Pure VC is a tumor type in which the cells have not invaded through
the basement membrane, and as such has an excellent prognosis. Although
the tumor can be large and locally destructive, complete surgical resection
is often curative because the tumor cells cannot spread beyond the local
site. Radiation is also an acceptable treatment, particularly in poor surgical
candidates. Conventional invasive SCC sometimes arises from VC. When
this occurs, the prognosis and behavior are the same as for conventional
SCC. Thus, the lesion must be thoroughly sampled histologically before
a diagnosis of pure VC is made.
d. Spindle cell carcinoma (SpCC} is the head and neck mucosal form of sarco-
matoid carcinoma, a variant of sec consisting of spindled or pleomorphic
tumor cells that simulate a true sarcoma. It is clear from ultrastructural,
immunohistochemical, and molecular studies that the sarcomatoid cells
represent a clone of poorly differentiated, or divergently differentiated,
carcinoma cells. SpCC has demographics similar to those of conventional
sec, occurring in the fifth to sixth decade, showing a strong association
with smoking and alcohol use, and having a very high male to female ratio.
It occurs most commonly in the larynx (particularly the glottis) followed
by the oral cavity, hypopharynx, and nasal cavity. Up to 20% of patients
have a history of previous radiation to the originating site, which is higher
than that for conventional sec.
Grossly, the vast majority of laryngeal and hypopharyngeal SpCC, and
approximately 50% of oral SpCC, have a polypoid growth pattern result-
ing in an exophytic mass with a smooth and extensively ulcerated surface.
Up to 75% of SpCC are biphasic tumors with areas of conventional SCC
admixed with areas of spindled and/or pleomorphic tumor cells (e-Fig.
1.28). The spindled component usually predominates. The conventional
squamous component may take the form of squamous dysplasia, carci-
noma in situ, or invasive carcinoma. Because the tumors are usually exo-
phytic with extensive surface ulceration, the noninvasive component may
be only a focal finding or may be effaced altogether. SpCC has a wide vari-
ety of architectural patterns including fascicular (e-Fig. 1.29), storiform,
lace-like, or myxoid areas of growth. On occasion, the tumor cells may be
widely spaced in an edematous stroma mimicking granulation tissue with
cytologic atypia, which is a major diagnostic pitfall. Approximately 5%
of tumors will have definable heterologous sarcomatous differentiation,
either osteo- or chondrosarcomatous. Immunohistochemistry in the spin-
dle cell component is positive for cytokeratins and/or epithelial membrane
antigen in approximately two-thirds of cases, and positive for p63 in a sim-
ilar percentage. Vimentin is always positive, and a significant minority of
tumors will be positive for smooth muscle actin.
Although any malignant spindle cell lesion of the mucosa of the upper
aerodigestive tract should be considered an SpCC until proven otherwise,
the differential diagnosis includes a true sarcoma, spindle cell melanoma,
nodular fasciitis, and ulcers and granulation tissue with reactive atypia
(particularly after radiation). When a conventional squamous cell carci-
noma component is present intermingled with the spindle cells, the diag-
nosis of SpCC is confirmed without the need for additional studies.
Because SpCC is inherently a carcinoma, current treatment recommenda-
tions are essentially identical to those for conventional sec, and taking all
patients together, the prognosis does not appear different from conventional
SCC. However, the prognosis is worse for oral cavity SpCC, and patients
with endophytic tumors do worse than those with exophytic tumors.
e. Papillary SCC is a rare variant of SCC that is defined as more than 50%
of the tumor having an exophytic, papillary growth pattern (e-Figs. 1.30
and 1.31). It is more common in the larynx than in the oral cavity and
oropharynx, and has a good prognosis.
f. Adenosquamous carcinoma is another rare variant consisting of a mixture
of sec and adenocarcinoma with true gland formation, often with mucin
production (e-Fig. 1.32). The oral cavity and oropharynx are less common
sites than the larynx. These tumors are typically more aggressive than
conventional sec.
g. Basaloid squamous cell carcinoma (BSCC) is a variant that is rare in the
oral cavity but is slightly more common in the oropharynx, larynx, and
hypopharynx. It is characterized by molded nests forming a "jigsaw" pat-
tern. The tumor cells are basaloid with hyperchromatic, round to oval
nuclei, and scant cytoplasm (e-Fig. 1.33 ). Extracellular mucoid or hyaline
material may be present. Abrupt squamous differentiation or overlying
squamous dysplasia is seen (e-Fig. 1.34). While generally this tumor is
an aggressive variant with high rates of distant metastasis, BSCC in the
oropharynx is frequently HPV positive and when so, has a better progno-
sis than BSCC at other sites.
B. Melanocytic. Melanoma is not uncommon in the oral cavity. It occurs in adults
with an average age of approximately 60 years, with a very even incidence
from age 20 to 80 years. Unlike cutaneous melanoma, in which sun damage
underlies the development of most cases, no major etiology has been identified
for oral lesions. There is a slight male preponderance and, also unlike cutaneous
melanoma, oral lesions occur relatively equally among numerous races. Oral
melanomas present as incidental pigmented lesions identified by a dentist or
physician, as masses arising in a preexisting pigmented lesion, or most frequently
as a new mass growing over a few months. The most common oral site is the
hard palate ("'40%), followed by the maxillary gingiva ("'25%) (e-Fig. 1.35),
the buccal mucosa, the mandibular gingiva, and the lip. Melanomas of the
oropharynx are rare.
Grossly, most tumors are heavily pigmented and heterogeneous, with a
brown, gray, or black color. There are sometimes satellite lesions without inter-
vening pigmentation. Microscopically, most melanomas are deeply invasive,
but two-thirds retain a surface in situ component as well. Architecturally, the
tumors consist of single cells and ill-defined sheets and nests of either epithelioid
cells, spindle cells, or both. The large epithelioid cells have abundant cytoplasm
(which can be eosinophilic to gray) and round to oval nuclei with a characteristic
single, large, cherry red nucleolus (e-Fig. 1.36). Melanin pigment is commonly
present. Spindle cells are less common and have cigar-shaped nuclei and mod-
erate dear to eosinophilic cytoplasm. Plasmacytoid and rhabdoid cells can also
be seen. Intranuclear cytoplasmic inclusions that are typical of melanomas at
other sites are uncommon in oral melanomas. By immunohistochemistry, oral
melanomas express the same proteins as other melanomas, namely Melan-A,
MART-1, HMB-45, S-100, and tyrosinase.
Treatment consists of radical resection with radiation, with or without
chemotherapy. Neck dissection is performed only for clinically detected dis-
ease. The prognosis is poor. Numerous studies have demonstrated a collective
5-year survival between 15% and 25%. Breslow thickness, unlike cutaneous
lesions, is of very limited prognostic utility. Approximately 40% of patients will
develop cervical lymph node metastases, and distant metastases are common.
Major sites include the lung, the liver, and the brain.
C. Neuroendocrine carcinomas are uncommon tumors in the head and neck region
in general, and are very uncommon in the oral cavity and oropharynx. They
are essentially all high grade with a small cell morphology, composed of cells
with scant cytoplasm, crush artifact, granular chromatin without nucleoli, and
extensive necrosis with brisk mitotic activity (i.e., morphologically identical
to small cell carcinomas of the lung and other organs) (e-Fig. 1.37 and e-Fig.
1.38). Some are mixed with a component of SCC. In the oropharynx, recent
studies have shown that roughly one-half of tumors have been shown to have
transcriptionally-active HPV. Carcinoid tumors (low-grade neuroendocrine car-
cinomas) almost never occur in the oral cavity and oropharynx.
The prognosis for high-grade neuroendocrine carcinoma is very poor, with
rapid progression of disease including the development of cervical lymph node
metastases and distant metastatic disease. This seems to still hold true for
patients with HPV-related oropharyngeal neuroendocrine carcinomas as well.
D. Vascular
1. Pyogenic granulomas (or lobular capillary hemangiomas) are benign lesions
of the oral cavity that are usually solitary, and are most common on the
lips, tongue, and gingival and buccal mucosa. They occur in patients of all
ages but have a particular predilection for the gingiva of pregnant women
and, thus, are often referred to as a "pregnancy tumor" in this setting. They
usually present as small (a few centimeters or less) exophytic masses that
bleed easily. They may grow rapidly and cause false clinical concern for a
malignant neoplasm.
Grossly, they are polypoid or pedunculated, pink to red, and have a
smooth surface. Microscopically, they consist of lobules of small capillaries
with plump endothelial cells that have round to oval nuclei and occasional
mitoses (e-Fig. 1.39), with larger central "feeder" vessels. Extensive surface
ulceration with associated fibrin is usually present. The cells are positive for
endothelial immunohistochemical markers such as CD34, CD31, and factor
Vlll-related antigen.
Pyogenic granulomas are benign neoplasms that are cured by simple exci-
sion. A small percentage may recur. Pregnancy-related lesions often regress
postpartum and thus may be hormonally driven.
2. Hemangioma and lymphangioma are benign tumors composed of abundant
blood or lymphatic vessels, respectively. Hemangiomas of the oral cav-
ity usually occur in adults. They occur most commonly on the lip, buccal
mucosa, and lateral tongue borders and present as painless, nodular or well-
circumscribed, red or blue masses (e-Fig. 1.40) measuring <2 em in maxi-
mal dimension. Microscopically, they consist of blood vessels ranging from
small capillaries to large cavernous spaces. The endothelial lining cells can
be plump and may have mitotic activity, a feature more common in children.
There are a number of named histologic variants, most of which have no
clinical significance.
Lymphangiomas (or cystic hygromas) are composed of dilated lymphatic
channels. About 75% occur in the head and neck and, when presenting in the
oral cavity, they are almost always found in children younger than 3 years.
Histologically, they typically consist of very dilated lymphatic channels lined
by bland, inconspicuous endothelial cells, with intraluminal eosinophilic
material, lymphocytes, and occasional red blood cells. They often are inter-
stitial aggregates of lymphocytes.
Both hemangioma and lymphangioma are benign lesions cured by conser-
vative excision. Hemangiomas can be treated by sclerotherapy as well. Large
lymphangiomas are often de bulked, often via serial resections to avoid major
morbidity.
3. Kaposi sarcoma (KS) is a locally aggressive tumor uniformly associated with
human herpesvirus 8 (HHV-8) that predominantly involves the skin but can
also involve mucosal sites. When KS involves the oral cavity, usually as a
complication of AIDS, it is most commonly found in the palate, followed by
the gingiva and dorsal tongue. Clinically, KS appears as purple, red-blue, or
brown macules or plaques. Later in the disease course, the lesions become
nodular and may ulcerate.
The three histologic stages of KS (patch, plaque, and nodular; all dis-
cussed in more detail in Chap. 39) can all be associated with oral KS (e-Fig.
1.41). There are frequently associated collections of extravasated red blood
cells and hemosiderin-laden macrophages. A characteristic feature that is
sometimes seen is the pale, eosinophilic hyaline globule, which probably
represents degenerating red blood cells. Immunohistochemistry is positive
for the common endothelial markers CD34 and CD31.
The behavior of oral KS is variable. It is generally indolent in nonim-
munocompromised patients but is more aggressive in patients with AIDS.
However, the mortality related to KS is highly dependent on other comor-
bidities such as opportunistic infections and systemic symptoms.
E. NeuraVneuroectodermal. Granular cell tumors are benign, slow growing tumors
of neural origin that occur at many anatomic sites. Approximately 50% occur
in the head and neck region, and half of these occur in the tongue. They also
occur in the buccal mucosa, floor of mouth, and palate, are twice as common in
women as men, and approximately 10% to 20% are multiple. Grossly, they are
smooth, sessile, and firm with a pink or tan-white color. Microscopically, they
consist of infiltrative nonencapsulated sheets and cords of bland cells with abun-
dant eosinophilic granular cytoplasm and indistinct cell borders (e-Fig. 1.42).
There is usually no significant stromal reaction. The nuclei are small, oval, and
hyperchromatic with minimal atypia and no mitotic activity. A common feature
is pseudoepitheliomatous hyperplasia of the overlying squamous epithelium,
which can closely mimic SCC. Conservative excision is the treatment of choice,
with a risk of recurrence of <10%. Malignant granular cell tumors (as covered
in the Soft Tissue chapter) are very rare but do occur.
F. Mesenchymal
1. Peripheral ossifying fibroma is a reactive proliferation of fibrous tissue on the
gingiva which shows focal bone formation. It occurs over a broad age range,
but young adults are most commonly affected. The lesions range from a few
millimeters up to 2 em in maximal dimension. They are essentially exclusive
to the gingiva, particularly along the incisors, and present as sessile pink
nodules, usually with surface ulceration. Microscopically, they consist of
randomly distributed plump (but not atypical) fibroblasts with foci of miner-
alization ranging from dystrophic calcification, to cementum-like material,
to well-formed bone (e-Fig. 1.43). Rare giant cells can be seen. The lesions
should be excised down to the periosteum but will recur in 15% to 20% of
cases.
2. Peripheral giant cell granuloma is another reactive proliferation of the gingiva,
particularly along the incisors, caused by chronic irritation. It presents over
a wide age range, particularly in middle-aged to older adults, as a solitary
broad-based nodule that is reddish or blue and <2 em in diameter. Micro-
scopically, it consists of a mixture of multinucleated osteoclast-like giant cells
and plump spindled to oval mononuclear cells (e-Fig. 1.44). Hemosiderin,
chronic inflammation, and foci of metaplastic bone are frequent. The differ-
ential diagnosis includes brown tumor of hyperparathyroidism, cherubism,
and central (intraosseous) giant cell granuloma. The lesion is treated by local
excision down to the bone. Recurrence occurs in approximately 10% of
cases.
3. Congenital granular cell epulis is a rare benign mesenchymal tumor that clas-
sically arises from the anterior alveolar ridge of a newborn. Girls are more
frequently affected than boys by a ratio of 9:1. The tumor presents as a
11 I SECTION I. HEAD AHD NECK
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smooth, nonulcerated mass about 1 em in size arising from the gingiva over
the lateral incisor/canine area of the maxilla, or less commonly, the mandible.
Grossly, it is polypoid and has a homogeneous firm pink or tan cut surface.
Microscopically, it consists of sheets of large cells that have abundant, gran-
ular, eosinophilic cytoplasm and round to oval, bland nuclei. The surface
squamous epithelium is intact and shows no hyperplasia. By immunohisto-
chemistry, the cells are positive only for vimentin, and specifically are negative
for S-100.
Congenital granular cell epulis is not the newborn equivalent of a granular
cell tumor and shows no neural differentiation. The tumor stops growing at
birth and regresses over time, but most cases still require surgical resection.
There is no recurrence, even after incomplete removal.
V. PATHOLOGIC REPORTING OF ORAL CAVITY AND OROPHARYNGEAL MALIGNANCIES
A. Staging. The American Joint Committee on Cancer (AJCC) staging guidelines
for oral cavity and oropharyngeal carcinomas are listed in Tables 1.3 and 1.4.
Staging is extremely important for clinical management and establishing prog-
nosis. These staging guidelines are applicable to all forms of carcinoma. Any
nonepithelial tumor type is excluded. A specific and very important point in stag-
ing involves bone involvement by tumor. To qualify as a T4lesion, the tumor
must erode through the bone cortex. Superficial bone erosion is not sufficient
for classification as T4.
It is important to note that mucosal melanomas of the head and neck, includ-
ing the oral cavity, have their own staging system.
B. Additional pertinent pathologic features. As with carcinomas at all upper aerodi-
gestive tract sites, margin status, tumor differentiation, and the presence or
absence of perineural or lymphovascular space invasion should be reported.
Perineural invasion is particularly common in oral cavity carcinomas and is
correlated with a poorer prognosis. The pattern of infiltration as well as the
presence or absence of a host inflammatory response should also be reported,
because both features have been correlated in many studies with a higher rate of
local recurrence, a poorer prognosis, or both. Some recent studies have devel-
oped formal grading systems for these features that have promise for clinical
implementation, particularly for early stage oral cavity sec, but these are not
ready for active implementation. Depth of invasion, particularly for T1 and T2
tumors, although not reflected in the staging system specifically, is important
for clinical management and prognosis, and so should be reported.
SUGGESTED READINGS
Adelstein DJ, Ridge JA, Gillison ML, et al. Head and neck squamous cell cancer and the human
papillomavirus: summary of a National Cancer Institute State of the Science Meeting, Novem-
ber 9-10,2008. Washington, DC, Head Neck. 2009;31:1393-1422.
Ang KK, Harris J, Wheeler R, et al. Human papillomavirus and survival of patients with oropha-
ryngeal cancer. N Engl] Med. 2010;363:24-35.
Bouquot JE, Muller, S, Nikai H. Lesions of the oral cavity. In: Gnepp DR, ed. Diagnostic Surgical
Pathology of the Head and Neck. 2nd ed. Philadelphia, PA: W.B. Saunders Publishers; 2009.
Chemock RD, El-Mofty SK, Thorstad WL, et al. HPV-related nonkeratinizing squamous cell car-
cinoma of the oropharynx: utility of microscopic features in predicting patient outcome. Head
Neck Pathol. 2009;3:186-194.
Chemock RD, Lewis JS, Jr, Zhang Q, et al. Human papillomavirus-positive basaloid squamous
cell carcinomas of the upper aerodigestive tract: A distinct clinicopathologic and molecular
subtype of basaloid squamous cell carcinoma. Hum Pathol. 2010;41:1016-1023.
Slootweg P, Eveson ]W. Tumours of the oral cavity and oropharynx. In: Barnes L, Eveson JW,
Reichart P, Sidransky, eds. Pathology and Genetics Head and Neck Tumours. Lyon, France:
IARC Press; 2005.
Thompson LDR, ed. Head and Neck Pathology, 1st ed. New York, NY: Churchill Livingstone;
2006.
Larynx
James S. Lewis Jr.
I. NORMAL ANATOMY
A. Macroscopic/gross. The larynx is a unique organ designed to produce phonation
by modulation of the respiratory airstream. It is composed of several cartilagi-
nous structures: the thyroid, cricoid, and arytenoid cartilages and the epiglottis.
The hyoid bone sits above and is connected to the larynx by the thyrohyoid
membrane (Fig. 2.1). The thyroid (and lesser so, cricoid cartilage) ossifies in
adults. In functional terms, the larynx is divided into three subsites. The glot-
tis includes the true vocal folds or cords, below, and the false folds or cords,
above. The space between them is called the ventricle, and its deeper recess,
the saccule. The true cords, with a hypocellular stroma called Reinke's space,
are designed to vibrate for phonation, and the ventricle amplifies this further.
The cords are manipulated by muscles that attach to and move the arytenoid
cartilages, which sit at the posterior aspect of the vocal folds. The larynx can be
divided into three compartments for tumor management and staging purposes:
the supraglottis, glottis, and subglottis. The supraglottis includes the epiglottis,
aryepiglottic folds, false cords, and ventricle. Glottis refers to the vocal cords
from the edge of the ventricle to the free edge of the vocal cord. Subglottis
refers to the area from the free edge of the vocal fold to the inferior border of
the cricoid cartilage but has a slightly different definition for American Joint
Committee on Cancer (AJCC) staging (see later).
B. Microscopic. The larynx is covered by a mixture of squamous and pseudos-
tratified ciliated columnar (respiratory-type) epithelium. In smokers, however,
often the entire endolarynx is covered by squamous epithelium. The true cords
themselves are always covered by squamous epithelium. They contain a lamina
propria area called Reinke's space, which lies between the epithelium and the
vocal ligament and which consists of loose connective tissue with few capillaries,
no lymphatics, and sparse seromucinous glands (e-Fig. 2.1).* The false cords
and ventricle are typically lined by respiratory-type epithelium.
A. Endoscopic biopsies. The majority of specimens from this region consist of endo-
scopic forcep biopsies. The tissue samples should be placed immediately into
10% buffered formalin or other appropriate fixative. These should undergo
gross examination and description documenting the exact number of pieces
present and their size, and then be entirely submitted with three levels cut from
each paraffin block for hematoxylin and eosin (H&E) examination.
B. Resections
1. Partial. These are widely variable specimens depending on the location of the
tumor. Standard procedures include vertical hemilaryngectomy and supra-
glottic or supracricoid laryngectomy. As a generalization, these specimens
need to be oriented, the soft tissue margins inked, and margins demonstrated
by shave or radial section followed by sectioning of the tumor relative to car-
tilage/bone and soft tissue margins. The use of either shave or radial sections
depends on the nature of the specimen. If the tumor is relatively distant from
a margin, 1- to 2-mm shave sections are preferred. If the tumor approximates
a margin to < 1 to 2 mm, radial sections are taken.
*All e-figures are available online via the Solution Site Image Bank.
19
Pre-epiglottic
space (fa t)
Thyroid
C~~~"c--iJ,_-Cricothyroid cartilage
ligament
Cricothyroid
muscles
Cricoid
cartilage
(anterior)
B
Figure 2.1 Larynx anatomy: (A) anterior view; (B) sagittal cross section.
Partial resections with a C02 laser under an operating microscope are

becoming more common because of their low morbidity. Because an inherent
part of this procedure is to cut into the tumor or to remove tumor in more
than one piece, surgeons ink the individual pieces themselves, as they alone
know what constitutes the true margin. In the pathology lab, the pieces
are measured, described, and submitted entirely in sections perpendicular to
the ink.
2. Total. For years, total laryngectomy has been the standard operation for
malignancy. However, partial resections with preservation of function are
increasingly common. Total laryngectomy is used most often now as salvage
therapy for recurrences after partial surgery or after definitive radiation and
chemotherapy. Occasionally, patients present with progressive disease that
makes total laryngectomy necessary as the initial surgery (e-Fig. 2.2).
The usual approach to grossing a total laryngectomy is to initially ink
the peripheral nonmucosal soft tissue margins and then open the larynx by
a posterior vertical midline cut with scissors, propping it wide open with a
small stick or portion of a wooden swab. After overnight formalin fixation,
the specimen is ready for prosection. After orientation and measurement
in three dimensions, the tumor is measured and described, specifically not-
ing what structures are involved. The specimen typically includes the entire
larynx and cartilages, small portions of hypopharyngeal mucosa bilaterally
adjacent to the aryepiglottic folds, and the hyoid bone anterosuperiorly. Stan-
dard sections (Fig. 2.2) are taken as follows. (i) Margins: shaved inferior tra-
cheal ring; shaved right and left hypopharyngeal mucosa; shaved postcricoid
soft tissue; radial sections demonstrating anterior, anterolateral, and base of
tongue inked soft tissue margins. (ii) Soft tissue/mucosa: bilateral vertical
glottis (including both true and false cords); vertical anterior commissure;
vertical midline epiglottis showing pre-epiglottic soft tissue space; bilateral
aryepiglottic folds; four sections of tumor if not included in the previous
sections. (iii) Cartilage/bone: vertical sections showing tumor and thyroid
cartilage (at closest or at sites of gross invasion); sections of right and left
wings of the hyoid bone (where closest to or involved by tumor). Typically,
all soft tissue and margin sections are taken first, and the entire specimen is
Chapter 2 • Larynx I 21
Midline epiglottis shoiNfng pre-epiglottic 80ft tissue
Right superior hypopharyngeal

muc::osal shaved margin
Right Inferior hypopharyngeal

muc:osal shaved margin
Distal tracheal shawct margin

Figure 2.2 Larynx grossing. Standard sections from a total laryngectomy
are shown. Additional sections include sections of tumor (4-5 total), right
and left postcricoid soft tissue shaved margins, and anterior soft tissue
margins (either shaved or radial). Post decalcification, additional sections
should include cartilage deep to the tumor showing involvement or nearest
approach, any surrounding lymph nodes in neck soft tissue, of the hyoid
bone (where closest to tumor or grossly involved by it). If necessary, sec-
tions of tracheostomy skin, thyroid lobe(s), and neck skin should also be
submitted.
bisected in the midline vertically and decalcified in toto overnight followed

by cartilage and/or bone section acquisition.
C. Frozen sections. These are a critical element of surgical therapy for tumors of
the head and neck region. Although practices vary, most institutions have mar-
gins taken as small pieces by the surgeon from the periphery of the surgical
defect after the tumor has been removed. Because laryngeal resections are quite
variable, the sites where frozen sections are taken are not standard; with total
laryngectomies, sometimes no frozen sections are clinically necessary. The sur-
geon may sample the tumor or suspicious sites to confirm and/or map the tumor,
and then take margins from the area of closest approach after resection of it.
The tissue pieces are submitted individually to pathology in saline and frozen
in their entirety, with two (and at our institution, three) H&E slides generated
at representative levels. It is critical to obtain sections that represent the entire
tissue submitted so that small foci of tumor are not missed by "sampling error"
(i.e., where tiny foci of tumor are not present on frozen section slides but are
seen on permanent sections, due to not "sampling" the tissue well at the time of
the frozen). This is best done by making sure that the second and third sections
are taken from deep into the tissue. The pieces should be evaluated grossly for
mucosa-typically shiny and pink-tan on one surface of the tissue; if present,
the specimen should be oriented to demonstrate this surface on one edge of the
section with the submucosa below. Additional sections should be cut if needed
to assure that two or three quality sections are obtained.
The tissue that remains after frozen section is submitted for evaluation by
permanent sections. This process can help resolve a number of issues from frozen
section including freezing and cautery artifact, amount of tumor represented,
and orientation or embedding issues. The margins of the main resection speci-
men are also evaluated throughout its entirety because the separate frozen sec-
tion specimens are small and almost never cover the entire margin of a resection.
The final margin status is then a conglomerate of all three sources: frozen section
slides, permanent slides of the frozen tissue, and the margins of the specimen
itself.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES
A. Inflammation and infection. Inflammation of the larynx (laryngitis) is quite com-
mon clinically, and can be divided into acute and chronic forms, which are
variable by age. Laryngitis rarely necessitates tissue biopsy for pathologic eval-
uation. Infections can be caused by a myriad of agents including viruses, bacteria,
fungi, and parasites. The pathologist must be alert to the possibility of infection,
and the immune status of the patient is helpful information, as many of these
patients will be immunocompromised.
Inflammation is essentially never present in the normal larynx, so the pres-
ence of inflammatory cells is a diagnostic clue. Depending on the organism, the
inflammation can take a number of different forms, almost all of which are typ-
ical for the type of organism when it presents in other locations. Examples of
some of the major infections include cytomegalovirus, herpes simplex virus,
tuberculosis, rhinoscleroma (Klebsiella rhinoscleromatis), candidiasis, histo-
plasmosis, blastomycosis, cryptococcosis, coccidiomycosis, or rhinosporidio-
sis (Rhinosporidium seeberi). The resulting inflammation may cause mucosal
ulceration, acute and chronic inflammation, necrosis, or granulomas. Special
stains (such as Gomori•s methenamine silver (GMS), acid-fast bacillus (AFB),
or periodic acid-Schiff (PAS)) should be utilized to look for, and characterize,
organisms.
B. Non-neoplastic lesions
1. Traumatic
a. Vocal cord nodules and polyps. These are non-neoplastic degenerative stro-
mallesions of Reinke•s space that are usually related to trauma due to
misuse or vocal excess. As such, they have been referred to as singer•s or
screamer•s nodules. They are more common in women and are commonly
bilateral, characteristically occurring at the junction of the anterior and
middle one-third of the vocal cord, as this is the point of maximal vibra-
tion during phonation. Macroscopically, they appear as gray or white
broad-based nodules or polyps (e-Fig. 2.3). Microscopically, they consist
of squamous mucosa with or without hyperkeratosis, are only rarely ulcer-
ated, and overlie a sparsely cellular myxoid, edematous, fibrous, fibrinous,
or vascular stroma. The myxoid appearance is most common and it may
demonstrate small cystic spaces (e-Figs. 2.4 and 2.5). So-called vocal cord
polyps are histologically identical but clinically present unilaterally, and
in men and smokers with more regularity.
b. Contact ulcer. Also referred to clinically as contact granuloma or just gran-
uloma, these occur on the vocal process of the arytenoids classically as a
result of forceful vocalization in individuals who must affect a low, deep,
forceful voice. However, they also may result after endotracheal intubation
or as a result of gastroesophageal reflux disease. They are more common
in men, can be unilateral or bilateral, and present as polypoid lesions.
Microscopically, they are essentially granulation tissue polyps with an

ulcerated mucosa and a stroma containing abundant small vessels in a
haphazard configuration with plump endothelial cells (e-Fig. 2.6). The
stroma may be rich in lymphocytes, plasma cells, neutrophils, or histio-
cytes (sometimes including giant cells).
2. Cysts. Laryngeal cysts may be divided into three categories: (i) ductal cysts,
(ii) laryngoceles, and (iii) saccular cysts. All are cured by simple excision.
a. Ductal cysts. These are the most common and result from obstruction of
a minor salivary gland duct. Ductal cysts are typically small "bumps" on
endoscopy and have a predilection for the cords, ventricle, aryepiglottic
folds, and epiglottis. The cyst lining may be squamous or oncocytic.
b. Laryngocele. A laryngocele is an asymptomatic dilatation of the saccule
(the deep aspect of the ventricle). It may remain internal and manifest as
a supraglottic mucosal bulge, or may herniate above the thyroid cartilage
to project externally into the neck soft tissue and present as a neck mass.
Microscopically, it is lined by respiratory-type mucosa (e-Fig. 2.7).
c. Saccular cyst. This represents a mucin-filled dilatation of the saccule, either
developmental or acquired. It is also typically lined by respiratory-type
mucosa, but the lining can be squamous or oncocytic on occasion. It is
easily confused with a branchial cleft cyst.
3. Metabolic
a. Amyloidosis. The larynx is a well-established, although infrequent, site of
localized amyloidosis. It most commonly involves the false cord, followed
by the true cord and ventricle. A subset of patients has multifocal dis-
ease, with approximately one-third having tracheal disease as well. The
majority of patients have only localized disease, but systemic disease also
occurs in a subset of patients. All patients should have a workup to rule
out systemic amyloidosis with or without an associated plasma cell dyscra-
sia. The amyloid in localized laryngotracheal amyloidosis is of the AL (or
immunoglobulin light chain type) type. It usually macroscopically presents
as a polypoid nodule covered by intact mucosa. Microscopically, it con-
sists of sheets and nodular masses of amorphous, hypocellular eosinophilic
material in the stroma (e-Fig. 2.8), in blood vessel walls, and in the base-
ment membranes of mucoserous glands. The diagnosis is confirmed by
Congo red special staining with "apple-green" birefringence on polarized
microscopy. Thioflavin T staining with examination under fluorescence
microscopy is also sometimes used.
C. Neoplastic lesions. The World Health Organization (WHO) classification of
tumors of the larynx, hypopharynx, and trachea is listed in Table 2.1.
1. Benign
a. Squamous papillomas are squamous proliferations caused by human papil-
loma virus (HPV) and are the most common benign tumors of the larynx.
They are most common in the larynx, but also occur in the trachea and
bronchi. They also occur occasionally in the oral cavity and pharynx.
There are two separate clinical settings: juvenile or juvenile-onset laryn-
geal papillomatosis UOLP) and adult (or adult-onset) laryngeal papillo-
matosis (AOLP). Juvenile papillomas most often begin before the age of
5 years and are much more likely than adult papillomas to be multifo-
cal and to have an associated clinical impact. In a significant minority of
patients, carpeting of the larynx occurs, requiring repeated laser excisions
and occasionally tracheostomy or even laryngectomy for control and air-
way management. Papillomas tend to recur rapidly, but the disease severity
usually regresses in early adulthood. In adult papillomas, the peak age is
between 20 and 40 years, disease is usually unifocal or limited, and even
when multifocal, it is less aggressive. Only occasionally does it present as
multifocal disease that recurs after excision.
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Squamous papillomas are strongly associated with the low risk HPV
types 6 and 11, which are thought to be transmitted from the mother
to the upper aerodigestive tract (UADT) of the neonate during vaginal
delivery, thus explaining the occurrence in childhood. There is minimal
risk of transformation to invasive carcinoma.
Regardless of the clinical context, squamous papillomas have a typi-
cal morphology. Grossly, they appear as exophytic, granular, and friable
pink, red, or tan lesions. Microscopically, they consist of arborizing, pap-
illary fronds of thickened but maturing squamous epithelium with slight
hyperplasia of the basal layer (e-Fig. 2.9). Cells in the midlayer often have
cytoplasmic clearing, but frank koilocytosis is not regularly observed. The
nuclei are slightly enlarged and irregular but do not appear overtly dys-
plastic, and mitotic activity, while present, is usually limited. Character-
istically, there is minimal surface keratinization (e-Fig. 2.10). Frank dys-
plasia can be seen in some lesions and should be reported and graded as
in nonpapillary squamous mucosa (see Section m.C.2). However, there
is no consistent correlation of overt dysplasia with the development of
subsequent invasive carcinoma.
b. Granular cell tumors are benign, slowly growing tumors of neural origin
that occur in a multitude of anatomic locations. They are particularly
common in the head and neck region with a minority occurring in the lar-
ynx. They are slightly more common in African Americans and typically
present with hoarseness. Grossly, they are smooth, white polypoid tumors
involving the posterior true cords, anterior commissure, false cords, or
subglottis. Microscopically, they consist of infiltrative, nonencapsulated
sheets and cords of bland cells with abundant eosinophilic granular cyto-
plasm and indistinct cell borders. There is usually no significant stromal
reaction. The nuclei are small, oval, and eccentric with minimal atypia and
no mitotic activity. A common feature is pseudoepitheliomatous hyperpla-
sia of the overlying squamous epithelium, which can be quite alarming,
can mimic squamous cell carcinoma, and should prompt a search of the
submucosa for granular cells (e-Fig. 2.11). Conservative endoscopic exci-
sion is the treatment of choice with a less than 10% risk of recurrence.
c. Paragangliomas are tumors recapitulating the paraganglia, specialized
organs of the autonomic nervous system derived from the neural crest.
They occur in numerous locations throughout the body, and their behavior
is largely dependent on site. Laryngeal paragangliomas are benign and are
divided into two groups: superior and inferior. Superior paragangliomas
are much more common and occur in the supraglottic larynx, from the
false cord up into the aryepiglottic fold. They are polypoid submucosal
lesions. Inferior paragangliomas occur along the cricoid cartilage in the
subglottic region and often present as dumbbell-shaped lesions with both
intra- and extralaryngeal components.
Microscopically, these tumors are identical to those arising elsewhere.
They consist of sheets and nests of polygonal to spindled cells with abun-
dant eosinophilic to slightly basophilic cytoplasm, and round to oval
nuclei with a slightly granular chromatin. They are very vascular with
stellate large vessels throughout (e-Fig. 2.12). Nuclear pleomorphism may
be striking, but there is minimal mitotic activity. They are arranged in
nests (classically termed zellballen) (e-Fig. 2.13). By immunohistochem-
istry, they are strongly positive for synaptophysin and chromogranin A,
virtually always negative for epithelial markers such as epithelial mem-
brane antigen (EMA) and cytokeratin, and show the typical sustentacular
(or supporting) cell staining for S-100 around the periphery of the nests
whereas the tumor cells themselves are negative.
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tumors with raised edges. Microscopically, the term "squamous cell car-
cinoma, keratinizing type" is used to describe the common version of the
cancer and to distinguish it clearly from the numerous variant squamous
cell carcinoma types that exist. Keratinizing-type squamous cell carci-
noma consists of nests and sheets of cells with abundant eosinophilic
cytoplasm and round to oval nuclei, often with prominent nucleoli. Well-
differentiated tumors retain abundant pink or clear cytoplasm and often
show keratin "pearl" formation (e-Fig. 2.9). Moderately differentiated
tumors have more pleomorphism and a higher nucleus to cytoplasm ratio
in many of the cells while still retaining moderate eosinophilic cytoplasm
(e-Fig. 2.10). Poorly differentiated tumors often have single cells or small
nests of cells with more mitotic activity and less cytoplasm (e-Fig. 2.11).
Grading should be performed, although it has not been shown to consis-
tently predict the clinical behavior of individual tumors. The majority of
UADT squamous cell carcinomas are moderately differentiated. Although
frank keratin formation is commonly seen in well- and moderately differ-
entiated carcinomas, its presence or absence does not have any clinical
significance for laryngeal carcinoma.
i. Verrucous carcinoma (VC) is a specific, well-differentiated, and non-
metastasizing variant of squamous cell carcinoma. First described
by Lauren V. Ackerman in the 1950s, it has thus also been called
"Ackerman's tumor." It occurs in the larynx and oral cavity and grossly
appears as a well-circumscribed, warty, and exophytic, broad-based
white, granular, and friable mass. Microscopically, VC consists of very
thick, dub-shaped papillae with broad pushing bases. These blunt and
downward-pushing projections have sometimes been likened to "ele-
phant's feet." There is usually prominent surface hyperkeratosis, and
the sheets of tumor have bland cells with abundant eosinophilic to clear
cytoplasm, sometimes described as "glassy" in appearance. There is
no cytologic atypia, mitotic activity is very low and basal, and there
is a smooth interface with the stroma. This latter feature is because
the individual tumor cells have not breached the basement mem-
brane. The stroma directly beneath the tumor typically demonstrates
prominent plasma cell rich chronic inflammation (e-Figs. 2.18, 2.19,
and 2.20).
The prognosis for pure VC is excellent. Although it can be large
and locally destructive, complete surgical resection is often curative.
Radiation is also an acceptable treatment, particularly in poor surgical
candidates. However, routine invasive squamous cell carcinoma some-
times arises from VC. When this occurs, the prognosis and behavior
are the same as for typical squamous cell carcinoma.
ii. Spindle cell carcinoma (SpCC} is the term for a poorly dif-
ferentiated carcinoma that adopts a sarcomatoid, spindled, or
mesenchymal-appearing morphology but is, nevertheless, of epithelial
origin/differentiation. It is frequently biphasic with a spindled com-
ponent and intermingled either in situ or invasive squamous cell car-
cinoma. The tumor has the same demographics as routine squamous
cell carcinoma.
Grossly, SpCC is characteristically polypoid with a smooth, ulcer-
ated surface. The glottis is the most common laryngeal site. Microscop-
ically, these tumors can be quite variable, but typically consist of sheets
of spindle cells mimicking a fibrosarcoma or malignant fibrous histi-
ocytoma (e-Fig. 2.21). Sometimes they consist of pleomorphic, hyper-
chromatic cells widely separated in an edematous stroma (e-Fig. 2.22).
There is usually brisk mitotic activity and necrosis. Foci of recognizable
sarcomatous differentiation such as chondrosarcoma, osteosarcoma,

or rhabdomyosarcoma sometimes occur. Most cases are "biphasic,"
showing some component of in situ or invasive squamous cell
carcinoma (e-Fig. 2.23 ), but it may take extensive sectioning to demon-
strate the routine squamous component.
If the tumor consists of spindle or pleomorphic cells only, immuno-
histochemistry for epithelial markers such as pancytokeratin, EMA,
and p63 is usually very helpful. These markers are positive in one-
fourth to one-third of cases. With markedly extended panels utiliz-
ing antibodies to more than 10 different individual keratins, approxi-
mately three-quarters of tumors will demonstrate staining; however, in
everyday practice, utilization of pancytokeratin (AE1/AE3) as well as
34,8£12 or 5/6 usually suffices demonstrate keratin expression. SpCC
stains for some mesenchymal markers as well, such as vimentin (every
case), smooth muscle actin, and muscle-specific actin. Whatever the
case, a malignant spindle cell neoplasm involving/arising along the
mucosa of the UADT should be considered an SpCC until proven
otherwise, because sarcomas are distinctly uncommon at these sites.
The differential diagnosis includes a granulation tissue polyp, true sar-
coma, or inflammatory myofibroblastic tumor. The positive epithelial
marker immunohistochemistry will distinguish SpCC from these other
entities.
iii. Basaloid squamous cell carcinoma {BSCC} is a variant of squamous cell
carcinoma composed almost entirely of basaloid cells giving it a "blue
cell" appearance. It has the same demographics as routine squamous
cell carcinoma but has a predilection for the supraglottic larynx, the
oropharynx, and the hypopharynx, particularly the pyriform sinuses.
It typically presents at high stage and is more aggressive than typical
squamous cell carcinoma stage for stage, with high rates of distant
metastasis.
Grossly, BSCC presents as a centrally ulcerated mass with thicken-
ing at the edges, and commonly with extensive submucosal induration
and spread at the periphery. Microscopically, there are two compo-
nents. The first is predominant and consists of rounded nests of basa-
loid cells with hyperchromatic round nuclei, inconspicuous nucleoli,
and scant cytoplasm. Comedo-type central necrosis is common. The
second component is typical keratinizing-type squamous cell carci-
noma, either in situ or invasive, which is always focal (e-Fig. 2.24). The
basaloid component of many tumors shows a characteristic hyalinized,
eosinophilic basement membrane-like stroma around the tumor cells,
and/or as small round nodules within tumor nests akin to that seen in
cylindromas of the skin (e-Fig. 2.25).
The differential diagnosis includes neuroendocrine carcinoma or
the solid variant of adenoid cystic carcinoma. The former expresses
neuroendocrine markers in most cases and is negative for high-
molecular-weight cytokeratins such as 34,8E12 and 5/6. Solid adenoid
cystic carcinoma shows some evidence of myoepithelial differentiation
upon staining for p63 manifested as scattered staining at the periph-
ery of tumor nests; in contrast, BSCC stains diffusely throughout the
tumor (as with squamous epithelium and squamous cell carcinoma,
which also show strong staining).
iv. Papillary squamous cell carcinoma (PSCC} is an uncommon variant of
squamous cell carcinoma which has the same demographics as typi-
cal squamous cell carcinoma. The larynx is among the most common
sites of involvement, the supraglottis and glottis in particular; only
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carcinoid tumor (low-grade neuroendocrine carcinoma), atypical carci-

noid tumor (intermediate grade neuroendocrine carcinoma), and small
cell carcinoma (high-grade neuroendocrine carcinoma). More recently,
large cell neuroendocrine carcinoma similar to that seen in the lung has
been defined in the larynx as well. There is a clear spectrum of behavior
among the subtypes ranging from very indolent to highly aggressive.
Most neuroendocrine carcinomas of the larynx occur in the sixth to the
eighth decade, and there is a strong male predominance. They are almost
exclusively supraglottic, and smoking is associated with atypical carcinoid
tumors and small and large cell neuroendocrine carcinomas only.
The histology of these tumors directly parallels those within the lung.
Carcinoid tumor is characterized by nests, trabeculae, and sheets of round,
regular tumor cells with moderate eosinophilic to partially clear cyto-
plasm. Nuclei are round, regular, and show the typical "salt and pep-
per" or stippled chromatin. Mitotic activity is less than 2 per 10 high
power fields. Atypical carcinoid tumor has a similar appearance but is
less organized and shows more cellular pleomorphism, focal necrosis,
and more prominent mitotic activity (3-10 per 10 high power fields).
Small cell carcinoma is high grade and consists of sheets of blue cells
with nuclear molding, crush artifact, and delicate chromatin with indis-
tinct nucleoli (e-Fig. 2.16). Large cell neuroendocrine carcinoma is also
high grade but has larger cells with generous eosinophilic cytoplasm and
frequently a more organized growth pattern with peripheral palisading.
There is brisk mitotic activity and prominent necrosis in both forms
of high-grade neuroendocrine carcinoma. All of these neoplasms stain
with neuroendocrine markers by immunohistochemistry (synaptophysin,
chromogranin-A, CD56/N-CAM), but there tends to be less staining with
the high-grade tumors. In particular, in small cell carcinoma, the staining
can become quite focal and sometimes is only present for one of the several
neuroendocrine markers.
Carcinoid tumor has an excellent prognosis despite frequent local
recurrence. Metastases are uncommon. Atypical carcinoid tumor is an
aggressive tumor with frequent cervical lymph node and distant metas-
tases. Surgery is the treatment of choice for both. The high-grade neu-
roendocrine carcinomas have a dismal prognosis with frequent neck nodal
and distant metastases. Patients are typically treated with radiation and
chemotherapy rather than surgery.
c. Mesenchymal tumors. A great variety of mesenchymal tumors occasion-
ally occur in the larynx, the most common of which are the cartilaginous
tumors chondroma and chondrosarcoma. Chondrosarcomas are the most
common sarcoma of the larynx, occur in older adults, and are decidedly
more common in men. They originate from the cricoid or thyroid lam-
ina (3:1, cricoid to thyroid) and tend to present differently by site of
origin with slowly progressive dyspnea for cricoid tumors, or an ante-
rior neck mass for thyroid tumors. Symptoms are usually present for a
long period of time (often several years). Grossly, the neoplasms consist
of well-circumscribed, shiny white or gray lobulated masses with gritty
areas of calcification and a glistening cut surface (e-Fig. 2.17). Micro-
scopically, they consist of lobules of cartilage which, relative to normal,
show increased cellularity and nuclear atypia with variability in size, more
than one nucleus in individual lacunae, and binucleated cells (e-Fig. 2.28).
The periphery of the tumor is typically pushing and not infiltrative. The
grading system is identical to that for chondrosarcomas elsewhere in the
body. With increasing grades, the cellular atypia increases with cells having
enlarged nuclei and nucleoli, but mitotic activity is only seen in high-grade
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F.,,. ~ SB, e,rd DR,

NV,~--
Com"""' OC:, .. tl,- AJCCClw8r511,py-.ol: 7111 ed. N'"' - ·
201G.llood- p e - .
SUGGESTED READINGS
Brandwein-Gensler MS, Mahadevia P, Gnepp DR. Nonsquamous pathologic diseases of the
hypopharynx, larynx, and trachea. In: Gnepp DR, ed. Diagnostic Surgical Pathology of the
Head and Neck. Philadelphia, PA: W.B. Saunders Publishers; 2009.
GaleN. Benign neoplasms of the larynx, hypopharynx, and trachea. In: Thompson LDR, ed. Head
and Neck Pathology. New York, NY: Churchill Livingstone; 2006.
Slootweg PJ, Richardson M. Squamous cell carcinoma of the upper aerodigestive system. In: Gnepp
DR, ed. Diagnostic Surgical Pathology of the Head and Neck. Philadelphia, PA: W.B. Saunders
Publishers; 2009.
Thompson LDR. Malignant neoplasms of the larynx, hypopharynx, and trachea. In: Thompson
LDR, ed. Head and Neck Pathology. New York, NY: Churchill Livingstone; 2006.
Thompson LDR. Non-neoplastic lesions of the larynx, hypopharynx, and trachea. In: Thompson
LDR, ed. Head and Neck Pathology. New York, NY: Churchill Livingstone; 2006.
Zidar N, Boffetta P. Tumours of the hypopharynx, larynx, and trachea. In: Barnes L, Eveson JW,
Reichart P, Sidransky D, eds. Pathology and Genetics Head and Neck Tumours. Lyon, France:
lARC Press; 2005.
Nasal Cavity,
Paranasal Sinuses,
and Nasopharynx
Heather N. Wright and James S. Lewis Jr.
I. NORMAL ANATOMY
A. Nasal cavity. The normal sinonasal region consists of the central nasal cavity,
paired bilateral paranasal sinuses, and the nasopharynx. The nasal cavity con-
sists anteriorly of the nasal vestibule, the small hair-bearing region just inside
the nasal ostia, with the remainder representing the nasal antrum; the nasal
cavity has four walls, a central dividing septum, and paired upper, middle, and
lower turbinates. The nasal vestibule lining is an extension of the surround-
ing facial skin, and as such has a stratified, keratinizing squamous epithelium
with associated dermal appendages and haiL It extends for 1 to 2 em into the
nasal cavity. The nasal antrum is lined by pseudostratified ciliated columnar
(respiratory-type) epithelium of ectodermal origin referred to as the Schneide-
rian membrane (e-Fig. 3.1).* The submucosa consists of minor salivary gland
mucoserous glands embedded in fibrovascular connective tissue with small ducts
that convey their secretions to the surface. The turbinates have a more richly
vascular stroma. The roof of the nasal cavity contains the cribriform plate with
olfactory mucosa, a modified respiratory-type epithelium with olfactory nerve
cells, and supporting cells.
B. Paranasal sinuses. The paranasal sinuses consist of the maxillary (largest),
frontal, sphenoid, and ethmoid sinuses. They drain into the nasal cavity and
are air filled, intraosseous, and open. The ethmoid sinuses are small and com-
plex (referred to as the ethmoid labyrinth or air cells). All paranasal sinuses
are in continuity with the nasal cavity so they have a similar, although thinner,
mucosa. The submucosa is also thinner, looser, and less vascular than in the
nasal cavity, although it does contain prominent seromucinous glands.
C. Nasopharynx. The nasopharynx is the most cephalad portion of the pharynx and
is a cuboidal structure. Its roof is formed by the pharyngeal tonsil. The lateral
walls are the most pathologically important because they contain the openings of
the Eustachian tubes, and a depression posterior to the torus tubarius called the
fossa of Rosenmiiller. The fossa of Rosenmiiller is the most common site of origin
for nasopharyngeal carcinoma (NPC). Since the nasopharynx is surrounded by
bone and vital structures, it is poorly accessible for surgery. The epithelial lining
consists of a mixture of stratified squamous, intermediate (or transitional), and
respiratory-type epithelium.
A. Endoscopic biopsies. Most of the specimens from this region consist of endo-
scopic forcep biopsies. The small tissue pieces should be placed immediately
into 10% buffered formalin or other appropriate fixative. If there is a suspicion
of lymphoma, a minimum of three biopsy passes should be submitted in saline
or RPMI medium and send directly for an appropriate hematopathology work-
up. Standard formalin-fixed specimens should undergo gross examination and
description documenting the exact number of pieces present, and then should
34
Chapter 3 • Nasal Cavity, Paranasal Sinuses, and Nasopharynx I 35
be entirely submitted for histologic examination (three hematoxylin and eosin

[H&E] levels cut per block). For very small specimens, or where few pieces
are obtained from clinical masses, it is strongly recommended that additional
unstained slides be cut from the block on initial submission for potential use for
immunohistochemistry or special stains.
B. Functional endoscopic sinus surgery (FESS; "sinus contents"). The tissue consists
of fragments of ethmoid and maxillary ostium sinus bone and mucosa, resected
inflammatory polyps, and nasal cavity and sinus tissue obtained by suction
devices. These samples should be described, measured, and submitted for histo-
logic examination. If firm or dense tissue fragments are identified, they should
be submitted as they may represent an unsuspected neoplasm. Otherwise, pieces
of intact tissue should be collected from the blood and fluid from the suction
device, inspected grossly, and measured in aggregate. Only one cassette of these
fragments need be submitted for histologic examination with decalcification in
EDTA or formic acid if gross pieces of bone are identified.
C. Resections. Surgical resections for sinonasal tumors are complex and varied,
and are guided by tumor location, extent, and type. They always consist of
soft tissue, mucosa, and bone. Margins are in large part guided by the separate
specimens submitted for frozen section. The main specimen should be oriented,
the tumor identified, and mucosal, soft tissue, and bone margins identified. The
main specimen should be described and measured; the soft tissue margins are
inked and then mucosal and soft tissue margin sections sampled. If the tumor is
relatively distant from a margin, 1 to 2 mm thick shave sections are preferred;
if the tumor approximates a margin (within 1 to 2 mm), radial sections are
taken. After this, the tumor is sectioned and sampled. Four to five sections
should be taken from tumors, or if small, tumors should be entirely submitted
for histologic examination. Often sectioning requires a saw to cut through bone
to demonstrate the relationship of the lesion to the adjacent structures. The
bone should be decalcified, and shave sections from the bone margins as well as
sections demonstrating tumor involving bone should be taken.
D. Frozen sections are a critical element of surgical therapy for head and neck
tumors. Although practices vary, most institutions have margins taken as small
pieces by the surgeon from the periphery of the surgical defect after the tumor
has been removed. The pieces should be evaluated grossly for mucosa (which is
typically shiny and pink-tan on one surface of the tissue); if present, the tissue
should be oriented to demonstrate this mucosal surface on one edge of the sec-
tion. Two (or at our institution, three) high quality sections are obtained with the
second and third levels taken deep into the tissue to ensure adequate sampling.
The tissue that remains after a frozen section should be submitted for evalua-
tion on permanent sections. This is done to further assure adequate sampling of
the tissue and to help resolve a number of issues from frozen section including
freezing and cautery artifact, amount of tumor represented, and orientation/
embedding issues. The final margin status is therefore a conglomerate of the
frozen section slides, permanent slides of the frozen tissue, and the margins of
the resection specimen itself.
A. Inflammation and infection. These processes are extremely common in the United
States, necessitating a large amount of medical care and surgery.
1. Acute rhinosinusitis. Acute sinusitis is rarely seen by the pathologist because it
is treated medically. However, typical histologic findings include neutrophils
migrating through, and present in, the respiratory-type mucosa with luminal
contents showing necrotic material, apoptotic neutrophil nuclear debris, and
mucin with abundant neutrophils.
2. Chronic rhinosinusitis. Chronic inflammation of the nasal cavity can result
from allergy, upper respiratory tract infection, or cystic fibrosis. Sinusitis is
thought to occur secondary to obstruction of the outflow of the paranasal

sinuses by myriad etiologies such as edema and inflammation, or anatomic
abnormalities, particularly in children. Some of the most common obstruct-
ing agents are inflammatory polyps, a deviated septum, or concha bullosa
(air pocket in the middle turbinate). Finally, rare genetic conditions such
as immotile cilia syndrome (Kartagener's syndrome) cause chronic sinusitis.
Complications include secondary bacterial infection and, in chronic allergic
sinusitis, the development of inflammatory polyps.
Specimens from FESS in chronic sinusitis typically show edema of the
submucosa with a mixture of lymphocytes, plasma cells, and eosinophils,
the latter sometimes being quite prominent. The abundance and distribu-
tion of eosinophils histologically do not have a clinical correlate other than
suggesting allergy as an underlying etiology for the sinusitis.
3. Wegener granulomatosis is an autoimmune disorder characterized by necrotiz-
ing vasculitis that affects the nasal cavity and paranasal sinuses, pulmonary,
and/or renal systems. Manifestations of nasal cavity/paranasal sinus involve-
ment include rhinorrhea, sinusitis, headache, nasal obstruction, anosmia, and
sometimes middle ear and mastoid symptoms if inflammation obstructs the
Eustachian tube.
Histologically, in biopsies of the sinonasal region, the diagnosis can be
quite difficult. Features include mucosal ulceration, acute and chronic inflam-
mation, necrosis, and granulomas. Wegener granulomatosis causes a vas-
culitis which is often obscured by the inflammation, so elastic stains such
as Verhoeff-van Gieson may be helpful to demonstrate the elastic fibers of
inflamed vessels. The vasculitis involves arterioles, small arteries, and veins
with changes ranging from fibrinoid necrosis with neutrophils and associated
extravasated red blood cells and fibrin thrombi, to granulomatous inflam-
mation with multinucleated giant cells and histiocytes (e-Fig. 3.2). It is very
important to correlate the histologic findings with clinical information and
laboratory investigation, which reveals cytoplasmic antineutrophil antibod-
ies (cANCA) in approximately 90% of patients.
4. Inflammatory polyps. Sinonasal inflammatory polyps are nonneoplastic
mucosal and submucosal projections that arise in longstanding chronic rhini-
tis, usually associated with allergy or asthma. They are seen most commonly
in adults but can be seen in children as well, particularly in those with cys-
tic fibrosis. Symptoms include headache, nasal obstruction, and rhinorrhea.
They are multiple, often bilateral, and most commonly arise from the lateral
nasal wall. Although nonneoplastic, they are capable of dramatic behavior
including deviation of the septum, destruction of bone, and extension into
the nasopharynx and rarely the orbit or cranial cavities.
Grossly, inflammatory polyps can measure up to several centimeters and
are boggy, gelatinous, and partially translucent with broad bases. Microscop-
ically, there is a highly edematous or lightly myxoid stroma with a mixed
inflammatory infiltrate of lymphocytes, plasma cells, and a variable number
of eosinophils. There are few small vessels and minimal, bland, spindled stro-
mal cells. Inflammatory polyps typically are devoid of seromucinous glands,
a feature that is particularly helpful in identifying them when they are in frag-
mented pieces (e-Fig. 3.3 ). The surface mucosa is typically intact and lined
by respiratory epithelium with a variably thickened basement membrane,
although it occasionally shows squamous metaplasia.
5. Fungal infections are relatively frequent and can involve any of the paranasal
sinuses. They can be broadly classified as "invasive" and "noninvasive"
(Table 3.1).
a. Noninvasive. Immunocompetent patients usually develop noninvasive fun-
gal disease, either allergic fungal sinusitis or mycetoma ("fungus ball").
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b. Invasive. Patients with invasive fungal disease are usually immunocom-

promised, frequently from diabetes mellitus, bone marrow or solid organ
transplantation, and occasionally human immunodeficiency virus (HIV)
infection, and are at great risk of morbidity and mortality.
Patients are often severely ill with fever, cough, nasal discharge,
headache, and mental status changes. They sometimes have ophthalmo-
logic symptoms or neurologic deficits due to orbital or cranial involve-
ment. On endoscopy, there are dark ulcers on the mucosa and associated
black, greasy necrotic tissue. The fungi are angioinvasive, so they cause
extensive hemorrhage and necrosis. In the viable tissue, the microscopic
findings include minimal inflammation and blood vessels filled with refrac-
tile fungal hyphae (e-Fig. 3.6). The order Mucorales is most common (e.g.,
Mucor, Rhizopus, Absidia), but other fungi including Aspergillus species
may be causative. The morphology of Mucor species includes bizarre,
angulated hyphal fragments and elongated hyphae that are wide, irregu-
lar, and thick walled. Typically there is 90-degree angle branching without
septation. Diabetics may also develop a chronic pattern of invasive fun-
gal disease, often presenting as a slowly progressive orbital mass; micro-
scopically, angioinvasive hyphae or granulomas are often present in these
patients.
B. Nonneoplastic lesions. Several lesions, developmental or mechanical, can occur
in the sinonasal region and can be mass-like and simulate true neoplasms.
1. Mucous impaction is an uncommon lesion that occurs mostly in children and
young adults with a long history of chronic sinusitis. It represents impaction
of a large amount of mucus within the maxillary antrum. Grossly, it con-
sists of translucent gray to pink material. Microscopically, it simply consists
of slightly basophilic to eosinophilic extracellular mucin with a mixture of
plasma cells, lymphocytes, and neutrophils with desquamated respiratory-
type epithelium.
2. Paranasal sinus mucoceles are chronic, nonneoplastic cysts that form from
the obstruction of the sinus outlet by any of a number of processes. They
occur most commonly in the ethmoid and frontal sinuses. Grossly, they con-
sist of a cyst filled with mucoid or gelatinous material. Microscopically, they
consist of extracellular mucin with a flattened respiratory-type epithelial lin-
ing that may have secondary squamous metaplasia.
3. Respiratory epithelial adenomatoid hamartoma (REAH) is a rare, benign, poly-
poid lesion characterized by an adenomatoid proliferation of respiratory-type
epithelium that occurs primarily on the posterior nasal septum. Microscop-
ically, it consists of a polypoid proliferation of variably sized, round to oval
glands lined by respiratory-type epithelium with a markedly thickened base-
ment membrane and no cytologic atypia or significant mitotic activity (e-Fig.
3.7). The gland-like structures are often in direct continuity with the surface
epithelium (e-Fig. 3.8). The stroma is edematous and can resemble that of
an inflammatory polyp.
C. Neoplastic lesions. A wide array of neoplasms occur in the sinonasal region
(Table 3.2).
1. Benign
a. Schneiderian papillomas. The ectodermally derived Schneiderian mem-
brane gives rise to three different types of benign papillomas: exophytic,
inverted, and oncocytic (Table 3.3). The distinction of these papillomas is
important because of their differences in behavior and risk of carcinoma
development. For this reason, it is critical to entirely submit all papilloma
tissues for microscopic examination.
i. Exophytic papilloma (fungiform papilloma). Most of these occur on the
nasal septum, particularly anteriorly, in adults 20 to 50 years old. They
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epithelium characteristically has abundant intraepithelial neutrophils,
singly and in clusters, as well as numerous mucin-filled microcysts.
Some inverted papillomas can have a prominent exophytic-appearing
component. However, any Schneiderian papilloma with a significant
inverted and/or downward pushing component (more than just rare
nests) should be diagnosed as an inverted papilloma.
Uncommonly, there is surface keratinization and 5% to 20% of
these lesions show varying degrees of dysplasia. Interestingly, dyspla-
sia by itself does not signify malignancy or a different clinical course
for the patient, but it does make it important to thoroughly evalu-
ate the lesion for coexisting carcinoma. Approximately 10% to 15%
of inverted papillomas are complicated by carcinoma, mostly squa-
mous cell carcinoma. In approximately 60% of cases, the malignancy
is synchronous and in 40% of cases, the malignancy is metachronous.
A significant number of inverted papillomas will recur, particularly
after conservative resection.
iii. Dncocytic papillomas are the least common Schneiderian papillomas
and have various alternate names including cylindrical cell and colum-
nar cell papilloma. They occur in the same sites as inverted papillo-
mas, namely, the lateral nasal wall and paranasal sinuses. Unlike other
Schneiderian papillomas, they have an equal male to female ratio, and
studies have not identified HPV in them.
Microscopically, they typically have a mixture of exophytic and
endophytic components and are lined by a unique two- to eight-cell
layer of tall columnar oncocytic cells that have abundant granular,
eosinophilic cytoplasm. The nuclei are slightly more atypical than in
other Schneiderian papillomas with a wrinkled, dark to slightly vesicu-
lar appearance with small nucleoli. The epithelium contains numerous
mucin-filled microcysts and intraepithelial neutrophils (e-Fig. 3.12).
Oncocytic papillomas have a risk of local recurrence and carci-
noma similar to that of inverted papillomas.
b. Verruca vulgaris (nasal vestibule}. As the nasal vestibule is essentially an
extension of the surrounding nasal skin, verrucae occur here and have the
same morphology as elsewhere (see Chap. 38).
2. Malignant
a. Sino nasal undifferentiated carcinoma (SNUC}. This tumor is a clinicopatho-
logically distinct, aggressive form of sinonasal carcinoma. Patients are
most commonly in their sixth decade and present with symptoms of short
duration, including nasal obstruction and epistaxis, and often have orbital
or cranial nerve deficits. The tumor most often arises in the nasal cavity,
particularly superiorly, or in the maxillary or ethmoid sinuses. It is usually
bulky, often involving contiguous sites.
There are no unique gross features. Tumors are usually larger than 4
em, poorly defined, and invade bone. Microscopically, different growth
patterns can be seen including nested, trabecular, or lobular, although the
nests are not usually well defined. The tumor cells are small to moderate in
size, have a small to moderate amount of cytoplasm with distinct cell bor-
ders, and have nuclei that range from hyperchromatic to vesicular, often
with prominent nucleoli. Despite being overtly malignant, the individual
tumor cells tend to be remarkably regular in size. There is abundant apop-
tosis, brisk mitotic activity, and often prominent necrosis (e-Fig. 3.13). By
definition, there is a lack of definable differentiation.
Immunohistochemistry is positive for pan-cytokeratin and epithe-
lial membrane antigen. More specifically, simple keratins such as 7, 8,
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different staging system that reflects this unique behavior (see AJCC stag-
ing below). Finally, despite presenting with advanced disease, nonkera-
tinizing NPC responds well to systemic therapy (5-year survival of approx-
imately 65% in the United States). Keratinizing NPC, although it more
often presents with localized disease, does not respond well to therapy
(5-year survival of approximately 20% to 40% in the United States).
c. Squamous cell carcinoma/nonkeratinizing carcinoma. Although squamous
cell carcinoma is the most common carcinoma in most head and neck
anatomic subsites, it constitutes only approximately 65% of carcinomas
in the nasal cavity and paranasal sinuses. When it occurs in the nasophar-
ynx, it is designated keratinizing squamous cell carcinoma (WHO Type 1;
see section III.C.2.b above and Table 3.4). Keratinizing, or typical, squa-
mous cell carcinoma of the nasal cavity and paranasal sinuses has the
same morphology as that occurring elsewhere in the upper aerodigestive
tract. Squamous cell carcinoma of this subsite, unlike other head and neck
subsites, has only a modest association with smoking. Howevet; tumors
have been related to other exposures including nickel, chlorophenols, and
textile dust.
A distinct variant of nonkeratinizing squamous cell carcinoma occurs
in this region, the so-called cylindrical cell (Schneiderian or transitional)
carcinoma. The recent WHO classification simply terms it nonkeratinizing
carcinoma. It has a papillary configuration with ribbons of invaginating
tumor composed of pleomorphic cells without keratinization. These rib-
bons have a smooth border without clear stromal infiltration, which makes
identification of frank invasion difficult (e-Fig. 3.19). This tumor type has
been associated with HPV and is also reported by several authors to have
a better prognosis, although the latter contention is still controversial.
d. Salivary gland-type tumors constitute a small percentage of sinonasal neo-
plasms. They arise from submucosal seromucinous glands. The most com-
mon are adenoid cystic carcinoma and pleomorphic adenoma, although
almost all other types have been described (e-Figs. 3.20 to 3.22). They are
morphologically identical to their counterparts elsewhere.
e. Adenocarcinoma. Primary non-salivary gland-type adenocarcinomas of
the sinonasal region are uncommon. The WHO classifies them as intesti-
nal and nonintestinal types, the latter of which are subdivided into high-
and low-grade tumors.
i. Intestinal type. Primary intestinal-type adenocarcinomas (ITACs) of the
sinonasal region typically arise from the ethmoid sinuses or high in the
nasal cavity. They have a strong association with long-term occupa-
tional exposures, specifically wood dust (in carpenters), leather dust,
nickel, or chromium compounds. The latency period is several decades.
Microscopically, these tumors recapitulate gastrointestinal adenocar-
cinomas, including very well-differentiated tumors resembling colonic
adenomas, typical "colonic-appearing" tumors with columnar glands
with or without mucinous differentiation (e-Fig. 3.23 ), and high-grade
tumors with a signet ring cell morphology.
By immunohistochemistry, they are positive for cytokeratin 20,
CDX-2, MUC2, and villin, similar to gastrointestinal adenocarcinoma.
Cytokeratin 7 is variably positive and carcinoembryogenic antigen
(CEA) is usually positive. As such, they stain essentially identical to
gastrointestinal adenocarcinomas, so it can be very difficult to separate
them from the rare metastasis to this region from a primary gastroin-
testinal adenocarcinoma.
ITACs should be graded from poorly to well differentiated, and the
type (colonic, mucinous, papillary, signet ring cell, or solid) specified, as
44 I sEcT I 0 N I: HEAD AND NEcK
both have prognostic significance. The mortality rate is approximately

50% overall.
ii. Nanintestinal type. These are uncommon tumors, most of which are
low grade and have a predilection for the ethmoid sinuses. Low-grade
tumors have a glandular or papillary growth pattern with numerous
uniform, small glands arranged back-to-back and lined by a single layer
of cuboidal to columnar cells with a moderate amount of eosinophilic
to dear cytoplasm and round nuclei. There is only mild nuclear pleo-
morphism, modest mitotic activity without atypical forms, and no
necrosis. Controversy exists as to what defines them as frankly malig-
nant, histologically. Infiltrative growth is usually a critical feature to
identify. High-grade tumors typically occur in the maxillary sinus and
have similar histologic features, but also have solid areas and show
moderate to severe nuclear pleomorphism with brisk mitotic activity
and necrosis.
By immunohistochemistry, nonintestinal-type adenocarcinomas are
positive for cytokeratin 7 and negative for cytokeratin 20, CDX-2, and
MUC2. Smooth muscle actin and p63 are negative. The prognosis for
low-grade tumors is excellent, but for high-grade tumors is quite poor.
f. Neuroendocrine carcinomas occasionally arise in the sinonasal region and
are morphologically identical to those arising in the lung.
i. Law-grade neuroendocrine carcinoma (i.e., carcinoid tumor) is
extremely rare in this region.
ii. Intermediate-grade neuroendocrine carcinoma (i.e., atypical carcinoid
tumor) is also extremely rare in this region.
iii. High-grade neuroendocrine carcinoma (small cell carcinoma) is a
rapidly proliferating, aggressive neoplasm that presents with epis-
taxis, nasal obstruction, and exophthalmos. It usually presents at an
advanced stage with extensive local disease and lymph node or dis-
tant metastases. Microscopically, it consists of nests and sheets of
small- to intermediate-sized blue cells with minimal cytoplasm, speck-
led chromatin, frequent nuclear molding, and crush artifact (e-Fig.
3.24). Apoptosis is abundant, and there is brisk mitotic activity and
necrosis. By immunohistochemistry, it is positive for pan-cytokeratin
with a "dot-like" paranuclear staining pattern (e-Fig. 3.25A). It is vari-
ably positive for the neuroendocrine markers CD56, synaptophysin,
and chromogranin A, but in most cases at least one of these markers
is positive on careful inspection (e-Fig. 3.25B). The tumor is negative
for S-100, CD99 (013), and cytokeratin 20.
g. Melanoma. Primary malignant melanoma of the sinonasal tract constitutes
approximately 1% of all melanomas. It is more common in the nasal cavity
than in the paranasal sinuses, and most patients are over 50 years old, with
tumors most often occurring on the middle or inferior turbinate, or ante-
rior septum. Although variable, the typical macroscopic appearance is a
sessile or polypoid lesion with mucosal ulceration. Most lesions are heavily
pigmented with a brown or black color. Microscopically, melanoma has a
wide range of appearances, although usually it is composed of high-grade
epithelioid cells. Some tumors have a mixture of epithelioid and spindle
cells. A junctional component with pagetoid spread of melanocytes in the
intact mucosa is also sometimes present.
Immunohistochemistry results are the same as for melanomas at other
sites, with virtually all tumors expressing S-1 00, HMB-4 5, and melan-A.
Cytokeratin reactivity, if present, is only focal.
Metastasis of melanoma to the sinonasal region from another pri-
mary site is not particularly uncommon and should always be considered.
Although metastases can recapitulate a primary lesion, a junctional com-

ponent in the surface mucosa essentially rules out metastasis.
The prognosis for sinonasal melanoma is very po01; with frequent local
recurrence and distant metastasis. Sinonasal melanomas have their own
AJCC staging system, and, unlike cutaneous melanomas, tumor (Breslow)
thickness has not been shown to predict behavior.
h. Olfactory neuroblastoma. This unique tumor arises almost exclusively in the
upper nasal cavity from the olfactory mucosa of the cribriform plate, upper
lateral nasal wall, and superior turbinate. The presumed cell of origin is
the reserve cell that gives rise to neuronal and sustentacular cells of the
olfactory mucosa. The tumor occurs at any age but most commonly in the
third and fourth decades, with symptoms of nasal obstruction, epistaxis,
and anosmia.
The neoplasm is usually a unilateral, polypoid mass, microscopically
composed of small, round cells that are slightly larger than lymphocytes.
The nuclei are round, with uniform to delicately stippled chromatin with-
out nucleoli. Many tumors have areas with fibrillary eosinophilic mate-
rial reminiscent of neuropil (e-Fig. 3.26). Some tumors have a very lob-
ulated and nested growth pattern, whereas others have a diffuse pattern.
Homer-Wright rosettes (that have no central lumen) are relatively com-
mon. Mitotic activity is low. Necrosis and dystrophic calcification are
uncommon. Some tumors have been reported to show significant nuclear
pleomorphism and high mitotic activity. However, this is very uncommon
and merits consideration of another neoplasm, particularly high-grade
neuroendocrine carcinoma. Rare findings include ganglion cells, melanin-
containing cells, and divergent differentiation including glandular, squa-
mous, and teratomatous features.
By immunohistochemistry, olfactory neuroblastoma shows strong
cytoplasmic staining for synaptophysin, chromogranin A, and NSE
(e-Fig. 3.27). S-100 often highlights sustentacular cells at the periphery of
tumor nests, but is negative in tumor cells themselves. Although unusual,
cytokeratin reactivity has been reported in up to 35% of cases, specifically
CAM 5.2 and less often AE1/AE3. The staining is typically weaker and
more focal than would be expected in a carcinoma. Recent reports have
shown calretinin staining to be positive in olfactory neuroblastoma and
negative in many other tumors in the differential diagnosis.
Grading systems, most notably by Hyams, have been proposed, but
their correlation with prognosis has not been consistently demonstrated.
i. Mesenchymal tumors
i. Angiofibroma {nasopharyngeal angiofibroma). This unique tumor is
thought to arise from a fibrovascular nidus in the posterolateral nasal
wall adjacent to the sphenopalatine foramen. It occurs virtually exclu-
sively in boys aged 10 to 20 years. The tumor often presents as a
nasopharyngeal mass due to its pushing, well-circumscribed border,
which bulges posteriorly into the nasopharynx as it enlarges, causing
nasal obstruction and epistaxis.
Grossly, the tumor is gray-white to tan, smooth, and lobulated,
with a homogeneous cut surface. Microscopically, it consists of abun-
dant vessels ranging from capillaries to large vessels, the latter often
assuming a "staghorn" appearance (e-Fig. 3.28). The vessels are lined
by a single layer of endothelial cells that can be flat to slightly plump
without cytologic atypia. The vessel wall is typically devoid of smooth
muscle and blends imperceptively with the stromal component of
the tumor, which has regularly distributed, plump, stellate spindle
cells embedded in a dense collagenous stroma. The stromal cells have
41 SECTION 1, HEAD AHD NECK
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50 SECTION 1: HEAD AND NECK
B. Additional pertinent pathologic features. Several pathologic features not reflected

in the AJCC staging have been proven important for head and neck squamous
cell carcinomas, as well as for numerous other tumor types, including perineural
invasion, lymphovascular space invasion, and positive margin status. All of these
features have been demonstrated in numerous studies to correlate with a higher
risk of local recurrence, a poorer prognosis, or both.
SUGGESTED READINGS
Barnes L, Tse LLY, Hunt JL, et al. Tumours of the nasal cavity and paranasal sinuses. In: Barnes L,
Eveson JW, Reichart P, Sidransky D, eds. Pathology and Genetics Head and Neck Tumours.
Lyon, France: IARC Press; 2005.
Brandwein-Gensler M, Thompson LDR. Non-neoplastic lesions of the nasal cavity, paranasal
sinuses, and nasopharynx. In: Thompson LDR, ed. Head and Neck Pathology. New York,
NY: Churchill Livingstone; 2006.
ChanJKC, Pilch BZ, Kuo TT, et al. Tumours of the nasopharynx. In: Barnes L, Eveson]W, Reichart
P, Sidransky D, eds. Pathology and Genetics Head and Neck Tumours. Lyon, France: IARC
Press; 2005.
Perez-Ordonez B, Thompson LDR. Benign neoplasms of the nasal cavity, paranasal sinuses, and
nasopharynx. In: Thompson LDR, ed. Head and Neck Pathology. New York, NY: Churchill
Livingstone; 2006.
Prasad, ML, Perez-Ordonez B. Nonsquamous lesions of the nasal cavity, paranasal sinuses, and
nasopharynx. In: Gnepp DR, ed. Diagnostic Surgical Pathology of the Head and Neck. Philadel-
phia, PA: W.B. Saunders Publishers; 2009.
Thompson LDR. Malignant neoplasms of the nasal cavity, paranasal sinuses, and nasopharynx. In:
Thompson LDR, ed. Head and Neck Pathology. New York, NY: Churchill Livingstone; 2006.
Tumors and Cysts
of the Jaws
David E. Spence and Samir K. EI-Mofty
I. NORMAL ANATOMY. Of all the bones of the skeleton, the jaws are uniquely distin-
guished by harboring the odontogenic apparatus of the deciduous and permanent
dentitions. The teeth germs are composed of three main components: the enamel
organ, the dental papilla, and the tooth follicle. The enamel organ is composed of
ectodermally derived epithelial cells, the ameloblasts, and is responsible for enamel
formation. The dental papilla is of ectomesenchymal origin and produces the den-
tine. The tooth follicle is also ectomesenchymal; it surrounds the developing tooth
and provides the supporting structures of the formed teeth, the periodontium. The
odontogenic tissues may also be a source of a bewildering array of odontogenic
cysts and tumors. Nonodontogenic cysts and tumors of the jaws will also be dis-
cussed in this chapter.
II. ODONTOGENIC AND NONODONTOGENIC CYSTS. Odontogenic cysts of the mandible
and the maxilla are relatively common and can present over a large age range.
Accurate diagnosis is simplified by location, radiographic correlation, and micro-
scopic examination (e-Fig. 4.1)."' Odontogenic tumors by contrast are uncommon.
They may be epithelial, mesenchymal, or mixed; may be noncalcifying; or they may
contain hard structures that mimic enamel, dentine, cementum, or bone. Although
malignant odontogenic tumors are extremely rare, odontogenic carcinoma, sar-
coma, and carcinosarcoma do occur.
A. Odontogenic cysts. With the exception of a few cysts that may develop along
embryonic lines of fusion (known as "nonodontogenic cysts"), most jaw cysts
are lined with epithelium that is derived from the odontogenic epithelium. Odon-
togenic cysts are classified as either developmental or inflammatory. Various
types of odontogenic cysts are listed in Table 4.1.
1. Developmental odontogenic cysts
a. Dentigerous cyst (follicular cyst) is a unilocular cyst that forms in associa-
tion with the crown of an impacted tooth and is usually associated with
a molar or canine. Patients usually present with an asymptomatic, well-
defined, expansive, radiolucent lesion. Microscopic examination shows
a thin layer of cuboidal to slightly flattened epithelial cells. Focal kera-
tinization, mucous cells, inflammation, and dystrophic calcification are
possible. Enucleation and excision of the associated tooth are the treat-
ment of choice. Eruption cyst is a subclass of dentigerous cysts associated
with the erupting primary or permanent tooth.
b. Keratocystic odontogenic tumor (odontogenic keratocyst) usually presents
as an asymptomatic unilocular cyst in the mandible or the maxilla. While
more frequent in the second to fourth decades of life, they may be seen at
any age. Their incidence is higher in Caucasians, especially in and men, and
roughly 5% of patients with odontogenic keratocysts have nevoid basal
cell carcinoma (Gorlin) syndrome. Gross examination typically yields a
cyst containing keratinous debris. Microscopic examination shows pal-
isading basal cells covered by a few layers of squamous cells under a
51
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Chapter 4 • Tumors and Cysts of the Jaws I 53
solid areas (e-Fig. 4.5A and B). Hyaline bodies, ghost cells, and
ciliated cuboidal eosinophilic cells may be present. Care should be
taken to differentiate between a glandular odontogenic cyst and a
central mucoepidermoid carcinoma. Surgical excision is the treatment
of choice.
2. Inflammatory odontogenic cysts. Radicular (periapical) cysts are associated
with a nonviable carious teeth. The usual location is the apical third of the
tooth root, with occasional cases involving the lateral root surface. The cyst
is more common in the mandible. The typical age of patients is the third to
sixth decades of life. The most common presentation is pain and swelling, but
presentation as an incidental finding on routine radiographic examination is
not unusual. Microscopic examination reveals an inflamed, nonkeratinizing,
stratified epithelium. Cholesterol crystals, foamy macrophages, dystrophic
calcifications, and intraepithelial hyaline bodies (Rushton bodies) may be
identified (e-Fig. 4.6A and B). The residual cyst is a variant of radicular cyst
that is seen at the site of an extracted tooth.
B. Nonodontogenic cysts. This class of lesions includes a group of epithelium-lined
cysts as well as non-epithelium-lined bone cysts. The epithelial-lined cysts are
believed to arise from epithelial remnants entrapped along embryonic lines of
fusion and are referred to as fissural cysts (e-Fig. 4.7).
1. Epithelial-lined nonodontogenic cysts {fissural cysts)
a. Nasopalatine duct cyst {incisive canal cyst) is the most common of the
fissural cysts. It is believed to arise from remnants of the nasopalatine
duct. The cyst can develop almost at any age, but it is most common
in the fourth to sixth decades of life. Most studies show a slight male
predilection. The most common presenting symptoms include swelling of
the anterior palate, drainage, and pain.
Radiographs usually demonstrate a well-circumscribed radiolucency,
in or near the midline of the anterior maxilla, between and apical to
the central incisor teeth. The lesion most often is round oval or pear-
shaped. Microscopically, the epithelial lining of the cyst may be strati-
fied squamous, pseudostratified columnat; simple columnar, or cuboidal.
Commonly, more than one epithelial type is present. Because the cyst
arises within the incisive canal, moderate-sized nerves and small muscular
arteries and veins are usually found in the cyst wall (e-Fig. 4.8). Surgical
enucleation is the treatment of choice.
b. Globulomaxillary cyst is believed to develop from epithelium entrapped
during fusion of the globular portion of the medial nasal process with the
maxillary process, although its origin continues to be a subject of debate.
The cyst classically develops between the lateral incisor and cuspid
teeth, although occasionally it has been reported between the central
and lateral incisors. Radiographically, the cyst presents as well-circum-
scribed unilocular radiolucency between and apical to the teeth. The radi-
olucency is often pear-shaped. As the cyst expands, tilting of the adjacent
teeth may occur. Microscopically, many of the cysts are lined with strati-
fied squamous epithelium. Occasionally, howevet; the lining epithelium is
of pseudostratified columnar ciliated type. Enucleation is the treatment of
choice.
c. Median palatal cysts (median palatine cysts) are rare fissural cysts. They
are believed to develop from epithelium entrapped along the embryonic
line of fusion of the lateral palatal shelves of the maxilla. The cyst may be
difficult to distinguish from the nasopalatine duct cysts, and some cases
may actually represent a posteriorly placed nasopalatine duct cyst.
Clinically, the cyst presents as a firm or fluctuant swelling in the midline
of the hard palate, posterior to the incisive papilla. It is more frequent in
young adults and is often asymptomatic. The average size is 2 x 2 em,

but these cysts may become quite large. Radiographs demonstrate a well-
circumscribed lucency in the midline of the hard palate. Microscopically,
the cyst is commonly lined by stratified squamous epithelium, but areas
of pseudostratified columnar epithelium may be seen. Surgical removal is
the treatment of choice.
2. Non-epithelial-lined nonodontogenic bone cysts of the jaws
a. Simple bone cysts are known by multiple names including unicameral
bone cyst, solitary bone cyst, progressive bony cavity, hemorrhagic cyst,
and traumatic bone cyst. The typical patient is younger than 20 years of
age and has a well-demarcated osteolytic solitary lesion in the posterior
mandible. Unusual cases have been seen in the maxilla. The cyst cavity is
lined by fibrovascular tissue with hemosiderin-laden macrophages. Reac-
tive bone and osteoclasts may be identified.
b. Aneurysmal bone cyst is a rapidly enlarging blood-filled cystic lesion usu-
ally identified in the first three decades of life. The majority are well-
demarcated, unilateral pseudocysts located in the mandible (60%) and
the maxilla (40%). Aneurysmal bone cyst may be identified alone or
in conjunction with another lesion (e.g., chondroblastoma, osteoblas-
toma.) CT and MRI studies show characteristic layering of blood cells
and serum. Microscopic examination shows a fibrotic stroma with giant
cells, macrophages, and hemosiderin granules (e-Fig. 4.9A and B). Areas
of ossification may be present. Treatment is curettage and enucleation,
and about 25% of the lesions recur.
Ill. ODONTOGENIC TUMORS. Odontogenic tumors are classified according to their com-
position into epithelial, mesenchymal, or mixed. Epithelial odontogenic tumors
are composed of only odontogenic epithelium, whereas mesenchymal odontogenic
tumors are composed principally of ectomesenchymal elements. Mixed odonto-
genic tumors contain epithelial and ectomesenchymal tissues. Inductive interactive
action between the epithelial and ectomesenchymal elements may mimic normal
odontogenesis and thus dental hard tissues may, on occasions, be found in these
tumors. As stated above, malignant odontogenic tumors are rare, but carcinomas,
sarcomas, and carcinosarcomas do occur.
Odontogenic tumors are listed in Table 4.2.
A. Epithelial odontogenic tumors
1. Ameloblastoma is the most common clinically significant odontogenic tumor.
Its relative frequency equals the combined frequency of all other odontogenic
tumors, excluding odontoma. Ameloblastoma is slow-growing, locally inva-
sive neoplasm. The tumor is encountered over a wide age range; it is rare in
children and is most prevalent in the third to seventh decades of life. There is
no gender predilection. Some studies show an increased frequency in blacks.
About 85% of ameloblastomas occur in the mandible, most often in the
molar-ascending ramus area. About 15% of ameloblastomas occur in the
maxilla, usually in the posterior region. Painless swelling or expansion of
the jaw is the usual clinical presentation. If untreated, the lesion may grow
slowly to massive or grotesque proportions. Pain and paresthesia are uncom-
mon, even in large tumors. Radiographically, the most typical feature is that
of a multilocular radiolucent lesion. Cortical expansion is frequently present.
Microscopically, the lesion may present several patterns; these microscopic
patterns have no bearing on the behavior of the tum01; and large tumors
often show a combination of patterns.
The follicular pattern is the most common and recognizable. It is com-
posed of nests of epithelium that resemble the enamel organ of the developing
teeth, dispersed in a mature fibrous connective tissue stroma. The core of the
nests is composed of loosely arranged angular cells that resemble the stellate
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from 1bt boo-eat m.cml><a<>e (to<alle4 rmtte4 poWily) (eo~ .oJ.lO). Cy1t
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Unless removed in its entirety, ameloblastoma has a high recurrence rate.

The tumor cells tend to infiltrate the surrounding marrow spaces, and the
actual margin of the tumor often extends beyond its apparent radiographic
or clinical margin. Marginal resection is therefore the most widely used
treatment, but it is still associated with recurrence rates of up to 15%, and
thus many surgeons advocate that the margin of resection should be at least
1.0 em past the radiographic limit of the tumm:.
2. Calcifying epithelial odontogenic tumor (Pindborg tumor) is a rare tumor that
presents as a slowly growing painless expansive lesion that favors the pos-
terior mandible. The peak age of incidence is the third to seventh decades
of life, and the male/female ratio is equal. The radiographic appearance of
the tumor changes over time. Early on, the tumor appears as a radiolucent
lesion that can be mistaken for a cyst. As the lesion ages, it develops a poorly
demarcated border and multiple radio-opaque foci. The lesion may be mul-
tilocular.
On microscopic examination, the tumor is characterized by clusters of
pleomorphic polyhedral epithelial cells with a well-defined cell border and
dense nuclear staining. The cells show mild to moderate nuclear pleomor-
phism, rare mitotic figures, and may contain multiple nuclei. Layered calcifi-
cations and amyloid-like globules are usually present (e-Fig. 4.12). Calcifying
epithelial odontogenic tumors are generally treated surgically. Because they
tend to be infiltrating tumors, treatment should include removal with a bor-
der of clinically and radiographically normal bone.
3. Adenomatoid odontogenic tumor (adenoameloblastoma) typically presents as a
slowly growing asymptomatic mass in the anterior portion of the maxilla or
the mandible in patients younger than 30 years. Women are affected twice as
often as men. Radiographic examination demonstrates a radiolucent, well-
defined lesion involving the crown of an unerupted/impacted tooth. Adjacent
teeth may show root divergence without root resorption. Gross examination
reveals a well-defined encapsulated mass of soft tissue with focal cystic and
granular areas. Microscopic examination shows a well-defined fibrotic cap-
sule surrounding a multinodular mass of eosinophilic spindle and polyhedral
cells. Scattered among these cells are amphophilic globules with variable lev-
els of calcification and lamination; these globules are periodic acid-Schiff
(PAS) positive and diastase resistant. In addition to the globules, small cys-
tic spaces lined by a single layer of cuboidal to columnar cells with foamy
cytoplasm and basally orient nuclei are present (e-Fig. 4.13). Treatment is
enucleation, and recurrence is extremely rare.
4. Squamous odontogenic tumors are benign lesions that occur in patients over
a wide age distribution. Patients generally present with tooth loosening in
the absence of periodontal disease. Radiology demonstrates a radiolucent
mass in the anterior maxilla or the posterior mandible with tooth root
involvement. Microscopic examination shows that the lesion is character-
ized by nodules of bland squamous cells with peripheral flattening of the
basal cells, separated by a fibrous stroma (e-Fig. 4.14). Treatment is surgical
excision with associated tooth removal.
5. Malignant ameloblastoma and ameloblastic carcinoma. Very rarely, ameloblas-
toma exhibits frank malignant behavior with development of metastasis. The
frequency of such an event is difficult to determine but probably occurs in
far less than 1% of all ameloblastomas.
By definition, malignant ameloblastoma is a tumor that shows histomor-
phologic features of a benign ameloblastoma, yet metastasizes. Metastasis is
most often to the lungs, which has been regarded as aspiration or implant
metastasis. In such cases, the first evidence of metastasis is often discovered
1 to 30 years after surgical treatment of the primary lesion.
The term "ameloblastic carcinoma" should be reserved for an ameloblas-

toma that has the cytologic features of malignancy in the primary tumor, in a
recurrence, or in any metastatic deposit (e-Fig. 4.15). These lesions typically
follow a markedly aggressive local course but metastasis does not always
occur.
B. Mesenchymal odontogenic tumors
1. Odontogenic myxoma is a benign tumor that has the potential for local infiltra-
tion with extensive bone destruction and a relatively high recurrence rate. It
is thought to be derived from the ectomesenchyme. Microscopically, it resem-
bles the dental papilla of a developing tooth. Myxomas are most common in
the second and third decades of life; although they occur in patients aged 5 to
72 years, they are uncommon in patients younger than 10 years or older than
50 years. Some studies show a female predilection. The lesion may be found
at any location in the jaws, although some studies show a predominance of
maxillary tumors.
Myxomas vary in their radiographic appearance, from small and uniloc-
ular to large and multilocular, with a "soap bubble" appearance. Tooth
displacement and cortical expansion are common in larger lesions. Maxil-
lary tumors often extend into the maxillary sinus. Microscopic examination
reveals a bland, monotonous, hypocellular proliferation of loose mesenchy-
mal fibrous tissue. The cells are spindled or stellate, with long cytoplasmic
processes. The nuclei are small and may be hyperchromatic. Mitoses are
scarce. Small nests of odontogenic epithelium may be present but are not
necessary for the diagnosis (e-Fig. 4.16A and B).
Because of their lack of encapsulation and infiltrative growth, myxomas
tend to extend beyond their clinically anticipated boundaries. Recurrence
rates are as high as 25%, and thus dose follow-up is recommended (recur-
rences are usually due to incomplete excision). Some recurrences occur years
after excision.
2. Benign cementob/astoma (true cementoma) is a distinctive mesenchymal odon-
togenic tumor which is intimately associated with the roots of teeth. It is
characterized by the formation of calcified cementum-like tissue deposited
on the tooth root, most commonly mandibular molars. Although the tumor is
detected in patients over a wide age range, it most commonly affects teenagers
and young adults. Pain is a frequent symptom. The radiographic appearance
of cementoblastoma is characteristic and is almost pathognomonic, namely,
a radio-opaque mass that obliterates the radiographic details of the root of
the affected tooth.
On microscopic examination, the peripheral part of the tumor resembles
osteoblastoma. Centrally, thick trabeculae of cementum, which are strongly
basophilic, are deposited on the intact or partially resorbed tooth root.
Peripherally, bone-like trabeculae are rimmed with plump cementoblasts.
The intervening fibrovascular tissue shows dilated vessels and occasional
dusters of multinucleated osteoclast-like giant cells (e-Fig. 4.17).
Cementoblastoma is a slowly growing benign neoplasm, but it may attain
a large size if not treated. The recommended treatment is surgical excision
with extraction of the affected tooth. Recurrence is usually a result of incom-
plete removal.
3. Cemento-ossifying fibroma (COF} of the jaws is synonymous with ossifying
fibroma and cementifying fibroma. It is a benign odontogenic neoplasm
that is limited to the tooth-bearing areas of the jaws. COF is more often
seen in the mandible (90%) and in women (83%). Most patients are in
their third to fourth decades of life, and they present with a small asymp-
tomatic expanding bone mass that does not erode the adjacent cortical bone.
Larger lesions may present with facial deformation and pain. Radiographic
examination demonstrates a well-circumscribed radiolucent lesion with

patchy focal radio-opaque areas that does not encase the teeth roots. Tooth
displacement, resorption, and root divergence may be seen.
Microscopic examination shows a lesion characterized by a fibrous
stroma with bone trabecula and associated variable mineralized material that
resembles dental cementum; either component rnay dominate in an individual
lesion. The stroma is usually hypercellular; the bone trabeculae are usually
woven but lamellar bone may also be seen. Osteoblastic rimming may vary
in extent (e-Fig. 4.18).
Most COFs are small tumors that can be shelled out or curetted out of
the jawbone with relative ease. Recurrence after adequate removal is seldom.
Incompletely excised tumors continue to grow slowly but may attain a large
SIZe.
c. Mixed odontogenic tumors
1. Odontomas represent the most highly differentiated of the mixed odontogenic
tumors. They are considered by some pathologists to be hamartomas. Two
types of odontomas are recognized, compound and complex. Compound
odontoma is composed of many, sometimes even dozens, of small miniature
teeth that are surrounded by a dental follicle, the same tissue that surrounds
a normal developing tooth. This form of odontoma shows the highest degree
of histodifferentiation and morphodifferentiation (e-Fig. 4.19). In contrast,
the complex odontoma is composed of a mass of intermixed enamel and
dentine with no resemblance to normal or miniaturized teeth (e-Fig. 4.20).
Compound odontoma occurs most often in the anterior segment of the
jaws, particularly the canine area in association with an impacted canine
tooth. It is the most common odontogenic tumor. It is found most often in
the second decade of life and is more common in males than in females by a
3:2 ratio.
Complex odontomas occur most often in the posterior segment of the
jaws, primarily in association with an impacted third molar tooth. They are
the second most common odontogenic tumors and are usually discovered in
the early third decade of life. Like compound odontomas, there is a male
gender predilection of 2:1.
Odontomas are typically discovered when radiographic examination is
performed because of a delay in eruption of a tooth. They may be mostly
radiolucent with areas of opacity and may be associated with an odontogenic
cyst (particularly dentigerous cyst) or with calcifying odontogenic cyst. They
are treated by surgical excision.
2. Ame/ob/astic fibroma is a true neoplasm composed of both epithelial and
mesenchymal types of tissues but without calcified structures. The tumor is
typically seen in adolescent patients. The average age is 14 years and there
is an equal male/female distribution. About 80% of patients present with
a well-defined unilocular or multilocular lesion in the mandible. Associa-
tion with an unerupted tooth is common. Gross examination demonstrates a
smooth well-defined lesion with a tan white cut surface. Microscopic exam-
ination demonstrates a lesion characterized by a background of immature
connective tissue that resembles dental papilla with cords, strands, and nests
of cuboidal to columnar epithelial cells. The epithelial nests are indistin-
guishable from those seen in follicular ameloblastoma, with a central stellate
reticulum-like component and peripheral palisaded columnar cells showing
reversed nuclear polarity. A prominent basement membrane separates the
epithelial cells from the stroma (e-Fig. 4.21). Treatment is by enucleation and
thorough curettage and, if necessary, extraction of the involved tooth. Recur-
rence is uncommon. Malignant transformation is extremely rare but has been
documented.
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be bone or cementum. Proper diagnosis requires correlation of historical, clini-

cal, and radiographic findings. The more important types of fibro-osseous lesions
of the jaws are discussed here.
1. Fibrous dysplasia is a skeletal anomaly in which normal bone is replaced by
poorly organized and inadequately mineralized immature woven bone and
fibrous connective tissue. Fibrous dysplasia is separated into two forms: the
polyostotic form involves multiple bones, whereas the monostotic form is
limited to a single site. Polyostotic fibrous dysplasia is less common, and a
few of these cases may be associated with skin pigmentation and endocrine
anomalies, a condition known as the "McCune-Albright syndrome." The
craniofacial skeleton is involved in 20% to 25% of cases of monostotic
fibrous dysplasia, particularly the mandible and the maxilla.
Craniofacial fibrous dysplasia is not strictly monostotic but may extend
by continuity to adjoining bones across suture lines. The maxilla is involved
more often than the mandible. Most patients present in their second to
third decades of life with a painless expanding bone mass. The mass does
not involve the overlying bone cortex and growth halts when the patient
reaches skeletal maturity. The radiographic appearance can be variable, but
the majority of patients present with a poorly defined lesion that is radiolu-
cent when small but becomes radio-opaque as it enlarges, often described as
having a ground glass appearance. The lesion blends imperceptibly with the
surrounding bone.
Microscopic examination demonstrates a bland fibrovascular stroma
with numerous irregular trabeculae of woven bone merging to form complex
shapes described as resembling Chinese letters. The bone trabeculae are not
rimmed with osteoblasts and lamellar bone is rarely identified (e-Fig. 4.23).
Repair of the deformity is usually attempted after the cessation of growth
with skeletal maturity. Radiation is contraindicated because of the elevated
risk of radiation-induced sarcomas.
2. Juvenile ossifying fibroma (JOF), also known as "juvenile active and juvenile
aggressive ossifying fibroma," is used in the literature to describe two dis-
tinct clinicopathologic entities: trabecular ]OF (Tr]OF) and psammomatoid
(Ps]OF).
a. Trabecular juvenile ossifying fibroma (TrJOF}. The great majority of the
patients are children and adolescents, with an equal gender distribu-
tion. Clinically, the lesions are characterized by progressive and some-
times rapid expansion. The maxilla is more commonly affected than the
mandible. Radiographically, the tumor is expansive and may be fairly
well demarcated, with cortical thinning and perforation. Depending on
the amount of calcification, the lesion may show varying degrees of radio-
density.
Microscopically, TrJOF is composed of a cell-rich fibrous stroma con-
taining bundles of cellular osteoid and bone trabeculae, without osteoblas-
tic rimming. Aggregates of multinucleated giant cells are invariably present
in the stroma (e-Fig. 4.24). Cystic degeneration and aneurysmal bone cyst
formation may occur. The clinical course of TrJOF following conserva-
tive treatment is characterized by recurrence; eventual complete cure can
be achieved by reexcision without resorting to radical surgery. Malignant
transformation has not been reported.
b. Psammomatoid juvenile ossifying fibroma (PsJOF}. Unlike TrJOF, PsJOF is
a lesion that affects predominantly the extragnathic skull bones, particu-
larly the periorbital bones. Occasional cases are encountered in the jaws,
particularly the mandible. Affected patients tend to be older than those
who have TrJOF. There is no sex predilection.
On radiographic examination, the tumor appears expansive with well-

defined borders that may be corticated. Sclerotic changes within the lesion
may impart a ground glass appearance. The tumors vary in size from 2
to 8 em. Cystic changes are not uncommon and present as areas of low
density in the CT scans.
Microscopically, the tumor is noteworthy for multiple, round, uni-
form, small ossicles that are basophilic and resemble psammoma bodies.
The psammomatoid structures are embedded in relatively cellular stroma
composed of stellate and spindle-shaped cells (e-Fig. 4.25). Aneurysmal
bone cyst formation is not uncommon. Surgical excision is the treatment
of choice; multiple recurrences are not unusual. No malignant changes are
observed.
3. CementD-osseous dysplasia of the jaws is a nonneoplastic, presumably dys-
plastic, fibro-osseous lesion of the tooth-bearing areas of the jaws. Two types
are recognized: periapical cementa-osseous dysplasia and florid cementa-
osseous dysplasia.
a. Periapical cemento-osseous dysplasia (PCOD), also known as "periapical
cementoma," is a relatively common condition, particularly in middle-
aged black female patients. The anterior mandibular teeth are typically
the site of this lesion. The condition is nonexpansive, asymptomatic, and
is typically identified in routine dental radiographs as "periapical radi-
olucencies," which becomes progressively mineralized in older lesions.
Microscopic examination shows a poorly demarcated lesion consisting of
fibrovascular stroma with trabeculae of bone and smooth globular masses
of cementum-like material (e-Fig. 4.26). The bone and cementum may
merge with adjacent bone but will not involve adjacent teeth. No treat-
ment is required. It is of importance to distinguish PCOD from periapical
inflammatory disease.
b. Florid cemento-osseous dysplasia (FCODJ is uncommon. It usually presents
in middle-aged or older black women, is usually asymptomatic, and may
be incidentally discovered on routine radiographic examination. Pain is
rarely manifested. Radiographically, FCOD is characterized by extensive
sclerotic areas often involving the posterior quadrants of the mandible and
the maxilla bilaterally in the tooth-bearing areas, usually symmetrically
(e-Fig. 4.27). Microscopically, FCOD and PCOD are analogous. However,
large sclerotic masses are hypocellular, extremely dense, and have small
marrow spaces and are more likely to form in FCOD. It is of importance
to recognize the clinical radiographic features of FCOD so that the patient
is not subject to surgical intervention. In fact, surgery is contraindicated
because it may result in local infection, pain, and a complicated clinical
course.
B. Giant cell lesions of the jaws. Giant cell lesions of the jaws are heterogeneous
clinical entities that share similar microscopic features.
1. Central giant cell granuloma (CGCG), or intraosseous granuloma of the jaws, is
also known as "giant cell reparative granuloma." It is a localized osteolytic
lesion of the mandible (66% of cases) and the maxilla (33% of cases). The
majority of patients are younger than 30 years, and women outnumber men
by a ratio of 2:1.
Giant cell granulomas are typically nonaggressive and present on routine
dental radiographs as a small radiolucent expansive masses that do not erode
into the cortex. The majority of CGCGs occur in the anterior mandible, com-
monly crossing the midline. Microscopically, the lesions are unencapsulated
and are composed of focal or evenly dispersed aggregates of multinucleated
osteoclast-like giant cells in a richly vascular stroma, with little collagen
deposition (e-Fig. 4.28). Two types of mononuclear stromal cells are iden-
tified: spindle-shaped fibroblastic cells and polygonal macrophage-like cells.
Areas of ossification and hemosiderin granules may be present.
A rare aggressive variant characterized by pain, rapid growth, and corti-
cal perforation with a marked tendency for recurrence may be an example of
"true" giant cell tumor of bone. The lesion may show increased mitotic activ-
ity and evenly distributed larger giant cells that have an increased number of
nuclei.
CGCG of the jaws is usually treated by curettage. Recurrent lesions often
respond to further conservative surgery. A number of alternative nonsurgical
approaches have been used in recent years, including intralesional corticos-
teroids injections, subcutaneous calcitonin injections, and interferon alpha
therapy.
2. Brown tumor of hyperparathyroidism is an osseous lesion that develops in bones
affected by primary or secondary hyperparathyroidism. It is currently less
frequently encountered since the diagnosis of hyperparathyroidism is now
often made on the basis of elevated serum calcium levels in asymptomatic
adults.
The lesions may be solitary or multifocal, and the mandible is a common
site of involvement. Radiographically, brown tumors are well-defined lytic
lesions that are microscopically identical to CGCG. Treatment is aimed at
correction of the hyperparathyroid state; complete resolution usually occurs
within 6 months after removal of a parathyroid adenoma.
3. Cherubism is a rare dominant genetic disease with complete male penetrance
and 50% to 70% penetrance in women. It typically presents as painless
bilateral symmetric jaw expansion in children aged 1 to 5 years, which
slowly increases in size until puberty; at puberty, the lesion undergoes vari-
able regression. The lesions may be unilocular or multilocular, with a "soap
bubble" appearance on radiographic examination.
Microscopically, the lesions are essentially similar to CGCG. However,
the giant cells in cherubism tend to be less numerous and placed in a less cellu-
lar stroma. The pathologic process in cherubism is self-limited and treatment
is dictated by cosmetic and functional needs. Curettage and contouring of
bone are the treatments of choice.
SUGGESTED READINGS
Delair D, Bejarano P, Peleg M, et al. Ameloblastic carcinosarcoma of the mandible arising in
ameloblastic fibroma: a case report and review of literature. Oral Surg Oral Med Oral Pathol
Oral Radio/ Endod. 2007;103:516-520.
El-Mofty SK. Bone lesions. In: Gnepp DR, ed. Diagnostic Surgical Pathology of the Head and
Neck. 2nd ed. Philadelphia: WB Saunders Co; 2009:729-784.
El-Mofty SK. Cemento-ossifying fibroma and benign cementoblastoma. In El-Mofty SK Guest
editor. Semin Diag Pathol. 1999;16:302-307.
El-Mofty SK. Psammomatoid and trabecular juvenile ossifying fibroma of the craniofacial skeleton:
two distinct clinicopathologic entities. Oral Surg Oral Med Oral Pathol Oral Radio/ Endod.
2002;93:269-304.
El-Mofty SK, Kyriakos M. Soft tissue and bone lesions. In: Gnepp DR, ed. Diagnostic Surgical
Pathology of the Head and Neck. Philadelphia: WB Saunders Co; 2001:505-604.
Nevill BW, Damm DD, Allen CM, et al, eds. Odontogenic cysts and tumors. Oral and Maxillofacial
Pathology. 3rd ed. Philadelphia: WB Saunders Co; 2009:589-642.
Philipsen HP, Reichart PA, Slootweg PJ, et al. Odontogenic tumors. In: Barnes L, Eveson ]W,
Reichart P, et al, eds. Pathology and Genetics Head and Neck Tumors. Lyon: IARC Press;
2005.
Slootweg PJ, El-Mofty SK. Ossifying fibroma. In: Barnes L, Eveson ]W, Reichart P, et al, eds.
Pathology and Genetics Head and Neck Tumors. Lyon: IARC Press; 2005.
The Eye
George J. Harocopos
I. DISEASES OF THE CONJUNCTIVA

A. Degenerative. Two benign degenerative lesions on the conjunctiva are the
pinguecula and the pterygium, which are manifestations of chronic actinic
damage to the interpalpebral bulbar conjunctiva. The pinguecula is confined
to the conjunctiva, appearing clinically as a yellowish nodule, whereas the
pterygium extends onto the peripheral cornea, appearing clinically as a vascu-
lar, wing-shaped lesion. Histologically, a pinguecula shows elastotic (actinic)
degeneration and may show variable degrees of chronic inflammation (e-Fig.
5.1).* The pterygium may show these same findings, but the most prominent
feature is congested vessels (e-Fig. 5.2).
B. Inflammatory. A variety of infectious and noninfectious conditions may cause
conjunctivitis.
1. Sarcoidosis often affects the conjunctiva, manifesting clinically as small, tan
nodules in the inferior forniceal conjunctiva, often in non-injected, asymp-
tomatic eyes. Sarcoidosis may also cause symptomatic inflammation in all
parts of the eye, including conjunctivitis, uveitis, retinal phlebitis, optic neu-
ritis, and so on. Conjunctival biopsy may provide the most expedient way
of diagnosing this systemic disease, even in cases where there are no visible
nodules clinically, though the diagnostic yield is highest when an obvious
nodule is present.
Histology shows noncaseating granulomatous tubercles in the stroma,
with a variable (but usually minimal) cuff of lymphocytes and plasma cells
(e-Fig. 5.3).
2. Ocular cicatricial pemphigoid (OCP} is a form of cicatrizing conjunctivitis that
typically also involves other mucous membranes and sometimes involves
the skin. When conjunctival biopsy is performed to establish the diagnosis,
half the specimen should be submitted in formalin for routine histology,
and half in Michel's medium or saline for immunofluorescence studies.
Histology shows epithelial bullae (or blebs) and a subepithelial band of
chronic inflammation composed predominantly of plasma cells (e-Fig. 5.4).
Immunofluorescence demonstrates lgG, lgA, and/or IgM immunoglobulins,
and complement (C3) positivity in the epithelial basement membrane zone.
The sensitivity of immunofluorescence may be as low as 50%, and accord-
ingly, a negative result does not rule out OCP.
C. Neoplasms of the conjunctiva fall mostly into one of the three categories: squa-
mous (surface epithelium), melanocytic, or lymphoid.
1. Neoplasms arising from the surface epithelium range from benign papil-
lomas (e-Fig. 5.5) to ocular surface squamous neoplasia (OSSN) which is
further subdivided into conjunctival intraepithelial neoplasia (CIN) (e-Fig.
5.6) versus invasive squamous cell carcinoma (e-Fig. 5.7). Histologic sec-
tions of a papilloma show finger-like projections of hyperplastic epithelium
draped over fibrovascular cores. The epithelium may exhibit loss of goblet
cells and surface keratinization if the lesion was exposed (i.e., not covered
adequately by the tear film due to its size).
63
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Chapter 5 • The Eye I 67
3. Lymphocytic lesions of the conjunctiva include lymphoid hyperplasia and

lymphoma, either of which may be unilateral or bilateral. Lymphomas can
range from primary localized lesions (even if bilateral), to lesions asso-
ciated with systemic disease. Most conjunctivaVorbital lymphomas are
low-grade B-cell lymphomas, with the single most common type being
extranodal marginal zone lymphoma (e-Fig. 5.11), which is generally local-
ized to the conjunctiva. Histologically, marginal zone lymphoma shows a
sheet of lymphocytes infiltrating the subepithelial region of the substantia
propria (stroma) without well-defined follicles; scattered lymphocytes may
extend into the epithelium. Immunohistochemistry forB- and T-cell markers
(e-Fig. 5.12) as well as in situ hybridization (ISH) for kappa and lambda
light chains (e-Fig. 5.13) are very helpful diagnostically. Other techniques
for establishing clonality such as IgH gene rearrangement testing by PCR
and flow cytometry are also useful, particularly in cases where ISH proves
insufficient. Fluorescence in situ hybridization (FISH) may also be used if
needed to test for specific genetic translocations.
Follicular and mantle cell lymphomas are also seen in the conjunctiva.
More rarely, diffuse large B-cell, Burkitt, Hodgkin, plasmacytoma, or T-cell
lymphoma may also occur. Lower-grade lymphomas are more often local-
ized to the conjunctiva, whereas higher-grade lymphomas are more likely
to be associated with systemic disease. If the lymphoma is localized to the
conjunctiva, the treatment is generally orbital radiation. Another treatment
option is subconjunctival injections of IFN or intravenous rituximab. In
contrast, if systemic lymphoma is present, then it is treated accordingly with
chemotherapy; if systemic remission is achieved, the conjunctivallesion(s)
will likewise resolve. The staging scheme for conjunctivaVorbitallymphoma
is given in Table 5.3.
4. Other neoplasms. Oncocytoma, also known as apocrine cystadenoma or
oxophilic cystadenoma (e-Fig. 5.14), is a benign lesion that arises most
commonly in the caruncle of elderly females. Histologically, it is an adenoma
composed of apocrine or accessory lacrimal gland epithelial cells which
exhibit distinctive eosinophilic cytoplasm and surround gland-like spaces.
Any neoplasm seen in the orbit may also occasionally arise in the con-
junctiva, including neural, vascular, fibrous, and muscular tumors. Addi-
tionally, metastatic lesions to the conjunctiva rarely occur.
II. DISEASES OF THE CORNEA. The host tissue from a corneal transplant procedure
is traditionally referred to as a corneal "button"; the surgical procedure itself is
a keratoplasty (KP). Most often, full-thickness cornea is removed, (i.e., penetrat-
ing keratoplasty or PKP). More recently, however, surgical techniques have been
developed such that, for certain disorders, only the diseased layer(s) of the cornea
need be transplanted, for example, deep anterior lamellar keratoplasty (DALK),
in which only the epithelium and stroma are removed, versus Descemet's strip-
ping endothelial keratoplasty (DSEK), in which only Descemet's membrane and
endothelium are removed. In the gross room, corneal buttons are bisected, and
each half is embedded with the cut side down. The most common reasons for ker-
atoplasty to be performed are Fuchs endothelial dystrophy, pseudophakidaphakic
bullous keratopathy, keratoconus, infectious keratitis (especially herpetic), and
graft failure.
A. Fuchs endothelial dystrophy (e-Fig. 5.15) exhibits an autosomal dominant inher-
itance pattern or may be sporadic, generally becoming symptomatic in middle-
aged to older individuals. The corneal endothelial cells diminish in number sig-
nificantly faster than is normal for aging, ultimately leading to chronic edema
of the cornea. If conservative therapy is unsuccessful, then keratoplasty (either
PKP or DSEK) is necessary. Histologically, Fuchs dystrophy demonstrates loss
of the endothelial cells and the presence of anvil-shaped excrescences, known
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an anterior segment procedure, but occasionally 50% or more of the endothe-

lial cells may be lost if the surgery is highly complex/traumatic. The endothe-
lium may then decompensate, either immediately if the residual endothelial
cells are too few, or sometime later, after the endothelial cell population
declines further with age. Keratoplasty (either PKP or DSEK) may ultimately
be required. Although cataract surgery is the most common intraocular pro-
cedure performed, bullous keratopathy may similarly occur following other
intraocular procedures, for example, multiple glaucoma surgeries or retinal
surgeries. Histologically, endothelial cell loss is seen, but without guttae of
Descemet's membrane; epithelial bullae are present.
C. Keratoconus is a condition generally presenting in teenagers and young adults
in which the cornea is more ectatic (i.e., cone shaped) than normal, causing
myopia and astigmatism. Both genetic and environmentaVacquired factors may
be involved, since keratoconus is associated with both hereditary connective
tissue disorders such as Marfan syndrome, as well as with excessive eye rubbing
such as in chronic atopic conjunctivitis or chronic blepharitis associated with
Down syndrome. In the early phases of the disease, the patient may be managed
with spectacle or contact lens correction, but if the condition progresses to the
extent that a corneal stromal scar forms at the apex of the cone (by which time
the patient is often middle aged), then keratoplasty (either PKP or DALK) is
required for visual rehabilitation. The corneal button shows central thinning;
breaks in Bowman's layer; and often loss of Bowman's layer with anterior
stromal fibrosis at the apex of the cone correlating with the apical scar seen
clinically (e-Fig. 5.17).
D. Corneal buttons with ulcerative keratitis may or may not reveal the offend-
ing microorganism when special stains are performed (e.g., Gram for bacte-
ria; Gomori's Methenamine Silver [GMS] and periodic acid-Schiff [PAS] for
fungi; GMS and PAS for Acanthamoeba). Lack of microorganisms on histol-
ogy despite a history of positive cultures is generally attributable to antecedent
anti-microbial therapy, as an attempt is generally made to sterilize the ulcer
prior to keratoplasty. This scenario is frequently encountered with bacterial
corneal ulcers, whereas fungi (e-Fig. 5.18) and amoeba! cysts (e-Fig. 5.19) are
often more difficult to eradicate with medical treatment and are therefore gen-
erally still present on histology. Herpes simplex keratitis has a characteristic
histopathologic appearance including patchy loss of Bowman's layer, stromal
fibrosis and vascularization, interstitial keratitis (consisting of plasma cells and
lymphocytes in the corneal stroma), and often granulomatous inflammation
near Descemet's membrane (e-Fig. 5.20).
E. When a PKP fails (e.g., due to immunologic rejection, infectious ulcerative ker-
atitis, or most commonly, simply gradual loss of endothelial cells over time), a
regraft may be necessary. On histology, the button may be identified as a regraft
by peripheral discontinuities in Bowman's layer and peripheral stromal scars
(e-Fig. 5.21) that represent the entry sites of sutures and sometimes the graft-
host interface (i.e., if the surgeon chose to make the regraft of a slightly larger
diameter than the prior graft). Residual suture material may be present, or may
be absent if all the sutures were removed at some point after the initial pro-
cedure. Chronic inflammatory cells are often present along the suture tracks.
The most essential histologic feature of graft failure is generally endothelial
cell loss. In many cases there is also a fibrous retrocorneal membrane.
F. Other rare entities for which a PKP is performed include hereditary stro-
mal dystrophic diseases, for example, macular dystrophy (due to acid
mucopolysaccharide accumulation), granular dystrophy (due to hyaline depo-
sition), lattice dystrophy (due to amyloid), and combined granular-lattice
(Avellino) dystrophy, in which both hyaline and amyloid deposits are seen
(e-Fig. 5.22).
Ill. VASCULAR, INFLAMMATORY, AND INFECTIOUS DISEASES AND TRAUMA. Eyes are often
removed when they become "blind and painful." The reasons for an eye becoming
blind and painful are numerous, but often the final common pathway involves the
development of secondary angle-closure glaucoma and/or chronic retinal detach-
ment, and optic nerve atrophy. The antecedent event can range from previous
accidental trauma or previous surgical trauma to specific disease entities such as
retinal vascular disease (including diabetes, or central retinal artery or vein occlu-
sion), intraocular infection (endophthalmitis), chronic inflammatory disease, or
primary retinal detachment. Surgical methods of removing an eye include enu-
cleation, in which the entire eye is removed intact, or evisceration, in which only
the intraocular contents (lens, retina, uvea) and possibly cornea are removed with
retention of the sclera in the eye socket to house the orbital implant. Exentera-
tion, in which the eye along with surrounding orbital soft tissues are removed, is
sometimes required to achieve complete excision of an eyelid/orbital tumor (see
later).
A. Secondary glaucoma. The key histopathologic finding in most cases of sec-
ondary glaucoma is an anterior chamber angle closed by peripheral anterior
synechiae. The term "closed angle" means that the anatomical angle normally
formed by the cornea and iris, and occupied by the trabecular meshwork (the
main outflow channel for aqueous humor), is occluded by the peripheral iris. If
this occlusion is chronic, then permanent adhesions form between the periph-
eral iris and the trabecular meshwork, that is, peripheral anterior synechiae
(e-Fig. 5.23). These adhesions may be induced by chronic inflammation, such
as in various forms of chronic uveitis, or by the abnormal proliferation of cap-
illaries (neovascular glaucoma), as seen in proliferative diabetic retinopathy or
retinal vascular occlusion (see later). Occasionally, in contrast to the closed-
angle appearance, post-contusion angle recession may be seen in cases of blunt
trauma. The retina in glaucoma exhibits atrophy of the nerve fiber layer and
loss of ganglion cells; the optic nerve shows cupping (e-Fig. 5.24) and atrophy.
B. Chronic retinal detachment. Retinal detachment may be idiopathic, trauma
induced, related to inflammation or infection, or secondary to proliferative
retinopathy of various etiologies (diabetic, post-vascular occlusion, and so on).
Often an attempt is made to repair the detachment surgically, but severe cases
in which surgery fails or the detachment repeatedly recurs may ultimately be
treated by enucleation (e-Fig. 5.25).
C. Retinal vascular diseases. Eyes with severe retinal vascular disease (most com-
monly diabetic retinopathy, or central retinal arterial or venous occlusion)
often exhibit both secondary angle closure due to neovascularization of the
iris (e-Fig. 5.26) and angle (neovascular glaucoma), as well as chronic trac-
tional retinal detachment due to proliferative retinopathy. In cases of diabetic
retinopathy, microscopic examination of the retina reveals the key features of
diabetes: lipoproteinaceous exudates, intraretinal hemorrhages, and neovas-
cularization of the inner surface of the retina (proliferative diabetic retinopa-
thy) (e-Fig. 5.27), often with tractional retinal detachment and hemorrhage
into the vitreous. In cases of old central retinal vein or artery occlusion, the
retina exhibits atrophy and cystoid degeneration of the inner two-thirds of the
retina, lipoproteinaceous exudates, and in the case of venous occlusion, often
a persistence of hemorrhage. There may also be a neovascular epiretinal mem-
brane with tractional retinal detachment and vitreous hemorrhage. As with
neovascular glaucoma of any etiology, eyes with severe diabetic retinopathy or
central retinal vascular occlusion also exhibit atrophy and possibly cupping of
the optic nerve.
D. Intraocular inflammation and infection. Endophthalmitis most commonly occurs
via an exogenous source (e.g., following a surgical procedure, trauma, or per-
forated corneal ulcer), or may arise from an endogenous source (e.g., secondary
to bacteremia or fungemia). Endophthalmitis is generally treated with intrav-

itreal antibiotics and/or antifungals, and possibly vitrectomy surgery to clear
the abscess. However, in severe cases treatment may not be successful, and
evisceration or enucleation may ultimately be performed. On histology (e-Fig.
5.28), such cases demonstrate intraocular abscess formation and retinal detach-
ment/destruction; depending on the degree of chronicity, optic nerve atrophy
rnay also be seen.
Other severe inflammatory or infectious diseases that can lead to a blind
and painful eye include diffuse uveitis of unknown etiology; juvenile rheuma-
toid arthritis-associated uveitis or uveitis associated with other collagen vascu-
lar/autoimmune diseases; diffuse granulomatous uveitis secondary to sympa-
thetic ophthalmia (e-Fig. 5.29), sarcoidosis, or toxoplasmosis; and necrotizing
retinitis (as seen in cytomegalovirus [CMV] retinitis or herpetic retinitis).
E. Trauma. Many eyes are enucleated because of severe trauma. If there is no
attempt at repair by the ophthalmologist because the rupture is too exten-
sive, the pathology specimen usually consists of a ruptured globe with massive
intraocular hemorrhage and total retinal detachment (e-Fig. 5.30). Often there
is loss of intraocular contents such as the lens and a portion of the uvea, and
possibly also the retina. If an attempt has been made by the surgeon to sal-
vage the ruptured globe, but the eye later has to be enucleated because it has
become blind and painful, the histopathologic findings vary depending on the
degree of chronicity; in cases of recent rupture, the findings may be the same
as described above (i.e., massive intraocular hemorrhage, total retinal detach-
ment, and loss of intraocular contents) or may also include additional findings
such as endophthalmitis. In cases with a more distant history of rupture, there
may be total retinal detachment with gliosis and loss of intraocular contents,
intraocular granulation tissue or fibroconnective tissue emanating from the
rupture site, and often osseous metaplasia of the retinal pigment epithelium
(see later). Angle closure or post-contusion angle recession may also be present
depending on the location of the rupture and the nature of the traumatic forces.
Optic nerve atrophy is generally also seen.
F. Phthisis bulbi. When an eye has undergone a previous insult such as severe
trauma, complications of surgery, proliferative retinopathy, or chronic inflam-
mation, and has a chronic total retinal detachment, a gradual degenerative
process eventually ensues in which the eye undergoes shrinkage, that is, phthi-
sis (from the Greek verb "to wither"). Clinically, the eye has hypotony (very
low intraocular pressure) and is visibly shrunken. Histologically, the retina is
totally detached, with marked diffuse atrophy and architectural distortion of
all layers of the eye including such severe retinal gliosis as to render the retina
barely recognizable. Often the retinal pigment epithelium undergoes osseous
metaplasia, and bone may occupy a significant portion of the intraocular cavity
(e-Fig. 5.31). Choroidal effusion may be seen. The optic nerve is atrophic.
IV. INTRAOCULAR NEOPLASMS
A. Gross room processing of enucleated globes follows a standard protocol to
generate the pupil-optic nerve (p.o.) section, as shown in Figure 5.1. Larger
cassettes are required for processing. The figure shows the standard transverse
plane that is often used for cutting the section, but the globe may be opened
along any plane that best captures the particular area of interest. For eyes
enucleated due to intraocular tumors, especially primary uveal melanoma, the
tumor may be localized by transillumination prior to cutting the eye. Transil-
lumination is achieved by shining onto the cornea a very bright light source
that is about 1 em or less in diameter. The normal uvea will not block the
transmission of light, and hence the sclera will glow. However, a uveal tumor
will block light transmission, thereby casting a shadow over the corresponding
portion of sclera.
Left Eye Right Eye

Posterior View
Figure 5.1 The globe is identified as to right eye or left eye using anatomic landmarks (upper
panel}. Two cuts (traditionally refell'ed to as the p.o. sections) are then made in a transverse plane,
passing in relation to the pupil and optic nerve as shown (lower panel}. The superior cap (also
known as the superior calotre) and inferior cap are usually not processed unless abnormalities are
seen or the eye has an intraocular tumor.
B. The most common primary intraocular neoplasm is melanoma of the uvea

(choroid, ciliary body, or less commonly, iris). Melanomas usually occur in
adults of light complexion. Melanomas of the iris have a better prognosis than
do melanomas of the ciliary body and/or choroid. Although some intraocular
melanomas can be treated by excision or brachytherapy, some patients do
not seek medical attention until the melanoma has become so large that the
only course of management is enucleation of the globe. If the diagnosis is in
doubt, fine-needle aspiration biopsy (FNAB) may be performed to confirm
the diagnosis (e-Fig. 5.32). In cases of medium-sized melanomas treated with
radioactive plaque brachytherapy, some ocular oncologists routinely perform
FNAB at the time of plaque placement so as to obtain a sample for gene
expression profiling, as this provides useful prognostic information.
Large uveal melanomas are generally treated with enucleation. Primary
uveal melanoma usually arises from the ciliary body or choroid as an ellipse or
almond-shaped mass (e-Figs. 5.33 and 5.34) which eventually breaks through
Bruch's membrane and becomes mushroom shaped (e-Fig. 5.35). Microscop-
ically, intraocular melanomas are composed of cells ranging from spindle to
>15.0 4 4 4
E' 12.1-15.0 3 3 I 4 4
-
E
9.1-12.0
6.1-9.0 2
3
2
3
2
3
2 I 3
3 3
3
4
4
3.1-6.0 1 1 1 I 2 2 I 3 4
:S;3.0 1 1 1 1 I 2 2 4
:S;3.0 3.1-6.0 6.1-9.0 9.1-12.0 12.1-15.0 15.1-18.0 >18.0
Largest basal diameter (mm)
Figure 5.2 Size categories for ciliary body and choroidal melanoma based on thickness and
diameter.
epithelioid, with the latter having a worse prognosis (e-Figs. 5.36 and 5.37).
Other histopathologic features that predict prognosis are size (e-Fig. 5.38),
extrascleral extension (e-Fig. 5.38), and location (those arising from the iris
have a better prognosis, whereas those involving the ciliary body [e-Fig. 5.39]
or peripapillary region have a poorer prognosis). Other factors associated with
a worse prognosis include necrosis and mitotic figures. The staging scheme for
melanoma of the uvea is described in Table 5.4 and Figure 5.2.
C. Retinoblastoma is the most common ocular neoplasm in children. The usual
age for clinical presentation in nonfamilial cases is 1 year, whereas familial
cases are generally screened within the first few weeks of life and therefore
present earlier. The usual clinical appearance is leukocoria (white pupil). In
nonfamilial cases, the tumor is generally large and unilateral and accordingly
treated with enucleation. In familial cases, tumors are generally seen bilaterally.
The tumor may be much larger in one eye than the other, in which case the
eye with the large tumor is enucleated, whereas the eye with smaller tumor(s)
may be salvaged via laser or cryotherapy applied to the tumor foci, possibly
with antecedent adjuvant systemic chemotherapy. A few centers utilize intra-
arterial chemotherapy (delivered to the ophthalmic artery) to treat unilateral or
bilateral tumors, thereby potentially salvaging eyes that would otherwise need
to be enucleated. Some centers utilize periocular injection of chemotherapy.
Histopathologically, retinoblastoma arises from the retina (e-Fig. 5.40) and
is composed of small blue cells intermingled with anastomosing pools of pink
necrosis giving rise to the traditional description of "islands of blue tumor
in a sea of pink necrosis" (e-Fig. 5.41). Scattered flecks of purple calcium
are often seen. Cytologically, the tumor is composed of cells with hyperchro-
matic, oval-shaped nuclei with scant cytoplasm; the nuclei are densely packed
together and thus show nuclear molding. There are many apoptotic bodies and
numerous mitotic figures. More differentiated retinoblastomas often exhibit
Flexner-Wintersteiner rosettes (e-Fig. 5.42). Extension into the optic nerve,
and especially presence of tumor at the surgical margin of the nerve, predicts
a poor prognosis (e-Fig. 5.43). The other factor predictive of poor progno-
sis is massive invasion of the choroid. If optic nerve invasion past the lamina
cribrosa or massive choroidal invasion is seen on histology, then the patient is
treated with adjuvant systemic chemotherapy. (Some practitioners have a lower
threshold for adjuvant chemotherapy than others; the optimal algorithm for
post-enucleation chemotherapy has not been established). The staging scheme
for retinoblastoma is given in Table 5.5.
V. DISEASES OF THE LACRIMAL GLAND AND LACRIMAL SAC
A. Lacrimal gland lesions are similar to lesions of the parotid gland (see Chap. 6).
Infiltrative lesions include dacryoadenitis, sarcoidosis, lymphoid hyperplasia,
and lymphoma. Intrinsic lesions of the lacrimal gland include pleomorphic
,.. I SECTION I. HEAD AHD NECK
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SUGGESTED READINGS
Eagle RC. Eye Pathology, an Atlas and Text. 2nd ed. Philadelphia, PA: Lippincott Williams &
Wilkins; 2011.
Font RL, Croxatto JO, Rao NA. In: AFIP Atlas of Tumor Pathology, Series 4. Vol5. Washington,
DC: ARP Press; 2006.
Smith ME, Kincaid MC, West CE. In: Krachmer JH, ed. The Requisites in Ophthalmology.
St. Louis, MO: Mosby; 2002; chapters 3-9,23.
Yanoff M, Sassani JW. Ocular Pathology. 6th ed. Philadelphia, PA: Elsevier/Mosby; 2009.
Salivary Glands
Joshua I. Warrick, Elise L. Krejci,
and James S. Lewis Jr.
I. NORMAL ANATOMY
A. Macroscopic/gross. The salivary glands are exocrine organs that secrete compo-
nents of saliva to both break down carbohydrates and lubricate the passage of
food. There are three major paired salivary glands: the parotid, the submandibu-
lar, and the sublingual. There are also numerous minor salivary glands located
in the submucosa of the entire upper aerodigestive tract (UADT), from the lips
and nasal cavity to the major bronchi.
The parotid glands are the largest major salivary glands and are located
between the ramus of the mandible and the mastoid process. Each gland is
composed of a superficial lobe and a smaller deep lobe, and the facial nerve is
intimately associated with both lobes. Each gland normally contains approx-
imately 20 intraglandular lymph nodes. The secretions of the parotid gland
empty via Stensen's duct into the buccal mucosa of the oral cavity near the
second maxillary molar.
The submandibular glands (also referred to as the submaxillary glands) are
located just medial to the body of the mandible, and are smaller than the parotid
glands. Their secretions drain through Wharton's duct into the floor of the
mouth.
The sublingual glands are the smallest of the major salivary glands and
are located in the floor of the mouth between the genioglossus muscle and the
mandible. Their secretions drain into the floor of the mouth directly via small
ducts or through the larger (Bartholin's) duct which drains into Wharton's duct.
B. Microscopic. The major salivary glands are enclosed by a connective tissue cap-
sule and divided into lobules composed of ducts and acini, whereas the minor
salivary glands are unencapsulated. The acini are composed of mucinous and/or
serous epithelial cells surrounded by a layer of myoepithelial cells which con-
tract to aid in the movement of glandular secretions. Myoepithelial cells are
inconspicuous in normal salivary gland sections.
A single acinus may be composed of a mixture of serous and mucous cells.
Serous cells are pyramidal in shape and contain basophilic periodic acid-Schiff
(PAS)-positive granules within their cytoplasm. Mucous cells are more rounded,
have basally oriented nuclei, and contain abundant clear mucin. The parotid
gland is almost exclusively serous, the sublingual gland almost exclusively muci-
nous, and the submandibular gland is mixed serous and mucinous (e-Figs. 6.1
to 6.3 ). * The acini secrete fluid into the intercalated ducts, which are lined by a
low simple cuboidal epithelium. Intercalated ducts are also the source of reserve
cells that can repopulate the acinar system. The intercalated ducts join to form
striated ducts, which merge to form interlobular ducts, which ultimately empty
into the large named ducts.
A. Biopsies of many salivary gland lesions are taken prior to surgery to characterize
the lesion and direct management.
1. Fine needle aspiration (FNA) is the procedure of choice for initial character-
ization of salivary gland lesions for many reasons (see cytology section that
follows).
81
2. Core needle and incisional biopsies of the parotid gland can damage the facial
nerve branches so are rarely performed.
B. Resections. Some salivary gland masses are removed in their entirety without a
previous tissue diagnosis. Superficial parotidectomy remains the initial proce-
dure of choice for benign parotid gland tumors. Submandibular gland resections
are usually performed without taking any significant periglandular soft tissue,
although more tissue may be included with the specimen for submandibular or
parotid tumors that are known or suspected to be malignant. The rare sublingual
gland tumors necessitate a resection of the floor of mouth that is qualitatively
similar to those performed for mucosal-based squamous carcinomas of the same
area.
In general, salivary gland specimens are small enough to prosect the day
of receipt. The gland should be described, and its dimensions recorded. The
deep (covered by muscle and fascia) and superficial (covered by subcutaneous
fat) surfaces of the parotid gland can sometimes be discerned, and should be
differentially inked. The specimen should be serially sectioned, and any mass
or focal lesion described including dimensions, color, texture, and distance to
the margins. Four to five sections of the tumor, including representative areas of
the closest inked margins, should be taken. At least one section of the normal
surrounding gland should also be taken. For the parotid gland, the surrounding
gland and any periglandular soft tissue should be thoroughly searched for lymph
nodes, which should be separately submitted.
A. Inflammation and infection. Sialadenitis, or inflammation of the salivary glands,
can be divided into bacterial causes, viral causes, and autoimmune disease.
1. Bacterial sialadenitis is rare and is generally a result of obstruction by stones
(sialolithiasis). The causative organism is usually Staphylococcus aureus. fol-
lowed by Streptococcus viridans and gram-negative rods. Surgical specimens
from bacterial sialadenitis are almost never seen. However, sialadenitis may
lead to an abscess that requires surgical drainage. Gross specimens may show
a purulent exudate or a relatively intact gland. Microscopic sections can show
abscesses, cystic degeneration, and/or bacterial colonies.
2. Viral sialadenitis is most commonly caused by mumps (paramyxovirus), but
can also be associated with Epstein-Barr, coxsackie, influenza A, and parain-
fluenza viruses. Surgical specimens are rarely encountered. Grossly infected
glands are boggy and edematous, and chronic inflammation may be seen
microscopically.
3. Autoimmune sialadenitis is relatively common, and usually presents as
Sjogren's syndrome, characterized by facial swelling, dry mouth, and dry eyes
(termed Sicca syndrome) with inflammatory arthritis and elevated autoan-
tibodies. On physical examination, bilateral, symmetric enlargement of the
salivary and lacrimal glands is seen. Diagnosis may be facilitated by a minor
salivary gland biopsy, typically taken from the inner lip.
Grossly, the major glands usually have a discrete, tan nodularity. Micro-
scopically, in early disease, there is a lymphoplasmacytic septal inflamma-
tory infiltrate with little to no abnormality of the parenchyma. The so-called
"focus score," a nodular collection of >50 lymphocytes, is considered diag-
nostic of autoimmune sialadenitis in the appropriate clinical context. In larger
glands or in more severe disease, there is typically an extensive lymphoid
infiltrate with germinal centers (e-Fig. 6.4). The acini are often atrophic,
and interstitial fibrosis may be present. In late disease, acini may be com-
pletely absent leaving only isolated residual ducts, termed "epimyoepithelial
islands," with an associated dense intraepitheliallymphocytosis. In Sjogren's
syndrome, the lymphocytic infiltrate is polyclonal, in contrast to the mon-
oclonal lymphoid proliferation seen in mucosa-associated lymphoid tissue
(MALT) lymphoma.
Chapter 6 • Salivary Glands I 83
4. Chronic sialadenitis (or chronic sclerosing sialadenitis) is usually unilateral

and clinically can mimic a true salivary gland neoplasm. When it occurs in
the submandibular gland, it is known as a Kuttner tumor. Chronic sialadenitis
most commonly results from sialolithiasis with obstruction, although some
cases may be due to radiation therapy or duct strictures. It can have no struc-
tural or obvious etiology and recently most of these cases have been shown
to be an autoimmune-related lgG4-related sclerosing disease similar to that
occurring in the pancreas. Grossly, it is characterized by a very hard, fibrotic
gland with lobulation and septation (e-Fig. 6.5). Histologic examination of
the gland early in the disease process shows dilated ducts filled with secretions
with an associated lymphocyte and plasma cell-rich infiltrate, occasionally
with germinal center formation. As the disease progresses, fibrosis surrounds
the ducts and results in lobulation of the gland (e-Fig. 6.6) with acinar atro-
phy and broad fibrous bands. Surgical excision may be required, either to
definitively rule out a neoplasm or just for symptom relief.
5. Necrotizing sialometaplasia is a rare inflammatory/destructive lesion that
simulates malignancy and is thought to occur because of ischemic injury.
Although it can occur anywhere along the UADT, the vast majority of cases
occur in the hard palate. Men are slightly more commonly affected, and the
average patient age is 46 years. Clinically, the lesion consists of a sharply
defined and deep ulcer that develops rapidly (over a few days) and can per-
sist for months. Grossly, the lesion consists of loose tissue with a surface
ulcer and no distinguishing mass lesion. Microscopically, the most typical
feature is coagulative necrosis of the minor salivary gland lobules with a
prominent associated inflammatory response. There is extensive squamous
metaplasia of the ducts (an alarming feature), but the lesion retains its overall
lobular architecture (e-Fig. 6.7). The surface squamous mucosa may show
pseudoepitheliomatous hyperplasia. The lack of peripheral infiltration and
the retained lobular architecture are keys to recognizing the lesion as benign.
No specific therapy is indicated because the lesion is self-healing.
B. Cystic lesions
1. Acquired immune deficiency syndrome (AIDS}-related parotid cysts (ARPCs} are
benign lymphoepithelial cysts. Patients present with unilateral or bilateral,
painless, slowly enlarging parotid masses. They sometimes have associated
cervical lymphadenopathy and/or nasopharyngeal swelling. Grossly, these
lesions have multiple cystic spaces usually containing serous fluid. Micro-
scopically, ARPCs consist of cysts that most often have a thin, mature squa-
mous epithelial lining, although the lining is sometimes cuboidal or columnar
with goblet cells. Below the epithelium, the cyst wall has dense lymphoid tis-
sue with germinal centers. Occasionally, lesions within the gland can lack
cystic change and appear similar to epimyoepithelial islands.
Unlike autoimmune sialadenitis, the vast majority of human immunode-
ficiency virus (HIV) patients with ARPC have no autoimmune symptoms, no
Sicca syndrome from glandular dysfunction, and no serum autoantibodies.
The etiology of ARPC is thus unclear. Patients usually have bilateral disease
by radiologic examination, even if there are no symptoms and no clinical mass
in the contralateral gland. If the diagnosis is established by radiologic imaging
and cytology, surgery is not necessary for other than cosmetic reasons.
2. Benign lymphoepithelial cysts are unifocal lesions that occur in patients in
their fifth and sixth decades. They most commonly involve the parotid gland,
but are occasionally seen in the oral cavity. There is no association with
systemic disease. Grossly, they consist of well-circumscribed unilocular cysts
with contents ranging from serous to mucoid to caseous; keratinous debris
may be present. Microscopically, they consist of a cyst lined by thin, mature
squamous epithelium without papillary projections. In rare cases, the lining
is cuboidal or columnar with goblet cells. Below the epithelium, the cyst
wall has dense lymphoid tissue with germinal centers (e-Fig. 6.8). Excision
is curative, and the lesion does not recur.
3. Mucoceles are the most common non-neoplastic lesion of salivary gland tis-
sue and are defined as pooling of mucin in a cystic cavity. Two types are
recognized. In the retention type, the mucin is within a dilated duct. In the
extravasation type, the mucin accumulates in the soft tissue. The lower lip
is the most common site for mucoceles, followed by the tongue, the floor of
the mouth (where the lesion is termed a "ranula"), and the buccal mucosa.
The peak incidence of mucoceles is in the third decade. Grossly, a mucocele
presents as a cystic cavity in the connective tissue that is filled with glistening
fluid. Microscopically, the cyst wall may or may not have an epithelial lining,
depending on the type. Those of the retention type have a lining and usu-
ally no surrounding inflammation, whereas those of the extravasation type
consist of mucin with surrounding inflammatory cells and no lining. Some
late lesions consist only of a collection of foamy histiocytes containing mucin
(e-Fig. 6.9), which can be confirmed by PAS or mucicarmine stains. Surgical
excision is usually curative.
IV. NEOPLASMS. Neoplasms of the salivary gland can be roughly classified based on
the cell type(s) of the normal salivary gland toward which they differentiate: aci-
nar, myoepithelial, or ductal. However, most neoplasms have dual differentiation
because almost all show of those with ductal differentiation also have some myoep-
ithelial differentiation. Most benign neoplasms have a malignant counterpart. The
World Health Organization (WHO) classification of tumors of the salivary gland
is listed in Table 6.1.
A. Benign neoplasms
1. Pleomorphic adenoma (PA) is the most common neoplasm of the salivary
glands. Ninety percent of PAs occur in the parotid gland and PAs represent
60% of parotid gland neoplasms; the majority of the remaining cases occur
in the hard palate or submandibular gland. The tumor occurs over a wide age
range, but the peak incidence is in the fourth and fifth decades. The tumor
presents as a slow-growing, painless mass that usually is between 2 and 5 em
in diameter. On gross examination, these tumors appear well-circumscribed
but are not usually encapsulated. They are never encapsulated when occur-
ring in minor salivary glands. Sectioning reveals a rubbery, myxoid, tan-
white mass (e-Fig. 6.10). The microscopic appearance, as the name suggests,
is highly variable, but always shows an intimate admixture of epithelial and
mesenchymal elements. The epithelial component consists of ductal struc-
tures with an associated myoepithelial layer but also may contain collections
of myoepithelial cells that can range from spindled to clear, plasmacytoid,
or basaloid. The mesenchymal, or stromal, component is typically myxoid,
hyaline, or chondroid (e-Fig. 6.11). Although defined subcategories have no
clinical significance, pleomorphic adenomas have been divided into a myxoid
type (>80% mesenchymal-type tissue), cellular type (>80% epithelial-type
tissue), and mixed or classic type (generally an equal mix of components).
Treatment requires complete excision because PAs are likely to recur if
tumor is left behind or transected. Multinodular growth is uncommon in pri-
mary tumors but is frequent in recurrent disease, yielding a so-called "buck-
shot" pattern (e-Fig. 6.12). Therapy for recurrence is consequently more
difficult.
2. Warthin tumor (papillary cystadenoma lymphomatosum) is the second most
common benign salivary gland tumor and is found exclusively in the parotid
gland. It is the most common bilateral or multifocal salivary gland tumor,
usually presents in the sixth or seventh decade, and is associated with smok-
ing. Grossly, it presents as a soft, brown or yellow mass that is composed of
cysts that are classically filled with viscous brown fluid. Microscopically, the
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mitotic activity is low. The epithelial cells express cytokeratin and S-1 00, but
are negative for calponin.
The differential diagnosis for canalicular adenoma includes pleomorphic
adenoma and basal cell adenoma. However, the pattern of growth of canalic-
ular adenoma is virtually always distinctive enough to make the diagnosis.
Simple local excision is performed with an attempt to obtain dear margins.
Recurrence is rare.
5. Myoepitheliomas are benign tumors composed of myoepithelial cells. These
tumors occur with approximately equal frequency in the parotid gland and
minor salivary glands, particularly the hard and the soft palate, and present
as slowly growing, painless masses in adults. Men and women are affected
equally.
Grossly, these neoplasms are well-circumscribed and encapsulated with a
tan to white cut surface. Histologically, they are composed almost exclusively
of sheets and cords of myoepithelial cells, though ductal cells may comprise
a small percentage ( <10%) of individual tumors. In this regard, myoepithe-
liomas are sometimes considered to lie at one end of a biologic spectrum,
with pleomorphic adenoma in the middle, and basal cell adenoma at the
opposite end. The tumors can be classified into four major subtypes, based
on myoepithelial cell morphology: spindle cell, hyaline, plasmacytoid, and
clear cell. All subtypes commonly have a collagenous or myxoid stroma. The
spindle cell type shows an interlacing, fascicular pattern of growth (e-Fig.
6.19). Loosely cohesive myoepithelial cells with eosinophilic cytoplasm and
eccentric round nuclei are present in the plasmacytoid variant (e-Fig. 6.20).
The clear cell type is composed of sheets and aggregates of dear cells (e-Fig.
6.21). The tumor cells express typical myoepithelial markers such as S-100,
pan-cytokeratin, smooth muscle actin, and p63.
Simple excision is generally the treatment of choice unless malignant fea-
tures, such as infiltrative growth or perineural invasion, are encountered,
in which the diagnosis is myoepithelial carcinoma. Myoepitheliomas have a
slightly lower recurrence rate than pleomorphic adenomas.
B. Malignant neoplasms
1. Mucoepidermoid carcinoma is the most common salivary gland malignancy.
The major salivary glands account for more than half of all cases, with most
arising in the parotid gland. These tumors also arise from minor salivary
glands in the oral cavity, particularly in the hard palate, buccal mucosa,
lip, and retromolar trigone. They may also develop intraosseously in the
mandible and maxilla, but such tumors are considered odontogenic in ori-
gin and have a different clinical behavior. Mucoepidermoid carcinomas are
slightly more common in women, and the mean age of affected patients is
approximately 45 years. The tumor also occurs in children and is the most
common pediatric salivary gland carcinoma. Patients usually present with a
painless, slowly growing mass, but may also present with a blue-red superfi-
cial nodule along the oral mucosa mimicking a mucocele or vascular lesion.
Grossly, both solid and cystic components may be present, often with
mucinous material within the cysts. Microscopically, the hallmark of these
tumors is the presence of the three different cell types: mucous, squamoid
(epidermoid), and intermediate, although frequently squamoid cell are incon-
spicuous. These three cell types are present in variable proportions in individ-
ual tumors and form sheets, nests, duct-like structures, or cysts. Intermediate
cells frequently predominate, and range from small basal cells with minimal
basophilic cytoplasm to larger oval cells with pale eosinophilic cytoplasm.
The mucous cells occur singly or in clusters and have pale, foamy cytoplasm,
distinct cell membranes, and eccentric small nuclei (e-Fig. 6.22). They fre-
quently line cystic spaces and are positive with mucicarmine and PAS stains.
Squamoid cells have abundant eosinophilic cytoplasm and vesicular nuclei
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solid nests, lobules, cribriform gland-like structures, duct-like arrangements,

or a characteristic concentric whorling of the cellular nests in a single file
arrangement that has been termed "the eye of the storm." Stromal hyaliniza-
tion with a grayish-blue hue is characteristic. The periphery of the tumor
shows markedly infiltrative growth, and perineural invasion is common
(e-Fig. 6.23). Cytologically, in contrast to the architecture, the tumor cells are
very uniform with moderate eosinophilic cytoplasm and characteristic round
to oval nuclei with open chromatin (e-Fig. 6.24). There is minimal mitotic
activity and no necrosis. The differential diagnosis includes pleomorphic
adenoma, which usually exhibits myxochondroid areas that are not seen
in PLGA, and adenoid cystic carcinoma, which consists of basaloid cells
with more hyperchromatic nuclei than those of PLGA and which has a much
"tighter" cribriform pattern with a much more obvious myoepithelial component.
Conservative resection is the treatment of choice. Recurrence occurs in
10 to 15% of patients. Neck dissection is only recommended for clinical
adenopathy or proven metastasis because lymph node metastases are uncom-
mon. Distant metastases are even less common. Patients have an excellent
long-term prognosis even in the presence of recurrent/metastatic disease, and
deaths due to PLGA are quite rare.
3. Acinic cell carcinoma accounts for only 1% to 3% of salivary gland tumors.
It occurs evenly through the second to seventh decades and can be seen in
childhood (in fact, it is the second most common childhood salivary gland
malignancy). About 80% of cases arise in the parotid gland, usually present-
ing as a slowly growing mass which is occasionally painful.
Grossly, acinic cell carcinoma presents as a single, usually circumscribed,
solid mass that can undergo cystic degeneration. Histologically, the architec-
ture can be solid/lobular, microcystic, papillary-cystic, or follicular. Mixed
architectural patterns are common as is a background inflammatory infiltrate
with germinal center formation (e-Fig. 6.25A). Small tumors can be easily
missed because the acinar cells may be very well differentiated. The acinic
cell is characteristic (and requisite for the diagnosis) and has the appearance
of a salivary acinar cell with abundant granular, basophilic cytoplasm and
a small, round, somewhat eccentrically placed nucleus (e-Fig. 6.25B). PAS
stains will highlight the cytoplasmic zymogen granules, which are resistant to
diastase digestion. A number of other cell types can also be present including
eosinophilic, clear, and vacuolated cells. The periphery of the tumor may not
be infiltrative, but this should not be interpreted as a finding of benignancy.
There is no acinic cell adenoma. The differential diagnosis differs depending
on the architecture and predominant cell type, and includes normal parotid
gland, oncocytomaloncocytic carcinoma, and clear cell carcinoma. The pap-
illary variant of acinic cell carcinoma must be differentiated from cystade-
noma/adenocarcinoma. In addition, a new entity, the mammary analogue
secretory carcinoma, has similar features but has no true serous granules
and has intraluminal secretory material.
Recurrence after excision occurs in approximately one-third of cases.
Although classically regarded as a low-grade malignancy, 10% to 15% of
these tumors will metastasize to regional lymph nodes or distantly, particu-
larly to the lungs and bones. This has been more clarified by recent literature
which shows that approximately 15-20% of cases with have high grade areas
with mitotic activity higher than 2 per 10 high power fields and/or with necro-
sis (e-Fig. 6.26). The higher grade areas usually lose their acinic cell (serous
granules) features. The older term for this was "de-differentiation" although
more recently, authors have favored the terms "high grade" or "with high
grade transformation." Tumors with these features consistently have pro-
gressive disease with poor outcomes and are the ones responsible for the 10-
15% of metastatic cancers noted above. For this reason, grading is actually
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5. 11Malignant mixed tumor" is a broad term that is used to encompass true malig-
nant mixed tumors, carcinoma ex pleomorphic adenoma, and metastasizing
mixed tumor.
a. True malignant mixed tumor (or carcinosarcoma) is a malignant neoplasm
that is composed of both carcinomatous and sarcomatous components,
and is exceedingly rare. The mean age is 58 years, and one-third of patients
have evidence of a preexisting pleomorphic adenoma. Approximately two-
thirds of cases arise in the parotid gland, 15% in the submandibular gland,
and 15% in the minor salivary glands of the palate.
Grossly, there is typically a firm, tan-white mass with hemorrhage,
necrosis, and, on occasion, grittiness or calcification. Microscopically,
there is an intimate admixture of the two components. The carcinoma-
tous component typically takes the form of high-grade salivary duct carci-
noma or undifferentiated carcinoma (e-Fig. 6.31). The sarcomatous com-
ponent is usually chondrosarcoma or osteosarcoma, but fibrosarcoma,
leiomyosarcoma, and even liposarcoma occur rarely. Treatment consists
of wide local excision combined with radiotherapy. The tumors are very
aggressive, with up to two-thirds of patients dying of disease, usually
within 30 months.
b. Carcinoma ex pleomorphic adenoma is defined as a mixed tumor in which
carcinoma is present. This tumor accounts for >95% of malignant mixed
tumors, and is most common in the parotid gland, followed by the minor
salivary glands, submandibular gland, and sublingual gland. Most patients
are in their sixth or seventh decade, which is approximately 1 decade older
than the age of most patients who have pleomorphic adenomas. The classic
history is a patient with a longstanding mass that undergoes rapid growth
over a period of several months.
Grossly, these tumors can reach up to 25 em in diameter, and the aver-
age size is more than twice that of pleomorphic adenomas. The carcinoma-
tous component is usually an infiltrative, hard, white to tan-gray mass with
hemorrhage and necrosis. Microscopically, the proportions of the carci-
noma and pleomorphic adenoma are quite variable, and the pleomorphic
adenoma component may be replaced by scarring or may be overgrown
by the malignant component. The malignant component most often is a
poorly differentiated adenocarcinoma not otherwise specified (e-Figs 6.32
and 6.33), salivary duct carcinoma, or undifferentiated carcinoma. Low-
grade carcinomas such as myoepithelial carcinoma occasionally occur.
Prognosis and management are highly dependent on the type of
carcinoma and extent of invasion. The malignant component should be
classified as noninvasive (intracapsular), minimally invasive (~1.5 mm
in greatest extent), or widely invasive (>1.5 mm). Wide resection is the
treatment of choice, with lymph node dissection and radiation therapy
reserved for widely invasive tumors or tumors with obvious cervical
lymph node metastases.
c. Metastasizing mixed tumor is the least common form of malignant mixed
tumor. These tumors have the same bland morphology of a pleomorphic
adenoma, but metastasize either to local lymph nodes or distantly, usually
to bone and lung. Often, there is a protracted clinical course with many
recurrences at the primary site before nodal or distant metastases develop.
Overall mortality due to the tumor is 40%.
6. Salivary duct carcinoma is one of the most aggressive primary salivary gland
tumors and accounts for <10% of salivary gland tumors. Men are more
commonly affected in a ratio of 4:1. Patients generally present in the sixth
decade with a rapidly growing parotid mass with facial nerve involvement.
Grossly, the tumors are solid and white with hemorrhage, necrosis,
and cystic areas. Infiltration of the surrounding tissue is usually apparent.
Microscopically, it resembles high grade ductal carcinoma of the breast. Duc-

tal carcinoma in situ is often present, and frequently shows a cribriform pat-
tern often with comedo-type necrosis (e-Fig. 6.34 ). The tumor's invasive com-
ponent consists of large cells with abundant eosinophilic cytoplasm and large,
round nuclei with vesicular chromatin and prominent nucleoli (e-Fig. 6.35).
The neoplasm shows marked tissue infiltration with stromal desmoplasia and
brisk mitotic activity, and vascular and perineural invasion are common. Sali-
vary duct carcinomas express low- and high-molecular-weight cytokeratins,
carcinoembryogenic antigen (CEA), androgen receptors, and human epider-
mal growth factor receptor 2 (HER2Jneu), for which the gene is amplified
in up to 1/3rd of cases. The differential diagnosis includes metastatic breast
carcinoma, poorly differentiated squamous cell carcinoma, and mucoepi-
dermoid carcinoma. The presence of intraductal carcinoma is an important
finding that argues strongly for a diagnosis of a primary salivary duct carci-
noma rather than metastasis.
Salivary duct carcinoma is the most aggressive salivary gland tumor.
Approximately one-third of patients develop local recurrence, half develop
distant metastases, and 65% of patients die of their disease, most within
4 years of diagnosis. Wide local excision with neck dissection and postoper-
ative radiotherapy is the treatment of choice.
7. Epithelial-myoepithelial carcinoma is a low-grade carcinoma that accounts
for 0.5% to 1% of salivary gland neoplasms. It arises in the parotid gland,
and occasionally from the minor salivary glands of the larynx or paranasal
sinuses.
Grossly, the tumor is firm and well-demarcated, averaging 2 to 3 em in
diameter. Microscopically, the tumor is usually partially encapsulated, with
invasion by tumor into the adjacent parenchyma. The classic morphology
is of ductal structures lined by an inner layer of eosinophilic epithelial cells
and an outer layer of clear myoepithelial cells (e-Fig. 6.36), which lie in an
eosinophilic, hyaline stroma. In rare cases, the myoepithelial cell component
may be present as large sheets or nests of cells with only focal ductal differenti-
ation. The individual tumor cells are bland with minimal mitotic activity. The
epithelial cells express low-molecular-weight cytokeratins, and the myoep-
ithelial cells express calponin, smooth muscle actin, and p63. The differen-
tial diagnosis includes pleomorphic adenoma, myoepithelioma/myoepithelial
carcinoma, and the tubular variant of adenoid cystic carcinoma.
Epithelial-myoepithelial carcinoma is a moderately aggressive tumor
with a recurrence rate of 40%. Metastases to lymph nodes, lung, or liver
occur in 15% of patients, and overall survival at 5 years is about 80%.
Treatment is wide local excision with or without radiotherapy.
8. Squamous cell carcinomas only rarely as primary tumors of the salivary gland.
Metastases to the intraparotid lymph nodes from primary skin cancers of
the head and neck, particularly of the scalp, eat; and face, are much more
common. Most patients are in their sixth to eighth decades. About 80%
of tumors arise in the parotid gland and 20% in the submandibular gland.
The tumors typically are high stage at the time of diagnosis. By definition,
the diagnosis of primary squamous cell carcinoma is restricted to the large
salivary glands, because tumors arising in the minor salivary glands cannot be
reliably distinguished from primary squamous carcinoma of the surrounding
mucosa.
Grossly, the neoplasm is a firm, white, infiltrative, and nonencapsulated
mass. Microscopically, it is identical to typical squamous cell carcinomas
of the UADT, although cases arising in a salivary gland tend to be well-
differentiated. Prominent desmoplasia is common, and perineural invasion
and extension of the tumor into periglandular soft tissue are frequently
present.
The differential diagnosis most importantly includes metastatic squamous

cell carcinoma. High-grade mucoepidermoid carcinoma can have a largely
squamoid appearance but lacks keratinization, has a more heterogeneous cell
population, and almost always demonstrates mucous cells.
Primary squamous carcinomas are aggressive tumors, and the 5-year sur-
vival of patients is approximately 25%. Treatment involves radical surgery,
neck dissection, and radiotherapy.
9. Malignant counterparts to benign salivary gland tumors are relatively uncom-
mon and microscopically resemble benign salivary gland tumors in differen-
tiation and cellular components but show aggressive features such as an inva-
sive growth, perineural invasion, lymphvascular invasion, cellular anaplasia,
necrosis, and/or metastasis. The more common types are basal cell adenocar-
cinoma (e-Fig. 6.37) and myoepithelial carcinoma (e-Fig. 6.38), although the
majority of benign salivary gland tumors have a reported malignant coun-
terpart.
10. Metastasis to the salivary glands or, more commonly, to intra- or periglan-
dular lymph nodes, is a frequent occurrence.
The parotid contains an average of 20 lymph nodes which drain the
scalp, face, ear skin, external auditory canal, and tympanic membrane. Skin
tumors such as squamous cell carcinoma and melanoma, therefore, account
for approximately 80% of metastases to the gland. The remaining metastases
are from non-head and neck primary tumors, most commonly lung, kidney,
and breast carcinomas.
The opposite distribution of metastases is seen for the submandibular
gland, which does not contain intraglandular lymph nodes. More than 85%
of metastases arise from infraclavicular primary tumors, most commonly
breast, kidney, and lung carcinomas, particularly pulmonary small cell
carcmoma.
11. Pediatric tumors are uncommon. Hemangioma is the most frequent, but most
salivary gland neoplasms that occur in adults can also occur in children. There
are two congenital tumors that are worth mentioning here.
a. Sialoblastoma is an extremely rare, potentially aggressive neoplasm that
recapitulates the embryonic stage of salivary gland development and
is thought to develop from retained blastomatous cells. Clinically, it is
seen in the perinatal to the neonatal period, usually involving the parotid
gland. This tumor may grow quickly and cause skin ulceration or airway
compronnse.
Grossly, the mass is lobulated and partially circumscribed. Microscop-
ically, it is composed of nests or nodules of basaloid cells with scanty cyto-
plasm, round to oval nuclei, and fine chromatin with small nucleoli. The
mitotic rate is highly variable and can be quite high. Complete surgical
excision with a rim of normal tissue is the treatment of choice. Although
local recurrence is relatively common, occurring in up to 30% of cases,
metastasis is extremely uncommon.
b. Salivary anlage tumor, also referred to as a congenital pleomorphic ade-
noma, occurs in male neonates in the first 2 weeks of life and is associated
with respiratory obstruction or difficulty feeding. Sometimes the mass is
spontaneously passed or inadvertently removed by airway suctioning. On
examination, a mass attached by a thin stalk to the posterior nasal mucosa
or nasopharynx is sometimes present.
Grossly, salivary anlage tumors are firm and tan-yellow with a smooth
surface. Microscopically, the tumor's surface shows nonkeratinizing squa-
mous epithelium, with a deeper stroma composed of bland spindled
cells and intervening squamous islands. The overall histology suggests a
hamartoma rather than a true neoplasm. The treatment is simple excision
of the mass, and the lesion does not recur or spread.
•• I SECTION I. HEAD AHD NECK
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V. PATHOLOGIC REPORTING OF MALIGNANT SALIVARY GLAND TUMORS

A. Staging: American Joint Committee on Cancer (AJCC). Clinical staging of salivary
gland cancers is important for prognosis and treatment decisions. The 2010
Tumor, Node, Metastasis (TNM) AJCC staging classification is provided in
Table 6.5. Minor salivary gland carcinomas are staged according to the anatomic
site of origin (e.g., oral cavity, sinus, larynx). Staging guidelines are applicable
to all forms of carcinoma. Any nonepithelial tumor type is excluded.
B. Additional pertinent pathologic features. Pathologic features such as tumor grade,
positive resection margins, and skin or bone invasion have been demonstrated
in numerous studies to correlate with a higher risk of local recurrence, a poorer
prognosis, or both, and so should always be reported. Perineural and lympho-
vascular space invasion should also always be reported when observed, because
they have been correlated with distant metastases in some studies, even though
they are not necessarily correlated with recurrence or poorer prognosis.
Cytopathology of the
Salivary Glands
Brian Collins
The parotid and minor salivary glands are superficially located and palpable, and so
salivary gland lesions are readily amenable to percutaneous FNA. A pattern approach
is typically utilized for the evaluation of FNA specimens for diagnosis, classification,
and clinical management of patients.
I. NON-NEOPLASTIC CONDITIONS
A. Chronic sialadenitis. Chronic inflammation can cause diffuse or focal enlarge-
ment which by palpation can be indistinguishable from a neoplasm. However,
chronic sialadenitis is very tender on FNA, which can be a helpful due to diag-
nosis. Aspiration smears are hypocellular with benign ductal epithelial groups
which have a minimal degree of nuclear atypia. Acinar elements can be scant
or absent due to the underlying destructive nature of the process. A variable
mixture of large and small lymphocytes is present. Acinic cell carcinoma is
an important differential diagnostic consideration when presented with bland
acinar-type cells and lymphocytes on aspirate smears.
B. Lymph nodes. The parotid is a common site of intra-parenchymal lymph nodes
which can be clinically difficult to distinguish from a neoplasm. Aspirate smears
will show a mixture of lymphocytes in the pattern typically seen with reactive
lymphoid hyperplasia.
II. BENIGN NEOPLASMS
A. Warthin tumor. This tumor commonly arises in the parotid gland and by aspi-
ration shows a mixture of lymphocytes and cohesive groups of oncocytic-type
epithelial fragments (e-Figs. 6.39 and 6.40). The lymphoid elements are inti-
mately admixed with the oncocytic cells, and the background usually shows
cystic change and with macrophages and granular debris.
B. Pleomorphic adenoma. Aspiration smears can show a wide variety of patterns
and cellular elements. Commonly, there is a mixture of bland epithelial, spin-
dled, and myoepithelial-like cells which occur as single cells and are intimately
admixed with a variable amount of fibromyxoid stroma (e-Figs. 6.41 and 6.42).
The relative proportions of these elements can vary significantly between cases.
Diff-Quik staining of smears helps to highlight the metachromatic fibromyxoid
stroma. When present in the customary pattern, a definitive diagnosis can be
rendered.
C. Basal cell adenoma. Aspiration smears are cellular and contain small uniform
basaloid cells with scant cytoplasm and round to oval nuclei with indistinct
nucleoli. The dense extracellular material present, usually less than that seen in
pleomorphic adenoma, tends to be arranged in globules and trabecular-tubular
configurations (e-Fig. 6.43). This pattern typically is classified as a basaloid neo-
plasm and the differential diagnosis includes adenoid cystic carcinoma, since the
cytomorphologic pattern on aspiration does not permit definitive classification.
Ill. MALIGNANT NEOPLASMS
A. Adenoid cystic carcinoma. Aspiration shows small basaloid cells with globules
of dense matrix material. The cells are monomorphic with round to oval nuclei,
even chromatin, indistinct nucleoli, and scant cytoplasm. Aspirates are usually
cellular with cell groups arranged in syncytial and three-dimensional fragments.
The extracellular matrix consists of dense amorphous material with tubular
and/or globular shapes (e-Figs. 6.44 to 6.46). Classically, the cells appear to
surround the round globules. Because of the cytomorphologic overlap with other
basaloid tumors, definitive classification is not possible.
B. Mucoepidermoid carcinoma. These neoplasms vary in their cellular composition,
and this difference is reflected in the patterns and cellular elements encountered
by FNA. For low-grade neoplasms, aspiration smears are hypocellular and con-
tain thick mucoid-like material with granular debris. The three cell types that
characterize the tumor (squamoid or epidermoid), mucous, and intermediate
are all present, and they can be arranged separately or be admixed. The nuclear
features of all three cell types are bland (e-Figs. 6.47 and 6.48). For high-grade
neoplasms, aspirate smears are cellular with groups and sheets of cells with a
squamoid appearance. Nuclei are large with coarse chromatin and prominent
nucleoli; the cytoplasm is dense, and moderate in amount with well-defined cell
borders. Mucin cells with intracytoplasmic vacuoles are usually few and can be
difficult to identify (e-Figs. 6.49 and 6.50).
C. Acinic cell carcinoma. Aspirate smears are cellular with a mixture of single cells,
small groups of cells, and three-dimensional clusters. The cells are intermediate
in size with monomorphic round bland nuclei with nucleoli. The cytoplasm is
critical in the diagnosis and will demonstrate features of acinar differentiation
with delicate vacuoles and granules (e-Figs. 6.51 and 6.52). While usually dean,
the background can show the features characteristic of cystic lesions and show
significant lymphoid elements.
SUGGESTED READINGS
Eveson ]w. Malignant neoplasms of the salivary glands. In: Thompson LDR, ed. Head and Neck
Pathology. New York, NY: Churchill Livingstone; 2006.
Eveson JW, Auclair P, Gnepp DR, et al. Tumors of the salivary glands. In: Barnes L, Eveson JW,
Reichart P, Sidransky D, eds. Pathology and Genetics Head and Neck Tumors. Lyon, France:
IARC Press; 2005.
Faquin WC, Powers CN. Salivary Gland Cytopathology: Essentials in Cytopathology. New York,
NY: Springer; 2008.
Gnepp DR, Henley JD, Simpson RHW, et al. Salivary and lacrimal glands. In: Gnepp DR, ed. Diag-
nostic Surgical Pathology of the Head and Neck. Philadelphia, PA: W.B. Saunders Publishers;
2009.
Layfield L. Cytopathology of the Head and Neck. Chicago: ASCP Press; 1997.
Powers CN, Frable WJ. Fine Needle Aspiration Biopsy of the Head and Neck. Boston: Butterworth
Heinemann; 1996.
Richardson MS. Non-neoplastic lesions of the salivary glands. In: Thompson WR, ed. Head and
Neck Pathology. New York, NY: Churchill Livingstone; 2006.
Torske K. Benign neoplasms of the salivary glands. In: Thompson LDR, ed. Head and Neck Pathol-
ogy. New York, NY: Churchill Livingstone; 2006.
Wenig BM. Major and minor salivary glands. In: Wenig BM, ed. Atlas of Head and Neck Pathology.
Philadelphia, PA: Saunders Elsevier; 2008.
The Ear
Peter A. Humphrey and Rebecca D. Chernock
I. NORMAL ANATOMY. The ear is composed of the external ear, the middle ear, and the
inner ear. The external ear is made up of the auricle, which leads to the external
auditory canal. The auricle has a supporting plate of elastic cartilage, which also
helps form the outer two-thirds of the external auditory canal. Skin covers both
the auricle and the canal; the main distinctive histologic features of this skin are
that the squamous lining of the inner half of the canal is thinned, and that modified
apocrine glands called ceruminal glands are present in the outer third of the canal.
The clustered ceruminal glands are lined by cuboidal epithelial cells that have
an eosinophilic cytoplasm that often harbors a granular golden-yellow pigment
(e-Fig. 7.1).*
The middle ear, or tympanic cavity, lies within the temporal bone. It is separated
from the external auditory canal by the tympanic membrane, a thin fibrous sheet
that has an external keratinizing squamous epithelial lining and an inner cuboidal
cell lining. The middle ear contains the three auditory ossicles (malleus, incus,
and stapes), ossicle ligaments, tendons of the ossicular muscles, the auditory tube,
the tympanic cavity itself, and the epitympanic recess, the mastoid cavity, and the
chorda tympani of the facial nerve (cranial nerve VII). The auditory or eustachian
tube connects the tympanic cavity with the nasopharynx. The tympanic cavity is
lined by a single layer of flattened to cuboidal respiratory epithelium, whereas most
of the auditory tube is lined by low ciliated epithelium.
The inner ear is located within the petrous portion of the temporal bone and
is composed of a membranous labyrinth surrounded by an osseous labyrinth. The
membranous labyrinth houses the cochlea and the vestibular apparatus, both of
which are supplied by cranial nerve VIII. There are several parts to the cochlea:
the cochlear duct with the organ of Corti (the end organ of hearing), and the
scala vestibuli and scala tympani, which hold the perilymph. The organ of Corti
has thousands of neurotransmitting hair cells. The vestibular apparatus, which
functions in motion and position sensing, consists of three semicircular canals and
the utricle and the saccule. The ampullae of the canals have a sensory end organ,
the crista ampullaris, with neurosensory hair cells. The utricle and the saccule
also possess a sensory end organ, the macula, which has neurosensory hair cells
and otoliths. There is also a blind sac in the membranous labyrinth known as the
endolymphatic sac, which is lined by tall columnar epithelium arranged on papillae.
II. GROSS EXAMINATION AND TISSUE SAMPLING
A. External ear. Biopsy and excision specimens should be handled as skin specimens
from other anatomic sites (see Chaps. 38-40).
B. Middle and inner ear. Samples from the middle ear are often obtained in cases
of suspected cholesteatoma. For these cases, it should be noted whether bone
fragments are present. Standard hematoxylin and eosin (H&E) slide preparation
is sufficient. Ossicles from the middle ear can be processed by gross examination
only unless microscopic examination is requested by the surgeon. For middle ear
and inner ear neoplasms, which are uncommon, use of ink to mark the peripheral
margins is not usually necessary because these specimens are typically received
as small fragments. In those rare cases in which the patient has a history of
97
lymphoma or lymphoma is suspected clinically, fresh tissue should be processed

according to the standard lymphoma work-up protocol. For all other middle ear
samples, all tissue should be submitted for histologic examination, with H&E
slide generation.
Ill. COMMON DISEASES OF THE EXTERNAL EAR
A. Nonneoplastic diseases. The common diseases of skin that involve the pinna and
the external ear canal are covered in the chapters on skin (Chaps. 38-40). Some
diseases have a particular predilection for the skin of the ear, including gout,
keloids (often secondary to ear piercing), relapsing polychondritis, angiolym-
phoid hyperplasia with eosinophilia (epithelioid or histiocytoid hemangioma),
and chondrodermatitis nodularis (the latter is discussed below).
1. Congenital anomalies of the ear that may be seen by the surgical patholo-
gist include accessory tragi, branchial cleft abnormalities, congenital aural
sinuses, and salivary gland ectopia.
a. Accessory tragi are found at birth and clinically and macroscopically are
most often solitary, sessile, or pedunculated polyps in the preauricular
area. Microscopically, skin, hair follicles, and a central fibrofatty core
with or without cartilage are observed (e-Fig. 7.2). They should not be
misdiagnosed as a papilloma, fibroma, or chondroma.
b. Anomalies of the first branchial cleft present near the ear as cysts, sinuses,
and fistulas. The epithelial lining can be squamous or respiratory; type I
or pure squamous cell-lined cysts can be confused with keratinous cysts
histologically. Lymphoid tissue in the wall (e-Fig. 7.3) is less frequently
found in the first branchial cleft cysts than in the much more common
second branchial cleft cysts, which arise in the lateral neck. Type II defects
can harbor skin including adnexal structures or cartilage (e-Fig. 7.4 ); asso-
ciated salivary gland tissue may also be present when the process extends
into or near the parotid gland.
c. Congenital aural sinuses are distinguished from branchial cleft anomalies
by location: Branchial cleft abnormalities are found in infra- or postauric-
ular sites, whereas congenital aural sinuses are present in a preauricular
location.
2. Chondrodermatitis nodularis helicis is a condition of uncertain etiology that
occurs on the skin of the external ear, usually on the upper part of the
helix. Clinically, middle-aged or olde.t;, typically male, patients present with
a small (<1 em) painful nodule that can be ulcerated and can exhibit a
crust. This appearance can clinically simulate actinic keratosis or squamous
cell carcinoma. Microscopically, there is a somewhat funnel-shaped ulcer
with associated dermal collagen edema and degeneration. There may be
surrounding pseudoepitheliomatous squamous cell hyperplasia (e-Fig. 7.5),
granulation tissue and fibrosis, and a predominantly lymphocytic inflam-
matory cell infiltrate, although the infiltrate can be mixed. A perichondri-
tis with destruction of cartilage can be seen but is uncommon. Superfi-
cial or shave biopsies may show only a few of the above findings. Curet-
tage and cautery are used for treatment, with recurrence in a minority of
patients.
3. Otitis externa is inflammation of the external auditory canal and/or pinna and
is very common in clinical practice. Biopsy is generally not indicated.
4. Necrotizing (malignant) otitis externa is usually caused by Pseudomonas aerug-
inosa infecting diabetic patients, but fungi can also be the causative agent.
Microscopically, the response is one of necrotizing inflammation.
B. Neoplasms of the external ear. The 2005 World Health Organization (WHO)
histologic classification of tumors of the ear is given in Table 7.1. The most
common neoplasms of the external ear are basal cell carcinoma and squamous
cell carcinoma of the skin of the ear. Of the tumors of the external ear, only
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2. Malignant tumors of ceruminous glands include adenocarcinoma, adenoid cys-

tic carcinoma, and mucoepidermoid carcinoma. The latter two are histologi-
cally identical to those arising in salivary glands. Ceruminous adenocarcino-
mas show infiltrative growth and range from cytologically bland to markedly
atypical with an increase in mitotic activity. Perineural invasion is uncom-
mon, but can be a useful diagnostic clue favoring adenocarcinoma when
detected, especially in small biopsy samples. Ceruminous adenocarcinomas
are locally aggressive.
3. Other unusual neoplasms and tumorlike conditions of the external ear include
malignant melanoma, benign fibro-osseous lesion, osteoma and exosto-
sis, and idiopathic pseudocystic chondromalacia (which is a nonneoplastic
swelling of the pinna due to fluid accumulation within the cartilage of the
ear).
IV. COMMON DISEASES OF THE MIDDLE EAR
A. Nonneoplastic middle ear diseases. The common disorders include choristoma,
inflammation, infection, cholesterol granuloma, cholesteatoma, and otosclero-
sis.
1. Choristomas (heterotopic tissues) in the middle ear are composed of benign
salivary gland, glial, or sebaceous gland tissue. Glial heterotopia is histologi-
cally identical to the more commonly occurring encephalocele; the two must
be distinguished clinically (e-Fig. 7.7A and B).
2. Otitis media is one of the most common diseases of childhood. Most acute
purulent cases are due to bacterial infection by Streptococcus pneumoniae
or Haemophilus influenzae. Tissue samples are not usually procured, but in
chronic cases tissue may be removed. Microscopically, granulation tissue,
scar tissue, chronic inflammation, calcific debris, and sclerotic or reactive
bone can be seen. Occasionally, neutrophils and foreign body-type giant cells
are present. There may be associated polypoid granulation tissue, choles-
terol granulomas, cholesteatoma, or tympanosclerosis. Entrapped metaplas-
tic glands should not be mistaken for neoplastic glands.
3. Cholesterol granulomas are found in a number of chronic ear diseases. Choles-
terol clefts, a foreign body-type giant cell reaction, and hemosiderin deposi-
tion are characteristic (e-Fig. 7.8).
4. Cholesteatoma may be congenital or acquired (Eur Arch Otorhinolaryngol.
2004;261:6). The congenital form is found in infants and young children, is
defined as occurring in the presence of an intact tympanic membrane, and
may result from an epidermoid cell rest (epidermoid formation). In the more
common acquired form, seen mainly in older children and adults, there is an
association with severe otitis media and a perforated tympanic membrane.
Histologically, there are three major elements: keratin (e-Figs. 7.9 and 7.10),
stratified squamous epithelium, and fibrous and/or granulation tissue. Since
squamous epithelium is not normally found in the middle ear, the diagnosis
is usually straightforward. Downgrowth of the epithelium into underlying
subepidermal connective tissue may be appreciated. Marked vascular con-
gestion and abscess formation may also be found. Cholesteatoma is not a
neoplasm, but can be locally destructive; ossicle(s) and the bony wall of the
middle ear can be eroded. There is an increased cell proliferation index in the
squamous epithelium of cholesteatoma (Acta Otolaryngol. 2003;123:377)
and overexpression of cathepsin enzymes (Laryngoscope. 2003;113:808),
abnormalities that may be related to the local growth but are not required
for diagnosis.
5. Otosclerosis is a disease of unknown etiology that leads to progressive fix-
ation of the stapes footplate and, as a result, conductive hearing loss. The
onset is typically in adulthood and women are more often affected than men.
Chapter 7 • The Ear I 101
The initial histologic changes include resorption of bone and replacement by

cellular fibrovascular tissue. Over time, this is replaced by dense sclerotic
bone (e-Fig. 7.11). Patients are treated with stapedectomy.
B. Neoplasms of the middle ear include paraganglioma, adenoma of the middle ear,
papillary tumors, meningioma, and squamous cell carcinoma (Table 7.1).
1. Paraganglioma (also known as glomus tumor, glomus tympanicum, or
chemodectoma) is the most common tumor of the middle ear (but is nonethe-
less still rare). It usually presents clinically with hearing loss or tinnitus
in patients in the fifth and sixth decades of life. The neoplasm can be
solitary or part of a familial paraganglioma-pheochromocytoma syndrome
caused by germline mutations in SDHB. SDHC. or SDHD genes; the head
and neck paragangliomas in this syndrome are bilateral/multicentric. Clin-
ically, paraganglioma is bulging, red, pink, or bluish (and not white like a
cholesteatoma). Biopsy may result in brisk bleeding.
Microscopically, sections usually demonstrate the classical "zellballen"
appearance with nests of small uniform epithelioid cells that have a periph-
eral layer of flattened cells (e-Figs. 7.12 and 7.13). Sclerosis and vascularity
can be pronounced in some tumors; in the former circumstance, the nested
pattern may not be apparent (e-Fig. 7.14). Immunohistochemical stains can
be confirmatory and particularly contributory in small biopsy samples, which
may display significant crush artifact. Chromogranin A and synaptophysin
immunostains are positive, whereas carcinoembryogenic antigen and ker-
atin immunostains are negative. Only a few S-1 00-positive peripheral sus-
tentacular cells may be present. These tumors grow slowly and can recur
after surgery and/or radiation treatment. Intracranial extension develops in
a small minority of patients, and about 1% to 2% of patients suffer from
metastatic spread. It is not possible to predict aggressive behavior based on
histopathologic features.
2. Adenoma of the middle ear is a benign glandular neoplasm with variable
neuroendocrine and mucin-secreting differentiation (Arch Pathol Lab Med.
2006;130:1067). The typical clinical presentation is in an adult (mean age in
the 40s) with muffled hearing, tinnitus, and/or a sensation of pressure and/or
fullness. The neoplasms are variably colored and only uncommonly penetrate
through the tympanic membrane. Microscopic architectural patterns include
closely packed small glands with solid and/or trabecular arrangements (e-Fig.
7.15). The glands lack a myoepithelial layer. Cytologically, the cuboidal to
columnar glandular cells are bland, with uniform small nuclei and rare nucle-
oli (e-Fig. 7.16). Mitoses should not be present. The immunoprofile includes
positivity for cytokeratin and neuroendocrine markers such as chromogranin
and synaptophysin. In the past, this immunophenotype was viewed as being
indicative of a carcinoid tumor of the middle ear, but it is now recognized
that expression of neuroendocrine markers is a characteristic feature of most
middle ear adenomas. Recurrence has been reported in a small minority of
cases, usually after incomplete surgical excision.
3. Papillary tumors ofthe middle ear include aggressive papillary tumor, Schneide-
rian papilloma, and inverted papilloma, although only a few cases of the latter
two neoplasms have been described. Aggressive papillary tumors are more
common (Adv Anat Pathol. 2006;13:131), and are also known as low-grade
papillary adenocarcinomas and endolymphatic sac tumors (Heffner tumor).
Typically, patients in their fourth decade present with hearing difficulty
and vertigo; 15% of patients have a family history of von Hippel-Lindau
syndrome. Microscopically, there are complex interdigitating papillae with
fibrous cores covered by cuboidal to columnar cells that have bland nuclear
cytology and eosinophilic cytoplasm. Mitoses are absent. The papillae may
overlie the granulation tissue causing diagnostic confusion with reactive pro-
cesses. Cystic spaces with a colloid-like material simulating thyroid follicles
can also be present. Immunostains show positivity for keratin, with variable
S-100, glial fibrillary acid protein (GFAP), and synaptophysin immunoreac-
tivity; thyroglobulin immunostaining is negative. Despite the bland histologic
appearance, the tumor is a slowly growing, locally aggressive, but nonmetas-
tasizing neoplasm. Temporal bone invasion is common, and the tumor may
extend into the cerebellum. Outcome is related to size of the tumor and ade-
quacy of excision; radical surgical excision affords the best chance for cure.
Recurrence is seen in about 20% of cases, with tumor-specific death in about
13% of patients.
4. Meningioma in the middle ear is a rare neoplasm that is more likely to rep-
resent secondary extension from an intracranial meningioma than from a
primary middle ear meningioma. Patients with primary middle ear menin-
giomas present at a mean age of 50 years with hearing changes and sometimes
otitis and pain (Mod Pathol. 2003;16:236). The histopathologic features
are similar to intracranial meningiomas, with meningothelial meningioma
predominating (e-Fig. 7.17). Vimentin, progesterone receptot; and epithelial
membrane antigen immunostains are positive, and cytokeratin immunostains
are negative. Meningiomas are slowly growing neoplasms and can recur fol-
lowing incomplete surgical excision. The 5-year survival is 83%.
5. Squamous cell carcinoma in the middle ear is uncommon and is typically
advanced at presentation. Its development is not clearly related to chronic
otitis media and is not related to cholesteatoma. Microscopically, the carci-
noma is keratinizing and displays a variable degree of differentiation. Out-
come is related to tumor extent and margin status at surgery, but not his-
tologic grade. The 5-year survival is roughly 50% (Int j Radiat Oncol Biol
Phys. 2007;68:1326).
6. Other rare neoplasms of the middle ear include embryonal rhabdomyosar-
coma (e-Fig. 7.18), lipoma, hemangioma, osteoma, ossifying fibroma, and
teratoma.
7. Metastasis to the ear is uncommon, accounting for only 2% to 6% of all
neoplasms of the ear. Most patients have known, widely disseminated can-
cer. The middle ear is the most common site for metastatic spread. Breast,
lung, and prostate are, in ordet; the common primary sites of origin for the
metastatic deposits.
V. NEOPLASMS OF THE INNER EAR. Vestibular schwannoma, lipoma, and hemangioma
are the most common neoplasms of the inner ear. Endolymphatic sac tumors also
arise in the inner ear (the aforementioned aggressive papillary tumor of the middle
ear is thought to represent an endolymphatic sac tumor with extension into the
middle ear).
A. Vestibular schwannoma (acoustic neuroma) is relatively common. Unilateral
vestibular schwannoma accounts for 5% to 10% of all intracranial tumors,
and has been found in about 1% of autopsies. Bilateral vestibular schwan-
noma, found in 5% of all vestibular schwannoma cases, is characteristic of
neurofibromatosis type 2 (an autosomal dominant disease due to mutations in
the NF2 gene on 22q12). Patients, usually in their 40s or 50s, present with pro-
gressive hearing loss and tinnitus. Grossly, the size range of the tumor is a few
millimeters up to 6 em in maximal dimension. The smaller tumors are round
to oval, whereas large tumors can assume a mushroom shape. The cut surfaces
are yellow, and can exhibit hemorrhage and cystic change. Microscopically, the
attributes are the same as in soft tissue schwannomas (e-Fig. 7.19), with Antoni
A regions with Verocay bodies, and Antoni B areas. Mitotic figures should be
rare. Degenerative nuclear atypia should not be taken as a sign of malignancy.
S-100 immunoreactivity is strong.
Chapter 7 • The Ear I 103
B. Lipomas of the internal auditory canal resemble lipomas at other anatomic sites,
except that cranial nerve VII or VITI or their branches may be present in the
lipoma.
SUGGESTED READINGS
Barnes L. The ear. In: Silverberg SG, ed in chief, Silverberg·s Principles and Practices of Surgical
Pathology and Cytopathology. Philadelphia, PA: Churchill Livingstone Elsevier; 2006:2268-
2287.
Barnes L, Eveson JW, Reichart P, Sidransky D, eds. Tumors of the Ear, Chapter 7. In: Pathology
and Genetics Head and Neck Tumours. Lyon, France: IARC Press; 2005.
Wenig BM. Diseases of the External Ear, Middle Ear and Temporal Bone. Chapter 9.ln: Barnes L,
ed. Surgical Pathology of the Head and Neck. New York, NY: Informa Healthcare; 2009:423-
474.
SECTION II
Thorax
Lung
Jon H. Ritter and Hannah R. Krigman
I. NORMAL ANATOMY. The lung is defined by airway branching, first into right and
left lobes, then segments, and finally, the functional unit of the lung, the lob-
ule. Arteries follow the airways, while veins and lymphatics flow toward lobular
septa and finally to the hilum and main pulmonary veins. Bronchi are lined by
a pseudostratified respiratory epithelium that includes goblet cells, which is sep-
arated by basement membrane from a delicate submucosa. Larger airways have
a smooth muscle wall that contains minor salivary glands and a cartilaginous
skeleton; bronchioles have lost the latter components. Each lobule has a cen-
tral bronchovascular bundle; the interstitial space between the pulmonary artery
branches and the bronchioles is eventually continuous with the alveolar intersti-
tium. Lymph nodes occur within the lung, and also are discontinuous along the
bronchi.
Progressive branching of the airways leads to the alveolar ducts, from which
alveoli spring. Normal alveolar walls are very delicate and have a fine elastic tissue
matrix. The barrier between the blood in capillaries and air in the alveolar space
consists of the endothelial cell, the basement membrane, the wispy alveolar inter-
stitium, epithelial cell basement membrane, and the flattened type 1 pneumocyte.
Type 2 pneumocytes can proliferate to replace injured type 1 cells.
The visceral pleura consists of an inner vascular layer that abuts the alveolar
tissue, a connective tissue layet; and an outer layer of mesothelium. The visceral
pleura is reflected back at the hilum and becomes continuous with the parietal
pleural layer that lines the chest cavity.
II. SPECIMEN HANDLING AND REPORTING
A. Samples. Biopsies of lung tissue for diagnosis are obtained endoscopically
(transbronchial or endobronchial biopsy), via radiologically guided procedures
(usually needle core biopsies), or for peripheral disease, via wedge biopsies.
Resections can involve a single lobe (lobectomy), two lobes (bilobectomy), or
an entire lung (pneumonectomy.)
B. Gross examination and sampling
1. Endoscopic biopsies are usually submitted in a single cassette. The fragments
are small; use of a nylon mesh bag or a filter paper wrap is preferable to
submission on biopsy sponges since tissue can be compressed or lost in
the pores of sponges. The number of fragments, range of size, and color
should be recorded. Three hematoxylin and eosin stained levels should be
examined. For smaller biopsies, additional unstained levels may be cut at
the time of initial sectioning in case special stains are needed.
104
Chapter 8 • Lung I 105
Core biopsies of nodules presumed to represent neoplasms are obtained

under CT guidance. They should be processed as for endoscopic biopsies.
2. Wedge resections are performed for either non-neoplastic or neoplastic pro-
cesses. The specimen should be measured in three dimensions and weighed.
Assessment of alveolar architecture may be improved by gently inflating
the specimen by injection of formalin at multiple sites. If the specimen is
inflated, the gross description should include this fact. Wedge biopsies gen-
erally have multiple staple lines, which should be cut off as close to the
staples as possible; ideally, sections are taken perpendicular to the staple
line. The entire specimen should be submitted in non-neoplastic cases; lev-
els are usually not necessary. If the specimen is obtained for diagnosis of a
neoplasm, the presence and dimensions of any masses should be described,
as well as the distance to the staple line(s).
3. Lung resections should be described as lobectomy, bilobectomy, or right
or left pneumonectomy. The specimen should be measured in three dimen-
sions and weighed. Any additional designation by the surgeon (e.g., sutures)
should be described. If a portion of chest wall is attached to the specimen, its
size in three dimensions and the number of ribs or other attached structures
should be noted. For lobectomy and pneumonectomy specimens, insuffla-
tion by injection of formalin into the bronchial orifices improves fixation
and visualization of changes. Occasionally, obstruction of bronchi by tumor
makes this technique ineffective, in which case the lung can be expanded
by injection of formalin at multiple sites. The pleural surface should be
described; areas of puckering, dullness, or adhesions should be noted, as
should adhesions of lobes together in multilobe resections. Areas of pleural
distortion over a mass should be marked with ink. The hilar area should
be examined and the number and size of bronchial stumps noted. The lung
can be sectioned in either parasagittal or in coronal planes; it is important
to choose planes to highlight the extent of tumor, pleural invasion, invasion
of adjacent structures, proximity to the hilum, and relationship to major
airways and vessels. At least four sections of tumor should be taken, includ-
ing one or two with the closest pleural surface (which may be at the hilum).
If tumor is invading structures such as a rib, chest wall, or mediastinal soft
tissue, a section that shows tumor in the lung in continuity with the involved
structure should be taken. Other sections should include normal appearing
lung, areas distal to the tumor that show obstructive pneumonia, and any
additional nodules or lesions. Vascular and bronchial margins, hilar lymph
nodes, and peribronchial lymph nodes should be submitted as well.
C. Adjunct Information. For both non-neoplastic and neoplastic samples, the radio-
graphic findings are an important adjunct to diagnosis. Either review the radio-
graph directly, or review its interpretation; old films may provide information
on the pace of disease. The distribution of abnormalities, the presence of lym-
phadenopathy, and the extent of disease all contribute to the final diagnosis. For
non-neoplastic cases, a review of the clinical history, including the presence of
systemic illnesses, medications, exposures; laboratory data including cultures
and serologic studies; and pulmonary function tests all assists in interpretation
of the findings.
D. Microscopic description
1. A description of small biopsies includes the number of pieces and their
constituent elements: alveolar tissue, bronchial wall, alveolar tissue, and
superficial detached fragments of epithelium. The number of fragments is
not insignificant; six fragments are considered to be representative of lung.
The low power impression of normal or abnormal alveolar architecture
should be included. Expansion of the alveolar septa may be by inflam-
matory tissue, cellular tissue, or acellular fibrosis. Alveolar spaces may be
106 I SECTION II: THORAX
empty, or filled with blood, histocytes, or exudates. The vasculature may

be unremarkable, thickened, show inflammation or vasculitis, or contain
tumor or thromboemboli. The bronchial epithelium can be columnar, squa-
mous, or dysplastic. The absence or presence and type of granulomas should
be described. Inflammatory cells should be noted and characterized as to
type and distribution, whether alveolar, septal, peribronchial, perivascular,
or diffuse. The results of special stains for organisms or fibrosis should be
reported.
Biopsies for a neoplasm should include the same quantitation of the
biopsy fragments as for non-neoplastic samples, as well as a description of
the neoplasm. An in situ or dysplastic component should be described, if
present. Detectable vascular space invasion should be noted.
2. The pathology report of wedge resections for non-neoplastic processes
should contain the same information as for smaller biopsies, but should also
include additional assessment of larger airways and vessels, and distribu-
tion of any non-neoplastic processes. Fibrosis, for example, can be diffuse,
peripheral, or sparing the periphery. Similarly, inflammation or granulomas
can be perivascular, peribronchial, or distributed along septa.
3. Definitive resections of neoplasms, from wedge resections to pneumonec-
tomies, should provide as much information as possible. Prior sampling
(endoscopic biopsy, mediastinoscopies, or previous wedge resections) and
prior treatment (chemotherapy or irradiation) should be included in the
pathology report. Margins should be evaluated, and the non-neoplastic
lung should be described.
Ill. NON-NEOPLASTIC DISEASE
A. Acute lung injury patterns. When evaluating lung biopsies taken for medical
diseases, identification of patterns of acute lung injury should be a major point
of emphasis. These patterns represent the response of the lung to an acute
injury, and immediately switch the diagnostic considerations away from the
chronic fibrosing interstitial lung diseases. The commonly recognized patterns
of acute lung injury include diffuse alveolar damage (DAD), bronchiolitis oblit-
erans/organizing pneumonia, and acute interstitial pneumonia (AlP). All three
processes share key findings: they all feature fibrosis characterized by loose,
fibroblast-rich, new fibrous tissue; they are temporally uniform; and they may
be potentially reversible, at least before significant collagen fibrosis develops.
1. DAD is the prototypical pattern of acute lung injury (e-Fig. 8.1).* It is the
pattern of histologic changes underlying the clinical syndrome of acute res-
piratory distress syndrome (ARDS), the clinical triad of diffuse lung infil-
trates, hypoxemia, and decreased pulmonary compliance. DAD has myr-
iad causes, as detailed in Table 8.1. DAD is caused by cellular injury to
pulmonary epithelial and endothelial cells. The initial insult causes edema
from leaky capillaries, and the sloughed cellular material and fibrinous exu-
date form hyaline membranes; there is often surprisingly little inflammation
in this early phase, known as the exudative stage. By six to seven days after
the initializing injury, the edema and hyaline membranes begin to disappear,
and hyperplastic and regenerative type 2 pneumocytes are prominent. By
seven days, the process of organization is well underway, with interstitial
and airspace fibroblastic proliferations; this stage is known as the prolifera-
tive or organizing phase. During organization, the alveolar exudates may be
incorporated into the alveolar wall if proliferating type 2 pneumocytes grow
on top of the exudate rather than along original alveolar basement mem-
brane. Likewise, alveolar collapse may lead to further remodeling of prior
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Chapter 8 • Lung I 10 9
of this disease. Acute inflammation is often restricted to the honeycomb

areas, which may contain mucoid debris. Spatial heterogeneity refers to the
finding that the most severe fibrosis is in the subpleural areas and along
lobular septa; more central parts of the pulmonary lobule typically show
less severe disease. Chronic inflammation is mild and patchy within areas of
fibrosis; autoimmune or connective tissue disorders (CfDs) should be con-
sidered in cases with more extensive chronic inflammation. Related changes
in the lung include secondary pulmonary hypertension and type II pneumo-
cyte hyperplasia. The clinical course both before and after diagnosis is vari-
able; most patients die within five years of diagnosis, although some patients
have a more protracted course. In the process known as acute exacerbation
of UIP, histologic sections show DAD superimposed on a background of
UIP; the precise etiology for acute exacerbation of UIP is not known (which
parallels the fact that the etiology of UIP itself is generally unknown).
2. Non-specific interstitial pneumonitis (NSIP} is the second most common form
of idiopathic interstitial pneumonitis. As with UIP, patients tend to be mid-
dle aged or older adults. Many patients have underlying CIDs such as
rheumatoid arthritis or lupus. Pulmonary function tests show a restrictive
pattern. CT scans disclose significant differences from those in UIP; hon-
eycombing is rarely a prominent feature in NSIP, but ground glass opac-
ities, linear opacities, and small nodular infiltrates are frequently present
(e-Fig. 8.4).
Microscopically, the process has a more homogeneous appearance than
UIP. Biopsies usually lack subpleural accentuation, but instead show more
uniform involvement of both central and peripheral parts of the lobule. The
process is generally more cellular than UIP with a mixed interstitial acute
and chronic inflammatory infiltrate. Cases may show features of organizing
pneumonia or BOOP, and small non-descript granulomas may be seen in
some cases. Honeycombing, if seen on microscopic sections, is usually more
focal and should not be a dominant pattern. The forgoing findings are char-
acteristic of the cellular or mixed patterns of NSIP; since they bear signifi-
cant resemblance to processes such as chronic hypersensitivity pneumonia
or unresolved or slowly resolving organizing pneumonia, it is not clear that
cases labeled as NSIP do not represent some unusual variants of the latter
groups. A third pattern of NSIP, namely the sclerotic variant, consists of
hyaline-like interstitial fibrosis. Although the full characterization of NSIP
is still evolving, the one unifying feature of NSIP is that the patients survive
longer, and with less disability than those patients whose biopsies show UIP.
3. Desquamative interstitial pneumonia and related lesions (DIP} is a rare form
of interstitial pneumonia characterized by abundant macrophage exudates
that fill the alveolar spaces. In most patients, this process is related to
cigarette smoking, although rare examples have been reported in non-
smokers; most patients are middle aged to older adults. Radiographic stud-
ies are dominated by ground glass opacities reflective of the filling of alveo-
lar spaces, and microscopic sections feature dramatic filling of the alveolar
spaces by macrophages, which tend to be light brown due to smoking-
related pigment (e-Fig. 8.5). DIP is spatially homogeneous in that virtually
any microscopic field from involved lung will show identical features. The
alveolar walls may be thin and delicate, or may show some mild inac-
tive hyaline fibrosis. Honeycomb changes are rare. DIP has an excellent
prognosis; in cases related to cigarette use, the primary therapy is smoking
cessation.
Less dramatic findings along the same spectrum of smoking-related
changes include respiratory bronchiolitis (RB) and respiratory bronchiolitis
associated interstitial lung disease (RB-ILD). RB shows similar macrophages
as in DIP, but limited to the lumen of terminal airways; it is often most

prominent in the upper lobes and is thought to be the precursor of cen-
triacinar emphysema (e-Fig. 8.6). RB-ILD is similar with the addition of
mild interstitial fibrosis surrounding the terminal bronchioles.
4. Lymphoid interstitial pneumonitis (LIP} is another rare idiopathic form of
interstitial pneumonitis. LIP can occur in patients of any age. When LIP
occurs in children, it is often a harbinger of HIV infection; similarly,
LIP has been described in immunocompromised patients as a manifes-
tation of EBV infection. Some cases of UP are related to Sjogren syn-
drome. Chest radiographs demonstrate infiltrates of a variety of patterns.
Microscopically, there is diffuse expansion of the pulmonary interstitium by
a mixed inflammatory cell infiltrate that includes small lymphocytes, plasma
cells, germinal centers, and histiocytes. There may be some associated inter-
stitial fibrosis. Many cases previously described as LIP likely represent
examples of pulmonary MALToma or related neoplasms. Immunostains
should show a mixed pattern of CD3-positive T-cells and CD20-positive
B-cells. In cases of suspected LIP, flow cytometry or molecular studies to
assess clonality are very useful to exclude lymphoma.
5. Giant cell interstitial pneumonitis is very rare. Most cases are now known
to represent a reaction to various heavy metals. Consequently, optimum
diagnosis is made by the combination of accurate history and spectropho-
tometric analysis of lung tissue.
6. Smoking-related interstitial fibrosis (SRIF} is a recent addition to list of
smoking-related lung diseases (Hum Pathol. 2010;41:316-325). SRIF is
often found in lobectomy specimens of resected lung carcinomas, in patients
with no pre-operative clinical symptoms or imaging diagnosis of interstitial
lung disease. The fibrosis typically has a hyaline or sclerotic appearance that
somewhat resembles the sclerotic form of NSIP. Also, the fibrosis is usually
superimposed on significant emphysema.
c. Other non-infectious, non-neoplastic pulmonary processes
1. Connective tissue disorders. Lung involvement in CTDs is extremely com-
mon and variable. Prominent patterns of involvement for selected disorders
are summarized in Table 8.3. There is significant overlap among these enti-
ties and assignment to a specific entity should not be made on the basis of
pulmonary findings. Also, many of the lung findings in CTDs overlap with
the idiopathic interstitial lung diseases.
2. Drug-induced pulmonary changes. Prominent pathologic findings associated
with drug reactions are listed in Table 8.4. It is clear from this list that
drug reactions can mimic virtually any non-neoplastic condition, which
emphasizes the need for accurate history and correlation with the clinical
setting.
a. Amiodarone. Perhaps 5%-10% of patients treated with amiodarone
experience a pulmonary complication. The drug inhibits phospholipase,
so the hallmark of exposure is accumulation of phospholipids, which
in the lung manifests as accumulations of foamy macrophages (e-Fig.
8.7); note that accumulation of foamy macrophages is seen in almost
all patients on the drug to varying degrees, and does not by itself indi-
cate toxicity. Toxicity occurs as early as 1 month after initiation of ther-
apy, with an average of 10-12 months, and is associated with higher
doses. Patients present with a variety of symptoms, such as cough, dys-
pnea, chest pain, feve.t;, and myalgias. Radiographs can show a mixture
of airspace infiltrates, interstitial disease, or even isolated collections.
Microscopically, toxicity has several manifestations, including a cellular
chronic interstitial pneumonitis characterized by chronic inflammation
in the interstitium, pneumocyte hyperplasia, and fibrosis; foamy alveolar
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produce peripheral infiltrates that simulate eosinophilic pneumonia, or

granulomatous vascular impingement that can simulate veno-occlusive dis-
ease. End-stage cases often are dominated by the presence of apical bul-
lous disease; the granulomas at this stage may be largely "burnt-out" and
replaced by hyalinized fibrous tissue that tracks along the lymphatic routes.
The differential diagnosis of sarcoid is always granulomatous infection,
anditisimportanttonote that up to 10%-15% of biopsies with granulomas
and negative special stains are culture-positive. Infection should always be
suspected if the granulomas are necrotizing. The differential diagnosis also
includes a drug reaction, berylliosis, and aluminum exposure.
5. Pulmonary eosinophilic granuloma (also called Langerhans cell histiocytosis,
or histiocytosis X) is a disease of adults, with most cases presenting in the
third and fourth decades of life. There is a history of cigarette smoking in
almost 90% of cases, indicating that this disease should be considered as
another facet of smoking-induced lung disease. Patients with pulmonary
eosinophilic granuloma (EG) have disease limited to lung; although some
are asymptomatic, most complain of cough, dyspnea, fever, or weight loss.
Pneumothorax is another documented presentation ofEG. Radiologic stud-
ies show an upper lobe predominance; cysts and small stellate nodules can
be seen by high-resolution CT scans. Because of the smoking history, there
is often co-existent emphysema, desquamative interstitial pneumitis, RB, or
RB-ILD.
The characteristic histologic picture of EG (e-Fig. 8.10) is a stellate inter-
stitial collection, often near small airways, of eosinophils and Langerhans
cells. Langerhans cells feature a unique convoluted nucleus and a moderate
amount of cytoplasm; some hi-nucleated forms may also be present. An
admixture of pigmented alveolar macrophages ("smoker's macrophages")
is also often present. Langerhans cells can be easily demonstrated by
immunostains for S-100 and CD1a; alveolar macrophages stain for CD68
but not S-100 or CD1a. In contrast to the cellular lesions just described,
resolved or "burnt out" lesions in chronic disease may consist only of hyalin-
ized stellate scars in the upper lung zones; immunostains may highlight a
few Langerhans cells in these scars, or may be completely negative. About
10%-20% of cases progress to fibrosis, but most patients improve with
cessation of smoking; rare patients develop severe pulmonary hyperten-
sion. Eosinophilic pneumonia is one important differential consideration in
small biopsies, but can be easily dismissed by correlation with radiographic
and clinical features; in addition, the macrophages in eosinophilic pneumo-
nia will be negative for S-100 and CD1a. Eosinophilic pleuritis may develop
after pneumothorax, and may raise concern for EG in the lung, but reactive
mesothelial cells are negative for S-100 and CD1a.
6. Lymphangioleiomyomatosis (LAM) is a disease that essentially occurs only in
woman of reproductive age. Although classified as an interstitial disease, it
features preserved lung volumes, unlike most fibrosing diseases. CT scans
show diffuse involvement of the lung by cysts of rather uniform size that
feature some mural thickening around the cystic spaces, a finding that serves
to distinguish LAM from processes such as emphysema. In fact, high res-
olution CT images are so characteristic that the first pathologic specimen
seen in many patients is explanted lungs. Patients present with obstructive
lung symptoms or spontaneous pneumothorax, and also may demonstrate
large chylous pleural effusions. Microscopic sections of LAM (e-Fig. 8.11)
show a proliferation of abnormal smooth muscle-like cells in the lung. In
many cases these cells seem to swirl away from the native smooth muscle of
airways and vessels. Vascular compromise is linked to microhemorrhages,
and so many cases show abundant hemosiderin within the lung. While
LAM is centered in the lung, it also involves lymph nodes in the pulmonary
hilum, mediastinum, and abdomen in most cases. The smooth muscle-like
cells stain with vimentin, desmin, and smooth muscle actin, and also may
express estrogen and progesterone receptor. LAM also shows cross-lineage
staining with melanoma markers including HMB-45 and melan-A. In this
regard, the cells of LAM share the staining attributes of the members of the
PEComa family (including the sugar tumor of lung), renal angiomyolipo-
mas, and some soft tissue tumors (see Chap. 46). LAM also shows overlap
with tuberous sclerosis (TS) in that some TS patients develop an identi-
cal cystic lung disease, and both LAM and TS share an association with
angiomyolipomas in the kidney and elsewhere. The course of the disease is
unpredictable; transplantation has been the only long term option for those
with severe disease.
7. Alveolar proteinosis refers to a peculiar accumulation of intra-alveolar
eosinophilic, granular PAS-positive protein and phospholipid. It was ini-
tially reported as an idiopathic process, but a relation to immune deficiency,
hematologic malignancies, infections, and various exposures is now recog-
nized. Classic exposure-related cases are associated with massive acute silica
exposure, which is believed to poison the alveolar macrophages and thus
inhibit their ability to dear alveolar debris.
Clinically, patients present with slowly progressive alveolar infiltrates
and complain of dyspnea, cough, or sputum production with fever; CT scans
show "crazy-paving" with alveolar infiltrates and septal line thickening. The
diagnosis is often apparent from the milky appearance of lavage fluid; biop-
sies show complete filling of the alveolar spaces by granular eosinophilic
debris that is PAS-positive and diastase-resistant (e-Fig. 8.12). Findings
often include cholesterol clefts (acicular clefts), globular eosinophilic debris,
and macrophages. In most cases the underlying alveolar structure appears
normal, although some chronic cases may eventually show fibrosis. Sec-
ondary infection of the fluid by Nocardia, mycobacteria, and fungi has
been reported. Pneumocystis infection can microscopically mimic alveolar
proteinosis, although the material in alveolar proteinosis lacks the frothy
appearance characteristic of Pneumocystis infection.
D. 11AIIergic" diseases
1. Eosinophilic pneumonia can be divided into acute and chronic forms. The
acute forms include the "simple form" also known as Loeffler syndrome;
this is an acute, self-limited process with fleeting infiltrates and peripheral
blood eosinophilia and is rarely biopsied. The tropical form is usually linked
to filaria infection, and also presents as an acute illness. The chronic form
is more likely to require biopsy for diagnosis.
Chronic eosinophilic pneumonia (CEP) has a variable presentation from
acute illness with fever, dyspnea, and weight loss, to vague respiratory com-
plaints. Many patients have a history of asthma, and laboratory tests reveal
elevated blood IgE and peripheral blood eosinophilia. Chest radiographs
show patchy non-segmental infiltrates, often peripheral, that may cross fis-
sures, a pattern that is sometimes described as the "photographic negative of
pulmonary edema." There are myriad underlying causes; major categories
include drugs (antibiotics such as nitrofurantoin, sulfonamides, penicillins,
anti-inflammatory agents, and chemotherapeutics), fungus (Aspergillus and
Candida), parasites, nickel vapor, and idiopathic cases. Histologic sec-
tions of CEP (e-Fig. 8.13) show an alveolar-filling process consisting of
a mixture of eosinophils and macrophages. Necrosis of eosinophils may be
present, forming an eosinophilic abscess. Charcot-Leyden crystals will also
be present, as well as interstitial and perivascular eosinophils, lymphocytes,
plasma cells, and areas of BOOP.
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is basophilic (e-Fig. 8.20). Epithelial cells, vascular cells, and even stromal
cells can exhibit inclusions. Immunohistochemistry for CMV will decorate
the enlarged cells; electron microscopy demonstrates virions in the inclu-
sions, while a PAS stain with diastase highlights the cytoplasmic deposits.
2. Herpesvirus (HSV). Both HSV types I and ll can induce pneumonitis, and
immunocompromised hosts are more prone to Herpes viral pneumonia.
Clinically, HSV pneumonia may result from extension of upperway disease
with primarily bronchiolar inflammation, or from systemic infection which
presents as multiple small perivascular inflammatory nodules. Three find-
ings are characteristic of HSV: necrosis, individual cells with inclusions (e-
Fig. 8.21) (featuring eosinophilic nuclear inclusions with perinuclear clear-
ing; amphophilic nucleoplasm in single cells may represent early inclusions
or giant cell formation), and herpes viral giant cells (which contain two or
more nuclei with ground glass central nuclear clearing and coarsely gran-
ular, sharply defined nuclear borders; the nuclei are often molded against
one another). Immunohistochemistry using antibodies to HSV decorates
the giant cells and the individual cells with inclusions.
3. Varicella zoster (VZV) is the causative agent for chicken pox and shingles.
Primary infection in healthy adults and immunocompromised children can
result in pneumonia. The underlying lung injury pattern can be either DAD
with hyaline membranes and proteinaceous exudates, or nodular inflamma-
tion with central necrosis that calcifies and persists on chest radiographs.
Biopsies of VZV infections show giant cells similar to those of HSV.
4. Adenovirus infection generally presents with symptoms typical of an upper
respiratory infection; pneumonia develops in a small percentage of healthy
and immunocompromised children and adults. Two patterns of infection
evolve. Some patients develop necrotizing bronchiolitis and pneumonia,
while others respond with DAD with hyaline membranes and exudates.
In adenoviral infections both alveolar lining cells and bronchial epithelial
cells may exhibit blurred and hyperchromatic nuclear chromatin (so-called
smudge cells) (e-Fig. 8.22). The bronchial damage of adenovirus may result
in fibrosis or constricting bronchiolitis.
5. Respiratory syncytial virus (RSV) affects primarily small children and infants;
premature infants and immunocompromised children are particularly prone
to infection. RSV infection exhibits some seasonality, and is more common
in fall and winter. RSV can induce bronchiolitis with symptoms of cough,
wheezing, and respiratory distress. Biopsies show necrotizing bronchiolitis
and/or interstitial pneumonia.
6. Measles virus. With the advent of vaccination, infection by measles virus is
rare, and evolution to pneumonia rarer. Most patients are immunocompro-
mised, and the characteristic skin rash is present. The underlying pathology
is DAD, with associated individual and giant cells with viral inclusions;
the alveolar spaces may contain exudates, and necrosis may be present.
Both eosinophilic intranuclear and cytoplasmic inclusions develop in both
alveolar and vascular lining cells. Measles pneumonia features a distinc-
tive multinucleate giant cell thought to derive from coalescence of type n
pneumocytes, the Warthin-Finkeldey giant cell, which contains up to 60
nuclei.
7. Parainfluenza and influenza generate non-specific patterns, consisting of
varying degrees of DAD, bronchial necrosis, and peribronchial inflamma-
tion.
B. Bacterial infections
1. Mycoplasma induces acute and chronic bronchiolitis, with necrosis and
denudment of bronchial epithelium. The bronchial lumina may contain
acute inflammatory cells admixed with denuded epithelium. Acute and
chronic inflammatory cells often traverse the bronchial wall. Alveolar spaces
may exhibit bronchopneumonia, with BOOP or DAD.
2. Mycobacterial infections are typically grouped into tuberculosis and other
(atypical) mycobacterial infections. The most common stain used to demon-
strate organisms in tissue sections is the Ziehl-Neelsen stain; immunoflu-
orescent (auramine-rhodamine) and immunohistochemical stains can also
be used to identify mycobacteria. PCR based genetic methods can also be
used to detect (and speciate) the organism in tissue sections.
a. Tuberculosis (TB). Multidrug resistance has emerged in this organism and
consequently the pathologic spectrum of tubercular infection is expand-
ing. The causative organism is Mycobacterium tuberculosis. which is
transmitted via inhalation of organisms. Histologically, the tubercular
granuloma is a classic palisaded necrotizing granuloma. Granulomas
may caseate and coalesce, creating nodules with central necrosis, or even
cavitary masses. Organisms may be found in multinucleate giant cells or
at the periphery of necrosis (e-Fig. 8.23). Lymph nodal involvement may
be present. Miliary tuberculosis is unlikely to be sampled in biopsy or
resection specimens.
b. Atypical mycobacterial infection. Mycobacterium avium intracellulare
(MAl) is the most common of the atypical mycobacterial infections, and
is more common among immunocompromised patients where it presents
with non-specific fever and malaise. Radiographs show diffuse or patchy
infiltrates. Histologic findings vary from more typical non-necrotizing
punctuate granulomas, to necrotizing granulomatous pneumonia, to dif-
fuse pneumonitis (in which abundant pneumocytes or interstitial cells
contain abundant organisms by acid fast stains).
C. Fungal Infections
1. Candida infection arises in several contexts. Mucocutaneous candidiasis can
arise in immunocompetent adults, but arises more frequently in immuno-
compromised patients; the trachea or bronchial tree can have plaques of
fungus admixed with desquamated cells and neutrophils. Impaired host
defenses potentiate invasive disease; in the lungs, vascular invasion results in
hemorrhagic necrosis. Direct inoculation into the bloodstream from iatro-
genic sources (catheters, surgery) or other inoculation (drug abuse) results in
disseminated disease. Transplant patients can develop infection at the sites
of anastomosis. C. albicans is the most prevalent species; other species more
often infect compromised hosts. All species exhibit the same basic morphol-
ogy: non-branching, aseptate pseudohyphae forming "box-car" like chains
of cells; yeast forms bud from pseudohyphae (e-Fig. 8.24). Yeast forms are
visible on H&E, PAS, or silver stains.
2. Mucormycosis. Several members of the Phycomycetes result in clinically and
morphologically identical disease, including Mucor, Absidia. and Rhizopus.
among others. Almost all cases occur in the setting of diabetic ketoacidosis
(sinonasal or rhinocerebral disease) or immunosuppression from hemato-
logic malignancies. Lung involvement may be the primary focus, or develop
secondary to head and neck disease. Lung lesions are typified by hemor-
rhagic pneumonia; fungal thrombi with distal infarction are often present.
The dual circulation of the lung (bronchial and pulmonary arterial systems)
results in perfusion of infarcted areas with resulting hemorrhagic necrosis,
associated with varying amounts of inflammation. A key to recognition of
the infection is identification of wide, ribbon-like non-septate hyphae with
irregular wide angle branching. The pseudohyphae are often described as
"empty" and stain poorly with most special stains.
3. Aspergillus species cause a wide spectrum of disease, dependent on both
host immune status and site of growth. Colonization with Aspergillus can
induce an allergic response, including ABPA, sinusitis, and hypersensitiv-

ity pneumonitis; Aspergillus can also colonize mucus or grow in a pre-
existing cavity, and sinus or cavitary lung lesions both can harbor fungus
balls. Transplant anastomoses are also prone to colonization by Aspergillus
species (e-Fig. 8.25 ). The characteristic lung lesion is the target lesion with a
sharply delineated hemorrhagic borde.r; which reflects the fact that the fun-
gus is frequently vasoinvasive and produces hemorrhagic infarcts. In well-
preserved areas, organisms have relatively uniform septa, which are thinner
than those of Mucor. The hyphae branch at "'45 degrees; the reproductive
form (fruiting body) is only rarely seen in tissues. Degenerate hyphae can be
mistaken for Mucor.. with empty or dilated forms; acute angle branching,
occasional septa, and more intact forms indicate the correct diagnosis.
4. Cryptococcus. The most common pathogen of this genus is C. neoformans.
which is usually an opportunistic infection but rarely also infects normal
hosts after massive exposure. The common portal of entry is the lungs;
patients remain virtually asymptomatic while the fungus spreads to other
sites. The organism is present in the lung as "naked masses of organisms," to
intracellular forms resembling those of histoplasmosis, to granulomas with
surrounding fibrosis (so-called cryptococcomas) (e-Fig. 8.26). The organ-
isms are much larger than Candida or Histoplasma with an average diame-
ter of 4-10 f.L; only yeast forms are found in tissue. The diameter of crypto-
coccal forms varies in large part with capsule thickness, a function of host
immune status, and mucicarmine stains the capsule strongly (generally con-
sidered a diagnostic feature). Capsule-deficient forms are common in cancer
or AIDS patients, and may be much more difficult to diagnose definitively.
5. Blastomycosis is generally seen in the middle of the United States. Infection
usually involves lungs and skin; spore inhalation is the mode of transmis-
sion for virtually all cases. Pulmonary blastomycosis takes several forms; the
most common is a solitary focus of infection with variable associated lymph
nodal disease, which usually heals and leaves a fibrous scar. Progressive dis-
ease is less common, in which infection spreads throughout the lung as mil-
iary foci which range from neutrophil-rich abscesses to tubercle-like gran-
ulomas. The causative agent maintains a variably sized yeast form in tissue
ranging from 5 to 25 f.t, and has a thick, refractile, double-contoured wall;
unlike Cryptococcus. this wall is negative or very weakly mucin-positive
(e-Fig. 8.27).
6. Histoplasmosis. In the United States, infection is usually caused by H. capsu-
latum from bird or bat droppings. In tissue, the fungus reverts to a primitive
yeast, 2-5 f.t in diameter with occasional unequal budding. Primary histo-
plasmosis produces a mild, self-limited febrile illness in most cases, with
hyalinized granulomas the result (e-Fig. 8.28) but can result in a progres-
sive, disseminated, fatal disease. Cases with active disease can also produce
a granulation-tissue like pattern, with vague granulomas. The organism can
also induce secondary scarring forms of inflammation, such as sclerosing
mediastinitis with calcified granulomas in which organisms are often not
identified.
7. Coccidiomycosis is caused by Coccidioides immitis. a soil borne saprophyte
typically found in the southwestern United States. Patients typically have
travel histories to that region, and present with an acute febrile illness (some
infections are asymptomatic). Infection develops in both immunocompetent
and immunocompromised individuals. The inhaled organisms transform
into spherules, thick walled sacs 6o-80 f.t in diameter containing multiple
endospores (e-Fig. 8.29); reproduction in tissue results from rupture of the
spherule with release of endospores. Microscopically, the organisms create a
nodule, typically with non-caseating granuloma formation; cavitation may

occur. Lymph nodal involvement may be present and disseminated infection
may result. The organisms can be demonstrated with GMS or PAS stains.
8. Pneumocystis carinii pneumonia {PCP). Pneumocystic carinii was formerly
considered a protozoan, but genetic analysis suggests that it is best classi-
fied as a fungus. Infection is generally seen among immunocompromised
patients. The chest radiograph classically shows diffuse infiltrates, which
correspond to the diffuse alveolar infiltrates that are almost diagnostic
microscopically (e-Fig. 8.30). Cytologically, the infiltrate exfoliates in lavage
specimens as alveolar casts. PCP can induce interstitial pneumonitis or gran-
ulomatous inflammation; these variant forms are often seen in chronic dis-
ease or with partially treated disease, may progress to cavitating or cystic
disease in the upper lobes, and are associated with extra-pulmonary dissem-
inated disease. The cysts stain well with GMS in most cases; in degenerate
cases, immunostains may be of some help. The organism has a helmet or
cup shape, often referred to as a "dented ping-pong ball."
D. Parasite Infections
1. Dirofilariasis. Dirofilaria are nematodes, the most common pathogen of
which is D. immitis, the common dog heartworm. While this parasite does
not have a life cycle in the human host, occasional infection can develop
with adult nematodes via transmission by a mosquito bite. Infection can
result in non-caseating granulomas presenting as a nodule in the lung (or
other organ). Cross sections of the organism's refractile cuticle (10-14 /.L in
greatest diameter) may be seen in histologic sections. Endovascular thrombi
with organisms have been reported as well.
V. SELECTED PNEUMOCONIOSES
A. Asbestosis. Asbestosis is an interstitial lung disease caused by asbestos. It usu-
ally occurs in workers heavily exposed, for a prolonged period of time. There
is often a long latency period, usually at least 15 years, between exposure and
the disease. Mild disease shows no symptoms; with increasing severity, patients
complain of dyspnea, dry cough, weight loss, and chest pain. Radiographic
findings are characterized by small irregular opacities, most prominent in the
bases of the lung. The American Thoracic Society has defined six clinical crite-
ria a clinical diagnosis of asbestosis, including exposure, a latency period, rales,
decreased lung volumes and DLCO by PFTs, and chest radiograph infiltrates.
Thankfully, pathologic criteria include just two required findings: the presence
of peribronchiolar fibrosis (fibroelastosis), and associated asbestos bodies (e-
Fig. 8.31). Grading of asbestosis can be performed: grade 1: fibrosis confined
to respiratory bronchioles; grade 2: fibrosis involves alveolar ducts, or two
tiers of alveoli; grade 3: fibrosis involves all alveoli between two bronchioles;
grade 4: honeycombing. The lungs show a high fiber burden: 98%-99% of
cases have at least 2000 asbestos bodies per gram of wet lung tissue; most have
10,000 to 100,000 or more asbestos bodies per gram of wet lung tissue which
correlates with at least several asbestos bodies per tissue slide in the majority
of cases (iron stains often help to demonstrate asbestos bodies). If pathologic
examination of biopsy or lung resections suggests asbestosis, or if there is clin-
ical suspicion for asbestosis, it is prudent to set tissue aside for fiber analysis
(fiber studies on either fresh lung tissue or formalin fixed and embedded tissue),
although it is not required for diagnosis in most cases. Classically, asbestosis is
required to attribute pulmonary carcinomas to asbestos exposure; asbestosis
plus smoking increases the risk for lung carcinomas by a multiplicative factor,
to perhaps SOX non-smoking, non-asbestotic controls. Pleural plaques and
mesothelioma, which are also asbestos-related pleuropulmonary lesions, are
discussed in the chapter on serosal membranes (Chap. 11 ).
B. Silicosis. Silicosis is produced by silica deposition in the lung. Occupations at

risk include sand-blasting, grinding, mining, plastering, and masonry, among
many others. Silicosis typically produces infiltrates in the mid-lung zones, in
contrast to other types of pulmonary fibrosis. Lymph nodes in the chest will
also show characteristic "egg-shell" calcifications on X-ray. The characteristic
lung lesion is the silica nodule; nodules have a lymphangitic distribution, so are
seen along the bronchovascular tree and in the pleura (e-Fig. 8.32). Early nod-
ules are cellular and composed of a swirling collection of fibrohistiocytic cells;
birefringent particles can be seen in these nodules with polarized microscopy.
Older lesions become progressively hyalinized; they may coalesce to form irreg-
ular masses that can mimic pulmonary neoplasms. The relationship of silicosis
to the development of pulmonary neoplasms is debated, but appears to be a
small risk, if present at all. Nodules may also include other material such as
iron or carbon, in which case a diagnosis of mixed dust fibrosis is appropriate.
VI. PULMONARY TRANSPLANTATION
A. Gross processing of transplant specimens. Pulmonary transplantation is now a
well established therapy for various end-stage lung diseases including emphy-
sema/chronic obstructive pulmonary disease, cystic fibrosis, LAM, pulmonary
hypertension, and pulmonary fibrosis of various causes. Examination of the
explanted, native lungs should include thorough documentation of the under-
lying disease process. A minimum of one section per lobe should be submitted
for diffuse processes, as well as sections of the hilar lymph nodes; cases of pul-
monary hypertension may require additional sections to document plexiform
lesions, and any mass or focal abnormality should be sampled thoroughly.
Examination should be comprehensive enough to exclude foci of malignancy,
or infections which may recur due to the immunocompromise of transplant
recipients.
The allograft is surveyed by transbronchial biopsy, and occasional wedge
biopsies, both at scheduled protocol intervals, as well as in response to changes
in clinical condition. A minimum of five pieces of alveolar tissue is considered
adequate for assessment in this setting. In general, three levels of H&E stained
sections are obtained; some institutions employ protocols with adjunct special
stains for infectious organisms on all biopsies.
B. Microscopic features of transplant biopsies
1. Preservation injury is the first change seen in post-transplant biopsies. This
change reflects ischemic damage that develops in the lung in the interval
between removal from the donor andre-implantation. Preservation injury
produces a picture of classic lung injury that may exhibit features of DAD
or BOOP (e-Fig. 8.33). The appearance sometimes suggests viral infection,
but for biopsies taken in the first 1-2 weeks after transplant, it is generally
too early for opportunistic infections to become manifest. One important
differential diagnosis for early acute graft injury is humoral or hyperacute
rejection (discussed below).
2. Opportunistic infections. Biopsies after the first week or two must be surveyed
for opportunistic infections, the most frequent of which is CMV, the features
which are detailed above (e-Fig. 8-20). Important clues of CMV infection
include interstitial neutrophils and alveolar fibrin exudates, findings which
warrant immunostains for CMV since transplant patients may not always
develop classic inclusions. CMV may produce perivascular inflammation,
a finding that mimics acute rejection (see below). Other viral infections
often seen include Herpes virus (e-Fig. 8.21) and Adenovirus (e-Fig. 8.22),
and a variety of non-specific pneumonitis patterns may actually represent
responses to other viral pathogens. The results of culture, serologic, and
molecular tests for infectious organisms must be integrated with biopsy
data by the transplant clinician.
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history suggests nodules or a mass. Cases usually fall into three general
categories: plasma cell hyperplasia, consisting of cases that show low-grade
findings; polymorphous PTLD, which is comprised of a mixed population of
atypical lymphoid cells; and monomorphous PTLD, consisting of cases that
consist of a monotonous population of high grade, malignant-appearing
cells (e-Fig. 8.37). Monomorphous cases are much less likely to regress
following diminution of immunosuppression as compared with the lower
grade lesions; thus, higher grade cases are likely to require cytotoxic therapy.
In allograft biopsies, if there is a question of PTLD versus a rejection-related
infiltrate, a simple panel of CD3, CD20/CD79a, and EBV immunostains will
provide dichotomous results: the infiltrate in severe rejection or infections
is almost always a CD3 predominant infiltrate, while the infiltrate in PTLD
is positive for CD20 and CD79a, as well as EBV, in the great majority
of cases. Adjunctive studies to indicate clonality may also be of value in
terms of diagnosis, classification, and prognosis. It is important to note
that rare examples of PTLD may have the morphology of Hodgkin disease,
and that late cases of EBV negative lymphoid neoplasms (4 or 5 years after
transplant) which resemble non-Hodgkins lymphomas can also develop.
7. Recurrent disease. With the exception of a few cases of sarcoid, recurrence
of the native lung disease in allografts is not a significant problem. There
are reports of occasional cases of carcinoma arising in transplanted lungs,
as well as in the native lung in the setting of unilateral transplant; these
patients fare poorly.
VII. NEOPLASMS. The WHO classification of lung neoplasms is presented in Table 8.12.
Malignancies of the lung are the most common cause of cancer-related mortality
in the United States. Despite decades of warnings, cigarette smoking remains the
predominant risk factor for the development of pulmonary carcinoma. There has
been speculation that a shift in location and cell type for pulmonary carcinomas
is related to changes in cigarette usage. Some data suggest that filtered cigarettes,
which remove larger tar particles, have allowed carcinogens to penetrate to more
distal parts of the lung and produce peripheral adenocarcinomas instead of cen-
tral squamous carcinomas or small cell carcinomas caused by larger particles.
Changes in tobacco formulation over the years may also have played some role
in these changes. Other risk factors for lung carcinoma, either proven or specula-
tive, include asbestos exposure, radiation, various chemicals, heavy metals, viral
infection, fibrosing lung diseases, immunosuppression, and genetic syndromes.
Outside of these causes, there remain some cases of lung carcinomas for which
there is no clear etiologic factor.
Up to 10% of patients with head and neck carcinomas may harbor a concur-
rent lung primary carcinoma; therefore, the finding of a lung nodule in a head and
neck cancer patient should not automatically be assumed to represent metastatic
disease. Similarly, 2%-5% of lung carcinoma patients present with apparent syn-
chronous lung primary tumors (which may be due to the greatly improved resolu-
tion of CT techniques which can now detect additional small nodules in patients
with a dominant lung masses); histologic sampling of these lesions often shows
alveolar proliferations with varying degrees of atypia, which may represent pre-
cursor lesions in the peripheral lung similar to squamous dysplasia in the more
central airways.
A. Pathologic reporting. The important data to be included in a diagnostic report
of a primary pulmonary neoplasm are summarized in Table 8.13. Generally,
tumor size is assessed by gross examination, and this measurement suffices
unless there is confounding fibrosis or peritumoral pneumonia. The extension
of tumor through the pleura covering the lung (visceral pleura) or into the
pleura lining the chest wall (parietal pleura) should be noted; an elastin stain
can delineate these layers in some cases. Lymphatic and venous invasion should
,, /,~,!I J! fll WltO Hld*lft~l Cluft:atloftO!TUiftOit ol111e ......

llol,pnhlllhlllltl-
~(';lf{l
PIpilary
Cleorcol
Smolleo!l
BI.Miold
-
Slnallt:flll-..
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hl..,...rctnol!lll, mlrod OllbCYI>o

Acllllltad""""",...,.m•
Popllary-mo
Bronclllolollillollr ardnoma
~muc!nou.a
Mudnwt
Mlllacl ratomv~and mudnOUIOf' ..ldcDtmN1D
&1111 llditlocordnotr~~~...th lllll<ti lltOduellon
Faaladonowdn"""'
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Slao""Bif-clnan.t
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U,..co/1-
Uoll!l<ell ,.,~,. cotel""""
Comb!fted larg& oel neuroentfo(:rt\a<::~~~rd"'oma
BI.Miold c:an:lnan.t
~plioopilllolomtol.. Cfthm.t
Cloer col ~!nwna
--
Uoll!l<eii<Orcr.orno~ 11\obd<*l llilen<Cli>G
~ CdJ1:Iir<Jmi>
Pioomorplilc ..,,...,.m,
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Giant oall <::e~a
Cl_,..ft\11
Pu"-Y lilolomo
CiHtlflolt1 """"'
'Pfl>bll-lndd
Alypbl a,...,.ld
SIJfwqfl{itnd liNnIn
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--
hllf\OI:I oY*<Ordnotr~~~
F~>iiii•H'n)ololihollol Clldnorna
Squ.-nova arc!l"'a''W In a!tu

Alypbl adonomllo<allyporplull
0tttuso ldlojlollllc pulm""ry """""'ndo1:11no ...u hyp!lp,g,
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(~
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''I.I II J:! fl I WltO HlttlliolfC~I C....Hicllloft ol TulftOit ol111e 1.11,. (CIIIIffiNIH.)

Q\ond""""
CW\pn\11 porlbrond!IBI m~bro-lllmour
0111uto pulmOfllryl',lm~ll
lhllomrrdlltV 11\Y'l11bro~ odc1Umou'
~pN ...~-
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a_.t.U:
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..... tpllllollll1:1 . . .
~ .......... poplkma
Eaoplljl!o
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A-...
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MUCO<Il.atond adenM>O
f'loomorglje odei'Jolno
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MolltlollMO lkollll!nDhomo of Che MALT"""
L,..pi\Gnl_ .,...,_Ia
DI!I'IM la,...lkool "'"PhM>O
LlnJIIIIN,. coil hiiiSi>qiDSII
... ~ ....... tl. .
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~homiiii&IM>o
O.r <:611:l.l"n0r
Gttm 011111 tum.:R
~~mllure
Inmltne
Oll>or..., .... tl.mot>
lrrtn~pul'noiiiJIIdl)momo
---MolafiOitlll
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lp: W\C Preoo; 2004. U&od Mfi pe-.
IIIMtt-. - .
'lll)m<nlllld-
be D.O!Cd ~!dy. iht «<llllt of cht; d!jl.pn. of~ <lbould be liad,

..-.! urr obtt~ in chc «dj-tla.ug ehollLI be <letcribcd. iht AjCC
poul>alosic ""'&i"'! ollw>a <:a!Cinamo.c upon.u.l in Tahle 8.14.
ID chc p.ul, dH:zon ofDDOHIIII&ll cell bq aarcin.oma.o (SO.C.) wularw:lT
iDdepmclall of bialogy. H""""""' """"""'~ill the a...r.PJ' -to
blotolo,llc clo.oo!ticat!OillmpottaAL 'I'hcnpy wltlr. pcmu......thu be= dc:m..,.
-ted to be **- in pulmoaaty ~aDd ~
12a I SECTION 11. THORAX
'' 1.I'! I J'! f II DIIIIJ!Odc r.tura To Be Reptllted wltll ~ C.CII-. ..._.._

• TUmorttD& • Melllllt•
• Grade • Aooocla1od ~lbla In tlor>N~Ia.ollo Lun1
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• PIMnlln-
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Cl'l••a• urc I 12•
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1X
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m...,..nt .... ,., "'l'//lln or tnodllal WMI!Inp bit not "'""llzod 1>J
1m,..« l:toh<iloooo!11'
TO N o - at roimii)IM>•>r
1k C.!d>oruln d:u
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-.u piMn, - u t Iron""""""'I : -tllnvulon """'
'"""*""11Nn t110 lol8' b<"onl:llua O.e., not In IIH> meln bfml:tlul)'
n. 111m..- 2.tm or lao>lh ·-dmonolorl
nb 11im«mor&thon2<m but$em0flouln _ _ d _
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porlalol ploural (PLJ) <hool wall ~clHina>Uporlor oulcut!Lmor>),

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may be~.., mymid ~ ....t ,.,...orulocrino
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important EGFR mutations; the clinical value of rigorous testing for

more uncommon mutations is currently less clear.
Crizotinib is another tyrosine kinase inhibitor that targets lung ade-
nocarcinomas harboring the EML4-ALK translocation. This transloca-
tion tends to occur in tumors with mucinous or signet ring histology, but
can occur more rarely in other histotypes. Overall, 4%-5% of lung ade-
nocarcinomas harbor this translocation, and detection by FISH analysis
is the current methodology of choice.
K -ras mutations are also found in a subset of adenocarcinomas. It has
been shown that K-ras, EGFR, and ALK mutations are essentially mutu-
ally exclusive, and thus some experts advocate the following approach:
Initial genetic analysis should consist of testing for K-ras mutations,
which can be easily and rapidly accomplished. If a K-ras mutation is
found, testing is essentially complete; if K-ras is wild type, EGFR testing
should be performed, since EGFR mutations are more prevalent than
ALK mutations. ALK testing can be performed in the K-ras negative,
EGFR negative cases. A number of other targets for directed therapy
are currently under investigation (such as COX-2, BRAF, met, IGF), but
at this time as there is no accepted therapy currently tied to these tar-
gets and such testing is, therefore, not part of routine clinical analysis
of lung adenocarcinomas. Similarly, multi-gene expression assays and
whole genome sequencing are under investigation and may in the near
future provide additional prognostic and therapeutic information, but
are not currently part of routine clinical analysis.
b. BAC is a specialized form of pulmonary adenocarcinoma. With the rel-
ative and absolute increase of peripheral lung carcinomas, as discussed
above, an increasing number of primary lung cancers present as ground
glass opacities on chest imaging. Clinicians and radiologists often charac-
terize these as BAC, but the morphologic diagnosis of BAC is somewhat
rare because the restrictive WHO guidelines demand that BAC maintain
an alveolar architecture. WHO criteria specify that BAC grow primarily
in a lepidic pattern; cases with any significant component of destruc-
tive invasion, as indicated by desmoplasia, should be classified as inva-
sive adenocarcinomas. Pure BAC, therefore, is rare; it develops relatively
more often in women and in non-smokers. It is much more common as
a peripheral component of an invasive adenocarcinoma, in which case
it is best reported as adenocarcinoma with bronchioloalveolar features.
Emerging thought suggests that the degree of invasion in peripheral ade-
nocarcinomas is an important feature. Tumors with no invasive compo-
nent are "pure BACs," and are analogous to in situ carcinoma. Cases
with a large BAC component and 5 mm or less of invasion also have an
excellent prognosis, while cases with >5 mm of invasive tumor behave
closer to classical invasive adenocarcinomas. Thus, reporting of the total
tumor size, and the constituent size of BAC and invasive components is
suggested.
BAC may present as a solitary peripheral nodule, multiple nodules,
with lobar consolidation that mimics pneumonia, or rarely with diffuse
involvement of the entire lung. BACs are well-differentiated neoplasms.
The tumor cell nuclei are often only modestly atypical, but are abnormal
by their monotonous nature, and may be differentiated from reactive
alveolar processes in part by this cellular uniformity. So-called sclerosing
BAC may have some expansion of the alveolar interstitium by fibrosis
without invasion, but many postulated examples of this tumor are actu-
ally invasive adenocarcinoma. Because BAC grows very slowly, there
may be a central elastotic scar that is degenerative or involutional and
1X2 I SECTION lit THORAX
,, 1,1,! I u Ul A\JIIIQII Nweolltl.,.,llll ~ llloftchloiMiveol.v tvd-•

IIJ!IIt~~l .llllul•lr.t!M .,~~~~a Inn diiii1111Md1 r 1:11 rei•-
SliD Smol uoual~ <5 mm
Cll'llcol~ Oftoll ~. maybe fouhd
I n - d.,...tt "nanNI'
_,~,.In - tillllliJj"'
Mldy otypaol, but poilmOII>h""'
and cavitation is a common radiologic finding. Squamous dysplasia and

carcinoma in situ not infrequently accompany sec and may be the only
material retrieved by endoscopic biopsy. The histology of SCC does not dif-
fer significantly from sec that occurs at other body sites; both keratinizing
and non-keratinizing variants occur, as well as basaloid and clear cell forms
(e-Fig. 8.43).
Poorly differentiated SCC should not be confused with small cell car-
cinoma; sec lacks the individual cell necrosis, crush artifact, and nuclear
molding seen in small cell carcinoma. The distinction between small cell
carcinoma and sec can be facilitated by immunostains for p63; sec gen-
erally reacts with antibodies to p63 while small cell carcinoma does not.
sec also seems to be somewhat less prone to nodal metastasis.
3. Large cell carcinoma is an undifferentiated epithelial neoplasm. Neither
squamous differentiation (keratin, cytoplasmic bridges) nor glandular fea-
tures (mucin, acinar formation) are seen by light microscopy, although ultra-
structural studies show both squamous and glandular elements. Exhaustive
and expensive attempts to subclassify undifferentiated large cell carcinoma
into a specific diagnostic category offers no clinical utility; the finding of
a tumor so poorly differentiated that standard sampling and diagnostic
techniques cannot demonstrate clear cut squamous or glandular differenti-
ation provides prognostic information in itself. These tumors are generally
aggressive. Large randomized clinical trials have shown that demonstration
of neuroendocrine markers in non-small neoplasms in general, and large cell
neoplasms in particular (outside of the defined neuroendocrine tumor types
delineated below), does not denote responsiveness to small-cell carcinoma
specific chemotherapeutic regimens.
4. Giant cell carcinoma is a unique variant of large cell carcinoma. The tumor
grows as a large bulky peripheral mass, and often invades the pleura and
chest wall. Giant cell carcinoma is composed of large epithelioid cells, many
of which are multi-nucleated (e-Fig. 8.44 ). The cells grow in large sheets and
are loosely cohesive. An intense acute and chronic inflammatory cell infil-
trate almost always permeates giant cell carcinoma. A spindle cell compo-
nent often mixes with the giant cell pattern, producing what has sometimes
been designated as pleomorphic carcinoma. Unique aspects of giant cell
carcinoma include a propensity to metastasize to abdominal sites including
the small bowel, and an association with leukemoid reaction. Immunore-
activity for epithelial markers including cytokeratin and EMA are gener-
ally maintained; documentation of these markers in a giant cell carcinoma
distinguishes this tumor from such possible mimics as malignant fibrous
histiocytoma, anaplastic large cell lymphoma, or choriocarcinoma.
5. Clear cell carcinoma does not represent a distinct primary form of pul-
monary carcinoma; rather, lung carcinomas with cytoplasmic dearing rep-
resent variants of other forms of non-small cell carcinoma. Clearing has
been reported in 25% or more of all pulmonary adenocarcinomas, squa-
mous carcinomas, and even large cell carcinomas (e-Fig. 8.45). Of course,
a pulmonary clear cell neoplasm should raise concern for a metastatic clear
cell carcinoma, typically of renal origin, as well as the rare pulmonary clear
cell tumor (see below).
6. Adenosquamous carcinoma is an uncommon variant of pulmonary carci-
noma. These tumors are generally associated with a smoking history. The
diagnosis should be reserved for tumors in which there is dearly recog-
nizable differentiation into squamous (intercellular bridges, and possibly
keratinization) and glandular elements (acini). Solid adenocarcinomas with
minimal squamoid differentiation or foci of cytoplasmic eosinophilia should
not be interpreted as adenosquamous carcinoma; similarly, sec s that
114 I SECTION lit THORAX.
WilD
Tlllllroalt,o
Clrdnotl
.. ...
~i(.1:!1j:l(il l'<liltura of Put_,.,- .... IOII*>cdllllleoplu!M
,,
AI tJii'RIIII:IN
GraoolNEC
Colli Slit
Moclum
toler•
_,.
llllrr:IMr
Non It
....
<2/hpf N..,.
lollallplll
llacttollt hlaltlal
'-
~I Gred&2NEC Mtdum Mlil 2-lMIDI Focol tntorrnodlllo
Co- to flllll'
Lilli" Col Nou· G-3NEC, Llrp MI!Ud >!MIDI Exllns!\lre Hfall
IQM'Idoc:tfne lMIIt Coil
c.rmoma tJpe
Small Col Nou· Graoo3NEC, Smllll Hl!;l Eans!'te Htpl
"""rlald
I'Ofllldocdno sm•eon
Cormoma typo
associated with the carcinoid syndrome, although cases of Cushing syn-

drome have been reported due to ACfH release.
Grossly, central carcinoid tumors usually present as yellow-tan poly-
poid intraluminal masses covered by normal respiratory tract mucosa; they
almost always measure <5 em. Microscopic sections show a variety of
architectural patterns including trabecula, rosettes, papillary formations,
and areas of solid growth (e-Fig. 8.48). The tumors have vascular stroma,
which can also feature elements including amyloid and bone. The individual
cells tend to have moderate to abundant cytoplasm, which is often granular
and eosinophilic. The tumor cells have nuclei that are regulat; and round to
oval; the chromatin is granulat; and often referred to as having a "salt and
pepper" character. Mitotic activity should be essentially absent; the most
recent WHO standard is ~2 mitosis per 10 high power fields. Necrosis
should be absent. Peripheral carcinoid tumors are morphologically identi-
cal to central tumors, although peripheral carcinoid tumors often exhibit a
spindled morphology. Tumors with a peripheral location and spindled pat-
tern should not be designated automatically as atypical carcinoid tumors
(see below).
Carcinoid tumors (which can also be referred to as well differentiated
neuroendocrine carcinomas) contain abundant neurosecretory granules,
and show strong expression of chromogranin and synaptophysin; they are
also immunopositive for epithelial markers such as cytokeratin. Carcinoid
tumors have a low malignant potential. Only about 5% of patients present
with lymph node metastases; <5% of cases develop distant metastases dis-
ease with spread to the livet; brain, bones, and skin. Long-term survival is
in excess of 95%.
a. Carcinoid tumorlets are microscopic proliferations (4 mm or less in size)
that are otherwise histologically and immunophenotypically identical to
carcinoid tumors (e-Fig. 8.49); distinction is based solely on size. Tumor-
lets tend to occur in distal airways, and may be single lesions incidentally
discovered in resection specimens. Rare patients may have hundreds of
these lesions in their distal airways. Multiple carcinoid tumorlets have
some association with chronic airway diseases. Tumorlets have little or
no clinical significance.
b. Paraganglioma features nested cells, a fine fibrovascular stroma, cells
with eosinophilic to granular cytoplasm and some spindling, and reg-
ular nuclei, and so it can be difficult to distinguish paraganglioma from
carcinoid tumor. Although paraganglioma shows immunopositivity for
chromogranin and synaptophysin, it is not reactive with antibodies to
cytokeratin; in addition, the tumor contains S-1 00 positive sustentacular
cells that are not present in carcinoid tumors.
c. Chemodectomas. Multiple pulmonary chemodectomas were once con-
sidered a variant of paraganglioma, but now are known to have no
relationship to paraganglioma. Chemodectomas consist of 1-2 mm stel-
late proliferations that fill the pulmonary interstitium and feature bland
epithelioid cells in a whirling pattern. These tumors share immunore-
activity for EMA and vimentin, and have been labeled as "minute
meningothelial-like nodules." They are of no clinical significance.
3. Atypical carcinoid tumor, better designated as grade 2 neuroendocrine carci-
noma, is more rare than carcinoid tumot; and more evenly divided between
central and peripheral locations (e-Fig. 8.50). Because more tumors are
peripheral, airway-related symptoms are less common. In addition, there
is a closer relationship to cigarette smoking. These tumors tend to be
slightly larger than grade 1 lesions at the time of diagnosis, but it is micro-
scopic features that distinguish grade 2 neuroendocrine carcinoma (atypical
carcinoid) from grade 1 neuroendocrine carcinoma (carcinoid). The tumor

cells tend to be slightly smaller than those of classic carcinoid tumors, and
some modest nuclear pleomorphism is also usually present. Spindling of the
cells is also more common than with grade 1 cases, and mitotic activity is
more frequent; up to 10 mitoses in 10 high power fields is an accepted cri-
terion. Necrosis may be present, but is usually limited. Because the cells are
moderately differentiated, neuroendocrine immunostains tend to be posi-
tive.
Atypical carcinoids show significantly greater malignant potential than
classic carcinoids. About 25% of patients with atypical carcinoid tumor
have lymph nodal metastases at presentation, and approximately 25% of
patients are dead of disease at 5 years after diagnosis.
4. Large cell neuroendocrine carcinoma (large cell NEC) can also be designated
grade 3 neuroendocrine carcinoma. It is an extremely lethal form of lung
carcinoma, and is the most recent variant of neuroendocrine carcinoma to
be described. The demographics of LCNEC are similar to those of other
lung carcinomas; almost all cases have been reported in cigarette smokers.
The vast majority of the tumors are found in the periphery of the lung, and
thus they tend to present with the same sort of non-specific symptoms as
do other lung carcinomas. The gross of appearance of the tumor is similar
to that of other lung carcinomas, and may include necrosis or cavitation
(e-Fig. 8.51).
The microscopic features of LCNEC are quite distinctive. The tumor has
an organoid appearance in which large solid nests of tumor are separated
by scant fibrovascular stroma. In some cases, the tumor seems to fill up the
preexisting alveolar spaces. Tumors may show palisading of tumor cells at
the periphery of the nests, and spindling or rosettes may also be present.
The dominant feature in most cases is extensive necrosis, which may make
up the bulk of the tumor mass. The tumor cells themselves are generally
polygonal and have a significant amount of cytoplasm. LCNEC can be
distinguished from small cell neuroendocrine carcinoma based on several
factors: at lower power microscopy, the nuclei of the tumor do not touch (as
do the nuclei in small cell carcinomas, see below); the nuclear-cytoplasmic
ratio is significantly lower than that of small cell carcinoma; the nuclear
chromatin may be vesicular and nucleoli are often prominent; and finally,
crush artifact and nuclear molding are not usually seen.
Mitotic activity is one of the key features of this tumor; the tumor
should feature at least 10 mitoses per 10 high power fields, although the
actual rate is generally far higher. Combined with the extensive necrosis
described above, it is generally relatively straightforward to separate large
cell NEC from lower grade tumors such as atypical carcinoid. In fact, large
cell NEC probably has greatest overlap with non-small cell carcinomas such
as poorly differentiated squamous carcinomas. Thus, immunostains can be
quite helpful in diagnosis. Large cell NEC shows reliable staining with pan-
cytokeratin antibodies (because of the increased amount of cytoplasm in the
tumor, cytokeratin stains do not show the dot-like pattern that is typical
of small cell carcinoma, but rather show strong circumferential cytoplas-
mic staining). The majority of cases of large cell NEC also show positive
staining for chromogranin, synaptophysin, CD 56, and CD 57, although the
more poorly differentiated nature of LCNEC is reflected in more focal reac-
tivity for these neuroendocrine markers. Large cell NEC often also shows
positive staining for p53.
The prognosis for large cell NEC is quite poor. Although the great major-
ity of patients present with node negative lesions at diagnosis, most patients
rapidly develop recurrent or metastatic disease. The overall survival at five

and 10 years is poor, approximately 20% and 10%, respectively.
5. SCLC is classically described as high grade neuroendocrine carcinoma. The
incidence of SCLC is decreasing, but SCLC still accounts for at least 10%
of all primary lung carcinomas. This tumor type has a very strong associa-
tion with cigarette smoking. At least 95% of patients present with a central
mass comprised of hilar and/or mediastinal adenopathy; the adenopathy
often is larger than any radiographically definable intra-pulmonary mass
(e-Fig. 8.52). The bulky tumor in the mediastinum leads to a variety of clin-
ical presentations, including cough, hemoptysis, lobar collapse, shortness of
breath from pleural effusions, chest pain, hoarseness from recurrent laryn-
geal nerve invasion, or superior vena cava syndrome. A significant number
of patients also present with signs or symptoms referable to distant spread,
such as neurologic symptoms (brain metastases), bone lesions, or symptoms
due to abdominal organ involvement. Paraneoplastic syndromes are com-
mon, including the syndrome of inappropriate secretion of anti-diuretic
hormone (Eaton-Lambert syndrome) and Cushing syndrome (related to
AC1H production).
Since small cell carcinomas are rarely resected, the gross features of the
tumor are seldom seen in the surgical pathology laboratory. These features
include a large central mass that tends to spread along the bronchial tree,
with involvement of hilar lymph nodes often in a contiguous fashion with
the primary tumor. Some cases show an endobronchial lesion; <5% of cases
present as a peripheral pulmonary nodule.
The microscopic appearance of small cell carcinoma varies with the
method of sampling (e-Fig. 8.53). Endobronchial biopsies commonly have
the so-called "oat cell" appearance characterized by small cells (abouttwice
the diameter of a resting lymphocyte) with dark hyperchromatic nuclei.
There is scant to barely visible cytoplasm, and crush artifact and nuclear
molding are prominent; nucleoli are not usually prominent. Individual
apoptotic cells are often seen, and more extensive confluent necrosis may be
present. Mitotic figures are frequent. The cells tend to stream through the
tissue in irregular sheets, although some cases may show rosettes, palisades,
or trabecular growth patterns. Small cell carcinoma often has a slightly dif-
ferent appearance in larger tissue samples, such as resected primary tumors
or lymph node biopsies. The cells are oftentimes slightly larger (up to four
times the diameter of a resting lymphocyte), but still feature dark hyperchro-
matic nuclei, often with nucleoli. Crush artifact and nuclear molding may
be less prominent. Necrosis is often widespread; one well-known feature
of small cell carcinoma with excessive necrosis is the so-called Azzopardi
effect, which is the coating of blood vessel walls by nucleic acid to produce
dark blue ring-like structures in the midst of otherwise eosinophilic necrotic
areas.
Immunostains can be helpful in the diagnosis of small cell carcinoma.
Cytokeratin immunostains often produce a dot-like perinuclear pattern of
positivity due to the small amount of cytoplasm and the condensation
of cytoskeletal elements in the tumor cells. Stains for CD45 can exclude
lymphoma. In cases with classic morphology, this simple panel (cytokeratin
and CD45) is sufficient for the diagnosis of small cell carcinoma. If addi-
tional immunostains are requested to confirm neuroendocrine differentia-
tion, it should be noted that small cell carcinomas have very few neurose-
cretory granules, and hence chromogranin stains are relatively insensitive.
Similarly, synaptophysin and CD57 stains are positive in only approxi-
mately 50%-60% of cases. Stains for NCAM (CD56) are more sensitive,
and stains TfF-1 are also positive in the majority of cases of pulmonary
small cell carcinoma. The cells of small cell carcinoma may also express
CD99, bcl2, p53, and CEA, although these stains generally are not obtained
in a diagnostic context.
Since small cell carcinoma has spread extensively at the time of diagnosis
in most cases, treatment is generally non-surgical. Survival remains poor.
Modem chemotherapy and radiation regimens produce significant disease
remissions in the majority of patients, but relapse within a few months is
common. OverallS year survival is in the range of 5% to 10%.
Neuroendocrine carcinoma may also be mixed with non-small cell car-
cinoma. This combination is rarely if ever seen with low grade lesions, but
is seen in a small minority of cases of large cell NEC and small cell car-
cinoma. Admixed non-small cell components are rarely seen in bronchial
biopsy specimens, but are detected in up to 10% of resected small cell can-
cer cases; this discrepancy is probably related to sampling volume. When
recurrences occur after treatment of small cell carcinoma, a dominant non-
small cell component may be present; it is likely that this shift in phe-
notype represents selective survival of a previously minor admixed non-
small cell component not targeted by treatment directed against small cell
carcmoma.
D. Sarcomatoid carcinoma and related neoplasms. A subset of primary pulmonary
carcinomas have sarcoma-like features. These tumors have been given a vari-
ety of names, including carcinosarcoma and spindle cell carcinoma; sarcoma-
toid carcinoma is the currently preferred designation for all carcinomas with
sarcoma-like features. Patients with sarcomatoid carcinoma have a similar age
and smoking history as those with other forms of pulmonary carcinoma. Sar-
comatoid carcinoma, however, presents in two distinct patterns. Some cases
present as a polypoid intraluminal mass within a large central airway; these
tumors tend to be small, most likely because patients present with airway-
related symptoms early in the course of the tumor's growth. In contrast, other
patients present with a bulky peripheral mass, often with pleural and chest
wall invasion. Microscopic sections show malignant spindled tumor cells (e-
Fig. 8.54) which may transition from areas of non-small cell carcinoma. Var-
ious heterologous elements such as malignant cartilage or osteoid may also
be seen.
The major differential is with true pulmonary sarcomas and sarcomatoid
mesotheliomas. In order to diagnose sarcomatoid carcinoma, the sarcoma-like
tumor must be proven to show some evidence of an epithelial lineage. This
can be accomplished by immunostaining; reactivity for cytokeratins, EMA,
p63, or other generic carcinoma markers such as carcinoembryonic antigen
will be present in the majority of (although not all) examples of sarcomatoid
carcinoma. Electron microscopy, with identification of epithelial features such
as cell junctions, may be quite useful in this context. In many cases, exten-
sive sampling is enough to demonstrate an epithelial component by showing
transition from sarcoma-like areas to classic areas of squamous carcinoma
or adenocarcinoma; the finding of such a transition provides definitive evi-
dence as to the nature of the neoplasm. Differentiation from mesothelioma
may be difficult in that immunophentoypes may overlap; Wf1 and calre-
tinin expression would favor mesothelioma. Clinical presentation as a single
intrapulmonary mass may be a critical feature that favors sarcomatoid carci-
noma, since mesothelioma tends to present as a diffuse pleural neoplasm.
The behavior of sarcomatoid carcinomas varies with the clinical presenta-
tion. Cases presenting as a small intraluminal polypoid airway lesion have a fair
prognosis. In contrast, those cases occurring as large bulky peripheral tumors
have a poor outcome since high stage disease at presentation is common.
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PPB is subclassified on the basis of gross features: predominately cystic (type

1 ), solid and cystic (type 2), or solid (type 3 ) (e-Fig. 8.5 7). When cystic, the
legion is often confused with a variety of benign cystic conditions including
cystic adenomatoid malformation; the misdiagnosis is often discovered when
the lesion recurs. Microscopic sections of the cystic cases show that the cysts are
lined by benign epithelium with an underlying stroma that can have a variety
of appearances, ranging from mature fibroblastic cells to overtly malignant
cells with a sarcomatous appearance. The more solid cases similarly show
sarcomatous-like malignant cells; rhabdomyoblasts and malignant cartilage
may be included. Unlike monophasic and biphasic pulmonary blastoma, PPB
does not include a malignant epithelial component. The majority of patients
with completely excised cystic lesions that are confined to the lung demonstrate
long-term survival; the prognosis is much more guarded for predominantly
solid lesions.
H. Salivary gland tumors. Minor salivary glands are present all along the tracheo-
bronchial tree. Consequently, the airway can be the site of any of the neoplasms
that can occur in the salivary glands. Although rare examples of pleomorphic
adenoma (e-Fig. 8.58), acinic cell tumor (Fechner tumor), and oncocytoma
have been reported, two salivary gland type neoplasms occur with a high
enough frequency to warrant further discussion.
1. Mucoepidermoid carcinoma. Low grade forms of this tumor tend to present
as a polypoid intraluminal mass, and cases have been described in both
adults and children. Histologic sections show a mixture of mucus producing
cells, dear cells, and squamous cells (e-Fig. 8.58). There is minimal mitotic
activity and necrosis is lacking. Many cases have abundant mucus-filled
cystic areas. The tumor is generally indolent; local invasion and local recur-
rence are the primary concerns, and local resection is often curative. The
major differential diagnostic considerations include mucus gland adenoma
(which lacks the locally invasive character of mucoepidermoid carcinoma)
and squamous carcinoma (which tends to show more extensive keratiniza-
tion and less mucus production). High grade forms of mucoepidermoid
carcinoma similar to those described in the salivary glands also occur in the
lung; since such high grade tumors have a prognosis similar to other forms
of non-small cell carcinoma, by convention they are generally considered
to represent adenosquamous carcinomas.
2. Adenoid cystic carcinoma (ACC) also occurs in the tracheobronchial tree. It
is more common in the trachea, but also arises in the larger bronchi. Most
tumors are found in middle aged to older adults, and many patients present
with a long history of airway symptoms such as wheezing; the long history
reflects the indolent nature of these tumors. ACC is usually 2 to 5 em at the
time of diagnosis. The tumor may grossly appear well circumscribed, but
microscopically most cases extend beyond the area of gross disease. The
histology of the tumor in the lung is identical to the analogous tumor of the
salivary glands, with uniform, modestly atypical polygonal cells arranged
in nests, cribriform patterns, and solid sheets (e-Fig. 8.59). Many of the
tumor nests contain characteristic eosinophilic matrix material. As in the
salivary glands, perineural invasion is common. Immunostains are consis-
tent with the proposed myoepithelial origin of this tumor, demonstrating
immunopositivity for cytokeratin, vimentin, actin, and SlOO. Many cases
are CD117 positive (it is not dear whether this finding has any therapeutic
implications). Positive resection margins are common because of the infil-
trative growth pattern; postoperative radiation therapy may help control
incompletely resected tumors for extended periods of time. Intra-pulmonary
metastasis eventually occurs in many cases as a late complication; even
these metastases tend to be slow growing and many patients survive with
metastatic disease for an extended period of time.
I. Sarcomas. Primary sarcomas of the lung are distinctly uncommon. Before con-
cluding that a tumor is a primary pulmonary sarcoma, primary pulmonary sar-
comatoid carcinoma and a sarcoma metastatic to the lung must be excluded. It
is, therefore, important to know details of the clinical history and radiographic
findings before labeling a lung tumor as a primary lung sarcoma. Fibrohistio-
cytic, fibroblastic, smooth muscle, and vascular sarcomas have been reported
as primary lung sarcomas, as have primary neurogenic, osteogenic, and car-
tilaginous sarcomas. Small blue cell tumors of the lung include members of
the Ewing sarcoma/primitive neuroectodermal tumor (EWS/PNET) family and
rhabdomyosarcoma. Lung sarcomas are essentially identical in appearance to
their soft tissue counterparts.
1. Synovial sarcoma is a tumor that has recently been recognized to occur in
both the lung and pleural spaces. It occurs in relatively younger adults than
do primary lung carcinomas. The tumor may show calcifications on radio-
graphs, but otherwise has few defining clinical or radiographic features
(e-Fig. 8.60). Histologic sections show features identical to synovial sarco-
mas of the soft tissue (see Chap. 46). Immunostains are tremendously help-
ful for averting a misdiagnosis; synovial sarcomas of the lung demonstrate
expression of cytokeratin and EMA staining in both the glandular and spin-
dle cell areas, and the tumor cells may also be positive for CD34 and CD99.
The differential diagnosis includes other biphasic pulmonary tumors such
as sarcomatoid carcinoma and biphasic mesothelioma. Molecular demon-
stration of a t(X;18) translocation can be a helpful aid in diagnosis. As with
synovial sarcoma arising in other locations, recurrence or metastasis may
take many years to develop, but the ultimate prognosis is poor, with the
majority of patients eventually dying of the disease.
2. Epithelioid hemangioendothelioma {EH} was originally termed intravascular
bronchioloalveolar tumor. EH presents most commonly in women of young
to middle age. Patients may be asymptomatic, or present with cough or
shortness of breath. Radiology typically shows multiple small pulmonary
nodules. Histologic sections show nodules containing pale-staining, hyaline
to myxoid stroma in which the preexisting alveolar structure is often still
apparent. The neoplastic cells are present within this stoma; they are small
and epithelioid and often contain an intracytoplasmic lumen which repre-
sents a primitive attempt at vessel formation (e-Fig. 8.61). Ultrastructural
studies show endothelial cell features such as Weibel-Palade bodies; the
cells also react with antibodies for vascular markers, including CD31 and
CD34. Although the tumor grows slowly, most patients eventually develop
progressive disease with respiratory failure.
3. Kaposi sarcoma (KS} is another vascular tumor that may involve the lung.
Pulmonary involvement occurs almost exclusively in the setting of immuno-
compromise, predominantly HIV/AIDS, and rarely in solid organ trans-
plant recipients. Pulmonary KS most often co-exists with cutaneous disease.
Radiographic studies show nodular infiltrates and a pleural effusion may
be present; symptoms include cough, fever, and hemoptysis. KS spreads via
lymphovascular routes and thus is seen along the bronchial tree, along pul-
monary vessels, and along the pleural and lobular septa. Histology shows
typical lesions of KS, with spindled cells arranged to form slit-like vascular
spaces, extravasated erythrocytes, and hemosiderin (e-Fig. 8.62). Patients
who have pulmonary KS have a poor prognosis, determined not only by the
response of KS to chemotherapy but also by the course of the underlying
HIV infection.
4. Pulmonary artery sarcoma is another rare but deadly lung sarcoma. Patients
tend to be middle aged or older, and typically present with shortness of
breath or signs of right-sided heart failure. Imaging studies often show
intravascular filling of the pulmonary artery trunk which may be inter-
preted as thromboembolic disease. The tumor may be situated in the main
pulmonary artery trunk or in one or both of the main right and left
artery branches; the sarcoma may extend distally into progressively smaller
branches within the lung. The histologic features of pulmonary artery
sarcoma are variable, including smooth muscle, fibrohistiocytic, endothe-
lial, and even chondroid or osteoid differentiation. The prognosis of pul-
monary artery sarcoma is poor; even in cases with complete resection,
distal recurrences within the ipsilateral lung are the rule. To date, radia-
tion and chemotherapy have not been particularly effective in treating this
sarcoma.
5. Thoracopulmonary small cell tumor, also known as Askin tumor, is now
known to be a member of the EWS/PNET family of neoplasms. This highly
malignant tumor most often occurs in children and young adults and is
typically very large at presentation; it may literally fill an entire hemitho-
rax (e-Fig. 8.63). The exact site of origin is often unclear, although most
tumors probably originate from the chest wall with secondary invasion into
the lung. The tumor consists of a classic small blue cell proliferation, grow-
ing in sheets or rosettes. Necrosis may be prominent. The tumor shows
membrane staining for CD99 as well as various neuroendocrine markers.
Most cases demonstrate a characteristic t(11;22) translocation {see Chap.
46).
J. MISCELLANEOUS NEOPLASMS
1. Pulmonary hamartoma (chondroid hamartoma) is a benign proliferation usu-
ally seen in adults. The tumor usually occurs as a solitary peripheral mass
with a radiographic appearance of a so-called "coin lesion." A minority
of cases involve the more central airways, or even the trachea. Chondroid
hamartomas may be part of a heritable syndrome in some cases; Carney
triad consists of pulmonary hamartomas, gastric stromal tumors, and extra-
adrenal pheochromocytomas. Grossly, the tumor is a well circumscribed,
nodular lesion (e-Fig. 8.64). Histologic sections disclose a mixture of benign
mesenchymal components, including cartilage, mature adipose tissue, and
smooth muscle. Bronchial type epithelium is usually present within the
lesion, although this is thought to represent entrapped tissue rather than a
true component of the proliferation. Radiographic diagnosis of this lesion
can be confidently made based on the presence of adipose tissue and calci-
fications within the cartilaginous component; for this reason, hamartomas
are often not resected.
2. Pulmonary clear cell tumor, or sugar tumor, is a unique pulmonary tumor
composed of cells with clear cytoplasm. The tumor generally arises in adults
and presents as a well circumscribed, nodular mass; most are found inci-
dentally in asymptomatic patients. The tumor usually measures <5 mm in
greatest dimension. Histologic sections show epithelioid cells with bland
nuclei and abundant clear cytoplasm (e-Fig. 8.65). The cells grow either in
nests separated by fine fibrovascular stroma or in a more sheet-like pattern.
PAS stains are positive due to abundant intracellular glycogen, as confirmed
by ultrastructural studies which may also show pre-melanosomes within the
tumor cells. Immunostains show a unique pattern of vimentin positivity, as
well as positivity with melanocyte markers such as HMB-45 and melan-A,
with expression of actin and CD117 as well.
The histogenesis of this tumor has long been debated. Most recently it
has been suggested that the tumor is a member of the family of tumors
known as perivascular epithelioid cell tumors, or PEComas. The most

important neoplasm in the differential diagnosis is metastatic renal cell car-
cinoma; in this regard, it is important to emphasize that pulmonary clear cell
tumors uniformly lack expression of epithelial markers such as cytokeratin
and EMA. The behavior of pulmonary clear cell tumor has generally been
considered to be benign, although rare tumors show malignant behavior.
3. Inflammatory myofibroblastic tumor (IMT) has traditionally been given a vari-
ety of names, including inflammatory pseudotumor and plasma cell granu-
loma. IMT is the most common benign lung tumor in children, but occurs
in adults as well. The most common presentation is as a relatively small
solitary peripheral nodule; IMT also occurs as an endobronchial lesion.
Patients may be asymptomatic, or may present with a variety of systemic
signs and symptoms, including feve.t; anemia, and polyclonal hypergam-
maglobulinemia; systemic manifestations usually resolve with removal of
the tumor. In most cases, the tumor is confined to the lung, although occa-
sional cases may exhibit more aggressive local behavior, including invasion
of mediastinal structures.
Microscopic sections show a proliferation of spindled myofibroblastic
cells with a haphazard pattern of vague fascicles (see Chap. 46) (e-Fig.
8.66). The spindle cells are characteristically bland and mitotic figures are
rare, without abnormal mitotic figures; necrosis is unusual. There is often
a marked inflammatory cell infiltrate in the lesion that may include plasma
cells, lymphocytes with lymphoid follicles, neutrophils, and eosinophils.
The stroma varies from myxoid to densely collagenized and keloid-like.
Immunohistochemical stains show that the spindle cells are positive for
vimentin and smooth muscle actin, and are negative for cytokeratin, CD34,
and desmin. ALK-1 is variably expressed in IMTs of the lung, corresponding
to those cases that harbor a rearranged ALK gene (see Chap. 46). The
plasma cells are polyclonal by light chain studies. The immunophenotype
of IMT can be used to rule out several tumors in the differential diagnosis
such as spindle cell carcinoma, solitary fibrous tumor, and fibrohistiocytic
neoplasms.
IMT of the lung is usually cured by surgery. Recurrence is rare, but
more common if the lesion has invaded adjacent structures at the time of
resection. Extremely unusual cases of malignant transformation to a high
grade fibroblastic or round cell neoplasm have been reported.
It must be emphasized that non-neoplastic inflammatory processes occur
in the lung, which are also capable of producing a mass-like lesion, such
as organizing pneumonia, infarcts, scars, and confluent granulomas. These
processes must always be included in the differential diagnosis of IMT.
K. HEMATOLYMPHOID LESIONS. Hematolymphoid neoplasms and pseudoneo-
plasms can involve the lung, both primarily and as part of systemic disease.
Fresh tissue should be set aside for flow cytometry whenever possible. In cases
for which only fixed tissue is available, a variety of molecular studies including
gene rearrangement analysis can also be used to evaluate donality. By defi-
nition, primary pulmonary lymphomas should not have evidence of disease
outside of the lung or hilar lymph nodes.
1. MALToma. Lymphomas of mucosal associated lymphoid tissue, so called
MALTomas, commonly arise in the lung and constitute the most com-
mon form of primary pulmonary lymphoma. MALTomas for the most part
correspond to marginal zonelymphoma, and many harbor a t(8;11) translo-
cation which may be relatively specific for lung MALTomas. Most cases
occur in adults; there is no specific relationship to infection or autoimmune
disorders. Pulmonary MALToma may have a nodular or diffuse appear-
ance, with an infiltrate comprised of small lymphocytes, monocytoid cells,
and plasmacytoid cells (e-Fig. 8.67) that may replace or disrupt the appear-
ance of pre-existing germinal centers. The cells also infiltrate the bronchial
mucosa with production of lymphoepitheliallesions as are seen with MAL-
Toma in other sites. The infiltrating cells are positive for CD19, CD20, and
bcl-2, but negative for CDS, CD10, CD23, and cyclin Dl.
2. Large Cell Lymphoma. Pulmonary large cell lymphoma generally presents
as a solitary mass lesion involving a single lobe of the lung. These masses
may develop central necrosis. Pulmonary large cell lymphomas are usually
either of diffuse large cell type or immunoblastic, and are of B-celllineage
(e-Fig. 8.68).
3. Lymphomatoid granulomatosis (LYG} is the name for a lung lesion now rec-
ognized to be a malignant lymphoma, now also known as angiocentric
immunoproliferative lesion (AIL). It is an Epstein-Barr virus (EBV)-related
B-cell proliferation analogous to post-transplant lymphoproliferative dis-
order or AIDS related lymphoma. In fact, patients with LYG often have
some underlying immunodeficiency. LYG usually is seen in middle aged
and older adults, and shows multiple nodules that may suggest metastatic
disease; concomitant skin and CNS involvement is quite common. Patients
have respiratory symptoms such as cough or dyspnea, or may have consti-
tutional symptoms. Microscopically, vasculocentric and angio-destructive
lymphoid infiltrates that involve all layers of the vessel are the hallmark
of LYG (e-Fig. 8.69); this vascular infiltration is hypothesized to result in
infarct-like necrosis of the lung seen in the higher grade cases. Grade 1
lesions have infiltrates composed mainly of small T-cells, plasma cells, and
histiocytes; CD20 and EBV stains show only rare atypical B-cells (<5 per
hpf). Grade 2 cases show a greater number of atypical B-cells (5-20 per
hpf). Grade 3 cases have abundant large, atypical B-cells.
4. Leukemic infiltrates. The lung may be the site of acute leukemic infiltrates
in cases of acute myelogenous leukemia (AML), representing a granulo-
cytic sarcoma or extra-medullary myeloid tumor. Infiltration of airways,
interstitial infiltrates, or nodular parenchymal masses are possible manifes-
tations. Histologic clues to a diagnosis of AML include blast-like cells and
the presence of immature eosinophilic precursors. Many cases are initially
thought to represent a large cell lymphoma; one of the first clues to the
correct diagnosis is the absence of expression of both B- or T-cell markers
in the presumed lymphomatous cells.
The lung may also show extensive involvement by chronic lympho-
cytic leukemia/small lymphocytic lymphoma. In this setting, the infiltrate
presents as nodular expansions along lymphatic routes in the lung.
5. Other hematolymphoid diseases. The lung has been the reported primary
site of a variety of hematolymphoid lesions, including Hodgkin disease,
intravascular lymphomatosis, plasmacytomas (e-Fig. 8.70), and Castle-
man disease. However, it should be reiterated that secondary involvement
of the lung in patients with advanced hematolymphoid disease is very
common, can occur with essentially all entities, and so must always be
excluded.
6. Pseudolymphoma. Small nodular lymphoid deposits in the lung have tradi-
tionally been called pseudolymphoma. However, with the advent of more
specific diagnostic techniques, many of these lesions have been shown to be
lymphomas, such as MALTomas. For lesions that can be demonstrated to
be polyclonal, the term lymphoid hyperplasia is more appropriate.
L. Metastatic tumors. It is prudent to include metastatic disease in the differential
diagnosis of every lung tumor. Metastases should be suspected when a tumor
type is encountered that would be unusual as a lung primary tumor. Also,
presentation as multiple nodules, the finding of a tumor that is predominantly
within lymphatics or vessels, and a tumor that appears very well circumscribed
without a host stromal and inflammatory reaction should also raise suspicion
for a secondary lung tumor. Clinical history, liberal use of special stains, and
radiologic consultation can all be used to evaluate the possibility of metastatic
disease.
Cytopathology of the Lung

Hannah R. Krigman
I. SAMPLE TYPE. Cytology is an appropriate diagnostic modality for both neoplastic

and non-neoplastic conditions of the lung. Exfoliative techniques include prepara-
tions of sputum, fluid from bronchial washing or lavage, and bronchial brushing.
Fine needle aspiration of pulmonary processes can be via a transthoracic, trans-
bronchial, or transesophageal approach. The sensitivity of cytopathologic evalua-
tion depends on the type of procurement technique used. Sputum specimens yield
the lowest quantity of exfoliated cells, FNA specimens yield the highest; endo-
bronchial and transthoracic FNA for central and peripheral lung masses, respec-
tively, both yield similarly cellular specimens.
II. SPECIMEN ADEQUACY AND PREPARATION
A. Exfoliate endoluminal samples (sputum, bronchial washing, bronchioalveolar
lavage, and bronchial brushing). Bronchial washing, brushing, and lavage speci-
mens are adequate if sufficient alveolar macrophages are present, or if diagnos-
tic findings are present. Sputum must contain macrophages to be adequate; if
sputum does not contain macrophages, then the cells are likely of oropharyn-
geal origin. For bronchial brushing specimens, bronchial epithelial cells are also
required (e-Fig. 8.71). Samples can be submitted fresh or in liquid fixative for
the monolayer technique. Mucus in sputum can be dispersed by mechanical dis-
ruption or by lytic agents. Airdried cytospins can be stained by a Romanowsky
technique; alcohol fixed samples can be Pap stained. Brushing samples can be
smeared directly on a slide, but care must be taken to fix smears immediately
or air-dry artifact will render the slides uninterpretable. Brushes may be fixed
in fluid, and cell block or Pap stained specimens can be made from material
dislodged from the brush into the fluid.
B. Fine needle aspirates of lung masses, whether obtained via endoscopic or
transthoracic approaches, are DiffQuik and Papanicolaou stained.
C. Pleural effusions are generally submitted fresh, in toto, from which a well-mixed
portion (generally 2-300 ml) is used to prepare both a cytospin DiffQuik and
ThinPrep Papanicolaou stained slide preparation. Benign mesothelial cells are
generally found in these specimens (e-Fig. 8.72), with or without inflammation.
Ill. DIAGNOSTIC CATEGORIES
A. Negative for malignancy. This diagnosis is rendered when the specimen shows
only alveolar macrophages, benign bronchial epithelial cells, and mixed inflam-
matory cells. This diagnosis is also used when fungal elements are identified, or
when viral cytopathic changes are seen.
B. Atypical cytology. This diagnosis is rendered when the specimen shows bronchial
epithelial cells which can be interpreted as reactive, but in which there is the
absence of evidence of an underlying lesion. A repeat aspirate or tissue biopsy is
usually suggested. An aspirate which shows markedly reactive bronchial epithe-
lial cells which may be interpreted as positive for carcinoma should be repeated
if the cells are adjacent to ciliated bronchial epithelium (e-Fig. 8.73).
C. Suspicious for malignancy. This diagnosis is rendered when rare malignant cells
are present, but the quantity is insufficient for a definitive diagnosis of malig-
nancy. This diagnosis usually prompts a repeat diagnostic procedure before
definitive surgical treatment.
D. Positive for malignancy. This diagnosis is rendered when both the quality and
quantity of malignant cells are sufficient for an unequivocal diagnosis of malig-
nancy. Diagnoses of malignancy are best considered a function of both quality
and quantity of atypical cells; either rare markedly atypical cells or an abundance
of only minimally atypical cells can suggest malignancy.
IV. NON-NEOPLASTIC CONDITIONS
A. Infections
1. Viral. The radiologic correlates to viral infection typically include diffuse
pulmonary infiltrates. Cytopathologically, viral changes are best appreciated
on bronchial washing and bronchioloalveolar lavage specimens. The viral
cytopathic changes seen in Herpes virus infections include multinucleate and
single infected cells that have large nuclei with clear to faintly basophilic cen-
ters and peripheralized chromatin; the nuclei are molded with one another.
CMV infected cells have both nuclear and cytoplasmic inclusions that are
PAS positive. Adenovirus, which yields a typical "smudged" appearance on
histologic section, shows a polygonal nuclear inclusion and multinucleate
cells. Measles shows a characteristic multinucleate giant cell.
2. Fungal. The chest X-ray of patients with fungal infection can exhibit either a
mass effect, multiple nodules, or, on occasion, diffuse infiltrates.
a. The cytologic features of Pneumocystis jiroveci are well described. Alve-
olar casts containing fibrin and the organisms have a bubbly or foamy
look on Papanicolaou-stained materials. Romanowsky stained prepara-
tions show a central organism and a dear coat. Methenamine silver stains
can highlight the organism, and the stain can be applied to formalin-
fixed paraffin embedded cell blocks, or to cytospins. Patients taking
prophylactic antibiotics for Pneumocystis may not have alveolar casts,
and their samples may have rare, cup shaped organisms located within
macrophages.
b. Fragments of Rhizopus or Aspergillus species can be seen in washings,
either from cavitary masses or in ABPA. In the latter, concretions of allergic
type mucin or Charcot Leyden crystals rna y be noted.
c. Histoplasma infection rarely shows detectable organisms; however, a
granulomatous reaction may be present in aspirations of pulmonary
masses or of involved lymph nodes.
d. Cryptococcus, Coccidiomycosis, and Blastomycosis can all induce solitary
masses, which can be sampled by aspiration. The yeast forms are best
appreciated on Romanowsky stains; often the organisms are clear. Again,
cell blocks, direct smears, or cytospins can be stained with traditional
techniques to better highlight the organisms.
3. Bacterial
a. The prototypical bacterial infection for which cytologic sampling is pur-
sued is tuberculosis. Infection with M. tuberculosis can yield both diffuse
infiltrates, as in military tuberculosis, or mass-like lesions, occasionally
with cavitation.
Washings or lavage may be pursued primarily for obtaining material
for culture, molecular laboratory test to detect the organism, or antibody-
mediated studies. Cytopathologically, washings or lavage fluid can show
no significant changes or may exhibit necrosis and acute inflammation.
Cavitary lesions can have secondary squamous metaplasia, which may
be atypical; for this reason, the possibility of a false positive cytology
should always be considered in the setting of a cavitary lung mass. Patients
with M. tuberculum infection can also have extensive hilar adenopathy,
which may sway the less confident reviewer to a spurious diagnosis of

malignancy. Finally, because tuberculosis can present as a mass lesion,
FNA may be obtained; frequently, multinucleate giant cells are present,
again in a setting of acellular necrotic debris and acute inflammation.
b. Community acquired bacterial pneumonias are less often sampled, but again,
fluids from such cases exhibit acute inflammation, macrophages, and res-
piratory epithelial cells.
B. Sarcoidosis. Sarcoidosis is a diagnosis of exclusion, and fine needle aspiration
from patients with infiltrates and adenopathy may be used as a minimally inva-
sive technique for evaluation. Washings and lavage do not have a high yield for
diagnosis. Aspiration of lymph nodes form patients with hilar adenopathy yields
fragments of granulomas, rare multinucleate giant cells, and a background of
mixed chronic inflammatory cells.
C. Pulmonary alveolar proteinosis (PAP). Bronchioloalveolar lavage is both diag-
nostic and therapeutic in patients with PAP. The gross features of PAP cytology
specimens are fairly typical in that lavage fluid is white to milky and may have
a surface lipid layer. Cytologically, washing samples contain granules and frag-
ments of PAS positive, diastase resistant material that is pale on Pap stain and
basophilic on Romanowsky stain. The background contains mixed acute and
chronic inflammatory cells, with variable numbers of eosinophils.
D. Hemosiderosis. Alveolar hemorrhage from any cause can result in the pres-
ence of hemosiderin laden macrophages, highlighted by iron stain. Detection
of hemosiderin pigment in 20% of macrophages on bronchoalveolar lavage
reportedly correlates with alveolar hemorrhage. Hemosiderosis requires a com-
bination of radiographic and clinical features and is best diagnosed on open
biopsy, after exclusion of other causes of alveolar hemorrhage.
V. NEOPLASMS
A. Squamous cell carcinoma (SCC), a type of non-small cell carcinoma, consists of
cells with varying degrees of keratinization, inconsistent size and shape, polyg-
onal to amphophilic cytoplasm, and dark pyknotic nuclei. The background
usually shows so-called dirty necrosis with abundant keratinized cellular debris
(e-Fig. 8.74).
B. Adenocarcinoma, another type of non-small cell carcinoma, shows cells with fine
foamy to vacuolated cytoplasm, and vesicular nuclei with prominent nucleoli.
There is typically no background necrosis unless the tumor is large (e-Fig. 8.75).
Immunohistochemistry has been proposed as an effective adjunct to cyto-
logic examination in separating SCC from adenocarcinoma. Performed on
formalin fixed, paraffin embedded cell blocks, immunohistochemical stains gen-
erally show expression of thyroid transcription factor 1 (TTF-1) in adenocar-
cinomas but not in SCC s. Care must be to distinguish between tumor groups
and incidentally obtained normal pulmonary epithelial elements which retain
TI'F-1 expression. In general, sec s express cytokeratin 5/6 and p63, while
adenocarcinomas do not express these antigens.
C. Neuroendocrine carcinoma. Small cell carcinomas and large cell neuroendocrine
tumors must be distinguished from non-small cell carcinomas since they respond
differently to chemotherapy.
1. Carcinoid tumor has both polygonal cells with rounded nuclei and spindle
cells with elongate nuclei. Nucleoli are not prominent. Aspirate smears can
have some nuclear molding and crush artifact, but aspirates should contain
some areas where the cells show the morphology of typical carcinoid.
2. Large cell neuroendocrine tumor has flattened cohesive groups of cells with
large round to polygonal nuclei that have prominent nucleoli and vesicular
nuclear chromatin. Cytoplasm is sparse.
3. Small cell carcinoma is composed of cells with only a small amount of cyto-
plasm. Nuclei show a molding pattern, fine granular chromatin, and stream-
ing. Apoptotic debris and cellular necrosis are prominent (e-Fig. 8.76).
D. Hematopoietic malignancies. Both high and low grade non-Hodgkin lymphomas

occur in the lung, either as solitary pulmonary nodules or associated with
adenopathy. The cytologic features are similar to those of lymphomas aspi-
rated at other sites. High grade lymphomas contain a discohesive single cell
population with fragmented lymphocytes in the background; individual cells
have sparse cytoplasm. Prominent chromocenters are present. Low grade lym-
phomas are composed of a monotonous mature lymphoid population. For most
lymphomas, some of the FNA passes should be reserved for flow cytometry to
characterize the malignant cell population. If the background inflammation is
mixed and contains eosinophils, material should be reserved for a cell block to
identify the Reed Sternberg cells of Hodgkin disease or for gene rearrangement
studies for T-celllymphoma.
E. Mesothelioma is usually seen in association with a pleural effusion. Cytologically,
the cells show variability in size, with those in clusters having scalloped edges. A
cell-in-cell arrangement is often present. Enlarged and convoluted nuclei, with
or without nucleoli, are typical (e-Fig. 8. 77). Appropriate immunohistochemical
stains to rule out adenocarcinoma should be performed. Asbestos fibers (e-Fig.
8.78) may be seen in bronchioalveolar lavage specimens from these patients.
F. Metastases to the lung are common and should be evaluated on the basis of the
patient's clinical history. The diagnostic approach to tumors of unknown origin
presenting as lung metastases is the same as for tumors of unknown origin
presenting at other sites (Semin Oncol. 1993;20:206).
SUGGESTED READINGS
Churg A, Green FYH, eds. Pathology of Occ"(Jational Diseases. New York, NY: lgaku-Shoin;
1988.
Churg AM, Myers JL, Tazelaar HD, et al. Thurlbecks Pathology of the Lung. 3rd ed. New York,
NY: Thieme; 2005.
Colby lV, Koss MN, Travis WD. AFIP Atlas of Tumor Pathology. Series Ill. In: Tumors of the
Lower Respiratory Tract. Armed Forces Institute of Pathology; 1996
Katzenstein AA. Katzenstein and Askin's Surgical Pathology of Non-Neoplastic Lung Disease, 4th
ed. Philadelphia, PA: WB Saunders Company; 2006.
Papanicolaou Society of Cytopathology Task Force on Standards of Practice. Guidelines of the
Papanicolaou Society of Cytopathology for the Examination of Cytologic Specimens Obtained
from the Respiratory Tract. Diagnostic Cytopathology. 1999;21:61-69.
Tomashefski TF, Cagle PT, Farver CF, et al. Dail and Hammar's Pulmonary Pathology. Vol. 1
Non-neoplastic Lung Disease, 3rd ed. Springer Verlag; 2008.
Travis WD, Brambilla E, Miiller-Hermelink HK, et al. Pathology & Genetics of Tumours of the
Lung, Thymus And Hearl (World Health Organization Classification of Tumours). WHO
Press; 2004.
Travis WD, Colby TV, Koss MN, et al. AFIP Atlas of Non-Tumor Pathology. Series 1: Non-
Neoplastic Disorders of the Lower Respiratory Tract. Washington, DC: American Registry of
Pathology and the Armed Forces Institute of Pathology; 2002.
Wick MR, Leslie KG. Practical Pulmonary Pathology: A Diagnostic Approach. 2nd ed. Churchill
Livingstone; 2010.
Cardiovascular System
Jochen K. Lennerz and John D. Pfeifer
HEART
I. NORMAL ANATOMY. The normal weight of the adult heart is 300 to 350 g (male) and
250 to 300 g (female). Cardiomegaly above a critical weight of 500 g is associated
with ischemic changes (see later) and is termed cor bovinum. The normal ventricular
thickness is 0.3 to 0.5 em on the right and 1.2 to 1.5 em on the left (e-Fig. 9.1),*
measured at the base of the papillary muscles (Fig. 9.1). The heart is composed
of three layers: the epicardium (including the serous or visceral pericardium, and
the main branches of the coronary arteries), the muscular myocardium, and the
endocardium (with an ill-defined subendocardial layer that contains many Purkinje
fibers).
Microscopically, the normal myocardium is a functional syncytium of myocar-
dial fibers (cardiac myocytes) that have centrally located nuclei (e-Fig. 9.1 ). Cardiac
myocytes are a specialized form of striated muscle; faint dark eosinophilic inter-
calated discs between the myocytes form the mechanical and electrical couplings.
Numerous capillaries with sparse interstitial tissue are found between the myocar-
dial fibers (e-Fig. 9.1).
The atrioventricular valves (mitral and tricuspid) are composed of an annulus,
leaflets, chordae tendineae, and papillary muscles. The semilunar valves (aortic
and pulmonic) are composed of three cusps (each with a sinus), which meet at
the three commissures (corpora arantii, e-Fig. 9.2). Valves are relatively avascular,
and are lined by endothelial cells on a thin layer of collagen and elastic tissue on
the atriaVarterial side, a thicker layer of dense collagen on the ventricular side,
and loose myxoid connective tissue (zona spongiosa) in between. The fibrous and
spongiotic regions are normally of equal thickness (e-Fig. 9.2).
The conduction system is composed of specialized myocytes, with fewer interca-
lated discs and higher glycogen content. Masson trichrome, Verhoeff-van Gieson,
and Alcian Blue stains can be used to demonstrate the conduction system (e-Fig.
9.3 ). Exact knowledge of the topographic anatomy and correct sampling techniques
are paramount (e-Fig. 9.4).
II. GROSS EXAMINATION AND TISSUE HANDLING
A. Endomyocardial biopsies are usually taken via a right-sided cardiac catheter;
the most common indications are monitoring of heart transplant rejection, and
grading of Adriamycin toxicity. To avoid sampling errors, a minimum of three,
preferably four, samples of myocardium are recommended (e-Fig. 9.5). The tis-
sue fragments should be counted and measured during gross examination; their
color and consistency should be noted. The tissue should be placed between
foam pads or wrapped in filter paper for routine processing. Examination of
at least three levels is recommended; some laboratories keep the intervening
sections for additional stains if required to assess myocyte damage and fibrosis.
Histologically, an adequate biopsy contains at least 50% myocardium, exclud-
ing previous biopsy sites (e-Fig. 9.5). Occasional cases require fresh frozen tissue
or glutaraldehyde fixation for special techniques such as molecular diagnostics
*Aile-figures are available online via the Solution Site Image Bank.
149
Coronary Arteries - Bypasses - Sectioning
B
LAD iii anterior
D RCA LVT
Figure 9.1 Coronary arteries, bypasses, sectioning of the ventricles. (A) The left coro-
nary artery branches into LCX (left circumtlex) and LAD (left anrerior descending),
the latter supplies the anterior septum via SP (septal perforators). The RCA (right
coronary arrery) supplies the AV node (not shown), branches into the AMB (acute
marginal branch) and the PD (posterior descending artery). The origin of the PD
derermines the distribution type (right vs. left). Examples of ACVB (aorta-coronary
venous bypass) and UMA (left internal mammary artery) grafts are displayed in gray.
(B) Slice from section illustrated in A. The myocardium displays the supplying arteries
for mapping of myocardial infarctions. The left ventricular thiclcness (LVT) is mea-
sured on the level of the anterior papillary muscle. The large septal square illustrates a
section to determine myocyte disarray. The small rectangle close to the right ventricle
(RV) illustrates a myomectomy specimen, which are sectioned perpendicular to the
endocardial surface; preferred is horizontal (a) or vertical (b).
150
Chapter 9 • Cardiovascular System I 15 1
or electron microscopy (see later), respectively. Adipose tissue between myocar-

diocytes is a normal finding and does not indicate ventricular perforation
(e-Fig. 9.5).
B. Cardiac valves are often removed because of calcific degeneration or perforation
as a sequela of bacterial endocarditis (e-Fig. 9.2). Most valves are received in
fragments; if possible, the description should include the distribution of vegeta-
tions (e-Fig. 9.6) and presence or absence of non-surgery-related leaflet destruc-
tion. In cases of calcific degeneration, slow acid decalcification after fixation
may be necessary. Sections are taken from the free edge to the annulus.
Prosthetic valves are typically removed because of thrombosis, anastomotic
or valvular leakage, mechanical failure, or infection. Evaluation of the prosthetic
valve ring attachment is, therefore, critical as infective endocarditis typically
affects this region. For most mechanical heart valves, it is not possible to submit
any tissue for histology, unless vegetations are present. For bioprosthetic valves,
however, the valve cusp is submitted.
Valves from patients treated with the appetite-suppressant drug Fen-Phen
(a combination of fenfluramine and phentermine) show patterns of changes
that resemble carcinoid valve disease with superficial layers of myofibroblastic
proliferation on otherwise normal valve architecture.
C. Myomectomy specimens, from ventricular aneurysm repair or septal myomec-
tomy procedures should be measured, weighed, and sectioned at 3-mm inter-
vals, perpendicular to the endocardial surface (Fig. 9.1). All layers of the heart
should be described. For cardiac tumors, appropriate sections should assess the
inked specimen resection margins (see later).
D. Heart explant specimens should be weighed, described, and dissected as outlined
(Fig. 9.2). In addition, the valves (circumference or diameter) and walls (Fig.
9.1) should be measured. The septal and ventricular configuration (concentric
vs. dilatative ventricular hypertrophy) should be described. Usually the inflow
tract of the donor heart is dissected to match the recipient's anatomy; thus,
fragments of donor tissue are frequently submitted in the same container.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF THE HEART
A. Disorders of the Endocardium
1. Infective endocarditis is characterized by bacterial colonization of the valve
forming vegetations that are red, irregular (e-Fig. 9.6), and composed
of granulation tissue and thrombus; their friability explains the propen-
sity for associated septic embolization (e-Fig. 9.2). The myocardium is
typically not involved. Staphylococcus aureus typically produces acute
endocarditis, whereas Streptococcus viridans produces subacute endo-
carditis. Several organisms normally found in the oral cavity are also
causative, and have been referred to as the gram-negative HACEK organ-
isms (Hemophilus aphrophilus. Actinobacilus actinomycetemcomitans. Car-
diobacterium hominis. Eikenella corrodens, and Kingella kingii). Staphy-
lococcus epidermidis also causes infective endocarditis, more common in
the setting of prosthetic valves. Healed infective endocarditis leaves residual
valve damage, often fenestrations, usually with a hemodynamic jet lesion and
adjacent endocardial fibrosis.
2. Nonbacterial thrombotic endocarditis (marantic endocarditis) produces small
(rarely >0.5 em), pink, bland, and sterile vegetations attached to the valve
surface at the lines of closure (e-Fig. 9.6). It is typically seen in cachectic
patients with a hypercoagulable state (e.g., Trousseau syndrome).
3. Libman-Sacks endocarditis is seen in 4% of cases of systemic lupus erythe-
matosus and is characterized by flat, pale tan, spreading bands of vegeta-
tions located on both surfaces of the valves or chordae tendineae (e-Fig. 9.6).
Affected, in order of frequency, are the tricuspid, mitral, pulmonic, and aortic
valves.
152 SECTION II: THORAX
Dissection Techniques of the Heart

A lnflow..Qutflow Method
Setting: Virtually any heart disease, explanls, autopsy
Method: (1) Open superior vena cava (spare sinus node)
to inferior vena cava. Valves are indicated by
arrowhead and point (can be spared In pediatric
cases) and are cut between their commissures
(2) Right ventricular inflow tract to right-apex and
right outflow tract to pulmonary arteries
2 (3) Open left atrium extending into appendage
(not shown)
(4) Left ventricular inflow tract to left apex and left
outflow tract bending pulmonary artery ventral and
cutting through the left coronary artery.
Short-Axis Method
Setting: Virtually any heart disease, infarction (territories,
see Fig. 9.1)
Method: Place diaphragmatic side of the heart on paper
towel and perform firm cuts parallel to the atrio-
ventricular groove (Fig. 9.1). Continue "slicing"
until the left anterior papillary muscle is visible.
Then continue with Inflow-Outflow Method (A).
c
Window Method
Setting: Previous heart surgery. display of known
malformation
Method: Dissect coronary vessels
Display ventricles. outflow tract(s)
atria, foramen ovale or valves according to
underlying anatomy/findings.
D · v1 Valve-Plane Method (Base-of-Heart Method)

f ~ . Setting: Valvular disease/repair/replacement
: Method: Dissect coronary vessels
A Remove atria and great vessels (dotted line)
' . .,_.. •/ Open ventricles, then continue w ith
j) Short-Axis Method (B).
·'l'
lJ '- E Four-Chamber Cut
Setting: Cardiomyopathy
~ethod: Place diaphragmatic side of the
hgart on paper tow€11. P€1rform one
firm cut in plane displayed, opening
both atria and ventricles.
Figure 9.2 Dissection techniques of the heart.

4. Rheumatic heart disease (RHO) is a sequela of rheumatic fever (RF) caused

by Streptococcus pyogenes (group A or P-hemolytic streptococcus). Aschoff
nodules (ANs) are a characteristic feature and appear as interstitial collec-
tions of plump mononuclear cells with occasional neutrophils arranged in a
granuloma-like formation, although the presence or the number of ANs does
not correlate with clinical course or activity of the rheumatic process. The
most characteristic cellular component of ANs is the Aschoff giant cell, which
has two or more nuclei with prominent nucleoli; another characteristic fea-
ture is the presence of Anitschkow cells, which are mononuclear histiocytes
that are often arranged in a palisade around the center of the granuloma.
The macroscopic pattern is variable (e-Fig. 9.6).
The valvular disease characteristic of chronic RHD is usually the result of
multiple recurrent episodes of acute RF, and typically develops many decades
after the initial insult. RHD is the most common cause of mitral stenosis, and
in up to 75% of cases, the mitral valve is the only valve affected. In 25%
of cases, the mitral and aortic valves are affected. Progressive fibrosis leads
to thickening of the valve and chordae that eventually leads to fusion of
the mitral leaflets at the commissures, producing the classic "fish mouth"
appearance.
5. Endocardial fibroelastosis is an uncommon condition that can result in restric-
tive cardiomyopathy. Pearly-white fibroelastic thickening, caused by accu-
mulation of collagen and elastic fibers typically in the left ventricular endo-
cardium (e-Fig. 9.3), is often associated with aortic valve obstruction. The
disease occurs either focally or diffusely in children from birth to 2 years of
age.
6. Loeffler endocarditis is also known as fibroelastic parietal endocarditis with
blood eosinophilia. Classically, three stages (acute necrotic myocarditis, orga-
nizing thrombus, and endomyocardial fibrosis) are distinguished. The cardiac
lesions are associated with dense eosinophilic infiltration of other organs, and
the disease is usually rapidly fatal.
B. Disorders of the Valves
1. Myxoid change is stromal accumulation of glycosaminoglycans as a sign of
degeneration (e-Fig. 9.2). The layered architecture is preserved; if the archi-
tecture is absent or distorted, the differential diagnosis should include RIID.
The chordae tendineae are thinned and elongated in myxomatous degenera-
tion, whereas in RHD they are shortened and thickened.
2. Mitral valves are removed for acquired post-inflammatory stenosis (e.g.,
RHD) and may show commissural fusion, cusp scarring, and dystrophic cal-
cification (e-Fig. 9.2). RliD vegetations are composed mainly of fibrin and
are usually no more than 2 mm in size. Cases of mitral insufficiency or myx-
omatous degeneration show a floppy valve with redundant and ballooned
leaflets with abundant myxoid change.
3. Tricuspid valves are most commonly removed for insufficiency or infective
endocarditis.
4. Aortic valves are removed for stenosis and are typically heavily calcified,
sometimes with commissural fusion (senile calcific aortic stenosis), post-
inflammatory scarring, or calcification due to a congenitally bicuspid valve
(present in 1% of the population).
5. Pulmonary valves are usually excised because of stenosis due to congenital
heart disease (most commonly as a component of tetralogy of Fallot).
C. Disorders of the Myocardium
1. Myocarditis is the underlying etiology in about 10% of patients with new-
onset cardiac dysfunction. If not fatal, myocarditis often proceeds to dilated
cardiomyopathy. Findings in subsequent biopsies, using the first specimen as
a reference point, include ongoing/persistent myocarditis, resolving/healing
myocarditis (damage substantially reduced), and resolved/healed myocardi-

tis (damage no longer present). The so-called Dallas criteria (Hum Pathol.
1987;18:619) are often used to categorize myocarditis mainly on histopatho-
logic findings, although it has been suggested that the criteria are no longer
adequate. The WHO defines myocarditis as a minimum of 14 infiltrat-
ing leukocytes per square millimetet; preferably T-cells, with as many as
four macrophages (also known as the Marburg criteria; see Circulation.
1996;93:841). The diagnosis of active myocarditis classically requires the
presence of an inflammatory infiltrate (usually lymphocytic) and myocyte
necrosis/degeneration or damage not characteristic of an ischemic event; bor-
derline myocarditis indicates the absence of necrosis or damage and can be
applied to any form of inflammatory infiltrate.
a. Primary viral myocarditis accounts for most cases of myocarditis in devel-
oped countries. Cardiac involvement typically follows the primary viral
infection by several days. The most commonly associated agents are
enteroviruses (Coxsackie A and B), adenovirus, echovirus, and poliovirus,
influenza viruses A and B, and IllY. The infiltrate is composed mainly of
lymphocytes with associated myocyte damage (e-Fig. 9. 7). Eosinophils are
typically not seen. Primary viral myocarditis includes four clinical patho-
logical manifestations: fulminate, chronic active, eosinophilic, and giant
cell myocarditis.
b. Fulminant myocarditis has a distinct onset within 2 weeks of presentation
of profound left ventricular dysfunction without dilatation. Biopsy shows
multiple foci of active inflammation and necrosis. Patients usually show
complete histologic and functional recovery, or die within 2 weeks.
c. Chronic active myocarditis has an indistinct onset with moderate ventricu-
lar dysfunction and active or borderline myocarditis. Ongoing inflamma-
tion and fibrosis may result in the development of restrictive cardiomy-
opathy with 2 to 4 years after presentation.
d. Eosinophilic myocarditis can be attributed to eosinophilic syndromes or
allergic reactions that result in left ventricular compromise. Eosinophils
and myocyte damage are present in the biopsy (e-Fig. 9.7). This entity
sometimes referred to as hypersensitivity myocarditis is linked to treat-
ment with methyldopa, antibiotics (penicillin, sulfonamides, and strepto-
mycin), anticonvulsants, and antidepressants, and also shows eosinophils
and occasional giant cells. The myocardium has little myocyte necrosis,
and the inflammatory infiltrate is lymphohistiocytic and predominantly
perivascular (e-Fig. 9.7). In some cases, the infiltrate is subendocardial or
appears as poorly formed granulomas.
The differential diagnosis of eosinophilia in the myocardium includes
parasitic infection, allergy, a hypereosinophilic syndrome, and hemato-
logic malignancies. Cytomegalovirus infection should enter the differen-
tial diagnosis in an immunosuppressed patient.
e. Idiopathic giant cell myocarditis, also known as Fiedler's myocarditis, is
associated with autoimmune diseases (e.g., inflammatory bowel disease,
hypothyroidism) and is rapidly fatal if untreated. It typically occurs in
young, healthy, white adults and presents as congestive heart failure. Dif-
fuse, geographic myocardial necrosis with a mixed inflammatory infil-
trate including eosinophils and multinucleated giant cells in the absence
of granulomas is typical (e-Fig. 9.8). The giant cells have the immunohis-
tochemical profile of histiocytes.
f. Other organisms associated with myocarditis include bacteria, fungi, spiro-
chetes (especially Borrelia burgdorferi), Rickettsiae, Chlamydia, para-
sites (including Toxoplasma gondii in immunocompromised patients), and
helminths (trichinosis).
g. Chagas disease, the most common form of protozoal myocarditis, is

caused by the hemoflagellate Trypanosoma c:ruzi and is uncommon in
the USA. However, in endemic regions of South and Central Amer-
ica, it accounts for 25% of all deaths of 25- to 40-year-olds; up to
80% of patients with Chagas disease develop myocarditis. Histologically,
myofibers contain parasites with an associated mild chronic inflammatory
infiltrate. In the acute phase, dense inflammation with myocyte necrosis
and trypanosome amastigotes in myocytes is characteristic, whereas the
chronic phase shows interstitial and perivascular lymphoplasmacytic infil-
trate without fibrosis.
h. Secondary myocarditis can occur in the setting of collagen vascular dis-
eases, RF, drugs, heat stroke, and radiation.
i. Granulomatous myocarditis (e-Fig. 9.9) can be seen in tuberculosis or sar-
coidosis.
j. Cardiac sarcoidosis shows non-necrotizing granulomas (e-Fig 9.9) in a
background of fibrosis and necrosis. Cardiac involvement, though present
in 25% of systemic cases of sarcoidosis, is usually patchy and, therefore, a
single negative endomyocardial biopsy does not exclude the disease. The
differential diagnosis in cases of suspected cardiac sarcoidosis includes
idiopathic giant cell myocarditis, amyloid, Chagas disease, and Fabry
disease.
2. Cardiomyopathy
a. Ischemic cardiomyopathy is usually secondary to severe coronary artery
disease (e-Fig. 9.10).
b. Hypertrophic cardiomyopathy is typically seen in healthy individuals less
than 30 years old, but can be seen at almost any age. Affected individuals
suffer from angina, exertional dyspnea, or sudden cardiac death as a result
of diastolic dysfunction due to ventricular thickening. In hypertrophic
obstructive cardiomyopathy, there is classically asymmetric ventricular
septal hypertrophy (with a wall thickness of 15 to 30 mm), with associ-
ated fibrous endocardial plaques and mitral valve thickening. Microscopi-
cally, disarray ofmyofibers (e-Fig. 9.11), myofiber hypertrophy, basophilic
degeneration, and interstitial fibrosis are characteristic, although nonspe-
cific.
Currently, over 450 disease-causing mutations in 16 genes encoding
myocardial contractile proteins have been implicated in hypertrophic car-
diomyopathy, a spectrum of genetic changes that suggests that so-called
next generation sequencing (see Chap. 60) is a viable diagnostic approach.
However, as isolated findings, many causal mutations have limited impli-
cations in risk stratification and prognostication, and so the role of DNA
sequencing for genetic screening remains unsettled (Eur] Clin Invest.
2010;40:360).
c. Dilated cardiomyopathy, also known as congestive cardiomyopathy,
presents as cardiac failure due to progressive cardiac dilatation with sys-
tolic dysfunction. Hypertrophy (increased weight with normal or reduced
wall thickness) and marked dilatation of all chambers is typical (e-Fig.
9.12). Histologic examination shows nonspecific abnormalities; in about
50% of the cases, leukocytic infiltrates are present in endomyocardial
biopsies. A significant number of cases are thought to be post-viral, or
associated with alcohol use or chemotherapeutic agents. Pheochromocy-
toma is also associated with dilated cardiomyopathy. Dilated cardiomy-
opathy occurring in the peripartum period (up to 6 months after delivery)
is known as peripartum cardiomyopathy. About 90% of familial cases
show autosomal dominant inheritance, and 5% to 10% are X-linked; in
addition to the genes affected by hypertrophic cardiomyopathy, additional
mutations in cytoskeletal, nuclear envelope, and mitochondrial proteins

have been found (Circulation. 2002;66:219).
d. Restrictive (obliterative) cardiomyopathy is uncommon in developed coun-
tries. The ventricles are normal or slightly enlarged and are not dilated; in
contrast, the atria exhibit relative bilateral dilatation. Patchy or interstitial
fibrosis is found histologically (e-Fig. 9.13). The eosinophilic form shows
an eosinophil-rich myocardial infiltrate, whereas the noneosinophilic form
(more common in USA) shows nonspecific findings. Restrictive cardiomy-
opathy is typically caused by endomyocardial fibrosis or hemochromato-
sis, but is often idiopathic.
e. Infiltrative cardiomyopathy is descriptive for a broad panel of metabolic
diseases, and can be assigned to any disorder that restricts ventricular
filling.
i. Cardiac amyloidosis histologically shows amorphous, eosinophilic,
extracellular material (e-Fig. 9.14). Cardiac amyloidosis is associ-
ated with restrictive features due to associated decreased ventricular
compliance and so presents with diastolic dysfunction. Grossly, the
myocardium appears stiff, and rubbery or waxy. The diagnosis of
amyloid is confirmed by demonstrating apple-green birefringence with
polarized light using a Congo red stain and/or electron microscopy
(e-Fig. 9.14), or by immunohistochemistry (e-Fig. 9.15).
ii. Hereditary hemochromatosis is a homozygous autosomal recessive disor-
der resulting from HFE gene mutations. The mutation results in unreg-
ulated uptake of iron in the small intestine, leading to iron deposition
in the liver (hepatomegaly), pancreas (diabetes mellitus), skin (hyper-
pigmentation), or heart (dilated or restrictive cardiomyopathy). In the
heart, myocytes and interstitial macrophages contain abundant brown
pigment (e-Fig. 9.16), which can be demonstrated to be iron by the
Prussian blue stain, but there is little correlation between the amount
of cardiac iron and systolic dysfunction. Increased cardiac iron must be
distinguished from lipofuscin; the latter is more finely granula.r; derived
from normal intracellular lipid peroxidation, and is not stained by
Prussian blue (e-Fig. 9.16). Iron overload is not specific for hereditary
hemochromatosis but can also be seen in the setting of thalassemia,
multiple transfusions, hemosiderosis, or hemolytic anemia.
iii. Other infiltrative cardiomyopathies include Loeffler endocarditis, endo-
cardial fibroelastosis, and mitochondrial myopathies.
f. Arrhythmogenic right ventricular cardiomyopathy is also known as right
ventricular dysplasia, parchment right ventricle, and Uhl's anomaly. This
uncommon variant of familial cardiomyopathy shows replacement of the
myocardium by adipose and fibrous tissue, predominantly in the inferior
and infundibular wall, without associated coronary artery sclerosis. The
genetics of the disease have recently begun to be characterized(] Cardia-
vase Electrophysiol. 2005;16:927).
g. Drug-/radiation-induced cardiomyopathy (Cancer Treatm Rev. 2004;30:
181) is caused by drugs such as Adriamycin and cyclophosphamide
and shows primarily subcellular changes that are best seen by electron
microscopy.
i. Adriamycin (doxorubicin) toxicity is characterized by dose-dependent
changes, predominately in the subendocardial region. It frequently
occurs after lifetime doses above 500 mg/m2 • Vacuolization of myocytes
(mainly due to marked dilatation of the sarcoplasmic reticulum) is
initially present (e-Fig. 9.17), followed by the appearance of typical
"adria cells" that show loss of cross striations, myofilamentous bundles,
and accompanying homogeneous basophilic staining (corresponding to
ultrastructural fragmentation of sarcomeres). There is no accompany-

ing inflammation. Since the microscopic features are not specific for
Adriamycin toxicity, clinical correlation is required (Environ Health
Perspect. 1978;26:181 and Int] Cardiol. 2007;117:6).
ii. Cyclophosphamide toxicity may produce hemorrhagic necrosis, intersti-
tial hemorrhage, extensive capillary thrombosis, fibrin deposition, and
necrosis of myocardial fibers.
iii. Radiation enhances the changes seen with chemotherapy. Constrictive
pericarditis, myocardial fibrosis, and coronary artery lesions are also
associated with radiation therapy.
3. Myocardial ischemia-ischemic heart disease
a. The appearance of a myocardial infarct is dependent on the age of the
infarct (e-Fig. 9.18). Following acute ischemia, the histologic changes
include waviness of fibers (after 1 to 3 hours), progressing to contrac-
tion band necrosis (after 4 to 12 hours) (e-Fig. 9.19) and infiltration by
neutrophils (after 2 to 24 hours). In cases of reperfusion, contraction band
necrosis can be seen after 18 to 24 hours as the cells begin to lose cross
striations and nuclear detail. Total coagulative necrosis can be seen by 24
to 72 hours.
b. Chronic ischemic heart disease culminates in diffuse myocardial atrophy
(brown atrophy) with patchy perivascular and interstitial fibrosis, with
progressive ischemic necrosis. The heart is small with chocolate-colored
myocardium that shows excessive lipofuscin deposition within the fibers.
c. Microscopic arteriopathy is a term used to designate the changes in periph-
eral coronary arteries that undergo sclerotic changes (e-Fig. 9.20) resulting
in a small lumen(> 75% reduction in cross-sectional area). The disease is
typically seen in chronic hypertension or with cocaine-induced cardiomy-
opathy, but will to some degree occur in chronic heart transplant rejection
where it becomes the rate-limiting step to long-term survival.
D. Disorders of the Pericardium
1. Acute pericarditis (e-Fig. 9.21) is idiopathic in 90% of cases, but can be caused
by viruses (Coxsackie B, echoviruses, influenza, mumps, Epstein-Barr virus)
or bacteria (Staphylococcus aureus, Streptococci, or Haemophilus influenza).
Acute serous pericarditis can be secondary to acute RF, connective tissue dis-
orders (e.g., systemic lupus erythematosus), uremia, metastatic malignancy,
and renal transplantation. In contrast, acute fibrinous or serofibrinous peri-
carditis can be secondary to myocardial infarction (typically after 1 to 3
days), uremia, chest radiotherapy, RF, systemic lupus erythematosus, cardiac
surgery, pneumonia, pleural infection, and cardiac trauma. Caseous peri-
carditis is usually due to Mycobacterium tuberculosis infection. Healed acute
pericarditis usually results in a focal pearly thickened epicardial plaque, also
known as a "soldier•s plaque."
2. Chronic pericarditis can lead to constrictive pericarditis where the heart is
encased by a thick layer of fibrous tissue. Constrictive pericarditis can follow
caseous pericarditis or radiotherapy, but is usually idiopathic.
3. Neoplasms. Although primary neoplasms of the pericardium are very rare
(including mesothelioma, germ cell tumors, and angiosarcoma), pericardia!
involvement is present in up to about 10% of patients with disseminated
malignancy.
4. Pericardia! effusions. Effusions can be as large as 500 mL in some settings,
such as congestive heart failure and hypoproteinemia. However, in acute
cardiac tamponade, rapid accumulation of as little as 200 to 300 mL can
cause cardiac compression and death.
IV. NEOPLASMS OF THE HEART. The four most common cardiac primary tumors (and
tumor-like conditions) are all benign and account for 70% of cardiac neoplasms.
Primary malignancies of the heart are very rare; involvement of the heart by a
malignancy is far more likely to represent metastasis by lung carcinoma, breast
carcinoma, melanoma, lymphoma, leukemia, renal cell carcinoma, and choriocar-
cinoma. In cases of metastatic spread to the heart, the pericardium is often involved.
A. Cardiac myxoma is the most common primary tumor of the heart. In the spo-
radic form, the tumor typically occurs in middle-aged women, and is grossly
a spherical, soft gray-white, gelatinous, lobulated tumor 1 to 10 em in maxi-
mal dimension, typically attached by a stalk to the left atrium near the fossa
ovalis (e-Fig. 9.22). In familial cases (e.g., Carney complex, NAME syndrome
[Nevi, Atrial myxoma, Myxoid neurofibroma, and Ephelides], or LAMB syn-
drome [Lentigines, Atrial myxomas, Mucocutaneous myxomas, and Blue nevi]),
the mean age of patients is mid-20s, and the tumor is more often attached to
the right atrium or is multicentric. Microscopically, myxomas consist of plump
spindled or stellate cells in abundant loose myxoid stroma (e-Fig. 9.22). Het-
erologous elements including cartilage, foci of ossification (petrified myxoma),
or gland formation (glandular myxoma) can be seen, but have no prognostic
significance (e-Fig. 9.22).
B. Papillary fibroelastoma occurs typically on the ventricular surface of the semilu-
nar valves or the atrial surface of the AV valves. The tumor accounts for 75% of
all valvular tumors, and can be up to 7 em in greatest dimension. In children, the
right side is predominantly affected. The branching avascular papillae are com-
posed of fibroelastic myxoid stroma and are lined by hyperplastic endothelium
(e-Fig 9.22).
C. Lipomas typically have a subendocardial location in the left ventricle.
D. Rhabdomyoma presents as a single (10% of cases) or multiple (90% of cases)
well-circumscribed gray-white firm myocardial nodule up to 6 em in size that
often protrudes into the ventricle. The tumor is often discovered in the first year
oflife, and is the most common cardiac tumor in the pediatric age group. Patients
usually present with heart failure or arrhythmias. Microscopically, the tumor is
composed of mixtures of round and polygonal cells with glycogen-rich vacuoles
(e-Fig. 9.22) that are separated by strands of cytoplasm radiating from the center
of the cell (the so-called "spider-cells"). Rhabdomyoma is mitotically inactive,
noninvasive, and nonmetastasizing; some tumors even regress spontaneously
after the first year of life. The tumor is thought to be hamartomatous and is
associated with tuberous sclerosis. Rhabdomyoma cells are immunopositive for
vimentin, desmin, actin, and myoglobin; focal HMB45 positive cells can be
present.
E. Intramural cardiac fibroma usually occurs as a single, white, rubbery lesion
(e-Fig. 9.22). The tumor cells are typically immunopositive for vimentin and
smooth muscle actin, indicating myofibroblastic origin; immunoreactivity for
the muscle-specific markers desmin and myoD1 is absent.
F. Other benign tumors include mesotheliaVmonocytic incidental cardiac excres-
cences (also known as cardiac MICE; e-Fig. 9.21), calcified amorphous tumor
of the heart (also known as cardiac CAT), lipomatous hypertrophy of the atrial
septum, mesothelioma of the atrioventricular node, adenomatoid tumor, epithe-
lioid or histiocytoid hemangioma, paraganglioma (extra-adrenal pheochromo-
cytoma), schwannoma, and granular cell tumor.
G. Angiosarcoma is the most common primary malignant tumor of the heart. It
typically involves the right atrium as a large mass with intracavitary extension,
and may also infiltrate the myocardium. Primary angiosarcoma of the heart is
typically more poorly differentiated than elsewhere (e-Fig. 9.22). Other rare pri-
mary cardiac sarcomas include Kaposi sarcoma, leiomyosarcoma, liposarcoma,
and rhabdomyosarcoma.
H. Carcinoid heart disease, seen in "'50% of patients with carcinoid syndrome, typ-
ically affects the heart's right side, particularly the ventricular outflow tract and
pulmonic valve. Gross findings include prominent hypertrophy and plaque-like

thickening of the endocardium. Microscopically, the valvular cusps show pro-
liferation of smooth muscle and collagen deposition, without valve destruction.
There are no carcinoid tumor cells in the lesion.
V. CARDIAC TRANSPLANTS
A. The most sensitive method for the evaluation of cellular rejection is microscopic
examination of an adequate myocardial biopsy. Often the sample will be taken
from a previous biopsy site and show healing foci of ischemic injury with vary-
ing degrees of inflammatory infiltrates, changes which should not be confused
with acute rejection. The revised and original grading scheme for acute cellular
rejection (Fig. 9.3) refers to the histologic findings (e-Fig. 9.23); however, the
presence or absence of myocyte necrosis should always be documented(} Heart
Lung Transplant. 2005;24:1710).
B. Antibody mediated rejection (AMR, also known as humoral or vascular rejection)
occurs in 10% to 20% of cardiac transplants, and is associated with hemody-
namic compromise, development of cardiac allograft vasculopathy (see later),
poor overall graft survival, and death in 20% to 50% of patients. Specific vascu-
lar and cardiomyocyte changes associated with AMR in endomyocardial biop-
sies have been described, but alone may not be a reliable method for diagnosis. In
contrast, strong staining of the endothelium of small vessels and the myocardial
capillary network (e-Fig. 9.24) by immunohistochemistry for the complement
component C4d has been shown to correlate with abnormal cardiac hemody-
namics due to AMR (}Heart Lung Transplant. 2008;27:372). The diagnosis
of AMR, therefore, rests on a combination of histopathologic findings (light
microscopic features and identification of diffuse capillary C4d immunostain-
ing) and clinical findings (clinical evidence of anti-donor (HLA) antibodies and
graft-dysfunction).
C. Quilty effect (e-Fig. 9.25) refers to the presence of a dense subendocardial lym-
phocyte infiltrate (Am] Cardiovasc Pathol. 1988;1:139), composed of predom-
inately B-cells. Quilty A lesions are limited to the endo/subendocardium, while
Quilty B lesions extend into underlying myocardium where there is often asso-
ciated myocyte damage (Cu" Opin Cardiol. 1997;12:146). There is no consen-
sus as to the pathogenesis or clinical significance of Quilty B lesions since up to
20% of post-transplant biopsies show this finding (also known as cyclosporine
effect). Quilty lesions have no known adverse prognostic effect, and are not
associated with EBV infection responsible for post-transplant lymphoprolifera-
tive disorders. Quilty effect is, therefore, classified as one of four nonrejection
findings.
Given the location of Quilty B lesions within the myocardium, and the poten-
tial for associated myocyte damage, it is often difficult to distinguish the lesions
from conventional cellular rejection. However, since Quilty effect is character-
ized by a collar of T-cells surrounding a central aggregate of B-cells, immunos-
tains for CD3 and CD20 can be used to help classify problematic cases (e-Fig.
9.26). It has recently been demonstrated that a compact network of follicular
dendritic cells is also present in the center of Quilty lesions (Am] Surg Pathol.
2006;30:1008) which can be highlighted by an immunostain for CD21 (e-Fig.
9.27).
D. Ischemic injury is the second nonrejection finding. It presents either early (up
to 6 weeks post-transplant) or late, and is related to allograft coronary disease.
Ischemic injury must be differentiated from preservation injury, which develops
as a result of the lack of organ perfusion between harvest and implantation.
Preservation injury is a common incidental finding in endocardial biopsies dur-
ing the week or two following transplantation and is characterized by necrosis
of the most superficial regions of the endocardium with associated overlying
organizing fibrin (e-Fig. 9.28). Similarly, artifacts of endomyocardial biopsy
....
en
Cl
Cellular Rejection in Heart Transplant Biopsies

Grade
2004 OR 1R 2R 3R
I Normal II .
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'-1 nfiltrate.J'\oiffuse \.~2 Foci 1\DiffuseAPoly-~
morphous
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Edema
substantial Extensive Hemorrhage
Vasculitis
Figure 9.3 Grading of cellular rejection in heart transplant biopsies. The grading is illustrated from left to right. Compar-
ison of original (1.9.90) and revised grading (2004) schemes is schematically illustrated (see also e-Fig. 9.23). Open circles
represent cardiomyocyte profiles, small dots represent vessels or in1lammatory inDltrate. The main diagnostic feature of
each grade is provided. The diagnostic features required for the 1990 grading are provided below the scheme. (Note:
diagnosis is based on the highest grade nndings present; see} Heart Transplant. 1990;9:587;} Heart Lung Transplant.
2005;24:1710; Heart Transplant Pathol. 2007;131:1169.)
processing that simulate contraction band necrosis (e-Fig. 9.19) must not be
confused with true ischemic injury.
E. Infection and lymphoproliferative disorders are the two other nonrejection find-
ings in biopsies, characterized by diffuse infiltration by small to medium-
sized lymphocytes in a pattern resembling rejection(] Heart Lung Transplant.
2005;24:1710).
F. The so-called transplant arteriopathy or cardiac allograft vasculopathy is char-
acteristic of chronic rejection. It features concentric luminal narrowing of small
vessels by intimal thickening and medial proliferation with relative preservation
of the internal elastic lamina, a pattern thought to represent an accelerated fonn
of atherosclerosis. In cases with complete vascular obstruction ischemic damage
can be found, although ischemic events are clinically silent due to the lack of
cardiac reinnervation after transplantation.
VESSELS
I. NORMAL ANATOMY. The luminal endothelial cell layer defines vessels and arteries
have three layers (e-Fig. 9.20): the intima (composed of the endothelium, internal
elastic lamella, and subendothelial connective tissue), media (smooth muscle), and
adventitia (connective tissue). Venous vessels have the same three layers, but a
thinner media and a thicker adventitia.
Endothelial cells are characterized by immunoreactivity for CD34, CD31,
vimentin, endothelin, and von Willebrand factor. Endothelium also stains for Fac-
tor VIII-related antigen and Ulex europaeus I lectin, both stronger in blood vessels
in comparison to lymphatic vessels. The smooth muscle cells of the media express
desmin. Depending on the anatomic site, pericytes and smooth muscle or glomus
cells are located along the outside of the vessel; these cells show immunoreactivity
for actin, vimentin, and myosin.
The size of arterial vessels is typically defined in relation to vessels in the kidney.
The aorta is categorized as a large artery, the renal and lobar arteries as medium-
sized arteries, and the arcuate and interlobular arteries as small arteries (Fig. 9.4).
The next smallest arterial vessels, arterioles, are defined by a media that has two
to five layers of smooth muscle cells, or as having a luminal radius that equals the
wall thickness.
II. GROSS EXAMINATION AND TISSUE HANDLING. Temporal artery biopsies are typically
about 2 to 3 em long. Because arteritis can have a patchy distribution with the so-
called skip areas (see later), proper tissue handling is essential to ensure a maximum
diagnostic yield from the biopsy. The external aspect of the vessel should be inked
(which is used to ensure that the microscopic sections include the entire wall);
the vessel should then be serially sectioned at 3-mm intervals. Mter processing,
embedding of the vessel segments should result in tissue sections with complete
ring-like profiles that have an inked external surface. At least three levels should
be examined. Orienting the vessel segments in agar before processing (Ann Diagn
Pathol. 2001;5:107) is a simple way to ensure that proper orientation is achieved
during embedding.
Although embolectomy specimens are easy to gross, the submission of all tissues
can have tremendous clinical impact, because the pathologist may ascertain the
exact source of an embolus (e.g., endocarditis, atrial myxoma) in a minute piece of
tissue.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES. Vascular diseases affect all organs
and contribute to the histopathologic presentation of a variety of diseases.
A. Vasculitides. Vasculitis is a noninfectious inflammatory disease of the vessel
wall and surrounding tissue. The Chapel Hill classification system for vasculi-
tis based on the size of the vessels (Fig. 9.4) is widely used (Arthritis Rheum.
1994;3 7:187).
Vascular Tree and Distribution of Typical Vasculitides
Aorta Artery Arteriole Capillary Venule Vein

elastic muscular
@ 0 D D
Large
Medium
Small
910-40y TAKAYASU'S
>50y&mm/h GCA ="temporal arteritis"
PAN; KD
WG; MPA; CSS
CG; CLV; HSP
Figure 9.4 Overview of the vascular tree and typical vasculitides. TA = Takayasu•s arteritis;
GCA = giant cell arteritis = remporal arteritis; PAN = Panarteritis nodose; KD = Kawasaki
disease; WG = Wegener's granulomatosis; MPA = microscopic polyangii.tis; CSS = Chw:g-Strauss
syndrome; CG = cryoglobulemia; CLV = cutaneous leucocytodastic vasculitis; HSP = Henoch-
Schonlein Purpura. Boldface type indicates ANCA-positi.ve. See also e-Fig. 9.20 and e-Fig. 9.29.
1. la1Je vusel vasculitides

a. Giant cell arteritis (also known as temporal arteritis). Three of the following
five diagnostic criteria by the American College of Rheumatology (ACR)
are required for a diagnosis of giant cell arteritis: over 50 years of age,
recent localized headache, temporal artery tenderness, ESR 50 mmlh, and
a temporal artery biopsy demonstrating vasculitis (e-Fig. 9.29). It is worth
noting that up to 60% of patients with clinical features of giant cell arteritis
show no evidence of vasculitis by arterial biopsy (Baillieres Clin Rheuma-
tol. 1991;5:387).
There are four key diagnostic features of the vasculitis associated with
giant cell arteritis: transmural inflammation, giant cells in dose relation to
disrupted elastic lamellae, intimal thickening, and marked intimal edema.
Giant cells are not required for the diagnosis, but typically are present if
a substantial or granulomatous inflammatory infiltrate is present. Non-
contiguous foci of inflammation, the so-called skip areas, are occasionally
present; patches of arteritis can be less than 0.3 mm long, which empha-
sizes the importance of microscopic examination of multiple tissue levels
(Arch Opbthalmol. 1976;94:2072).
The diagnosis of healed (e-Fig. 9.29) or subacute cranial arteritis is an
indication for prolonged steroid therapy, and so is an important differen-
tial diagnosis. Focal aggregates of lymphocytes and/or macrophages in the
media, irregular fibrosis and scarring of the media, breaks of the internal
elastic lamella (involving up to 25% of the circumference), and irregular

intimal fibrosis are the histologic hallmarks. Since the media does not con-
tain blood vessels in normal arterial vessels, medial neovascularization is
a useful indicator of previous inflammation.
Normal changes in the arteries of the elderly can complicate diagnosis.
However, arteriosclerosis typically does not include inflammation, and
the associated intimal and medial fibrosis is concentric and not irregular.
While the internal elastic lamella may show fragmentation, long breaks
are uncommon.
b. Takayasu's arteritis. Clinical findings and vascular distribution are neces-
sary to distinguish Takayasu's arteritis from giant cell arteritis because
both diseases show identical morphologic features. In more than 80%
of cases, Takayasu's arteritis affects women in the age range of 10 to 40
years. The aorta and typically the left mid to proximal subclavian artery
are affected, although in 50% of patients the pulmonary arteries and
abdominal aorta are involved. In contrast, giant cell arteritis occurs in
patients over 50 years of age and typically involves the external carotid
artery branches.
2. Medium vessel vasculitides
a. Polyarteritis nodosa is a rare systemic, necrotizing vasculitis that is not
associated with glomerulonephritis. The lesions are segmental, and may
be only partially circumferential. The inflammation may cause weakening
of the arterial wall, with subsequent aneurysmal dilatation and localized
rupture.
b. Kawasaki disease is febrile illness of childhood of unknown etiology that
is characterized by a self-limited acute vasculitic syndrome. Microscop-
ically, the vasculitis consists of an acute necrotizing arteritis similar to
polyarteritis nodosa. Differentiation from polyarteritis nodosa is based
on the distinctive clinical picture and age at presentation.
3. Small vessel vasculitides. This group of diseases is subclassified on the basis of
the presence or absence of anti-neutrophil cytoplasmic antibodies (ANCA).
Since cytoplasmic ANCA (c-ANCA) mainly recognize proteinase 3 and peri-
nuclear ANCA (p-ANCA) mainly recognize myeloperoxidase, the terms PR3-
ANCA and MPO-ANCA, respectively, are now in common use (Arch Intern
Med. 1996;156:440). ANCA-associated small vessel vasculitides are the most
common vasculitides in adults.
a. ANCA-positive. There are four ANCA-positive small vessel vasculitides,
Wegener granulomatosis, microscopic polyangiitis, Churg-Strauss syn-
drome, and drug-induced small vessel vasculitis.
The absence of granulomas defines microscopic polyangiitis. When
granulomas are present, the distinction between Churg-Strauss syndrome
and Wegener granulomatosis is made on the basis of the presence or
absence of asthma and eosinophilia, respectively.
b. ANCA-negative. There are three main ANCA-negative small vessel vas-
culitides, Henoch-Schonlein purpura, cryoglobulinemia, and the so-called
non-ANCA small vessel diseases.
The presence of lgA-dominant immune deposits in small vessels (N
Engl] Med. 1997;337:1512) is indicative of Henoch-Schonlein purpura,
the most common vasculitis in children (e-Fig. 9.29). The absence of
IgA deposits, together with the presence of serum cryoglobulins is diag-
nostic of cryoglobulinemia. In the absence of both lgA and cryoglobu-
lins, the differential diagnosis includes various non-ANCA small vessel
vasculitides including paraneoplastic small vessel vasculitis, inflamma-
tory bowel disease vasculitis, and immune complex small vessel vasculi-
tis (a category which itself includes lupus vasculitis, rheumatoid arthritis,
Goodpasture's syndrome, Sjogren disease, drug-induced immune complex

vasculitis, Beh~et disease, and infection-induced immune complex vas-
culitis). The diagnostic criteria for this group of vasculitides have been
well described (Am Pam Physian. 2002;65:1615 and N Engl] Med.
1997;337:1512).
B. Amyloid angiopathy. The deposition of waxy, extracellular, amorphous, weakly
eosinophilic material in the absence of an inflammatory reaction, without inti-
mal myofibroblasts or collagen deposits, is the hallmark of amyloid angiopathy.
IV. NEOPLASMS OF THE VESSELS
A. Benign
1. Leiomyoma is the most common benign tumor of veins, and usually arises
in the peripheral veins. Leiomyomas that arise in the inferior vena cava have
a prominent luminal component, and often represent extension of a uterine
leiomyoma in the setting of intravascular leiomyomatosis.
2. Rare benign neoplasms of the large arteries include inflammatory pseudo-
tumor and benign fibrous histiocytoma. Paragangliomas occur within the
aortic adventitia.
3. Benign lesions of the endothelium are covered in the chapter on soft tissue
tumors (see Chap. 46).
B. Malignant
1. Leiomyosarcoma is the most common malignant neoplasm of veins, and is
thought to arise from the smooth muscle cells of the media (e-Fig. 9.30). The
tumor usually shows extension into the adjacent soft tissues; only rarely is
the tumor confined to the vascular lumen. Most cases arise in the inferior
vena cava, in women (the female to male ratio is over 4:1) in their sixth
decade. Microscopically, the tumor has the same morphologic features as
leiomyosarcomas that occur at other sites.
2. Aortic intimal sarcoma is the most common malignant neoplasm of large
arteries, and is thought to arise from the pluripotent mesenchymal cells of
the intima. By definition the tumor is luminal (e-Fig. 9.30), although some
cases show focal extension into or through the media. Most cases arise within
the abdominal aorta in patients in their seventh decade. Microscopically,
the tumor is poorly differentiated and shows myofibroblastic or fibroblastic
differentiation, although rare cases show specific histologic differentiation
such as angiosarcoma or osteosarcoma. Cytologically, the tumor cells are
usually spindle-shaped with marked atypia and pleomorphism.
An analogous rare tumor, intimal pulmonary sarcoma, involves the pul-
monary arteries, usually in patients in their fifth decade who present with
symptoms suggestive of recurrent pulmonary emboli. As with aortic intimal
sarcoma, a subset of cases has the morphology of a specific sarcoma type.
SUGGESTED READINGS
Anderson RH, Becker AE. Cardiac Anatomy: An Integrated Text and Color Atlas. London: Gower
Medical Publishing; 1980.
Bharati S, Lev M. The Pathology of Congenital Heart Disease. New York, NY: Futura Publishing
Company; 1996.
BloomS, ed. Diagnostic Criteria for Cardiovascular Pathology: Acquired Diseases. Philadelphia,
PA: Lippincott-Raven Publishers; 1997.
Bowker TJ, Wood DA, Davies MJ, et al. Sudden, unexpected cardiac or unexplained death in
England: a national survey. Q J Med. 2003;96:269.
Cooper, LT. Myocarditis. N Engl] Med. 2009;360:1526.
Silver MD, Gotlieb AI, Schoen FJ. Cardiovascular Pathology. 3rd ed. Philadelphia, PA: Churchill
Livingstone; 2001.
Mediastinum
Louis P. Dehner
I. GROSS ANATOMY. The mediastinum is located in the thoracic cavity and is gen-
erally divided into superior, anteriot; middle, and posterior compartments and is
bounded by the pleura laterally. Generally, the first rib defines its superior limit
and the diaphragm its inferior border. The sternum, ribs, and thoracic vertebrae
(T1 through T11-12) constitute the skeletal confines of the mediastinum. The thy-
mus, heart and great vessels, lungs, and esophagus are among the most obvious
organs which occupy the anterior (thymus), middle (heart), and posterior (esopha-
gus and aorta) mediastinum. The aortic arch and the proximal segment of the aorta
(ascending and proximal aorta) are located in the superior mediastinum, which is
bounded by the manubrium sterni anteriorly and thoracic vertebrae 1 through 4.
The embryological aspects of the mediastinum are basically those of the organs
and structures which occupy this compartment.
The definition of the mediastinum relates to the structures and organs with
observable pathology on imaging studies and the associated differential diagnosis.
For instance, the anterior mediastinum is the site of the thymus with its varied asso-
ciated pathology from Hodgkin and non-Hodgkin lymphoma (NHL), to thymoma,
to germ cell neoplasms. The pathology of the posterior mediastinum is dominated
by a variety of neurogenic neoplasms and bronchoenteric developmental cysts.
A. Fine needle aspiration {FNA) biopsy. FNA is generally performed as an image-
guided or endoscopically directed procedure on suspected pathology in the ante-
rior or middle mediastinum. These specimens are processed in the same manner
as other FNA specimens, and are commonly examined for adequacy so that the
results can be transmitted contemporaneously with the procedure. Given the
broad range of pathologic processes in the anterior and middle mediastinum,
an advanced level of experience is recommended for FNA interpretation of spec-
imens from these sites.
One of the more common specimens is a lymph node with the differential
diagnosis of an infectious and/or granulomatous process, metastasis, or lym-
phoma. Metastasis, usually a carcinoma of the lung or elsewhere (most often
squamous cell carcinoma of the head and neck, papillary thyroid carcinoma,
or renal cell carcinoma), accounts for over 50% of the diagnoses. NHL (lym-
phoblastic lymphoma and mediastinal large B-celllymphoma) and Hodgkin
lymphoma (HL) are the most common primary malignant neoplasms of the
mediastinum; howevet; while strong suspicion about an NHL can be voiced in
the case of a lymphoblastic lymphoma, both the large B-celllymphoma and HL
of the nodular sclerosis subtype have a considerable fibrous component, which
may complicate the ability to obtain a sufficiently cellular FNA specimen for
diagnosis.
Biopsy. Tissue samplings from the anterior mediastinum are generally small
(<1 em) and often consist of multiple fragments which have been obtained
via mediastinoscopy. These specimens, commonly intrathoracic lymph nodes,
are submitted for intraoperative frozen section consultation to ascertain their
metastatic status for purposes of operability of a nonsmall cell carcinoma of the
lung. Often biopsies that show no evidence of malignancy contain granulomas
in varying stages of activity and type (including the so-called naked granulomas
of sarcoidosis), or simply a carpet of pigmented macrophages.
165
Mediastinal biopsies in those cases with a clinical suspicion of a disease

process other than the metastatic carcinoma present the intraoperative dilemma
of performing a frozen section or not when the biopsy consists of only a small
amount of tissue; howeve.t;, a discussion with the surgeon is helpful in these cases.
When lymphoma is suspected, tissue should be set aside for flow cytometry,
but necrosis and fibrosis often limit the evaluation. When the lymphoma is a
suspected HL, every fragment of the tissue is critical in the search for Reed-
Sternberg cells and their subsequent confirmation by immunohistochemistry (in
this case, prospective sectioning through the entire formalin-fixed paraffin tissue
block is recommended, with mounting of the unstained sections for additional
studies; howeve.t;, the alternative approach of returning to the block later results
in a high risk of loss of potentially diagnostic tissue during refacing of the block).
Both benign and malignant processes in the mediastinum may be accompa-
nied by a substantial fibroinflammatory reaction, which encases the underlying
pathology. It is therefore necessary in these cases to recommend a re-biopsy
when the only findings are those of chronic inflammation and fibrosis.
B. Resection. Surgical resections of mediastinal contents are restricted in most cases
to mass lesions in the anterior mediastinum with thymic-related neoplasms, the
thymus gland in cases of myasthenia gravis or a germ cell neoplasm (which
may or may not be associated with the thymus). An enlarged substernal ade-
nomatous thyroid or parathyroid adenoma may also present in the anterior
superior mediastinum; howeve.t;, the examination of these latter two specimen
types should follow the recommendation in Chapters 24 and 25, respectively.
The other compartments with resectable specimens include foregut cysts of the
middle mediastinum (most commonly a bronchogenic cyst) and enteric dupli-
cation cysts of the posterior mediastinum, as well as the entire morphologic
spectrum of neurogenic neoplasms from neuroblastoma to schwannoma and
paraganglioma (as discussed below).
A resected thymus may be represented by nondescript fibroadipose or adi-
pose tissue which upon sectioning fails to demonstrate any mass lesion (Thorac
Surg Clin. 2011;21:191). On the other hand, a mass lesion may be clearly evi-
dent by its size, shape, and weight; these three characteristics should be noted
upon the initial gross examination before any sectioning takes place. The exter-
nal surface should be described as to whether it is smooth and/or glistening,
or irregular by virtue of apparent fibrosis; the latter may reflect the presence of
adhesions between the mass and contiguous structures such as the pericardium,
lung, or pleura which may be included as part of the resection specimen. Because
surgical margins are important in pathologic staging, especially in the case of a
thymoma, the surface of the tumor should be marked in such a manner that the
resection margins can be identified microscopically. If the superior and inferior
poles of the specimen can be identified, then the specimen can be bisected along
that plane and the surface exposed to describe the salient features including
any apparent capsule or pseudocapsule; circumscription or lack thereof; dif-
fuse or lobulated appearance; uniform or heterogeneous character; solid, solid
and cystic, or cystic appearance; hemorrhage or necrosis; and any identifiable
portion or remnant of uninvolved organs. The selection of blocks for micro-
scopic section should include a thorough sampling of the margins; sections of
the apparent tumor should include any regional variations in the appearance of
the mass.
If the mass is predominantly cystic, the differential diagnosis is teratoma,
thymic cyst, or cystic thymoma. A solid, or solid and cystic, mass may represent
a thymoma, thymic carcinoma, seminoma, or mixed germ cell neoplasm, HL,
Castleman disease (CD), mediastinal large B-cell lymphoma, Langerhan cell
histiocytosis, granulocytic sarcoma (acute myeloid or monocytic leukemia), or
localized sclerosing-fibrosing mediastinitis.
Chapter 10 • Mediastinum I 167
Ill. MEDIASTINAL SOFT TISSUES

A. Inflammation (mediastinitis}
1. Acute and chronic inflammation. Acute mediastinitis with a purely neutrophilic
reaction is a consequence of a contiguous infection, rupture-perforation
of the esophagus, penetrating trauma, congenital duplication, or foregut
cyst (Thorac Surg Clin. 2009;19:37). A peritonsillar abscess, suppurative
thyroiditis, periodontal abscess, and post-sternotomy infection, notably by
methicillin-resistant Staphylococcus aureus, are other causes. In addition to
the acute inflammatory reaction, the tissues may have a necrotizing appear-
ance especially in those cases with the spread of an infection from the head
and neck region into the mediastinum by the so-called acute descending
necrotizing mediastinitis (Heart Lung Circ. 2008;17:124). With the pas-
sage of time, acute inflammation is accompanied by a mixed inflammatory
response with macrophages, a fibroblastic reaction, and microvascular pro-
liferation.
2. Chronic fibroinflammatory process (fibrosing-sclerosing mediastinitis}. This
uncommon but well-documented clinicopathologic entity comes to atten-
tion with a persistent cough and fever in young to middle-aged adults (Semin
Respir Infect. 2001;16:119). A mass lesion is usually present in the right
paratracheal or subcarinal region, often associated with punctuate calcifica-
tions. Less frequently, the presentation is as more diffuse infiltrative mass,
which is no longer confined to the middle mediastinum. An abnormal host
response to the antigens of Histoplasma capsulatum is thought to account
for cases in regions endemic for the infection. A needle or wedge biopsy is
the usual type of specimen for pathologic evaluation. Infrequently, fibroscle-
rotic lesions may be present elsewhere in the mesentery or retroperitoneum as
manifestations of the IgG4-related diseases (Am] Surg Pathol. 201 0;34:211 ).
There are several microscopic stages through which this fibroinflamma-
tory process evolves from a reactive fibroblastic stage with an edematous
background resembling nodular fasciitis, to a dense hyalinized collagen stage
of the interstitium and thickened blood vessels showing similar hyalinized
features (e-Fig. 10.1).* A dispersed population of lymphocytes and plasma
cells is present throughout the biopsy. Granulomas are not a feature in most
cases despite the association with Histoplasma. Dystrophic calcifications may
or may not be present. A similar pathologic process occurs in the lung as pul-
monary hyalinizing granulomas.
The differential diagnosis includes HL and NHL, inflammatory myofi-
broblastic tumor, calcifying fibrous pseudotumor, and fibromatosis (desmoid
tumor). Appropriate immunohistochemical studies are helpful in the differ-
ential diagnosis if diagnostic or suspected cells are found in the biopsy (Table
10.1).
3. Granulomatous mediastinitis. A number of infectious etiologies are responsible
for granulomatous inflammatory reactions in the mediastinum. It is gener-
ally the case that other sites in the thoracic cavity including the lungs and
regional lymph nodes also harbor the particular infection, which is usually
either tuberculous or fungal in nature. The active, infectious granulomas
show the presence of caseous necrosis. The granulomas are hyalinized with
or without dystrophic calcifications when the infection is inactive. The most
common organism in addition to Mycobacterium tuberculosis is H. capsu-
latum; other rare causative fungal organisms include Cryptococcus, Blasto-
myces, Coccidioides, and the Rhizopus group. Sarcoidosis typically involves
hilar lymph nodes without direct involvement of the mediastinal soft tissues,
,•• I SECTION lit THORAX
I' i.1:! I I(.!\ , .,..DtllttoeiMrnkll Phi now- .t Flbfofnftlun~ W.lof\t

' _ ~fhUidlutf...,
Fl~>roo~rc modlosllntis :!:

Ho<I!Hn llmP\Oino + + :!:
~~~~-1!1)0)11bloble.Sie tu111or - + +
Oole&'jln,lbrou poou<lo!urntlf ± +
M6cl1 tlnllllrp 8-cel tymphonw +
Fl'"""-tmla (d"""'*l +
Cltlptet 10. Nldieetii\I.IM I ,••
fltlhlllll ~~- Yolk •oll.mor

Th)1n..,.. Clab:ordnome
-r,pe A(lpNia c:dl; rnodu!luyl Temomo, moMo
1'J1>o /lJJ <mlo>dl Temomo,lmm~"'
1'J1>o 81 (lympk-lt:ll; tjmphoeyli<;
predon\Nrdt oo-\ orpnold) GeTs ef rnmlllan """hbttlop:al\YPI
-r,pe 82 (l:oo1lall) er-ccn
-r,pe 83 (lp!ll\olll; lltyrlbl; II<IIIOtnoill; GCTo w«<l...,<li:>IJpe mol.,..,.,
till clthwil'ltlldad thymic cudnoma) OOTo w«<l......trllod h~
...... ...,,..•
M~ill1lt/lnoma
rndg)laney
Motal>lldc d\ll'nomo lllolloolhiii.IIIIIIOioa.,.
M-plelllyiMna ~
Scli!n>Uiil dl)morna lk>illftnphomo
Upollbrao611noma T-eollftnfilome
Th)1nlc ....:!noma (lndudhiJ-• Hodt<Jn lftnpk..,.. oflho modlldnurn
epllldlal tumo1> ol 11>o tlljmusl "'G"'f"""'• - · Hodpln and
$qUIII'lOII4 col CIII'Ci1101110 non.fol~ t~mpllornaa
8t.llllold eare!l"'l:r'''':::l
Mu...ptlomtold c.a-• Hlllloq1li: •nd-..ntumo,.
Mll>lol:l ..rcomo end -rnd!IIIY
~~corcln~m~ llCIIa ln'!lliol:lleukolrio
s.-tllld ..- . (catl....rcoma)
car 0D1 (;:lfdnOI'l'lia - ....1111-tftla.,_n
AdllllOCI.....,mo Mlllolfl•
PIIDiotYodo,_l1101110 Thymol_.,rn•
Oo.....,rna \Oiih l!IS.I!Il tran..,,.,.,. Ll!>omo of111o mod~~a~m
...,_,..,... ollho modlldnurn
Wtii-<!III!Willl8d nM~roendoalne
..,.,..,.... (CRindd lllmo!O)
Sdllluy tbi'OUI; 111,....
~W'l:OMI
-r,plei)OIIdndd
VUcullir n...,~a~ms
A~CIRttold
Rhe.bclorn)IOWII'OOme
Pl>otit dl'lllll1111otod neu-.doalno
....,,....,.. ~oloU>llimoto
~ Clll! nllllLiroendoalne a.rdn~ 1\UI'rOI'O ot Clio DOIII>herd 110rvoo
Smd ail cwc!l"'a''\f, MC.rmendocm.&~ &ne-nrlltla . , _
Und!l!'--me [doplo tum,.. al lire U.,..uo
Cllrn~nod lll)mlc eplllloltolll.mo~>, [doplo llljl1>ll tumors
lncll<ln1 110u100Moeltto «<cl""""" Edot>lo DOlo11!l!'old111nore
1om ealt" - (11:00 aftla••ltnH
GCTa ol one llilli>ll!ll<ai!JPe (purt CCTI)
..........to..,_. ...-.
Mlllall••
s.m-.ome
E'lnbr!l>nll Clldnomo
1'10 I SECTION lit THORAX
''/.I,! I J (.!)1 WltO Clullll:atloft of~ (A. Bl, Ill. Bs. All lid C .,m.)
r.,. 'li.Tallll 'JI.IIwlllilla llla•~:etlc ......
"
Ill
a
I&
10
45
htlln~ma ..ko -I'll,
Si>lndlo" IM>Id e*">llol .... \01111 <lffllso«
110 tr'rnl>hooYioo
R....mllleo norrnll~ua..tii-W111r.. or
lrnrnollr"' tlryrnr>qW and ...,!Wid .piiM!III aolls,
wl!lar-..H....,.oorpusdos
82 28 70 Lobirlol<>lll'll' potf&ONIOIIfii\Oiol cdlueperalodllf
lmmotuR> T.Jtm~
83 II 8S Lobirlol "' 1811!> potf&ol\01 eollllelol <Olio ... ~ ""' •
mlrlmll tr'rnP~ ..,.,_-of mild
oplllol~lftlplrl rn11 ,.ba tile poalbll~ Ill' lll)ml:
Cll't!noma
AB 31 Lobirlol- mbo!d ptdlamsof~poA 01mJII>oclta pmr)
01\d IY1>e B O!m~ rtoill p01tomo..tllornol
Glindledto oodd oiiiPOd 6lill\oli!l coli>; OWIOI(
c 5
---·-""'""""·
ond B oq~ '''"'"""'tid or OM
Arf1 petlarn OIQOr<:tlomO- tqLIIrnoull,

-m
tr'rnphoeyls&"""" num.,... 1M\ In typo A; 1y1>0 A
mer
lymplloopll>olol, otlor cdl, mu....,ldormold, -let
..,,..,oat'*!, pei>laJII, llld muct.ouo foot!~""'
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~ Cld '&.,., ~TumQI.4 ~4,f,(j ~ Tumwa<t/11\el.f.ll¥ PJata2t,. ~1.19
1t1111 HMrt.I.Joft: lARC P,-.; :D)&. Lll!led Mtl:t'l ,_.,IIIIIDI\.
Cltlptet 10. Nldieetii\I.IM I 1,1
''I.I II Jt!t' I Clll:op~t!HIIOI!c ........ of lltii...,. ~~~~~- .....

S11J1 llllllf.llltt-I'M Si'Nr •1'111¥11 00
I ~DIIMIY -~ \lltfloijlfn""""' of C$1Uie 9S-100
II O,_lnYIOb'l~ouriOQidlnsooft~or.........,l -.as
plelnl•ndlor m""""""'*' In-n mo """"'"'
111 GI'OSII......,n <ll ~<fum, "nc.
and~- 60-70
(Mio-l>fblopsyor.-onorlrblo~
....nrmstb!J
IVt D'-nN1ioh'>IIOffcanlllln 81ulkr Dlwra v.tt..tl 40-50
ooMia\IOUO e>reod e>lh- Ill
IVb Dllllnt a i i L I - ~nr. ""*'•
bono, I...,) 2>30
112 SECTION lit THORAX.
:: 1"""-· lltmlc Cllcl-.

Cl7 Cllll ann COla m-1 CIII:OS NIXIII
~ + +' + ....
1ltftrolc '""'""'""'
+ + + + :1:
Lllrc.....mma :1: + :1:
Chapter 10 • Mediastinum I 173
I. A thymoma can be rarely present in an ectopic site, most commonly in

the neck, where the ectopic thymoma may or may not be associated with
a cyst, with or without accompanying parathyroid.
3. Neuroendocrine carcinoma. This primary neoplasm of the thymus con-
stitutes ~5% of all thymic epithelial neoplasms (Semin Diagn Pathol.
2005;22:223 ). Cushing syndrome is one of the known clinical presenta-
tions, as is a manifestation of multiple endocrine neoplasia type I. These
tumors are typically large and have usually invaded into the surround-
ing mediastinal tissues. The histologic features (e-Fig. 10.10) are those of
neuroendocrine carcinomas elsewhere including organoid profiles and/or
rosette-like formations of uniform cells with finely distributed nuclear chro-
matin and central coagulative necrosis to a small cell carcinoma resembling
its pulmonary counterpart (in the latter case, imaging studies are a reliable
method to localize the mass to the hilum or thymus). It is worth noting
that rosette-like profiles are seen in thymoma and mediastinal large B-cell
lymphoma. The combination of thymoma and neuroendocrine carcinoma
patterns in the same tumor argues to the point that the latter neoplasm is
fundamentally derived from thymic epithelium.
B. Germ cell tumors (GCTs). The mediastinum, typically the anterior compartment, is
one of the more common extragonadal primary sites for this group of neoplasms
accounting for 10% to 15% of all the GCTs (Adv Anat Pathol. 2007;14:69).
GCTs represent approximately 15% to 20% of all primary mediastinal neo-
plasms, as a category of malignancies that includes seminoma, endodermal sinus
tum01; embryonal carcinoma, choriocarcinoma, or a mixture of these patterns
with or without teratomatous elements. Individuals with Klinefelter syndrome
are at an increased risk for mediastinal GCTs of various histologic types from
teratomas to mixed GCTs, which may be heralded by the development of pre-
cocious puberty. Primary malignant GCTs of the mediastinum have a marked
male predilection (80% or more of the cases), whereas mature and immature
teratomas do not have a similar male preference. Metastatic GCTs from the
testis more often than the ovary can present as an apparent primary mediastinal
neoplasm.
1. Teratomas, usually of the mature cystic type, and seminoma (germinoma)
are the two most common single pattern GCTs arising in the mediastinum,
and together represent 50% to 60% of all the mediastinal GCTs. Endodermal
sinus tumor (yolk sac tumor) is next in frequency as a pure pattern GCT. Pure
choriocarcinomas occur almost exclusively in males. The remaining tumors
have a mixture of teratomatous, seminomatous, and nonseminomatous fea-
tures in a fashion similar to malignant mixed GCTs of the testis, and thus
it is important to widely sample any GCT of the mediastinum. The medi-
astinum is one site in which a GCT may engender a sarcomatous component
including embryonal rhabdomyosarcoma (ERMS), angiosarcoma, and other
sarcomatous patterns. Finally, granulocytic sarcoma and other hematologic
malignancies (true malignant histiocytosis) are also known to occur in this
pathologic setting.
Mature cystic teratoma presents over a broad age range from the neonatal
period into early adulthood. The pathologic findings are as with the ovarian
counterpart (e-Figs. 10.11 and 10.12). As with teratomas elsewhere, espe-
cially in young children, the somatic components may have immature or fetal-
like features, most commonly found in the neuroepithelium with embryo-like
neural tubes and neuroblastic foci; however, this finding should not be viewed
with any more concern than as in a sacrococcygeal teratoma in an infant. In
an older child, adolescent, or young adult, a more cautious approach to the
same finding is appropriate.
Seminoma, unlike teratoma, can present a diagnostic dilemma from

other somewhat similar appearing neoplasms in the anterior mediastinum.
Sheets of uniform polygonal tumor cells with a central round nucleus and
clear cytoplasm accompanied by lymphocytes and granulomas is the classic
microscopic appearance of a seminoma in the mediastinum, as in the testis.
However, the lymphocytic infiltrate or granulomas can obscure the tumor
cells (e-Fig. 10.13). The differential diagnosis can include mediastinal large
B-celllymphoma, HL, sarcoidosis, and primary or metastatic clear cell car-
cinoma.
Immunohistochemistry can be extremely helpful in most cases since
mediastinal seminoma has a distinctive phenotype including CAM5.2 and
vimentin (dot-like pattern, 70% to 80%), SALL4 (100%), placental-like
alkaline phosphatase (80% to 90% ), CD117 (> 70% ), CD30 (variable), and
OCI4 (90% to 100%) positivity. Cytokeratins AE1/AE3 and 7 are expressed
in only approximately 5% of the seminomas. If the seminoma is immunoposi-
tive for one or another cytokeratins; if the seminoma has arisen in the thymus;
or if a thymoma, thymic carcinoma, or non-small cell carcinoma of the lung
are other possibilities, then the diagnostic evaluation is more problematic (as
discussed above).
C. Lymphoid neoplasms (lymphomas). Lymphomas of all categories account for only
approximately 15% of all the mediastinal neoplasms overall, but for 50% to
60% of malignancies in the mediastinum (Histopathology. 2009;54:69).
1. HL of the nodular sclerosis type is the most common, with a particular
predilection for adolescent and young adult females (e-Figs. 10.14 and
10.15). When there is extensive sclerosis-fibrosis, a definitive diagnosis can
be difficult to establish as previously discussed above in the section on
sclerosing-fibrosing mediastinitis.
2. Lymphoblastic lymphoma in children presents most commonly as a mediastinal
mass (50% of the cases) (Semin Pediatr Surg. 1999;8:69). With the exception
of infrequent precursor B-celllymphoblastic lymphomas, nearly all the cases
are examples ofT-lymphoblastic lymphoma. If the bone marrow is involved,
there is generally no need to biopsy the mediastinal mass.
3. Mediastinal large B-celllymphoma (MLBCL) is specific to the mediastinum and
is thought to be derived from thymic medullary B-cells (Arch Pathol Lab
Med. 2011;135:394). It is a neoplasm which, like HL, has a preference for
young females. In fact, the differential diagnosis from HL may not be entirely
clear as is indicated by the small subset of cases referred to as "mediastinal
gray zone lymphoma" that have hybrid features of nodular sclerosing HL
and MLBCL. CD23 positivity discriminates HLBCL from HL (lnt] Surg
Pathol. 2010;18:121).
4. The unicentric form of casdeman disease (CD) (70% to 75% of all the cases)
has two microscopic patterns: hyaline vascular and plasma cell types (Cu"
Opin Hematol. 2007;14:354). Approximately 6% to 10% of all the unicen-
tric cases present in the anterior mediastinum as a well circumscribed mass
consisting of one or more matted lymph nodes resembling nodular sclerosis
HL (e-Fig. 10.16). Microscopically, a penetrating small blood vessel extends
into a germinal center composed of follicular dendritic cells surrounded by a
mantle zone of concentrically arranged small lymphocytes in the hyaline vas-
cular type of CD. In contrast, the follicles have hyperplastic germinal centers
and mature plasma cells occupying the interfollicular zone in the plasma cell
variant. The histologic features of multicentric CD resemble the plasma cell
variant. The mande zone lymphocytes are known to harbor HHV-8.
D. Neurogenic and neuroblastic neoplasms. These neoplasms in aggregate account
for 20% to 25% of all the mediastinal neoplasms, and virtually all types present
in the posterior mediastinum. In children, these tumors are neuroblastic and
Cltlptet 10. Nldieetii\I.IM I 1, .
...
~i(J:]I Jt.t.JI P&*lotk l)pelofNuoblutfc lumen
l.lndll!'ellllldotod N8 IUFl
~~~~~~- ... ,...
lt!h I1IJd& moi!NM ""hd ¢Ill ,.,...,101111
hlltl MIO 01\d need f>or l'nmii\ONot-
odlonllll!ytod-fl'llmotller
molpnt round colllumon
Pcorl)'diiii!Willl8d NB (UF or FH on- MllpntcollumdoriNn uhdllll,.._
of qDtnd MKU NB, with nsutonb,...JY p10c:o:naa.. vartetM
r-loWo<hW. MIO.IIId ........,of
""""l!lllti>U>orsa-ntan oaomo
Dlfll.o6or I!Dimbd p!Wionauro-.. lndlotlullMm orlobu~rfoel ofnouro-
(FH) will promlnonl-bfllilly-
Pl',ljbn coli dlffMMrtlo!Son, and
1101.10mlllouutn•ne.
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....,_101111 hWI « l:>w MIQ In a
~IM.
-~ngp!Wtonouroma !FHJ M.,_plefod<>l-ln •• Cllfl.....,
mowropl\lllonouromo
"""''" Pftli~urorne (fHJ A-..oof1101.10-
1'J& I SECTION lit THORAX
~i (.1:! I J l•fJI c,.tl and C,.UC LtiiMt <If tM Mtdlutfnum

l!mnclqoonlc: c:ysl (!iOo8)l(, a1 CONS) -~ qot
Ei11<r1:: <*Jpl- (l'onoiPIII c:ysl ThyrapOIW duct <)'II
Parli:ftlol (""'lomlc:) qot Moui)Omol bono 0)01 (dlost well
EclllfiOCoc<el qot Mlllllowar ll!ymlc: cytt
Thoradeductcytt CyoC~c11>Yrnomo
--<)01
PonerM11c: ~
MLIIIIIIn (H-o qol
CyodcadenomalOidtnnor
PlelraP<J-I'f bluumo (t,'pO I or 10
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An ...mic d...-lion cyll of lllo - - . . io on iDtnm.urd nm1oo,Jor q1t

lined by a "'!IWII,.. or mle:IX type ,.....,, Abo"' 20% of all dLe oale:IX
clo.plicatl011.t o.tcfo.md.ID.the~ ha tn>eoifotqp.rt lll&lfwmallo.ll, thoqrt
mt.Y be ..oQaa:.l wid! ancm>olict of 1be 1\l)pet' aitwq iodclding a ~
cytt, cr~ fiaoula, or uomm'll.ltic:alionofOl!Ct'fl'tOt ao.o!her wid> dLe
....,_.;,
VI. 10fT nnUE IEII'LAIIIIL Soft 1iMue """PJ.... llllldated co the heart or t._
!rot p:rac:zcliag .mhin the llH>ruic amty iD ooe of the mediePinal a>mplrtl..-
""" ....,.dho;&Jy .,.......,..... Tbat lurrio.g ._, DOled, virtuolly - aoft done
-plum of beniF, ittdelemrina•t, Ot !XlfligNot cype bu been rq><x!U4 itt dLe
~ ua tioa1ccue 01-n«:<ief.
Lipoma ADd schW'Mnnrn!l ueth.t most<:OmiXLOD heaip ftlm.O:ta •riring, rapu·
lively, in 1be .......;or IUl.C! ~or medieeQaum. Lipo-. invvl•aDCZCt of dLe
1byam.c -.111 i111be 110-<>&!led. d!,molipoma "'*'- poll!.011-.il u & ...........
tom& ot D<49laam ....,.... IIIIC:dtiiD.. M70icl. cello h&vc beoa ohaemd !A thlo
lco!o:a, whlcb l!l:.o lipoma,'"" ltiO!A ....,.., di._!l...,, Both~
....t h-~- of ft<iout ~U'btypct ue alto found io 1be mc<li.utinum. M$1ig·
IWll p<riph.eml"""" sbo:.&th """"; ""Bi-ltiam,....._mo,li,posarcoma,
B...U."""amo~lltme>e<:to<ftmlalt11me!r(H-..io.-I.S..CdJ'l'h...
2()01);1..•U1), cmlipot ri>olxloicl CWDDr !P~ lldiol 201)!1o,l9:31!1), clc:smi>-
pluD<: .....n
row cell tllm01; ao.cl E1lMS .... jg.n- of the aoft ciDue &&ra>m.U
!bat ba.c bcCil ~ ID. the mcdlutlactm (~ 2002;22:621). !A the
-ofl!RMS,the~~oaitiedudeaplatt~blut-(PI>B)
m• chilo~ u.adcr lOrura of ago:, or a •!)<'~( ••uinoil "''"'""""'Cdltllli<'l ~ mcdi·
...mw ll:rat<miA iD aD al.t.r dci!d, •h" .... "" J'O'ODI1 aclult. S.,.ci-riAI """""""'is
wdJ doamrnted u e. pz:imaty .a.eoplum ..;ring iD. thtdustu e.I!H'ZT1i~ media«i-
.W Dllll, ollboo&hit&Ctll&ll,.<>riai'"ta ~ 1be plc:un. .....U..r hloa (IIMio~
k:l•.2006;16;923). Sollwy £iht<roo t11m0t bu olio bcCill'Cpo.!Ud !A tho mcdlutlaol
toft w.- ....t lhymua (1-....a c.~. Tltonlc; $Mrg. 2.010;1l;382). Ia •
chilol6 ,_. oht or leta, the .olid or t.c>lid ....t cytDc: Pn wid> itt <»"'P'ex cm:dli·
palll!nl.l:<ls.ucoma may eo:!md iDto..,. or"""" moc!Wtmol=mponmm!L
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Jon H. Ritter and John D. Pfeifer
I. INTRODUCTION. The serosal membranes are derived from the mesoderm, and form
the visceral and parietal surfaces of the pleural cavity, peritoneal cavity, peri-
cardium, and tunica vaginalis testis. Histologically, the serosal membranes consist
of a single layer of flat mesothelial cells that rest on a basement membrane, below
which is a poorly delimited connective tissue layer. The parietal surfaces of the
serosal membranes are perforated by numerous narrow stomas, the so-called lym-
phatic lacuna, that connect with the extensive lymphatic plexus which drains the
enclosed cavities. Under an electron microscope, mesothelial cells show character-
istic long, slender surface microvilli; their demonstration can be used to support a
mesothelial origin for a neoplasm that is indeterminate by other histopathologic
methods.
II. SPECIMEN PROCESSING
A. Biopsy samples, from procedures performed for diagnosis or in the context
of staging procedures, are usually small tissue fragments in the range of 1
to 5 mm in maximal dimension. Detailed gross descriptions are unnecessary,
although documentation of the number and size of the fragments is important
to ensure that they are adequately represented on the slides. The tissue should
be submitted in its entirety, and three hematoxylin and eosin (H&E) stained
levels should be examined microscopically.
B. Excision specimens, from procedures performed for benign or malignant dis-
eases, include tissue from pleural decortication procedures (stripping proce-
dures to remove thick visceral pleural peels that encase the lung and decrease
ventilatory function), debulking procedures, and resections. The aggregate
size of the tissue should be described, as well as its color and texture. The pres-
ence of gross lesions should also be documented. Gross abnormalities should
be thoroughly sampled. When no gross lesions are identified, as a general rule,
at least one section per centimeter of aggregate tissue should be submitted for
microscopic examination.
Ill. NONNEOPLASTIC LESIONS OF THE SEROSAL MEMBRANES
A. Acute serositis
1. Acute pleuritis is usually infectious in origin and is most commonly associ-
ated with pneumonia. Gram-positive bacteria are most commonly isolated,
although a wide variety of pathogens can be responsible. Spontaneous bac-
terial pleuritis occurs occasionally in patients who have cirrhosis. Autoim-
mune pleuritis, although sterile, can produce clinical and pathologic findings
that resemble infectious pleuritis.
2. Acute peritonitis is usually associated with a perforated viscus. If it is due to
gastric, biliary, or pancreatic rupture, it has a chemical etiology; on the other
hand, if it is due to intestinal rupture, it has a bacterial etiology. Spontaneous
bacterial peritonitis also occurs, usually in children, immunocompromised
patients, or patients with cirrhosis. Localized acute peritonitis is a feature
of pelvic inflammatory disease.
3. Acute pericarditis can have an infectious etiology or can be a manifestation
of autoimmune disease.
B. Granulomatous serositis can present in a number of different patterns; stud-
ding of the serosa by innumerable small nodules can be especially worrisome
clinically for disseminated tumor.
177
1. Infectious. Although special stains can often demonstrate the offending

pathogen, microbiologic cultures are a more sensitive and specific method
for identification of the causative organism. Common causes include
mycobacteria, fungi (including Histoplasma, Cryptococcus, and Coccid-
ioides), and parasites (including Schistosoma, Echinococcus, and Ascaris).
2. Noninfectious etiologies include a reaction to foreign material from a prior
surgical procedure (such as starch granules and sutures) or from a perfo-
rated organ. In women, additional causes include retrograde introduction of
foreign material through the fallopian tube (e.g., douche fluid, lubricants,
radiographic contrast agents), and spillage of amniotic fluid following a
Cesarean section.
Peritoneal granulomas can form as a response to implants of keratin
produced by a neoplasm of the female reproductive tract, including mature
cystic teratoma, endometrioid adenocarcinoma with squamous differenti-
ation (of either endometrial or ovarian origin), squamous cell carcinoma
of the cervix, or even atypical polypoid adenomyoma of the uterus. Micro-
scopically, laminated deposits of keratin (sometimes including the so-called
ghost squamous cells) are present in the granulomas, but in the absence of
viable tumor, these granulomas have no prognostic significance.
3. Autoimmune causes include Crohn disease and sarcoidosis.
4. Meconium peritonitis in a neonate can lead to a serosal granulomatous reac-
tion.
C. Mesothelial Hyperplasia is commonly seen in response to chronic serosal injury.
Microscopically, hyperplasia has a number of different patterns, including
solid, tubular, trabecular, papillary, or tubulopapillary, and often shows limited
extension into the underlying connective tissue (e-Fig.11.1).* The hyperplastic
cells are often disbursed in linear, parallel, or thin layers in associated organiz-
ing fibrinous tissue. Cytologically, mild-to-moderate nuclear pleomorphism is
present, and mitotic figures and even occasional multinucleated cells can be
identified.
Given these architectural and cytologic features, mesothelial hyperpla-
sia can be difficult to distinguish from well-differentiated diffuse malignant
mesothelioma (DMM), especially epithelioid mesothelioma. The distinction is
based on the degree of cellular proliferation and atypia. Mesothelioma should
be suspected when deep infiltration of the underlying soft tissue is present, or
when areas of necrosis are present. Knowledge of the clinical setting can be
used to guide the diagnosis, although it is well established that slowly grow-
ing mesothelioma can initially present as a lesion that cannot be distinguished
from mesothelial hyperplasia.
D. Metaplasias are predominantly a feature of the peritoneal serosal surfaces in
women. Most originate from the so-called secondary Mullerian system, which
by convention includes the pelvic and lower abdominal mesothelium and
underlying mesenchyme. The close embryologic relationship of the mesothe-
lium in these areas and the Mullerian ducts (which arise from invaginations
of coelomic epithelium) provides an explanation for the fact that many of
the metaplasias produce tissues that are a normal component of the female
reproductive tract.
1. Endometriosis is thought to arise via a metaplastic process, through ret-
rograde implantation of menstrual endometrium (the so-called metastatic
theory), or as a developmental anomaly. Rare cases of pleural endometriosis
have been reported, as have cases of endometriosis in men who have been
Chapter 11 • Serosal Membranes I 17 9
treated with long-term estrogen therapy (usually in the setting of adenocar-

cinoma of the prostate).
When endometriosis develops in association with the viscera, such as
the wall of the intestine, adjacent to the ureter, the wall of the bladder,
and so on, it can clinically present with signs and symptoms that resem-
ble malignancy. Microscopically, the findings include endometrial glands
and stroma, often associated with chronic inflammation, hemosiderin laden
macrophages, dense fibrosis, and adhesions (e-Fig. 11.2). Since a num-
ber of different malignancies, most commonly endometrioid adenocarci-
noma and clear cell adenocarcinoma, can develop in endometriosis, areas
of endometriosis in biopsy and excision specimens must be carefully exam-
ined.
2. Endosalpingiosis typically occurs in women during their reproductive years.
Microscopically, multiple dilated cysts lined by a single layer of fallopian
tube-type epithelium are present. The lack of endometrial-type stroma dis-
tinguishes endosalpingiosis from endometriosis.
3. Endocervicosis, consisting of benign glands with an endocervical type
epithelium, and squamous metaplasia, are both rare. Both occur in women,
and are primarily metaplasias of the peritoneal mesothelium.
4. Ectopic decidual reaction is an incidental finding in women who are pregnant
or on high-dose progestogen therapy. Most lesions are not evident grossly,
but when they are, they consist of small gray-white nodules which may
be hemorrhagic, and often stud the peritoneal surfaces. Microscopically,
the metaplasia involves the submesothelial stroma, and consists of large
epithelioid cells with prominent cell borders and abundant amphophilic
cytoplasm (e-Fig. 11.3) morphologically identical to the cells comprising
the decidual reaction characteristic of the fallopian tube, cervix, and upper
vagina in pregnant women. Diagnostic difficulty can arise on the rare occa-
sions when the decidual cells assume a signet-ring appearance.
5. Walthard nests, usually found on the serosal surfaces of the fallopian tubes
or in the mesovarium as yellow-white nodules, are usually only several mil-
limeters in the greatest dimension. They may show cystic change, and are
usually lined by mesothelial cells that have undergone transitional (urothe-
lial) metaplasia.
6. Disseminated peritoneal leiomyomatosis (leiomyomatosis peritonealis dis-
seminata) is an uncommon multifocal proliferation of smooth muscle-like
cells that is thought to represent a hormone-induced metaplasia of the mul-
tipotential submesothelial mesenchymal cells of the peritoneum. Grossly,
it appears as widely scattered nodules that often suggest metastatic malig-
nancy. Microscopically, the lesion is characterized by cytologically bland,
benign spindle cells centered in the submesothelial connective tissue (e-Fig.
11.4 ). A conservative approach to treatment is indicated, since the condition
tends to spontaneously regress.
E. Fibrosis
1. Pleura
a. Reactive pleural fibrosis is usually a consequence of prior inflammation
or surgery. Often, the fibrosis is associated with formation of dense adhe-
sions. Because reactive mesothelial cells are entrapped within the fibrous
tissue, careful microscopic examination with knowledge of the clinical
history is required to avoid over-interpretation as mesothelioma.
b. Pleural plaques, which primarily occur on the parietal pleura of the tho-
racic cavity, are raised, discrete, white to gray-white lesions that range
from several millimeters to over 6 em in diameter. When pleural plaques
are bilateral, they are almost always related to prior asbestos exposure,
even very low fiber levels. Causes of unilateral plaques include asbestos,
as well as any process that features pleural chronic effusions. Micro-

scopically, they consist of pauci-cellular dense collagenous connective
tissue with a basket-weave pattern, sometimes associated with overlying
organizing fibrinous deposits. Asbestos bodies are essentially never seen
within the plaques. Mesothelial cells are not a prominent component of
the lesion; any significant cellularity in a putative pleural plaque should
raise concern for desmoplastic mesothelioma. Finally, since mesothe-
lioma and plaques may occur in the same individual, it is not uncommon
for blind biopsies to sample plaques; in this setting, additional biopsies
are indicated if there is strong clinical suspicion for a pleural malignancy.
c. Diffuse visceral pleural fibrosis has a number of etiologies. It is a fea-
ture of several occupational exposures (e.g., silicosis), and occurs as an
advanced hypersensitivity reaction, as a component of connective tissue
diseases, and as a sequela of bacterial pneumonia (especially as a result of
empyema). Grossly, diffuse visceral pleural fibrosis may be difficult to dis-
tinguish from desmoplastic mesothelioma. Microscopically, the fibrosis
does not infiltrate the subjacent soh tissue and has a zonated appearance,
with more cellular areas near the surface while the deeper tissues tend to
be more paucicellular; mesothelioma has the reverse pattern. Nonethe-
less, careful microscopic examination, often accompanied by immuno-
histochemical studies, can be required to exclude mesothelioma.
2. Peritoneum
a. Reactive peritoneal fibrosis is usually a consequence of recurrent bouts of
peritonitis (often associated with long-term peritoneal dialysis), decom-
pensated cirrhosis, or surgery, and is often associated with formation
of dense adhesions. As is true with reactive pleural fibrosis, reactive
mesothelial cells entrapped within the fibrous tissue must not be over-
interpreted as mesothelioma.
b. Localized plaques, composed of dense hyalinized fibrous tissue, are fre-
quent incidental findings on the splenic capsule.
c. Sclerosing peritonitis is due to hyperplasia of submesothelial mesenchy-
mal cells, and manifests as diffuse sheets of white, thickened visceral
peritoneum that encase the small bowel and also involve the diaphrag-
matic, hepatic, and splenic peritoneum. Known etiologies include peri-
toneal dialysis, infections, autoimmune disorders, therapy with the beta
adrenergic blocker practolol, and the carcinoid syndrome. Many cases
are idiopathic.
F. Cysts
1. Emphysematous bulla are the most frequent cystic lesion that involves the
pleural cavity.
2. Peritoneal inclusion cysts characteristically occur in the peritoneal cavity in
women of reproductive age (although they also rarely occur in males, and
also rarely occur in the pleural cavity). They are usually incidental findings
at the time of surgery, and consist of single or multiple, thin-walled, translu-
cent, unilocular cysts lined by a single layer of bland, flattened mesothelial
cells.
3. The so-called pericardia} cyst is the most common cyst associated with the
pericardium. It can achieve dimensions of 15 em or more. Microscopically,
it is lined by bland mesothelial cells.
G. Splenosis is an incidental finding, and usually represents implantation of splenic
tissue as a result of traumatic splenic rupture. Grossly, innumerable red-blue
nodules ranging from several millimeters to over 5 em in diameter are scattered
widely through the abdomen.
H. Eosinophilic peritonitis arises in the context of a variety of medical diseases
including childhood atopy, autoimmune disorders (especially collagen vascular
Chapter 11 • Serosal Membranes I 181
diseases), and the hypereosinophilic syndrome. It also occurs in association

with lymphoma and metastatic carcinoma. Other causes include a ruptured
hydatid cyst, and in association with peritoneal dialysis.
IV. BENIGN SEROSAL NEOPLASMS
A. Adenomatoid tumor is of mesothelial origin, and usually arises in the peritoneum,
also rarely from the pleura. It most commonly involves the serosal surfaces of
the uterus or fallopian tubes, or paratesticular regions. Grossly, the tumor
usually forms a tan 1 to 2 em well-circumscribed nodule. Microscopically,
the tumor is composed of tubular and slit-like spaces lined by a single layer
of flattened cuboidal cells with bland cytology (e-Fig. 11.5). The cells are
immunopositive for cytokeratin, calretinin, WT1, and vimentin expression,
but immunonegative for factor VIII-related antigen and CD31 expression, a
profile that can be used to distinguish the tumor from metastatic carcinoma and
vascular tumors. Adenomatoid tumor is clinically asymptomatic and complete
excision is the appropriate management.
B. Multicystic peritoneal inclusion cyst usually arises in the pelvis in women and
is typically associated with lower abdominal pain. It forms a palpable mass
adherent to the pelvic organs that can grossly be indistinguishable from a cys-
tic ovarian tumor, although a subset of cases arises in the upper abdominal
cavity, in a hernia sac, or even in the retroperitoneum. Most cases are associ-
ated with a history of previous abdominal operation, endometriosis, or pelvic
inflammatory disease (Obstet Gynecol Surg. 2009;64:321).
Microscopically, the neoplasm consists of numerous thin-walled cysts lined
by a single layer of bland, flat-to-cuboidal mesothelial cells. The septa and walls
between the cysts are composed of loose fibrovascular connective tissue. The
constitutive cells are immunophenotypically identical to other mesothelial cell
lesions.
Some confusion exists regarding the proper classification of multicystic
peritoneal inclusion cyst, as demonstrated by the fact that the lesion is also
known as multicystic mesothelioma. Tumors in which the mesothelium has
bland cytologic features with no significant atypia have an indolent course
(Cancer. 1989;64:1336), although very rare cases may progress to conven-
tionalmalignantmesothelioma (Am] Surg Pathol. 1988;12:737;] Surg Oncol.
2002;79:243). However, cases in which the cysts are lined, even focally, by
markedly atypical mesothelial cells and/or that harbor areas of conventional
malignant mesothelioma are best considered low-grade mesotheliomas from
the outset (see below).
V. MALIGNANT PLEURAL NEOPLASMS. The World Health Organization (WHO) clas-
sification of tumors of the pleura is shown in Table 11.1.
A. Mesothelial
1. Diffuse malignant mesothelioma (DMM). The WHO recommends the termi-
nology DMM when referring to malignant neoplasms arising from mesothe-
lial cells. The association of the tumor with asbestos exposure is well estab-
lished (Ann Occup Hyg. 2000;44:565). There is usually a long latency
period between asbestos exposure and the onset of mesothelioma, usually
30 to 40 years. While sequences from the highly oncogenic SV40 virus have
been reported in some cases (Clin Lung Cancer. 2003;5:177), an associa-
tion with latent viral infection has yet to be established. Rare cases may be
related to therapeutic radiation exposure or chronic pleural infections.
Patients with mesothelioma usually present with dyspnea, chest wall
pain, and a significant pleural effusion. Constitutional symptoms include
weight loss, malaise, chills, sweats, weakness, and fatigue. While the tumor
may begin as multiple small nodules on the parietal and visceral pleura, it
eventually encases the lung, invades the soft tissue of the chest wall, and
often extends into the mediastinum with encasement of the pericardia! sac
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cytoplasm, although in some cases the cells have more anaplastic fea-
tures. Architecturally, sheet-like, microglandular (adenomatoid), and
tubulopapillary patterns are common (e-Figs. 11.6 and 11.7). Psam-
moma bodies are occasionally encountered.
b. Sarcomatoid mesothelioma is composed of spindle cells that have a
haphazard distribution (e-Fig. 11.8). Some cases resemble fibrosar-
coma, others have a pattern that resembles undifferentiated pleomor-
phic sarcoma. Immunohistochemically, sarcomatoid mesothelioma is
less likely to express cytokeratin 5/6; areas of chondrosarcomatous
or osteosarcomatous differentiation may show positive staining for
actin, desmin, vimentin, and/or 5100. Many cases retain expression
of calretinin. The potential overlap of histologic and immunohisto-
logic features of sarcomatoid mesothelioma with sarcomatoid carci-
noma and various soft tissue sarcomas highlights the necessity for
correlation with clinical and radiographic findings; a solitary mass
should raise concern for another nonmesothelial sarcomatoid pro-
cess. Sarcomatoid mesothelioma is generally thought to have a more
aggressive course than the epithelioid varieties, and so patients with
this subtype are generally excluded from consideration for surgical
therapy.
c. Desmoplastic mesothelioma. By definition, this type of sarcomatoid
mesothelioma consists of scattered atypical cells in a storiform or non-
specific pattern in >50% of the tumor, set in a dense collagenous back-
ground. This subtype is the most likely to be misdiagnosed as organizing
pleuritis in small biopsy specimens.
d. Biphasic mesothelioma. This subtype contains a combination of the other
patterns, in most cases a combination of the epithelioid and sarcomatous
patterns (e-Fig. 11.9). By definition, each component should comprise at
least 10% of the tumor.
2. Well-differentiated papillary mesothelioma is a rare type of mesothelioma
that occurs in a wide range of patients, although most patients are elderly.
An association with asbestos exposure has not been established. Patients
usually present with dyspnea or a recurrent pleural effusion, but rarely with
chest pain. At presentation, the tumor may be either solitary and localized,
multifocal, or widespread.
Microscopically, the tumor features fibrovascular cores (that often have
a myxoid stroma) covered by a single layer of bland, cuboidal-to-flattened
mesothelial cells. Focal areas of limited stromal invasion may be present. In
cases with widespread invasion, DMM with papillary architecture must be
excluded. The distinction is important, since when strictly defined, well-
differentiated papillary mesothelioma has an indolent course with pro-
longed patient survival.
3. Localized malignant mesothelioma is a circumscribed nodular lesion attached
to the parietal or visceral pleura that is usually < 10 em in the greatest dimen-
sion. It is usually discovered incidentally on imaging studies. Microscopi-
cally, the tumor has architectural patterns that are identical to DMM. Some
cases are cured by surgical excision. It is interesting to note that recurrent
tumors often metastasize in a pattern more typical of sarcomas, without
spread along the pleura surfaces.
B. Mesenchymal
1. Epithelioid hemangioendothelioma (termed intravascular bronchioloalveolar
tumor when it arises in the lung) is a low-grade malignant neoplasm of
endothelial cells that can develop at virtually any anatomic site. Primary
cases arising from the serosal surfaces occur, albeit rarely (lnt] Surg Pathol.
2006;14:257).
Microscopically, the lesion is characterized by cords, short strands, and

solid nests of bland, round-to-slightly spindled endothelial cells that have
an epithelioid or histiocytoid morphology and a low mitotic rate (e-Fig.
11.10). Endothelial differentiation is evident by the formation of intracyto-
plasmic lumina (said to "blister" the cells), but distinct vascular channels
are rarely formed. The neoplastic cells are classically embedded within a
chondroid-like to hyalinized stroma. Immunohistochemically, epithelioid
hemangioendothelioma typically expresses a variety of vascular antigens
including CD31, CD34, and Ulex Europaeus antigen; expression of von
Willebrand factor is more variable. Of note, 25% to 30% of the cases
show focal cytokeratin expression, which can lead to an incorrect diag-
nosis of metastatic signet-ring cell carcinoma. In problematic cases, elec-
tron microscopy can be used to confirm the tumor's vascular origin by the
demonstration of Weibel-Palade bodies.
2. Solitary fibrous tumor is classically considered a pleural tumor, although it
is now recognized to occur at virtually any anatomic location. Most cases,
in fact, occur in extrapleural sites. The tumor is most common in patients
between 20- and 70 years old. It is classified as a tumor of intermediate
(rarely metastasizing) biologic potential (see Chap. 46).
Grossly, pleural solitary fibrous tumors can be >20 em in the great-
est dimension, although most tumors are < 8 em. The tumor is usually
well circumscribed although not encapsulated, and has a firm white cut
surface which may show hemorrhage and areas of myxoid degenera-
tion. Microscopically, the tumor is composed of bland plump spindled
cells with a so-called patternless architecture that surround branching
blood vessels of the type typically associated with hemangiopericytoma
(e-Fig. 11.11). The cellularity often varies within individual tumors, and
the background stroma can show areas of myxoid change or fibrosis.
Immunohistochemically, the tumor cells express CD34 and CD99; in a
subset of tumors, the cells also show immunoreactivity for smooth muscle
actin, BCL2, and epithelial membrane antigen; focal immunoreactivity for
desmin, cytokeratin, and/or S100 may even be present. Significant cytok-
eratin expression should raise concern for sarcomatoid mesothelioma or
carcinoma.
Malignant solitary fibrous tumors show an increased mitotic rate (~4
mitoses per 10 high power fields), areas of necrosis, increased cellularity,
and focal marked cytologic atypia, usually with infiltrative margins (Am]
Surg Pathol. 1998;22: 1501). However, the clinical behavior of an individual
tumor is not absolutely correlated with its histologic features.
c. Lymphoid
1. Primary effusion lymphoma is a subtype lymphoma that has a distinct clinical
pathologic setting, presenting as an effusion without an associated tumor
mass. It is defined by the presence of human herpesvirus 8 (Adv Cancer Res.
2001;80:115) and most cases arise in immunodeficient individuals in the
setting of HIV AIDS. Immunohistochemically, primary effusion lymphoma
is usually of null phenotype, although occasional cases express B-cell or
T-cell markers (Cancer. 2007;111:224).
2. Pyothorax-associated lymphoma is a diffuse large B-celllymphoma that usu-
ally presents as a pleura mass in elderly individuals. As the name implies,
it occurs in patients who have a longstanding history of pyothorax, usu-
ally in the setting of pulmonary tuberculosis or tuberculous pleuritis, and is
strongly associated with Epstein-Barr virus infection. Immunohistochemi-
cally, representative B-cell markers other than CD20 are frequently negative,
while aberrant expression of T-cell markers such as CD2 is present (Adv
Anat Pathol. 2005;12:324).
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F!Otne T,..... 'tWD. 8/IIT!tet E. MDIIer-HeriNII .... HI(. Kan1a:CC,«ia:. tt\:lrt1 He!IJJA ~
t'!lfee~tRT..mouta. ~llftd~ T~d'ftl.9£~ fllelw~ 71IJoll"kd.m1HiMtt
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indolent course (Cancer. 1990;65:292). Howevet; those cases that show

evidence of invasion of organ walls or fat (emphasizing the need for thor-
ough microscopic sampling) are associated with progressive disease and a
worse prognosis, and so should be classified as DMM.
2. Multicystic mesothelioma is also known as multicystic peritoneal inclusion
cyst. As suggested by the two very different names, there is confusion regard-
ing the biologic potential of the neoplasm. Tumors in which the cysts are
lined by mesothelium with bland cytologic features with at most only reac-
tive atypia have an indolent course, consistent with a designation as mul-
ticystic peritoneal inclusion cysts (as discussed above). Howevet; cases in
which the cysts are lined, even focally, by markedly atypical mesothelial
cells and/or that harbor areas of conventional malignant mesothelioma are
best considered low-grade mesotheliomas (Hum Pathol. 1991;22:856).
3. DMM arising in the peritoneum is rare; the ratio of pleural to peritoneal
mesothelioma is approximately 10:1 in the United States.
a. The epithelioid subtype is most common (e-Fig. 11.12). Rare cases,
designated the deciduoid type, have a morphology that resembles an
exuberant ectopic decidual reaction (Am ] Surg Pathol. 2000;24:285 ).
Immunohistochemically, peritoneal epithelioid mesothelioma expresses
calretinin, thrombomodulin, WT1, and cytokeratin 5/6, but does not
express MOC-31, Ber72.3, Ber-EP4, CA19-9, and CD15 (Leu-M1); this
panel of markers is the most useful for distinguishing peritoneal epithe-
lioid mesothelioma from peritoneal and ovarian serous carcinomas (Am
] Surg Pathol. 1998;22:1203; Mod Pathol. 2006;19:34; Cytopathology.
2011;22:5).
b. The sarcomatoid and desmoplastic subtypes are very uncommon in the
peritoneum.
B. Epithelial tumors of Miillerian type. The architectural and cytologic features
of these tumors are identical to those of their counterparts that arise within
the ovary, fallopian tube, endometrium, and cervix. Their morphology is
thought to represent another manifestation of the close embryologic relation-
ship between the mesothelium and the secondary MUllerian system.
The criteria for diagnosis of a tumor as of primary peritoneal origin include
the following. First, the ovaries must be of normal in size, or enlarged only
as a result of a benign process. Second, the extraovarian involvement must be
greater than the surface involvement of either ovary. Third, ovarian involve-
ment must be absent, confined to the ovarian surface epithelium without stro-
mal invasion, or involve the cortical stroma with a maximal tumor dimension
of less than 5 x 5 mm2 (Cancer Res. 2000;60:1361).
Primary peritoneal epithelial tumors are currently staged according to the
scheme used for ovarian tumors (Table 31.3).
1. Primary peritoneal carcinoma occurs virtually only in women; the mean age
of the affected patients is the seventh decade. The most common type is
serous adenocarcinoma (e-Fig. 11.13), but clear cell adenocarcinoma (e-
Fig. 11.14 ), endometrioid adenocarcinoma, transitional cell carcinoma, and
even squamous cell carcinomas occm:. Primary peritoneal carcinoma should
possibly be included as a phenotype in familial breast and ovarian cancer
syndromes, although the pattern of genetic abnormalities in primary peri-
toneal carcinomas seems to be distinct from the pattern that is characteristic
of ovarian tumors.
2. Primary peritoneal borderline tumors (tumors of low malignant potential) are
diagnosed by the same criteria as for their borderline counterparts arising
in the ovary (see Chap. 31). Serous borderline tumors are by far the most
common histologic type.
C. Uncertain origin. Desmoplastic small round cell tumor was originally described
as a peritoneal tumor that arises in young men in their second or third decade.
~~IJto"' 111m«
Ma..,.llt rr.othelomtl
<Wm o.!lt.rrnota
-ltic pofialrd~llll ....
Chapter 11 • Serosal Membranes I 18 9
the same histologic types as tumors arising from the pleura. The development
of pericardia! mesothelioma is also associated with asbestos exposure.
B. Germ cell tumors. Intrapericardial germ cell tumors are rare, but occur over
a wide age range, from neonates to the elderly. Intrauterine presentations are
increasingly being recognized in second and third trimester gestations due to the
widespread use of prenatal ultrasound examination. Germ cell tumors of the
pericardium, as with extragonadal germ cell tumors at other sites, are thought
to arise from germ cells that lodge in midline structures early in embryogenesis
along the normal route of migration from the yolk sack to the gonad.
Teratomas account for the vast majority of pericardia! germ cell tumors.
Over 75% of the cases occur in children under the age of 15 years. Teratomas
can achieve remarkable sizes, up to 15 em in the greatest dimension. Grossly,
they usually have a lobulated, smooth surface. Histologically, the vast majority
are mature teratomas that resemble their counterparts arising in the gonads or
mediastinum, in which case the differential diagnosis includes a bronchogenic
cyst. While teratomas are benign, tumors that contain other germ cell elements
(e.g., embryonal carcinoma, choriocarcinoma, endodermal sinus tumor) are
malignant. They are exceedingly rare, and most cases arise in adults.
C. Secondary neoplasms. Metastases are the most common tumor of the peri-
cardium. In a significant percentage of cases, a biopsy (often performed to
establish the cause of pericarditis or life-threatening tamponade), provides the
first evidence that the patient has a malignancy. The most common primary
tumors that metastasize to the pericardium, in decreasing order of frequency,
are carcinoma and adenocarcinoma of the lung, breast, and thyroid; lym-
phoma; and sarcoma. Although lymphatic or hematogenous spread is the most
common route of involvement, direct extension (e.g., by pleural mesothelioma
or malignant thymoma) also occurs.
VIII. MALIGNANT TUNICA VAGINALIS TESTIS NEOPLASMS
A. DMM occasionally arises from the tunica, and rarely, even from hernia sacs.
Grossly, the tumor forms nodules or papillary excrescences. Microscopically,
the most common pattern consists of a prominent papillary architecture with
associated tubular and solid areas.
B. Epithelial tumors of Mullerian type have been reported.
C. Secondary neoplasms. The tunica, as well as the lining of hernia sacs, can be
involved by metastatic carcinoma. In some cases, the involvement of the tunica
or hernia sac is the first manifestation of metastatic disease.
SUGGESTED READINGS
Battifora H, McCaughey WTE. Tumors of the Serosal Membranes. Atlas of Tumor Pathology, 3rd
Series, Fascicle 15. Washington, DC: Armed Forces Institute of Pathology; 1995.
Clement PB. Diseases of the peritoneum. In: Kurman RJ, ed. Blaustein's Pathology of the Female
Genital Tract, 5th ed. New York: Springer; 2002.
Tavassoli FA, Devilee P, eds. Pathology and Genetics of Tumors of the Breast and Female Genital
Organs. World Health Organization Classification of Tumors. Lyon: IARC Press; 2003.
Travis WD, Brambilla E, Miiller-Hermelink HK, et al. eds. Pathology and Genetics of Tumors of
the Lung, Pleura, Thymus and Heart. World Health Organization Classification of Tumors.
Lyon: IARC Press; 2004.
SECTION Ill
Gl Tract
The Esophagus
Dan ielle H. Carpenter and Elizabeth M. Brunt
I. NORMAL ANATOMY. The esophagus, a tubular structure that connects the pharynx to
the stomach, is composed of cervical, thoracic, and abdominal segments. It begins
at the level of the cricoid cartilage and ends at the gastroesophageal junction (GEJ).
The GEJ is the junction of the tubular esophagus and the saccular stomach, and
is distinct from the squamocolumnar junction.
By endoscopy, the esophagus is measured beginning 16 em distal to the incisors
and extends 35 to 40 em to the GEJ. Precise anatomic location within the esoph-
agus is a significant parameter in the differential diagnoses of various pathologic
processes, as well as for staging squamous carcinoma.
The esophageal mucosa is composed of stratified squamous epithelium that
extends distally to the squamocolumnar junction; this overlies paucicellular lam-
ina propria and is delimited by thin muscularis mucosae that have a rich network
of lymphatics (the latter allows for early metastases of relatively superficial malig-
nancies). The deeper submucosa also has a rich lymphovascular network as well as
submucosal glands connected to the lumen by ducts. The deep muscularis propria
is composed of an inner circular layer and outer longitudinal layer; the proximal
third of the muscularis is striated, the distal third is smooth muscle, and the mid-
dle third is a mixture. There is no serosal surface on the esophagus, but rather an
adventitia.
A. Endoscopic biopsy. The standard endoscopic biopsy consists of several small
(1 to 5 mm) unoriented pieces of mucosa with varying amounts of attached
muscularis mucosae. In some cases, the endoscopist may use "jumbo forceps"
to obtain larger fragments (4 to 8 mm); submucosa may be present in these
biopsies. All fragments are submitted and three hematoxylin and eosin (H&E)-
stained slides are examined.
B. Endoscopic mucosal resection {EMR}. EMR is a more conservative approach
than esophagectomy for resection of superficial malignant and premalignant
lesions. These en bloc resections of 1 to 2 em lesions are obtained by elevation
of the mucosa with submucosal saline injection, followed by removal of the
mucosa with variable amounts of attached submucosa. The specimens range
from 1 to 4 em in the longest dimension and up to 1 em in thickness.
EMR specimens are carefully pinned flat for fixation, all deep and radial
margins inked, and the specimen serially sectioned and submitted entirely to
assess not only the mucosal lesion, but also the deep and radial margins (Fig.
12.1). Initially, at least three H&E levels are evaluated.
190
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192 I SECTION Ill: Gl TRACT
Proxrdimal
sh:margin
---- ------ --; '
Uninvolved
stomach
Stomach
Distal shave margin

Figure 12.2 Sectioning of esophagectomy specimens excised for malignancy.
esophagus. Histologically, it can be antral, fundic, or cardiac type mucosa
and is often inflamed (e-Fig. 12.1).*
3. Ectopic tissue. Other ectopic tissues have been described in the cervical
esophagus including thyroid and parathyroid. Sebaceous glands have been
described at all levels of the esophagus; they may be a type of metaplasia
rather than ectopia.
B. Mucosal injury
1. Reflux asophagitis. In patients with gastroesophageal reflux disease (GERD),
the distal esophagus may have a range of appearances from minimal eryw
thema, to hemorrhage with ulceration. Histologically, the damage from the
GERD is seen as intraepithelial inflammation with reactive changes (e-Figs.
12.2 to 12.4) characterized by an increased basallayet; elongation of the
papillae, and intra- and intercellular edema; in severe cases, howevet; ero-
sion can be observed as well.
Chronic gastroesophageal reflux with mucosal injury can also lead to the
development of intestinal metaplasia (Barrett esophagus); this is discussed
under section on "Epithelial Neoplasms and Preneoplasms."
2. Chemical injury. Secondary to exposure to damaging or caustic agents, chem-
ical injury is seen as mucosal damage ranging from mucosal erythema and
*All ewfigures are available online via the Solution Site Image Bank.
,.,. l!o.lll~llllc aao,..&tl•
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b. Herpes simplex virus (HSV). The classic lesion of HSV esophagitis is shal-
low ulceration, ranging from millimeters to centimeters in diameter. Viral
cytopathic change, characterized by multinucleation, molding of nuclei,
and margination of chromatin, are seen most frequently at the edges of
the ulcer (e-Figs. 12.16 to 12.18). Immunohistochemistry (IHC) is con-
firmatory in this regard.
c. Cytomegalovirus (CMV). Focal, discrete ulceration in the distal esophagus
occurs in CMV esophagitis. Granulation tissue from the ulcer base (as
opposed to the edge in HSV) is optimal for identifying viral cytopathic
change in endothelial and stromal cells. Large eosinophilic nuclear and/or
cytoplasmic inclusions can be seen on routine H&E stained sections,
although immunostains for CMV can highlight or confirm the virus.
IV. BENIGN POLYPS
A. Fibrovascular polyps are elongate, often large ("giant fibrovascular polyp"),
pedunculated intraluminal growths that can fill the esophagus and present as
an intraoral mass if regurgitated. Histologically, these polyps are composed of
an edematous, loose stroma with a rich vascular network covered by benign
squamous mucosa.
B. Inflammatory polyps. Often related to GERD, inflammatory polyps are present
near the squamocolumnar junction and may endoscopically resemble adeno-
carcinoma. Howevet; histologically they are benign and composed of inflamed
squamous and/or foveolar mucosa.
C. Squamous papilloma. Also often related to GERD, squamous papillomas can
have a filiform appearance microscopically, if not grossly. Delicate fibrovascular
cores are covered by benign reactive squamous epithelium with varying degrees
of inflammation (e-Fig. 12.19). Although rare cases of virus-related papillomas
of the esophagus have been reported in immunocompromised patients, squa-
mous papillomas are most often seen in patients with GERD.
V. EPITHELIAL NEOPLASMS AND PRENEOPLASMS. The current World Health Orga-
nization (WHO) histologic classification of esophageal tumors is given in Table
12.2. The AJCC Staging guidelines for esophagus and esophagogastric junction
are applied to primary esophageal and esophagogastric junction mucosal tumors,
including the proximal 5 em of stomach (Table 12.3 ).
A. Squamous dysplasia. Endoscopic evidence of squamous dysplasia may be subtle
and most likely identified in association with plaque-like, mass-forming lesions.
Dysplastic squamous mucosa has cells with enlarged, hyperchromatic nuclei
and increased nuclear to cytoplasmic ratios, as well as overall dysmaturity of the
epithelium. Unlike squamous carcinoma, dysplasia is limited by the basement
membrane; however, unlike benign reactive changes, dysplasia lacks cytologic
uniformity and orderly epithelial maturation.
B. Squamous cell carcinoma (SCC) has a range of gross appearances, from ery-
thematous eroded mucosa, to plaque-like growths, to mass-forming exophytic
lesions. It is most commonly found in the middle third of the esophagus, fol-
lowed by the lower then upper thirds. By definition, neoplastic cells extend
through the basement membrane into the lamina propria ("carcinoma in situ"
is a term no longer used; it has been replaced by "high grade dysplasia")
(e-Figs.12.20 and 12.21). Depending on tumor grade, there are varying degrees
of keratin formation in SCC of the esophagus (e-Fig. 12.22), although the
various subtypes of SCC (basaloid, spindle cell, and verrucous) often seen in
other body sites can also occur in the esophagus. In addition to tumor, lymph
node, metastasis, and grading characterizations, tumor location also factors
into tumor staging (Table 12.3).
C. Barrett esophagus. A diagnosis of Barrett esophagus entails endoscopic find-
ings of salmon-colored mucosa of any length above the GEJ with correlat-
ing histologic findings of a specialized columnar epithelium with goblet cells.
a..~ u. ne. ~· I ,••
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NET Gl CCI/dnoldlllln <2 rrlt/10 hill$, :s2%1<167 Unllo!Tn blond timor colh; "-'lu
Will" ..,._ I>01I8m
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10671n- pattetn, nii!CtOitl
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10671nllal" tmek:IO!Iisltures
MANECf<ay rare >20 mi/IO hpll, >20'll'. Neu~ne and a.danoc:udnoma
10671ndox" or sec oomb!ned
Chapter 12 • The Esophagus I 19 9
D. Gastrointestinal stromal tumor (GIST}. In contrast to the stomach, where leiomy-

oma is rare and GIST is common, in the esophagus leiomyoma is common and
GIST is rare. Esophageal GISTs typically present as intraluminal masses in the
distal esophagus. GISTs are KIT(CD117)-positive spindle-cell neoplasms, with
KIT mutations similar to those in the stomach (e-Figs. 12.52 to 12.54).
E. Other benign and malignant mesenchymal tumors. Rare benign mesenchymal
tumors in the esophagus include glomus tumor, lipoma, and Schwannoma.
Rare malignant mesenchymal tumors include rhabdomyosarcoma, synovial sar-
coma, and Kaposi sarcoma. All such tumors have features identical to corre-
sponding primary tumors of soft tissue (see Chap. 46).
VII. OTHER NEOPLASMS
A. Melanoma. Primary melanoma of the esophagus occurs as a pigmented, poly-
poid lesion in the middle-to-distal esophagus.
B. Lymphoma. Primary lymphoma of the esophagus is rare. The most common type
is diffuse large B-celllymphoma, although Mucosa-associated lymphoid tissue,
lymphoma and T-celllymphomas have been described.
C. Metastasis. Although uncommonly a site for metastatic disease, breast, lung,
and melanoma are the most common malignancies to metastasize to the esoph-
agus.
Cytopathology of the Esophagus

I. INTRODUCTION. Indications for esophageal cytologic examination include a sus-

pected neoplasm, an infection, or for surveillance of Barrett esophagus. Mucosal
abnormalities are best sampled with endoscopic brushing for circumferential sam-
pling; endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) may be
employed to sample targeted mucosal lesions, intramural masses, and for staging
of esophageal carcinoma via transmural sampling of paraesophageallymph nodes.
EUS-FNA is the accepted nodal staging modality for esophageal cancer and has an
81% to 97% sensitivity, 83% to 100% specificity, and 83% to 97% accuracy for
malignancy (Ann Thorac Cardiovasc Surg. 2003;9:2).
II. INFECTION/REACTIVE ATYPIA. Brushings are employed in the setting of infection, usu-
ally in a patient with esophagitis, erosions, or ulcers. As in other squamous epithelial
sites, reactive and reparative atypia are characterized by cohesive two-dimensional
(2D) sheets of cells that have slightly enlarged nuclei, smooth nuclear contours, and
conspicuous nucleoli. Features worrisome for dysplasia such as marked nuclear
crowding or a single-cell distribution pattern are lacking. A background of acute
inflammation and some debris are typically present. GERD is generally not an indi-
cation for brushings, but some reactive atypia related to epithelial repair may be
noted.
Reactive atypia is also present in radiation esophagitis. However, radiation
esophagitis will also show the features characteristically present after radiation
therapy, such as large, bizarre cells, multinudeation, and biphasic cytoplasm.
A. Candida! esophagitis displays pseudohyphae and yeast form (e-Fig. 12.55). Oral
contamination should be excluded, especially in the absence of inflammation.
B. Herpetic esophagitis (almost always HSV-1) and shows herpes viral cytopathic
effect, characterized by ground glass chromatin, thick nuclear membranes, mult-
inucleation, and nuclear molding (e-Fig. 12.56). Cowdry type A viral inclusions
(distinct eosinophilic nuclear inclusions surrounded by a dear halo and a thick
nuclear membrane are present (e-Fig. 12.57).
C. CMV esophagitis demonstrates significantly enlarged mononuclear cells with a

large intranuclear inclusion separated from a thickened nuclear membrane by a
clear halo. Occasional cytoplasmic granular inclusions are also present.
Ill. BARRETT ESOPHAGUS. Cytological diagnosis relies on the identification of goblet
cells within sheets of benign glandular cells. The goblet cells exhibit a single large
cytoplasmic vacuole that displaces the nucleus, creating a crescent-shaped nucleus.
The diameter of the vacuole is at least three times the width of a normal columnar
cell (Am] Clin Pathol. 1988;89:493; Hum Pathol. 1997;28:465).
For dysplasia in Barrett esophagus, brushing cytology as a screening tool offers
the advantage of sampling a wider area of abnormal mucosa as compared with
biopsy. The presence of dysplasia and malignancy is evaluated based on architec-
tural irregularities, cell cohesion, and cytological atypia. As interobserver discrep-
ancy is high, any dysplasia identified in cytology specimens should be confirmed by
biopsy (Hum Pathol. 1997;28:465). Dysplastic changes are categorized as low
grade and high grade; high-grade lesions contain more severe cytologic atypia
including an increased nuclear:cytoplasmic ratio, nuclear enlargement, hyperchro-
masia, and nuclear contour irregularity.
IV. NEOPLASMS
A. Intramural neoplasms include GIST, Schwannoma, and leiomyoma. The cellular-
ity of the specimen is usually low. Findings include microfragments composed of
spindle-shaped cells with oval-to-spindled nuclei, fine chromatin, and delicate
cytoplasm. The distinction among these low-grade spindle cell neoplasms rests
on lliC (Am] Clin Pathol. 2003;119:703).
Granular cell tumor may present as a circumscribed or infiltrative process,
is characterized by squamous hypertrophy of the overlying mucosa. The tumor
cells themselves are polygonal with granular, eosinophilic cytoplasm. Immunore-
activity for S-100 may be helpful in establishing the diagnosis (] Gastrointest
Cancer. 2008;39:107).
B. Adenocarcinoma. The specimen is highly cellular, consisting of haphazardly
arranged 3D clusters and abundant isolated atypical cells (e-Fig. 12.58). The
nuclear atypia and pleomorphism are marked. Necrosis may be present. The
distinction from high-grade dysplasia is difficult by cytomorphology alone,
and is quantitative rather than qualitative (Cancer. 1992;69:8; Hum Pathol.
1997;28;465 ).
C. Squamous cell carcinoma (SCC). Well-differentiated SCC displays abundant iso-
lated cells with hyperchromatic and pyknotic nuclei, keratinized cytoplasm with
sharp cytoplasmic borders, spindle- and tadpole-shaped malignant cells, and
necrosis (e-Fig. 12.59). Poorly differentiated SCC shows crowded groups and
isolated cells with enlarged nuclei and coarsely clumpy chromatin; distinction
from poorly differentiated adenocarcinoma may be difficult due to lack of ker-
atinization.
D. Secondary tumors and metastases. The most common form of secondary tumor
spread to the esophagus is by direct extension of a thyroidal, pulmonary, or
laryngeal carcinoma. Rarely, hematogenous metastases to the esophagus have
been reported, usually arising from the stomach, breast, larynx, or pancreas.
The diagnostic work-up rests on recognition of cytomorphology not native to
the esophagus coupled with immunohistochemical studies.
The Stomach
Kathryn M. Law and Elizabeth M. Brunt
I. NORMAL ANATOMY. The stomach, a distensible, ]-shaped organ, is traditionally

divided into five regions: cardia, fundus, body, antrum, and pylorus. The cardia
is a poorly defined region extending up to 3 em distal to the gastroesophageal
junction. The fundus is the region that curves lateral and superior to the level of the
gastroesophageal junction. Below the gastroesophageal junction in continuation
with the cardia and fundus is the body, which extends to the incisura. The antrum
encompasses the distal third of the stomach, begins at the incisura, and extends to
the pylorus. This region grossly and endoscopically has more flattened and firmly
anchored mucosa than the fundus or the body. The pylorus is a muscular zone of
the stomach controlling passage of food into the duodenum.
The mucosal lining is of two types: antral (antrum, pylorus, cardia) and oxyn-
tic (fundus, body). Antral-type foveolar mucosa demonstrates a 1:1 relationship
of surface to deeper mucus secretory glands separated by lamina propria (e-Fig.
13.1).* Oxyntic foveolar mucosa has a 1:4 pit to gland relationship (e-Fig. 13.2).
The mucus neck cells midway along the foveolar fold harbor the regenerative cells
of the stomach. The underlying glands are composed of gastric parietal cells, which
produce acid and intrinsic factor, and chief cells, which produce pepsinogen. In
the antrum, endocrine cells are located just below the surface foveola and include
gastrin-producing G-cells, serotonin-producing enterochromaffin (EC) cells, and
somatostatin-producing D-cells. In the fundus, endocrine cells are located at the
base of the crypts and consist of histamine-producing enterochromaffin-like (ECL)
cells and a small number of EC cells. The lamina propria of the stomach normally
contains a few inflammatory cells including lymphocytes, plasma cells, eosinophils,
and mast cells. The muscularis mucosae, submucosa, muscularis propria, and
serosa of the stomach are histologically similar to that of the intestines.
A. Endoscopic biopsies. Endoscopic findings and pertinent history assist in
histopathologic interpretation. The gross description should include the num-
ber of fragments and overall specimen dimensions to ensure all material has
been analyzed microscopically; three levels stained by hematoxylin and eosin
(H&E) should be prepared for routine microscopic analysis. Some laboratories
order Helicobacter pylori stains at the time of processing; others prefer to eval-
uate the specimens for specific features of infection (see below) before ordering
special stains.
B. Gastrectomy. Gastrectomies may be partial (often including a portion of the
duodenum or esophagus) or total. When the specimen is received, the serosal
surface should be examined for evidence of tumor penetration and the area
overlying the tumor inked (e-Fig. 13.3). After opening longitudinally along the
greater curvature without transecting the tumor (e-Fig. 13.4), the specimen
should then be pinned on Styrofoam (with the mucosal surface facing out) and
fixed in formalin overnight to ensure well-fixed and oriented sections. Standard
measurements include the length of the greater and lesser curvatures, circum-
ferences at the resection margins, and wall thickness. If a tumor is grossly iden-
tified, its location, shape, maximal dimension, and distance from the margins
*All e-figures areavailable online via the Solution Site Image Bank.
201
should be recorded. The tumor should be cross-sectioned and an estimate of the

depth of invasion recorded (e-Fig. 13.5). The presence of any other mucosal
abnormalities should also be recorded. Three to four sections of the tumor
should be submitted to include the deepest invasion and relationship of tumor
to uninvolved mucosa. Sections from other mucosal lesions, and uninvolved
antrum and body, should also be submitted.
If a previously diagnosed adenocarcinoma cannot be confirmed by a grossly
visible lesion, careful examination of the gastric mucosa should be performed to
identify subtle mucosal alterations including erosions and effacement of folds.
Multiple sections should be taken of any of the abnormalities noted, and a
diagram of the sections constructed for later reference.
If the tumor is distant from the margins, a single shave section from the
proximal and distal margins is adequate. However, if the tumor approaches the
margins, multiple radial sections demonstrating the relationship of tumor to
margin should be taken. A careful lymph node dissection should be performed;
all perigastric lymph nodes are considered regional lymph nodes.
Ill. DIAGNOSTIC FEATURES OF NONNEOPLASTIC CONDITIONS OF THE STOMACH
A. Gastritis. The classification of gastritis lacks universal standardization. The most
recent attempt at standardization is the updated Sydney System, which com-
bines topographical, morphologic, and etiologic information to arrive at a
theoretically reproducible and clinically usable diagnosis (Am ] Surg Pathol.
1996;20:1161). However, in most settings, there is not enough information or
sampling to fulfill the requirements of this classification system. Nonetheless,
the system describes several parameters that should be evaluated in a gastric
biopsy including the presence and degree of neutrophilic (active) inflammation,
chronic inflammation, and atrophy (intestinal metaplasia). Additional features
to be noted are surface epithelial damage, lymphoid follicles, foveolar hyperpla-
sia, pseudopyloric metaplasia, pancreatic metaplasia, and endocrine cell hyper-
plasia.
1. Chronic gastritis is the most common pathologic diagnosis in gastric biop-
sies. The diagnosis is associated with a lymphoplasmacytic infiltrate in the
lamina propria, and in many cases is attributable to H. pylori. However,
the number of lymphocytes and plasma cells normally present within gas-
tric biopsies varies with different patient populations, and it is therefore
difficult to determine how much inflammation constitutes clinically signifi-
cant chronic gastritis. Certain histologic features suggest a diagnosis of H.
pylori associated gastritis, including lymphoid follicles with germinal centers
and the presence of neutrophils within the lamina propria, epithelium, or
gland lumens (which is termed "active" rather than "acute" gastritis) (e-Fig.
13.6). Patchy or focal activity may be seen with H. pylori but can also be seen
in inflammatory bowel disease, especially in children with Crohn's disease
(e-Fig. 13.7).
H. pylori can best be identified on routine H&E stain in pits near foci
of exocytosis or surface mucin (e-Fig. 13.8). However, if needed, histo-
chemical (Giemsa) and/or immunohistochemical (IHC) stains can be used
to enhance detection of the organisms (e-Figs. 13.9 and 13.10). The chance
of finding organisms diminishes in the absence of active inflammation or
in regions of intestinal metaplasia, as the organisms do not colonize this
type of epithelium. Additionally, in patients on proton pump inhibitors
(PPis), antral inflammation is milder and H. pylori may relocate deeper
in the oxyntic glands. Finally, treatment with other antibiotics, while not
eradicating the organism, may nonetheless hinder the ability to detect
H. pylori.
After treatment for H. pylori, neutrophils disappear within 6 to 8 weeks.
However, the lymphoplasmacytic infiltrates can persist for longer (up to a
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5. Granulomatous gastritis. Granulomas in gastric mucosal biopsies are not spe-

cific findings. Although they may be associated with gastric involvement in
Crohn's disease (e-Fig. 13.17), they can also be found in a number of other
conditions including sarcoidosis, infections, foreign/endogenous material,
and less commonly with vasculitis or xanthogranulomatosis.
6. Infectious gastritis. A number of bacterial (e.g., mycobacteria, Treponema
pallidum), viral (e.g., cytomegalovirus, Epstein-Barr virus), fungal (e.g.,
Candida, Histoplasma, Mucoraceae), and rarely parasitic (e.g., Cryp-
tosporidium, Giardia, Strongyloides, Anisakis) organisms can infect the
stomach (e-Figs. 13.18 and 13.19). Some of these infections can be associ-
ated with eosinophilic infiltrates and granulomas.
7. Ischemic gastritis. As the stomach has a rich vascular supply from five arter-
ies, ischemic gastritis is very rare. It can be seen in the setting of hypoper-
fusion with findings of erosion, hemorrhage, necrosis, or ulceration (e-Fig.
13.20).
8. Graft versus host disease (GVHD} in the stomach, as in other parts of the
gastrointestinal (GI) tract, shows gland apoptosis and destruction (e-Fig.
13.21).
B. Miscellaneous conditions
1. Foveolar hyperplasia is not a specific finding. It may be seen in antral biopsies
of chronic gastritis and is characterized by edematous, elongated, villiform
foveolar lining of the mucosa (e-Fig. 13.22).
2. Reactive or chemical gastropathy is characterized by foveolar hyperplasia
with tortuous gland outlines and smooth muscle fibers within the lamina
propria (e-Fig. 13.23 ). Although focal activity may be present, inflammation
is typically mild to absent. However, these histologic features are nonspecific
and can also be associated with bile reflux or with medications, particularly
nonsteroidal anti-inflammatory drugs (NSAIDs), which can also cause ero-
siOns.
3. Menetrier's disease is endoscopically characterized by giant gastric folds
mainly involving the body and fundus. The histologic findings include
marked foveolar hyperplasia and glandular atrophy (loss of oxyntic mucosa
in the body and fundus), and may be reminiscent of hyperplastic polyps
(l-IPs). Clinically, patients present with protein-losing gastropathy and low
acid production.
4. Parietal cell hyperplasia due to PPI use or Zollinger-Ellison (ZE) syndrome
presents secondary to hypergastrinemia, with an increase in size and number
of parietal cells resulting in enlarged mucosal folds in the body and fundus
(e-Fig. 13.24). Parietal cells can be found as high as the junction with the
foveolar epithelium. Some of the glands may be cystically dilated reminis-
cent of fundic gland polyps. Foveolar hyperplasia is absent. Hypergastrine-
mia also causes endocrine cell hyperplasia and can result in neuroendocrine
tumors (NETs). The final distinction of PPI use or ZE syndrome is based on
the clinical setting.
5. Gastric antral vascular ectasia, also known as watermelon stomach because
of its endoscopic appearance, is characterized by dilatation of mucosal cap-
illaries in the gastric antrum with or without fibrin thrombi (e-Fig. 13.25),
with associated mucosal smooth muscle fibers. Similar changes can be seen
in portal hypertension. However, portal hypertensive gastropathy involves
the body and fundus and does not classically show fibrin thrombi.
6. Mucosal calcinosis. Calcifications are usually found incidentally in mucosal
biopsies from patients with renal failure or in organ transplant recipients
(e-Fig. 13.26).
7. Pseudomelanosis results from iron deposition in mucosa in patients taking
ferrous sulfate (surface mucosa) or in patients with hemochromatosis (full
Chapter 13 • The Stomach I 2 05
thickness or deep) (e-Figs. 13.27 and 13.28). In iron pill gastritis, there may
be mild inflammation or erosions (e-Fig. 13.29).
8. xanthelasma or xanthoma is characterized by collections of lipid-laden foamy
macrophages in the lamina propria and is significant for its potential con-
fusion with poorly differentiated signet ring cell adenocarcinoma on biopsy
(e-Fig. 13.30). In problematic cases, the diagnosis can be clarified with lliC
stains; positivity for pan-cytokeratins and CDX2 is seen in most adenocar-
cinomas, whereas positivity for CD68 and CD163 would favor xanthoma.
IV. DIAGNOSTIC FEATURES OF COMMON GASTRIC POLYPS
A. Hyperplastic Polyps (HPs) are composed of elongated, dilated, branching foveola
in association with inflamed, edematous lamina propria. These polyps can show
surface erosions, regenerative changes, and intestinal metaplasia (e-Fig. 13.31).
In addition, foci of dysplasia or invasive carcinoma are present in ""'2% of HPs.
The likelihood of these findings increases with polyp size, especially in those > 2
em. HPs may occur in a subset of patients with Osler-Weber-Rendu syndrome.
B. Fundic gland polyps are composed of cystically dilated spaces partially lined
by parietal cells which may be hyperplastic or attenuated (e-Fig. 13.32). They
can be syndromic, where they represent the most common gastric lesion of
familial adenomatous polyposis (FAP), or sporadic. A significant proportion of
syndromic fundic gland polyps are associated with low grade or indefinite dys-
plasia. However, high-grade dysplasia and invasive carcinoma are exceedingly
rare. Sporadic fundic gland polyps are sometimes associated with PPI treatment
for gastroesophageal reflux.
C. Inflammatory fibroid polyp (IFP} is most commonly found in the gastric antrum
and consists of a submucosal collection of bland spindle cells, which are charac-
teristically CD34 positive and c-kit negative, in a background of dilated vascular
channels and mixed inflammation, which often contains abundant eosinophils
(e-Figs. 13.33 to 13.35).
D. Hamartomatous polyps can be seen in patients with Peutz-Jeghers syndrome,
juvenile polyposis, and Cowden's disease. The histologic features of many of
these polyps may be reminiscent of hyperplastic gastric polyps.
V. DIAGNOSTIC FEATURES OF COMMON NEOPLASMS OF THE STOMACH. The World
Health Organization (WHO) histologic classification of gastric tumors is given
in Table 13.2. The American Joint Committee on Cancer (AJCC) staging schema
is given in Table 13.3. Molecular tests used in the work-up of stomach specimens
are listed in Table 13.4.
A. Adenoma and dysplasia. Unlike colonic adenomas, most gastric adenomas are
not sporadic but rather arise in the setting of chronic gastric injury. Adenoma
and dysplasia are distinguished by architecture: polypoid = adenoma, flat =
dysplasia. Both can contain either low-grade (e-Fig. 13.36) or high-grade dys-
plasia (e-Fig. 13.37) defined by cytologic features and architectural complexity.
There are three types of stomach adenomas: intestinal type (e-Fig. 13.38),
resembling those in the colon and containing goblet cells and/or Paneth cells,
and two gastric types, foveolar- and pyloric-type adenomas. Foveolar-type ade-
nomas are composed of foveolar epithelial cells containing neutral mucin with
apical mucin caps, stain positively for MUC5AC, and are sometimes associated
with FAP. Pyloric gland adenomas are composed of tightly packed pyloric gland-
type tubules with a single layer of cuboidal to low columnar epithelium that
shows round nuclei and pale to eosinophilic ground glass cytoplasm, and that
stains for MUC5AC and MUC6 (e-Fig. 13.39). Pyloric gland adenomas have
been shown to occur more commonly in women and are more frequently asso-
ciated with autoimmune atrophic gastritis (Am] Surg Pathol. 2009;33:186;
Virchows Arch. 2003;442:317). Pyloric gland adenomas, as is the case with
intestinal-type adenomas, are also associated with a higher rate of dysplasia
and malignant transformation than gastric foveolar-type adenomas (Am] Surg
Pathol. 2002;26:1276).
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>5 em, >5 mlosaiiSO HPf1 HW> nl""'l't pot.enU.~ , _ , . ntl6ortunwof41111ed
ITIOflltay,4').&!1
mu.tatloao, whlchtfi=l~~< ~ .....u. tbat.cqulo.te cclJ ~

~ II>Oiilier, aDd tum.d Stlldict ht.vt ehown vuyiog ~ ~ tht;
ncr imt.tilill> ~ (XI>
""~"""".-,
-*
tWW~; Hit_, 11 ~ p:rWet 'beutr
I'DGF.RA ....,18 .,....,. thow primory t<Sisw-, aD4 oob:r
d:it &D.C! I'DGI'ltA m - f.U in btnotcU.
IDdi.ridaalo wilb fwmiliol t>htmut&liom4m:IJ>p GIST• atat!ighd.r,_
qc wt may ba.c .a:roltl£ocol .U...... GiSI'..., aloo bt ...., ill padcata w:!dl.
Camqll:lad/4yad ....t ~lotio Type I, wt ! l a o - t=.d ~he
,...,.. _m
1X Pttr.lfytl.l"n()r CUt not b&Ma.e!!Md
ro No oMienoo"' 1JIIrna.ytumor
T1 Tumor2anoriMI
T2 TUtnOf >2 Q1l blt I"Mrt >5 em
l3 TUrr>Of>5cmi>IAnot>l0<:m
T4 1\lmor >10 an 1'1 .,..,..,. dl""""""
...,..., ....... -Oil
~ Rl!PNIIJmJilmc!es C1111nol b o -
NO Noreafonll tlfnpllnoclo-
Nl RIPNIIJmJil,_ -.Ia
. _..., ..... 011
MQ
Ml
III.P ....... IIIIIII!UIItl
Slqj:IIA TlcwT2 NO 1110 Low
SQIIB T3 NO 1110 Low
SQIII Tl NO 1110 Hl!h
T2 NO 1110 Hi&ll
T4 NO 1110 Low
Slqti!IA l3 NO 1110 HIIJII
SIQ:II!IB T4 NO 1110 Htpl
SQIIV AIIYT Nl 1110 Any""'
AIIYT NO Ml Any""'
C111ptw n • T~t-t l!itMe~e~t I 211
<-~tit mlll&tion Jl<llllift. Palia1Zi<: G!Sfr, lhov,sll Gftm ohow pooicift for c-IO:t
_.,..;on by IHC, olao laid CD be <-~tit mutaJiDo noptm:.
Muoy p.oU!c GISI'a bchm: full o-.uaD.lll.dobt ..,•• ...., ....t thctc <2.""
ue~ll!li~~H~GISTadoluvc«~CD-·
cuize, whlch caD C)("«t m:n 10 CD 20 J'Wf d"!cr iaiiW fU111ClV• A number of
"""""""'h.&.. hem clm:lopal CD rille malify GIST, and proposal pidrtinrs
o.....t oo o.ltlldy of > 1SOO 1!1111D< GISTa wilhi""B"U:zm C..U.,.,...lll' ..., lirted
ill Toble 13.6 tAm J Smg Pli/JxJ. 200S;l~..S2). The <\I'CC lll4l&illll tw ~
GISTalllD. Table 13.7.
!. ,,..,....,_. kolll ~JIIIa- at••--'-"IJ'IIIIIolllllluue{IW.f)~·
il«~~c:rtr;~•odallJmllh=oompottdof~yhta:t~-n
Jl.celiJ (..l'ijJ. US). U,pto CII.Oo<bird ofpdli<: MAmmay ohow plumoaqtr>id
diff~. The GI INCt i& llu. moot """"""n oil!> m MALT 111DPhom.a
(SO% of.,....) &lldlhen0D!.41d! i.e th.e_n...,_ Gl w(5S% of cua).
Table 13.8 dcoc:dbca ~ u.oc:ful lA d!mngnlehl . . MAll' lym•
phOil'A from cbR.olc gaotr1t1o (..l!lc. U.54). Go.auic MALT lymph""'" uo
typiully poo«iti.. tw GD20 (..V.., U.S$), CD~ CDll, &l1d GD3S, b'atn.cg·
- &.r GDS, Cli1Q, ADd GD23. Som.e pm;.: MAil" lympli.CIDIAI are CI>43
politi.. (..F;s. U.S'6). D~ of li8ht choin ramclicm is lu:lpful ill
divri..WID!g MAll' lymphoma fmm .-:Uvt .....,.. G:ar1zi<: MALT lym-
ph.oma.o boo"' bem 1uociaa:d with H. /11lari ia.foai.cm; bowevtt, the d,C!a' of
dttt.elio&H. nkr/dc=aaeowith~CD~bomo,...diOIIICIU'OJ'OO•
iii.. ~ will be~.. fo< fl. ~ ia hia!O!"tho. ~The
"""* pollIll
llltlll!lfc1'lltlr.
l""pl\olll folll<io
"'*•llrc:dontr.rtson
ln-lbl~r ~photy!al
MN.T IJIIII!G•
f""'U""
Mlybop-.,.nl
SINIto lnt.wmed11111n lb,
~
Mq
-
I>&-
S..ll•na round, n'lllura
fmolp.jlr n u - - .
~
Pl'odcmnont ..-nt In S!>ll110, uoual~llmbd to
~phold l>lklou!'od \rm~ loll-, do not
lrnslfolkullr- may 0001<1>-coo
-C043
T ~pllo<:jQo (poon"" lor Vlrkbla In nLITiber, K:IIUenlcl
-le. . bon-""'-
Pn!domNnt. dltflnoly '""'""
CD3) the krrina ~and
lntelfdli::IJ\ar • -
In nt.mbor, UIWIJt ~It ll"'''"N!!I, diiPu,.,Y
pn!ISI!II'It., tki!!tlunfna
lnJn.ltplll\llklm, -ll&trt proprll,llck 11atrt chlln
dloln- NllllrlciSon
Ron> ond '"""""""""'~ t11o
USIIol\r poo'lnont tho
111111:ra111& \rm!>hold .... lnlh- ~phold «>lh
uo B«>liund folm •ro T .....nd lnd~lt
........., ala!'od"'r distl1bWd, ~·"'
-...:tli>n- d - nol404dont
-pylori Miybop-...nl M1Jb6~
mk::nxqlnbml
ln111h11on of mUCJ\ali>
mu..,_ bY~Phold...,
most common recurrent cytogenetic abnormality in gastric MALT is t(11;18),

which has also been associated with resistance to H. pylori eradication therapy.
MALT lymphomas tend to be indolent, slow to disseminate, and responsive to
radiation therapy. Even involvement of multiple sites and the bone marrow do
not portend a worse prognosis. However, if solid areas or sheets of large cells
are present, the tumor is more appropriately diagnosed as diffuse large B-cell
lymphoma.
Cytopathology of the Stomach

I. INTRODUCTION. The indications for cytologic sampling of the stomach include the
presence of an inflammatory process or a neoplasm. Mucosal lesions can be sampled
by endoscopic brushing cytology, and intramural lesions by endoscopic ultrasound-
guided fine needle aspiration (EUS-FNA). Brushing cytology for malignancy has
a sensitivity of 85% to 93% and a specificity of 99%, both comparable to the
sensitivity and specificity of endoscopic biopsy. However, brushing cytology and
biopsy are best considered complementary for detection of malignancy (Acta Cytol.
1988;32:461; Acta Cytol. 1990;34:217). EUS-FNA for malignancy has a lower
sensitivity and specificity (Gastroenterology. 1997;112:1087), which most likely
reflects limitations of the technique due to inadequate sampling.
II. INFLAMMATORY PROCESSES. Antral mucosa brushing cytology with Papanicolaou
stain is a sensitive, accurate, and simple procedure for investigating the presence
of H. pylori infection in cases of gastritis. The bacteria presents as curved and S-
shaped rods with basophilic staining properties (World] Gastroenterol. 2005;11:
2784).
Ill. NEOPLASMS
A. Adenocarcinoma.
1. The smear of intestinal-type adenocarcinoma is hypercellular, consisting of
haphazardly arranged three-dimensional cell groups and atypical single cells.
The malignant cells show nuclear enlargement, hyperchromasia, and irregu-
lar nuclear membrane contours (e-Fig. 13.57). A necrotic, dirty background
is often present.
2. The cytologic diagnosis of diffuse-type adenocarcinoma is difficult due to
scarcity of the malignant cells; when present, the characteristic signet ring
cells demonstrate an intracytoplasmic vacuole that indents the nucleus into a
concave shape (e-Fig. 13.58), with associated nuclear hyperchromasia. The
differential diagnosis of the atypical cells in diffuse-type adenocarcinoma
includes histiocytes and goblet cells (Diagn Cytopathol. 2006;34:177).
B. Gastrointestinal stromal tumor (GIST). The smear shows microfragments and
sheets of spindle cells with moderate to high cellularity (e-Fig. 13.59), intact
single spindle cells, and abundant stripped nuclei. The spindle cells have spindle
to oval nuclei, fine chromatin, and abundant delicate cytoplasm with indistinct
borders (e-Fig. 13.60). Nuclear atypia, mitosis, and necrosis may be identified
occasionally. The epithelioid variant demonstrates large epithelioid cells with
round nuclei and distinct cell borders. GIST cannot be graded based on cyto-
logic specimens. Immunostains are required for a definitive diagnosis to exclude
other submucosal spindle cell neoplasms that possess similar cytomorphology
(Cancer. 2001;93:269; Am] Clin Pathol. 2003;119:703 ).
C. Neuroendocrine Tumor G1 {Carcinoid). The cytomorphology of carcinoid tumor
(low-grade neuroendocrine carcinoma) is identical to that of the tumor at other
sites. The smears are cellular and composed of loosely cohesive clusters and
Chapter 13 • The Stomach I 2 13
isolated cells with characteristic salt-and-pepper chromatin. Focal and variable

endocrine atypia are easily identified.
D. Malignant lymphoma. The cytomorphology varies among the subtypes of gastric
lymphoma. In general, the smears show isolated lymphoid cells exhibiting differ-
ent degrees of atypia and monotony (the detailed cytomorphology of different
lymphomas is discussed in the cytology section of Chap. 43). Precise diagnosis
and classification require ancillary studies, which can be applied to EUS-FNA
material (Acta Cytol. 1994;38:169).
The Intestines,
Appendix, and Anus
ILKe Nalbantoglu and Elizabeth M. Brunt
I. NORMAL ANATOMY
A. The small intestine is 6 to 7 m long and divided into the duodenum, jejunum,
and ileum. It begins at the distal gastric pylorus and ends at the ileocecal valve
and is lined throughout its length by villous mucosa. The individual villus is a
slendet; fingerlike projection with a variable length-to-crypt ratio ranging from
3:1 to 5:1 (e-Fig. 14.1).* The epithelium consists predominantly oftall, colum-
nar absorptive cells that have basally situated nuclei, eosinophilic cytoplasm,
and an apical brush border. The absorptive cells rest on a visible, refractile
terminal bar. Other cell types of the intestine include goblet cells, crypt cells,
basal cells, Paneth cells, and endocrine cells; the granules of Paneth cells are
refractile, eosinophilic, and supranuclear, whereas those of endocrine cells are
smallet; eosinophilic but nonrefractile, and infranuclear. The lamina propria
contains mixed inflammatory cells including plasma cells, although neutrophils
are restricted to vascular channels. Peyer's patches, which are lymphoid aggre-
gates, are distributed throughout the small intestine mucosa. Intraepithelial
lymphocytes (IELs) are normally rare (no more than one per five enterocytes
at the tips of the villi), although an increased IEL density may be seen in epithe-
lium overlying lymphoid aggregates or Peyer's patches in the distal ileum where
the associated villi may be shortened or even flattened. Shortened and broad-
ened villi may also be seen in duodenal mucosa overlying Brunner's glands.
B. The large intestine, or colon, is 1 to 1.5 m long and consists of the right and
left colon. The right colon is further subdivided into the cecum, ascending,
and proximal transverse colon; the left colon consists of the distal trans-
verse, descending, and sigmoid colon, and the rectum. The mucosa contains
evenly spaced, nonbranching crypts arranged perpendicularly to the lumen
and extending from the surface to the muscularis mucosae (e-Fig. 14.2). Occa-
sional branching crypts or slight crypt architectural distortion may be seen in
the rectum and sigmoid colon, and in areas adjacent to lymphoid aggregates.
Paneth cells may be seen until the mid-transverse colon and, as noted earliet;
IELs can be prominent in epithelium overlying lymphoid aggregates. Howevet;
there are no villi, the lining epithelium has no microvilli and does not rest on
a terminal bar, and goblet cells are more numerous (particularly in the left
colon). The lamina propria components are similar to those of the small intes-
tine, although the lamina propria is denser in the right colon, and muciphages
(mucin-containing macrophages) are more common in the lamina propria of
the left colon.
The entire small and large bowel mucosa rest on muscularis mucosae. This
smooth muscle layer delineates the lamina propria from the submucosa which
comprises loose fatty tissue with a rich angiolymphatic supply. The inner and
outer layers of the muscularis propria lie below the submucosa and are sep-
arated by the ganglio-neuronal Auerbach plexus. The entire surface of the
214
Chapter 14 • The Intestines, Appendix, and Anus I 2 15
intestines is covered by visceral peritoneum (serosa) up to the distal portion of

the rectum.
C. The appendix is a tubular organ that extends from the cecum. It has an average
length of 7 to 10 em and has a mucosa that is similar to the large intestine,
except for the presence of more prominent lymphoid aggregates which often
have well-formed germinal centers. The appendix also has a poorly developed
muscularis mucosae that may be interrupted by lymphoid aggregates.
D. The anal canal, the terminal 3 to 4 em of the gastrointestinal (GI) tract, is an
anatomically complex area (e-Fig. 14.3). This chapter employs the surgical
anal canal terminology as defined by the most current WHO classification of
tumors (discussed later). The mucosa lining the upper portion of the anal canal
is a direct extension of the rectal mucosa (colorectal mucosa). The mucosa
lining the middle portion of the anal canal (the so-called anal transitional zone
(ATZ), a 0.5- to 1-cm segment above the dentate line) has the features of both
metaplastic squamous mucosa and urothelium. Submucosal and intramuscular
anal glands open into the ATZ via anal ducts that are also lined by ATZ
epithelium. The mucosa of the distal anal canal, which extends from the dentate
line to the anal verge, consists of specialized nonkeratinizing squamous mucosa
with melanocytes. It is distinguished from the perianal skin by the lack of skin
appendages.
II. GROSS EXAMINATION AND SPECIMEN HANDLING
A. Endoscopic biopsy. When processing the specimen, it is important to record
pertinent clinical history and endoscopic findings. Biopsies are typically small
fragments of mucosal tissue in the range of 1 to 5 mm in greatest dimension
that do not need to be inked or subdivided. Important gross descriptors are the
number, size, and the size range of the biopsy fragments. In cases where numer-
ous fragments are present, an estimate for the number and the dimensions in
aggregate should be given (documentation of the number and size is important
to ensure that the biopsies are adequately represented on the slides). Routine
microscopic examination of endoscopic biopsies usually entails examination
of three hematoxylin and eosin (H&E)-stained levels.
B. Suction biopsy of the rectum, which makes possible sampling of the submu-
cosa, is used for evaluation of Hirschsprung's disease. After processing, the
tissue is serially sectioned in its entirety, but initially only every third level is
H&E-stained; if no ganglion cells are identified in these slides, the remaining
sections are stained and examined. In some labs, frozen sections are performed
for histochemical staining by the acetylcholinesterase reaction to identify pro-
liferating nerve fibers in the lamina propria and muscularis mucosae.
C. Polypectomy specimens should be described and measured. The need for inking
of the resection margin is a controversial topic; in practice, it is often difficult
to do since the stalk retracts and thus may be hard to identify grossly (although
the cauterized base can be easily identified microscopically). The specimen is
bisected or serially sectioned depending on its size, and entirely submitted. Sec-
tioning should follow the vertical plane of the stalk to maximize the evaluation
of the polypectomy margin. At least three H&E levels are examined.
D. Endoscopic mucosal resections can be single or multiple fragments. All dimen-
sions are recorded and the mucosal surface described. Inking of the margins
is a matter of choice since cautery artifact will be noted at the time of micro-
scopic evaluation for deep and mucosal margins, and since ink may actually
artifactually extend along non-marginal mucosa. The entire specimen is serially
sectioned, and the fragments are oriented and entirely submitted.
E. Bowel resection.
1. Neoplastic. Tumor resections include segmental resection of a portion
of small or large bowel, ileocolectomy, low anterior resection (LAR),
abdominoperineal resection (APR), total colectomy, and total mesorectal
excision (TME). The portion of resected bowel is oriented, and the length,
diameter (or circumference), and wall thickness are measured. The length
and diameter of the appendix and the dimensions of mesentery are also mea-
sured, if present. The external surface (serosa in most cases) of the bowel is
inspected for tumor involvement, perforation, adhesion, and fat wrapping.
For TME specimens, the grossly observable completeness of the mesorec-
tum is evaluated as "complete," "near complete," or "incomplete" before
opening the bowel (see College of American Pathologists Cancer Protocols
for Colon and Rectum at www.cap.org). The bowel is opened longitudinally
along the antimesenteric border, unless this would mean cutting through the
tumor.
The maximal size of the tumor and the distance to the proximal and
distal resection margins, or to the closest margin in unoriented specimens,
are documented. After fresh tissue is collected for biobanking (as needed),
the specimen is pinned out on a wax board (mucosal side up) and fixed
by submerging in 10% formalin overnight. The tumor is then sectioned
to assess the depth of invasion; blocks for microscopic examination are
taken to include the area of deepest penetration and the relationship to
adjacent, grossly nonneoplastic mucosa. Additional sections include prox-
imal and distal resection margins; if tumor approximates the margin, such
as in APR or LAR specimens, the margin should be inked and multiple
sections perpendicular to the margin submitted; if the inked radial margin
is not included in the tumor sections, one separate radial margin section
should be submitted. One random section from normal-appearing bowel,
and sections from any additional gross lesions (such as separate polyps),
should also be submitted. If the appendix is present, it is handled as an
appendectomy specimen as described later.
The mesentery and soft tissue are also dissected for lymph nodes (many
nodes are located along large vessels), and the number and size range of
identified nodes are recorded. Small lymph nodes can be submitted in toto
without sectioning. Larger nodes are serially sectioned and the cut surfaces
examined; if metastatic carcinoma is grossly appreciated, as evidenced by
a white and hard cut surface, the size of the metastatic deposit should be
recorded, and one representative section from each grossly positive node
should be submitted. If the cut surfaces of the nodes are tan, soft, homo-
geneous, and lack gross evidence of metastasis, the entire node should be
submitted for microscopic evaluation. Although a minimum of 12 nodes is
required by established staging criteria, all nodes that can be found should
be submitted. Fewer nodes may be acceptable for small specimens, for cases
that have received preoperative chemoradiation, and for APR or LAR spec-
imens (because lymph nodes are less numerous below the peritoneal reflec-
tion). However, when fewer than 12 nodes are identified, a second attempt
to dissect lymph nodes is strongly recommended (and should be documented
in the pathology report).
2. Nontumor bowel resections
a. For polyposis specimens, pinning and gross examination are similar as
for tumor specimens. Sampling focuses on the largest lesions, lesions with
a distinct or worrisome gross appearance (including firmness, ulceration,
and adherence to the wall), and flat and depressed areas of mucosa.
b. Resections for inflammatory bowel disease (IBD), particularly for ulcera-
tive colitis (UC), require sequential sections spaced every 10 em. The sec-
tions include transition regions between normal-appearing and diseased
segments, distal and proximal margins, and representative inflamma-
tory polyps. Any focal lesions (such as areas with raised mucosa), fistula
tracts, and strictures are sampled. The appendix, if present, is handled
as an appendectomy specimen, as described later. Representative lymph

nodes are submitted, but there is no need for extensive sampling unless
a carcinoma is suspected or identified.
c. Miscellaneous resections. In the case of ischemic necrosis, the mesen-
teric vessels should be carefully examined and sampled to evaluate the
possibility of thrombosis, embolization, or vasculitis. For penetrating
traumatic injuries, inspection for possible entry and exit wound sites is
important. It is also important to grossly and microscopically examine
the proximal and distal resection margins for tissue viability.
When proctectomy or rectosigmoid resection is performed for
Hirschsprung's disease, the distal margin is usually indicated by the sur-
geon. Sequential sections every 1 to 2 em from distal to proximal should
be submitted to achieve an accurate estimation of the aganglionic region.
F. For appendectomy specimens, the length, diameter, surface appearance, and
dimensions of the specimen, including the mesoappendix, are recorded. For a
nonneoplastic appendix, one half of the longitudinally bisected tip, the proxi-
mal margin, and two cross-sections are submitted in a single cassette, to include
the mid-portion and any possible perforation. For a neoplastic appendix, after
photography and gross description, the specimen is bread loafed and submit-
ted in its entirety. In cases of pseudomyxoma peritonei, any associated mucin
is also submitted in toto.
G. Anal biopsy is treated similarly to other GI biopsies.
H. Endomucosal resection {EMR} specimens are processed as are esophageal EMR
specimens (as discussed earlier).
I. Hemorrhoidal excision requires one section.
Ill. DIAGNOSTIC FEATURES OF NONNEOPLASTIC CONDITIONS OF THE SMALL INTESTINE
A. Congenital anomalies
1. Heterotopic gastric mucosa typically presents as a small nodule or sessile
polyp in the duodenal bulb and consists of full-thickness fundic-type oxyntic
mucosa. It differs from foveolar surface metaplasia in which the surface
epithelium of the duodenal mucosa is replaced by gastric foveolar cells
(e-Fig. 14.4), and which is often associated with duodenitis secondary to
Helicobacter pylori infection.
2. Heterotopic pancreas presents as a mass lesion in the duodenum and is
composed of ducts and acini, with or without islets.
3. Meckel's diverticulum results from persistence of the proximal portion of
the vitelline duct and is always located on the antimesenteric border of
the ileum. Associated heterotopic pancreatic tissue or gastric mucosa is
common. Congenital diverticulum is a rare occurrence in the duodenum
and jejunum (e-Figs. 14.5 and 14.6).
4. Malrotation, stenosis, atresia, duplication, and defects of the musculature are
rare. Duplications can be cystic or tubular; about 75% are not contiguous
with the lumen of the associated bowel segment. Duplications contain all
the layers of the segment from which they have arisen, although mucosal
heterotopias may occur. Any segment of the GI tract may be involved, and
duplications are not associated with other anomalies. Neuroenteric rem-
nants most commonly occur in the cervicothoracic or lumbosacral regions
and also contain all the layers of the originating segment; however neuroen-
teric remnants originate from the dorsal midline and are associated with
other congenital anomalies. Neural elements may be observed, primarily in
lesions approximating the spinal cord.
B. Malabsorptive disorders
1. Celiac disease, also known as gluten-sensitive enteropathy, celiac sprue, or
nontropical sprue, is an immune-mediated disorder secondary to hypersen-
sitivity to a-gliadin. Classic histologic features include villous atrophy, crypt
21. I SECTION llh <II lAACT
'' 1,1,! I ltlb I CoftdltlriM llllt can llllrnle GlulleoterMitiM Ellt.ropa!IIJ

Ct!Gd !E~I.It6r;Mesi:M)t~
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Alllolmml.llo......,pclhy
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--~~~lnlllrntniiiDIJ df11P
-P1fo/llnlocllon
apparent. A complete lack of goblet cells and/or Paneth cells may be seen
in some cases. Although some patients have anti-enterocyte and/or anti-
goblet cell antibodies, serologic tests are not routinely employed. A clinical
response to steroids may help establish the diagnosis.
5. Eosinophilic gastroenteritis involving the small intestine exhibits histologic
features similar to those described for eosinophilic gastritis (e-Fig. 14.11).
There may or may not be villous blunting, but IELs are usually not increased.
Parasitic infestations, food allergy including cow's milk protein intolerance,
a drug reaction, connective tissue disorders, and a neoplasm should be
excluded.
6. Common variable immunodeficiency is characterized by the absence of lam-
ina propria plasma cells (e-Fig. 14.12). Other features may include a vari-
able degree of villous blunting, intraepitheliallymphocytosis, and lymphoid
aggregates. Infectious agents, particularly Giardia, should be searched for
in these biopsies.
7. Microvillus inclusion disease is a rare autosomal recessive disease causing
intractable diarrhea in infancy. The hallmark of the disease is the loss of
a normal brush border on the luminal surface of the enterocytes. Instead,
the brush border is incorporated into the cytoplasm as apical microvillus
inclusions. The microscopic features can be best demonstrated by periodic
acid-Schiff (PAS) stain, electron microscopy, and immunostains for car-
cinoembryogenic antigen, CD10, or villin. Diffuse villous atrophy is also
present, but an inflammatory response and intraepithelial lymphocytosis
are not evident.
8. Lymphangiectasia, either primary (congenital) or secondary (due to obstruc-
tion), may present as a localized mass lesion or diffusely involve the bowel
(e-Fig. 14.13). The presence of secondary lymphangiectasia is concerning
for an unsampled underlying mass lesion as the source of obstruction, which
should be mentioned in the report.
9. Abetalipoproteinemia features lipid accumulation in enterocytes giving rise
to a clear or foamy appearance. The normal villous architecture is well
preserved.
C. Infectious diseases
1. Tropical sprue and bacterial overgrowth simulate celiac disease but may
involve the entire small intestine with more severe disease distally. Clin-
ical history, including any travel history, is important in establishing the
diagnosis.
2. Giardiasis does not induce significant villous architectural change or an
inflammatory response. The diagnosis is based on the identification of pear-
shaped trophozoites at the luminal surface of normal-appearing mucosa (e-
Fig. 14.14), which can be mistaken as cytoplasmic debris. The organisms
can be highlighted by trichrome and Giemsa stains.
Whipple disease exhibits distended villi due to lamina propria accumula-
tion of foamy macrophages stuffed with the diastase-resistant, PAS-positive
(e-Fig. 14.15), rod-shaped bacterium Tropheryma whippelii. The microor-
ganisms can also be detected by polymerase chain reaction (PCR) analysis
and electron microscopy. Gomori's methenamine silver (GMS), acid-fast
bacilli (AFB), or Fite stains should be performed on these biopsies to rule
out fungal (histoplasmosis) or mycobacterial (due to Mycobacterium avium
intracellulare) infections because the morphology of these infections are
quite similar to Whipple disease on H&E stain.
3. Cryptosporidiosis is characterized by uniform, spherical, 2- to 4-~m bodies
attached to the brush border that appear bluishonH&E stain (e-Fig.14.16).
The organisms may be confused with mucin droplets.
4. Strongyloidiasis is diagnosed by identification of larvae, eggs, and adult

worms embedded in the crypts (e-Fig. 14.17). Eosinophils, sometimes with
Charcot-Leyden crystals, may be prominent. The nematodes most com-
monly infect the small intestine, but also rarely infect the stomach and
colon. Gastric strongyloidiasis may sometimes be associated with infection
by human T-lymphotropic virus type 1, a virus that causes adult T-celllym-
phomalleukemia.
IV. DIAGNOSTIC FEATURES OF POLYPS AND NEOPLASMS OF THE SMALL INTESTINE.
The WHO classification scheme of tumors of the small intestine is given in
(Table 14.2)
A. Brunner's gland hyperplasia, hamartoma, and adenoma may actually be variants
of the same process and consist of expanded lobules of benign Brunner's glands
separated by delicate fibrous septa. They are typically located in the submucosa,
but penetration into the mucosa is common. Cystic degeneration may occur,
which has been termed Brunner's gland cyst.
B. Peutz-Jeghers polyp, while most common in the small intestine, also occurs
in the colon and stomach. It is a hamartomatous polyp characterized by an
arborizing network of smooth muscle supporting benign-appearing mucosa
that may be hyperplastic (e-Fig. 14.18). Most polyps occur as part of an inher-
ited cancer syndrome, but sporadic cases may be encountered. Because syn-
dromic polyps carry an increased risk of cancer, they should always be assessed
for dysplasia.
C. Adenomyoma of the ampulla of Vater exhibits an orderly arranged lobular pattern
of benign pancreaticobiliary ducts in a background of proliferating smooth
muscle. It may coexist with heterotopic pancreas.
D. Adenomas are rare in the small intestine and usually occur in the duodenum.
Multiple adenomas are almost always associated with familial adenomatous
polyposis (FAP). Histologically identical to their colorectal counterparts, they
are classified into tubular, tubulovillous, and villous types. The differential
diagnosis includes gastric surface metaplasia and reparative change.
E. Adenocarcinoma of the small intestine is rare, accounting for only 2% of all
primary GI tumors despite the fact that the small intestine constitutes about
75% of the length and about 90% of the mucosal surface of the GI tract
(Table 14.3). Adenocarcinoma of the small intestine is morphologically indis-
tinguishable from colorectal adenocarcinoma, but most cases are cytokeratin
7 (CK7)-positive which may help resolve the differential diagnosis.
F. Ampullary carcinoma is actually a heterogeneous group of tumors. It arises in the
vicinity of the ampulla of Vater and includes the most common intestinal-type
adenocarcinoma as well as the pancreaticobiliary type. The former has a more
favorable outcome than the latter, although the overall survival of ampullary
carcinoma is better than that of pancreatic ductal carcinoma (which probably
reflects differences in respectability). However, distinguishing the site of origin
is sometimes a challenge (Table 14.4).
G. Neuroendocrine tumor {NET) accounts for one-third of small intestinal tumors.
Duodenal NETs are derived from endocrine cells of the foregut and tend to be
<2 em in greatest dimension and asymptomatic. Gastrin-producing NETs are
associated with ZES (Zollinger-Ellison syndrome) in 40% to 50% of cases.
Distal jejunum and ileal NETs are derived from cells of the midgut; 25% to
30% are multifocal, and clinically they are more aggressive than proximal NET.
Microscopically, small intestinal NET are similar to NET arising elsewhere
(e-Fig. 14.19) and have a very bland cytomorphology; invasion into or beyond
the muscularis propria and/or distant metastasis are the main criteria for deter-
mining malignant behavior. In contrast, poorly differentiated NET exhibit
overt histologic features of malignancy. Tables 14.5 and 14.6 summarize the
current WHO classification and AJCC staging schemes, respectively.
4
C!Nipt• 14 • liM: ltrtMtines, ~i.l ud ANa I 2 21
''I.I Illt!J I WltO H~tto~ot~c Cllulllcaltluf r - of 1M tllllll lnt..Urtt

(lllbllll ~~
Ad....l!lll
Tubular
Woua
TubulcMibus
illol>- 0-ptilollalnooplula~ I<Mrpde
illol>- 0-ptilollalnooplula~ hW! ga.d&
Hoi!III-
Peiii:Hoa!IOI'8I>CtJI>
J.-'lllo llCtJI>
c..t!ol!lll
Ad--~~~~
Mudnow:.tdlnOCIII'Cttoma
S\!I>Ot·ltiJJ: oeii ..reN<n•
Squ""""" ooi.....,I""""
Ad"""'lUI/nOII4 CIICI""""
Meduii!Y<ttdlomo
Und!ll'oAII'ilobid.,..mno
IINIMIM<II• ....IMM
,_urcorlllcemo tum« CNE:rl
NET, Gl C01n:h>ld)
NET,G2
Nourcondccmo .,..nm, (NEl:l
~CIIIINEC
Stn .. CIIII Nre
Mlcod ed.,.,.,"""'*""*no l!lldn""" (l.tANEC)
£C...a.-M~NET
Ollll!loc!Cie 1>0111!1/11llcwne
...
Galrl,...,.
L<el~ ~~-~IIz poplido1nd PPIPI'Y·"""'uotla NETs
..... ,.,...
Som-.pR>dulila NET
u-
Ltlo~
GulrolrboUnllalromal tumor
Ltlomyou"""""
Anp""""•
KaDCOI ..,.,....
Olton
a, Jb nn
8ultlt t.omplloma
8...,.1 bl'nDholllll, uhCIMolllel*>•...t~~-.oltllonmodiiiD botoo>on <lti'U.oo ..,., 8...,.1
~plloma and Bultlt bl'npilomo
Olll'ullo l a r p - ~phoma
Inrn_..,_ tmallln-11 d!w.M lh:Iuc!M ""h""'Ychaln d!Muo)
"'*"llr ~plloma
Mlr&NI"""' bl'nplloma o f m u - ~pldd tl&iwe (MALT ~pllomal
Mlnlle ooll.wntp""""
T....l bl'nDholllll
·-""-
E'IIWC~ ...,..loled T-<ooll,wntph,.... (E'ATI.)
I'1."! t! I
Ij I ~ lhlutallt (1'NIIO . . . kllemt hw tllllll lnttttflllll
"'-'a-m
lX Prm., t1.mo<..nnot be .......~
TO r i o - ol polmuy tvmor
11> Cor<*\Oino 1'1 .au
Tie 1\Jm« I'IO!Id•lomlna _,.
Tlb 'nJMOf fniJII!ldM 1\JbMI.IOO:I&t
T2 TumOf ln\lades muscuiwtl proplta
T3 Tum«-· tlirOI.IIIII the .......,IIJtl Pf'lllN lrbtlle su-..., «Into
nonpofllon- portnu...,ler U..uo ~or rdlo~l
lOti\_,.., 2tm « _..
T4 1\Jmor ~111o ..w:.tal """"*''"or dii"OOdl/ln¥0doo o111or «111/10
or_,... tN!ud• dller ""'"'ohtrtdl-, ,_,,,..
...,....._m em,
>2 lind ahdomlr'oll,... blf""Yol....,..;for
duodenum only, I n - of 1>11\C,.... Of bloducl)
...,....,.,.,. llllttlll)
KIC Relllonollfroph IIOdoo tonnot be ..,. "'.S
r«l rio..,.,.! tlmril n O l l e -
Nl -.llslnl-.lr..,_tlmrii-
N2 -.llsln4ormmr...,...tjmph-
. _..., ..... 011
MQ riodlllol!t m-Ilo
Ml ~nt rruaatu!a
III.P .......
SQIO 11> NO l,f.)
fllqol T1 NO l,f.)
T2 NO l,f.)
SlqtiiA T3 NO l,f.)
SlqtiiB T4 NO l,f.)
Slqj:IIIIA AnyT Nl l,f.)
SQ>I!IB AnyT N2 l,f.)
SQIIV AnyT M:tN Ml
,.-tt
........ ~IW!b::le.tt _.tll'dCI.6It t:Wtua !e. tarJej:unutn al'ld lteu"n. ofh tr "llef'itl\d, tot
53 3
d\JCIC5enutn II\ •.a:wt'«eael'OIM 1a .. ea-c, pert dt:M t~lotaJ/1\.

F<>,. ~ SB, B,rd DR, Cool"""
~. 5rlfl-; 21)10. tJood-" ' - ·
0C:, .. 01, d . A!CC Ciw8r~- 7111 ed. NM ll>1<,
,,, , . . . . (!)
1X Pr!muytumwai'\I"Mrt bo IIJIIIfJIN
ro No-ofp"'""l)'tllmOf
11> Cote!1101110 In ollll
n Tt.mOr lm!...s" ompulo o f - . , spllll'odWof Odd!
T2 Tt.mOr-~1..,;
l3 n.mor_..,.,...
T4 Tl.mor -lllfl...,.,- soft-or Gilt Of lll)looniOfliiN"'
..., ,,.,..._{I, wlnu:tuA>I
Nit R""'""l t)mpll noclotcoMal tou...-

NO No • • I I '.!mph node melallals
Nl Ropnol t)mpll noc1a-
IIGrt••s•••t•
MQ
(1111)
Nodlalanlme-
Ml Dllllnlmo-
S'llltii'M.....
SQIO 11> NO lot)
Slqa lA Tl NO Ill)
SlqoiB T2 NO Ill)
Slqo IIA l3 NO Ill)
5lqo liB Tl Nl Ill)
T2 Nl Ill)
l3 Nl lot)
Slqa Ill T<l AnyN Ill)
Slqa IV ArryT AnyN loll
Cis! M IIIII cr.tt DIIIIIIUDn

~1!)1 b~n.:t. mliJ>IIcoount <2 por 10
llf&tl-noldi(HPFs)•-2'l!. ,;KI
671nd ...
II ~:alirb~nd. mllotio<~>unl~pcr
10 HPf..ndlorHClli IQ 671'1doa
,_uroondooltlo CIII'CI"'""' INECl Ill Mltoei:COIIIt >20 110r 10 HPf"""""'
>2!l!r. IQ 671nd&x
MANEC Tumor db~JIIout 30l(. al
~rpialll: •nd snneoplasllc- od""""-aorNEC
Mr•r tnnor Qlnnot bo -IOd

No Mlan"' <II pffmery tum Of
1\Jmor IWadeo lo/rlnap!Oift oroubmuoos,.nd sl em• lrulm (lmo!l
III1Dolr..ltnnolll; 111n« sl em CemouliriYturncnl
1\Jmor l'iW.deo m~U~<~JioJ1o lli'Olft or >I em., a <orno!llnteo1fnol1umortl;
tumtlf >l ""'(llmpulorytl.mOIII)
T3 TUmDI'- tlrliu&li IIi& lfti*U~ilo Pi'Opffa ln1i> IUiioiiroallllsuo-
~n<il~- ~l\lllttt llultumol'l)tttlnlllld• pcnc:IMS
w ,..,_.,...,,. (ompu!luy 0< duodonol turnort) or Ho
nonpolllu""bsd-
1\Jmor I'IW.deo.,._ol D<doneum C-> ttt lnlladoo dlror .,..,,.
F<>r illiY T, .rid Cml ftr m Llilr* tum""
"Tu,...llrnbd t.o 1111po.jllof- r for ampullly lllJIIjtrqllc; . .rap!111jkima
1-.fM.tlliiiiJII IINIOD
r« floPnol tlmril ·-"'motbea!I•I&Od
NO No ft!j!jonal t,.rrph nod• rnall.I1IOSio
Nl """'""I tlmril noclo m o -
--·llillalll 01)
MO No dllllnt"""""""'
..........
Ml
Slqj:IO
SQII n
Dtawrtrrw•ttWa.
111' NO
NO
1110
1110
SQIIIA '12 NO 1110
fllqo liB T3 NO 1110
SlqtiiiA T4 NO 1110
SlqtiiiB AnyT Nl 1110
Slqj:i IV AnyT AnyN Ml
,;2 an. , ; 5 - HPFa l!cin_,., n - """or turn.....llilod m<floll\r. 0

>2~6~m. <6m~HPFs ~ Olllll,lnilnt pcAotrilal,-""" ...
1\imor-relelod mor1alk\f; 4'11
>S~lOanJ~rn~ Mtdenrte maiQNnt ~\ rnetutuk ra&or
HPfl 111m,...-""'"'ll:y; 25!1.
>lOan, or >5m~HPFt Hl&fi n.II&Nnt pot.llntl.e\ miilld•+ ndoor
tvrnwoflllidDd mor1allf;l; fiO.I30%
4
C!Nipt•14 • liM: ltrtMtines,~i.l udAtlua I 22&
I' I.I II J t! 'l l'IIIMII!Wt. IWattalllll (TNIll) . . .kllllmt CMIIcllllllltl..l

' _ SWolal Tu_. (b:ludi~W StoMa~
,,, , . . . . (!)
1X Pr!muytumwai'\I"Mrt bo IIJIIIfJIN
ro No-ofp"'""l)'tllmOf
n Tl.mor 2 em ......
T2 Tl.l'n()r >2 an but rttt >San
l3 Tl.l'n()r >San but rttt >lOem
T4 T11n0r >lOan In """""'ld-slon
bill
r«
,,.,..,...on R<w.~Mol tll'nllil ll«ieo(;Onl\(it be •.......:!
NO No re.t~~onol tjmph n o d a -
Nl R""'""l tll'n!lil n<>clo-
MO No dian~ rno10IIIaoil
Ml Dlllont-
Mie Lung
Ml b 0111• dlllenll..,
hilt. . . ?lllollutlt
SQIIA Tl«T2 NO MO Lew
Slqa II 111 NO MO Lew
Slqo II~ T1 NO MO Hf&tl
T4 NO MO Lew
5lqo IIIB T2 NO MO Hf&tl
T3 NO MO Hf&tl
T4 NO MO Hlilh
Slqa IV AnyT Nl MO Arri-
AnyT AnyN loll Any-
ll<:ulo'<UCulorccdullon
Throm-
Einbolsm
Noftocduot.oo m..,.ntolle la:llemll.
Pl.r bol
-~...........1111
v.....-
and
~loblollltal
-PIIII•
Drllaal!'odo
Voowlllco<m>looolon
Vollub
""'!ttlMN(Jttoft
Colll<:"""'comp-
WIIcllon•
Aln)lolibll
Rtdldloll clam ...
Dloba rnoillu•
C!Nipt• 14 • liM: ltrtMtines, ~i.l4 ud ANa I 2 27
I'1.1 I II
I J tit(. :nu•IDI.IInoelt .r Hlttcqfc Alldllp., M-lllcllemle
Ob&tltiNnlciJI!!clloa>llls
Eil!cn>hMIUihiiiJ<: Escllorlc/lir aol
NSA!Dd.,_
Crolill't<>Oik
Rlllio11Gnet>lilt
Oollqonou&-
or inactive. Treated IBD may be normal. Cytomegalovirus (CMV) inclusions

have been described in refractory UC, which otherwise mimics active disease
(e-Fig. 14.29).
1. Crohn's disease may involve any portion of the GI tract. Roughly, about
40% of patients have small bowel disease only, about 40% have small and
large bowel involvement, and about 20% have colonic disease only. On
the basis of the clinical behavior and pattern of disease, Crohn's disease is
divided into three categories: inflammatory, fistulizing, and fibrostenotic.
Grossly, the involved bowel segment typically has a rigid, strictured, or
thickened wall with creeping fat. Upon opening, the segment usually grossly
maintains its cylindrical shape (e-Fig. 14.30). The mucosa may show cob-
ble stoning due to linear and transverse ulcers with intervening edematous
mucosa. Deep fissuring ulcers and fistula tracts are common. The muscle
layer is thickened.
The microscopic hallmark of Crohn's disease is transmural inflamma-
tion with a lack of homogeneous involvement, that is, skip lesions (areas
of active disease separated by normal bowel) are present. In resection spec-
imens, lymphoid aggregates may be present in all layers of the bowel wall
but are characteristically located in the subserosal fat along the vascula-
ture in a "necklace" pattern (e-Fig. 14.31). Granulomas, seen in up to 40%
of cases, may be found in the mucosa, submucosa, and subserosa. In the
mucosa, Crohn's granulomas are typically small, well-formed, nonnecrotiz-
ing, and lack multinucleated giant cells (e-Fig. 14.32). A diagnostic pitfall
is the so-called crypt granuloma, which represents a pericryptal histiocytic
response to mucin from ruptured crypts (e-Fig. 14.33), that occasionally
includes foreign body-type giant cells. Granulomas within the muscularis
mucosae can be overlooked because of similarity to smooth muscle bundles.
In the subserosa, granulomas can be larget; can contain giant cells, and are
frequently associated with lymphoid aggregates (e-Fig. 14.34).
It should be emphasized that Crohn's disease is a clinical diagnosis.
Although supportive histopathologic findings in mucosal biopsies include a
lack of uniformity of involvement of all fragments, or a lack of uniformity
within of a given fragment, focal colitis is not specific for Crohn's disease.
Focal colitis is also commonly seen in other conditions, including infectious
colitis, drug toxicity (particularly with nonsteroidal anti-inflammatory drug
[NSAID] treatment), and partially treated UC. Consequently, it is prudent
to avoid labeling a patient with Crohn's disease at the first biopsy, but
rather to give a descriptive diagnosis (such as focal active colitis) and to
provide a differential diagnosis. Appropriate clinical and genetic workup,
and subsequent biopsies, usually resolve the diagnostic dilemma.
2. Ulcerative colitis classically involves the entire colon but not the small bowel
and has a tendency to be more severe distally. In some cases, the disease
involves only the rectum (ulcerative proctitis) or presents as left-sided col-
itis with discontinuous involvement of the cecum (cecal patch), ascending
colon, and/or appendix. Grossly, the affected colon often has a thin and flac-
cid wall that flattens upon opening. The mucosa loses its normal folds and
is granulat; friable, erythematous, and ulcerated (e-Fig. 14.35). Microscop-
ically, the disease is characterized by diffuse crypt architectural distortion
and inflammation. In contrast to Crohn's colitis, crypt architectural distor-
tion is more dramatic, and inflammation is usually limited to the mucosa
and immediate submucosa. In severe active disease with broad-based ulcers,
the inflammatory infiltrate extends into the submucosa and the muscularis
propria in the ulcerated areas. Interestingly, in children with UC, the dis-
ease may be inhomogeneous at initial presentation. Treated UC may also
llw•llh ealllt Creh'ld...._..
illllrllutlon atrua. oonUnuou1 foool <sillp), - " "
Dec~~~~ of_,~ Muoooe.eubm.- 'h 1J511\U181
,.,_.•_1111<01
-
lll1IIU~r ul:ola. triable, ~IIY CoiJblooll:t\1'1.1
B"""IMII 'Tilt> 'Titlebnod "'norrnol
C!Mjllna m. .ntMc: fd Common
Sl!lclure U sultj•- ~.~.,..
flllull Uouoly•- M<Jlle

Floouq Uouoly•- Common
lloolln..,tftlnellt ..:1~ lboo!Moh) Common
Uppor Gllmol•••"nt Usuoly no ~.~.,..
-
RtdOI-11\Mt IOOlf. ~ISl!.
Anll-M'II 5-IO'lf. -75'lr.
Wei~--""""'
Tronwnunoll)mpllold ...,.,.....
Common
Common
,.._ ..,_ Clllllll M uelllr ~ c!MIIa.lll

Po!lant..,. CldOf{>liO) You-(<!iO)
DloeMo adMI; ftdtt8 « ..,_ l!ouallfadM
DloeMo dura11on Sllol1!lr. uowl~ <lOli>OFt L«<..llr. uouallf>l0)'001'8
-""'1!1-
---
EJ>d<*>)ple 11>-"""" IrI'<I!)Jolr
Clondl R.oaulu a>nfl.lpnllol\ lr"''l''oroon!Vr-
su- otaDn noer -..mNI arrte.ce
Mbllu"'al nDm""''laslfund
I\OOiliOA pndo
D\OIO'oDiilc
Nudol
"'*
<Oih 1\ore
Sl:rdttfldld, UMI!!t iMI&I
''""'*''
FreaUOirt
- " ' · ..rb.l!; ....,.
Domon:o11Gnfl<m Sllarp Gradurrlthlnolllon
&lrrc&l ndfiW Ml.fctlll Mld~dbtatod Prom..,....ll'CIIIor:lliral dlllort!on
~"'""'of"""'""".,. No Yos
m""""
PFOII'Inont lom!FIO DI"<IC>rlo.
,.,..,uclerlr...,
PFOII'Inontlomlno Dr<~C>III +
nlllln>jlllb
PFOII'Inont pfi311y No
lm...,nohlllloch..-y
lloll.-ln
4 """"'& ..!eum.& _,.be <lliliC11lt. lD """"'""""'!be clisrinction may be

1111-" "1"""""
a DALM """ be a<lr:qgatdy-...! by liiDple pofnr..,.
tomy an.cl e<mciD1>041 ~ u ..nh • opcr:ra4ic admom.&. How<ttt;
bcaiiiC the cllaleolco:ucq"""""' of die cl!a,pocll oi d}'~Plul& "*" lad oxic
ioo:twe4 ~or t'I'CI> ~.,.,. t'Crifie&lioo of dlcdi~oti• by"'
<"\\)Critnocd Gl pd>olotift ia ~~to.t!gly.--mmeaoled. ~·
>:ol marlo:n in Aid for 4iHumtial d....oN of ciy1p1u». ..,... 01p0..&
•elm-.. ""'beiQII"-ll>ped an.ctareu. .... ;, .....,._, (o-Pip.143t
"""- 14.40). Tol:rk 14.12.
r.c..... cmli<OIId)
.,..,.ri_
,.._...,.lied.
aom.e of the pocmliol r!iff....,cioliag
I. ,_hills t<ic.n 10!d·-·tlo.D.oid!.cl!W.pcrod;-.1 aftct a totaloolco-

~ u.u.liy f»r UC. ~ 1>0IId>itit eh.owl mixed ewrophilic an.cllym·
phoplumac,1i<: iDiiltNtain theihal mu- <WI eroDa.a.o or uloo:m!iaas in
!lUI"'............... o.r..a;.....,...,....:~~ .... .,;u.,.,. ~ diltoftiJ>D,
.mo.... ctzophy, &114 PJI=;c; 1111tapla&i&, -.ylre ..... iD laot!-G:nll &-.
An~ difluaalial di"ii'oci';. .........,1 Crolm'l> cli.aaoe ill a"""'
p;reoriolllly di"'P'oted u UC. ~tloll oi a p;l<'ricw oolcctomy oped.·
-~Ire •coe.a'l' if granlllomu, a fittllla, a ain~~t tn.ct, or a fiMuring
llktt ia <1-=d ia • pG'IIdl.
D. Ghllrtlalt:GIIIII rdoxo oo"" mf!.umnam'l' ""P"""' in the bliad ...,......, u.nr.·
olly dr.e -~- (H&riiiWm pol>dr.) followiag a,'""""' .,. ...
~
DolL The p - i a tbo,.Ptto be coo...! by a ddki...., of abm:-dlain fany
mmr....
ac:ldt beca.... dr.e Oflomy ~=l...tco dlct<dWI>. from thct..:.l atrcam.
Tbe dfMic: finclinp - t ~ frial>lt ~M wid> m•~ 11"'ph.oid
11-plc liD
CIYI>CIIIl o< CIYI>t•booa"''' :!:!Focoll l'lunlhelrt
C!Yptd-n :!:(F<all Promlrwrt
BUll plwnUjttJOI& + ++
Buol tJ'mpllold liiii'IIIIIM :!: ++
P.,atll ... rnotaplulll + +
p,..,Mnl IJmphooyllc or plu.....,UC ++ +
111111/M\111Soft
,_~~~r<e>NI& 1'1 ltlmn 1110P11a :!: ++
p.,.,Mnt ...tntJI)kll& +(co} :!:
Thlcllonod or I,....,fir 111bejilltolol a>llqen loyef
Ulooror..U..
++locl
± (Rire)
* tl'<>all)
++(Common)
H. Infectious colitis is caused by a wide range of microorganisms including bacte-

ria, viruses, parasites, and protozoa.
1. Bacterial infection, for example by Campylobacter. Shigella. and
Salmonella. results in acute self-limited colitis (ASLC) characterized by neu-
trophilic infiltration of the lamina propria and epithelium with associated
cryptitis and crypt abscesses. The inflammation is accentuated in the lumi-
nal portion of the mucosa and is accompanied by damage of the surface
epithelium with erosion and flattening. The crypt architecture is preserved.
2. Pseudomembranous colitis is a potentially fatal mucosal reaction to tox-
ins produced by Clostridium difficile. Most cases are associated with prior
antibiotic exposure which results in loss of normal bacterial flora. Pseu-
domembranous colitis is characterized grossly by adherent yellow-white
plaques; microscopically these are parallel arrays of polymorphs within fib-
rin and mucin. The underlying crypts are ruptured, giving rise to the classic
appearance of "volcano" lesions (e-Figs. 14.46 and 14.47). It is important
to note that pseudomembranous colitis is a morphologic diagnosis that
can occur in C. difficile infection or in ischemic colitis; likewise, ischemic
colitis can occur as a result of infection such as with Escherichia coli
0157:H7.
3. Intestinal spirochetosis is not uncommon in HIV-infected patients but can be
an incidental finding in immunocompetent individuals. It may also be inci-
dentally detected in appendectomy specimens. It is characterized by a fuzzy,
purplish or bluish band of organisms carpeting the mucosal surface (e-Fig.
14.48). The organisms are more easily recognizable on silver stains, such
as the Warthin-Starry stain. There is no associated inflammatory response
or epithelial damage.
I. Drug-induced colitis presents with a wide spectrum of histologic findings.
NSAIDs are the most common medications to induce injury and can be asso-
ciated with diaphragm disease in small intestines, and lymphocytic infiltrates,
apoptosis, and microscopic colitis in the large intestine. Kayexalate, cocaine,
and amphetamines are among the agents that are associated with ischemic
colitis. Excessive laxative use can cause melanosis coli (Nat Clin Pract Gas-
troenterol Hepatol. 2007;4:442). Agents used for bowel preparation may cause
mucosal edema, hemorrhage, surface epithelial detachment, neutrophilic cryp-
titis, and increased apoptotic activity.
J. So-called diaphragm disease in the small intestine is another example of drug-
induced injury. It is caused by NSAIDs, and the lesion is characterized by
multiple mucosal webs that result in luminal narrowing. Biopsies are rarely
obtained in this condition.
K. GVHD commonly affects the skin, liver, and gut. Histologic hallmarks in the
intestinal tract include a paucity of inflammatory cells in the lamina pro-
pria, apoptosis, and crypt dropout. (e-Figs. 14.49 and 14.50), and the severity
of acute colonic GVHD is graded accordingly (Table 14.14). CMV infection
shares the finding of "exploding crypt cells" (apoptosis), and thus needs to be
excluded. Mycophenolate mofetil toxicity can also result in crypt apoptosis,
and thus communication with the clinician is important (e-Fig. 14.51).
L. Miscellaneous conditions. Melanosis coli (pigmented macrophages in the lam-
ina propria) is a common finding in patients due to excessive laxative use (e-Fig.
14.52).
Brown bowel syndrome, in contrast, is a unique condition believed to be
caused by vitamin E deficiency, which leads to mitochondrial dysfunction and
lipofuscin accumulation in smooth muscle cells.
Irritable bowel syndrome (ffiS) is a common cause of abdominal pain and
chronic diarrhea. It is a clinical diagnosis, and colonic biopsies from these
patients are entirely normal on H&E stain.
C!Nipt•14 • liM: ltrtMtines,~i.l udMA 4 I 2aa
''I.I Illtl£!: 'I Hlttolotfc Gfldlltl'lf Coloftle C1'1ft ,_l·lbt Dr.ut (GVHD)
lladt ~IIMIIIflll: fNIIrtt
1 lhdWuolcol -~~~a IWt¢YilCO, eDOroo tlmi>IIO<:YII: 1'111-
2 AI>OIII* CIYI>ta. CIYI>t•bac"" ..• e1ld rnbtld lomlha I>I'OI>IIo 1'1111:ra>
$ Laud lndli'odual <rnJ~a, •reo of mu""""- oi"'YP'", fooll ulcet-.
4 Wldoip'eod ""'"'a)'l)11, and oulflueplllolllm wllll m._ldonudlllon and
ulcollllm
2J4 I SECTION llh <II lAACT
'' 1,1,! I lt!iU I WHO llltolollc Claullcalloft ol TUIIIOf'Uf * Coloft lnd Rectum
(tllloiiii-
Adonorno
Tubullr
VIlola
Tubub¥111ouo
----
ll)opi.Ua (1-pilllollllnooploslo), """•-
Diotc>I.Ua (1-lihollll IIOOI>Iooil), tqjl ll"ldf>
ltyp&IJ)iulle llOttP.IIII..,.IUiilypeo
'hdlttNIMii!lted
- .." - 00i>r'<1nllpolnl
liarniJ1x>lnololul
adertOII"'''Il
{!M->
~1111d PQin>
JUI'IIIIIIo PQin>
Pau~potn>
o..,.,.,...
""'-''*'"',.
C~blffonn
.. -~ a.rlen..,..l,..,.
MeWlilY <t~rdnotn~~
MlcraP«~>IIuyaul:lr,..,,
Mudftoute.rdftOI'nll
~., .... .,.,.,._
St:Nr•B:I ~rdnOil'liD
Adiii!OOQ 1111110ua CIII'CIIIOinll

Spindle col CIICiftM'II
&IUIIToOUa ceii<O......i!\11
Undllollllllilled .,......,m•
Ne!raontloe!lrl& •"''>l&amt
fW'canclocrlno tumor (NEI)
N£TGI (<Jrdnold)
N£TQ2
Nl!roontloerlnoco.....,m• (NEC)
Lerpeo!INEC
Mlood -..u-
Smoll<l!ll NEC
EC col-n-.~roduct\.1 N£T
.,..I""""'
_..,..._""
L cal, alu""''f\'h Pllllillund PP/PW~roclu~ N£T•
u-
U.iorn'J'>I!III
-t.Mlll'llllll,..,lll!.mot
I.e~
Anp,...,..
l<iopool..,..,.
&rwr~awa
lk.oll !ifnpi\Gm.. lll<iudllhl&, wllll _,,..lntsmw!ID bol>.loon d-lq&
B-alli !lmpi\Gma •nd Buillllt lfmJ:IIoma
Bullllt tjmpi\Gm&
Dlll'lloe IIIIo 8-col b'rn~homo
Marilo ... b'rnpllan.t
M&lfJNI"""' tjmpi\Gml dmi_H.,.,....... tjmphold-.., (MAI.T
t,'mpllome)
$tc IIJUWI
villous, and tubulovillous adenoma if the villous component is between 25%

and 75% (e-Fig. 14.59). By definition, adenomas contain at least low-grade
dysplasia characterized by nuclear stratification; nuclear enlargement, elon-
gation and hyperchromasia; and cytoplasmic mucin depletion. Paneth cells,
neuroendocrine cells, and squamous cell clusters may occur in adenomas.
E. Mixed hyperplastic and adenomatous polyp is a synonym for SSAIP with cyto-
logical dysplasia.
F. FAP is an autosomal dominant disorder caused by germline mutations of the
APC gene located at 5q21. A minimum of 100 colonic adenomas is required
for the diagnosis, but an attenuated form with a reduced number is not uncom-
mon. Adenomas of the upper GI tract, particularly in the ampullary region,
and fundic gland polyps are also common findings in FAP patients. Progres-
sion to colonic adenocarcinoma approaches 100% by midlife if prophylactic
colectomy is not performed (e-Fig. 14.60).
Gardner and Turcot syndromes are considered variants of FAP. In addition
to adenomas of the GI tract, extra-GI tumors are seen in these patients including
osteoid osteoma, epidermal cysts, and intraabdominal fibromatosis (desmoid
tumor) for the former; and tumors of the central nervous system for the latter.
G. Lynch syndrome {LS), previously known as hereditary nonpolyposis colorec-
tal cancer (HNPCC) syndrome, is an autosomal dominant disorder with an
increased risk of colorectal cancer as well as an increased risk of extrain-
testinal epithelial malignancies including of the endometrium, ovary, stomach,
small intestine, upper urinary tract (and others). It is caused by defects in DNA
mismatch repair (MMR) genes including MLH1. MSH2. MSH6. and PMS2
that lead to MSI. Defects in MMR can be evaluated indirectly by immunohisto-
chemical stains for DNA mismatch repair proteins or by DNA-based molecular
analyses for MSI, or directly by DNA sequence analysis of the MMR genes
themselves.
Immunohistochemical analysis is used in many centers to identify the most
likely mutated MMR genes; the mismatch repair proteins are normally found
in human tissues, thus loss of nuclear staining may represent the presence of a
genetic abnormality. The commonly tested proteins are MLH1, MSH2, MSH6,
and PMS2. MLH1-PMS2 and MSH2-MSH6 form dimers, the latter member
of each pair regulating expression of the former. Thus, if there is a mutation
in MSH6. expression of both MSH2 and MSH6 proteins will be lost, as will
nuclear staining by immunohistochemistry. However, the loss of expression
has different implications for each protein pair. If there is loss of MSH2 and
MSH6, or of MSH6 and PMS2 alone, the patient is more likely to have LS and
therefore should be further referred for genetic counseling. On the other hand,
loss of MLH1 and PMS2 can be due to sporadic mutations as well as a rare
form of LS, so additional testing for BRAF mutation is indicated; the presence
of a BRAF mutation indicates a sporadic mutation rather than LS.
MSI analysis is currently recommended for all patients that fit revised
Bethesda criteria (Table 14.16).
H. Hamartomatous polyps can occur sporadically or as part of juvenile polypo-
sis syndrome, Peutz-Jeghers syndrome, Cowden syndrome, and Cronkhite-
Canada syndrome. All these syndromes have an autosomal dominant inheri-
tance pattern, except for Cronkhite-Canada which is a nonhereditary disorder.
Juvenile polyps feature cystically dilated and tortuous crypts with edema-
tous and inflamed lamina propria (e-Figs. 14.61 and 14.62). Dilated crypts
often contain neutrophils and/or mucin, hence the name "retention polyp."
The surface of the polyp may be eroded or ulcerated, with granulation tissue
and epithelial regenerative changes. Juvenile polyps are not restricted to young
individuals. The polyps in patients with Cowden and Cronkhite-Canada syn-
dromes closely resemble juvenile polyps.
~i(.1:!1 Jtl(l( ,...lettn111l crtt.fatM ... PCC ClpctlS)ucio.)

1. CRCd.-lnapollontof<50_.ataao
2. Pr......,ol.,<:!ll•no•lt.rndaf;lnch.....,..,.IOI'fldoiOf-ri.S--tlunoro.
'"llnlllm of Ill"
S. CRC 1«1 MSI·H' ~ dla&nooed at <lii>)'OOIOof ...
4 P - . - CRC••d alht"'- - • • ~tl.lmr,- one oftha<O...,..
d~ b!lot& t!IUJ110ol50 )Ol!llra
S. P-1-CRC\OIIIItwoormmlht~Ofa-.....r.. --acynch
~miMI!IIIIod tlnnor, "''IUdllm<>l..,
r-. ~~oc~~t, Mt~M~allt (l'HIIO hetrw kll••• fof c.ct~t- ot
h td,. aftdl Reelllm
,,, , . . . . (!)
1lC PrtmOIY111m«..,IIOt be-*
TO No-ofp-,tumOf
lis Oin:l,..,...ln ollv: lnlraapilllollal Ofln- of lunlne p""""•
n nmor .,...,._ subrrn.rc:t~~M
'12 Tl.mor IWodoo mUOCIIIollo 111'01>11a
'13 Tt.mOr IO'oiSdedt"""" Ill& m....,lalt& proP/It tnt> pOIIoolontdalllow ...
T4e Tl.mor l"'"dfiolbt loll>o ..,,_of¥1Kml poollonoum•
T4b Tt.mOrciiOCIIy ln\0\ldM tr I t _ , 10011181 - · • .rldrUe!U""'
llillnii .... IIOdltOII
Nit R""'""l t)mplt nodotcoMalbu...-
NO NoiOifonoHr•mphi\Ode~
Nl * - h In 1-$ ......I lymph MdM
Nla lo'loltO\IIoslo In 1 r'lllonoll:ymplt IIOdlo
Nib *M!•h In 2-$ ......I lymph MdM
Nlo TIIIICir dapoollts) In llteou-...., ~. « nonpa-llzod parloolc
Of~ lluuM -~~~ I'Oflonel nodel m - I t
lt2 IIWnJMb In 4 Of mo"' . n i l lymph , _
1t2o """""'""'In 4-6 nllllonotlymph I\Od.,
lt2ll IIWnJMb In 7 Of mo"' . n i l lymph,_
IIGrt••s•••t• (1111)
MO Nodllt.ont-
Ml Dlllontme-
Mla
Mlb """""-In
lo'loltO\IIoslo <mil nod to
~~~treMQI\0
""'"''Jill
_nl.._.
or*
Of111e Dflltoneum
S'llltii'M....
SlqoO lis NO MO Sllillo IllS "13-T4o NIINic 1110
$QII Tl NO MO T:!-'13 N21 MO
T2 NO MO Tl-T2 N211 MO
5lqo IIA T3 NO MO Stiee IIIC T4o N2e MO
Slqa liB T4o NO MO T3-T4o N211 MO
5lqo IIC T411 NO MO T4b NI-N2 MO
$QIIIIA Tl-'12 NI/Nic MO 5\epn.:<l AnyT AllyN Mle
Tl ~ MO Stoee 1\19 Nry T MjN Mlb
I
..
2aa SECTION 111. 01 lAACT
~i (.1:! I Jf8!:1( AI•••• nt Plllrnetea forComp""""' of M...,tw•

._...
IJIIIo IM.Ik b> tile """"'iidl.m
Od'Dob In ll>o """"""""m _,to
tile muO<ulorlt ptQptlo
•-'MY
_, ........
Mr.t trill_... -...~L 111• ctwrnw-1 mii'B!h
-11tbl.lk1Dtllon-.
m>e~~ior
mt;lllllrlyoftllo m...,llldlltutlecelOIIII d ->5 mm, but nona -nato

111& mUOCIJ\ali> I>IOI>!Io
No ...... ohtslbllb cf111o mUOCIJiali> 1>101lda- at111ol'looil1blob•UIIo
leiMbr ani muad!tll:
C'JEIWt
lnloctbl.lk;( m-A>C:tim willa-h..,,_
on~mlwi~OI\IIOJ1IIoootllle ,_odol ..,rf...,
No 011lflloo c1e11>:1o >S mrn •~ doi>Ch
NoecninJ!:-llla-1 n11J11n dth&-lman
MtM trill...,.. ....tsonlna,lltactOJmletlntlll miiJin ·--~~~
,,, , . . . . (!)
1X Prmoy 11/morcon not be • -
ro No - " " " " polmel)' tunv
n 1\Jmor lhw.deo 10/nlhO D~®!la Of •ubm- ••d a :S2 <tn
ne 1\Jmor 11m 2cmwttll mulond~mln•
p<Opola"' subm...,..
Tumor - vrou&~~ lila m.....,.,. propolo ln1D " ' - Oflnlo
11011~1md llil!lcdloor 1>011.-111-.
1\Jmor lhw.deo """"""urn «OCher 0111/111
!'« "''Y,&dd Cml ftr multiple bJmOfl
fbtl'oaoi.,.._OII
r« R"""'lbm,., · -canld be·~
NO No rUonol tJrnDIIIIO<Io ..............
Nl R9NII)m,., nod& m-.olo
IWI1t•1t1111dt OIQ
MO No dlltllnt - ·
ftql_,...
Ml
Slqo 0
Dlslant - ·
11l' NO MO
51oio I n NO MO
51oio IIA l2 NO MO
SloiO 118 13 NO MO
Sileo 1114 T4 NO MO
Sileo 1118 AeyT Nl Ml
Sileo IV AllY T AllYN MO
acid phosphatase, an immunomarker that is also positive in prostatic ade-

nocarcmoma.
3. Mixed adenoneuroendocrine carcinomas (MANEC} contain at least 30% of
histologically recognizable adenocarcinoma or squamous cell carcinoma,
and NEC. Both components should be graded.
B. GIST of the colon accounts for about 5% of all GISTs and is most commonly
seen in the rectum (e-Fig. 14.69). Colonic GISTs tend to have an aggressive
biologic behavior, and tumors with mitotic activity can recur and metastasize
despite a small size of <2 em.
C. Leiomyoma is the most common mesenchymal tumor of the colon. It arises from
the muscularis mucosae and presents as a well-demarcated nodular expansion
in the submucosa. It is typically hypocellular, lacks cytologic atypia, and lacks
c-kit expression (e-Fig. 14.70).
D. Submucosal lipoma can be associated with hyperplastic change in the overlying
colonic mucosa (e-Fig. 14.71).
E. Inflammatory polyps are a heterogeneous group of polypoid lesions and include
the pseudopolyps seen in IBD. Inflammatory polyp features inflamed lam-
ina propria and damaged or distorted crypts, and granulation tissue may be
present. Inflammatory polyps may be microscopically indistinguishable from a
juvenile polyp if cystic dilation of the crypts is prominent; the appropriate diag-
nosis relies on clinical information. Occasionally, pseudosarcomatous stroma
is noted, particularly in eroded or ulcerated areas, characterized by bizarre or
multinucleated stroma cells simulating sarcoma (e-Fig. 14.72).
F. Mucosal prolapse occurs most commonly in the rectum but can be seen any-
where in the colon (the alternative term for this entity, solitary rectal ulcer, is a
misnomer). The characteristic histologic features include a fibromuscular lam-
ina propria with vertical extension of the muscularis mucosae into the spaces
between crypts, and elongated hyperplastic distorted crypts (e-Fig. 14.73).
However, since lamina propria smooth muscle proliferation can be seen in
any polypoid lesion, the diagnosis of mucosal prolapse should not rely solely
on this finding. It should also be noted that smooth muscle fibers are normally
present in the duodenal mucosa, and thus should not be viewed as evidence of
mucosal prolapse when they are present in that location.
G. Inflammatory cap polyps are located mostly in the sigmoid and rectum. These
lesions are characterized by hyperplastic, tortuous crypts with abundant
inflammation in the lamina propria. The "cap" is composed of an exudate.
H. Colitis cystica profunda is a benign condition in which cystically dilated crypts
are misplaced in the submucosa and/or deeper layers of the bowel wall (e-
Fig. 14.74). The distinction from well-differentiated adenocarcinoma lies in
the lobular arrangement of the displaced crypts, lack of dysplastic features,
lack of desmoplasia, presence of surrounding lamina propria components, and
presence of hemorrhage or hemosiderin.
I. Mucosal ganglioneuroma resembles a neurofibroma and features a bland
Schwann cell proliferation with nerve fibers that expands the lamina pro-
pria, with individual or nests of ganglion cells embedded in the spindle cell
background (e-Figs. 14.75 and 14.76). Multiple (ganglioneuromatous polypo-
sis) and diffuse (ganglioneuromatosis) lesions are usually seen in patients with
multiple endocrine neoplasia (MEN) 2B or type 1 neurofibromatosis (NF1).
J. Inflammatory fibroid polyps are usually sessile lesions. They contain an abundant
fibromyxoid stroma and an inflammatory infiltrate rich in eosinophils (e-Figs.
14.77 and 14.78). The lesion is usually submucosal but can also involve the
mucosa.
K. Mucosal folds and prominent lymphoid follicles can resemble polyps endo-
scopically. On biopsy, a mucosal fold consists of completely normal colonic
mucosa.
Chapter 14 • The Intestines, Appendix, and Anus I 241
L. Miscellaneous polypoid lesions. Pneumatosis coli is characterized by gas accu-

mulation in colonic wall. It most commonly affects the sigmoid colon. Micro-
scopically, cystic spaces surrounded by histiocytes and giant cells are seen
(e-Fig. 14.79). Endometriosis, pseudolipomatosis, and xanthomas can also
present as polypoid lesions.
VIII. DIAGNOSTIC FEATURES OF COMMON NONNEOPLASTIC AND NEOPLASTIC CONDITIONS
OF THE APPENDIX
A. Acute appendicitis usually occurs as the result of luminal occlusion (such as
by a fecalith, lymphoid hyperplasia, or Enterobius vermicularis), followed by
bacterial infection. Diverticulosis of the appendix is a rare cause. Microscopi-
cally, acute appendicitis is characterized by transmural neutrophilic infiltration
(e-Fig. 14.80). Abscess formation, gangrenous necrosis, and perforation may
ensue. When inflammation extends into the mesoappendix and the serosa,
periappendicitis should be diagnosed.
B. Cystic fibrosis involving the appendix is characterized by thick, eosinophilic,
inspissated mucoid material in the lumen and in the crypts (e-Fig. 14.81).
C. HP of the appendix is histologically similar to that of the colorectum but tends
to be sessile. Hyperplasia can also diffusely involve the appendiceal mucosa
(mucosal hyperplasia).
D. SSAIP, the most common serrated lesion of the appendix, is histologically sim-
ilar to SSAIP of the colorectum and has an intact underlying lamina propria.
E. Adenoma of the appendix is also similar to that of the colorectum. There is no
associated mucin in the wall of the appendix.
F. Adenocarcinoma of the appendix has recently been reclassified (Table 14.20),
and a staging scheme separate from that for colorectal adenocarcinoma has
been developed (Table 14.21).
1. Low-grade appendiceal mucinous neoplasm (LAMN}. The appendix is gener-
ally enlarged and filled with mucin. The lining epithelium is villous, serrated,
and undulating and comprises a single layer of columnar or cuboidal cells
with low-grade dysplasia. In contrast to adenoma, the neoplastic epithelium
rests on fibrous tissue with no underlying lamina propria (e-Fig. 14.82), but
no definitive invasion or desmoplasia is present. The associated mucin may
be acellular, and lesions are mostly associated with low-grade pseudomyx-
oma peritonei.
2. Mucinous adenocarcinoma. These tumors have> 50% extracellular mucin as
with their colonic counterparts. The tumor cells have high-grade cytologic
features. Malignant glands invade the appendiceal wall and a desmoplastic
response is present.
3. Signet-ring cell carcinoma. These tumors have a >50% signet-ring cell com-
ponent.
G. Pseudomyxoma peritonei is primarily a clinical diagnosis, and its prognosis
depends on the associated mucinous neoplasm. Although appendiceal lesions
are responsible for the vast majority of the cases, mucinous tumors of other
sites such as the ovary can rarely be the cause.
1. Peritoneal adenomucinosis (low-grade pseudo myxoma peritonei} is defined by
the presence of mucin-containing epithelium that is benign or shows only
mild cytologic atypia.
2. Peritoneal mucinous carcinomatosis (high-grade pseudomyxoma peritonei}
features the presence of frankly malignant epithelium similar to moderately
and poorly differentiated adenocarcinoma in a background of mucinous
ascites.
H. Neuroendocrine neoplasms are the most common neoplasms of the appendix
and include a heterogeneous group of lesions with variable biologic behavior.
1. NETs are histologically identical to those seen in the other parts of the GI
tract and are typically found in the distal third of the appendix. However,
2C2 I SECTION llh <II lAACT
'' 1,1,! I lt!j.j I WHO llltolollc Claullcalloft ol TUIIIOf'Uf * .nclll

(tllloiiii-
P,..,..I,Inont-.
""'""'""'
1\Jbulll
VIlola
1\Jbulrw!lk>ol;
---
l>)opi.Ua {1-pillloiiiJIIIOpioslo), - · -
l>)opi.Ua {1-pillloiiiJIIIOpioslo), lilltl fi'IC!6
lf.IWP!Mdc I>O~P
Seoolooemotod~
'hdlttNIMii!lted adertOII"'''Il
c.rm.m.
Jlolonocarmom•
MucfnC~~~.atdanocarcfnoma
~~~~~~ mudnwsnooplonm
~n1...a Clldnomo
UN!IIo...,tii!Od .,.,_m•
rwr..ntloe!lrl& •"''>l&amt
-ntloellno tum" {N£1)
NETGI (clrdnold)
NETG2
Nelrcontloe!h<O.....,mo (NEI))
Uollll«>ll NEC
Smolleo!INEC
EC <Ill, -n-.i>rodU~ NET
Goblllco!ICJ-
~c:dl, l!luoapHI<e popl»o 1nd PPII'I'Y'flroduclnJ NET
_._,.........
1\Jbulll Cllolrdd
Nllrom•
u-
u..._....,.
~lom!omt
l<iii>Oll..,..,.
&rwr~awa
IK151FT.,_
1'1.' IIJtlflll
,,, , . . . . (!)
Clf"$JWI
1X Pltni/Y tumot' <:~nrd be a nllnid
TO No e\4i:lence d l>llh'IIIIY 111nor
..... C.-eln d:u: lntra.optilellalor lrwulon tl ~rnN PfOI>IIs'
Tl 1\rm« fnilll!ki'i!ll&UbrnuQO,Isll
l2 Tuma- fnillOICies muiCUirt& ~rta
l3 Tum .. tnllldosiiiiOII,III 1M mu~a~flrla pi'Ojrilln!D p o -
UIIuao
1\irnor - - 1 0 lllo ot.nce of.......-.J l>flltMourn. lncblha;
rnuel110114 Pl.....,.ltlrnorl«lln lllo r1&tlt lower QuadiOnt llldJOf
dlred:ty lrtw.dM ctl'lll!lr t:l!p nt or .strud:l.lr_.,"
1\rm~» pertl!lthlbl: 'ltloi!in.l ~ttlllllm, ftteUj!fW rnueftoua
petltlntwil
tum<rwllllln!M 1\!111- Quadrant
T4b Tumor dhdlt l - o r Is adhonnliD allllr _ . , mudlns
C.rcfllll
1X Pitn::l!ity 1:l.l"nnr Cl mot bll • see !llk'ld
TO NotMdenoo tl PflftWYl!l""'r
Tl
Tla
Tlb
Tum"~~ em In.,_"•test
Tumor 2 em or les$"
Tumor >lan butnat>2em

dlmomb1
dlmllnslon
T2 1\imor >2 <1n ~no ,_.<tn orv.!lh _...,10 lllo """"m

'D 1\imCf >4<111 or\01111-......101118 leurn
TA 1\irn,. d~~ 1_011\.,ld)ocoK "'11'M or ''""''"""'• e.g.,
abdomlnii\IOII!and -..musdl'
111111111,..... IIOdlltOU
Clrsb
Nl(
NO
~nil t,omph nodlaa """""'
No ....,...lljmJII node m"""'*
""-'*"
Nl MIICIIIolbln 1-.3 "''ll:>nal tlmlil n -
N2 MIICIIIolbln4«""'"' "''ll:>nal tlmliln-
C.-I
NO
Nl
Rlll'onlll.lmph node--
No ....,...lljmJII node m"""'*
Clrsbn'
MQ Nodlslont ,...,..,... (no palllobfle MO,'""' elnlool M ll>"""P"*'
.........p)
Ml Dktlnl: m"'utnls
Mia ln1r""',_' rnallubula bo)oond rWrt_. qllldrant,ln-na
c-udorrf;xor•• J)eltonea
Non~lm_,.,
,....,....D)
~-nt I!'DIInb (no palllol>lll= MO• ...., elnlool M lbQOI!Ip'*
Ml ObeortmotaD.Gio
__.....,__
Slqj:IO n. 1'10 MO
SIQ:II n 1'10 MO
1'2 NO MO
fllqo I!A 13 NO MO
fllqo liB T4a NO MO
Slqti!C T~b NO MO
Slqti!IA n Nl MO
T2 I'll MO
13 Nl MO
T4 Nl MO
AnyT N2 MO
AnyT NO MlaGl
AnyT NO Mla G2,3
AnyT Anyl'l Mlb AnyG
AnyT N2 MlaAnr Q
AnyT AnrN Mlb AnyO
C..illid.lbp ........
SIQ:II n 1'10 MO
Slqj:ll! T2. T3 1'10 MO
Sla.ellll T4 NO MO
AnyT Nl MO
fllqo IV Any T AnyN Ml
C!Nipt• 14 • liM: ltrtMtines, ~i.l4 ud ANa I 2•a
,, I.I IlltjfU( WIIO Hlttoktlc Clllll'kalloft o!TIIIWIIf ol111t Mill C.nal
ljtlhlllll ~~
p..,...,ant-
Anll ~olol """'*Ill' (d)oj:losla), low .,..so
Anll~olll,.....~.hllhlll>do
a-nd.....
-IN
l'<rilnoll !qUIIMI&In1ruc>llfl...l noq>l&als
f'aeotd-
SquiJl"'IOU.l <::11 c:arc!nana
Verntcou> <Orcttorno
Ad""""'.....,m"
Mudnoutlldi8f\OC:IImotrtlll
Naurcondccmo """'""""
Naurcondccmo tumor (NET)
NET Gl (.."'*""'J
NETG2
NaUICOIIdccltto -""""' INECI
~MINEC
StnoiMINEC
·-""-
.... . ,. . . . .Ill
, •• I SECTION llh <II lAACT
I'Biillf tnnor Qlnnot bo -IOd

No Mlan"' <>I pffrnery tum Of
Cor<tlomt1 In oltl ca-n'o d'- hl!h1JI'Ido OQIIIIInOU>Intreo~~l
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X. MISCELLANEOUS LESIONS OF THE MESENTERY

A. Mesenteric fibromatosis is the most common primary tumor of the mesentery
and most often occurs in the mesentery of the small intestine. Even though most
cases are sporadic, some are associated with FAP/Gardner syndrome; men are
affected more commonly than women. Grossly, the lesion is well-circumscribed
and usually measures >10 em.
Histologically, spindle cells are dispersed in a densely collagenous stroma,
and myxoid change of the stroma can be present. The borders of the lesion
are infiltrative despite the well-circumscribed gross appearance. Immunohisto-
chemically, the tumor cells show nuclear beta-catenin positivity. Patients with
Gardner syndrome show similar histologic findings, even though the myxoid
stroma can be more prominent.
Due to the infiltrative nature of the lesion, complete excision is difficult,
and recurrence is common. The clinical course is more aggressive in Gardner
syndrome patients; in fact, fibromatosis is the second most common cause of
death in this patient group.
Sclerosing in mesenteritis is in the differential diagnosis of mesenteric fibro-
matosis. The lesion is usually solitary and most often arises from the mesentery
of the small intestine, although in some cases diffuse involvement by multiple
masses may occur. Histologically, the lesion contains areas of fibrosis, inflam-
mation, and fat necrosis in varying proportions. By immunohistochemistry, the
tumor cells are smooth muscle actin-positive but nuclear beta-catenin-negative.
B. Inflammatory myofibroblastic tumor {IMTI involves the mesentery and omentum
as the most common non pulmonary sites. Being the most common in children,
IMT is multinodular or lobular with a rubbery, tan-white cut surface. A variety
of histologic patterns can be seen in different lesions or within a single tumor.
One common pattern resembles nodular fasciitis with spindle to stellate shaped
cells embedded in a myxoid stroma. Other tumors show spindle cells arranged
in a storiform or a fascicular growth pattern. Prominent lymphoid aggregates,
or lymphoplasmacytic infiltrates, are seen in most cases. Immunohistochem-
istry is positive for smooth muscle actin and desmin. Since rearrangements of
the ALK gene are characteristic of the tumor (see Chap. 46), immunostains for
the ALK protein are especially helpful.
C. Desmoplastic small round cell tumor is an aggressive malignancy most com-
monly seen in young boys and patients present with an abdominal or pelvic
mass. The tumor is composed of nests of small round blue cells embedded
in a desmoplastic stroma separated by fibrous tissue. Rhabdoid features are
commonly seen. The tumor cells are vimentin and desmin positive; perinu-
clear keratin positivity is also present. The t(11:22)(p13; q12) translocation
characteristic of the tumor results in an EWS-WTI fusion (see Chap. 46).
D. Mesenteric inflammatory venoocclusive disease (MIVD) most commonly occurs
in men in the pericolonic soft tissue and subserosa. The presenting symptoms
are those of ischemia, thus the clinical differential diagnosis is usually quite
broad. Idiopathic myointimal hyperplasia is a characteristic feature of the dis-
ease, consisting of a concentric proliferation of smooth muscle cells within
the small- to medium-sized veins of the affected mesenteric segment. Since the
involved veins can be mistaken for arteries, an elastic stain can be helpful in
diagnosis (e-Figs. 14.92 and 14.93). Trauma and phlebitis have been ques-
tioned as the underlying etiology of MIVD. The patients follow an indolent
course after surgery.
The Liver
I. NORMAL ANATOMY. The largest solid organ of the body, the mass of the adult liver
is 1200 to 1600 g. The right, left, and caudate lobes are subdivided into segments
on the basis of inflow blood supply (e-Fig. 15.1).* The liver has dual inflow supply,
with approximately two-thirds from the low pressure, low 0 2 portal vein, and
one-third from the systemic pressure, high 02 hepatic artery. Pressure equalization
occurs in the sinusoids, along with the nutrient and 02 gradient from portal tracts
to terminal hepatic venules. The venous return is via the left, right, and middle
hepatic veins which join to form the inferior vena cava as it enters the heart at the
right atrium. Bile duct blood supply is entirely from the hepatic artery plexuses.
Microscopically, the hepatic cords are lined by reticulin fibers and separated
by sinusoids. The parenchyma is subdivided into acinar units of Rappaport which
reflect an oxygen/nutrient gradient from most (zone 1) to least (zone 3 ), respec-
tively; zone 2 is an ill-defined area in between. The anatomy of Rappaport's units
underlies many pathologic processes. The lobule, often used interchangeably with
acinus, is a term based on the concept of the hexagon in which hepatic cords radi-
ate from the central vein toward the portal tracts. Each portal tract is a fibrous
matrix that contains a branch of the hepatic artery, a portal vein, a bile duct, and a
poorly visualized lymphatic (e-Fig. 15.2). Larger portal tracts contain autonomic
nerve fibers. Inflammatory cells are typically lacking, or are few in number. The
parenchyma is separated from the portal tract at the limiting plate.
II. GROSS EXAMINATION AND SPECIMEN HANDLING
A. Needle core biopsy. Specimen handling depends on the reason for liver biopsy.
After measuring and description of the number of cores, liver biopsies are
wrapped in lens paper and fixed overnight; use of sponge pads is strongly dis-
couraged because of the artifacts created during sectioning. Protocol "special"
stains and six additional unstained sections are recommended for medical
liver biopsies at initial preparation. Stains include three hematoxylin and eosin
(H&E), a stain for collagen (trichrome or picrosirius red), reticulin, periodic
acid-Schiff after diastase (PAS-d), and modified Perls' for iron. Additional stains
that must be available include copper or copper binding protein (rhodanine,
orcein or Victoria blue; orcein is also useful to differentiate passive septa of col-
lapse from active elastic fiber deposition in fibrosis); and Verhoef£ van Gieson
(VVG) for vessel wall architecture (Semin Diagn Pathol. 2006;23:190).
1. Tumor. Processing of biopsies for diagnosis of tumors includes three levels
for H&E and six unstained for possible additional immunohistochemistry
(IHC).
2. lmmunocompromised patients. Biopsies from immunocompromised patients
(typically solid organ or bone marrow transplant patients) may or may not
require "rush" processing; clear communication with the submitting clin-
icians is required in these cases and fixation, grossing, and processing are
tailored to the clinical needs.
3. Medical liver biopsy. The reason(s) for the liver biopsy, that is, diagnosis
or confirmation; grading and staging of hepatitis; or other possible medical
questions should be clearly understood prior to sign out.
248
Chapter 15 • The Liver I 24 9
4. Frozen section. Indications for frozen sections of liver core or wedge biopsies
include donor liver evaluations for quantity of steatosis and/or portal inflam-
mation; acute fatty liver of pregnancy (AFLP) for microvesicular steatosis
detection by oil red 0 stain (from sections of liver biopsies at any stage of
processing prior to xylene clearing, cut onto charged slides). Evaluation of
intraoperatively encountered lesions is done from fresh tissue submitted on
saline-moisturized gauze; cores are best sectioned with as little handling as
possible, sectioned at 90 degrees to the long axis. Wedge biopsies may require
breadloafing before sectioning. Frozen artifact creates spaces that may be
challenging to distinguish from fat; thus conservative estimates of the degree
of steatosis are recommended from frozen sections. If electron microscopic
examination is expected for a potential metabolic disease, additional tissue
should be fixed in 3% buffered glutaraldehyde.
5. Miscellaneous. Iron and copper tissue quantitation can be performed in ref-
erence laboratories directly from tissue in the paraffin block.
B. Wedge biopsy Dr excision. Wedge biopsy is not typically recommended for the
evaluation of diffuse liver parenchymal diseases, as the subcapsular parenchyma
contains fibrous extensions for 3 to 5 mm that may mimic fibrosis (e-Fig. 15.3).
The subcapsular regions tend to show parenchymal collapse; elastosis may occur
in this location as well as a result of chronic ischemia. Wedge excisions for super-
ficial, circumscribed lesions are managed similarly to resections, as described
below.
C. Segmentectomy, lobectomy, or panial hepatectomy are performed for large lesions
that are not amenable to wedge excision. The surgery may or may not fol-
low anatomic boundaries, and thus before sectioning, it is important to under-
stand the procedure that was done; review of the imaging studies and reports
is invaluable. After the type of surgery, mass, and dimensions of the speci-
men are recorded, the resection margin is inked, the appearance of the capsule
noted, and the specimen is sliced in the axial plane at about 0.5 em incre-
ments. Gross examination of the lesion(s) and nonlesionalliver parenchyma
should include color(s) (nutmeg; tan; bile-stained; hemorrhagic; yellow), via-
bility (necrotic, nonnecrotic), texture (firm; hard; soft; spongy), and presence
of nodularity. If a tumor has been preoperatively embolized via transarterial
chemo-embolization (TACE), the percentage of tumor necrosis grossly should
be assessed; however, only after complete submission and evaluation micro-
scopically can it be adequately reported. Tumor sections (at least three) should
demonstrate relationship of lesions to liver parenchyma, grossly visible vessels
or ducts, margins (if close), and any variable areas within the tumor. Sections
of nonneoplastic liver and inked resection margin(s) are submitted to evaluate
underlying liver disease, vascular alterations, and margin status. One nontumor
section of normal liver should be submitted and evaluated by routine special
stains.
D. Total hepatectomy (explant}- performed for end-stage chronic liver disease, ful-
minant hepatic failure, or metabolic disorders- is followed by orthotopic liver
transplantation. Radiology reports must be consulted before processing the spec-
imen to ensure that radiographically detected lesions are sampled. The total
weight, dimensions of each lobe, and capsule appearance are recorded. The
hilum is completely removed, breadloafed, and submitted in toto proximally to
distally without dissection. If a TIPSS stent has been placed, no attempt should
be made to remove it as the spring-opened wires are not protected; rather, the
hilar tissue should be dissected away. The liver is then placed on the cutting
board facing up, and sectioned axially cephalad-caudad in about 0.5 em incre-
ments. Each slice is carefully examined fresh and after fixation (especially in
cirrhotic livers) for bulging, large, or discolored nodules (which are all gross
features of dysplastic nodules [DNs], or small HCC) that require documentation
2!0 I SECTION Ill: Gl TRACT
which includes location, number, size, color, and relation to the capsule or hilum.
In their absence, two random sections from both the left and the right lobes are
submitted. Only one section is necessary for routine special stains. The native
and donor gallbladders are submitted as for routine cholecystectomies.
Ill. DIAGNOSTIC FEATURES OF COMMON NONNEOPLASTIC CONDITIONS. In the approach
to liver biopsy, knowledge of the clinical information is essential. The adequacy of
the biopsy should be assessed, which must be judged on the basis of the nature of
the question(s) being asked. Grading and staging chronic hepatitis ideally involve
a 1.5 em core, and up to 11 portal tracts; <5 portal tracts is not optimal (Semin
Diagn Patbol. 2006;23:132) (see Figs. 15.1 and 15.2).
Grading and staging schema were initially developed for comparisons of treat-
ment for autoimmune and "nonAnonB hepatitis" trials, but they quickly tran-
sitioned to apply to chronic hepatitis (Hepatology. 2000;31:241). All systems
share the assessment of portal and lobular necroinflammation for grade and fibro-
sis for stage (e-Figs. 15.1 and 15.2). One published system (Am] Surg Patbol.
1995;19:1409) is simple to apply and communicate clinically; this system can be
applied for any form of chronic hepatitis, but is not meant for cholestatic or vas-
cular diseases, or alcoholic or nonalcoholic fatty liver diseases (NAFLD).
Patterns of collagen deposition and architectural remodeling are frequently sug-
gestive of the precedent injury: hepatitic, biliary, vascular, alcoholic, etc. Viral hep-
atitis and alcoholic hepatitis differ as the former is portal-based, and the latter is
centered in zone 3, is perisinusoidal initially, and results in nodules the size of the
acinus (e.g., micronodular cirrhosis). The portal-portal fibrosis of the chronic bil-
iary diseases often results in cirrhotic remodeling with maintenance of the terminal
hepatic venule in its central location; a "jig-saw" pattern is therefore suggestive of
biliary disease.
• •
A B
•
• •
• •
c
• •
•
•· •
•
..·e• •.•
• •• • • •
• • • •
• •
•
• • • •
• • •• • .. . . . . •
• •
~ •••
• • • •
•
•••••
Figure 15.1 Ludwig and Batts Grading of chronic hepatitis. A: Stage 1. B: Stage 2. C: Stage 3.
D: Stage 4. From: Am] Surg Pathol. 1995;19:1409. With permission.
Chapter 15 • The Liver I 2s1
•
-~
A B
• •
(it •
(;)
• • •
~
• -~
~ • •
• • • •
Figure 15.2 Ludwig and Batts Staging of chronic hepatitis. A: Grade 1. B: Grade 2. C: Grade 3.
D: Grade 4. From: Am] Surg Pathol. 1995;19:1409. With penni.ssion.
A. Infectious liver diseases

1. Viral hepatitis refers to one of the hepatotropic viruses: HAV, HBC, HCV,
HDV (with superinfection or coinfection with HBV), and HEV.
a. Acute viral hepatitis is commonly diagnosed clinically by serologic and clin-
ical tests. Pathologists, therefore, have relatively little experience with this
form of liver disease. Histopathologically, acute viral hepatitis is char-
acterized by simultaneous hepatocyte injury and regeneration, chronic
inflammation, and no fibrosis. The changes include swollen hepato-
cytes (hydropic degeneration), apoptotic (acidophil) bodies, lobular spotty
necrosis, lobular greater than portal inflammation, sinusoidal cell reaction
(Kupffer cell and endothelial cell hypertrophy), and bi- and multinucle-
ated hepatocytes. Collectively, these findings result in "lobular unrest"
and "disarray" (e-Fig. 15.4 ). The inflammatory infiltrates consist predom-
inantly of mononuclear cells, with occasional plasma cells and eosinophils.
In severe cases, confluent perivenular, bridging (zone 3 to zone 3), sub-
massive (panacinar) or massive (multiacinar) hepatic necrosis may occur
(e-Fig. 15.5). The ductular reaction, a form of regenerative response, may
be accompanied by mononuclear or polymorphonuclear cells (e-Fig. 15 .6).
Trichrome and reticulin stains may be confusing as the reticulin collapse
may be extensive and the collapsed sinusoids may seemingly react with
the stains for collagen. In these cases, the orcein stain for elastic fibers is
helpful, as elastic fibers are only found in fibrosis (e-Fig. 15.7). Zone 3
canalicular cholestasis may be present; if prominent, a diagnosis of acute
cholestatic hepatitis is given (e-Fig. 15.8). Careful evaluation of portal
tracts will exclude biliary obstruction (see later).
Fulminant hepatic failure or complete resolution may occur in any

form of acute viral hepatitis. HAV may have zone 1 confluent or bridging
necrosis and numerous plasma cells or may resemble cholestatic obstruc-
tive hepatitis. HBV, HCV, and HDV may evolve to chronic hepatitis. It
is important to note that the histopathologic differential diagnoses for
acute viral hepatitis include autoimmune hepatitis (AIH), Wilson's dis-
ease, AFLP, drug-induced liver injury (DILl), and rarely, ischemic hepati-
tis. Cryptogenic acute hepatitis, for which no clinical cause is found, is
reported as such (e-Fig. 15.9).
b. Chronic viral hepatitis, defined by persistently elevated abnormal liver tests
for more than 6 months, is caused by HBV, HCV, or HDV coinfection
with HBV, and is histologically characterized by mononuclear cell infil-
trates rich in T lymphocytes, greater in the portal tracts than in the lob-
ules. Plasma cells, macrophages, and eosinophils are also present, but
in smaller numbe1:. The portal inflammation is accompanied by varying
degrees of interface activity (previously referred to as piecemeal necro-
sis) (e-Fig. 15.10), lobular activity with acidophil bodies (e-Fig. 15.11) or
spotty necrosis, and portal-based fibrosis (e-Fig. 15.12).
Some features are characteristic of the specific types of chronic viral
hepatitis. Chronic HBV is recognized by the presence of ground glass
intracytoplasmic inclusions; this is HB S Ag in expanded smooth endo-
plasmic reticulum (e-Fig. 15.13). HB SAg IHC has three patterns: mem-
branous, cytoplasmic, and inclusion (e-Fig.15.14); intranuclear inclusions
require HB C Ag lliC for identification (e-Fig. 15.15). In HBV- and HDV-
coinfected cases, the necroinflammation tends to be more severe, and delta
antigen can be demonstrated in nuclei by immunostaining.
The portal lymphoid aggregates of HCV are not pathognomonic
because they may occur in HBV and AIH, but they are common and
suggestive (e-Fig. 15.16). Steatosis is common: if in zone 1, it is most
likely due to HCV; and if in zone 3, it is most likely due to host fac-
tors (metabolic or alcoholic). Acidophil bodies, bile duct injury, and sinu-
soidallymphocytosis may be seen in HCV; HCV genotype 3 often has
marked macrovesicular steatosis. Crystalline material is often present in
substances that are abused by intravenous injection, and may be evident
as polarizable material within portal macrophages.
c. Epstein-Barr virus hepatitis is characterized by a sinusoidal infiltrate of
atypical lymphocytes in a "beads on a string" pattern (e-Fig. 15.17). The
diagnosis can be confirmed by in situ hybridization for viral RNA and
serologic tests.
d. Cytomegalovirus hepatitis in immunocompromised patients commonly
induces microabscesses surrounding infected cells with characteristic
intranuclear and intracytoplasmic viral inclusions (e-Fig. 15.18). Thus,
microabscesses should prompt immunostaining if inclusions are not evi-
dent. CMV infection in immunocompetent patients may show microgran-
ulomas; in this setting, viral inclusions are not usually present. Multinu-
cleation of hepatocytes can occur in neonates.
e. Adenovirus hepatitis, uncommon in immunocompetent hosts and in adults,
causes nonzonal foci of coagulative necrosis with a minimal inflammatory
response. Nuclei of infected hepatocytes are hyperchromatic and smudgy,
with chromatin margination (e-Fig. 15.19). Confirmation is by immuno-
histochemical staining.
f. Herpesvirus hepatitis includes pyknotic debris and nonzonal "punched
out" necrosis; nuclear ground glass (Cowdry A) viral inclusions can
be found in syncytial nuclei at the periphery of necrosis, or in other
cells within the liver (e-Fig. 15.20). lliC is confirmatory. HSV is not
more common in pregnancy. Rapid diagnosis and treatment may be

lifesaving.
2. Bacterial infections involve the liver in various ways: space-occupying abscess,
toxic cholangitis (e.g., toxic shock syndrome), granulomatous inflammation
(Mycobacterium spp.), and peliosis (bacillary angiomatosis in AIDS). Sepsis
may result in microabscesses, zone 3 canalicular cholestasis, and/or ductular
cholestasis (e.g., cholangitis lenta) (e-Fig. 15.21).
3. Fungal infections are due to systemic infections such as candidiasis, aspergillo-
sis, and histoplasmosis. The biopsy may show organisms or features of sepsis
(see above), or DILl.
4. Parasitic infections, including hydatid cyst, amebic abscess, and schistosomi-
asis, each with characteristic histopathologic features, require a high index
of clinical and pathologic suspicion for diagnosis.
B. Metabolic and toxic liver diseases
1. Alcoholic liver disease (ALD} encompasses a spectrum of fatty liver, alco-
holic hepatitis and steatohepatitis, alcoholic foamy degeneration, and alco-
holic cirrhosis. Steatosis, steatohepatitis, and cirrhosis may be indistinguish-
able from NAFLD. Steatosis, reversible with abstinence, is predominantly
macrovesicular and initially involves zone 3. The hallmarks of alcoholic
hepatitis are hepatocytes with Mallory-Denk bodies (MDBs) and satellitosis
(neutrophilic infiltration around hepatocytes containing MDB), commonly
embedded in dense pericellular fibrosis (e-Fig. 15.22). The pattern of fibrosis
in alcoholic hepatitis/steatohepatitis is characteristic; it begins in zone 3 as
perisinusoidaVpericellular collagen deposition and eventually there is dense
"chicken-wire" fibrosis that may involve the entire acinus (e-Fig. 15.23).
Canalicular cholestasis may be seen. Perivenular fibrosis is common. Oblit-
eration of the terminal hepatic venule occurs in severe alcoholic hepatitis
and results in sclerosing hyaline necrosis, a poor prognostic feature (e-Fig.
15.24). Alcoholic cirrhosis is typically micronodular. Obliterated outflow
veins may be found, as may copper in periseptal hepatocytes. Steatosis is
variably present in ALD.
2. NAFLD may resemble some of the histologic findings of milder forms of ALD,
and occurs in patients who are not heavy drinkers but who have features
of metabolic syndrome (obesity, hypertension, abnormal glucose tolerance,
and dyslipidemia) (Clin Liv Dis. 2010;14:591). NAFLD may also occur with
a variety of medications. Unlike alcoholic hepatitis, however, a diagnosis
of nonalcoholic steatohepatitis (NASH) requires the presence of steatosis;
NASH does not have cholestasis and has less prominent MDB. Central scle-
rosing hyaline necrosis has not been reported. Minimum features of adult
NASH are zone 3 macrosteatosis and hepatocyte ballooning, and lobular
inflammation. Ballooning rnay be confirmed with KS/18 immunostaining (e-
Fig. 15.25). Portal inflammation may be present in all forms of NAFLD,
but if disproportionate, should raise concern of a second process such as
HCV. Pediatric NAFLD may show greater steatosis, less zone-3 zonality,
and more portal inflammation and fibrosis than adult NASH. The pattern of
fibrosis in NASH resembles that of ALD, in that it is zone 3 perisinusoidal
initially. Advanced fibrosis includes periportal fibrosis, bridging, and cirrho-
sis. As with ALD, steatosis may be absent in cirrhosis resultant from NASH;
"burnt-out" NASH is considered the most likely cause of "cryptogenic" cir-
rhosis, however, other considerations include AIH and alcoholic cirrhosis
(e-Fig. 15.26).
Grading and Staging NAFLD (Table 15.1). The NAFLD Activity Score
(NAS) (Hepatology. 2005 ;41: 1313 ), validated and developed for therapeutic
trials from the original Brunt proposal for NASH, is the unweighted sum of
scores for steatosis, lobular inflammation, and ballooning. The numeric NAS
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5. a1-Antitrypsin deficiency, the most common genetic pediatric liver disease,

is characterized by accumulation of intracytoplasmic eosinophilic glob-
ules of varying sizes in zone 1 hepatocytes, easily demonstrated by PAS-D
(e-Fig. 15.30). The diagnosis is confirmed by characteristic peripherallliC
positivity, and serum electrophoresis phenotyping. The hepatocyte globules
signify a Z allele, or other rare alleles of M or S; whether or not heterozygos-
ity is causative or enhances liver disease, is an ongoing debate (Semin Diagn
Pathol. 2006;23:182). In children under 2 years of age, globules may not be
present; thus, this disease is included in the differential diagnosis of giant cell
or neonatal hepatitis (e-Fig. 15.31). Biliary atresia and Alagille syndromes
are mimics, and it is important to remember that globules may also occur
in benign and malignant liver neoplasms. The differential diagnosis of non-
a1AT globules includes polyglucosan inclusions in polypharmacy; Lafora
bodies; cyanamide therapy; HB SAg; adaptation; and fibrinogen inclusions.
6. Wilson's disease, which can occur as a result of over 200 mutations of biliary
copper transporters and ceruloplasmin formation, results in copper accumu-
lation in the livet; brain, cornea, and kidney, and presents between 5 and
45 years old. Hepatic histology is as protean as the clinical disease. Wilson's
disease can be queried for otherwise unexplained macro or microsteatosis,
chronic hepatitis, cirrhosis, or submassive necrosis in a young adult. Peri-
portal glycogenated nuclei and MDBs are common. Excessive copper may
not be visible by staining as it largely remains intracytosolic; diagnostic test-
ing requires quantitation from the paraffin block (>250 1-Lg/g dry weight).
Chronic cholestatic diseases also result in copper deposits in the eyes (Kayser-
Fleischer rings) and periportaVperiseptal hepatocytes, but the increase in cop-
per does not reach the quantitative levels of Wilson's disease (Semin Diagn
Pathol. 2006;23:182).
7. Glycogen storage diseases, or glycogenoses, are characterized by abnormal
accumulation of glycogen in hepatocytes giving rise to a pale, distended, and
mosaic appearance. PAS stains and electron microscopy may help confirm
the diagnosis.
8. Lysosomal storage diseases, the most common of which is Gaucher's disease,
are characterized by distended Kupffer cells. The cytoplasm of "Gaucher
cells" is finely striated as with "wrinkled tissue paper."
9. Hematologic disorders: Lymphoma, leukemia, hemophagocytic syndrome, and
sickle cell disease. Lymphoma and leukemia are discussed further below.
Macrophage activation syndrome (e-Fig. 15.32), characterized by Kupffer
cell erythrophagocytosis, reflects systemic malignancy, viral infection, or col-
lagen vascular disease. Serum ferritin levels are extremely elevated. Portal and
parenchymal CDS+ lymphocytes are common. The sickled cells in sickle cell
disease cause microthrombi, erythrophagocytosis, and increased hepatocel-
lular iron (e-Fig. 15.33).
10. Reye's syndrome (e-Fig.15.34) and other mitochondriopathies are more com-
mon in children than adults. The liver shows pauci-inflammatory microvesic-
ular steatosis. Ultrastructural examination highlights mitochondrial alter-
ations. Similar changes occur in alcoholic foamy degeneration.
11. Total parenteral nutrition (TPN} results in steatosis or steatohepatitis in adults
and cholestasis with a ductular reaction and fibrosis in children. In both,
bridging fibrosis and cirrhosis may occur.
12. Amyloidosis commonly involves portal tract arteries and may be inconse-
quential, and can occur as the result of several disease entities (Clin Liver
Dis 2004;8:915). Howevet; hepatic involvement is considered terminal when
amyloid replaces the sinusoids and results in compression and atrophy of the
hepatocytes (e-Fig. 15.35). Trichrome stain shows the characteristic gray
color of amyloid, and polarized Congo red shows the apple-green birefrin-
gence of amyloid.
13. Cystic fibrosis has hepatic manifestations in about 20% of patients. CF may
present as a mimic of biliary atresia or neonatal hepatitis, or later as por-
tal hypertension, due to bile duct mucus plugging and fibrosis. The histo-
logic hallmarks are dense eosinophilic inspissated mucous in dilated ducts,
cholangitis, ductular reaction, chronic inflammation, and fibrosis (e-Fig.
15.36). Focal biliary fibrosis occurs in up to 70% of adults and may warrant
liver transplant.
14. Drug- and toxin-induced liver injury (DILl), commonly in the differential for
unexplained liver test elevations, can be direct (predictable, intrinsic) or indi-
rect (unpredictable, idiosyncratic). Direct toxicity involves agents known to
produce liver damage in a dose-dependent manner; methotrexate, antibi-
otics, and chemotherapeutic agents are examples. Indirect toxicity is immune-
mediated and dose-independent; granulomatous and eosinophilic inflamma-
tion typifies this type of injury. Acute injury may lead to cholestatic hepatitis,
bland cholestasis, interlobular duct damage, acute hepatitis, and massive
necrosis. Chronic injury may assume the form of chronic hepatitis, granulo-
matous hepatitis, steatosis, steatohepatitis, vascular injury, fibrosis, cirrhosis,
or neoplasia. Oxaliplatin injury is discussed below in sinusoidal obstruction
syndrome (SOS).
C. AIH and bile duct disorders Df the liver
1. Autoimmune hepatitis {AIH} can present in adolescent or postmenopausal
females, and is associated with hypergammaglobulinemia (lgG) and high
titers of antinuclear and antismooth muscle antibodies in adults, and anti-
liver-kidney microsomal type 1 antibodies in girls. The diagnosis should only
be made after other metabolic diseases have been excluded, and after neg-
ative viral serologies have been demonstrated (Hepatology. 2008;48:169).
In classic cases, there is a dense portal and lobular mononuclear cell infil-
trate enriched in plasma cells (e-Fig. 15.37). Marked interface hepatitis, cen-
trilobular confluent or bridging necrosis, and hepatitic resetting are present.
Advanced fibrosis may be found at presentation. Cholestasis is rare but sig-
nifies severity. Alli may rarely present as fulminant hepatic failure. Likewise,
mild chronic hepatitis and/or cirrhosis may be the initial findings.
2. Primary biliary cirrhosis (PBC}, a progressive cholestatic disease of middle-
aged women, results in the destruction of intrahepatic bile ducts. lgM and
serum cholesterol are elevated. The early stage (Table 15.2) has mixed portal
inflammatory infiltrates and the pathognomonic florid duct lesion consist-
ing of granulomatous or lymphocytic infiltration of duct epithelium (e-Fig.
15.38). The granulomas in PBC are epithelioid, may be portal or lobular, and
present in any stage of disease. Stains for fungal and acid-fast organisms are
appropriate at the time of initial diagnosis. The disease is inhomogenous, but
ductular reaction, interface hepatitis, chronic cholestasis, and biliary piece-
meal necrosis develop with progression, with eventual bridging necrosis, sep-
tal fibrosis, and biliary cirrhosis. Chronic cholestasis (cholate stasis) is char-
acterized by periportal edema, ductular reaction, MDBs, lobular foam cells,
copper (e-Fig. 15 .39), and cholestatic rosettes. Nodular regenerative hyper-
plasia (NRH)-like parenchymal features may occur in any stage of PBC and
may explain the clinical findings of noncirrhotic variceal bleeding.
Autoimmune cholangiopathy and anti-mitochondrial antibody (AMA)
negative PBC are synonymous terms for seronegative PBC that is otherwise
clinically and histologically identical to PBC (and is distinct from overlap
syndrome as discussed below). ANA testing is often positive in this setting.
3. Primary sclerosing cholangitis {PSC), a progressive fibre-obliterative disorder
primarily of young men, is of unknown etiology. PSC affects the extra- and
intrahepatic biliary tree leading to biliary strictures and ectasias, and cirrho-
sis. PSC is strongly associated with ulcerative colitis. The definitive diagnosis
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cirrhosis indistinguishable from PSC. Biliary atresia with extensive lobular

injury may simulate neonatal or giant-cell hepatitis.
6. Idiopathic adulthood ductopenia (lAD) is, by definition, paucity of intrahep-
atic bile ducts as defined by a ratio of < 0.5 interlobular ducts to portal
tracts (the normal range is 0.9 to 1.8). This poorly understood condition
is pauci-inflammatory, insidious in presentation and progression, and rarely
is familial. Immunostains for keratins 7 or 19 aid in identifying hypoplas-
tic or absent ducts and progenitor cells. Unlike extrahepatic biliary atresia
or obstruction, ductular reaction (i.e., proliferation of the progenitor cell
compartment) is absent in lAD. In pediatric patients, paucity may be syn-
dromic (Alagille syndrome) or nonsyndromic; the latter is associated with
other causes of neonatal hepatitis. In adults, the diagnosis requires exclusion
of PBC, PSC, chronic allograft rejection, GVHD, sarcoidosis, and untreated
Hodgkin's disease.
D. Major vascular disorders of the liver
1. Venous outflow obstruction can result from cardiac disease (congestive heart
failure) or large vessel disease (occlusion of large hepatic veins or the IVC,
i.e., Budd-Chiari syndrome). Both involve zone 3 sinusoidal dilatation (con-
gestion) and may show red cell extravasation into the space of Disse with
hepatocyte loss. With chronicity, cord atrophy, and withering occur, and
fibrous replacement results. Ductular reaction may be prominent. Cardiac
sclerosis/cirrhosis with reverse lobulation has become an uncommon find-
ing. Caudate lobe hypertrophy is a feature of Budd-Chiari syndrome due to
separate venous drainage.
2. Sinusoidal obstruction syndrome (SOS} occurs unpredictably following pre-
conditioning for bone marrow transplant, and following chemotherapy reg-
imens containing oxaliplatin for colorectal metastases. Grossly, the disease
is characterized by a "blue livet." Formerly referred to as veno-occlusive
disease, the process has been renamed to reflect the fact that the location
of initial injury is the sinusoids, which are denuded. Downstream, debris is
deposited in the outflow vein branches, which results in subendothelial fibro-
sis of outflow veins. In the bone marrow transplant (BMT) setting, the onset
is soon after BMT; acute abdominal swelling and elevated bilirubin herald
SOS, and hepatocyte necrosis in zone 3 is a recognized feature in this set-
ting. VVG is helpful in visualization of affected veins in the area of necrotic
hepatocytes (e-Fig. 15.42). In oxaliplatin-related SOS, seemingly randomly
scattered foci of ectatic sinusoids, some nearly peliotic, may be seen.
Hepatocyte anisonucleosis in H&E-stained sections is a diagnostic clue
(e-Fig. 15.43).
3. Noncirrhotic ponal hypenension (idiopathic portal hypertension), usually the
result of presinusoidal causes, may also be due to hepatic causes such as
schistosomiasis, sarcoidosis, or congenital hepatic fibrosis. Clinically, normal
liver synthetic function is maintained in IPH.
a. Nodular regenerative hyperplasia (NRH) is a condition related to aber-
rant flow that commonly results in noncirrhotic portal hypertension. The
parenchyma is diffusely nodular, but without fibrosis. The nodularity is
appreciated at low magnification and by reticulin stains (e-Fig. 15.44),
which highlight the regenerative cords outlined by atrophic cords.
b. Hepatoponal sclerosis is due to intrahepatic portal vein injury and scarring.
The extrahepatic portal vein is patent, at least initially. The parenchyma
shows a variety of alterations: abnormally approximated portal tracts,
periportal enlarged vessels in direct contact with hepatic cords, and
multiple ectatic sinusoidal structures that resemble angiomatoid struc-
tures. Portal and/or sinusoidal fibrosis may be present. Portal veins may
be inapparent or show marked wall thickening with luminal narrowing
(e-Fig. 15.45).
4. Portal vein thrombosis (PVT} may result in subtle or gross parenchymal atrophy
characterized by approximation of vascular structures (i.e., the infarct of
Zahn). Biliopathy may also result from PVT.
5. Osler-Weber-Rendu syndrome (hereditary hemorrhagic telangiectasia; HHT}
is an autosomal dominant disorder that results in multisystem angiodys-
plasias. Of the four known genetic subtypes, liver lesions are in the I-n-IT 2
group of alk1 gene mutations on chromosome 12; final diagnosis rests with
fulfillment of three of five Curacao criteria, which include hepatic AVMs as
well as family history(] Med Genet. 2011;48:73). Nosebleeds are common.
The hepatic malformations may be insidious and range from ischemic bil-
iopathy, to focal nodular hyperplasia (FNH) with high output cardiac failure,
to noncirrhotic portal hypertension. Hepatic lesions include intraparenchy-
mal thick-walled veins with adherent arteries, scattered dilated sinusoids,
and abnormally sized portal tracts with extruded dilated vascular channels.
Commonly, grossly observed subcapsular enlarged vessels are present.
E. Miscellaneous
1. Granulomas of various sizes occur in the liver. Underlying etiologies are as
variable as the morphology of the granulomas. Stains for infection organ-
isms and evaluation under polarized light are useful in the evaluation of true
epithelioid types. Considerations specific to liver include PBC, sarcoidosis,
DILl, HCV, fungal or mycobacterial infections, foreign bodies, and hepa-
tocellular adenoma (HCA); rarely, granulomatous hepatitis is a bona fide
clinico-pathologic diagnosis.
2. Pregnancy is associated with various mitochondrial alterations. AFLP occurs
in the late third trimester and is potentially fatal to both mother and fetus;
emergent delivery is the treatment. While the histologic hallmark is zone 3
or diffuse microvesicular steatosis, oil red 0 stain on a frozen section may be
required since hepatitic features and extramedullary hematopoiesis may be
present. Endophlebitis is common. The affected hepatocytes appear swollen,
and the microsteatosis gives an appearance of cytoplasmic reticulation (e-Fig.
15.46). Preeclampsia/eclampsia and HELLP syndrome (hemolysis, elevated
liver enzymes, and low platelets) may cause zone 1 hemorrhage, necrosis,
and fibrin deposition.
3. Ductal plate malformation (DPM) results from developmental arrest with per-
sistence of the embryologic ductal plate, which assumes an anastomosing
ring-like structure lining the periphery of the portal tracts (e-Fig. 15.47).
DPM may manifest as von Meyenburg complexes, congenital hepatic fibro-
sis, Caroli's syndrome or disease, or polycystic liver disease.
a. Congenital hepatic fibrosis (e-Figs. 15.48 and 15.49) is a significant cause
of noncirrhotic portal hypertension. The abnormal portal tracts are
expanded, have an increased number of aberrant duct profiles, show
hypoplastic or absent portal veins, and numerous hypertrophic hepatic
artery branches. Bridging is noted but portal-central relationships are
maintained, as is synthetic function. Inspissated bile may be noted in
ectatic ducts.
b. Caroli's disease is characterized by segmental cystic dilatation of the larger
intrahepatic ducts, usually accompanied by recurrent bacterial cholangitis
and biliary lithiasis. When associated with congenital hepatic fibrosis, it
is termed Caroli's syndrome.
c. Polycystic liver disease is a debilitating process due to massive enlargement
with cystic replacement of the parenchyma (e-Fig. 15.50). Liver function
is maintained, but transplantation may be needed for quality of life. Berry
aneurysm of the middle cerebral artery is a significant complication.
4. Ascending cholangitis is diagnosed clinically. Histologically, intraluminal neu-
trophils are present within the interlobular bile ducts. Other features of
biliary obstruction are usually noted (e-Fig. 15.51).
IV. TRANSPLANTATION PATHOLOGY

A. Donor liver evaluation is performed on frozen sections to evaluate steatosis, or
portal inflammation and fibrosis in an HCV donor. For the former, percentages
of the core involved by large droplet steatosis are estimated; the presence of
>30% macrovesicular steatosis is commonly accepted as a cut-off potentially
associated with poor graft function in the immediate posttransplant period,
whereas >50% is unacceptable. Pitfalls of frozen artifacts include the tiny spaces
in hepatocytes and sinusoidal spaces that may be misinterpreted as large fat vac-
uoles. The use of oil red 0 stain is discouraged as it results in an overestimation
of steatosis due to small droplet fat that is not associated with graft dysfunction.
For HCV positive donors, portal inflammation, lobular activity, and fibrosis as
seen in H&E-stained sections are documented. Coagulative necrosis is a worri-
some finding in donor biopsies, and is worth both verbal communication and
written documentation.
B. Preservationlreperfusion injury is related to harvesting, transportation, and reper-
fusion of the graft. The relevant findings are located in zone 3; specifically, hep-
atocyte ballooning and cholestasis. Frank necrosis may occur in more severe
cases. Resolution is expected within 2 weeks.
C. Humoral (hyperacute) rejection rarely occurs with current patient management
paradigms. Historically, humoral rejection was associated with changes in
the liver, including coagulative and hemorrhagic necrosis or portaUperiportal
edema, neutrophilic portal infiltrates, and prominent ductular reaction.
D. Acute (cellular) rejection is directed primarily at antigens on duct epithelium
and venous endothelium. It can occur anytime immunosuppression is reduced
or discontinued, but is most common between 5 and 30 days after transplan-
tation. The classic histologic triad consists of mixed portal chronic inflamma-
tion, bile duct damage, and endotheliitis. The portal infiltrates consist of lym-
phocytes admixed with eosinophils, histiocytes, plasma cells, and occasional
neutrophils. Bile duct damage is characterized by inflammatory intercala-
tion into ductal epithelium; it may be accompanied by cytoplasmic vac-
uolization and other cytologic alterations. Nuclear pyknosis and cytoplasmic
eosinophilia are signs of ischemic injury, however (e-Fig. 15.52). Endotheli-
itis is defined as subendothelial lymphocytic infiltration with lifting, detach-
ment, and sloughing of endothelial cells; attachment of lymphocytes to the
luminal aspect of the endothelium is insufficient for diagnosis (e-Fig. 15.53).
Endotheliitis most frequently involves portal veins, but central veins can be
similarly affected. Centrilobular necrosis is sufficient for a diagnosis of severe
rejection.
Centers differ for treatment thresholds, thus, careful description and stan-
dard evaluation is recommended for optimal clinical communication. Acute
rejection may be graded using the Banff schema recommended by an interna-
tional panel (Table 15.3).
E. Chronic rejection (CR) is a misleading moniker as the characteristic feature, specif-
ically loss of bile ducts, may occur any time after transplantation. The diagnostic
criteria are: (i) bile duct loss affecting >50% portal tracts, or (ii) obliterative
arteriopathy by foamy histiocytes. Biliary epithelial senescence with eosinophilic
cytoplasm and nuclear hyperchromasia is considered an early feature of CR. The
arterial lesions mainly involve large and medium size vessels and are rarely seen
in small vessels sampled by percutaneous biopsy. Therefore, the diagnosis of
CR (Table 15.4) is primarily based on the evaluation of bile ducts and has been
divided into early and late stages (Hepatology. 2000;31:792). The early stage
typically shows perivenular hepatocyte dropout and central perivenulitis (e-Fig.
15.54); the late stage features duct loss in ~50% of portal tracts with variable
perivenular fibrosis (e-Fig. 15.55). Portal inflammation and ductular reaction
are typically insignificant in the late stage as the progenitor cell compartment
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G. Technical complications usually occur during the first few months after trans-
plantation. Hepatic artery or PVT or stricture may cause zone 3 hepatocyte
necrosis, but infarction is rare. Ischemic damage of the biliary tree is a sign
of hepatic artery complication, with protean manifestations including abscess,
cholestasis, obstructive changes, stricture, or loss of the bile ducts. Thus, CR
cannot be diagnosed without demonstration of a patent hepatic artery. Hepatic
vein thrombosis or stricture results in changes of venous outflow obstruction.
Stenosis or obstruction of the bile duct anastomosis causes morphologic changes
similar to those of biliary obstruction or biliary cirrhosis.
H. Recurrent diseases. Most diseases recur in the transplanted liver but the time-
frame and severity vary; diagnosis is complicated by the fact that recurrent dis-
eases share histopathologic features with rejection or technical complications.
Histologic evidence of recurrent HCV initially is acidophil bodies; portal
and lobular chronic inflammation occur later. Overlapping features with mild
acute rejection include bile duct damage, endotheliitis, and mixed infiltrates;
thus, a final diagnosis should include, if possible, description of the balance
of damage due to hepatitis versus rejection. HBV recurrence is documented by
protocol evaluation ofHB Sand C Ag testing on every follow-up allograft biopsy
(e-Fig. 15.56).
Fibrosing cholestatic hepatitis (FCH) (e-Fig. 15.57) is a rare but progressive
disease seen in both recurrent hepatitis B and C, with rapidly rising bilirubin
that may result in graft loss. FCH's histologic features include marked portal and
periportal perisinusoidal fibrosis with ductular reaction, canalicular cholestasis,
and nonzonal hepatocyte ballooning; the latter may be the earliest clue to diag-
nosis. Inflammatory changes are generally mild. In FCH-B, HB Sand C Ag are
highly expressed in reinfected hepatocytes.
PBC and PSC may recur several years after transplantation. Even with gran-
ulomatous duct lesions, recurrent PBC is difficult to diagnose. Recurrent PSC
needs to be distinguished from technical complications due to hepatic artery or
biliary stricture, or biliary obstruction. Recurrent steatosis and steatohepatitis
are a challenge to distinguish from de novo occurrence due to persistence of
the patient's metabolic syndrome and/or the medications utilized for allografts.
Recurrent HCC or cholangiocarcinoma are typically rapidly lethal complica-
tions.
I. Acute GVHD (e-Fig. 15.58) following bone marrow or stem cell transplantation
shows duct damage, and less frequently endotheliitis. Hepatitic forms of GVHD
rnay coexist. The differential diagnosis includes viral infection and DILl. Chronic
GVHD typically occurs after 100 days and simulates ischemia or ductopenic CR.
Skin and GI GVHD are commonly concurrent.
V. DIAGNOSTIC FEATURES OF COMMON NEOPLASTIC AND TUMOR-LIKE CONDITIONS. The
current World Health Organization (WHO) histologic classification of tumors of
the liver and intrahepatic bile ducts is given in Table 15.5. The 2010 American
Joint Committee on Cancer (AJCC) tumor, node, metastasis (TNM) staging is in
Table 15.6.
The most common tumor type in noncirrhotic livers is metastatic; neoplasms
that commonly metastasize to the liver include carcinomas of colorectal, pancreatic,
renal, pulmonary, and breast; melanoma; and neuroendocrine tumors. Metastases
usually present as multiple nodules, in contrast to the single nodules of primary
liver tumors. Metastases to a cirrhotic liver are very uncommon.
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is not caused by oral contraceptive use, but instead is a polyclonal
regenerative response to a local vascular injury. FNH has no malignant
potential. The nodularity of the lesion underlies the original moniker of
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Ill. lllla••muy;~ll-.• (lA), alaobown u relo,.:.ctolio: 1011-(45%
to 60%), cfo.c co IIIUIIdoll of IIP130 .!D. the majority oi a.oc&, hm:
~on oi !he i•ft•mrn.tex>ry m,trlrert SAA &lid. CliP by IHC.
Fo<lrl(:dy tt;nnt,d ~ I'NH,"lA ht.vt beco eh.owa to 'be-
adenomas by a variety of assays, and are associated with obesity and

alcohol use. IA are often hemorrhagic, have foci of sinusoidal dilatation
and peliosis, and have abnormal vessels; chronic inflammatory infil-
trates are common along fibrovascular septa with periseptal ductular
reaction (e-Fig. 15.63). lA may be confused with FNH by imaging and
initial gross and microscopic evaluation, but glutamine synthetase is
not "map-like."
iv. Unclassified adenoma (1 0%), which is without specific genotypic or phe-
notypic features.
2. Premalignant. DN, a well-defined premalignant lesion arising in the back-
ground of cirrhosis (Semin Liver Dis. 2005;25:133; Gastroenterol Clin N
Am. 2007;36:867), is diagnosed by the presence of a nodule with abnormal
architecture, clonal hepatocytes with an increased N/C ratio, and nuclear
atypia. DNs are usually multiple in livers with chronic liver disease or
cirrhosis. Additional features include gross differences from surrounding
nodules including bulging above the background, expansile growth with
pushing borders, small cell change (nuclear crowding and sinusoidal align-
ment), resistance to iron accumulation in an otherwise iron-loaded cirrhosis
(e-Fig. 15.64), clear cell change or deep eosinophilia, and the presence of
isolated artery branches. Unlike HCC, DNs do not have cords >3 nuclei
thick or stromal invasion. Macroregenerative nodules with intralesional
portal tracts are considered low-grade DN. DN may contain small foci
of nonencapsulated HCC within them, referred to as "nodule-in-nodule"
(N-1-N).
DNs are rarely >2 em in size. When the lesion is <0.1 em, it is termed
a dysplastic focus. Large cell change, although associated with the presence
of HCC, is not itself considered premalignant. Large cell change (LCC) is
characterized by cellular and nuclear enlargement with nuclear atypia, but
a normal N/C ratio. Small cell change (SCC), on the other hand, is strongly
associated with progression to HCC.
3. Malignant epithelial neoplasms
a. HCC, more common in middle aged and older men, usually but not exclu-
sively occurs in chronic liver disease with a cirrhotic background. Chronic
viral infection accounts for up to 85% of HCC; alcohol abuse is the most
significant nonviral cause. Hereditary hemochromatosis (HH) carries a
significant risk. Tobacco use is an additive factor. Obesity and the asso-
ciated complications of diabetes and steatohepatitis are also recognized
risks, and multiple risk factors increase the overall risk.
HCC is usually encapsulated in cirrhotic livers, but not in noncir-
rhotics. Prior to the advent of advanced imaging for surveillance, several
grossly observable types of HCC were documented: the single nodular
type, the multifocaVmulticentric type, the diffuse (cirrhotomimetic) type,
and the uncommon pedunculated type. Tumor nodules are soft, fatty (yel-
low) or green (bile) stained, and bulge above the cut surface. Central necro-
sis is not rare, particularly in TACE or radio frequency ablation (RFA)
treated HCC. With advanced surveillance in known cirrhotics, detected
tumor nodules are often small (< 1 to 2 em) and subtle.
Microscopically, HCC assumes a variety of patterns: trabecular, pseu-
doglandulat; acinar, compact, or scirrhous. Unattached trabeculae are
referred to as "floating trabeculae" and are characteristic of HCC. Mixed
patterns are common, particularly with increased tumor size. Cell plates
are >3 nuclei wide, a feature of importance in distinguishing DNs from
early HCC (discussed later). Isolated arteries noted within the tumor
can be detected with trichrome, aSMA, and CD34 IHC. HCC receives
blood solely from systemic arteries, thus CD34 positive tumor sinusoidal
endothelium differs from CD34 negative endothelium of nontumor liver.

Reticulin stain highlights widened cell plates and shows reduction or loss
of normal reticulin fibers (e-Fig. 15.65). Tumor cells share many char-
acteristics of nonneoplastic liver cells, for example, intranuclear inclu-
sions, eosinophilic or basophilic cytoplasm; small droplet, large droplet,
or microvesicular steatosis; clear cell change; MDBs; PASd inclusions; fib-
rinogen inclusions; and HB S Ag. Bile production by tumor cells must be
distinguished from trapped hepatocytes, but can be useful to identify the
tumor as hepatocellular; polyclonal CEA and CD10 react with canaliculi
and are equally specific, as hepatocytes are the only cells that contain these
structures and secrete bile (e-Fig. 15.66). Arginase-1, another marker of
hepatocellular origin, may be useful in selected cases. The presence of
stromal invasion is diagnostic, but uncommon; vascular invasion is an
important parameter for tumor staging.
Small HCC (<2 em) may or may not be well differentiated; most
HCC are moderately or poorly differentiated. Well differentiated HCC
can be difficult to distinguish from adenoma in noncirrhotic liver or from
DNs in cirrhosis; if present, stromal invasion and vascular invasion are
diagnostic of HCC. Loss of K7 or K19 positive ductular reaction at the
periphery of an encapsulated lesion may signify stromal invasion, and thus
malignancy. The oncofetal protein glypican-3 (GPC-3) may be useful when
positive; the reactivity is inhomogenous in a cytoplasmic, membranous,
and/or canalicular pattern in up to 70% to 80% ofHCCs, but is negative
in HCA, although a small fraction of dysplastic and cirrhotic nodules
have been reported as immunopositive for GPC-3. Glutamine synthetase,
positive only around central veins normally, is commonly diffusely positive
inHCC (e-Fig.15.67), and heat shock protein 70 (HSP70) is positive in the
majority of HCC. Some investigators advocate the use of a panel of GPC-
3, glutamine synthetase, and HSP70 as diagnostic tools in difficult cases
(] Hepatol. 2009;50:746). The presence of >2% positive K19 tumor cells
has been proposed as a poor prognostic marker in HCC, although the
specificity of the finding remains unsettled.
Other variants of HCC are listed in Table 15.5. By international con-
sensus, early HCC is a low-grade, low-stage, small HCC (Hepatology.
2009;49:658) that has grossly and microscopically indistinct borders from
the surrounding cirrhotic liver, and may be difficult to distinguish from a
high-grade DN. Early HCC may be fatty. Nodules of encapsulated HCC
that are distinct from the background are not categorized as independent
early HCC since they represent biologically advanced tumors.
b. Fibrolamellar HCC (FL-HCC) occurs in noncirrhotic livers and is character-
ized by large eosinophilic polygonal tumor cells with prominent nuclei
and nucleoli in a background of paucicellular lamellar fibrous bands. FL-
HCC may bear resemblance to FNH by imaging and grossly. Some of the
tumor cells may contain cytoplasmic pale bodies due to the presence of
fibrinogen (e-Fig. 15.68). The slightly better prognosis of FL-HCC likely
is a result of occurrence in nondiseased, noncirrhotic livers in patients
younger than is common for HCC. The primary distinction in biopsy is
with the scirrhous variant of HCC; the morphology of the hepatocytes is
key. Clinical information and serum markers may be helpful. In contrast
to HCC, serum AFP is rarely elevated in FL-HCC; however, vitamin B12,
vitamin B12 binding capacity, and serum carcinoembryonic antigen may
be elevated in FL-HCC.
B. Malignancies of mixed cellular origin
1. Combined hepatocellular-cholangiocarcinoma (HCC-CCa). According to the
2010 WHO classification, combined HCC-CCa is distinguished from
collision tumors which contain typical HCC and typical CCa separately in
the same liver. In contrast, in combined HCC-CCa, the elements are admixed
within a single tumor. Combined HCC-CCa is further subdivided into a
"classical type," in which all elements are typical by light microscopy and
rnc, and subtypes with stem-cell features including "typical," "intermedi-
ate," and "cholangiocellular." Notably, in the "typical" HCC-CCa, either
intermediate cells (with features of both hepatocellular and biliary differen-
tiation) or stem-like cells may be found at interfaces of the two components
of the HCC-CCa. The greater the biliary differentiation of HCC-CCa, the
more abundant the sclerotic stroma. The outcome of combined HCC-CCa is
worse than traditional HCC, but lack of uniform terminology has prohibited
large and comparative series.
2. Hepatoblastoma, a primary malignant blastomatous neoplasm of several cell
lineages, is rare but is nonetheless the most common liver tumor of children.
Up to 80% to 90% of cases present by 5 years, and prematurity and low
birth weight are associations. Most present as symptomatic masses in the
right lobe.
a. The wholly epithelial type consists of four subtypes: fetal, mixed fetal
and embryonal, macrotrabecular, and small cell undifferentiated (SCUD).
Fetal cells, resembling adult hepatocytes but smaller in size, contain vari-
able amounts of cytoplasmic fat and glycogen, giving rise to an alternating
light-and-dark pattern (e-Fig. 15.69). Embryonal cells have a higher N/C
ratio and may form nests, rosettes, or small tubules. The macrotrabecular
has a growth pattern reminiscent of HCC.
b. The mixed epithelial-mesenchymal (MEM) type, with or without teratoid
features, contains malignant mesenchymal components such as cartilage
and osteoid in addition to the epithelial elements.
c. Hepatoblastoma, NOS, is the final category.
The outcome of hepatoblastoma has improved significantly with
the improved ability to shrink tumors prior to surgical extirpation.
Chemotherapy induces a variety of predictable tumoral alterations. Hepa-
toblastoma staging systems are based on pretreatment features and postre-
section findings.
C. Tumors of the biliary origin
1. Benign
a. Biliary cysts, whether solitary or as components of the ductal plate malfor-
mation (DPM) (described previously), are lined by a single layer of benign,
flattened, cuboidal or columnar biliary-type epithelium, surrounded by a
variable amount of fibrous tissue.
b. The von Mayenburg complex (e-Fig. 15.70) represents persistence of the
ductal plate; it is a common finding in normal livers, and may dilate to be
the origin of solitary cysts or cysts of polycystic livers. The VMC is usually
adjacent to a portal tract and appears as dilated duct-like structures within
mature fibrous stroma.
c. Ciliated foregut cyst {e-Fig. 15.71), while rare, may be found by imaging
or incidentally during surgery. It is subcapsular and often in segment 4.
The lining is composed of ciliated pseudostratified columnar epithelium
lying on a basement membrane.
d. Bile duct adenoma/peribiliary gland hamartoma is composed of closely
packed, well-formed, and relatively uniform small ducts that form a 1
to 20 mm diametet; sharply demarcated nodule (e-Fig. 15.72). Careful
microscopic examination is necessary to exclude the angulated architec-
ture, desmoplastic stroma, cytologic atypia, mitotic activity, and evidence
of invasive growth that are indicative of cholangiocarcinoma or metastatic
adenocarcinoma.
Chapter 15 • The Liver I 269
2. Premalignant
a. Biliary intraepithelial neoplasia, grade 3 (BILIN3) is a precursor of intra-
hepatic cholangiocarcinoma (ICC) characterized as flat dysplasia with
multilayering of nuclei and micropapillary intraluminal projections. The
morphologic findings are identical to of extrahepatic ducts.
b. Intraductal papillary neoplasm (lPN) is another precursor of ICC. This cat-
egory of neoplasms has replaced lesion previously known as biliary papil-
loma and papillomatosis (e-Fig. 15.73). Synchronous or metachronous
IPNs may develop throughout the biliary system. IPNs are classified as
low, intermediate, or high grade on the basis of cellular and nuclear
features; the parallels with pancreatic intraductal lesions are many, and
up to one-third are mucin producing. Most of the intrahepatic papil-
lary neoplasms are lined by biliary epithelium, although intestinal, onco-
cytic, or gastric type may also be present. Ducts may be massively
expanded by the villous growth and distinction with MCN requires thor-
ough evaluation of the stroma (e-Fig. 15.74). Invasive carcinoma may
be found in association with lPN; perineural invasion is common. Col-
loid carcinoma is the type of malignancy that occurs with intestinal-type
lPN.
c. Mucinous cystic neoplasm, formerly known as biliary cystadenoma as in
the gallbladder and pancreas, occurs in women, does not communicate
with the biliary tree, and consists of cystic dilated structures lined by a
layer of mucin-producing columnar cells on a spindled-type mesenchy-
mal stroma (e-Fig. 15.75). It may additionally show low- or high-grade
intraepithelial neoplasia and is graded on the basis of the highest compo-
nent; thus, numerous sections need to be evaluated. Cases associated with
an invasive carcinoma occur in older patients.
3. Malignant Intrahepatic biliary neoplasms
a. Intrahepatic cholangiocarcinoma (ICC) is a nonencapsulated mucin-
secreting adenocarcinoma usually arising in noncirrhotic livers. Cirrhosis
or chronic liver disease cannot be used as evidence to exclude the diagno-
sis, however. The tumor is characterized by three growth patterns that may
coexist: mass-forming, periductal, and intraductal; it is the mass-forming
pattern that results in the prominent desmoplastic background with the
characteristic firm, white or gray gross appearance.
The tumor is an adenocarcinoma with a variety of architectural pat-
terns resembling canals of Hering, cholangioles, bile ducts of varying sizes,
or peribiliary glands. In larger ICC, the central areas may be mostly stroma
and paucicellular, while the peripheral regions may show a growth pattern
that replaces and incorporates portal tracts. Greater than 90% express
keratins 7 and 19, and about 40% also express keratin 20; most cases are
also positive for expression of CEA (using a monoclonal antibody) and
CA19-9. ICC may be indistinguishable from metastatic ductal adenocarci-
noma of the pancreas by histologic and immunohistochemical evaluation.
ICC involving only the peribiliary glands may be challenging to distinguish
from reactive atypia.
The prognosis of ICC is dismal due to the lack of effective treatments.
The exceptions are small peripheral ICCs found incidentally, the muci-
nous intraductal growth ICC that are completely resected, and the highly
selected cases of perihilar cholangiocarcinoma arising in PSC treated with
protocolized neoadjuvant chemotherapy and liver transplantation. The
2010 AJCC TNM Staging for carcinomas of the intrahepatic bile ducts is
in Table 15.7.
b. Cystadenocarcinoma occurs in men as commonly as in women, and no
spindled stroma is present. These tumors likely arise from lPN within a
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Chapter 15 • The Liver I 271
is characterized by small vascular channels lined by a single layer of plump

endothelial cells (e-Fig. 15.78) in scant stroma with trapped ducts.
4. Epithelioid hemangioendothelioma is a low- or intermediate-grade malignancy.
The epithelioid tumor cells have abundant eosinophilic cytoplasm, and may
be difficult to discern initially as they are arranged as single cells or small
dusters of cells in a dense or myxoid fibrous stroma, and may occasionally
be dendritic or spindled. Some tumor cells contain an intracytoplasmic vas-
cular lumen simulating signet-ring-cell carcinoma (e-Fig. 15.79). Sinusoidal
growth of the tumor cells at the periphery of the lesion is characteristic, often
accompanied by hepatocyte atrophy. The tumor may grow into and occlude
outflow veins. The tumor cells express at least one endothelial marker such
as CD34, CD31, or factor VIII-related antigen.
5. Angiosarcoma, a highly aggressive tumor, is the most common mesenchymal
malignancy of the liver. The characteristic feature of the tumor is a sinu-
soidal growth pattern by hyperchromatic, plump, malignant endothelial cells
with little associated stromal response (e-Fig. 15.80). Solid growth patterns
may pose diagnostic challenges, but immunostains demonstrate the vascular
nature of the tumor.
6. Kaposi sarcoma consists of a pure spindle-cell proliferation with slit-like
spaces containing extravasated red cells, and is restricted to patients with
human immunodeficiency virus (HIV).
Cytopathology of the Liver

Julie Elizabeth Kunkel and Hannah R. Krigman
I. INTRODUCTION. Fine needle aspiration of the liver is typically prompted by the

presence of a mass and can be achieved via percutaneous ultrasound or CT-
guided fine needle aspiration; lesions in the left lobe can be aspirated by endo-
scopic ultrasound-guided FNA (EUS-FNA) (Gastrointest Endosc. 2002;55:859).
Hepatic FNA is a very safe procedure; fatal complications are rare, hemorrhage
being the most common which occurs at a rate of 0.006% to 0.031% (Radiology.
1991;178:253). The sensitivity for malignancy ranges from 76% to 95%, with
a specificity dose to 100% (Diagn Cytopathol. 2002;26:283; Diagn Cytopathol.
2000;23:326).
As with all aspirates, during interpretation it is critical to bear in mind the inter-
vening tissue the needle must pass through, which rna y be a source of confusion and
misdiagnosis. Examples complicating hepatic FNA interpretation include benign
or malignant cells originating from the pleura, peritoneum, and lung. Familiarity
with and recognition of normal hepatic elements, including bile ductular cells,
Kupffer cells, and endothelial cells, are also crucial to avoid a false-positive diag-
nosis of malignancy (World] Surg Oncol. 2004;2: 1 ). Reactive hepatocytes in par-
ticular may pose a diagnostic challenge in the distinction from a well-differentiated
hepatocellular carcinoma (HCC) (Arch Pathol Lab Med. 2002;126:670).
II. BENIGN NON-NEOPLASTIC MASSES. Hydatid cysts, abscesses, granulomas, bile duct
adenoma/von Meyenburg complexes, focal nodular hyperplasia (FNH), and regen-
erative nodules are examples of benign non-neoplastic masses.
A. The aspirate of an abscess shows abundant neutrophils and necrotic debris
(e-Fig. 15.81). Amebic abscesses contain more necrotic debris and fewer inflam-
matory cells. Culture and special stains are usually necessary to identify
microorganisms (e-Fig. 15.82).
For cases of suspected abscess, it is always necessary to rule out the pres-
ence of a necrotic or infected neoplasm by extensive sampling and careful cyto-
logic evaluation for the presence of viable tumor cells which may be obscured
by the inflammation. In the instance of suspected tumoral necrosis or well-
differentiated HCC, re-biopsy with attention toward the periphery of the lesion
should be recommended, such that the interface of the lesion and surrounding
normal parenchyma may be examined (Radiology. 1997;203:1 ).
B. In the case of hydatid cyst, the aspirated fluid may be clear or turbid. Lami-
nated cyst walls, scolices, and hooklets are observed; a neutrophilic background
is sometimes present. Although aspiration of a hydatid cyst poses a risk of
anaphylactic reaction, successful procedures are the norm (Diagn Cytopathol.
1995;12:173).
Ill. VASCULAR LESIONS
A. The aspirate of a hemangioma usually shows abundant blood. Scattered stro-
mal fragments with bland elongated spindle cells are characteristic (Diagn
Cytopathol. 1998;19:250). Cell block preparations are helpful to identify the
vascular channels.
B. Aspirates of epithelioid hemangioendothelioma are paucicellulat; containing
single cells and small tissue fragments, and display a spectrum of cytomorphol-
ogy from small bland-appearing epithelioid and spindle cells, to malignant large
tumor cells. The epithelioid cells have abundant cytoplasm and may contain
characteristic intracytoplasmic lumina or sharply defined intranuclear cytoplas-
mic inclusions (Acta Cytol. 1997;41:5).
C. In angiosarcoma, the aspirate shows abundant blood in which there are iso-
lated cells and loose clusters of cells. The malignant cells are spindle-shaped to
epithelioid, and have hyperchromatic nuclei and abundant but ill-defined cyto-
plasm. Necrosis is present. Scattered malignant cells may show hemosiderin-
laden cytoplasm or erythrophagocytosis (Diagn Cytopathol. 1988;18:208).
Vasoformative structures such as intracytoplasmic lumina, microacinar lumen
formation, and vascular channels are identified inconsistently (Anat Pathol.
2000;114:210).
IV. FNH AND HCA. These entities typically occur in distinct clinical scenarios as a solitary
nodule and as such require clinical, pathologic, and radiologic correlation for the
correct diagnosis. In both cases, the diagnosis rests on evaluation of the presence
or absence of cytologically benign liver elements; it is therefore critical that only
lesional tissue is sampled (World] Surg Oncol. 2004;2).
A. FNH. Aspirates of FNH show both abundant benign hepatocytes and benign
biliary epithelial cells (Acta Cytol. 1989;33:857).
B. HCA aspirates consist of benign hepatocytes without biliary epithelial cells (Acta
Cytol. 1989;33:857).
V. HEPATOCELLULAR CARCINOMA (HCC). The most characteristic and specific features
are thickened hepatocyte trabeculae rimmed by spindle-shaped endothelial cells
(e-Fig. 15.83), hepatocyte tissue fragments with well-defined traversing capil-
laries (e-Fig. 15.84), increased nuclear to cytoplasmic ratio (e-Fig. 15.85), and
frequent atypical naked nuclei (e-Fig. 15.86) (Diagn Cytopathol. 1999;21:370;
Cancer. 1999;87:270). Poorly differentiated HCC demonstrates loose nests, three
dimensional fragments, and occasional gland-like structures of malignant hepato-
cytes with marked pleomorphism, macronucleoli, necrosis, and numerous mitoses
(Cancer. 2004;102:247). Features that favor hepatocytic origin include polygonal
shaped cells with centrally placed nuclei, abundant granular cytoplasm, and bile
pigment (e-Fig. 15.86); however, the distinction from cholangiocarcinoma and
metastatic adenocarcinoma is challenging and may require immunostains (Arch
Pathol Lab Med. 2007;131).
The fibrolamellar variant of HCC has a distinct cytomorphology which
includes poorly cohesive clusters of cells and singly dispersed large monotonous
cells with abundant granular cytoplasm, a low nuclear to cytoplasmic ratio, promi-
nent nucleoli, and intracytoplasmic hyaline globules. Fragments of lamellar colla-
gen bands with benign spindle-shaped cells are present. The thickened trabeculae
typical of classic HCC are not identified (Diagn Cytopathol. 1999;21:180).
VI. CHOLANGIOCARCINOMA. The aspirate is composed of cells arranged in crowded
sheets, three-dimensional clusters, acinar structures, or as singly dispersed cells.
The malignant cells (e-Figs. 15.87 and 15.88) show a high nuclear to cytoplasmic
ratio, irregular nuclear membranes, prominent nucleoli, and occasional intracyto-
plasmic mucin (Cancer. 2005;105:220). Poorly differentiated carcinoma displays
marked nuclear pleomorphism and necrosis.
Combined hepatocellular-cholangiocarcinoma is a hi-phenotypic tumor arising
from the canal of Hering cells with features of HCC and cholangiocarcinoma. The
tumor bridges these two entities both morphologically and by immunohistochem-
istry. These tumors (e-Fig. 15.89) are cytologically malignant but usually require
immunostains for correct categorization as the malignant cells exhibit of spectrum
of differentiation from hepatoid to glandular (Acta Cytol. 1997;41:1269).
VII. METASTATIC MALIGNANCY. Metastasis from an extrahepatic primary tumor is the
most common malignancy of the liver (Diagn Cytopathol. 2000;23:326). The most
common primary tumors that metastasize to the liver are adenocarcinomas aris-
ing from the colon (e-Fig. 15.90), lung, pancreas, breast, and kidney. Comparison
with the primary malignancy in cases of metastases is essential for diagnosis, as is
appropriate immunohistochemical characterization. Carcinomas with polygonal
cell morphology, such as neuroendocrine tumors, renal cell carcinoma, adrenocor-
tical carcinoma, and others must be differentiated from HCC on the basis of their
immunoprofile (Arch Pathol Lab Med. 2007;131:1648).
The Gallbladder and
Extrahepatic Biliary
Tree
I. NORMAL ANATOMY. The gallbladder, comprised by the fundus, body, and neck, is
covered by serosa, except the portion in the liver fossa which merges with liver
parenchyma. The lining mucosa, a layer of folded columnar epithelium and lam-
ina propria of loose connective tissue, directly rests on muscularis propria which
consists of longitudinally oriented, to irregularly arranged bundles of smooth mus-
cle with overlying subserosa and serosa. No muscularis mucosae or submucosa
are present. Secretory mucous glands in the neck and extrahepatic bile ducts are
arranged in a lobular pattern (e-Fig. 16.1)."'
The extrahepatic ducts include the right and left hepatic ducts, which join to
form the common hepatic duct in the porta hepatis; when the common hepatic
duct is joined by the cystic duct, the common bile duct is formed. A single layer of
columnar cells lines the ducts and rests directly on dense connective tissue; from
proximal to distal, there is a variable periductal smooth muscle fiber investment,
intermingled with collagen bundles.
II. GROSS EXAMINATION
A. Cholecystectomy is most commonly performed for cholelithiasis. After the
gallbladder is measured and opened longitudinally, the following should be
described: serosal, mural, and mucosal appearances; cystic duct integrity; and
consistency, quantity, and color of stones. Full-thickness sections should be sub-
mitted from the fundus, body, neck, and duct; the cystic duct margin should also
be submitted, as well as any lymph nodes. For a suspicious lesion, the overly-
ing serosal surface or hepatic bed should be inked, the lesion breadloafed, and
sections taken to demonstrate relevant anatomic relationships.
The gross finding that the gallbladder wall is uniformly firm with an asso-
ciated flattened mucosal surface suggests the diagnosis of a so-called porcelain
gallbladder. After the specimen is photographed, at least one section per em
should be submitted (if not the entire specimen) to exclude adenocarcinoma
(e-Figs. 16.2 and 16.3).
B. Biopsy of the common bile duct is performed for stricture or overt neoplasm
during endoscopic retrograde cholangiopancreatography (ERCP). The num-
ber and dimensions of specimens should be recorded to ensure that the biopsy
fragments are adequately represented; inking is not needed. At the time of ini-
tial histologic sectioning, preparation of three hematoxylin and eosin (H&E)
stained slides together with six additional unstained slides avoids resurfac-
ing the block if subsequent deeper levels or special stains are required for
diagnosis.
C. Frozen section. Evaluation of bile duct margins by frozen section during pan-
creatoduodenectomy, or liver resections for bile duct adenocarcinoma, is often
performed. The tissue should be frozen in its entirety, oriented in the frozen sec-
tion block to obtain enface sections, and cut deeply to obtain sections that
274
Chapter 16 • The Gallbladder and Extrahepatic Biliary Tree I 2 75
represent the entire margin so that small foci of tumor are not missed by
inadequate sampling. The tissue that remains after frozen section should be
submitted for evaluation by permanent sections, which helps assure adequate
sampling.
Ill. DIAGNOSTIC FEATURES OF COMMON NONNEOPLASTIC CONDITIONS
A. Cholecystitis is associated with cholelithiasis in > 90% of the cases. Acute chole-
cystitis is characterized by full thickness edema, congestion, and an associated
fibrinopurulent serosal exudate. Hemorrhage, transmural necrosis (gangrenous
cholecystitis), and/or perforation may occur (e-Figs. 16.4 and 16.5).
Chronic cholecystitis is variably characterized by mural hypertrophy or atro-
phy with fibrosis and chronic inflammation (e-Fig. 16.6). Intestinal, pyloric, or
foveolar surface metaplasia may occur. Rokitansky-Aschoff sinuses, which are
herniations of the lining mucosa into the muscle layers, are common. Adeno-
myoma represents exaggerated herniations in the fundus accompanied by mus-
cular hypertrophy and may appear as a gross deformity (e-Figs. 16.7 to 16.9).
Both xanthogranulomatous cholecystitis (due to rupture of Rokitansky-Aschoff
sinuses) or mucosal ulceration from stones may be transmural with associated
bile extravasation and accumulation of foamy macrophages. Acalculous chole-
cystitis may be acute or chronic. Follicular cholecystitis may be associated with
primary sclerosing cholangitis (PSC) (e-Fig. 16.10).
B. Cholesterolosis (strawberry gallbladder) is characterized by yellow mucosal
specks grossly and lipid-laden macrophages in the lamina propria microscopi-
cally (e-Fig. 16.11). It is an incidental finding of no clinical significance.
C. Choledochal cyst, a form of fibropolycystic disease, results in fusiform or spheri-
cal dilatation of the common bile duct. Following photographic documentation,
the entire lesion should be submitted for microscopic examination to exclude
biliary intraepithelial neoplasia (BillN) or adenocarcinoma (e-Fig. 16.12).
D. Biliary atresia is a congenital process in which the extrahepatic ducts and gall-
bladder may be completely absent, or replaced by fibrous cords with no or only
a very small lumen.
E. PSC involving the extrahepatic biliary system is an idiopathic disease diagnosed
by cholangiography (discussed in more detail in Chap. 15).
F. Secondary sclerosing cholangitis, histologically indistinguishable from PSC, has
a variety of obstructive and nonobstructive etiologies, including tumors, toxins,
ischemia, and infections (including AIDS cholangiopathy).
IV. DIAGNOSTIC FEATURES OF COMMON NEOPLASMS AND PRECURSOR LESIONS (Table
16.1)
A. Adenoma is a single, small, and incidentally found polypoid lesion, and is char-
acterized by a tubular, papillary, or tubulopapillary architecture. A pyloric or
intestinal type epithelium is more common than a biliary type epithelium; squa-
mous morules, Paneth cells, and neuroendocrine cells may be present. By def-
inition, all adenomas are low grade, but larger adenomas may harbor foci of
high-grade intraepithelial neoplasia or invasive carcinoma and thus should be
entirely submitted for microscopic examination.
B. Biliary intraepithelial neoplasia (BiliN) is a classification nomenclature intro-
duced in 2010 by the World Health Organization (WHO). BiliN-1 and BiliN-2
(low and intermediate grade lesions) are incidental and without established clin-
ical significance. BiliN-3 may be associated with invasive carcinoma, and thus if
present in the gallbladder, thorough sampling (including of the cystic duct and
margin of excision) is necessary to exclude invasive carcinoma. If no invasive
carcinoma is present, and the surgical margin of the cystic duct is not involved,
cholecystectomy is considered curative. A distinguishing characteristic between
BiliN-3 and reparative atypia is the abrupt transition noted in the former (e-Fig.
16.13) compared with the gradual alterations and heterogenous, widespread
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bile ducts.
2. IPMNs of the extrahepatic bile ducts share the epithelial types characteristic
of pancreatic IPMNs, including pancreatobiliary, intestinal, oncocytic, or
gastric. Papillomatosis, a recurrent and potentially multicentric condition of
the entire biliary tree, is considered a subset of IPMN.
3. Mucinous cystic neoplasm (MCN) has replaced the term cystadenoma. More
common in the extrahepatic ducts than gallbladdex; MCN may grow to
20 em in maximal dimension. As in its hepatic counterpart, the lesion
contains estrogen receptor and progesterone receptor positive mesenchymal
stroma. As with papillary neoplasms, MCN lesions may be associated with
invasive adenocarcinoma.
D. The classification of adenocarcinomas of the gallbladder and cystic duct is shown
in Table 16.1. Tumors of both sites have similar staging schemes (Table 16.2).
1. Adenocarcinoma of the gallbladder may grossly result in mural indura-
tion and mimic chronic cholecystitis, or grow as an intraluminal poly-
poid lesion (e-Fig. 16.14). In order of decreasing frequency, the common
subtypes are biliary, intestinal (tubular or goblet), and gastric foveolar
(e-Fig. 16.15).
2. Biliary adenocarcinomas may contain mixed cell types including intestinal,
goblet, and neuroendocrine cells. The tumors composed of intestinal type
cells are usually 1<20 and CDX2 immunopositive; the goblet cell variant
may also have Paneth and neuroendocrine cells. The gastric foveolar type is
usually well-differentiated (e-Figs. 16.16 and 16.17).
E. Adenocarcinomas of extrahepatic bile ducts have been reclassified by the AJCC as
perihilar or distal (Fig. 16.1), and each has its own staging scheme (Tables 16.3
and 16.4). Perihilar tumors arise proximal to cystic duct and account for 60% of
the adenocarcinomas; the distal tumors arise between the junction of the cystic
duct and common bile duct and the ampulla of Vater, and account for 30% of the
adenocarcinomas (e-Fig. 16.18). Grossly, there may be only subtle thickening of
the duct wall, and thorough sampling is required to evaluate margins and local
extension.
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Cytopathology of the Gallbladder

and Extrahepatic Biliary Tree
Brian Collll\8 and Julia Elizabeth Kunkel
I. IAWILAIIIIU
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Chapter 16 • The Gallbladder and Extrahepatic Biliary Tree I 2 81
a benign/inflammatory stricture shows flat sheets of ductal epithelium with

minimal crowding and overlap. The nuclei are round to oval with fine chro-
matin, nucleoli, and minimal pleomorphism (e-Fig. 16.20). The background
can show granular debris of a postobstructive process. Single atypical cells
are not seen. It is important to know if a stent is in place since stents can cause
significant reactive atypia which can be confused with neoplasia.
2. PSC. Patients with PSC can have multiple strictures throughout the biliary
tree that are sampled and followed by bile duct brushing (Clin Liver Dis.
2010;14:349). When the strictures are inflammatory in nature, they typically
show a florid repair appearance consisting of nuclear enlargement, promi-
nent nucleoli, and cellular crowding and overlap with some loss of polar-
ity. Intraepithelial neutrophils can be present (e-Figs. 16.21 and 16.22). A
spectrum of reactive atypia is present without a distinct second population.
Overall, the findings do not meet the criteria required for the diagnosis of
carcinoma as detailed below. Howeve.r; the epithelial atypia present cannot
always be clearly categorized, and in these instances a descriptive diagnosis
of "epithelial atypia" can be appropriate.
B. Neoplastic. Cholangiocarcinoma is most commonly encountered in bile duct
brushings, but the advent of endoscopic ultrasound-guided fine needle aspira-
tion (EUS-FNA) is providing more access and better sampling of this neoplasm in
the biliary tree. Nonetheless, the lesion's underlying sclerotic nature can present
a challenge in obtaining sufficient diagnostic cellular elements U Clin Oneal.
2005;23:4561). By bile duct brushing, there is typically a mixed population of
benign/reactive ductal epithelial cells and a second population of ductal carci-
noma cells. The ductal carcinoma component shows small syncytial groups and
sheets of cells with crowding and overlap, and single cells. The nuclei are enlarged
with granular chromatin, nuclear membrane irregularities, anisonucleosis, and
nucleoli (e-Figs. 16.23 and 16.24). The application of UroVysion FISH to bile
duct brushing specimens can assist in the diagnosis (e-Fig. 16.25). By FNA, the
aspirate smears can be variably cellular and show small to intermediate groups
arranged as syncytial fragments and flat sheets. The nuclei possess nuclear mem-
brane irregularities, coarse chromatin, nucleoli, and show anisonucleosis.
The Pancreas
Dengfeng Cao and Hanlin L. Wang
I. NORMAL ANATOMY. The pancreas is located in the retroperitoneum. In adults, the

pancreas measures 15 to 20 em in length and weighs 85 to 120 g. The pancreas is
divided into four parts: the head (including the uncinate process), neck, body, and
tail. The vascular supply to pancreas is from branches of celiac trunk and superior
mesenteric arteries. The lymph nodes draining the pancreas consist of two major
systems: those ringing the pancreas, and those near the aorta from the level of celiac
trunk to the origin of the superior mesenteric artery. Microscopically, the pancreas
consists of lobules that include both exocrine and endocrine components.
The vast majority of the exocrine component is the acinar epithelium. The acinar
cells are large and polarized, with basally situated nuclei; the apical cytoplasm is
eosinophilic due to the presence of abundant zymogen granules, whereas the basal
portion is basophilic and contains abundant rough endoplasmic reticulum. The
second component of the exocrine pancreas is the duct system that begins with
the centroacinar cells. The centroacinar cells then drain into intercalated ducts,
intralobular ducts, interlobular ducts, and then into the main pancreatic duct of
Wirsung and the accessory duct of Santorini. The duct system is lined by cuboidal
to low columnar cells with no visible cytoplasmic mucin by hematoxylin and eosin
(H&E) staining.
The endocrine component, constituting 1% to 2% of the pancreas in adults,
is composed of islets of Langerhans and extrainsular endocrine cells scattered
among the acini and ducts. The islets of Langerhans contain four major cell types:
insulin-secreting {3 cells (60% to 70%), glucagon-secreting a cells (15% to 20%),
somatostatin-secreting li cells, and pancreatic polypeptide-secreting PP cells.
A. Biopsy, fine needle aspiration, and brushing cytology specimens may be obtained
percutaneously, intraoperatively, or via endoscopic retrograde cholangiopancre-
atography (ERCP). Needle core biopsies should be immediately fixed in 10%
formalin. The number and the length, or length range, of the biopsies should
be recorded. Documentation of the number and length is important to ensure
that the biopsies are adequately represented on the slides. Three H&E-stained
slides from each block are prepared for microscopic examination. Aspiration
smears are processed with alcohol fixation for Papanicolaou staining or air dried
for Diff-Quik staining. Brushings are handled by routine liquid-based cytology
methods.
B. Distal pancreatectomy specimens consist of the pancreatic tail (usually with
attached spleen) and a portion of the pancreatic body. After orientation, each
organ should be measured separately. An externally visible mass or lesion should
be documented for its size, extent, and location. The proximal and peripan-
creatic margins should be inked. Before sectioning through the pancreas, the
proximal margin is typically taken either as a shave margin or as a series of
perpendicular sections, depending on the location of tumor. The method of sec-
tioning the pancreas depends on personal preference. One method is to bivalve
the pancreas using a probe in the main duct as a guide; another method is to
section the pancreas in a breadloaf fashion. The size and characteristics of any
lesions (masses or cysts), along with their relationship to the main duct, spleen,
and peripancreatic soft tissue resection margins should be documented. If the
282
Chapter 17 • The Pancreas I 2 83
lesion is cystic, the cyst contents and the cysts relationship to the main duct
should be also documented.
Sections from the lesion should include samples that demonstrate the rela-
tion of the lesion to margins and uninvolved pancreas. If the pancreatic lesion
is cystic and small, it should be submitted in its entirety. For large lesions (e.g.,
>5 em), at least one section per centimeter is recommended, which should focus
on thickened, solid, or irregular areas. One or more perpendicular sections from
the inked posterior (retroperitoneal) soft tissue margin should be submitted. One
to two representative sections from the spleen are sufficient unless gross abnor-
malities are detected. Finally, the peripancreatic and splenic hilar soft tissue is
searched for lymph nodes; all identified lymph nodes should be submitted in
their entirety for microscopic examination.
C. The Whipple procedure, performed for tumors of the pancreatic head, common
bile duct, ampullary, or periampullary region, involves excision of a composite
specimen usually consisting of the pancreatic head, a portion of the common
bile duct, the duodenum, the distal stomach, and the gallbladder. Mter orien-
tation, the dimension of each organ should be measured. The various surgical
margins of the pancreas (pancreatic neck, posteriot; portal vein groove, and
uncinate) should be inked with different colors in the operating room with the
surgeon's aid; frozen section evaluation of the pancreatic neck and bile duct mar-
gins is almost always requested intraoperatively. The stomach is opened along
the greater curvature and the duodenum is opened along the aspect opposite
the pancreas to avoid the ampulla. The pancreas can be sectioned along the
plane defined by the pancreatic duct and bile duct using probes as a guide, or
cut perpendicularly. The size, location, and nature of the tumor are recorded, as
is the relation of the tumor to the margins. If the tumor is cystic, the cyst con-
tents and the relationship of the cyst to the main duct should be documented.
The specimen is then pinned out and fixed in 10% formalin before grossing. In
general, one section per centimeter of the tumor is submitted; if the tumor is an
intraductal papillary mucinous neoplasm (IPMN) or mucinous cystic neoplasm
(MCN), and is noninvasive in the initial sections, it is necessary to return to the
gross specimen and submit the entire lesion to ensure that invasive tumor is not
missed. Additional sections should demonstrate the relation of the tumor to the
various margins of excision, uninvolved pancreas, ducts, ampulla, duodenum,
and soft tissue; one section from the proximal gastric resection margin, and
one from distal duodenal resection margin should also be submitted. One sec-
tion from the uninvolved pancreas and one from uninvolved ampulla are also
submitted if not already sampled in the tumor sections. All identified lymph
nodes in the soft tissue should be submitted for microscopic examination; there
is no need to separate the nodes into groups because they are all considered
regional. Microscopic examination of a minimum of 10 to 15 nodes has been
recommended for Whipple specimens.
Ill. DIAGNOSTIC FEATURES OF PANCREATITIS
A. Acute pancreatitis is an inflammatory process in which pancreatic enzymes
autodigest the gland. It is most commonly associated with biliary tract disease
(such as gallstones) and alcohol abuse. The pancreas is swollen and edematous,
and hemorrhagic in more severe cases. Chalky white fat necrosis may be evi-
dent. Histologically, mild pancreatitis is characterized by interstitial edema and
leukocytic infiltration; the pancreatic parenchyma may be well preserved, with
only limited necrosis. In severe cases, extensive necrosis and hemorrhage are
seen (e-Fig. 17.1). * Calcification and secondary infection may occur.
B. Chronic pancreatitis is an inflammatory process characterized by irreversible

destruction of the exocrine component with acinar atrophy, mixed inflammatory
cell infiltrates, and fibrosis with relative sparing of the islets of Langerhans (e-
Fig. 17.2). Dilatation of the pancreatic ducts with proteinaceous, often calcified,
secretions is characteristic. In later stages, the endocrine component may also
be destroyed.
1. Etiology. Most cases are associated with chronic alcohol abuse. Other eti-
ologies include bile duct and pancreatic duct obstruction (due to lithiasis),
hyperlipidemia, hyperparathyroidism, autoimmune disorders, and heredi-
tary chronic pancreatitis [due to germline mutations in cationic trypsinogen
(PRSS1 ), cystic fibrosis transmembrane conductance regulator (CFTR), chy-
motrypsin C, calcium-sensing receptor, and anionic trypsin (PRSS2)] (Dig
Dis. 2010;28:324).
2. Autoimmune pancreatitis {AlP) is a distinct type of chronic pancreatitis fre-
quently associated with systemic autoimmune diseases, including inflam-
matory bowel disease and primary sclerosing cholangitis. AlP may mimic
pancreatic cancer radiographically and endoscopically when it presents as
a mass lesion in the pancreatic head and/or with strictures of the pancre-
atic and common bile ducts. Histologically, AlP shares many features of
conventional chronic pancreatitis, but is distinguished by a dense lympho-
plasmacytic infiltrate, particularly in the periductal region (e-Fig. 17.3 ), and
obliterative phlebitis (e-Fig. 17.4). In addition to various autoantibodies,
selective elevation of serum immunoglobulin G4 (lgG4) level helps estab-
lish the diagnosis (Arch Pathol Lab Med. 2005;129:1148; J Gastroenterol.
2006;41:613). The number of lgG4 plasma cells in the pancreas is typically
>10 per high-power field (HPF), and >50 per HPF is highly specific for the
diagnosis (Adv Anat Pathol. 2010;17:303). Patients often respond well to
steroid therapy.
A recent study has suggested that AlP can be further divided into type
1 and type 2 diseases (Am J Surg Pathol. 2011;35:26). Type 1 AlP is the
pancreatic manifestation of lgG4-related systemic disease, whereas type 2 is
confined to the pancreas; the serum IgG4level is higher in patients with type 1
disease than in patients with type 2 disease. Pathologically, type 1 and type 2
AlPs share periductal inflammation, however, interlobular inflammation and
fibrosis is unique to type 1, and obliterative phlebitis is much more common
in type 1 than type 2 (91% vs. 6% ). The number of lgG4-positive plasma
cells in the pancreas is much higher in type 1 disease than type 2 disease
(>50 per HPF in 88% and 7% of type 1 and 2 diseases, respectively; > 10
in 100% type 1 and 50% type 2 diseases, respectively). On the other hand,
ductularllobular abscesses and ductal ulceration are much more common in
type 2 than type 1 AlP.
IV. CYSTIC LESIONS AND TUMORS OF THE PANCREAS. Cystic lesions of the pancreas
include non-neoplastic and neoplastic types. The former mainly include pseu-
docyst, lymphoepithelial cyst, ductal retention cyst, mucinous non-neoplastic
cyst, and paraampullary duodenal wall cyst (groove pancreatitis). Neoplastic
cystic lesions include serous cystic neoplasm, MCN, IPMN, intraductal tubu-
lopapillary neoplasm (ITPN), solid pseudopapillary neoplasm, and cystic aci-
nar neoplasm. Pancreatic neuroendocrine tumors (islet cell tumors) can also be
cystic.
A. Nonneoplastic cystic lesions
1. Pseudocyst is the most common cystic lesion of the pancreas and usually
develops as a result of pancreatitis or trauma. About two-third of pseudocysts
are located in the tail. The cyst is lined by granulation or fibrous tissue without
an epithelial lining, with a wall thickness ranging from several millimeters to
several centimeters (e-Fig. 17.5).
2. Lymphoepithelial cyst is usually lined by squamous epithelium, typically ker-

atinized, with abundant underlying lymphocytes that occasionally form ger-
minal centers (e-Fig. 17.6). The lining epithelium may be of other types such
as flat, cuboidal, or transitional. Rarely sebaceous and mucinous cells may
be also seen in the wall, but skin adnexal structures are not found.
3. Ductal retention cyst (also named simple cyst) is caused by obstruction. Micro-
scopically, it is a unilocular cyst lined by a layer of simple epithelium. When
the lining epithelium becomes mucinous, it is termed a mucinous nonneo-
plastic cyst.
4. Paraampullary duodenal wall cyst (groove pancreatitis, cystic dystrophy of the
duodenal wall, paraduodenal pancreatitis) is located within the "groove"
between the head of the organ, the duodenum, and the common bile duct
(Hepatogastroenterology. 1982;29:198). It most likely arises from the sub-
mucosal pancreatic tissue associated with remnants of the minor papilla, and
is caused by obstruction of the minor papilla. It causes segmental chronic
pancreatitis affecting the groove area (hence the name groove pancreatitis).
Microscopically, it consists of dilated ductal structures within the duode-
nal wall; the lining of the ducts is often partially denuded and may contain
mucinous epithelium and reactive changes (e-Fig. 17. 7). The cyst wall is com-
posed of inflamed fibrous tissue containing lymphocytes, plasma cells, and
neutrophils, and the adjacent pancreatic tissue shows atrophy, fat necrosis,
and fibrosis.
B. Neoplastic cystic lesions
1. Serous cystadenoma is a benign cystic neoplasm usually found in the body
or tail of the pancreas in elderly patients. The vast majority is sporadic but
the lesion can be associated with the von Hippel-Lindau syndrome. It is
usually multilocular (microcystic cystadenoma), but occasionally unilocular,
oligocystic (macrocystic), or even solid. Grossly, the tumor has a spongy
appearance with a stellate central scar (e-Fig. 17.8) with cysts that are filled
with dear serous fluid. Microscopically, the individual cysts are lined by a
single layer of flat to cuboidal epithelial cells, with pale to clear glycogen-rich
cytoplasm (e-Fig. 17.9). Focally papillary structures lined with clear cells may
be seen. Immunohistochemically, the lining cells are positive for cytokeratin
and in most cases are also positive for a-inhibin and MUC6 (Am] Surg
Pathol. 2004;28:339). The solid variant can mimic metastatic clear-cell renal
cell carcinoma (e-Fig. 17.10).
Serous cystadenocarcinoma is exceedingly rare in the pancreas and is mor-
phologically indistinguishable from serous cystadenoma. The diagnosis is
established by the presence of distal metastasis, but vascular invasion and
invasion into adjacent structures have been observed.
2. MCN tends to occur in the tail or body of the pancreas, predominantly in
women about 50 years of age with a female to male ratio of 20:1. The
neoplasm is a solitary, multilocular (rarely unilocular) cystic mass filled with
mucin (e-Fig. 17.11) or mucin admixed with necrotic material. The cysts
do not communicate with the ductal system. The cyst lining, which may
be partially denuded, consists of tall columnar mucin-producing cells with
characteristic underlying ovarian-type stroma (e-Fig. 17.12). The ovarian-
type stroma surrounding the cysts is the defining feature of MCN and is
useful for distinguishing MCN from IPMN (Table 17.1). The stromal cells
are frequently positive for estrogen and progesterone receptors and inhibin,
and can be luteinized.
On the basis of the cytologic and architectural features of the lining epithe-
lium, noninvasive MCN can be divided into three categories: MCN with low-
grade dysplasia (mucinous cystadenoma) (e-Fig.17.13 ), MCN with moderate
or intermediate grade dysplasia (borderline MCN) (e-Fig. 17.14), and MCN
1'1."!'
'
JDUI DfldleUon !ktwMn lttladiiCIIII ,.,. ,-IIIICIIIolllNH!II- (liMit)
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patients with an invasive colloid carcinoma arising in association with an

IPMN have a better prognosis than those with an invasive conventional duc-
tal carcinoma arising in association with an IPMN (AnnSurg. 2001;234:313;
Ann Surg. 2004;239:400).
4. ITPN is a grossly visible solid nodular tumor obstructing the duct system (Am
I Surg Pathol. 2009;33: 1164 ). Architecturally, the tumor forms tubulopapil-
lary structures composed of cell with little cytoplasmic mucin (e-Fig. 17.21).
The cells uniformly show high-grade dysplasia, and some cases have associ-
ated invasive carcinoma. ITPN s do not harbor KRAS or BRAF mutations.
Expression of DPC4 is retained in most cases.
5. Intraductal tubular adenoma, pyloric gland type, is an uncommon tumor con-
sisting of pyloric-type glandular structures with mild to moderate atypia (Am
I Surg Pathol. 2005;29:607) (e-Fig. 17.22). Intraductal tubular carcinoma is
characterized by high-grade cytology and can have an associated invasive
carcinoma component (Anticancer Res. 2010;30:4435). Some authors con-
sider intraductal tubular carcinoma as part of the spectrum of ITPN (Am I
Surg Pathol. 2009;33:1164).
6. Cystic acinar cell cystadenoma is an extremely rare unilocular or multilocular
cystic mass lined by a single layer of cells cytologically resembling acinar cells.
In acinar cell cystadenocarcinoma, the cysts are lined by layers of cells that
exhibit more cytologic atypia with easily identifiable mitotic figures.
7. Cystic pancreatic neuroendocrine tumors are probably more common than
previously recognized. In one study, 17% of the pancreatic neuroendocrine
tumors were cystic (one-third purely cystic and two-thirds were partially
cystic)(] Am Coll Surg. 2008;206:1154).
V. SOLID TUMORS OF THE EXOCRINE PANCREAS. The current World Health Organiza-
tion (WHO) histologic classification of tumors of the exocrine pancreas is given in
Table 17.2. The 2010 American Joint Committee on Cancer (AJCC) tumor, node,
metastasis (TNM) staging schema is given in Table 17.3.
A. Pancreatic ductal adenocarcinoma (PDA) accounts for 85% to 90% of pan-
creatic neoplasms and is most frequently found in the head of the pancreas in
patients 60 to 80 years of age. Factors associated with an increased risk include
tobacco smoking, chronic pancreatitis, and gastrectomy. Several syndromes are
also associated with an increased risk for pancreatic carcinomas including Lynch
syndrome, familial atypical multiple mole melanoma syndrome (FAMMM),
Peutz-Jeghers syndrome, hereditary pancreatitis, mutations in BRCA2 and other
Fanconi anemia component genes, and familial pancreatic cancer syndrome.
Common somatic genetic alterations that have been detected include activa-
tion of the KRAS oncogene (by point mutations) in >90% of cases, Her2/neu
overexpression in 70% of cases, and inactivation of the tumor suppressor genes
TP53 in 50% to 70% and DPC4 in 55% of cases. Recent genomic sequencing
revealed an average of 63 genetic alterations in cases of pancreatic adenocarci-
noma (Science. 2008;321:1801), although most mutations are likely passenger
mutations with no role in pathogenesis.
Grossly, ductal adenocarcinoma is usually poorly demarcated, firm, and yel-
lowish gray or white due to its infiltrative growth and associated strong desmo-
plastic response. On the basis of glandular differentiation, mucin production,
mitosis, and nuclear features, PDAs can be divided into well-differentiated, mod-
erately differentiated, and poorly differentiated forms. Most tumors are well to
moderately differentiated and characterized by glandular structures haphaz-
ardly distributed in a desmoplastic stroma (e-Fig. 17.23). In well-differentiated
tumors, the neoplastic glands are well formed and usually large or medium sized;
however, the neoplastic glands may show rupture or be incomplete, features that
are not observed in normal ducts. Mucin production may be evident. The most
important histologic features that can be used to distinguish well-differentiated
, •• I SECTION llh <II lAACT
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CK20 is typically negative or only focally positive. Other markers that are
expressed in PDAs include claudin-4, cyclooxygenase (COX)-2, mesothelin,
KOC (K homology domain containing protein overexpressed in cancer), SlOOP,
and maspin. These markers may be useful in distinguishing ductal from nonduc-
tal pancreatic neoplasms and from benign pancreatic ducts, but are less useful in
distinguishing adenocarcinomas of nonpancreatic origin. Approximately 55%
of PDAs harbor mutations in DPC4, but this is not specific as loss of DPC4
expression is also observed in cholangiocarcinoma (Hum Pathol. 2002;33:877)
and colonic adenocarcinoma (Mutat Res. 1999;406:71).
PDA is associated with a dismal prognosis, and the overallS -year survival is
only 3% to 5%. Resection improves survival as shown by 10% to 20% 5-year
survival in patients treated with curative resection, but only 10% to 20% of
tumors are resectable at the time of diagnosis.
B. Variants of ductal adenocarcinoma
1. Adenosquamous carcinoma consists of neoplastic components with both duc-
tal and squamous differentiation (e-Fig. 17.27). Diagnosis requires the pres-
ence of a squamous component exceeding 30% of the neoplasm. Patients
with adenosquamous carcinoma have a poorer prognosis than those with
pure adenocarcinoma.
2. Colloid carcinoma (mucinous noncystic adenocarcinoma) is almost always
associated with intestinal-type IPMN, and rarely with a MCN. Colloid car-
cinoma is believed to have a better prognosis than conventional ductal ade-
nocarcinoma. Histologically, the tumor is similar to mucinous carcinoma
in other locations and is defined by mucin pools comprising >80% of the
tumor.
3. Hepatoid carcinoma is an extremely rare tumor in the pancreas. It can
have a pure form or be associated with ductal adenocarcinoma (Am Surg.
2004;70:1030), acinar cell carcinoma, or a neuroendocrine tumor (Cancer.
2000;88:1582; Am] Surg Pathol. 2007;31:146; Gut Liver. 2010;4:98). Mor-
phologically, hepatoid carcinoma consists of large polygonal cells with abun-
dant cytoplasm (e-Fig. 17.28). Immunohistochemically, the tumor cells are
positive for hepatocyte-specific antigen (hepar 1); CD10 and pCEA highlight
a canalicular pattern, and most cases are also positive for alpha fetoprotein.
Data on the prognosis of hepatoid carcinoma are very limited. Hepatoid
carcinoma should be distinguished from hepatocellular carcinoma arising in
ectopic liver in the pancreas (Virchows Arch. 2007;450:225) and metastatic
hepatocellular carcinoma from liver.
4. Medullary carcinoma is characterized by its distinct morphology that fea-
tures poor differentiation with limited gland formation, a syncytial growth
pattern, and a pushing border (Am J Pathol. 1998;152:1501). Some cases
are also associated with infiltration by prominent CD3+ T lymphocytes.
Medullary carcinoma arises sporadically or in association with Lynch syn-
drome. Immunohistochemically, medullary carcinoma often shows loss of
at least one DNA mismatch repair protein (Hum Pathol. 2006;37:1498).
Prognostically, patients with medullary carcinoma do better than those with
ductal adenocarcinoma.
5. Signet-ring cell carcinoma is an extremely rare variant with an extremely poor
prognosis. Metastasis from a gastrointestinal tract or breast primary should
always be excluded.
6. Undifferentiated/anaplastic carcinoma histologically exhibits little epithelial
differentiation, although some or most of the tumor cells express cytoker-
atins by immunohistochemistry. Morphologically, three variants have been
described: anaplastic giant cell carcinoma (e-Fig. 17.29), sarcomatoid carci-
noma, and carcinosarcoma (e-Fig. 17.30). The prognosis for this tumor is
extremely poor; the average survival is only 5 months.
7. Undifferentiated carcinoma with osteoclast-like giant cells, which should be

separated from undifferentiated carcinoma, is characterized by the presence
of nonneoplastic osteoclast-like giant cells (which are histiocytic in origin)
within the tumor (e-Fig. 17.31). In most cases, there is an associated in situ
or invasive adenocarcinoma or MCN. The prognosis is poor (mean survival
12 months).
C. Acinar cell carcinoma accounts for 1% to 2% of adult exocrine pancreatic neo-
plasms. Most tumors occur in late adulthood, but 6% occur in children. Approx-
imately 10% to 15% of the patients develop a lipase hypersecretion syndrome
characterized by extrapancreatic fat necrosis, polyarthralgia, and peripheral
eosinophilia.
Acinar cell carcinoma frequently presents as a well-circumscribed mass with
or without necrosis and cystic degeneration. Microscopically, acinar and solid
patterns are most common, although the tumor may grow in a trabecular or
gyriform pattern. Intraductal, papillary, or papillocystic variants of acinar cell
carcinoma have also been reported (Am J Surg Pathol. 2007;31:363; J Oncol.
2010;242016). The tumor usually lacks a desmoplastic stroma. The individual
tumor cells typically have basally located nuclei, single prominent nucleoli, and
moderate amounts of amphophilic or eosinophilic granular cytoplasm (e-Fig.
17.32). Mitotic activity and nuclear pleomorphism are variable. Most acinar
cell carcinomas are composed of cells that contain zymogen granules in their
cytoplasm, which can be demonstrated by periodic acid-Schiff (PAS) stain with
diastase or by electron microscopy. The tumor cells also exhibit abundant rough
endoplasmic reticulum by electron microscopy.
Immunohistochemically, the tumor cells are reactive with antibodies against
trypsin (>95% cases) (e-Fig. 17.33), chymotrypsin (>95% cases) (e-Fig.17.34),
and lipase (""'70% of cases). In more than one third of acinar cell carcinomas,
there are scattered tumor cells immunohistochemically positive for chromo-
granin A or synaptophysin. If >25% of the tumor cells show neuroendocrine
differentiation by immunostains, the tumor should be designated as a mixed
acinar-endocrine carcinoma.
Acinar cell carcinomas are aggressive tumors but their outcome is gener-
ally better than stage-matched ductal adenocarcinomas. Available data suggest
that the prognosis of acinar cell carcinomas in children is better than that in
adults.
D. Pancreatoblastoma is the most common pancreatic neoplasm of childhood, usu-
ally occurring in the first decade of life (mean age: 4 years), although the neo-
plasm also rarely occurs in adults. Approximately 25% of patients have associ-
ated elevated serum a-fetoprotein. Pancreatoblastoma is a highly cellular tumor
composed of sheets or islands of small monotonous cells divided by cellular stro-
mal bands. The tumor cells may exhibit acinar differentiation and form small
acinar lumina, although little endocrine and ductal differentiation may be also
seen. A histologic hallmark of pancreatoblastoma is the presence of squamoid
corpuscles, which are present in virtually every case (e-Fig. 17.35); they serve
as a useful feature to distinguish the tumor from other pancreatic neoplasms,
particularly acinar cell carcinoma. Squamoid corpuscles consist of an aggregate
of plump epithelioid cells, a whorled nest of spindle cells, or a cluster of frankly
keratinized squamous cells, and are usually located in the center of the tumor
lobules; the cells forming the squamoid corpuscle are usually larger than sur-
rounding tumor cells. The stroma in pancreatoblastoma can also be neoplastic,
and may contain heterologous elements such as bone and cartilage. The stroma
in adult patients, however, is often less abundant and less cellular than that
in pediatric patients. The prognosis of pancreatoblastoma is better in pediatric
patients than that in adults, likely due to an increased frequency of localized
and encapsulated tumors in children.
E. So.. palliiDJIIIIIIIIIJ -pilllll i a -

- - (IIIUJl "'I" 28 ,....., 90% iD -~
._.....u,....,
!aqcat the limo of dlopoolo but II typicolly well~ udllll.y b e -
RI-.1. Gto.ly, it exhibit$ nttablt toW! ud ~ UCU with h.em.o~
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in adi>l""""' girl.t ....t
be qgi"'
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plume~ of llbeetsof.....U ..t..i...ty"UIIifo"" <wD.Or..U., ll<l.lllr:lim.a .....
.......u...
cltlicate, oftm h~ lilm••ua.Jor""""' ... m.rm
_.w,p,.pillu
(..r~~~~o17.U~ The tiiii!Or <><1lt have .....,philic ot dJ:u .......t&te<lqtoplum,
bodcctcd ot Bl'Oomi auc:ld, ....t ~lllauclcol! (.,.l!ls. 17.3~ la.tra·
""'~w ~ ~rcftc.nt PAS-politi.. hrdi<>e giobulcl
may be t'l'ident. P<W117 <die, <>elll with d>.~ ..,..W., 411<1 fo:rclF bo~y
p t ..U. Dl.lf be pra<llt within dLe MILO~ In """'otic ..,..., dLe t1lma1' aillt
- du:ir ~ """_,.l!l!dnp>qlli<; dqp:umui= Mi...C.: fiaw,:a...,
""""The mot! frcqu.catly poll!I!... ,_,...uw~<cn fA..lld'1'1Q1dopopilluy ......
plum - ~ --.cr& t110luc, "1~ os-u;ticb.ymo-
lrJllU, ~ """l'(J'~ clawllll.t S ud 7, plcria 3, C)'diA D1, CI)tO
<..r.. t73Sl, an<~ crQC:!w ~'""''eniD
iti"' in I!.!.IDM <>elll but d>.r~ A is
<o-rr.,a(.,..,_
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argaliY'e. Posit!.. CI)10 -t
""'*"r 6--c•....;, n.&iDs are'llll<ful iD th.e cli!ljncriaa £<am ,...,.,..tic mcloaiDe
-plum.o (PilNo~ Ap-iwcozlr SO% of oolid. ~pi!l&ry 1100plum.o
ohow c-klt c:J:pi'COiio.ll by '-•mohlltocllcmiotey llrith011t u OMOclatcd mutt.·
Cio:o.ID the KIT- (Moll Ptlt);Ql. 2~1"11m.
Poti-wilh~l'-'pilla.ry-plum .wlybaYCan("lo:Dent~·
ooliJ if the 1:UIII.Or it compktdy em...!, u oa::om in 8S% "" 95% o f - .
Appmoimot<!y S% to 1S% oi....,. dunv m<mnuis, ......Jiy ""dLe li..r-'
paj-m, but ""'wuli• itl«hol ill ollly a lllllNt of po.oica11. IW<ly, oolid.
~pi!Wy-ploam.tlum: ...... dlfltt=d-.lotNl'<Ollll.told.""""l>-
(AM I S.,PIIIIbol. 200S;l!I-.S12). Bloloskolir 188'""'11«: tw110,. typ!ca1ly...,.
clcq> txlta~ --~ -w.r or peria.ellra1 U....uicll, tipifiaont «1·
lulu~ •udur aqp~ and u..:-.e.t mitotic 41Cti'rity.ln fiC"e<a~
~papillary """floum is~ o.l.,.....odo: molilll'""cY·
f. P.ancrNI!c lrt~a~pltlllllal _ , . . . (P.aniMI il oo....w.:...l a_......,, 10 invui:ft
~ dw:cal adm...,.,;....,,. Mozpboloaicoll.r, I'WN i.o a mic:mla>pK;
(typ:dly d mm.ID. o!tc) pa~ or llal qidleliol ""'f'lum ......t;ned 10 the
paocl'eldo do.ct .,......,. 0.. the lwll of dLe eytolosic ud ~ otnl&,
P&o!N1 ue iurthu .tlvidc.t i<rtolbrce ~one.: P&UIN-l (P&!IIN·lA, P&!IIN·
lB), P&!IIN·l, an.cl P<&!IIN·) !Am I Swz Plltbol. 1004;23>977) !Table 17.S).
Ie ·•••-.-= [j>idermold rnotap!ult, mutlloyelod m.a,_.

hill·lA: PJioflc: &lind """"lull, 110111111 coli mlllaj)ladt, ,..,..,.., hypeltrap!O)\ mucl....,.
ductalllyperplul.t, mu- c:.oll ~plalo, rnL>OCIIillrlnsfoom.atson, limpit llyperplull,
flit duot - · wtlllout aiJple. fllltdud.UII;rpotplulo, dudil hyp!IJIIaslo pol,
nonDOI'IMY _,111\elal ~p!O)\ 1101\I>Oi>laiY dueliiii'CIWP!aolo
holl·t•I'III>IIIIY ~ wiMY duclollrJI>o,.., 1>0Dia1Y dudllloolon-..tholtl
"'YJllt,ducalhypotplosM~2,adenonutlo<.l!:ducal~lldMOM
Irfl*plula
holl·t: Atypialllrfl*plul.t, poplluy duct- w111o atypia, """«r.tdo .t,.plosl.t, any PinIN
-
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Cl'l•• 11 • T~t-t PII/ICI"EEiii9. I 21a
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I Pill
Rtdlo!V"''hlc:oltf llal«t.oollo Yot
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A. Well-differentiated neuroendocrine tumors. Approximately 30% to 40% of well-

differentiated neuroendocrine tumors are nonfunctional (no clinical hormonal
syndrome although serum hormone levels can be elevated). Functional tumors
produce clinical symptoms corresponding to the hormones they produce, allow-
ing clinical diagnosis even when they are quite small. Among the functional
tumors, insulinoma is the most common type, and is benign in 90% to 95% of
the cases. In contrast, gastrinoma, glucagonoma, somatostatinoma, and VIPoma
tend to be malignant. By definition, pancreatic neuroendocrine micro adenomas
(<0.5 em in size) are nonfunctional.
Well-differentiated endocrine tumors are often well circumscribed, soft,
homogeneous, and yellow to pink lesions. Cystic spaces may be present in some
cases. Histologically, they often grow in an organoid fashion including solid,
nesting, trabecular, glandular, tubuloacinar, gyriform, and pseudoglandular pat-
terns. Hyalinized fibrovascular stroma, sometimes with amyloid deposition, is
characteristic (e-Fig. 17.44). The tumor cells are relatively uniform with finely
granular amphophilic to eosinophilic cytoplasm and a centrally located round to
oval nucleus with characteristic salt-and-pepper chromatin pattern. Other types
of cells described include clear cells (e-Fig. 17.45), lipid-rich cells (e-Fig. 17.46),
oncocytes, and rhabdoid cells (e-Fig. 17.47). Sometimes the tumor cells may be
pleomorphic. By definition, well-differentiated tumors have no more than 20
mitoses per 10 HPFs (typically no more than 10 per 10 HPFs). Necrosis may be
present but it is typically focal. Well-differentiated neuroendocrine tumors are
sometimes part of the multiple endocrine neoplasia type I (MEN I) syndrome.
The vast majority of well-differentiated neuroendocrine tumors are immuno-
histochemically positive for synaptophysin and chromogranin A. Other markers
that are expressed include protein gene product 9.5, CD56, islet-1, and PAX8
(Am I Surg Pathol. 2010;34:723).
Stage and grade are the most important prognostic factors. On the basis of
mitotic figures, well-differentiated neuroendocrine tumors are further divided
into low grade (0 to 1 mitotic figures per 10 HPFs) and intermediate grade
(2 to 20 mitotic figures per 10 HPFs) (J Clin Oncol. 2002;20:2633). Ki-67
proliferation index is also a prognostic factor; tumors with a Ki-67 proliferation
index <2% (low grade) do much better than tumors with an index of ~2% but
<20% (intermediate grade) (Clin Cancer Res. 2008;14:7798). Recently, KIT
and CK19 expression have also been shown to be of prognostic value for well-
differentiated endocrine tumors (Am I Surg Pathol. 2009;33:1562; Am I Surg
Pathol. 2006;30:1588).
B. Poorly differentiated neuroendocrine carcinomas typically show ill-defined bor-
ders. The tumor cells are often arranged in tightly packed nests or diffuse sheets.
There is often extensive necrosis. Using similar criteria as in the lung, poorly
differentiated neuroendocrine carcinomas are further divided into small cell
carcinomas and large cells neuroendocrine carcinomas. By definition, mitotic
figures numbering >20 per 10 HPFs and/or the Ki-67 proliferation index is
>20%. Given their rarity, a metastasis should be always ruled out before ren-
dering a diagnosis of primary pancreatic poorly differentiated neuroendocrine
carcmoma.
Cytopathology of the Pancreas

I. INTRODUCTION. Cytopathology has an important role in the evaluation of pancre-
atic lesions. In fact, due to a variety of factors, the cytopathologic sample will
frequently be the only diagnostic tissue sample obtained. The advent of endoscopic
ultrasound-guided fine needle aspiration (EUS-FNA) has greatly expanded the num-
ber and type of cases seen. When an abnormality is identified or suspected, EUS
makes it possible to visualize the pancreas by ultrasound, directly visualize the
lesion, and then perform an FNA.
By imaging, pancreatic lesions can be broadly categorized into two main
categories, solid and cystic. However, heterogeneous lesions with mixed solid-
cystic imaging features and other confounding factors that obscure a clear
classification also occur. The clinical approach and differential diagnostic con-
siderations vary significantly between these broad categories, and thus it is
important to ascertain the imaging characteristics of a pancreatic lesion in
order to appropriately evaluate and diagnose EUS-FNA specimens from the
pancreas.
II. SOLID LESIONS
A. Inflammatory
1. Chronic pancreatitis can present a significant clinical and pathologic chal-
lenge. Aspirate smears will usually be scant to minimally cellular, show dense
fragments of connective tissue with bland spindle cells, and fragments of
bland ductal and acinar epithelial elements which typically have only minimal
reactive cellular changes (e-Fig. 17.48). The presence of underlying chronic
pancreatitis does not alter the cytomorphologic findings required for a diag-
nosis of malignancy (Diagn Cytopathol. 2005;32:65).
B. Neoplasms
1. Adenocarcinoma accounts for the vast majority of pancreatic neoplasms. The
majority of patients have disease at clinical presentation that is not resectable
and thus EUS-FNA provides the definitive diagnosis. The cytopathologic
features of adenocarcinoma vary on the basis of the degree of differentiation;
individual cases that have a spectrum of differentiation are also encoun-
tered.
a. Poorly differentiated adenocarcinoma is less cohesive with single cells and
small groups of cells with overlap and crowding. Nuclei are round to
oval with evidence of pleomorphism, and have large nucleoli. The cells
usually have a moderate amount of granular cytoplasm with only few
intracytoplasmic vacuoles (e-Fig. 17.49).
b. Moderately differentiated adenocarcinoma shows more cohesion and flat
sheets of cells, with fewer single cells. The sheets frequently show a disor-
dered "honeycomb" pattern. The cells possess nuclei with size and shape
variation as well as prominent nucleoli. The cytoplasm is more volumi-
nous with intracytoplasmic vacuoles (e-Fig. 17.50).
c. Well-differentiated adenocarcinoma accounts for roughly 20% of aspirates
and can be diagnostically challenging. Aspirates show large cohesive frag-
ments with very few background single cells. The main diagnostic features
are nuclear enlargement, nuclear membrane irregularities, anisonucleo-
sis (at least 4x size variation), and cellular crowding and overlap (e-Fig.
17.51 and 17.52). However, these features can vary within aspirate smears
(Cancer. 2003;99:44).
d. Mucinous noncystic carcinoma shows abundant extracellular mucin with
single cells and groups of epithelial cells that have the typical cytomor-
phology of moderately differentiated adenocarcinoma, but they can be
scantily represented (e-Fig. 17.53).
e. Signet-ring adenocarcinoma presents a predominance of single cells and
small groups of cells that have a large intracytoplasmic vacuole compress-
ing the nucleus (e-Fig. 17.54).
f. Undifferentiated {anaplastic} carcinoma is cellular with numerous dispersed
single cells that are epithelioid and show marked pleomorphism, with
scattered intermixed giant tumor cells (e-Fig. 17.55).
g. Undifferentiated carcinoma with osteoclast-like giant cells shows an undif-

ferentiated pattern with intermixed benign multinucleated giant cells
(e-Fig. 17.56).
2. PEN presents as a solid mass usually in the body or tail. On aspiration, PEN
is very cellular and characteristically shows a loosely cohesive aspirate with
single cells and loose dusters of cells (e-Fig. 17.57). The individual cells
have round to oval nuclei with variable nucleoli, eccentric nuclear place-
ment imparting a plasmacytoid appearance, are often hi- and tri-nudeated,
and show neuroendocrine-type chromatin (Diagn Cytopathol. 1996;
1:37).
3. Solid-pseudopapillary neoplasm is typically seen in young women and is often
solid by imaging. On aspiration, the smears are typically cellular, although
the predominant cystic component can make the diagnosis difficult. Along
with a variable cystic background of histiocytes and granular debris, the
epithelial elements consist of large tissue fragments with a papillary pattern.
The cells are round to oval, small, uniform, and have a moderate amount
of cytoplasm (e-Fig. 17.58). Nuclei are eccentrically located and can have
grooves and pseudoindusions (Am] Clin Pathol. 2004;121:654).
4. Acinar cell carcinomas are most often solid but can have a cystic compo-
nent. On aspiration, these tumors are cellular with a variable pattern that
ranges from cohesive fragments to more dispersed single cells. The cohe-
sive fragments tend to be arranged as monolayers with a sheet-like pattern
with varying degrees of vascularity. The cells show monomorphic round to
oval nuclei, with nucleoli and an even chromatin distribution (e-Fig. 17.59).
There is typically a moderate amount of granular cytoplasm imparting an
acinar appearance, and small groups of cells can be arranged in acinar-like
configurations (Diagn Cytopathol. 2006;34:367).
Ill. CYSTIC LESIONS. EUS-FNA is only one factor in the overall assessment of a cystic
pancreatic lesion; a variety of clinical features (age, sex, size, shape, duct findings)
and concomitantly measured fluid properties (CEA, enzyme levels) also contribute
to the evaluation. The variable cellularity obtained by EUS-FNA can significantly
limit classification based on the cytomorphologic findings alone (Surg Clin N Am.
2010;90:399).
A. Pseudocysts can be present in the background of known acute or chronic pancre-
atitis, although the clinical history and imaging findings are not always definitive.
On aspiration, a variable to abundant amount of green-brown bile-tinged thick
material is obtained which consists of granular debris with variable amounts of
intermixed bile pigment, macrophages, neutrophils (usually a more acute phase
component), and scant benign ductal and acinar tissue fragments (e-Fig. 17.60).
The epithelial elements can show a reactive appearance, however, the atypia
does not approach the quantitative and qualitative features of adenocarcinoma
(e-Fig. 17.61).
B. Serous cystadenoma. By aspiration, these lesions are predominantly hypocellular.
The lining cells that are present are usually grouped as small to intermediate sized
cohesive flat sheets, and have round nuclei and a moderate amount of cytoplasm
which tends to be nondescript. The aspirate alone is often not sufficient for
diagnosis (Cancer. 2008;114:102).
C. MCN and IPMN. Cytologically, the neoplasms of this category share a variable
amount of extracellular mucin, sheets of glandular epithelial cells with vary-
ing degrees of cellularity, and a spectrum of cytologic atypia. These tumors
can have varying degrees of complexity of cell groups; papillary configurations
can be present in IPMN (e-Figs. 17.62 and 17.63). Invasion cannot be reliably
determined on the basis of aspiration smears (Cancer. 2004;102:92).
SUGGESTED READINGS
Ali SZ, Erozan YS, Hruban RH. Atlas of Pancreatic Cytopathology with Histopathologic Corre-
lations. New York: Demos Medical Publishing, LLC; 2009.
Bosman FT, Carneiro F, Hruban RH, et al., eds. Tumors of the pancreas. In: Pathology and Genetics
of Tumors of Endocrine Organs (WHO Classification of Tumors). Lyon, France: IACR Press;
2010:279-334.
Centeno BA, Pitman MB. Fine Needle Aspiration Biopsy of the Pancreas. Woburn, MA:
Butterworth-Heinemann; 1998.
Chhieng DC, Stelow EB. Pancreatic Cytopathology. New York: Springer Science+ Business Media,
LLC; 1997.
Hruban RH, Pitman MB, Klimstra DS, eds. Tumors of the pancreas In: Atlas of Tumor Pathology,
Fourth Series. Washington, DC: American Registry of Pathology; 2007.
Thompson LD, Heffess CS. Pancreas. In: Mills SE, ed. Steinberg's Diagnostic Surgical Pathology.
5th ed. Philadelphia: Lippincott Williams & Wilkins; 2009:1432-1491.
SECTION IV
Breast
Breast Pathology
Souzan Sanati, Omar Hameed, Joshua I. Warrick,
and Craig Allred
I. INTRODUCTION. This chapter provides a systematic approach to breast pathology,

which enables pathologists to effectively play their role in today's interdisciplinary
care of patients with breast disease, especially cancer. It is not intended to be a
compendium of histologic alterations occurring in the breast, which is enormous
and covered in many excellent textbooks on the subject (see Suggested Readings).
Instead, it is based on a template addressing the most common alterations, in
order of clinical importance, ranging from potentially lethal cancers to common
benign changes (e-Appendix 18.1).* Diagnoses are conveyed in a concise, stan-
dardized manner addressing specific issues important to other specialists-such as
correlating histologic with mammographic findings for radiologists, status of sur-
gical margins for surgeons, tumor, node, and metastasis (TNM) staging [essential
elements of which include: tumor size (T), nodal status (N), and distant metastasis
(M)] for oncologists, and so on. Although templates cannot be all inclusive, and
must be updated to stay current, their advantages far outweigh their limitations.
This chapter begins with a discussion of the methods used in grossing breast
specimens. A discussion of normal breast follows, because a comprehensive under-
standing of normal is necessary to appreciate what is abnormal. This is followed
by discussions of invasive carcinomas, noninvasive carcinomas, prognostic and
predictive biomarkers, common benign lesions, and reporting results.
A. General approach. The primary goals of grossing in breast pathology are to (i)
identify the specimen, determine its orientation, and dimensions; (ii) identify
the presence, location, and dimensions of lesions (masses, calcifications, etc.);
(iii) estimate the distance of lesions from surgical margins; and (iv) take small
samples for more precise microscopic evaluation. Secondary goals include tak-
ing samples from various other locations depending on the type of specimen
(e.g., nipple, all quadrants, and lymph nodes associated with mastectomies).
There are many methods of grossing breast surgical specimens; all accept-
able methods adequately address the goals listed above even if their specific
strategies vary. This section provides a basic grossing strategy to manage most
surgical breast specimens, followed by more detailed discussion of the most
common types of samples. More detailed information on grossing can be
* Online a vail ability

298
Chapter 18 • Breast Pathology I 299
found in specialized texts (Manual of Surgical Pathology. 2nd ed. Philadelphia:

Elsevier Churchill Livingston; 2006).
The central element of the basic strategy is a generic grossing template
(e-Appendix 18.2), which can accommodate specimens of almost any size,
ranging from small lumpectomies to large mastectomies. Utilization of the
template creates a permanent record (diagram) of the most important features
of a specimen (e.g., size, orientation, location of samples, location of mass
lesions, and distance of lesions from margins); the small amount of extra time
required to create the diagram has several additional benefits, including (i)
providing relatively precise information on the size and distribution of lesions
that are not apparent grossly; (ii) enabling better control of margins; (iii) assis-
tance in taking additional samples if necessary; and (iv) facilitating succinct,
comprehensive, and standardized gross dictations.
The main steps of grossing a surgical breast specimen are as follows:
1. Identification
2. Orientation
3. Dimensions
4. Inking of margins
5. Sectioning into thin slices to facilitate fixation. Small specimens are usually
cut (from 2 to 4 mm in thickness) into a few slices and entirely submitted
in a corresponding number of cassettes for formalin fixation. Larger spec-
imens are usually cut into slices (5 mm thick and hinged at the bottom to
maintain intact orientation), and allowed to fix before submitting samples
in cassettes.
6. Fixation in 10% neutral buffered formalin (NBF) for a minimum of 8 to
12 hours. The recommended maximum fixation time is 72 hours.
7. Production of a diagram of the specimen on the template. Diagrams are
more informative and useful than gross dictations alone.
B. Ancillary information. Most patients with breast pathology present with a clin-
ically or radiologically detected mass and/or a mammographically detected
abnormality, most often in the form of microcalcification. The manner of
presentation often dictates the approach to specimen processing. Because
most breast specimens lack natural anatomical landmarks, careful specimen
processing-especially margin assessment-is crucial for accurate pathologic
interpretation. In addition, evaluation of specimen radiographs represents an
integral part of examination of breast specimens, and should be reviewed when-
ever available. Review provides valuable information as to the nature of the
lesion (ill-defined vs. well-defined mass; microcalcification); location of the
lesion(s); and assists planning of the sectioning of the specimen.
C. Specimen types. Most breast specimens received for pathologic evaluation are
in one of the following forms.
1. Needle core biopsies have almost totally replaced fine needle aspiration
(FNA) biopsy specimens for the initial pathologic evaluation of localized
breast lesions in most centers. They are obtained to diagnose palpable breast
masses or nonpalpable breast lesions detected by screening mammography,
such as stellate densities or suspicious microcalcifications. For nonpalpa-
ble lesions, biopsy is usually obtained using image guidance (ultrasound
guided, stereotactic, or MRI guided). Vacuum-assisted biopsies increase the
volume of tissue obtained for microscopic examination. If biopsy is per-
formed for calcifications, a radiograph of the specimen is often obtained to
confirm that the calcifications have been adequately sampled. After describ-
ing the shape, size, and color of the tissue cores, they should be aligned in
parallel and placed in the cassette between two sponges (preferable) or
wrapped in tissue paper; similar to biopsies from other organs, overstuffing
of cassettes should be avoided. Formalin fixation time is absolutely critical,
300 I SECTION IV: BREAST
as underfixation and overfixation both may result in altered biomarker

results by immunohistochemistry. Current College of American Patholo-
gists/American Society of Clinical Oncology (CAP/ASCO) guidelines rec-
ommend that core needle biopsy samples be fixed in formalin for 6 to 48
hours prior to processing. At least three histologic levels should be obtained
from each paraffin block to ensure adequate representation of the lesion(s).
For all other breast specimens, the histologic findings should always be cor-
related with the clinical and radiologic findings, and, if discrepant (such as
when microcalcifications are not seen in original sections of a biopsy per-
formed for microcalcifications) additional deeper sections from the block,
and/or radiographic images of the paraffin block need to be examined to
resolve the discrepancy.
2. Excisianal biapsy/lumpectomy specimens are either oriented or nonoriented.
Nonoriented specimens are those in which evaluation of the status of specific
margins is not required by the surgeon (such as excisions of benign lesions or
malignant lesions with separate margin specimens); nonetheless, these spec-
imens should be inked. Oriented specimens need to be inked differentially
to facilitate specific margin orientation. A four-ink color approach is useful
to orient the margins (the superior, inferior, anterior, and posterior margins
will be inked by four different colors); the medial and lateral margins are
amputated, sliced, and completely submitted in separate cassettes. Another
acceptable approach is to use six ink colors to orient all the margins.
For these specimens, the gross dimensions and weight should be
recorded. Then, the specimen should be serially sectioned as soon as possi-
ble to allow for adequate penetration of fixative. Such sectioning is usually
performed perpendicular to the long axis of the specimen; however, sec-
tioning may be influenced by the shape and proximity of the lesion(s) to
particular margins. Gauze should be placed between the sections, and the
specimen should be fixed in formalin for 6 to 48 hours before submitting
samples in cassettes.
a. If excision is performed far mass lesions. The size of the mass (accurate
to nearest millimeter), consistency (gelatinous, rubbery, firm, or hard),
growth pattern (well circumscribed, infiltrative, pushing), and distance
from margins should all be described. The presence and size of a biopsy
cavity should also be noted. For well-circumscribed lesions thought to be
benign (such as fibroadenomas), one section per centimeter of the lesion
is usually sufficient. Ill-defined and suspicious lesions need to be entirely
submitted if possible. If the mass is too large to be completely submit-
ted, at least one section per centimeter of the mass should be submitted.
Margins are microscopically examined by submitting a perpendicular
section of the mass with each margin (superior, inferior, anterior, poste-
rior). If the margins are distant from the mass, representative-shave sec-
tions suffice. Medial and lateral margins are usually amputated, sliced,
and completely submitted in labeled cassettes (Fig. 18.1 ). Representative
sections of grossly noninvolved breast should additionally be submitted
with particular attention to fibrous areas of the breast.
b. If excision is performed far imaging-detected micracalcificatians. Most of
these specimens show no significant gross pathology. These specimens
are usually oriented (by two perpendicular sutures or dips), contain a
guiding wire, and are accompanied by a specimen radiograph. Similarly,
a surgical dip may have been placed at the area of previous core biopsy.
The guiding wire, which is placed at the site of imaging-detected abnor-
mality, should be used in conjunction with the specimen radiograph to
preferentially sample areas of abnormality, immediately adjacent areas,
as well as the margins. For smaller specimens, it is preferable to entirely
Chapter 18 • Breast Pathology I 30 1
Large Lumpectomy
Superior
B D F H J
C E G I K
Inferior
B D F
c E G K
Figure 18.1 Schematic representation of "bread-loafing,. and sampling of mass lesions. A:
Specimen was serially sectioned perpendicular to its long axis, and an irregular mass was iden-
tified in the center of the specimen. Sections A and L would be submitted as medial and lateral
margins, respectively, that are perpendicularly sectioned and entirely submitted. Given the size
of the specimen and the mass, submission of sections B through K would appear to adequately
represent the entire mass, as well as the immediately adjacent superior, inferior, anterior, and
posterior (deep) margins.
submit the specimen in consecutive sections, as this facilitates accurate

estimation of the extent of disease. In larger specimens, wide sampling
of the area of the wire tip and adjacent tissue is advised. If possible, the
specimen should be sectioned in a way that facilitates some inference of
the three-dimensional aspects of the lesion when histologic sections are
evaluated. This can be achieved by labeling the cassette numbers on a
generic diagram that can be used to determine the relationship of tissue
blocks containing abnormalities with each other (see e-Appendix 18.2).
If there are multiple lesions, their location and relationship to each other
should be documented; in addition to sampling each lesion individu-
ally, sections from normal-appearing areas in between the lesions can
be used to determine whether they are individual or multiple masses.
Representative sections from grossly noninvolved breast with attention
to fibrous areas should also be submitted (Fig. 18.2). In situations where
gross examination and initial set of sections do not show a histologic
abnormality, the entire specimen, or at least all the fibrous parts of the
specimen, should be submitted for microscopic examination.
Small Lumpectomy
Superior
CD E F G
:.. .
......... .:.... . .
.·. .. .
• • • I • .•
Medial : Lateral
Inferior
8 c D
~ E F G
Figure 18.2 Schematic representation of "bread-loafing" and sampling of a specimen excised

for microcalcifications. Especially, given the scattered widespread distribution of the m.i.crocalci-
fications and their proximity to several margins, the specimen should be entirely submitted for
histologic examination. The medial and lateral margins are amputated, perpendicularly sectioned,
and entirely submitted. The remaining tissue is serially sectioned and entirely submitted as marked
on the diagram.
3. Ma11in and biopsy raaxcisians. These cases range from unorientated flat por-
tions of tissue that should be laid flat in a cassette (shave margin), where the
presence of any malignancy seen histologically would thus be indicative of a
positive margin; to larger, variably oriented specimens that should be inked
preferentially, bread-loafed perpendicular to the new margin (oriented), and
submitted in a manner to examine the new margin. For reexcisions follow-
ing invasive carcinoma, the status of gross residual disease should be evalu-
ated as well as its relationship to different excision margins; in the absence
of gross disease, the specimen need only to be representatively sampled.
For reexcisions following a diagnosis of in situ carcinoma, the specimen
may need to be more widely sampled to detect residual disease and/or asso-
ciated invasive carcinoma. Complete reexcision of the cavity created by a
prior lumpectomy may also be performed; these specimens should be inked
with respect to orientation, and serially sectioned as in a lumpectomy. The
blood-filled biopsy cavity should be entirely submitted, in particular with
respect to new margins to evaluate the residual disease and status of surgical
margm.s.
4. Mastectomy specimens range from simple skin-sparing mastectomy (which
removes the breast only covered by the nipple-areola complex and a nar-
row rim of surrounding skin), to simple mastectomy (mastectomy without
axillary tissue), to modified radical mastectomy (mastectomy with axillary
lymph nodes). Radical mastectomy (modified radical mastectomy with pec-
toral muscles), is rarely performed in current practice. Mastectomies may be
prophylactic or therapeutic. When the specimen is received for pathologic
examination, it should first be measured and weighed. Then, the specimen
should be oriented and the surgical margins inked; it should then be serially
sectioned perpendicular to the skin at 5 mm intervals from the posterior
aspect, packed with gauze, and fixed in formalin overnight. The size of any
masses, including their growth pattern, location, and distance from all sur-
gical margins, should be recorded; the same principles discussed above (for
excisional biopsy/lumpectomy specimens also apply to grossing mastectomy
specimens. Masses should be entirely submitted if small, in a manner that
demonstrates the relationship to the surgical margin. If a mass is predomi-
nantly composed of ductal carcinoma in situ (DCIS) with focal microinva-
sion, submitting the entire mass is required to exclude a larger size of inva-
sive carcinoma. In addition, at least one section of nipple, grossly normal
tissue from each of the four quadrants of the breast, skin, and dermal scars
from previous biopsies should be submitted for microscopic examination.
If the specimen is a modified-radical mastectomy, the axillary tail should
be removed and dissected for lymph nodes, which should all be submitted
for microscopic evaluation (see below). A search for lymph nodes should
always be performed even in simple mastectomies and if lymph nodes are
found, they should be sampled accordingly. It is important to correlate the
gross and microscopic findings with imaging studies to optimize patient
care; for nonpalpable lesions, radiographing the specimen with submission
of the entire area of radiologic abnormality is recommended.
5. Mammary implants should be documented, photographed, and inspected
grossly for any evidence of leakage. One or two sections are usually suffi-
cient to evaluate the surrounding fibrous "capsule" that is usually submitted
with the implant(s).
6. Sentinel lymph nodes. The sentinel lymph node(s) is (are) the first lymph
node or group of lymph nodes that drain the breast into the axilla and in
most cases they are the first lymph node(s) involved in metastatic carci-
noma. Only rarely does cancer "skip" the sentinel node and metastasize to
a nonsentinellymph node. As such, specimens designated as sentinel lymph
nodes should be dissected carefully. Intraoperative evaluation (frozen sec-
tions and/or touch preparations) is usually limited to lymph nodes grossly
suspicious for malignancy; in these situations, intraoperative imprint cytol-
ogy of the lymph node can provide results at the time of surgery. For perma-
nent sections, each node is serially cut at 2 mm intervals and, in the absence
of gross evidence of metastatic carcinoma, entirely submitted for histologic
examination. There are many different protocols for microscopic evaluation
of sentinel lymph node, but one common approach involves examination
of three hematoxylin and eosin (H&E)-stained sections from each paraffin
block. The use of cytokeratin-stained sections-if initial H&E sections are
negative-is optimal, but not required. In the event of a positive sentinel
lymph node, axillary dissection is indicated.
7. Nonsentinel (axillary} lymph nodes. Axillary lymph node dissection may be
performed in conjunction with lumpectomy in a patient with a positive
sentinel lymph node, or may be performed as part of a modified radical
mastectomy (see above). After carefully dissecting grossly benign appear-
ing nodes from the fat, nodes <0.4 em can be submitted intact, whereas
larger nodes need to be bisected or trisected and submitted in a manner
that permits accurate enumeration (either by submitting them in individ-
ual cassettes or by differential inking). If possible, the size of the largest
grossly positive node or metastatic deposit should be measured, and the
presence of extranodal extension sampled in one section. It is recommended
that a minimum of 10 lymph nodes should be identified in an axillary
dissection (although this number is dependent on the surgical technique
and the extent of axillary lymph node dissection during surgery). Atten-
tion should also be directed to identification of possible intramammary
lymph nodes, which are usually identified in the upper outer quadrant of the
breast.
D. Examples of reporting template. To demonstrate the organization of a pathol-

ogy report on breast samples using the discussed diagnostic template (see e-
Appendix 18.1), a few examples are shown in e-Appendix 18.3.
Ill. NORMAL BREAST. An understanding of normal breast histology is essential for
accurate histologic evaluation of breast specimens. It should be noted that what
constitutes normal varies based on gende.t; age, menstrual phase, pregnancy, lac-
tation, and menopausal status.
The breast represents a modified skin adnexal structure composed of major
lactiferous ducts that originate from the nipple, progressively branching, until
eventually inducing grape-like dusters of secretory glands as lobules. Breast devel-
opment starts during the fifth week of gestation, at which time thickenings of
ectoderm appear on the ventral surface of the fetus extending from axilla to the
groin (mammary ridges or milk lines). The majority of this thickening regresses as
the fetus develops, except an area in the pectoral region. Failure of this regression
results in ectopic mammary tissue or accessory nipple. The most common location
for accessory nipple is axilla.
The adult female breast consists of a series of branching ducts, ductules, and
lobulated acinar units embedded within a fibroadipose stroma. Terminal duct
lobular units (TDLUs) are composed of lobules, which are groups of alveolar
glands, embedded in loose intralobular connective tissue that connect to a sin-
gle terminal ductule. These are the structural and functional units of the breast
and most pathologic processes arise within them (e-Figs. 18.1 and 18.2). The lin-
ing throughout the duct/lobular system of the breast is composed of two distinct
layers: an inner (luminal) epithelial layer with a cuboidal to columnar appear-
ance, and an outer (basal) myoepithelial layer (e-Fig. 18.3). The myoepithelial
cells have variable morphologies ranging from flattened, to epithelioid with dear
cytoplasm, to a myoid appearance. Identification of these two cell layers is very
important in the assessment of breast lesions as they are almost always preserved
in benign lesions, as well as in noninvasive malignant lesions, but are absent in
invasive carcinomas. Immunohistochemistry can also be used to identify myoep-
ithelial cells, as myoepithelial cells are usually positive for calponin, p63, CD10,
and smooth muscle myosin heavy chain, among other markers (e-Fig. 18.4). A
panel-based approach of two or more markers is recommended (Arch Pathol
Lab Med. 2011;135:422). In addition, a proportion of luminal epithelial cells
almost always expresses estrogen and/or progesterone receptors. The intralobu-
lar stroma is usually sharply demarcated from a denser, collagenized, paucicellular
interlobular stroma. The proportion of dense stroma to adipose tissue is variable,
with younger women having denser connective tissue (which partially explains
why mammography is less sensitive in younger individuals). Breast lobules can be
classified on the basis of their morphology into three major types. Type 1lobules
are the most primitive and rudimentary, and are usually seen in prepubertal and
nulliparous women. Type 3 lobules are the most developed, and are usually seen
in parous and premenopausal women. The progression from type 1 to type 3 is
accompanied by additional branching and increased number of alveolar buds.
Type 1lobules also predominate in postmenopausal women and premenopausal
women with breast cancer (Dev Bioi. 1989;25:643; Cancer Epidemiol Biomark
Prev. 1994;3:219; Breast]. 2001;7:278).
After birth and in the premenstrual period, breast development starts with
puberty and cyclic secretions of estrogen and progesterone. The ducts elongate
and branch primarily due to estrogen stimulus and lobulocentric growth advances
primarily under the influence of progesterone. In addition, the breast undergoes
various physiologic changes during menstruation, pregnancy, and lactation. It is
prudent to be aware of these physiologic changes since they can be mistaken for
pathologic processes by an inexperienced observer. Cyclic menstrual changes in
the breast tissue are subtle in comparison with other sites such as endometrium.
The follicular phase of the menstrual cycle is characterized by simple acini and
collagenized stroma, while the luteal phase is characterized by apical snouting
of the epithelial cells, prominent vacuolization of myoepithelial cells, and loose
edematous stroma. The epithelial cells show peak mitotic activity in the late luteal
phase. During pregnancy, there is progressive epithelial cell proliferation resulting
in an increase in both the number and the size ofTDLUs. By late pregnancy, lobular
myoepithelial cells become inconspicuous while the cytoplasm of the luminal
epithelium becomes vacuolated as secretions accumulate in the expanded lobules.
After parturition, florid changes including the frequent presence of luminal cells
with atypical nuclei protruding into the lumen (hobnail cells) can be seen, which
can be alarming to the inexperienced observer (e-Fig.18.5). Gradually and slowly,
the lobules involute to their resting appearance.
In postmenopausal women, the lobules undergo involution and atrophy char-
acterized by reduction in size and complexity with an increase in fat (type 1
lobules) (e-Figs. 18.6 to 18.8).
In contrast to women, due to lack of hormonal stimulation, TDLUs do not
develop to a significant extent in men, and male breast consists of branching ducts
within a fibroadipose stroma.
IV. INVASIVE BREAST CARCINOMA. Establishing the diagnosis of invasive breast can-
cer (IBC) is the first and most critical responsibility of pathologists (Table 18.1).
Most invasive carcinomas present as a palpable mass and/or as a mammographic
abnormality. However, in some cases the primary tumor is occult, and the patient
may present with lymph node or distant metastasis. The purpose of the pathology
report is to communicate all the diagnostic, prognostic, and predictive findings to
a multidisciplinary team of surgeons, oncologists, and other specialists; some find-
ings are strong prognostic factors (histologic type, histologic grade, lymph node
status), some determine the likelihood of response to specific treatment (hormonal
therapy, Trastuzumab), and some determine the need for additional surgical pro-
cedures (margin status). Determination of pathologic stage (Table 18.2) is vital
to determine prognosis and to guide the therapy. In addition to pathologic stage,
the prognosis of breast carcinoma is greatly dependent on additional prognostic
and predictive factors that are mandatory and should be evaluated and reported
for all breast carcinomas (15), and are most significant in lymph node-negative
breast carcinoma.
A. Prognostic and predictive factors
1. Histologic subtype. Histologic typing remains the gold standard for classi-
fication of breast carcinoma and provides useful prognostic information.
Five major types of IBC are currently recognized. Four are characterized
by relatively unique/uniform histologic features and, thus, they are referred
to as "special" histologic types. Collectively, the special types account for
about 25% of all the IBCs, and include the so-called invasive lobular, tubu-
lar, mucinous, and medullary carcinomas (approximately 15%, 5%, 2%
to 3%, and 1% to 2%, respectively) (Breast Cancer Res. 2008;10:54). The
remaining ""'75% of the breast carcinomas are histologically and prognos-
tically very heterogeneous and are referred to as invasive ductal carcinomas
(IDCs), no special type, or, not otherwise specified. Except, perhaps, inva-
sive lobular carcinoma (ILC), all the special type carcinomas have more
favorable prognosis compared with IDC, although the degree of improved
outcome is variable for different types. In addition, some pathologists rec-
ognize other less common special types of invasive carcinoma that have
a favorable prognosis. These include invasive cribriform (1 %), papillary
(<1 %), and adenoid cystic (0.1 %) carcinomas. Different experts have used
variable criteria for diagnosing special type carcinomas, which is partially
IDS I SECTION IV. BREASl
r,lhllal tu.all II.I'MIIIIbllalluiMII

m..M ducal CO/dftotr~~t, 1101 ~ eDOCIIod ,..,.l>tilellooio
m..M lobular <ttr<*\Oino Ad-~1111 ade•oole
1\Jbulu wei-
- crl-nn atdnoma
Mocllllly cordnoma
Mucfnwt cerdn00"18 and rSrtod tumcn
Nelrco•lloe'*"> 111m01'8
m..M r>aDII~IY <lll'<ho11111
__.......
Ad-ptilo!IOIM
MeiiiNnl m)O)Iplholon\a
Hemlllllorno
An&tomo!ooh
- m~-plleiY.,...I"""" Hem~
Fl---1\'0)
Apoctil'lll atdnoma l'llolldOlJIII>m-llnmllllypofpioala
,_ploo!Scca-u M)'aftbnli>-
Upld<fi::!ICW<:Inoma
SloctOCOIY <Or<*>orno lh1lo rnm~~o~y ln)0)1Ibloblu1lc 111m or
~cordftGtno u - 01\d Ol\lloii>OII'III
Adonold ejOIIe Olrell"""' Gn!iftu!.ar eel tumor
Aelnle eoll01rcmrna -bn>IM
~llch c!Mrcelatdnoma
~"'' Cllll'dnome
Hllmllllloly..- .
Lobular 1100Dioelo
Lobular cordftGtno lh du
-""""'
Anp"""""
UllOM"""""
Rho~
o.-rcomo
~ucal.,...,..,._ lesi<N IA!om)o>IM
lllwll-lllypofpiosla Ltlom)<loarama
All <lllfll\1111 aiJDII
Alnbl dudllllflMOplule AIM'IIf....lll . . . .
DCIS Flbroodenomo
Ml:toift..... ooil!lll PIIY!Iodootnnor
~ucal po~ry ,...ram, P•uti ucalllramal......,.ma, low fl'lld•
Paplloml MemiTIIry hi!NI!Dma
Alypbl Pll>loma
HroduebV..bl<)'lllc popllluyam:tloma -of-IWII
Ad0110111u N~I>IOlde•orna
11Jbulu
~.., ...,..
"""""""'
"-1M ulonomo
Sfltw>""""'•odOIIOI!III
f"''ll't'& d " -of !II& nlptlle
Ploomorpll~ odonom•
ilu<:tll od..,me
•uellfc..._
C - 1 8 . - l'rt!lololfi I 107
1'1.1 ]IJ!fll ~IWI.ttalll(TNIIO...,klllmefdreut

,,, ' ... llf$)
1X Prtnllly timor e~nnot bo· -
ro No _n,,..l pffrnery tumor
11> Cor<tlomo In o!IU
'lk(DCIS) Duetlli t~~~n::!l'll:r'M In alb.J
'lk(WS) Lobllor "'Jd>or!\lln d:u
Til (Poi,JIIIl P..,.rs d-ofthe rlppl& nol_,.,lllld wlllll-«ln du cartlnoiNI
n Tu""",;2anln~r""""<l"'"""""'
nml:: Ml::tdn- ~0.1cm In -">ssdlnlonolon
na '1\1""" >{).1 <111 t.n ;SO.S <mIn 111'00180td.....,..n
nb '1\1""" >{).5 <m t.n :sl<m In . l l - dim......,
ne '1\111\01' >l ""'bt.t 9..., I n - -d -
T2 Tu- >2 an bl.t ,;!; an In-""" d-
T3 Tu""" >5 an In,- dlmllllllon
T4 Turno< ollny....,_ dlr«t--., to c:tloltWillondlorsl<ln, mil'""
T<la
T4b
-bod !dow
·-n
Elilonoioll10 d!eot\lllil, nollnoludl'wmll' -111> mlllldo
Ellem.a (inell.ldlnJliJ&d• d!nlolill t l
IIIIIIa ol$1 nod""' ollll&!liln oflho b.....,"'
CMftnod tD tile 111!\6 -~~~~~-not mMI 1M
-~~~for lnllommololy ar<tlom1
T4<: Beth T4a and T4b
bil
T4d
,,.,..._(Jill'
lrrflemrnalorye~r<tloma
pNX Rf~Sb\al t)mJi' ntldal e~nnot bumrt'd

pNO No I1IP>niii,Ympll nodo - I l l or tlfnpll nodi mr!llllllll <0.2 rrm.
pNO.., bl fl.rth• dolstflld u
pNIXI+), ITO. (..0.2 mm. or <200 nonconfluonlc:dla on ono '*'Po
pNIXlrd+), 1\lnor
(--~""'lnr-)
...,d_
hllldoalc Olldlolll d-.1 b!f H&I «lmrnurdlllli>Cileln~
on!I' by IJQIIh molecl.lorftndlnp
pN1 ""' - I n 1.$&>1111Jyljmpll nodoa,•ndlorlnl-1 rn...,.rynod&

will mi<ros<oplc d_.W.....,IIy _ , tlfnpll nodo '"""""bl.t ncl
dlrl<llll' "'IP""nl ( n o t - dnl:dyor by non!- lmOfllna
DNlml
pNla
pN1b
pN1c
-nlqUOI)
"'"""""""' (>0.2 rnm •hd <2.0 rnml
""' - In 1.$&>1111Jy ljmpll nodoa (It loot OM
~, !et:h lnftlerrtl!ll mamtn~~~rynodM (rnkroor~··'"*l
rM->2.0 nvn)
Mrllnlllllln 1-ilolluy ljmpll n-lnd In InborN! mo.nvn.trylympll

-dlllldedlly _,_, tlfnpll nodi '""""" bl.t not clnlclltt 1ppouon!.
~ ....,.!rllod- >3 ,.-odlarylymph nod.., tile ln!Dmlt marrmrry
I\Odooare cluallod"' P~ 10 re1lact lnoreaoodlllm« bullilfl
DN2 ""'*'llleloln 4-!IOJIIIIIIY ljmpll nodoa or In <llnlcot;oi>I>OI'Oit l'llomol
mornnwy """"" •-In 11\ubalrooo rlod~ryt,ompll nodo ..........a.
DN3
pN2a
pN21>
·-of
Mrllnlllllln4-9oli1Jyljmpll • - ( I t lout.,.dr!pool >2mm)
Mrllnl11llln clnlclltt•ppnntlramll nwnmary tlfnph n-ln 1M
"""*' ulby .,.,.ph nodo m-.la

ll:lelo In ~ 10dary lymJ>IIIIOdao. «In l'lfrl>clallcuir ttrnl!llnodoo. or
In dnlcotJ OPP0111Miidl!Drelln!Dmol marnrnory lymph nodooln 11\o
r--n""rli!:1 paoltlvouJIIolytlfnpll nodos;orln >S-rylymph
- wllllcllnlcdy neplloe rnl:rallcoplc , . _ In Intimol mllnlt'IIIY
lymph nodD; «In "*'""' lllpnocll.llcutor tJ>mpll nodal
aaa I SECTION rv. BREASl
l'l.t!l jlfll l'IIIMII!We.llelutalllt(1'NIIO .._kllemehwlllreut

' - Cl!el- (t.WIIIIIMl
pN31 ._,,..,In ;,10-ry flmph- (II -one611poo~ >2 mm), «
"'-llslotllotnfr.-~r flmph nod..
pNi!b - In dnlcoli1•PpotCI1! lpsloCotoilntzlmel mommorJ flmph - I n
Ill& Dl_.... d 0!:1 DOOIIM>Olllloly IJm~ nodeoo « 1'1 >~dalY tr'rnPh
nod•...,.
nodeund In lrnernol rnamrnorytr'rnph m~ d"-"
- by-nel tr'rnph node dla&edlln bill nOidlnlodyepparent
pN3c ._llllolnlpol1111nolsu....,.._r!1mph nodos
.........,,. . . (10
MX ~nt-•comotboeOI&O!Od
MO ~diiWrt-olo
MO (I+) lltj>osb of m-.IIJ!yor rrk""""'~...,...,.,. tumor oekln - n g
bbod, bona~ or oth• non~l nod.el t:I.Due ~2 mm In a
pctlont- no dlrkol or nodloqlc: - o f dllllntmot""' '1°
Ml lllllonl-•111, dlllennlnod bycll'llcal•nd radlol:lit: m...., «
hlltdop:tly""""" >0.2 mm
Me ..........
fllq&O 'II> NO MO
S~q&IA n NO MO
SlqtiB TO Nlml MO
n Nlml MO
TO Nt• MO
n Nl' MO
T2 NO MO
fllq& 119 T2 Nl MO
l3 NO MO
Slqj:IIIIA TO N2 MO
n N2 MO
T2 N2 MO
T2 Nl MO
T2 N2 MO
SlqtiiiB T4 NO MO
T4 Nl MO
T4 N2 MO
AnyT ~ MO
AnyT M:tN loll
lldd- l'l'IIIIUIIt
('If,) Cll1l1:1~ lllllp M!ctoK~fle *-ru ,.....ta I'JJII:II "-art
ti-lo An:lllloc!urol dlrlorllon, Untonn """'"'""""'...._ ER (+). P(R (+), HER2 (-)
di!Ntylmd~"*Ol. -rllleradol
MUillb:ollibiiiiWOII ~C!IlDDllomle limn.
...,
Indlron ~11\ir. laiJIIII<>Id
..
--
, _ lubullr 5 Sptulrrlld m..., •moll""" M.l*fad ~-will Yos ER (+), P(R (+), HER2 (-)
.-,. llponol oncls, opon llmlno.
I~ miiCII'lOUO 2-$ W<lkl~<jjmorcd\od 1:>1>161bod 1\mor<lei&~"' POOh of Yeo ER l+l.I'I!R 1+1. HER2 H
.,..-,, m..., eolall"""""'-*
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, _ nw!ulllly
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1-.1! ro..,_,.,
eaJ/foc$
-
wolld1:UI!'ISCl1bed ,..,
...
Synejtill..,-, Pill~
bonllll, tlmp/IOplosn\I<!Yife
Y•Ofp\1'8) ER (-), 1'11'1 H, HER2 (-);
(lrlplo nop1!\'rl)
..
1 - m"""'"'~Iflly
.-,.
1-.1! $lnlor t1> 100, NST
ln1!1reta, !\WI NICIII>r ll'lllll
T~~mr _ . , wtilln om~
'l>'l""'· ,...... '""""'· ,..,

~bto¥M=Qer¢01'M
H..,r 1111ool
LNm-•
V.rlilblo
100, NST 7G-'7S S!>tula18d muo. Wllllblo varlilbr&. ~ otOIIOCiel Vamblo varlilbr&
Mllllploob: .,.,.-,, tjp& -~- ...
 T11110r tnonslo,_lo ~LN ER (-), 1'11'1 (-), HER2 (-);
noreond* (sq-111)
0< rr..... ocl>fmalolomonls
_.,......
m~~~C~ste . . (lrlplo ._tNo)
ER IUlrl 1'11'1 <On b8 ~
-" lnll!andu~r...,.rto
....
0
IDC, ~dl.dlltlteii'IOMII; HST, M:~l t,or.a.
110 I SECTION IV. BREASl
Elllon Ell Ia Modlb11oft of SaltHI- R~ Gla1Dq

S7diiM llof a,_..
c.c1-
sen
MI)OfiJ(>75liO)oft.mo< 1
..-...........
hlodOIOIO U0...7Sll0) oftl.mOf 2
IJUio « IlOilo (<1010 of u.mor 3
Smol. r...,llr, urlform nuclol 1

hlod111'111a - I n a.•nd '<Oflll~lt;, 2
Malbd ""!lob 3
11-aolllt,r
Flllcl dl.tm..., (mm) OA4 O.Sll 0.63
--
Flolcl""" (mm2J O.l!i2
().0
6-10
>11
0.274
().Q
10.1!1
>20
().312
0.11
12-:!2
>ZI
I
2
3
is a better tool to assess the prognosis in any given tumor (Pathol Oncol
Res. 2008;14:113).
c. Surgical margin. The distance of tumor from the margin of surgical exci-
sion is also important because it is inversely related to the likelihood
of local recurrence, especially in tumors removed by lumpectomy. Wide
margins with grossly distinct tumors can be adequately measured with a
ruler, whereas close margins require microscopic measurement. A mar-
gin is considered positive only if the ink that was applied to the margin
at the time of gross examination transects tumor cells. The distance of
the closest margin, as well as its location, should be reported. The extent
of margin involvement can be relayed as unifocal (<4 mm), multifocal
(more than one focus), or extensive (5 mm or greater).
d. Others
i. (MI) is a measure of the capability of the tumor cells to divide and
replicate and is determined by calculating the average number of
mitotic figures in a minimum of 10 consecutive high-power fields
(HPFs) in the most mitotically active part of the tumor, usually at the
periphery of the tumor. Only the clearly identifiable mitotic figures
should be counted.
ii. The presence of lymphvascular space invasion (e-Fig. 18.12) is an
important and independent prognostic factor, particularly in node-
negative patients with IBC. The presence of lymphatic channel inva-
sion recognizes a subgroup of node-negative patients who are at
increased risk of lymph node metastasis or distant metastasis (Ann
Oncol. 2005;16:1569; Diagn Pathol. 2011;13:18). There is a high
degree of interobserver variability in diagnosing lymphvascular inva-
sion (LVI); therefore, adherence to strict criteria is recommended in
order to reduce this interobserver variability. It is recommended that
the focus of LVI should be outside the borders of invasive carcinoma.
In addition, identification of an endothelial lining, and lack of con-
formation of tumor emboli to the shape of vascular space, help to
differentiate a vascular space from artifactual stromal clefting. LVI
present within lymphatic spaces in the dermis is often correlated with
the clinical features of inflammatory carcinoma (diffuse erythema and
edema involving one-third or more of the breast). In the absence of
the clinical features of inflammatory carcinoma, this finding remains
a poor prognostic factor but is insufficient to classify a cancer as
inflammatory carcinoma.
iii. Skin and nipple involvement. Skin can directly be involved by under-
lying invasive carcinoma (with or without ulceration). This usually
is seen in association with large tumors, or in tumors that are small
but very superficial. In addition, tumor cells from underlying lact-
iferous ducts of the nipple that are involved by DCIS can percolate
through the epidermis without breaking through the basement mem-
brane (Paget's disease).
Recently, gene expression profiling studies have classified breast
tumors on the basis of their gene transcription patterns into differ-
ent types or classes with different prognostic implications (Nature.
2000;406:747). These new technologies provide important informa-
tion that can be integrated into routine patient care (see below), but
histologic typing of breast tumors still remains the gold standard
for classification of breast carcinoma and provides useful prognostic
information.
B. IDC, no special type is the most common subtype of breast cancer, comprising
70% to 75% of the IBCs. This subtype is composed of a heterogeneous mixture
of tumors with different morphologies, clinical findings, and patient outcomes.

These tumors are predominantly devoid of histologic features that would allow
categorization of these tumors in a special type category, and usually present
as a spiculated mass, with or without calcifications, or as a mammographic
abnormality. Grossly, the tumors usually form a hard mass, with tan-white to
gray, gritty cut surface that is a manifestation of dense stromal desmoplasia.
The morphology of these tumors is quite heterogeneous with regard to growth
pattern, cytologic features, mitotic activity, and extent of associated in situ
carcinoma. The growth pattern can be solid, trabeculat; in cords, in tubules,
or as single cells (e-Figs. 18.13 to 18.25). Most often, heterogeneity can also
be noted within different parts of the same tumo.t As mentioned earlier, the
ESBR grading system is an important prognostic tool for categorizing these
tumors into groups with good versus poor outcome.
c. Special subtypes
1. Invasive lobular carcinoma (ILC} is the second most common type of IBC,
and comprises about 15% of all the invasive carcinomas with a report-
edly increased incidence in patients receiving postmenopausal combined
hormone replacement therapy (Breast Cancer Res. 2004;6:R149). These
tumors are associated with a higher rate of multifocality in the ipsilat-
eral breast, and are also more often bilateral than other subtypes. They
may present as a palpable mass, but more commonly they present with an
area of density or architectural distortion on mammogram. Some tumors
do not show any mammographic abnormalities. Grossly, either they may
form a mass, which is indistinguishable from a mass of IDC, or they may
show only an area of firm, rubbery breast tissue. Sometimes, carcinoma
is only revealed upon microscopic examination. Histologically, ILC has
a distinct morphology and pattern of infiltration within the stroma. The
classical form is characterized by small, uniform, noncohesive tumor cells
infiltrating in a single "Indian" file pattern and forming linear strands (e-
Figs. 18.26 to 18.28) that may concentrically surround benign epithelial
elements (targetoid pattern of growth) (e-Fig. 18.29). Typically, the tumor
cells infiltrate without provoking a stromal reaction and without destroying
normal architecture. Cytologically, the nuclei are small and uniform, and
located eccentrically within a cytoplasm that occasionally shows intracy-
toplasmic lumina (an intracytoplasmic vacuole containing an eosinophilic
mucin droplet). Mitotic activity is sparse. Variant forms of ILC differ from
the classical type with regards to architecture or cytology. While occasional
signet-ring cells can be seen in classical lobular carcinoma, in the signet-ring
cell variant the tumor shows a prominent population of signet-ring cells (e-
Fig. 18.30). In the solid and trabecular variants, tumor cells are cytologically
similar to the classical form; howevet; they have a different growth pattern
of either large sheets (e-Figs. 18.31 and 18.32) or nests of 20 cells or more,
which are separated from each other by stroma (e-Fig. 18.33), respectively.
In the pleomorphic variant, the tumor cells are larget; and show more sig-
nificant pleomorphism increased mitoses (e-Fig. 18.34 ). Most cases of ILC
show reduced or no expression of E-cadherin by immunohistochemistry. As
suggested by their higher grade, pleomorphic ILCs do not share the some-
what better outcome (compared to IDC-NST) that characterizes classic ILC.
Other differences from IDC-NST include a lower incidence of parenchymal
metastasis, and isolated tumor cell (lTC) metastatic patterns in minimally
involved axillary lymph nodes (as opposed to the usual finding of tumor
cell dusters in the subcapsular sinuses). ILCs have characteristic metastatic
spread to certain organs such as the ovary and stomach (where the metas-
tases can resemble primary signet-ring cell carcinoma). Up to 95% of the
ILCs are immunopositive for ER and PgR expression. ILCs typically do
not overexpress the HER2 gene product, except the pleomorphic variant,
which may show HER2 overexpression.
2. Tubular carcinoma. This tumor type constitutes 5% of the invasive
carcinomas, even higher on screening detected tumors (] Clin Oncol.
1999;17:1442), and is associated with limited metastatic potential and an
excellent prognosis. As a result of increased use of screening mammogra-
phy, these tumors present as a nonpalpable mammographic abnormality,
or are incidentally found on biopsies for other abnormalities. Grossly, they
form a spiculated mass. Microscopically, these tumors are characterized by
a haphazard arrangement of angulated tubules with tapering ends and open
lumens. The tubules are lined by a single layer of epithelial cells that often
display eosinophilic to amphophilic cytoplasm with apical snouts and oval
nuclei, with only mild to moderate nuclear pleomorphism (e-Figs. 18.35 and
18.36). A cellulat; often, fibroelastotic stroma is characteristic. The lack of
a myoepithelial layer is a helpful feature in distinguishing this tumor from
benign sclerosing lesions of the breast (sclerosing adenosis, radial scat; com-
plex sclerosing lesions). Estrogen and progesterone receptors are expressed
in >90% of these tumors (] Clin Oncol. 1999;17:1442), but they rarely, if
ever, overexpress HER2. Axillary lymph node metastasis is uncommon, and
if present, occurs only with larger tumors and is limited to the low-axillary
lymph nodes; axillary lymph node is not associated with the same adverse
outcome as IDC, NST tumors(] Clin Oncol. 1999;17:1442).
3. Mucinous carcinoma. Depending on the series and stringency of diagnos-
tic criteria, this variant comprises 1% to 6% of all the invasive carcino-
mas. This tumor is also associated with a favorable outcome. Grossly, the
tumor is well circumscribed and lobulated, with a soft gelatinous consis-
tency, and a glistening cut surface. Microscopically, the tumor is composed
of tumor cell clusters and trabeculae floating within lakes of extracellular
mucin (e-Fig. 18.37). Nuclear grade characteristically is low to interme-
diate. These tumors typically are immunopositive for ER (>90% of the
tumors) and PgR biomarkers, and negative for HER2 overexpression (]
Clin Oncol. 1999;17:1442). A significant proportion of mucinous carcino-
mas show neuroendocrine differentiation (Mod Pathol. 2004;17:568).
Mucinous carcinoma should be differentiated from other mucin-
producing lesions of the breast. Mucocele-like lesions are characterized by
mucin filled benign ducts and cysts that often rupture and result in extrava-
sation of mucin into surrounding stroma. Sometimes, the ducts show pro-
liferative changes, and portions of ductal epithelium may become detached
and float within the mucin pool. Howevet; the linear configuration of epithe-
lial cells and presence of myoepithelial cells favor a diagnosis of mucocele-
like lesion. Whenever the distinction between the two is not possible, espe-
cially in core needle biopsies, excision should be recommended for definitive
evaluation(] Clin Pathol. 2008;61:11).
4. Medullary carcinoma. This tumor comprises 1% to 2% of all the invasive
carcinomas. There is an association between medullary carcinomas and
carcinomas with medullary features with familial mutations in the BRCA1
DNA repair gene. Medullary carcinoma is usually diagnosed in younger
women and presents with a palpable mass. On mammogram, the tumor is
seen as a well-circumscribed mass usually without calcifications. On gross
examination, the tumor is a well-circumscribed, soft, tan-brown to gray
masses, with a bulging cut surface. Hemorrhage, necrosis, or cystic change
may be noted. Microscopically, these tumors are characterized by a syncy-
tial growth pattern in> 75% of the tumot; an intense lymphoplasmacytic
infiltrate, pushing borders, highly pleomorphic nuclei (nuclear grade 2 to
3), and a lack of glandular differentiation (e-Fig. 18.38). Tumors that show
most but not all of the above criteria are recognized as variant medullary car-
cinomas. Medullary carcinomas are generally are negative for all biomark-
ers (triple-negative tumors). Pure medullary carcinomas have a favorable
outcome, however, variant types do not share this prognostic advantage.
5. Other rare, but clinically significant subtypes
a. Invasive papillary (predominantly micropapillary) carcinoma. In a "pure"
form (> 75% of the tumor), invasive micropapillary carcinoma accounts
for < 1% of the invasive carcinoma; however, it is much more frequently
a minor component of IDC, NST tumors. Histologically, this variant is
composed of small solid clusters of malignant cells floating within clear
stromal spaces (e-Fig. 18.39). These cell clusters lack true fibrovascu-
lar cores and show reverse polarity (the apical surfaces of the cells are
polarized to the outside) (e-Fig. 18.40). The importance of recogniz-
ing this variant of invasive carcinoma (as well as mixed tumors with a
micropapillary component) is the associated high incidence of axillary
lymph node metastasis. It should be noted, however, that when matched
for stage, this variant does not necessarily have a worse prognosis than
IDC-NST (Mod Pathol. 1993;6:660; Am ] Surg Pathol. 2009;33:202;
Breast. 2010;19:231).
b. Metaplastic carcinomas are rare and comprise< 1% of the breast carcino-
mas. They include a heterogeneous group of tumors in which a portion of
the malignant cells have undergone transformation into a different cell
type: nonglandular epithelial (squamous), or mesenchymal cell (chon-
droid, osseous, muscle, spindle cell) types. These tumors are similar to
IDCs of no special type with regard to clinical presentation except that
they are usually larger at the time of presentation.
Tumors with squamous differentiation show a spectrum of differen-
tiation from well to poorly differentiated. Low-grade adenosquamous
carcinoma, frequently associated with papillary and sclerosing lesions,
is characterized by angulated glands embedded in a cellular stroma com-
posed of cells with low-grade cytologic atypia and focal squamous differ-
entiation (e-Fig.18.41). In contrast to higher-grade tumors that resemble
adenosquamous carcinoma elsewhere, low-grade adenosquamous carci-
nomas rarely metastasize. Metaplastic spindle cell carcinoma includes
carcinomas with abundant spindle cell transformation (e-Fig. 18.42).
The diagnosis is relatively straightforward when glandular elements are
evident, but when absent, diagnosis may only be made by demonstrat-
ing cytokeratin immunoreactivity in the spindle cells. It should be noted
that spindle cell carcinoma can be histologically bland and show min-
imal cytologic atypia (such as in the so-called fibromatosis-like spindle
cell carcinoma). The key to correct diagnosis is wide sampling of the
tumor to identify areas with clear glandular elements, and/or the use of
a panel of immunohistochemical markers, particularly high-molecular
weight/basal cytokeratins, myoepithelial markers, and p63 to demon-
strate the epithelial nature of the tumor (e-Fig. 18.43) (Histopathology.
2008;52:45). Metaplastic carcinomas with heterologous differentiation
are most commonly composed of foci of chondroid or osseous differ-
entiation; however, other types of mesenchymal differentiation such as
fibrosarcoma, rhabdomyosarcoma, and other sarcomas can also be seen
(e-Figs. 18.44 to 18.46). Despite the lower relative rate of lymph node
metastasis (considering their larger size at the time of diagnosis), meta-
plastic nonetheless carcinomas have a high rate of LVI and a poor prog-
nosis overall. ER, PgR, and HER2 are usually negative, but can be focally
expressed in the glandular component.
c. Inflammatory carcinoma is a clinical-pathologic entity characterized by
diffuse erythema, induration, and tenderness of breast skin associated
with non pitting edema (peu d'orange) involving one-third or more of the
skin of breast. A breast mass may or may not be evident. Histologically,
there is extensive dermal lymphatic invasion by tumor emboli, which
is believed to be the underlying mechanism for the clinical picture. A
tissue diagnosis is necessary to demonstrate invasive carcinoma in the
underlying breast tissue. The presence of dermal lymphatic tumor emboli
without the skin changes does not qualify as inflammatory carcinoma.
These tumors are classified as T4d by the TNM staging system.
d. Microinvasive carcinoma is defined as extension of cancer cells beyond the
basement membrane of terminal duct-lobular units to an extent :::;1 mm.
Microinvasion is usually seen in association with high-grade DCIS. In this
setting, the presence of a dense periductal stromal reaction and periductal
lymphoplasmacytic infiltration should prompt the pathologist to search
for foci of microinvasion that occurs as single cells or small clusters infil-
trating outside the confines of TDLUs (e-Figs. 18.47 and 18.48). The
focus of invasion should be obvious and care should be taken to avoid
interpretation of DCIS involving the lobules (a so-called cancerization
of lobules), or DCIS involving sclerosing lesions, as microinvasion. In
this setting, immunostains for myoepithelial markers and examination
of deeper levels of the block can be extremely helpful. Of note, if there
are multiple foci of microinvasion, the number of foci and the size of
the largest focus should be noted; separate individual foci of microinva-
sion should not be added together (http://www.cap.org/appsldocslcom
mittees/cancer/cancer_protocols/2009/InvasiveBreast-09protocol.pdf).
e. Other rare types of carcinoma include, among others, adenoid cystic carci-
noma, invasive cribriform carcinoma, apocrine carcinoma, myoepithe-
lial carcinoma, sebaceous carcinoma, lipid-rich carcinoma, clear celV
glycogen-rich carcinoma, acinic cell carcinoma, and oncocytic carci-
noma.
V. PATHOLOGY OF TREATED INVASIVE BREAST CANCER. Neoadjuvant chemotherapy
and radiation are increasingly being used in the management of breast cancer,
and so it is important to be familiar with radiation-induced changes in normal
breast to avoid interpreting them as malignant or premalignant disease. Likewise,
familiarity with chemotherapy/radiation-induced changes in tumors is required
to evaluate response to treatment.
Posttreatment changes in normal breast tissue usually include lobular atro-
phy, hyalinization, and nuclear atypia with degenerative changes, both nuclear
and cytoplasmic. Response to treatment is often categorized clinically as com-
plete, partial, or no response, and is a strong prognostic factor for disease-free
and overall survival. During gross examination, careful attention to identifying
and evaluating the tumor bed is necessary. In addition, exhaustive sampling of
the tumor bed region is necessary before making a diagnosis of complete treat-
ment response. Treatment may affect the morphology of the primary tumor and
lymph nodes involved by metastatic disease producing various histologic changes
in tumor morphology including necrosis, chronic inflammation, foamy histiocytic
collections, decreased cellularity, cytoplasmic eosinophilia, nuclear alterations,
fibrosis, and hemosiderin deposition (e-Figs. 18.49 to 18.51). Invasive carcino-
mas with a minor response may show little change in size; with greater degrees of
response, the carcinoma shows decreased cellularity or may show multiple foci
of invasion scattered over a larger tumor bed. When evaluating treated carcino-
mas, the size of the tumor is determined by the largest focus of contiguous tumor
and/or the number of foci involved over the tumor bed, not by the area of tumor
bed. Most carcinomas are of the same grade after treatment, but in some cases a
change in the grade of tumor occurs. Changes in biomarker status rarely occur
after treatment. Many systems have been developed for grading of pathological
response to treatment (Breast. 2003;12:320;] Clin Oncol. 2007;25:4414).
Lymph nodes with treatment response usually show fibrous scarring and
foamy histiocytic aggregates in the place of previous tumor (complete response),
or may show small tumor deposits within an area of fibrosis. A lymph node with
complete response should not be reported as "positive" for carcinoma.
VI. NONINVASIVE CARCINOMA. Noninvasive carcinomas have historically been divided
into two major categories: ductal and lobula:r; on the basis of the misconception
that they arise from ducts versus lobules, respectively. However, now it is recog-
nized that both of these lesions arise from TDLUs.
A. Ductal carcinoma in situ (DCIS) encompasses a heterogeneous group of lesions
with highly variable clinical presentations, histologic findings, biomarker pro-
files, genetic abnormalities, and clinical potential. They have in common a mon-
oclonal proliferation of neoplastic epithelial cells confined to the ductal-lobular
system, typically in a segmental distribution, without extension through the
basement membrane. DCIS is a nonobligate precursor of invasive cancer. It
expands and unfolds TDLUs, and may grow into larger ducts or into adja-
cent lobules, giving an appearance referred to as cancerization of lobules, or
may even grow further and expand into spheres, a source of the misnomer
"ductal" carcinoma. The most common clinical presentation ofDCIS is as cal-
cifications seen on mammogram (e-Figs.18.52 and 18.53) although up to 30%
of the detected DCIS is not associated with calcifications; in this latter scenario,
DCIS is seen as densities and architectural distortions on mammogram. Less
commonly, DCIS may present as a mass.
Grossly, most cases of DCIS are not visible. If palpable, they may form a
mass with cords of tissue extruding a paste-like material from their cut sur-
faces (a gross correlation to the material seen on histologic sections as comedo
necrosis, see below). Morphologically, DCIS is seen as proliferation of neoplas-
tic cells with variable nuclear cytology, and in a variety of growth patterns,
which usually grow over and obliterate the luminal space of the ducts and
TDLUs. High-grade DCIS may incite a desmoplastic stromal response mixed
with chronic inflammation which raises the concern for IBC (e-Fig. 18.54).
Myoepithelial markers may be useful in such cases as they highlight the myoep-
itheliallaye:r; thus providing evidence that proliferation has not yet breached
the ductal-lobular system. DCIS may become displaced during previous nee-
dle biopsy, leading to carcinoma within a biopsy tract that may be confused
with IBC in the subsequent excisional specimen. DOS may show a variety
of morphologies including apocrine (e-Figs. 18.55 and 18.56) and clear cell
(e-Fig. 18.57) differentiation. While familiarity with these morphologic pat-
terns is helpful in recognizing the lesion, most of these morphologic variants
do not bear clinical significance independent of their nuclear grade.
For clinical purposes, a few prognostic and/or diagnostic characteristics of
in situ carcinomas should be reported in a pathology report. This information
rna y affect the clinical decision making.
1. Size/extent. If the lesion is only present on one slide, microscopic mea-
surement of the focus of DOS will be the most accurate measurement. If
the lesion is present on multiple slides, the most accurate measurement is
achieved by correlating the microscopic sections containing DCIS with the
gross diagram of the breast specimen showing location and relationship of
different sections within the specimen (see e-Appendix 18.2). By correlat-
ing the two, an estimate size of the lesion can be provided. In rare cases
where an accurate size estimate cannot be given, extent of the lesion can be
reported as the fraction of slides involved by DOS.
2. Nuclear grade. On the basis of nuclear features, DCIS is classified into three
grades by SBR criteria: low, intermediate, or high grade (Table 18.5). Low-
grade DCIS consists of a proliferation of small, monomorphic, and evenly
spaced luminal epithelial cells with homogeneous chromatin distribution
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myoepithelial layer (e-Figs. 18.73 to 18.75), which can be demonstrated

by a lack of staining with myoepithelial markers such as p63 and
calponin. Some markers such as smooth muscle actin (SMA) not only
highlight the myoepithelial cells, but also stain the vascular walls within
the fibrovascular cores of papillary DCIS, which should not be overin-
terpreted as evidence of a myoepithelial layer.
f. lntracystic papillary carcinoma (IPC) is considered by most experts as a
variant of DCIS that consists of proliferation of monomorphic neoplastic
cells within an expanded enlarged cystic space with an outer fibrotic wall
(e-Figs. 18.76 and 18.77). The intracystic tumor usually shows scattered
fibrovascular cores lined by atypical epithelial cells that are not supported
by a myoepithelial layer (e-Figs. 18.78 to 18.80). Various architectural
growth patterns of DCIS, including micropapillary, cribriform, and solid,
may be present within the lesion. Care must be taken to avoid overinter-
pretation of malignant cells in the fibrous wall as a result of distortion
of the wall secondary to fibrosis, or at the site of previous core biopsy,
as the criteria for invasive carcinoma require the cells to be outside of
the fibrous wall (e-Fig. 18.81). The presence of adjacent hemorrhage or
hemosiderin-laden macrophages can be a helpful clue to the presence of
displaced tumor cells secondary to previous biopsy.
4. Extent of comedo necrosis. Comedo necrosis can be seen in any grade DCIS
but is more commonly is seen in high-grade lesions, and is characterized
by calcified necrosis within the center of a duct which is involved by DCIS
(e-Fig. 18.54). This can be conveyed as an estimate of the percentage of
surface area of the ducts involved by DCIS, which contain comedonecrosis.
Some studies have shown that abundant comedonecrosis is associated with
increased local recurrence following lumpectomy for DCIS.
5. Skin involvement (Paget's disease}. Paget's disease of the nipple can be seen
when neoplastic cells percolate through the underlying lactiferous ducts
to the epithelium without breaching the basement membrane. It is usually
associated with underlying high-grade DCIS (e-Fig. 18.82). It is important
to distinguish the Paget cells within nipple squamous epithelium from other
clear cells that occasionally can be seen within the nipple, mainly Toker cells.
Toker cells are incidentally found clear cells within the nipple, which are
smaller in size compared to Paget cells and do not show significant atypia.
A HER2 immunohistochemical stain can be useful in difficult cases, as the
cells of Paget's disease are positive for HER2 while Toker cells are negative
(Histopathology. 2009;24:367).
6. Surgical margin status is of particular importance and has an inverse rela-
tionship with the incidence of local recurrence. Since DCIS usually is not
grossly visible, margins should be microscopically examined, and the loca-
tion of the closest margin and its distance from DCIS should be mentioned
in the report. As with invasive carcinoma, a margin is considered positive
only if carcinoma is present at the inked margin. The extent of involved
margin (focal, multifocal, extensive) should be reported. If in situ carci-
noma is mixed with invasive carcinoma, the volume of in situ carcinoma in
the tumor (reported as the fraction of total tumor) should be mentioned,
as extensive DCIS mixed with invasive carcinoma has a higher rate of local
recurrence, particularly when DCIS is close to or present at the margins.
7. Presence of associated microcalcifications should be noted, as DCIS that is
first detected by microcalcifications will frequently recur as calcifications.
8. Biomarker status. Assessment of ERIPR status is an essential factor in the
evaluation of DCIS for adjuvant hormonal therapy. Although ER/PR status
can provide prognostic information, their major clinical value is to assess
the likelihood of response to hormonal therapy (see below).
B. Lobular neoplasia. This term refers to the entire spectrum of atypical epithelial
proliferations arising in TDLUs composed of loosely cohesive uniform cells
with small nuclei and indistinct nucleoli, which expand TDLUs and grow in a
Pagetoid fashion underneath the epithelial layer. Traditionally, LCIS and atypi-
cal lobular hyperplasia only differ in the degree of involvement and expansion
of the lobules; however, most authorities now consider such lobular prolif-
erations under the rubric of "lobular neoplasia" or "lobular intraepithelial
neoplasia." Many studies show that there is a direct relationship between the
extent of the disease and risk of developing IBC.
1. Lobular carcinoma in situ (LCIS} is characterized by a neoplastic proliferation
of small, loosely cohesive, uniform cells, with homogeneous chromatin and
inconspicuous nucleoli. The cells may contain intracytoplasmic vacuoles
containing eosinophilic mucin globules (e-Figs. 18.83 and 18.84). The pro-
liferating cells fill (and often distend) most of the lobules (>50%) in the
involved TDLUs. Although Pagetoid extension of neoplastic cells into the
major ductal system is usually seen with LCIS, it can also be seen in associa-
tion with DCIS; such extension undermines the normal epithelial layer and
produces a clover leaf-like appearance (e-Figs. 18.85 and 18.86). Differen-
tiating LCIS from intermediate and high-grade DCIS is usually straightfor-
ward. However, distinguishing LCIS from low-grade DCIS--particularly
with a solid growth pattern-can be challenging. In contrast to solid LG-
DCIS, the cells of LCIS are usually small and discohesive, and often display
intracytoplasmic lumina. The loss of membranous expression of E-cadherin
protein by immunohistochemistry has been recognized to be characteristic
of lobular carcinoma (cytoplasmic expression may still be evident) (e-Fig.
18.87), and retention of expression is characteristic of ductal carcinoma.
However, about 10% to 15% of the cases of lobular carcinomas retain E-
cadherin expression and a minority of ductal carcinomas lose E-cadherin
expression; therefore, the two should be distinguished mainly on histologic
grounds (Am J Surg Pathol. 2010;34:1472). Most cases of classic LCIS are
managed by steroid hormone receptor antagonists, and most data in the lit-
erature do not support reexcision for LCIS present at the surgical margins.
2. Pleomorphic LCIS is a morphologically distinct type of LCIS which shares
with classic LCIS distension of the TDLUs by discohesive malignant cells;
however, as the name indicates, it is composed of poorly differentiated cells
with significant pleomorphism that show two- to threefold nuclear size
variation, nuclear membrane irregularity, and prominent nucleoli. Central
comedo necrosis and calcifications may be observed (e-Fig. 18.88). The
main differential diagnosis for pleomorphic LCIS is intermediate- or high-
grade DCIS, a differential confounded by the not infrequent presence of
central comedo-like necrosis in all these lesions. The discohesive nature of
neoplastic cells can be very useful in confirming the diagnosis. Similar to
classic LCIS, pleomorphic LCIS can also spread along the duct system in
a Pagetoid fashion. Given the clinical, radiologic, and pathologic similar-
ities of pleomorphic LCIS to DCIS, including the frequent extension into
larger ducts, it is managed in a manner similar to DCIS; nevertheless, it is
still important to correctly identify pleomorphic LCIS due to the occasional
coexistence of ILC that can be focal and quite subtle (e-Fig. 18.28), espe-
cially on needle biopsy. In such difficult cases, the use of E-cadherin can be
potentially useful (Future Oncol. 2009;5:233). Intraepithelial macrophages
can mimic lobular neoplasia; however, microscopic examination at higher
magnification can resolve these issues (e-Fig. 18.89).
VII. THE ROLE OF BIOMARKERS IN BREAST CANCER. The role of biomarkers in estab-
lishing the prognosis and the management of patients with breast cancer can-
not be overemphasized. Established biomarkers such as estrogen (ER) and
progesterone (PgR) receptors have been used both as prognostic factors and
as predictors of response to endocrine therapy in patients with breast cancer.
More recently, human epidermal growth factor receptor 2 (HER2) was added
to this list as a prognostic factor (marker of poor outcome), and as a predic-
tor of response to certain chemotherapeutic regimens including trastuzumab (a
monoclonal antibody against the HER2 receptor). Currently, evaluation of these
biomarkers in patients with invasive carcinoma is considered standard of care.
Additional biomarkers have changed breast cancer treatment in the past decade
and have the potential of enabling individualized therapies to different molecular
subgroups. The shift toward an earlier diagnosis of breast cancer due to improved
imaging methods and screening programs highlights the need for biomarker dis-
covery to quantify the residual risk of patients to indicate the potential value of
novel treatment strategies. With the introduction of high-throughput technolo-
gies, numerous multigene signatures have been identified that have the potential
to outperform traditional markers (Endocr Relat Cancer. 2010;17:R245).
A. Estrogen and progesterone receptors. The estrogen receptor belongs to a fam-
ily of nuclear hormone receptors that function as transcription factors when
they are bound to their respective ligands. Estrogen and progesterone recep-
tors are parts of complex signaling pathways, which interact with multiple
survival and proliferation pathways in the cell and play a critical role in the
development and progression of breast cancer. They have proven usefulness as
prognostic factors, and more importantly as predictive factors, in the clinical
management of breast cancer. There is growing evidence that patients with
endocrine-responsive breast cancers benefit less from adjuvant chemotherapy.
Assessment of ER and PgR status is an important task in the evaluation of
breast cancer and is mandated in every primary carcinoma, as well as metastatic
tumors if the result could influence decision making. Approximately 70% to
80% of the breast carcinomas are ER/PgR positive. Immunohistochemistry
is the first-line attempt in evaluating hormone-receptor status, although it
can be highly affected by a variety of preanalytic factors, including time of
tissue fixation and the antigen retrieval method(] Clin Oneal. 2010;28:2784).
The expression of PgR is strongly dependent on the presence of ER and is
reflective of a functioning ER pathway. Tumors expressing PgR but not the ER
are uncommon and represent < 1% of all the breast cancer cases in some large
series; therefore, retesting of the ER status in this setting is recommended to
eliminate false ER negativity. In rare cases of solely PgR-positive tumors, the
patients still benefit from endocrine therapy. Recently, quantitative methods
for RNA-based assays (21-gene assay) have been established for ER and PgR
quantification. These quantitative assays have potential advantages compared
to rnc methods and may become the assays of choice in the future.
Several studies have shown excellent concordance rate between evaluation
of biomarkers on needle core tissue and excisional specimen (Acta Oneal.
2008;47:38; Ann Oncol. 2009;20:1948; Clin Breast Cancer. 2010;10:154;
Cancer Sci. 2010;101:2074). Because of better fixation of the needle core tis-
sue, and the potential for guiding subsequent therapy, immunohistochemical
assessment of hormone receptors and HER2 status is best performed on needle
biopsy material if available. In cases of negative hormone receptors on core
biopsy material, reevaluation of the excisional specimen may be warranted to
exclude a negative result due to tumor heterogeneity. To minimize the effect of
preanalytic variables on test results, the ASCO/CAP recommended using only
10% NBF as a fixative and controlling the formalin fixation time between 6
and 48 hours.
The ASCO/CAP has recommended an algorithmic approach to assess hor-
monal status. To standardize immunohistochemical evaluation of breast car-
cinoma, several scoring systems incorporating both intensity and percentage
Chapter 18 • Breast Pathology I 321
of staining have been established. In general, to obtain benefit from hormonal

treatment, a sample should demonstrate nuclear staining in at least 1% of the
tumor cells (e-Fig. 18.90) (/ Clin Oncol. 2010;28:2784).
B. Human epidermal growth factor receptor 2 {HER2}. ERBB2 protein is a receptor
thyrosine kinase, which is a member of the epidermal growth factor receptor
(EGFR) family of thyrosine kinase proteins. It is a membranous protein that
is expressed in all epithelial cells at low levels. The HER2/neu oncogene is
involved in the regulation of cell proliferation, survival, motility, and invasion,
and is overexpressed in about 20% of the breast cancers. ERBB2 amplification
is an independent prognostic marker of poor outcome in the absence of adju-
vant treatment and has been associated with an increased rate of metastasis,
decreased time to recurrence, and decreased overall survival. As a predictive
marker, it has been associated with responsiveness to anthracycline-based ther-
apy and trastuzumab. Immunohistochemistry and in situ hybridization are the
most common methods to evaluate HER2 status; they determine protein over-
expression and gene amplification, respectively. Immunohistochemistry is a
simple, rapid, and inexpensive method to assess HER2 protein overexpres-
sion on formalin-fixed tissues, and the ASCO/CAP has recommended an algo-
rithmic approach whereby cases are initially tested by immunohistochemistry.
Positive for overexpression (3+) is characterized by a uniform, intense mem-
branous staining of >30% of the invasive tumor cells (e-Fig.18.91). A negative
result is an lliC staining of 0, defined by lack of any membranous staining, or
1+, weak partial membranous staining (e-Fig. 18.92). Equivocal results (2+)
are defined as complete membranous staining that is either nonuniform or weak
in intensity but with obvious circumferential distribution in at least 10% of the
invasive tumor cells (e-Fig. 18.93), or intense, complete membranous staining
of 30% or fewer of tumor cells. As with hormone receptors, tissue handling,
fixation, and processing can greatly affect immunoreactivity of tissue samples.
However, well-calibrated immunohistochemistry can identify the majority of
cases as positive or negative. According to this algorithmic approach, inde-
terminate IHC results (2+) are evaluated by fluorescence in situ hybridization
(FISH) to determine gene amplification status (e-Fig. 18.94) (Arch Pathol Lab
Med. 2007;131:18; Arch Pathol Lab Med. 2009;133:775) (since the initial
guidelines were published, chromogenic in situ hybridization (CISH) has also
been approved by the Food and Drug Administration (FDA) for the same
purpose). There should be a high concordance rate (>95%) between ERBB2
protein overexpression by immunohistochemistry and gene amplification by
FISHICISH.
C. Other immunohistochemical prognostic markers. Among other markers, prolifer-
ation markers such as Ki67/MIB1 have been used as markers of poor outcome.
K.i-67 is reported as the percentage of tumor cell nuclei that are positive, how-
ever, the lack of standardized methodology and specific cutoff values limits its
value. TP53 tumor suppressor gene mutation has also been associated with a
worse outcome. Neither test has been recommended as a prognostic marker
for routine use.
D. Multiparameter-based markers. Recent expression profiling studies of breast
cancer have indicated the existence of at least four molecularly distinct types
of breast cancer, which may originate from different cell types (Nature.
2000;406:747). These subtypes differ in regards to their patterns of gene
expression, clinical features, response to treatment, and outcome, and are
termed luminal A, luminal B, HER2, and basal-like, respectively.
Luminal A and B cancers (accounting for approximately 70% of the breast
cancers) are characterized by expression of ER. They also express cytokeratin
8 and 18, typical of the mammary gland. Luminal A tumors are mostly his-
tologically low grade, while luminal B tumors tend to be of higher grade; in
general, compared with other subtypes, they have a better prognosis. Some of
the luminal B cancers may overexpress HER2. Both luminal A and B cancers
tend to respond to hormonal therapy, but luminal B cancers show a better
response to chemotherapeutic agents than luminal A.
HER2-associated cancers (accounting for 15% of the breast cancers) show
high expression of HER2 and low expression of ER and ER-regulated genes.
They are usually ER and PgR-negative and are more likely to be high grade
and involve axillary lymph nodes.
Basal-like cancers show high expression of basal epithelial genes (basal
cytokeratins such as CK 5/6 and CK 17) and low expression of ER, PgR, and
HER2 genes, the reason for calling these tumors triple-negative carcinomas.
This subtype is especially common in Mrican-American women, has a poor
prognosis, and is the most common phenotype in BRCA1-associated breast
cancers. These tumors are not amenable to treatment by endocrine therapy or
trastuzumab (Nature. 2000;406:747).
These results are significant in the sense that they show, although breast
cancer shows significant heterogeneity, from a biologic point of view many
tumors can be classified into particular groups on the basis of genetic similar-
ities, with gene signatures that correlate with clinical outcome and response
to chemotherapy. On the basis of genomic profiling data, several genomic
tests have been developed with the intent of providing even stronger prog-
nostic information. A 70-gene signature has been developed to predict the
risk of recurrence within 5 years in node-negative, ER-positive, or negative
patients; this test is able to accurately classify tumors into good or poor prog-
nostic categories (Nature. 2002;415:530; N Engl] Med. 2002;347:1999), is
performed on fresh frozen tissue, and has FDA approval for clinical use as
a prognostic test. Similarly, a 21-gene signature assay has been designed to
predict the risk of distant recurrence in patients withER-positive early breast
cancer (stage I and ll) who are receiving tamoxifen; it also predicts the benefit
from chemotherapy treatment in node-negative, ER-positive patients. This test
is done on formalin-fixed paraffin-embedded tissue and is based on real-time
polymerase chain reaction (PCR) measurement of the expression of 16 genes
with known significance in breast cancer; the results are used to calculate a
recurrence score (RS) that is predictive of overall survival independent of age
and tumor size, which classifies patients into groups with a low (< 10% ), inter-
mediate (10% to 30%), or high (>30%) risk of 10-year distant recurrence.
Another 76-gene assay has been developed for use on node-negative breast
cancers (PNAS. 2003;100:10393; Expert Rev Mol Diagn. 2004;4:169; N Eng
] Med. 2009;360:790) the 76 genes used in the assay do not overlap with the
genes used in 70-gene or 21-gene assays.
The above assays can only be performed by specific companies that have
patented the test, and quality assurance must be maintained within the com-
pany. Nonetheless, in current practice, histologic typing of breast tumors still
remains the gold standard for classification of breast carcinoma.
VIII. INTRADUCTAL PROLIFERATIVE LESIONS. Intraductal proliferative lesions that carry
an increased risk include atypical ductal hyperplasia (ADH) and atypical lobule
hyperplasia.
A.
1. Atypical ductal hyperplasia (ADH) is a term used to describe an intraductal
proliferation with some of the cytologic and/or architectural features of
low-grade DCIS, which qualitatively or quantitatively falls short of diag-
nosis of DCIS. Examples of ADH include a duct partially involved by a
uniform cell population resembling LG-DCIS (e-Figs. 18.95 and 18.96), or
a duct that is focally expanded by a uniform cell population with geometric
spaces (e-Figs. 18.97 and 18.98). Quantitatively, there are a variety of arbi-
trary criteria for distinction of ADH from LG-DCIS (<2 mm or <2 duct
spaces) (Am] Surg Pathol. 1992;16:1133; Hum Pathol. 1992;23:1095).
While there is agreement on the importance of the extent of the lesion for
this distinction, there is no widely accepted size cutoff (e-Fig. 18.99). While
a diagnosis of DCIS confers a 10-fold risk of later developing IBC, and
ADH confers a four- to fivefold risk, they are both on a continuum of the
same neoplastic process. A diagnosis of ADH made on a core needle biopsy
is usually followed by excision, as studies have shown that depending on
the technique of the biopsy and size of the needle, a follow-up excision is
associated with an in situ or invasive carcinoma in 20% to 40% of these
cases (Adv Anat Pathol. 2003;10:113; Breast. 2011;20:50). It should be
noted that a diagnosis of ADH is only conferred to lesions with low-grade
nuclear cytology; intermediate and high-grade DCIS should be diagnosed
as such regardless of their size.
2. Aypicallobular hyperplasia (ALH) differs from LCIS with regard to the degree
of lobular involvement and expansion (e-Figs. 18.100 to 18.102). ALH
and LCIS are also on a continuum of the same neoplastic process. While
ALH is associated with sixfold increased risk of invasive carcinoma, LCIS
is associated with 12-fold increase in risk.
B. Several intraductal proliferative lesions do not carry an increased risk.
1. Usual ductal hyperplasia (UDH) is characterized by proliferation of a het-
erogeneous epithelial cell population with a tendency to bridge across, fill,
and distend duct lumens. Haphazard placement of cells of variable size
and shape (both epithelial and myoepithelial), variable spacing of the cells
often with prominent streaming and/or swirling, indistinct cell borders, and
irregular and usually peripheral secondary spaces are all features of UDH
that distinguish it from low-grade DCIS (e-Fig. 18.103). Bridges formed
in UDH are usually not rigid, and show stretching with a central attenua-
tion. Immunohistochemically, UDH is usually positive for high molecular
weight keratin (e-Fig. 18.104) unlike most examples of DCIS. UDH is not
a direct precursor of breast cancer; however, if florid, it is a marker of low
increased risk of breast cancer (1.5- to 2-fold), and it is usually not acted
upon clinically.
2. Columnar cell hyperplasia (CCH). The enlargement of normal TDLUs by
hyperplastic epithelial cells is one of the most common abnormalities of
growth in the adult female human breast (Adv Anat Pathol. 2003;10:113;
SeminDiagnPathol. 2004;21:18;AmJ Pathol. 2007;171:252;Histopathol-
ogy. 2008;52:11). These lesions have been called by many names over the
years (Semin Diagn Pathol. 2004;21:18), but currently they are most com-
monly referred to as columnar cell lesions (CCLs) or CCH. CCHs are often
multifocal, bilateral, and up to 100-fold larger than the TDLUs they evolve
from. The majority of CCH are lined by one or two layers of monotonous,
crowded columnar epithelial cells (e-Figs. 18.105 to 18.107), but many
exhibit more diverse histologic features contributing to the complex termi-
nology that has evolved to describe them.
It is currently unknown whether the cytologic atypia occasionally
observed in CCH is associated with significantly higher risk for developing
breast cancer than the majority of cases that are without atypia, although
in some preliminary studies up to 20% of the cases with atypia identified on
core biopsies are associated with cancer in follow-up excisions, a worrisome
association that has lead some authors to advocate follow-up excisions in
this setting (Am] Surg Pathol. 2005;29:734; Histopathology. 2008;52:11).
However, not all studies find a significant relationship between CCH with
atypia and cancer. It seems likely that some CCH may indeed represent
relatively high-risk lesions, but it is possible that histologic features alone

may not always be sufficient to identify them.
IX. COMMON BENIGN ABNORMALITIES. The breast can be involved in a large number of
benign abnormalities. While these findings do not harbor clinical importance in
terms of breast cancer risk, familiarity with their morphologic features is impor-
tant to distinguish them from more clinically significant abnormalities. Only the
most common findings are briefly discussed.
A. Fat necrosis. Although fat necrosis can result from trauma, most cases are
idiopathic, or are secondary prior to surgery or radiotherapy. It clinically and
mammographically can mimic invasive carcinoma. Histologically, a cellular
inflammatory response composed of foamy macrophages and foreign body
giant cells is seen infiltrating the fat (e-Figs. 18.108 and 18.109). In later stages,
fibrosis and dystrophic calcification may be seen.
B. Duct ectasia usually presents in middle-aged women as pain and nipple dis-
charge, with or without an associated mass lesion. It is characterized by dila-
tion of the major duct system, often with luminal amorphous material and
complete filling and distention by intraepithelial foamy macrophages (e-Fig.
18.110). There is usually associated fibrosis and periductal chronic inflamma-
tion.
C. Microcysts and apocrine changes. Cystic dilation of the breast TDLUs (as
opposed to the major duct system in duct ectasia) is quite common and can
produce marked expansion and, especially when associated with microcal-
cification or fibrosis, can result in mammographically detectable and/or pal-
pable lesions. Histologically, these cysts are usually lined by flat nonatypical
epithelium (e-Fig. 18.111). Apocrine metaplasia is a very common change that
occurs in these cystic lesions (as well as normal TDLUs), usually as an inci-
dental finding; it is usually found in premenopausal women and is often part
of the so-called fibrocystic changes. Histologically, apocrine change is char-
acterized by eosinophilic cells with abundant finely granular cytoplasm and
rounded nuclei, often with a prominent nucleoli (e-Fig. 18.112). A papillary
architecture is sometimes evident.
D. Fibroadenoma is a very common benign lesion of the breast that presents as
a mass or radiographic abnormality, more commonly in younger women. On
mammogram, it usually presents as a round, well-circumscribed mass. Grossly,
fibroadenomas are grayish-white, firm, well-circumscribed, lobulated masses.
Histologically, they are composed of a biphasic growth of variably cellular
spindle cell stroma, with deft-like (intracanalicular) (e-Fig. 18.113) or tubu-
lar glandular (pericanalicular) (e-Fig. 18.114) growth patterns. The glandular
elements are composed of two cell layers, an inner epithelial layer and an
outer myoepithelial cell layer (e-Figs. 18.115 and 18.116). Some fibroadeno-
mas, especially those arising in the second decade of life, can grow rapidly
and appear quite cellular (an appearance that overlaps with benign phyl-
lodes tumor); such lesions have been termed cellular fibroadenomas. Myxoid
fibroadenomas display prominent myxoid changes in the stroma and rarely
may be a component of Carney syndrome (primary adrenocortical hypercorti-
solism, skin hyperpigmentation, and a variety of nonendocrine and endocrine
tumors). Fibroadenomas may undergo secondary changes such as infarction
or prominent hyalinization of the stroma (e-Fig. 18.117), with or without
calcification (the former is usually seen with pregnancy, whereas the latter is
more often seen in elderly patients with a longstanding lesion). In addition,
almost all of the epithelial changes that arise in the breast can secondarily
develop in fibroadenomas including various metaplasias, epithelial hyperpla-
sia, sclerosing adenosis, DCIS, LCIS, and invasive carcinoma (e-Figs. 18.118
and 18.119). Fibroadenomas with significant epithelial proliferation have been
termed complex fibroadenomas. The main differential diagnosis of fibroade-

noma is a benign phyllodes tumor.
Phyllodes tumor is much less common than fibroadenoma, accounting for
<3% of the fibroepitheliallesions. As mentioned above, the main feature dis-
tinguishing phyllodes tumor from fibroadenoma is the presence of the charac-
teristic leaf-like architectural pattern produced by extensive branching of the
epithelial component. Stromal hypercellularity is the rule, often with accentua-
tion near the epithelial clefts. Phyllodes tumor can be divided into benign (e-Fig.
18.120), borderline (e-Fig. 18.121), and malignant (e-Fig. 18.122) categories;
the first two are only distinguished by the degree of cellular atypia and mitotic
activity. A focally infiltrative border can be seen in borderline phyllodes tumor.
Malignant phyllodes tumor shows a prominent infiltrative border, unequivocal
sarcomatous areas, and stromal overgrowth (areas of stroma devoid of epithe-
lium). Heterologous sarcomatous elements may also be occasionally present
in malignant tumors. Overall, about 20% of the phyllodes tumors recur (rang-
ing from 17% for benign tumors to 27% in malignant tumors) and 10%
metastasize (0%, 4%, and 22% of benign, borderline, and malignant tumors,
respectively). Recurrences can be associated with grade progression. Phyllodes
tumors are extremely rare and their comprehensive diagnostic features can be
found in more specialized texts.
E. Intraductal papillomas usually show an arborizing growth pattern of fibrovas-
cular cores covered by two layers of cells, one layer of myoepithelial cells
overlying a layer of nonatypical epithelial cells (e-Figs. 18.123 to 18.125).
Intraductal papillomas are generally categorized into two groups: central papil-
lomas, which are usually solitary papillomas, which involve large lactiferous
ducts of the nipple; and peripheral papillomas, which are smaller papillomas
that involve TDLUs and are usually multiple. Central papillomas generally
present as a mass or with nipple discharge. Peripheral papillomas present as
incidental findings or sometimes as a mammographic abnormality. Secondary
hemorrhagic infarction, squamous or apocrine metaplasia, and extensive scle-
rosis (sclerosing papilloma) can sometimes be seen (e-Figs.18.126 and 18.127).
Intraductal papillomas and related lesions can secondarily be involved by other
different intraepithelial proliferations including UDH, ADH, DCIS, and LCIS.
In the absence of any secondary neoplastic proliferation, primary excision is
an adequate treatment for papillomas.
F. Sclerosing adenosis. This relatively common lesion is often incidental and
admixed with proliferative lesions. However, it can present mammographi-
cally with calcification and/or architectural distortion, or clinically as a mass
termed "adenosis tumor" or "nodular sclerosing adenosis." Sclerosing adeno-
sis is characterized by a lobulocentric proliferation of tubular glands (adeno-
sis) accompanied by a fibrotic (sclerosing) stromal proliferation (e-Fig.18.128).
The fact that the glands/tubules are markedly compressed and often obliterated
renders identification of a dual cell layer sometimes difficult (e-Fig. 18.129).
Accordingly, the main differential diagnosis is invasive (usually tubular) carci-
noma, which may be difficult to exclude on small biopsies, especially in areas
in which sclerosing adenosis has a pattern mimicking perineural invasion, or
is secondarily involved by a neoplastic intraepithelial proliferation such as
ADH, DCIS (e-Figs. 18.130 and 18.131), or LCIS. Elongated and compressed
(as opposed to angulated) tubules, lack of a cellular desmoplastic stroma, and
a lobulocentric pattern of growth are useful diagnostic features. If in doubt,
immunohistochemical demonstration of myoepithelial cells can additionally
exclude an invasive process.
G. Radial scar/complex sclerosing lesions are characterized by a central
fibrous/fibroelastotic scar from which a stellate arrangement of benign
ducts/lobules radiates (e-Figs. 18.132 to 18.134). Associated hyperplastic or
neoplastic epithelial proliferations are often identified. In addition to their

microscopic pseudoinfiltrative nature, these lesions may also be clinically, radi-
ologically, or grossly confused with invasive carcinoma due to their fibrotic
nature and their characteristic stellate/spiculated appearance. Nevertheless,
excisional biopsy is still recommended after a diagnosis of radial scar of needle
biopsy because of the occasional presence of associated unsampled DCIS or
invasive carcinoma at the periphery of the lesions.
X. LYMPH NODE STATUS. The presence of metastatic tumor deposits in axillary lymph
nodes as determined microscopically is a highly unfavorable prognostic feature,
as are a high number of involved nodes and (to a lesser degree) large size of the
deposits. Nodal status (N) is so powerful prognostically that it plays a major role
in determining therapy.
The pathology report should include the total number of examined lymph
nodes, the number of positive lymph nodes, size of metastasis, and the presence
or absence of extranodal extension by tumor deposits (the presence of extran-
odal extension is an indicator of tumor recurrence and its presence may dictate
additional radiation therapy). According to the TNM staging system, assessment
of the size of metastasis is important in determining the stage: ITCs are clus-
ters of tumor cells <0.2 mm or <200 cells (e-Fig. 18.135); micrometastasis is
defined as metastatic deposits that measure between 0.2 and 2.0 mm, or >200
cells in one cross section; macrometastasis is defined as a metastatic focus >2.0
mm (Table 18.2). For staging purposes, lymph nodes with a micrometastasis are
counted towards total number of positive nodes if at least one lymph node with
macrometastasis is present; nodes that only show ITCs should not be counted
in the total number of positive nodes. Cancer nodules in axillary tissue without
histologic evidence of classical lymph node are classified as regional lymph node
metastasis unless they are surrounded by breast tissue or DCIS to imply a separate
focus of invasive carcinoma.
Histologically, involvement of axillary lymph nodes by metastatic carcinoma
is most frequently manifested within the subcapsular sinuses with or without
sinusoidal involvement (e-Figs. 18.136 and 18.137). However, metastases in ILC
most commonly appear as scattered individual tumor cells within the parenchyma
and sinusoids. In all of these situations, the size of the largest deposit should be
measured and reported.
There are a few pitfalls in the evaluation of axillary lymph nodes for metastatic
carcinoma. Awareness of these pitfalls will minimize the risk of overdiagnosis.
Heterotopic epithelial elements including heterotopic breast tissue are rare find-
ings that can occasionally be seen in axillary lymph nodes (e-Fig. 18.138). The
heterotopic tissue is subject to all changes that can occur in breast tissue in the
mammary gland itself. The presence of myoepithelial cells and sometimes special-
ized stroma can be helpful diagnostic features. Nevus cell aggregates are capsular
aggregates of nevus cells that are infrequently identified in axillary lymph nodes
(e-Fig. 18.139); they usually present as nests of epithelioid cells within the lymph
node capsule. The diagnosis of nevus aggregates can easily be substantiated by a
combination of S-100 and/or HMB45 immunoreactivity and a negative reaction
with cytokeratin antibodies.
Cytopathology of the Breast

Lourdes R. Ylagan
I. METHODS OF SPECIMEN PROCUREMENT
A. FNA of palpable and nonpalpable breast lesions through mammographic guid-
ance is currently accepted as a cost-effective, reliable, and accurate tool in the
evaluation of breast lesions. The combination of mammographic features, clin-

ical findings, and cytologic evaluation of breast FNA specimens (the so-called
triple test) has considerably decreased the false diagnosis rate of breast cancer.
B. Ductal lavage has recently been employed as a screening method in women
with a personal history of breast cancer, but the sensitivity and accuracy of the
approach for detecting premalignant lesions of the breast ductal epithelium is
still under investigation (Clin Lab Med. 2005;25:787; Am] Surg. 2006;191:57).
II. SPECIMEN ADEQUACY. There is no uniform criterion on which specimen adequacy
can be determined, even in the presence of well-preserved and well-visualized breast
epithelium (e-Fig. 18.140), without taking into consideration the experience of the
aspirator and/or interpreter, clinical presentation, and mammographic findings of
the individual mass.
Ill. DIAGNOSTIC CATEGORIES
A. Negative for malignancy. This diagnosis is generally rendered for benign breast
lesions, including inflammatory or infectious lesions, that are without clinical
or mammographic findings suspicious for malignancy.
B. Atypical cytology. This diagnosis is rendered on cellular lesions showing some
degree of nuclear atypia; the cells generally maintain a well-organized pattern
(e-Fig. 18.141).
C. Suspicious for malignancy. This diagnosis is rendered when the aspirate shows
cells with worrisome cytologic features that fall short of those required for a
diagnosis of malignancy.
D. Positive for malignancy. This diagnosis is rendered when both the quality of the
cytologic changes and the quantity of the malignant cells are sufficient for an
unequivocal diagnosis of malignancy.
IV. CYTOLOGIC FEATURES OF COMMON BREAST LESIONS
A. Fibroadenoma is a clinically well-circumscribed nodule with a homogeneous
mammographic appearance. Cytologically, it is composed of benign ductal cells
arranged ina staghornpattern with abundant myoepithelial cells (e-Fig.18.142).
Abundant stromal fragments may be seen.
B. Gynecomastia is clinically a well-circumscribed and often painful subareolar
lesion in a man. The lesion has mammographic and cytologic findings that are
similar to those of a fibroadenoma (e-Fig. 18.142).
C. Fibrocystic changes typically present as a palpable, ill-defined lesion, which
may have mammographically detectable microcalcifications. Cytologically, the
lesion is composed of apocrine, ductal cells, mucus, and muciphages in varying
amounts (e-Fig. 18.143).
D. Subareolar abscess is a clinically painful, palpable subareolar mass typically
associated with lactation. Cytologically, it consists of neutrophils and benign
anucleate squamous epithelium (e-Fig. 18.144).
E. Ductal adenocarcinoma usually presents as a clinically palpable, mammograph-
ically suspicious mass, which cytologically shows a cellular smear containing
large ductal cells, which maybe poorly cohesive, without a myoepithelial com-
ponent. The cells have pleomorphic nuclei, vesicular chromatin, and prominent
nucleoli. Individual cells may have intracytoplasmic vacuoles containing inspis-
sated material (e-Fig. 18.145).
F. Lobular adenocarcinoma may not be clinically palpable, but presents mammo-
graphically as an ill-defined mass lesion. Cytologic preparations show singly
dispersed small plasmacytoid cells with vacuolated cytoplasm often containing
inspissated material. The nuclei are uniformly small, round-to-oval, and not
much bigger than a neutrophil (e-Fig. 18.146).
V. SPECIAL STUDIES. Immunocytologic evaluation of estrogen and progesterone recep-
tor studies can be performed on cytospin slide preparations. Immunocytologic eval-
uation of HER2 on cytospin slide preparations is limited due to the requirement of
an intact cell membrane for proper assessment. Immunohistochemical evaluation
of these markers can also be performed on paraffin-embedded cell block samples

prepared from fine needle aspirates. FISH for HER2 gene amplification can be
performed on cytospin slide preparations or paraffin-embedded cell blocks.
SUGGESTED READINGS
Dabbs DJ. Breast Pathology. Philadelphia: Elsevier; 2012.
Mckee GT. Cytopathology of the Breast with Imaging and Histologic Correlation. Oxford: Oxford
University Press; 2002.
O'Malley FP, Pinder SE, Mulligan AM. Breast Pathology: A Volume in the Foundations in Diag-
nostic Pathology series. 2nd ed. Philadelphia: Elsevier Saunders; 2011.
Page DL, Anderson TJ. Diagnostic Histopathology of the Breast. Edinburg: Churchill Livingstone;
1987.
Rosen PP. Rosen's Breast Pathology. 3rd ed. Philadelphia: Lippincott Williams & Wilkins; 2009.
Schnitt SJ, Collins LC. Biopsy Interpretation of the Breast. Philadelphia: Lippincott Williams &
Wilkins; 2009.
Tavassoli FA. Pathology of the Breast. 2nd ed. Chicago: McGraw-Hill; 1999.
Tavassoli FA, Devilee P. World Health Organization Classification of Tumors: Tumors of Breast
and Female Genital Organs. Lyon: IARC press; 2003.
Tavassoli FA, Eusebi V. AFIP Atlas of Tumor Pathology, Series 4, Tumors of the Mammary Gland.
Silver Spring, MD: ARP press; 2009.
Zakhou H, Wells C, Perry NM. Diagnostic Cytopathology of the Breast. London: Churchill Liv-
ingstone; 1999.
SECTION V
Urinary Tract
Medical Diseases of the

Kidney
Joseph P. Gaut and Helen Liapis
I. MEDICAL RENAL BIOPSY HANDLING AND PROCESSING. This chapter covers med-
ical renal biopsies; biopsy for diagnosis of renal masses is covered in Chap-
ter 20. Renal allograft biopsy pathology is added at the end of this chapter.
Renal biopsies are usually received in transport media, allowing distribution of
tissue for light microscopy (LM), immunofluorescence (IF), and electron micros-
copy (EM).
A. Light microscopy. A minimum of three hematoxylin and eosin (H&E), one
trichrome, two periodic acid-Schiff (PAS) stains, and one Jones silver-stained
section is required. A minimum of seven glomeruli and one artery is required
for adequate LM evaluation.
B. Immunofluorescence. A minimum of two glomeruli is required. A direct IF
method is routinely used employing a panel of antibodies, including anti-IgG,
IgA, IgM, C3, Cl q, fibrinogen, albumin, and kappa and lambda light chains. It
is important to document the staining pattern (linear vs. granular) and distribu-
tion (mesangial, loop, or combined).
C. Electron microscopy. Ultrastructural evaluation of two glomeruli is recom-
mended. EM allows for evaluation of cellular and extracellular abnormalities in
the glomeruli, tubules, interstitium, and vessels. EM is very useful in confirming
the presence and distribution of electron-dense deposits, which can be located
in the mesangium or the glomerular capillary basement membrane. In the base-
ment membrane, electron-dense deposits can be subepithelial, subendothelial,
or intramembranous (surrounded by basement membrane).
II. GLOMERULAR DISEASES. Glomerular diseases may be primary or secondary (associ-
ated with systemic diseases). Patients can be grouped into those that present with
nephrotic syndrome (>3 g urine protein/day), nephritic syndrome (proteinuria+
hematuria), or isolated hematuria. It is important for the pathologist to have access
to patient•s clinical and laboratory data. For example, minimal change disease
(MCD), focal segmental glomerulosclerosis, and membranous glomerulonephri-
tis (GN) usually present with nephrotic syndrome. In contrast, postinfectious GN
usually presents with nephritic syndrome.
A. MCDifocal segmental glomerulosclerosis (FSGS). MCD and FSGS are the most
common causes of nephrotic syndrome in children and are also common in
adults, affecting about 10% to 20% of patients with kidney disease. Primary
329
330 I SECTION V: URINARY TRACT
FSGS may be idiopathic, secondary, or familial (Am J Nephrol. 2003;23:353).

Less than nephrotic range proteinuria may be present in advanced cases. Some
patients have concurrent hematuria and hypertension (HTN).
The classic findings in MCD (Table 19.1) are as follows:
LM: Normal-appearing glomeruli
IF: Negative
EM: Extensive foot process effacement (e-Fig. 19.1).*
The diagnostic features of FSGS are as follows:
LM: Segmental sclerosis in some but not all glomeruli. Accurate diagno-
sis of FSGS depends on the extent of the disease and the number of glomeruli
present in the biopsy. Diagnosis may be missed because of sampling error, par-
ticularly with the smaller needles currently used. It has been estimated that in a
biopsy containing 10 glomeruli, there is a 35% chance of missing FSGS. Notably,
even one glomerulus with FSGS is sufficient for diagnosis. The corticomedullary
glomeruli are the first to be sclerosed; therefore, needle biopsies should opt to
sample this region.
IF: Is either entirely negative or has focal and weak mesangial C3 or IgM
immunoglobulin deposits.
EM: Shows focal foot process effacement (e-Fig. 19.2), the degree of which
may depend on the degree of proteinuria. Other EM findings in MCD/FSGS
include microvillus transformation of foot processes, endothelial cell edema,
podocyte detachment, and glomerular basement membrane (GBM) wrinkling
(see e-Fig. 19.2F).
Beyond this classic presentation, light and electron microscopic findings may
be similar in MCD and FSGS. For example, glomeruli may appear normal in
FSGS and focal foot process effacement may be present in MCD.
Glomerular hypercellularity and enlargement (glomerulomegaly) is thought
to represent an early lesion of FSGS. A recent FSGS classification scheme
describes various histologic patterns with significantly different prognosis (Kid-
ney Int. 1990;38:115). Glomerulomegaly is rare (seen in only about 3% of
FSGS), but the following variants are more frequent: perihilar FSGS (26% ), tip
lesion (17%), usual type not otherwise specified (NOS) (42%), and collapsing
FSGS (11%) (see e-Fig. 19.2A-E) (Am J Kidney Dis. 2004;43:368). The tip and
cellular lesions have the best prognosis and collapsing FSGS the worst (Kidney
Int. 2006;69:920). Tubulointerstitial damage and vascular thickening indicate
chronic disease. Pathologic similarities between MCD and FSGS initially sug-
gested that they were one disease with a spectrum of findings, but recent clinical
and molecular studies have demonstrated that FSGS has worse prognosis and
different pathogenesis.
B. Collapsing FSGS is a distinct FSGS variant considered by some to be an entirely
different disease.
LM: Characterized by segmental glomerular capillary collapse and podocyte
hypertrophy, often accompanied by microcystic tubular dilatation and intersti-
tial inflammation (see e-Fig. 19.2E). The main difference from usual FSGS is the
collapse of the loops versus sclerosis, and podocyte proliferation versus podocyte
loss.
IF: Nonspecific.
EM: Shows proliferating podocytes and wrinkled/collapsed capillary loops.
Clinically, it is characterized by black racial predominance, a high incidence of
nephrotic syndrome, and rapidly progressive renal failure. First identified in lllV
patients, collapsing FSGS was later recognized in association with viruses such
as Parvovirus 19 and hepatitis Band C, and with pamidronate chemotherapy
I"IIJ!I Jli!U llljof81emiNirPalbou•- - l o i i X . . , -. . IJii.M IIIII:-
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'lllln momtni'IO Cei!L61r""" 1Do11'10Dhi'OI*IIY AIDOlt
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.olOll'.•
-ON
E!Jtj """'IliA
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..-.-
-!lor
COI!Ipsq
Clq ,.pii/'Oj)llhy
QA:G P>m"'*'J)ftlly
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~ ,..,.,_,11111 ~~ ON.II1""""""01'hrbo; HOllo rd «! ..

<llorp- ; MPGN, mon .. '"""'o!- --!lied.
IQ, ..,munOIII:>WI1: HSP.
Jl)cmoNio""'hr1111; MillO, --111111111'101lcC>ulln - - · - ·
..,iidt4'<:-"""""" Mel), m'*"'l
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(Semin Diagn Pathol. 2002;19:106). It is an aggressive and difficult to treat dis-

ease (Semin Nephrol. 2003;23:209), and may also involve the allograft kidney.
Pathogenesis involves podocyte cell cycle dysregulation resulting in prolifera-
tion.
C. Mesangial proliferative glomerulonephritis {IGA, lgM, lgG and C3). Mesangial
hypercellularity is defined as more than three mesangial cells per glomerular seg-
ment; many glomerular diseases may have increased mesangial cells (Table 19.1)
including variants of MCD and FSGS. Mesangial hypercellularity indicates
mesangial immune deposits and/or reactive proliferation. The most common
disease is IgA nephropathy, clinically characterized by micro or macrohema-
turia and varying proteinuria (rarely nephrotic syndrome or crescentic GN)
(Nephrol Dial Transplant. 2001;16 Suppl 6:77).
LM: Mesangial hypercellularity varies from focal to diffuse.
IF: Diagnosis is made by the presence of predominant or co-dominant IgA
mesangial deposits (e-Fig.19.3). Mild lgGandigM deposits may also be present,
particularly in Henoch-Schonlein purpura (HSP), which is thought of as the
systemic form of IgA.
EM: Shows mesangiaVparamesangial deposits and occasionally capillary
loop, subendothelial deposits that may extend to the GBM and cause splitting
(more common in HSP), or "humps."
Histological parameters which negatively affect prognosis in IgA disease
include glomerular sclerosis, capillary wall IgA deposits, and vascular and tubu-
lointerstitial fibrosis. Glomerular sclerosis is the best independent predictor of
adverse outcome and renal failure. HSP mimics IgA pathologically, but clinically
is a systemic disease that presents with skin rash, arthritis, and abdominal pain
in addition to nephritis. IgA is the most common glomerular disease worldwide
with variable prognosis (""30% develop end-stage renal disease [ESRD]). HSP
tends to be self-limiting with only about 18% of patients progressing to ESRD.
Both may recur in transplant kidneys, but clinical symptoms are mild despite
IgA deposition.
Mesangial hypercellularity not infrequently accompanies various types of
GN. For example, MCD and/or FSGS with mesangial hypercellularity are gen-
erally thought to have a worse prognosis. Focal IgM deposits are seen in some
such cases. Rarely, diffuse lgM deposits are detected (which have raised consid-
erable debate whether they represent a separate entity named lgM nephropathy).
An entity known as Clq nephropathy is characterized by predominant
C1q deposits and is considered a variant of MCD/FSGS (Am] Clin Pathol.
1985:83:415). C1q nephropathy is primarily a disease of children and young
adults. Other glomerulopathies characterized by isolated C3 or IgG mesangial
deposits in patients without lupus stigmata were described recently and named
accordingly. IgG glomerulopathy is a pediatric disease that may be an aberrant
manifestation of lupus nephritis (LN), and it has been proposed that it should
be treated as such UASN 2002;13:379).
Recent studies have clarified the entity known as C3 glomerulopathy, which
may present with mesangial hypercellularity or minimal changes. Immunofluo-
rescene demonstrates isolated, granular, mesangial C3 deposits without IgG or
C1q staining (e-Fig. 19.4). Interestingly, the disease is associated with dysregula-
tion of the alternative complement pathway secondary to defects in complement
factor H (Nat Rev Nephrol. 2010;6:494; Kidney Int. 2009;75:1230).
D. Membranous GN is the most common cause of nephrotic syndrome in adults
(30% to 50% of cases). It may occur at any age, but accounts for <5% of
childhood nephrotic syndrome. Most cases are idiopathic, but ""10% are asso-
ciated with identifiable causes such as malignancy, autoimmune diseases (e.g.,
systematic lupus erythematosus), drugs, and infections (hepatitis B, syphilis).
Glomerular lesions resemble those seen in Heymann nephritis, an animal model
in which antibodies react with the Heymann antigen, a complex of megalin and
Chapter 19 • Medical Diseases of the Kidney I 3 33
the receptor-associated protein. expressed in the tubular brush border and the
basal surface of the visceral epithelial cells.
LM: Membranous GN is a diffuse process in which the glomeruli are not
hypercellular but usually exhibit thickening of the capillary basement membrane
while maintaining luminal patency. In Jones silver-stained sections, the base-
ment membrane can show "spikes" projecting from the epithelial side of the
basement membrane. Spikes result from the presence of subepithelial electron-
dense deposits (silver stain negative) and deposition of basement membrane-like
material on the sides of the deposits (e-Fig. 19.5). Glomeruli appear essentially
normal in early cases.
IF: Diffuse, granular staining for IgG and C3 is present along the glomerular
capillary loops by IF.. Other immunoglobulins can be present but have lower
intensity staining.
EM: At early stages, the electron-dense deposits are subepithelial. As the
disease progresses, deposition of basement membrane-like material at the sides
of the electron- dense deposits occurs so that with time, the electron-dense
deposits are surrounded by basement membrane and thus becomes intramem-
branous. The deposits eventually become electron-lucent, suggesting resolution
(e-Fig. 19.5).
E. Postinfectious glomerulonephritis {PIGN). PIGN is a classic complication of strep-
tococcal pharyngitis and presents acutely with nephritic syndrome. However,
classic PIGN is currently infrequent; most cases follow staphylococcal skin infec-
tions and other bacterial, viral, fungal, or parasitic infections, and are more
frequently chronic or atypical (Hum Pathol. 2003;34:3).
LM: The pathology is unique, characterized by white cells in the
glomeruli (predominantly neutrophils in the acute phase), and lymphocytes or
macrophages in chronic cases.
IF: There are large granular IgG and C3 deposits along capillary loops (e-
Fig. 19.6). Occasionally, deposits are located predominantly in the mesangium
(instead of in the loops) and are C3 or lgA/IgM instead of IgG (Semin Diagn
Pathol. 2002;19:146).
EM: Shows characteristic bell-shaped deposits (humps). Erythrogenic toxin
type B is thought to be the target antigen for immune complexes that are
implanted in the GBM.
F. Membranoproliferative glomerulonephritis {MPGN). Patients with MPGN can
present with features of nephrotic and/or nephritic syndrome, and most patients
have a low serum C3level. Although MPGN can affect patients of all ages, it is
more common in children. Three types of MPGN have been described; because
all types can have similar histologic findings, EM is used to differentiate them.
Type I is the most common type, followed by types ll and Ill. It is important
to note that MPGN type I can be associated with other diseases such as viral
hepatitis, so patients should be worked up for secondary causes of MPGN when
the pathologic diagnosis is established.
LM: MPGN is a diffuse glomerulopathy with endocapillary proliferation
that results in lobular accentuation (e-Fig. 19.7). The GBMs are thick, and the
capillary lumens are not evident. The Jones silver stain reveals double GBM con-
tours, also known as "tram-tracking." These findings are more commonly seen
in MPGN type I. MPGN type II tends to have a less consistent histologic pattern.
IF: A strong and diffuse granular staining for C3 is observed along the
glomerular capillary walls and the mesangium. Approximately 60% of type
I MPGN also exhibit IgG and/or C1q immunostain. Negative immunostains for
immunoglobulins and Clq are usually observed in MPGN type II.
EM: This is the most useful tool for differentiating MPGN types. MPGN
type I shows mesangial interposition (extension of mesangial cell cytoplasm
into the capillary wall) and subendothelial electron-dense deposits. When the
mesangial cell cytoplasm extends into the glomerular capillary wall, basement
membrane-like material is laid down by the mesangial and endothelial cells, cre-
ating a second "new" basement membrane. This process results in the "tram-
tracking" observed by LM. MPGN type II is also known as dense deposit disease
because it has ribbon-like electron-dense deposits along the capillary walls, often
replacing the lamina densa. These deposits are not necessarily present in all cap-
illary loops and may only be present in some segments and the mesangium. The
composition of these electron-dense deposits remains uncertain. The ultrastruc-
tural findings of MPGN type III are similar to those of MPGN type I, but with
subendothelial and subepithelial electron-dense deposits (e-Fig. 19.7).
G. Crescentic glomerulonephritis (crescentic GN}. The term "crescentic GN" refers
to the presence of cellular crescents in >50% of glomeruli available in a renal
biopsy. Its usual clinical presentation is that of rapidly progressive GN. Patients
can have a renal limited or systemic disease.
LM: Cellular crescents are identified in >50% of glomeruli (e-Fig. 19.8).
Necrotizing glomerular lesions are commonly seen. Additional morphologic
findings are dependent on the type of renal limited or systemic disease.
Immunofluorescence is used to further classify crescentic GN.
IF: In anti-GBM disease or Goodpasture•s disease, glomeruli exhibit smooth
linear IgG staining along the capillary basement membrane. Immune-complex
GN has a granular staining pattern for one or more immunoglobulins. Pauci-
immune GN has negative immunostain for all immunoglobulins.
EM: The smooth linear IgG staining of anti-GMB or Goodpasture•s dis-
ease does not have a morphologic correlation that can be detected by EM. The
glomerular changes noted by EM correspond to the necrosis, disruption of the
GBM, and crescents that are seen in all crescentic GN. Similar findings are
also identified in pauci-immune GN. Cases of immune-complex GN will show
electron-dense deposits (e-Fig. 19.8).
H. Lupus nephritis (LN}. Systemic lupus erythematosus (SLE) is a multisystemic
autoimmune disorder with a peak incidence in the second and third decades of
life and a female predominance (male-to-female ratio of 1:9). It is more common
in African-Americans. The clinical diagnosis of lupus is based on clinical and
laboratory criteria established by the American Rheumatism Association. Renal
involvement by the disease is relatively common; approximately half of lupus
patients develop lupus nephritis during the first year of the disease. Although
this chapter emphasizes the glomerular findings of (LN), interstitial, tubular, and
vascular lesions can also accompany the glomerular changes. Use of the ISN/RPS
classification of LN, which is a modification of the WHO classification (Kidney
Int. 2004;65:521), is recommended.
LM: LN can present with various light microscopic patterns (Tables 19.2
and 19.3). With time and/or treatment, the renal lesion can evolve into a dif-
ferent class. Membranous LN can occur in combination with Class lli or IY.
The following are considered glomerular active lesions: endocapillary hypercel-
lularity with/without leukocyte infiltration and with substantial luminal reduc-
tion; karyorrhexis; fibrinoid necrosis; rupture of the GBM; cellular or fibro-
cellular crescents; wire loops; hyaline thrombi (e-Fig. 19.9). The glomerular
chronic lesions are segmental or global glomerulosclerosis, fibrous adhesions,
and fibrous crescents.
IF: A glomerular IgG-positive immunostain is almost universal in LN; the
term "full house" is used when IgG, IgA, and IgM immunostains are positive.
C3 and C1q are usually present.
I. Diabetes mellitus (DM}. DM is a disorder of carbohydrate metabolism that affects
multiple organ systems. In the kidney, it increases the propensity to pyelonephri-
tis, papillary necrosis, arteriosclerosis, and glomerular disease. Hyperglycemia
in these patients induces biochemical changes in the GBM, nonenzymatic glyco-
sylation of proteins, and hemodynamic changes with glomerular hypertrophy.
Cl't8ptet 19 • Medical OieeM:Ieeot th& Kid""Y I II&
,,(,I ]IJ(fll taUIIPS Claaltlcii!Oiuf L11p1111Nepiii1U.
-·
Claal
Claall
Mlrlmal mmnJPI LN
-..,11"'1,.,_ LN
f<><111 LN ~of <!iOlCo
!IleAl
Ill we!
Ill (C)
"""'""I)
fotOI D10llfera1MI LN (DUrei/-IGolonel
Fa pmlllar.o1No and .llderoalng LN (..- ond
dlronle IMiona)
Fa -.uwLN (chronic:-- pmo!Wor
sail)
Claal¥ Dlluoo LN (Aoltfa Oflne- dlluto, ""'"'"""" , _ pobel end-lilly
01\d/ot -li!IBIYQN m.:Mn1 ~or m010 abnoru!D
IV-$ lAlor IV-<l CAl Dlllll.. -GMIII « eSoi:lol pr<lferat.'o LN
IV-S(M:l«IV-GWCl Oll!llaa~Ofgjabelp--and
111-S(C)«IV-G(C) scllrooirc LN
CltftWa ~"' alab<l sdoroolna LN
Mamb111nou.t LN
A<h11>011d ocloroslrc LN (!10!!; or m«e J,bnorUI l)ol>oltyocla.-)
c11a1 u- IIII:fOICIIIY ........ •-•m ~- llii!:~D~C~~~r

M!nlmalmooonllol ~+ ~~d~
II
Ill lna--
,.,.....,..,ldj
-rc•I .. ·~ ~~~lt:yond
EnOo<:o!illuy and/or
-~~~~IJ pr<lfcnllon In
<50% of alamerull
~+
~and loop+ ,..,.,..! and
tuban-olal d.,....
Enclocoli!IBIY •ndlor ~andloop+ ~l•nd

-~~~~IJ pr<lf-n In tubandoiii ...Id""'*
50%«,.,. of pmo<UI
v TNck lc:opund ,....,..! Numnus:subep!thelll
lrypoo -~or~~~ a n d - miiOIII,IIol
"-b
polymerized proteins forming beta-pleated sheets and stains red with Congo
red.
LM: Varies from GBM thickening to nodular or segmental sclerosis. Amor-
phous material is often apparent in the mesangium or the capillary loops. A
Congo red stain is diagnostic revealing apple green fluorescence under polariz-
able filters (e-Fig. 19.11).
EM: This shows characteristic randomly arranged fibrils of 4 to 12 nm
(e-Fig. 19.11C). It shows either finely granular or filamentous deposits in the
mesangium or the GBM (Semin Diagn Pathol. 2002;19:116).
IF: May show K or l chains. Some deposits of fragments of IgG, IgM, or
lgA immunoglobulin heavy chains or light chains do not form amyloid; in these
cases, a Congo red stain is negative and the term "congo red negative amyloi-
dosis" is used, or the equivalent term "monoclonal immunoglobulin deposition
disease" (MIDD).
The differential diagnosis of MIDD includes other nodular GN such as
diabetes and fibrillary and immunotactoid glomerulopathy. The latter may
have overlapping IF, but EM findings are distinct from amyloid (Kidney Int.
2002;62:1764). The other pathologic findings in MIDD are as follows:
LM: In MIDD, glomeruli may appear nodular or have nonspecific changes.
Diagnosis is usually (but not always) made by IF.
IF: Identifies the specific type of immunoglobulin fragment as either heavy
or light chain UAm Soc Nephrol. 2001;12:1482) (e-Fig. 19.12).
K. AI port syndrome/thin basement membrane disease. Classic X-linked (X-L) Alport
syndrome presents with microscopic or gross hematuria and deafness. Deafness
and/or proteinuria are indications of severe disease. Such symptoms are usually
absent in young children. Mutations in COLIVAS cause X-L AIport; COLIVA4
mutations cause autosomal dominant (AD) and COLIVA3 mutations cause
autosomal recessive (AR) Alport. As many as lS% of X-L Alport patients have
no family history and are thought to represent a new mutation. Patients with
AR Alport are clinically similar to X-L Alport; however, AD Alport is rare.
The involved organs in Alport syndrome reflect the sites where these collagen
IVa chains are normally expressed (a3, a4, and aS are exclusively found in
the kidney, eye, and ear). Renal biopsy is performed for diagnosis as well as
assessment of disease progression.
LM: This is not specific; it varies from unremarkable glomeruli, to FSGS,
to chronic scarring. Foamy interstitial cells were once thought characteristic of
Alport, but they are in fact seen in many types of proteinuria.
IF: Routine stains are negative. Diagnosis of Alport is facilitated by collagen
a3-S chain immunostaining. The aS chain of collagen IV is distinctly absent
in X-L Alport, and it is accompanied by the absence of a3 and a4 chains in
glomeruli. (Collagen IV monomers are incorporated into basement membrane
as triple helices to form the structural meshwork [Kidney Int. 2004;6S:1109];
when one chain is mutated, all three chains may degenerate because they become
susceptible to enzymatic proteolysis.) Typically, women with X-L Alport have
mosaic linear staining with collagen IVaS in the glomeruli or skin (e-Fig. 19.13)
(Hum Pathol. 2002;33:836). However, some women may have positive stain-
ing, making it difficult to distinguish from thin membrane disease (TMD). A
screening test for families with potential X-L Alport is skin biopsy (IVaS is the
only chain found in skin); the diagnostic pattern is absence of collagen IVaS in
men and mosaic staining (linear interrupted positivity) in X-L women carriers
(e-Fig. 19.13B-D).
EM: The cardinal pathologic findings ofX-LAlport are seen on EM and con-
sist of abnormal splitting, widening, or thinning of the GBM with degeneration
of collagen IV in the lamina densa known as "bread crumbs" (e-Fig. 19.13).
The lesions may involve tubular basement membranes as well. However, the
specificity of the EM findings is moderate (Kidney Int. 1999;56:760); for exam-

ple, young children and women with Alport nephritis may only manifest uni-
formly thin GBM and thus be indistinguishable from TMD (Semin Nephrol
2005;25:149). About 30% of Alport heterozygotes have uniformly thin GBM,
but have collagen IVa3-5 mutations by genetic testing. Others show only GBM
lamellation in thin segments. Routine genetic testing is not only technically dif-
ficult (several different genes must be evaluated and mutational hot spots do not
exist), but also does not strictly correlate with pathology or prognosis. A good
family history of close relatives is very helpful.
TMD is defined as GBM thinning <200 nm. It is debated as to whether GBM
thinning must be diffuse or focal, though most cases have focal GBM thinning
involving <50% of loops.
LM: Usually unremarkable.
IF: Routine stains are negative. Collagen 1Va3-5 immunostains are positive.
EM: Shows GBM diffuse or segmental thinning without lamellation (e-Fig.
19.13F).
Ill. TUBULOINTERSTITIAL DISEASES
A. Infectious processes. Infectious agents can result in tubulointerstitial nephri-
tis by colonizing the renal parenchyma, or by triggering a systemic immuno-
logic response that will target the renal tubules and interstitium. Bacterial
agents are responsible for most cases of acute pyelonephritis. Most cases of
acute pyelonephritis are ascending infections caused by gram-negative bac-
teria, particularly Escherichia coli. When bacteria reach the kidney using a
hematogenous route, Staphylococcus aureus is usually the responsible agent.
Acute pyelonephritis is characterized by an abundance of neutrophils in the
lumen of tubules and the interstitium. Neutrophils are usually accompanied by
other inflammatory cells.
Viral agents are also associated with tubulointerstitial nephritis. Polyoma
virus and cytomegalovirus (CMV) are particularly important in renal allo-
grafts. The polyoma BK virus is a DNA virus with high prevalence dur-
ing childhood, causing respiratory infections. Respiratory infection follows
hematogenous spread, reaching organs like the kidney, where it remains dor-
mant; when the patient later becomes immunosuppressed, the virus reactivates.
Kidneys with either of these viral infections show a chronic interstitial inflam-
mation, sometimes associated with tubulitis, a pattern that is difficult to dis-
tinguish from acute cellular rejection. Both viruses elicit cytopathic changes
that are seen in endothelial and tubular epithelial cells; polyoma virus forms
an intranuclear basophilic inclusion, and CMV forms cytoplasmic and intranu-
clear basophilic inclusions with a perinuclear halo. It is useful to confirm the
morphologic findings of viral infection using antibodies against the specific
microorganism.
A granulomatous inflammatory response is usually seen with mycobac-
terial or some fungal infections. Histochemical stains like acid-fast bacil-
lus (AFB), PAS, and silver stains are needed to visualize the micro-
orgamsms.
B. Acute tubular necrosis (ATN). ATN is the most common cause of acute renal fail-
ure and can be caused by toxins or ischemia. The histologic changes are mainly
confined to the tubules with necrosis of the lining epithelium. Regardless of
the cause, the proximal tubule is the main target in ATN. Epithelial necrosis is
more extensive and confluent in the toxic type of ATN, but tends to be patchy
in the ischemic type. Since necrotic tubular epithelium will exfoliate into the
tubular lumen, focal denudation of the tubular basement membrane is com-
mon (e-Fig. 19.14). Epithelial cells can also exhibit regenerative changes with
nuclear enlargement, prominent nucleoli, and mitotic activity. Interstitial edema
is usually present in ATN.
C. Allergic interstitial nephritis. Most cases of allergic interstitial nephritis are drug
related; renal manifestations develop "'2 weeks after drug exposure. The typ-
ical presentation includes feve.t;, skin rash, and eosinophilia, but this triad is
only present in approximately one-third of the patients. Interstitial eosinophils
are the most important histologic clue for diagnosis. Eosinophils are accompa-
nied by other inflammatory cells, mainly lymphocytes, and may only be seen
focally (e-Fig. 19.15). Tubulitis and tubular injury usually accompany the inter-
stitial changes. Nonsteroidal anti-inflammatory drugs (NSAIDs) can also cause
allergic interstitial nephritis, but interstitial eosinophils are usually rare. More-
over, patients taking NSAID can also develop nephrotic syndrome, and their
glomeruli exhibit changes indistinguishable from MCD.
D. Cast nephropathy. Cast nephropathy is a renal complication of multiple
myeloma. Casts are usually located in the collecting ducts, but due to retro-
grade filling, casts can even be seen in the proximal tubules. Casts are composed
of light chains and Tamm-Horsfall protein, and tend to have a fractured
appearance. A granulomatous response with multinucleated macrophages and
reactive epithelial cells is typically found at the periphery of myeloma casts
(e-Fig. 19.16); other inflammatory cells, such as neutrophils and lymphocytes,
can also be part of the inflammatory process. Nonspecific changes such as
tubular atrophy and interstitial fibrosis are commonly present. IF studies
exhibit a monoclonal light chain in the casts in most cases.
IV. VASCULAR DISEASES
A. Vasculitides. The kidney has a dense arterial, venous, and lymphatic network.
Vasculitides affect all these vessels, but most commonly the arteries. They are
usually classified as large, medium, and small vessel vasculitides on the basis
of the predominant vessel size affected, but although the classification is con-
venient, it is controversial because there is variability in the size of affected
vessels. Vasculitides are also classified by the underlying pathogenetic mecha-
nism as either immune-complex mediated or pauci-immune (absence of immune
deposits). The following are the most common vasculitides affecting the kidney,
grouped by size:
1. Takayasu's aortitis affects large branches of the abdominal aorta and the renal
arteries. Mural inflammation with giant cells is the distinct feature.
2. Polyarteritis nodosa, scleroderma, lupus, rheumatoid arthritis, and Wegener's
granulomatosis affect medium- to small-sized vessels. These are systemic dis-
eases and, with the exception of lupus, are pauci-immune systemic vasculi-
tis. Kidney pathology is variable and includes interstitial granulomas, arte-
rial acute inflammation, fibrinoid necrosis, and thrombotic occlusion. The
inflammation can lead to aneurysm formation or fibrous wall thickening.
Involvement of glomerular capillaries leads to crescentic GN. The diagnosis
largely depends on the clinical syndrome, not the pathological findings. For
example, Wegener•s granulomatosis is associated with serum autoantibodies
against components of neutrophils known as circulating antineutrophil cyto-
plasmic antibodies (ANCA). These antibodies correlate with disease activity
and underscore the pathogenesis of vascular injury (see Chap. 9).
3. Of the small vessel systemic vasculitides, some are referred to as thrombotic
microangiopathies(TMA). A typical presentation is with hemolytic anemia
and thrombocytopenia, known as hemolytic uremic syndrome (HUS). Enti-
ties included under TMA are diarrhea associated (classic HUS), thrombotic
thrombocytopenic purpura, eclampsia, pre-eclampsia, thrombosis due to
drugs (oral contraceptives, chemotherapy drugs such as bleomycin), cryo-
globulinemia, idiopathic hypereosinophilia, antiphospholipid syndrome,
hereditary deficiency of blood clotting factors, and malignant HTN (] Am
Soc Nephrol. 2003;14:1072). Renal biopsy shows arterial intimal thicken-
ing (called mucoid degeneration) and luminal fibrin thrombi (e-Fig. 19.17A).
Chapter 19 • Medical Diseases of the Kidney I 339
All of these entities may also show characteristic arteriolar thickening (also
known as onion skinning, e-Fig. 19 .17B), endothelial swelling, and/or lumi-
nal thrombosis.
B. Systemic HTN. Vascular diseases of the kidney are often associated with systemic
HTN. The most common cause of HTN is idiopathic, which, despite modern
pharmacologic therapies, remains a primary cause of renal failure. High blood
pressure damages parenchymal arteries causing wall thickening; the arterioles
show characteristic acellular eosinophilic material due to increased intravascular
pressure and endothelial injury that allows leaking and deposition of plasma pro-
teins. Glomeruli beyond damaged arterioles undergo sclerosis (e-Fig. 19.17B).
1. Malignant HTN, defined asdiastolic pressure> 120 mmHg, complicates ""10%
of patients with benign HTN, presents acutely with papilledema, nau-
sea, vomiting, convulsions, and coma. Renal function declines rapidly and
patients develop gross hematuria, albuminuria, and hemolytic anemia. Onion
skinning is characteristic (e-Fig. 19.17C).
2. A small fraction (5% to 10%) of benign HTN is secondary to specific
causes, some of which may be amenable to surgical therapy, that together
are known as renovascular HTN. Most common is renal artery stenosis
due to atherosclerotic aneurysms in older mostly diabetic men. Patients with
abdominal or renal artery atherosclerosis may present with acute renal failure
or proteinuria due to cholesterol embolism in parenchymal arteries (e-Fig.
19.17D). Fibromuscular dysplasia, an idiopathic mural thickening of the
renal artery, is seen in young women (Am] Kidney Dis. 1997;29:167).
V. PATHOLOGY OF THE ALLOGRAFT KIDNEY
A. Processing of renal allograft biopsies. Renal biopsy is a key component of man-
aging transplant recipients since clinical diagnosis is changed in rv40%, and
therapy in ""60% of cases following biopsy (Am] Kidney Dis. 1998:31:515).
Transplant biopsies in most centers are performed when clinically indicated
(indication biopsies) or at standardized time intervals (protocol biopsies). Crite-
ria for indication biopsies include proteinuria, acute renal failure, and increased
creatinine levels, among others. Adequacy criteria of the renal allograft biopsy
were established at the BANFF 1997 Consensus Conference (Kidney Int.
1999:55:713).
It is recommended that two 16- or 18-gauge needle core biopsies are sub-
mitted for pathologic evaluation: one for LM and the other divided for IF and
EM. The core for LM is submitted in 10% buffered formalin; the IF core should
be submitted in refrigerated transport media; and a small fragment from cortex
should be submitted for EM fixed in 2% glutaraldehyde. For LM, 10 glomeruli
and 2 arteries are required for definitive diagnosis; 7 to 10 glomeruli and 1
artery are considered marginal, while <7 glomeruli and no arteries are an inad-
equate sample. The paraffin blocks should be sectioned at 3 to 4 Jtm thickness;
seven slides including three H&E stains, three PAS or silver stains, and one
trichrome stain should be prepared (Kidney Int. 1999;55:713). For IF, a min-
imum of two glomeruli is required for the evaluation of glomerular disease;
however, IF evaluation for IgG, IgA, IgM, C3, fibrinogen, albumin, and C4d is
standard. For electron microscopic evaluation, at least one glomerulus is rec-
ommended. See Table 19.4 for BANFF diagnostic categories for renal allograft
biopsies.
B. Transplant rejection. Major complications with specific pathologic findings in
the allograft biopsy are acute and chronic rejection (which are subdivided into
cellular and humoral rejection), and recurrent and de novo disease (see Table
19.2).
1. Acute T-cell-mediated rejection. Tubules, arteries, and the interstitium are
the principle targets of acute T-cell-mediated rejection. The BANFF 2005-
updated classification of renal allograft rejection (see Table 19.2) is based on
aco I SECTION v. URINARY TRAel
1'1,1,!' JttS 1~;:.w ~ cu,or~u for ~~o~n~~ Alh~'lft llloP'IIeHI-" ' "
1.-1
2. Arili>ody<nod- ro)od!on
Quo lo clocumon1Dd ...-n., ..tlbocl;r, C4it 1nd .,q1lll prll>i>l:v
kl/1» •Mibo<tr'-rnodllllod reJedloh
'l'fl>oo (plo)
I. ATN-II<.t m!rlmalln!lam-, C4<1+ In portll.JbiJ~r ..plliulo!; ()"'''C)
11. C.pililly and!« P>mot\llu ln1!11!1mrllon anc!lof llin>m-., 04d+ In PTC
Ill. Al1oulll- .e, C4<l+ In f'TC
Chronic anlllloctl-- ,.,_,
04d+, p.-ceol d1Wo1fnuh1Hioll« •hllbodilo, ..,.Illlor dou'*> -ll'e•hdlor
DOII!ubulorCIIDiaiY- memb11110 muhllo>iol1,. arod/of ln1orodlol ~brooM!Jbular
....plly. .d/orlbiOI.Illr-.t""l thldcln!ngln •~ta~lol&
a llml•lht ella- •swpl&:lcus" ftlf""""' TGiklwfllled r<ljeclbn
Thl& ...,....,.. uNd.m., no lnllmala-1& p,_nt, but lllor•a1'6fad ofblb~ll Ql, 12, «
13) 101111 mlnllf ln1Dmlllollnflomtnlltion (10 Of II)"' ln1Drlllllol!nllomrn<11on will mill tubuli>
lUl.tl <4, 12•4-10, G;e: 10intoeokh..,lt,m~pertobularo,_-.
4. N.ob T....a.l!lfdo1ed relodbo
'l'fl>oo (plo)
lA. SfVII!aonl 1-llnllllmDI(>2S%!>'~1,12orla)and!Odai..-111A
blbullla (12)
18. _,lloant ln1Dmlllollnfllrwllon (>20% pnndrjmo.l2« G) •nd fad of -..tllb!Jlll&
!13)
IIA. Mid'> 111Cdemeln1imo10Mtll4 Ml
liB. s - . 1 - •"'ltii<Om1Jflst\& >25% ollte klmlnal"""' (.:!)
Ill. 'IIINmurol-ltllftlditlfanonallbo!nold ""'•IIO•rod .-otmedlal.srnooth m.-
CIIIIIwttll .....,poft)1ngl.lmplloq1lc !nlllmrnlllon (v3)
Chronic T.....modiii1Dd r$CIIOn
~blooll.- · .,_ol
Chronic aJoa7of\ artcrlo!oallr.r (u!crlellnllmolfllmibwttll monon....., ... lnfl-ln
S. lnllll11lllol ~blooll 01\dtobular o1roi>IIY !IFTAl, 110 Oltlencao!OIIOdllc eti>e'l

Ill. s-.IFTA (>SI)'Jj, o f - - . )
6. otllao en,_ nd -redID bo d&a to Nljocllon_. . -d11ml:
scored based on the degree of intimal inflammation as follows: mild (v1;

<25% luminal diameter), moderate (v2; >25% luminal diameter), and
severe (v3; transmural ± fibrinoid necrosis). The presence of even mild
endotheliitis is significant and warrants a diagnosis of at least grade llA
rejection (see Table 19.2).
2. Acute humoral (antibody-mediated) rejection may occur shortly after implan-
tation, or months to years following transplantation. At present, the diag-
nosis requires at least two of the following: circulating anti-donor specific
antibodies, diffuse C4d+ immunostaining of peritubular capillaries, and
morphologic evidence of acute tissue injury (ATN, peritubular capillaritis,
glomerulitis, thromboses, and/or arteritis) (Am] Transplant. 2010;10:464).
The earliest feature of acute antibody-mediated rejection (AMR) may be
ATN, but as the lesions develop, peritubular capillaritis becomes promi-
nent. Peritubular capillaritis is defined by the BANFF 2007 working group
as dilation of peritubular capillaries by marginating inflammatory cells (Am
] Transplant. 2008;8:753). A minimum of 10% of peritubular capillaries
must be involved (normally, peritubular capillaries contain no more than
two mononuclear cells), and the composition (neutrophils, monocytes, etc.)
and the extent (focal [<50%] vs. diffuse [>50%]) of peritubular capillaritis
should be mentioned in the pathology report (e-Fig. 19.21).
C4d immunostaining is another important feature of acute AMR. Depo-
sition of C4d in peritubular capillaries is used as a surrogate marker for
anti-donor antibody activity within the allograft, and IF is considered the
gold standard for C4d evaluation (Kidney Int. 1993;44:411). Normal kid-
neys show diffuse glomerular C4d+, acting as an internal control. Diffuse
C4d staining is defined as bright, linear peritubular capillary wall staining
involving >50% of either cortical or medullary capillaries (e-Fig. 19.20).
Focal (10% to 50%) C4d+ is also clinically significant (Am] Transplant.
2009;9:812).
3. Chronic rejection. Repeated graft injury from episodes of cellular rejection
may manifest as nonspecific tubulointerstitial fibrosis (IFTA) or glomeru-
lar fibrosis (BANFF category 5). The only histological lesion highly suspi-
cious of chronic cellular rejection is transplant arteriopathy (e-Fig. 19.22),
defined as fibrointimal hyperplasia (thickening of the intima) (Kidney Int.
1999;55:713). Elastin staining may help to distinguish transplant arteriopa-
thy from arterial thickening due to HTN since the stain will highlight intimal
proliferation with multilamination in HTN, a finding absent in transplant
arteriopathy.
4. Chronic antibody-mediated rejection. GBM duplication, interstitial fibrosis,
tubular atrophy, arterial intimal thickening, serum donor-specific antibodies,
and peritubular capillary C4d reactivity are all features of chronic AMR. At
least two of these features are required for the diagnosis (Am ] Transplant.
2010;10:464). GBM duplication is believed to be the most specific indicator
of chronic AMR, although this finding may be seen in other chronic kidney
diseases such as lupus. EM is a valuable aid in diagnosing chronic rejection;
GBM and/or peritubular capillary basement membrane multi-lamellation is
highly characteristic.
C. Recurrent glomerular disease occurs in up to 36% of cases with a mean inter-
val of 5 years, and is currently the third leading cause of graft loss. The most
common recurrent glomerular diseases are focal segmental glomerulosclerosis
(10% to50%),IgAnephropathy (13% to 46%), MPGNtypell (80% to 100%),
MPGN type I (20% to 25%), membranous GN (10% to 30%), anti-neutrophil
cytoplasmic antibody (ANCA) vasculitis (17%), diabetes (8% to 30%), and
amyloid (5% to 30%). MPGN type II and FSGS have the highest rates of graft
loss at 15% to 30% and 13% to 20%, respectively.
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3. Calcineurin inhibitor (CNI) toxicity. The CNI (e.g., cyclosporine, tacrolimus)

has markedly improved short-term allograft survival but are, however,
nephrotoxic.
a. Patients with acute CNI toxicity typically present with signs of acute renal
failure, which manifests on renal biopsy as isometric vacuolization of the
proximal tubular epithelial cells and/or ATN. The changes are reversible
with decreased exposure to the offending agent. Importantly, isometric
vacuolization and ATN are nonspecific findings requiring clinical corre-
lation. Osmotic nephrosis, for instance, may also produce isometric vac-
uolization.
b. Chronic CNI toxicity, in contrast to acute CNI toxicity, is irreversible
(Transplantation 1989;48:965). Biopsy findings include stripe or skip
interstitial fibrosis and arteriolar hyalinosis (e-Fig. 19.27). Subendothe-
lial beaded arteriolar hyalinosis is particularly characteristic, but since
arteriolar hyalinosis may also occur secondary to HTN or diabetes, clin-
ical correlation is warranted. Tubulointerstitial calcium deposits are also
part of CNI toxicity.
4. Infection. Infectious agents may be encountered, albeit rarely, in the trans-
plant renal biopsy. The opportunistic fungi Candida. Aspergillus. Crypto-
coccus. and Zygomyces account for "'5% of all renal transplant infections
(Transpl Infect Dis. 2001;3:203 ). White blood cell casts, granulomatous
inflammation, and crescentic GN are all seen in association with fungal infec-
tions, typically occurring within 6 months posttransplantation.
Viral infections, most commonly polyoma virus (BK and JC) infections,
may also be encountered. Polyoma virus typically infects distal tubules and
collecting ducts, manifesting histologically with homogenous intranuclear
inclusions, clumped nuclear chromatin, or bubbly inclusions (e-Fig. 19.28).
Cells with these features in urine cytology specimens are known as decoy cells
because of their similarity to malignant cells. CMV infects the renal allograft
but is currently rare due to CMV prophylactic therapy applied as a standard
of practice.
Surgical Diseases of
the Kidney
NORMAL ANATOMY
In the adult, the normal kidney weighs about 115 to 155 g in women and 125 to
170 g in men. Anatomically, the kidneys are composed of an outer cortex and an
inner medulla that has 8 to 18 pyramids. The base of each pyramid is at the corti-
comedullary junction, and the apex forms a papilla where the collecting ducts open
into the renal pelvis. The minor calyces receive the papillae and in turn join to form the
major calyces that are the dilated upper portion of the ureter in the renal pelvis. Histo-
logically, the components of the kidney include glomeruli, tubules, blood vessels, and
interstitium.
Maldevelopment and
Nonneoplastic Cystic Diseases
Johann D. Hertel, Peter A. Humphrey, and Helen Liapis
The developmental abnormalities and benign cystic diseases of the kidney that are most
likely to be seen by the surgical pathologist are presented here.
I. DEVELOPMENT ABNORMALITIES
A. Renal dysplasia is seen in malformed kidneys where there is abnormal differentia-
tion of metanephric elements. Most cases are unilateral and sporadic, but a wide
variety of genetic diseases, malformation syndromes, and chromosomal disor-
ders have been linked to renal dysplasia (Stocker ]T, Dehner LP, Husain AN,
eds. Stocker and Dehner's Pediatric Pathology, 3rd ed. Philadelphia: Wolters
Kluwer/Lippincott Williams and Wilkins, 2011). Bilateral renal dysplasia is less
frequent and is associated with renal failure at birth. Renal dysplasia is a com-
mon cause of an abdominal mass mimicking neoplasia in children < 1 year of age
but can also be diagnosed in older children and adults. There is an association
with congenital urinary tract obstruction in about 50% of cases.
B. Grossly, multicystic dysplasia is characterized by a slightly enlarged kidney, or
a small and irregularly cystic kidney (e-Fig. 20.1)* Aplastic dysplasia is typified
by a small, solid remnant of the kidney. Segmental dysplasia occurs when the
collecting system is duplicated; microscopically, there are immature tubules or
ducts surrounded by collarettes of condensed mesenchyme (e-Fig. 20.2), and
islands of immature-appearing cartilage (e-Fig. 20.3 ), cysts of varying sizes that
have a flattened epithelial lining, and islands of normal glomeruli and renal
tubules between the dysplastic areas. Rare cases of Wilms tumor and renal cell
carcinoma (RCC) have been reported in multicystic dysplastic kidneys.
344
Chapter 20 • Surgical Diseases of the Kidney I 34 5
II. NONNEOPLASTIC CYSTIC DISEASES

A. Autosomal dominant polycystic kidney disease {ADPKD) is an autosomal dominant
disease with complete penetrance but highly variable expressivity. The disease
results from a defective copy of the PKD1 gene in 85% to 90% of cases (on
chromosome 16p13.3) or PKD2 gene in 10% to 15% of cases (on chromosome
4q22.1). The genes encode polycystin-1 and polycystin-2, respectively, the loss
of which causes failure to appropriately assemble cilia in the renal tubules. While
the defect is present in utero and initiates cyst formation, a second mutation is
required for cyst enlargement U Am Soc Nephrol. 2007;18:1374). The disease
typically manifests in adulthood (mean age of onset 30 years), although there
are ADPKD cases manifesting in the neonatal period which usually present with
glomerular cysts (Arch Pathol Lab Med. 2010;134:583). Clinically, ADPKD
presents with renal failure, hypertension, hematuria, and flank pain. ADPKD is
a systemic disease; the connective tissue abnormality is also associated with liver
and pancreatic cysts, intracranial aneurysms, and mitral or aortic insufficiency.
Macroscopically, the involved kidney is dramatically enlarged with loss of
its reniform structure (e-Fig. 20.4). About 1% to 3% of nephrons are affected
by cysts which are usually large and oval in shape, distributed throughout the
medulla and cortex, and involve any part of the nephron. The cysts are typically
sac-like structures containing fluid ranging from clear and yellow to brown and
turbid. The cysts vary in size from a few millimeters to several centimeters in max-
imal dimension. Microscopically, the cysts are lined with columna.r; cuboidal, or
flattened epithelium (e-Fig. 20.5) surrounded by a thickened basement membrane
laye.r; and micropapillary hyperplasia (e-Fig. 20.6) and small intracystic polyps
may be present. Some studies have suggested an association between ADPKD
and RCC but none has shown a definitive increased risk of malignancy (Clin I Am
Soc Nephrol. 2009;4:1998). In the end stage, the kidney is hugely enlarged and
shows severe interstitial fibrosis, chronic inflammation, and severely thickened
vessels, although the glomeruli appear relatively unaffected.
B. Autosomal recessive polycystic kidney disease {ARPKD) is an autosomal recessive
disease with complete penetrance due to mutations in both copies of the PKHD1
gene encoding for fibrocystin. Fibrocystin is a transmembrane protein expressed
in cilia in many organs including the kidney, liver, and pancreas; how precisely
fibrocystin mutations cause ciliary dysfunction and subsequently cyst formation
is still under investigation (since the mutant proteins causing cyst formation in
ADPKD and ARPKD are localized to cilia, the proposal to classify cystic kid-
ney diseases as ciliopathies is currently favored by some investigators). ARPKD
usually develops in utero causing oligohydramnios and in utero renal failure,
but it may present later in childhood or adulthood (Clin I Am Soc Nephrol.
2010;5:972); in any event, progressive renal failure leads to end-stage renal dis-
ease during the first decade of life. Pulmonary hypoplasia secondary to oligo-
hydramnios is a common cause of mortality in the perinatal period. ARPKD is
also associated with congenital hepatic fibrosis, and the pancreas can also be
involved resulting in pancreatic failure. Macroscopically (e-Fig. 20.7), the kid-
neys are symmetrically enlarged and generally maintain their reniform appear-
ance. The kidney has a sponge appearance caused by innumerable small, smooth
lined cysts throughout the cortex and medulla. The cysts are typically cylindri-
cal (as opposed to the spherical cysts seen in ADPKD) and about 1 mm to 2
mm in size; fusiform dilatations of the collecting ducts run radially through the
cortex and medulla. Microscopically, the kidneys show ectatic dilated collecting
ducts (e-Fig. 20.8). The cysts are generally lined by a single cell layer of cuboidal
epithelium, although areas of hyperplasia may be seen.
C. Medullary sponge kidney is usually asymptomatic unless complicated by urinary
tract infection, nephrolithiasis, or hematuria. The condition is typically detected
radiologically during examination of an adult for stones. The kidney is not
enlarged. Microscopically, there is ectasia of the papillary collecting ducts in

the renal medulla, with intraluminal microliths commonly observed.
D. Medullary cystic kidney disease (MCKD) complex is an autosomal dominant adult
onset kidney disease. The specific gene abnormality has not been identified but
has been mapped to 1q22 for MCD type I and to 16p13 for MCD type II by
gene linkage analysis. Both types have a similar phenotype, although MCD type
II has an earlier age of onset (fourth decade) than MCD type I (seventh decade).
Both MCD I and IT present as progressive nephropathy and progress to end-
stage renal disease. Macroscopically, the kidneys appear normal to shrunken
but maintain their reniform appearance. On sectioning, numerous small cor-
ticomedullary cysts are seen. Microscopically, the kidney shows focal tubular
atrophy and dilatation with cyst formation caused by disintegration of the tubu-
lar basement membrane. The cysts are typically at the corticomedullary junction,
few in number, small, and lined by columnar, transitional, or metaplastic squa-
mous epithelium. The renal parenchyma reveals a modest interstitial lymphocytic
infiltrate and fibrosis indistinguishable from other tubulointerstitial processes.
E. Nephronophthisis is a hereditary disease of childhood and early adulthood with
gross and microscopic pathology indistinguishable from MCD. Some investiga-
tors view juvenile nephronophthisis and MCD as part of the same disease com-
plex. The disease is caused by mutations in at least 12 different nephrocystins-a
mutation in any of which leads to a loss of cilia function and development of
cystic renal disease. Grossly and microscopically, the findings are nonspecific and
similar to those found in MCD.
F. Simple renal cysts, or retention cysts, are asymptomatic incidental findings seen
on abdominal imaging. Simple renal cysts affect approximately 7% to 10% of the
general population and are increasingly common with advancing age. Surgical
pathologists typically see tissue only if there is suspicion of malignancy or if there
are symptoms (such as pain, hematuria, or infection). On imaging and by gross
examination, the cysts are usually unilateral, and unilocular, but can be bilateral
and multiple. The size of the cysts can vary dramatically. Microscopically, the
cysts have a flattened epithelial lining.
G. Acquired cystic kidney disease occurs in the setting of chronic hemodialysis. The
cysts are generally asymptomatic and are seen as incidental findings on abdomi-
nal imaging. On gross examination the kidney is normal to shrunken and shows
multiple smooth-lined cysts filled with clear fluid. Microscopically, the cysts are
generally lined by flattened cuboidal epithelium (e-Fig. 20.9), although hyper-
plastic and even dysplastic epithelium has been reported; in this setting, however,
no criteria presently exist for separating a region of hyperplastic epithelium from
a microscopic papillary RCC.
H. Cysts can be associated with malformation syndromes including tuberous sclero-
sis and von Hippel-Lindau (VHL) disease (see section on renal cell carcinoma).
Tuberous sclerosis complex is autosomal dominant and caused by mutations in
TSC1 (on chromosome 9q; encodes for hamartin) or TSC2 (on chromosome
16p; encodes for tuberin). Renal angiomyolipomas (discussed later) and cortical
cysts are common. These distinctive cysts vary in size and are lined by hyper-
plastic epithelium with eosinophilic cytoplasm, with multilayering and papillary
growth seen.
SUGGESTED READINGS
Agarwal MM, Hemal AK. Surgical management of renal cystic disease. Curr Urol Rep. 2011;12:
3-10.
Bonsib SM. The classification of renal cystic diseases and other congenital malformations of the
kidney and urinary tract. Arch Pathol Lab Med. 2010;134:554-568.
Eknoyan G. A clinical view of simple and complex renal cysts. JAm Soc Nephrol. 2009;20:1874-
1876.
Pediatric Renal Neoplasms
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tumors; howevet; intraoperative biopsies are discouraged due to the risk of tumor
spillage unless the diagnosis will alter operative management.
Pediatric renal tumors are often large, friable tumors that bulge beyond the nor-
mal renal contour. The capsule should be carefully examined for sites of rupture
and inked before incised. The initial plane of section is taken to demonstrate the
relationship of tumor to the capsule and renal sinus. At this point, fresh tissue from
each tumor nodule, nephrogenic rests, and normal kidney is snap frozen for proto-
col studies. Tissue may also be taken for cytogenetics, flow cytometry, and electron
microscopy, depending on clinical history. Following initial cuts to facilitate fix-
ation, overnight fixation is recommended before histologic sampling to decrease
tumor friability and capsule retraction. Tissue for histology should be mainly taken
from the tumor's periphery, as histology of the interface between tumor and renal
parenchyma is often important in identifying the type of tumor. Each separate
tumor nodule should be sampled, with at least one section per centimeter of tumor
diameter. Sections should also demonstrate the relationship of tumor to the capsule
and the renal sinus. The hilum is a common route of tumor spread, necessitating
adequate sampling of this area as well. Ureteral and vascular margins, hilar lymph
nodes, nephrogenic rests, and normal kidney are also sampled. Tumor sections are
mapped, using either a diagram or photograph of the specimen; mapping is nec-
essary for nephroblastoma, as the presence of diffuse versus focal anaplasia alters
type of treatment (Arch Pathol Lab Med. 2003;127:1280).
Ill. DIAGNOSTIC FEATURES OF PEDIATRIC RENAL TUMORS. Pediatric renal neoplasms are
notorious for their variable histologic patterns, which often show overlap between
the various entities; thus, there should be careful examination of the tumor-kidney
interface for pattern of infiltration, which is characteristic for each tumor type.
Correct diagnosis depends on knowledge of both classic and variant histology,
along with adequate sampling and correlation of the microscopic findings with
patient age and other clinical information. While immunohistochemical studies
may be useful for establishing the diagnosis (e.g., for INI1 in rhabdoid tumor) or
for excluding a diagnosis in specific cases, immunohistochemical stains are usu-
ally not needed. Molecular studies can also be used to confirm specific diagnoses
(Table 20.2).
A. Nephroblastoma (Wilms tumor) and nephrogenic rests. Nephroblastoma (Wilms
tumor) accounts for 85% of pediatric renal tumors. Most patients present before
10 years of age, with a peak between 2 and 5 years. Rare examples have been
reported in adults (] Clin Oncol. 2004;22:4500). The tumor is more common
in children of African descent than of other races and is slightly more common
in girls than boys.
Although genetic abnormalities have been identified in only a minority of
nephroblastomas, multiple genetic abnormalities and syndromes are associated
with the tumor. Abnormalities of WT1 on chromosome 11p 13, a gene involved
in renal and gonadal development, occurs in aniridia and genital anomalies
syndrome (WAGR), Denys-Drash, and Frasier syndromes, all associated with a
high risk for nephroblastoma. Molecular alterations of imprinted genes at the
WT2 locus on chromosome 11p15 are associated with Beckwith-Wiedemann
syndrome, which has an increased risk for nephroblastoma. Both WT1 and WT2
locus gene alterations are found in a minority of sporadic tumors. About 1%
of patients with nephroblastoma have a family history of nephroblastoma; two
familial genes, FWT1 and FWT2, have been identified in this setting in which the
disease shows autosomal dominant transmission with variable penetrance (Cu"
Opin Pediatr. 2002;14:5). Other molecular alterations that are associated with
tumor progression or aggressiveness have been identified; TP53 is implicated in
progression to anaplastic nephroblastoma, and loss of heterozygosity of 1 p and
16q is associated with poor prognosis in favorable histology nephroblastoma
(] Clin Oncol. 2005;23:7312 and] Clin Oncol. 2006;24:2352).
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Nephroblastomas are usually solitary masses; however, 10% are multicentric

and 5% are bilateral at presentation (e-Fig. 20.10). The cut surface is typically
pale gray and friable and may be hemorrhagic or cystic. Stromal-predominant
tumors often have a myomatous appearance. The tumor is derived from nephro-
genic blastema and is composed of varying proportions of blastema!, epithelial,
and stromal elements (e-Fig. 20.11). Nephroblastomas have a pushing border
surrounded by a fibrous pseudocapsule; one exception, however, is the diffuse
blastema! type, which infiltrates adjacent renal parenchyma. Other blastema!
types include serpentine, nodular, and basaloid patterns. Epithelial differen-
tiation includes tubular, papillary, and glomeruloid patterns; squamous cell,
mucinous, and neural differentiation can also occur. The stroma may be primi-
tive mesenchyme or show differentiation into skeletal muscle or, less frequently,
smooth muscle, adipose tissue, and cartilage. Tumors with prominent heterolo-
gous elements are sometimes referred to as teratoid Wilms tumor.
Nephroblastomas are designated favorable or unfavorable on the basis of
the presence and distribution of anaplasia rather than type of differentiation.
Anaplasia is defined by the presence of bizarre or multipolar mitotic figures
and large hyperchromatic nuclei (three times the size of other tumor nuclei)
(e-Fig. 20.11F). Focal anaplasia is defined as one or morefocal areas of anaplasia
surrounded by nonanaplastic tumor and limited to the kidney. Anaplasia that
does not meet the definition of focal is considered diffuse anaplasia. Anapla-
sia in a biopsy is also considered diffuse. Approximately 5% of tumors have
diffuse anaplasia, which is the sole criterion for unfavorable histology. The
incidence of anaplasia is higher in posttreatment nephrectomies U Clin Oncol.
2006;24:2352).
Nephrogenic rests are the precursor lesions of nephroblastoma (e-Fig.
20.10B and e-Fig. 20.12). Perilobar nephrogenic rests are located at the periph-
ery of the renal lobule, are well demarcated from adjacent renal parenchyma,
and have predominantly blastema! and epithelial elements. The cells in hyper-
plastic perilobar rests are cytologically identical to malignant tumor cells; how-
ever, rests tend to be ovoid in shape rather than spherical, and they are not
surrounded by a tumor pseudocapsule. Intralobar nephrogenic rests can occur
anywhere in the kidney and have a prominent stromal component that inter-
mixes with normal renal parenchyma. The presence of multiple nephrogenic
rests or nephroblastomas is consistent with nephroblastomatosis and increases
the risk for tumor in the contralateral kidney, especially in infants. In diffuse
hyperplastic perilobar nephroblastomatosis. the renal parenchyma is exten-
sively replaced by nephrogenic tissue (e-Fig. 20.10C). The diagnosis is made
by imaging studies and is treated without biopsy. Tumors that grow despite
chemotherapy are removed with kidney-sparing surgical procedures (Pediatr
Develop Pathol. 2009;12:237). Ectopic nephrogenic rests occur rarely (usually
in the inguinal canal or intrapelvic sacrococcygeal regions) and can be associated
with extrarenal nephroblastoma U Pediatr Surg. 2009;44:e13).
Nephroblastomas with prominent cysts are designated cystic nephroblas-
tomas. Multiloculated renal cysts with microscopic areas of nephroblastoma in
the cyst walls, without any expansile septal mass, are designated partially differ-
entiated cystic nephroblastomas (e-Fig. 20.10D). Multilocular cysts with only
mature elements and no expansile nodules are designated cystic nephroma (CN)
and, in children, are thought to represent end-stage differentiation of nephrob-
lastoma (Semin Diagn Pathol. 1998;15:2, and Cancer. 1989;64:466). Familial
and bilateral CN are associated with DICER1 mutations and pleuropulmonary
blastoma UMed Genet. 2010;47:863).
Following chemotherapy, biopsied or partially resected (usually bilateral)
nephroblastomas are categorized according to histologically observed treat-
ment effect (Pediatr Dev Pathol. 2005;8:320) (summarized in Table 20.3).
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Lt. ~IJCD!Ir. I\I!XI.Ort -111it1W (l()!Xlpk(c _ . . , ;, whi<:h cut tuq;ical
lftlldio11""" bepafo..-.1 w:itbautadJiiticm.dcho:moth.erApy. JW:.psy~
4em ODitlati.q WOuisome histntotPc fu.tu:ra such AI prt.dunin ID '* f1l 'b:l.o..naDaJ
o(...,.,,. or -luio oftc:r iaili&lllU1m:llt ""' nri1che4 to """'' -mitt
dl..c::m.Dcherapeu1ic ngi ram'
t. lllliiiMplrfo 111•11 arc wcl.hl!ffuclllltcd IICI'hto!llu1lc - """''''"'"'
nryiog proportioaf of q>iche&l ....t ~ cdla. ~ ll:!m<Xrt ue
beftigll; ~bod! ~bluci>D14 and p•pillaq> RCC ht.vc beta rcpottcd
iD ll:!mOrt with t:pith<lial elm-.
1. M ••JIIIe llr.aJ III•DFI 001'1) CCC1Il' t!uov,shov;t childl.ood, lro.c-
""""""l'ly ill iDfaa<y (..Pis- 20.13}. lfill»zi<>ally, tlu:se """""' ......, .,..,.
a!dctcd to be """"'h.Wdc .D.ePlu:omu (MN) tdloco....t later) bm uc .D.aW
..,.cbod a dlrtiDct ca1!ty, MST It ......U:, aollluy ....t c:xltl!da out flom.
cht; ~ ~ U .,_ ~·-Pf'!llatcd 111U1 tht.t.ln9' be aolid Ot eyt1i(:.
Tbc 1M""""""' i& li!m .....! " ' f - ~p;.:..!ly, the <IIDIJ>r i& .,..,.
poud.ofspilldled .,..,.u... ..U. w:ich jdisrinct.,.,.plum. U.ader l.,...p....,
mX:!ooa>py, cht; - hu. diwri!l<X Jll>da1ar · - clue ... .Jtem.t-
lllg atel.t d outwud to ccttap adjaOCIIt ~
w cub'alea, ru'll&iagi». cyN"""q>i!belid~ 4IDII ~
hyperplasia. Distinguishing features include concentric cuffs of spindled cells

around blood vessels and renal tubules ("collarettes") and angiodysplasia of
arterioles with epithelioid transformation of smooth muscle. Heterologous
elements such as cartilage and glial tissue are infrequently present (Am] Surg
Pathol. 2000;24:917).
2. Metanephric adenofibromas {MAFs) occur in both children and adults. In chil-
dren, the tumor has been reported as early as 5 months of age. MAFs are
centrally located and contain varying proportions of stroma resembling MST
as well as the epithelial nodules of metanephric adenoma. The peripheral
stromal component merges with normal renal parenchyma in a manner sim-
ilar to intralobar nephrogenic rests (see section on adult tumors) (Am] Surg
Pathol. 2001;25:433).
3. Metanephric adenoma occurs most frequently in adults, although the tumor
has been reported in children as young as 5 years of age (see section on adult
tumors). In children, the main differential diagnosis is epithelial nephrob-
lastoma. Unlike nephroblastoma, however, mitoses are rare, and there is
no pseudocapsule, blastema! component, or vascular invasion. Immunohis-
tochemical stains are helpful in distinguishing metanephric adenoma from
papillary RCC, but not for distinguishing metanephric adenoma from epithe-
lial nephroblastoma.
C. Mesoblastic nephroma {MN) is a distinct neoplasm of infancy which may present
antenatally as fetal hydrops (e-Fig. 20.14). Tumors diagnosed in a child >2
years most likely represent MST, clear cell sarcoma of the kidney (CCSK),
or other tumor. The tumors are unencapsulated, solitary, and they tend to
infiltrate the renal sinus. Microscopically, they are characterized as cellular,
classic, or mixed histology. Classic MN consists of intersecting fascicles of
spindled cells resembling infantile fibromatosis. Long fascicles of tumor cells
extend into the adjacent renal parenchyma, entrapping tubules and glomeruli
and resulting in cysts and epithelial embryonal metaplasia. Dysplastic change
with cartilage may be present in adjacent parenchyma. Consistent molecular
abnormalities have not been identified with classic histology tumors. Cellu-
lar MN is composed of plump cells with vesicular nuclei, variable amounts of
cytoplasm, and increased mitoses. The tumor has a well-demarcated interface
with adjacent renal parenchyma, although it lacks a pseudocapsule. Cellular
MN is histologically similar to infantile fibrosarcoma, both of which have the
t(12;15)(p13;q25) chromosomal translocation that produces the fusion gene
ETV6-NTRK3. Mixed MN contains both cellular and classic areas. MNs are
treated by complete surgical resection, with adjuvant chemotherapy for positive
margins. The recurrence rate is 5% to 10%, and the tumor rarely metastasizes,
usually to lung (Adv Anat Pathol. 2003;10:243).
D. Clear cell sarcoma of kidney {CCSK} represents 4% of pediatric renal tumors. It
is a primitive mesenchymal neoplasm not associated with any consistent chro-
mosomal or genetic abnormality, although t(10;17) and del(14q) are frequently
seen (Arch Pathol Lab Medi. 2007;131:446). CCSK occurs usually from 1 to 4
years of age but is also seen in infants (including stillborns) and rarely in adults.
It is a high-risk neoplasm, with a tendency for metastases (lung, bone, brain,
and soft tissue) and late recurrence. Boys are affected more often than girls.
The tumor is unilateral, solitary, well-circumscribed, and located in the renal
medulla. Its cut surface is typically tan-gray and mucoid, although the surface
may have a firm, whorled appearance (e-Fig. 20.15). Cysts are often present.
Microscopically, areas with classic histology contain nests and cords of uni-
form polygonal to spindled cells that have ovoid nuclei with finely granular to
vesicular chromatin, inconspicuous nucleoli, and indistinct cytoplasm, in a back-
ground of clear extracellular matrix. A delicate, arborizing fibrovascular net-
work separates groups of tumor cells. The tumor interface is well circumscribed
under low-power microscopy; however, a pseudocapsule is not present, and

tumor extends a short distance into adjacent parenchyma, entrapping tubules
that may be cystic or have epithelial metaplasia. Almost all tumors have at least
focal classic histology; however, numerous variant histologies including myxoid,
sclerosing, cellulat; epithelioid, palisading, spindle cell, pericytomatous, stori-
form, and anaplastic patterns may also be present and may predominate (e-Fig.
20.15). CCSK tumor cells are positive for vimentin and negative for epithe-
lial markers. Although immunohistochemical stains may be useful in ruling out
other tumors on an individual basis, there are no specific markers for CCSK
(Am] Surg Pathol. 2004;24:4 and Virchows Arch. 2005;446:566).
E. Rhabdoid tumor of the kidney (RTK) is a highly aggressive malignant neoplasm
of infancy, accounting for 2% of pediatric renal tumors (e-Fig. 20.16). Most
patients present at < 1 year of age with metastatic disease, and almost all patients
present by 3 years....... 10% to 15% of RTKs are associated with rhabdoid tumors
of the central nervous system. Both renal and extrarenal infantile rhabdoid
tumors contain molecular alterations of the hSNF5/INI1 gene at chromosome
22q11. Grossly, the renal tumor is pale tan, unencapsulated, and arises from the
renal medulla. Multicentric or bilateral tumors are considered metastatic lesions.
In areas of classic histology, there are sheets of discohesive tumor cells with
large vesicular nuclei, prominent nucleoli, and abundant eccentric cytoplasm
containing large eosinophilic inclusions. Ultrastructurally, these inclusions con-
sist of whorls of intermediate filaments that are characteristic but not specific
for the tumor. Cells with smaller nuclei and less cytoplasm may sometimes
predominate. Variant histology includes sclerosing, epithelioid, spindled, and
lymphomatoid patterns. Tumor cells have a polyphenotypic immunostaining
pattern, with diffuse vimentin positivity and patchy positivity for other mark-
ers, including epithelial markers. There is absent nuclear staining for INil (Adv
AnatPathol. 2003;10:243,Am] SurgPathol. 1989;1313:439,Am] Surg Pathol.
2004;28:1485).
F. Pediatric renal cell carcinoma {RCCs). Although pediatric RCCs bear a strong
morphologic resemblance to their adult counterparts, they often possess unique
clinical, pathologic, and genetic features that distinguish them from the adult ver-
sions (Adv Anat Pathol. 2003;10:243). The mean age at presentation is between
9 and 10 years, and the sentinel symptoms include a palpable mass, flank and/or
abdominal pain, hematuria, polycythemia, and hypertension. Children tend to
have a slightly better prognosis than adults, primarily due to their presentation
at earlier stages; metastatic disease has a poor prognosis (< 10% 5 year survival)
in both groups.
1. In adults, clear cell RCC is the most common malignant epithelial renal
neoplasm, but in children this tumor is less common and probably occurs
only in patients with von Hippel-Lindau syndrome or another predispos-
ing genetic background. The histopathologic features are identical to those
in adult dear cell RCC (see section on adult neoplasms). Many previously
reported/diagnosed cases of pediatric clear cell RCC are now thought to be
associated with X p 11.2 translocations and therefore likely represent different
entities (discussed later).
2. In children, papillary RCC is the most common malignant epithelial neo-
plasm of the kidney (Am] Surg Pathol. 1999;23:795). The histologic, molec-
ulat; and genetic findings are identical to those of the adult tumors. The tumor
is microscopically composed of a single layer of columnar cells lining a pap-
illary stalk, often with clusters of foamy macrophages, hemosiderin, and
necrosis in the surrounding background. The neoplasm is often surrounded
by a pseudocapsule with an associated lymphoid infiltrate. The tumor cells
are strongly positive for CK7 and EMA expression, which can help differen-
tiate papillary RCC from Wilms tumor and metanephric adenoma.
3. Recently, the category of translocation RCCs has been developed to describe

a group of neoplasms that, despite diversity in their histologic features,
share common molecular and genetic abnormalities (Pediatr Dev Pathol.
2005;8:168, Proc Natl Acad Sci USA. 1996;93:15294, Am] Clin Pathol.
2006;126:349, Med Pediatr Oncol. 1998;31:153). Although a variety of
chromosomal alterations underlie these tumors, the common end result is
a chimeric fusion gene involving a member of the TFE3/MiT gene family,
a group of basic helix-loop-helix transcription factors. Histologically, these
translocation carcinomas exhibit voluminous cells with clear to eosinophilic
cytoplasm arranged in papilla or nests with thin intervening fibrous septa
(e-Fig. 20.17). Psammoma bodies and hyaline nodules are frequently seen.
The tumor cells show variable immunopositivity for cytokeratins and EMA,
as well as CD10, a-methylacyl-coenzyme A racemase (AMACAR), and RCC
marker antigen (Am] Surg Pathol. 2008;32:656). The most common mim-
icker, epithelioid angiomyolipoma, shows positivity for HMB-45 and Melan-
A, which is absent or weakly expressed in the translocation carcinomas.
The most consistent and specific finding in translocation carcinomas is
nuclear positivity for the rearranged TFFJMiT protein. Xp11.2 translocation
RCCs express TFE3, which can be detected by antibodies directed toward
the C-terminal end of the protein, which is retained in the various gene
fusion products. The morphologic similarity of these renal tumors to alve-
olar soft part sarcoma (ASPS) is not surprising as the latter entity also has
a characteristic translocation involving TFE3. In fact, the ASPL-TFE3 rear-
ranged renal carcinomas (which harbor the t(X;1)(p11.2;p34) translocation)
bear the greatest resemblance to ASPS (Am] Pathol. 1997;21:621). Other
TFE3 fusion partners in this group of carcinomas include PRCC (in which
t(X;1)(p11.2;q21) is present) andPSF (in which t(X;1)(p11.2;q25) is present).
Recently, melanotic Xp11.2 translocation tumors bearing the PSF-TFE3
translocation have been described with melanin-containing epithelioid cells
arrayed in nests and sheets separated by prominent vascularized septa; the
tumor cells are immunopositive for HMB45, Melan A, and TFE3 expres-
sion, but immunonegative for expression of S100 protein, cytokeratins, and
renal tubular markers (Am] Surg Pathol. 2009;33:609, Am] Surg Pathol.
2009;33:1894). These melanotic Xp11.2 translocation tumors share mor-
phologic features of melanomas, PEComas, and translocation RCCs, but
appear to be molecularly and immunophenotypically distinct, and may carry
a worse prognoSis.
Another subset of tumors in this category are renal tumors with t(6;11)
translocations that involve the TFEB gene, and that have similar morphology
including large polygonal clear and eosinophilic cells, in nests and acini,
with intervening hyaline material (Am] Surg Pathol. 2005;29:230). The
other members of the TFEIMiT gene family, namely TFEC and MiTF. have
not been implicated in the pathogenesis of translocation RCCs, although
TFEC is preferentially expressed in kidney and small intestine, and MiTF
is involved in melanoma and other neural crest disorders (Nucl Acids Res.
2004;32:2315).
The translocation carcinomas appear to be unique among renal neo-
plasms in expressing not only TFE3 or TFEB, but also cathepsin K, a cysteine
protease upregulated by TFE proteins. Immunohistochemical stains for these
markers appear to be highly specific, but variably sensitive (Am] Surg Pathol.
2010;34:1295, Mod Pathol. 2009;22:1016).
Overall, children with translocation carcinomas appear to have a bet-
ter prognosis than adults, and translocation-positive tumors behave more
aggressively than translocation-negative ones. However, the best predic-
tors of outcome and the true prognosis for pediatric patients are yet to be
determined. Interestingly, all the translocation carcinomas have also been

reported as secondary malignancies in patients with a history of receiving
chemotherapy for other tumors.
4. Renal medullary carcinoma is an extremely rare, highly malignant neoplasm
which occurs only in patients with sickle-cell hemoglobin trait (e-Fig. 20.18)
(Mod Pathol. 2007;20;914). The etiologic relationship between medullary
carcinoma and sickle cell trait is unknown. Patients typically present as
teenagers or young adults, usually at late stages with widely disseminated
disease; survival is measured in weeks or months.
Renal medullary carcinoma grows in an aggressive, infiltrative, and
often sarcomatoid pattern, often entrapping native structures. Intrarenal
spread occurs hematogenously and gives the appearance of multifocallesions.
Necrosis and hemorrhage are prominent. The tumor can be morphologically
heterogeneous with yolk sac-like, cribriform, microcystic, and solid patterns;
it often has rhabdoid cytology. Common mimics include collecting duct car-
cinoma (CDC), endodermal sinus tumor (EST), and RTK; however, CDC
typically occurs in older patients, and EST has a distinguishing immunopro-
file. Like RTK, renal medullary carcinoma shows loss of nuclear hSNF5/INI1
staining, but the immunoprofile of renal medullary carcinoma is otherwise
nonspecific in that the tumor cells usually express cytokeratin and are vari-
ably reactive for carcinoembryonic antigen (CEA) and EMA.
5. Postneuroblastoma RCC is exceedingly rare, occurring in children and
teenagers 3 to 12 years old who have previously been diagnosed with neu-
roblastoma (Pathology. 2003;35:499 and Am I Surg Pathol. 1999;23:772).
Interestingly, postneuroblastoma RCC does not appear to be secondary to
treatment; no common chemotherapeutic or radiotherapeutic regimen has
been implicated, and at least two cases have been reported in patients with
previously untreated, spontaneously resolving stage NS neuroblastoma. The
pathogenesis of this neoplasm therefore remains unclear.
These tumors exhibit morphologic diversity including solid and papil-
lary growth patterns, a characteristic oncocytic cytology, and areas resem-
bling clear cell RCC with occasional psammoma bodies and/or collections
of foamy histiocytes. Immunohistochemically, the tumors are positive for
vimentin, EMA, and cytokeratin CAM 5.2 expression. No specific genetic
aberrations have been identified. Invasive and metastatic behavior is com-
mon.
6. The other renal epithelial neoplasms of adulthood-oncocytoma, chromo-
phobe RCC, and others-have all been rarely reported in children (Pedi-
atr Develop Pathol. 2007;10:125). In addition, clear cell RCC and CDC
have been reported in renal adult allografts in pediatric transplant patients
(Pediatr Transplant. 2008;12:6000, IntI Urol. 2008;15:175). Although the
histopathologic features are identical to those in adults, these entities occur
too infrequently in the pediatric population to ascertain whether the clinical
course is also similar.
G. Other small round cell tumors. Small round cell tumors that are usually
extrarenal but can occur as a primary renal tumor include neuroblastoma,
Ewing sarcoma/primitive neuroectodermal tumor (EWS/PNET), synovial sar-
coma, desmoplastic small round cell tumor (DSRCT), rhabdomyosarcoma
(RMS), and lymphoma. All can be mistaken for one of the relatively more
common pediatric renal tumors.
1. Undifferentiated neuroblastoma and EWS/PNET can mimic blastema! pre-
dominant nephroblastoma, and neural pseudorosettes may resemble primi-
tive tubule formation in nephroblastoma (e-Fig. 20.19). Features favoring
neuroblastoma over nephroblastoma include diffuse infiltration, hemor-
rhage, calcification, and nonoverlapping nuclei with "salt and pepper"
chromatin (Lab Invest. 2003;83:4P). EWS/PNET is distinguished from

nephroblastoma by diffuse infiltration; immunoreactivity for CD99 and
FLit, but nonreactivity for WT1; and by chromosomal translocations involv-
ing the EWS gene detected by fluorescence in situ hybridization (FISH)
or reverse transcription-polymerase chain reaction (RT-PCR) (Am I Surg
Pathol. 2001;25:133, Am I Surg Pathol. 2002;26:320). A novel PUS-ERG
fusion transcript has also been reported in renal EWS/PNET (Cancer Genet
Cytogenet. 2009;194:53).
2. Synovial sarcomas present in older adolescents. In the kidney, they are usually
monophasic and may resemble blastema! nephroblastoma or PNET. Synovial
sarcomas have the t(X;18) translocation resulting in a SYT-SSX fusion tran-
script (Am I Surg Pathol. 2000;24:1087, Urology. 2008;72:716).
3. Desmoplastic small round cell tumor {DSRCT) has been reported as a rare
primary renal tumor in both older children and adults. Primary renal
DSRCT may lack the characteristic desmoplastic reaction and can be con-
fused with other small round cell tumors. The tumor cells have nuclear
positivity for WT1, perinuclear dot-like positivity for vimentin, and vari-
able perinuclear dot-like positivity for desmin and cytokeratin. DSRCTs
have the t(11;22)(p13;q12) translocation resulting in the EWS-WTl fusion
transcript.
4. Rhabdomyosarcoma {RMS) and undifferentiated sarcoma are rare primary
renal neoplasms that can occur from early to late childhood. In a recent
Children's Oncology Group Report, all renal RMS were classified as embry-
onal subtype, the majority with anaplastic features. (Pediatr Blood Cancer.
2008;51:339).
5. Non-Hodgkin lymphoma rarely presents in the kidney as bilateral or multicen-
tric masses. Both B-and T-celllymphomas have been reported in children.
Immunohistochemical markers for lymphoma are used for diagnosis (Pediatr
Blood Cancer. 2007;48:711).
H. Miscellaneous tumors
1. Angiomyolipomas can occur in children, almost always in association with
tuberous sclerosis. Epithelioid angiomyolipoma (pure epithelioid PEComa)
with metastases has been reported in older children and adolescents (Am
I Surg Pathol. 2011;35:161). The main differential diagnosis of epithelioid
angiomyolipoma in children is translocation carcinoma (see section on adult
renal neoplasms).
2. Juxtaglomerular cell tumors have been reported in older children and ado-
lescents (Pediatr Blood Cancer. 2008;50:406) (see section on Adult Renal
Neoplasms).
3. Ossifying renal tumor of infancy is an extremely rare tumor that typically
presents as a calcified abdominal mass in male infants with hematuria. The
tumor is attached to the renal papilla and protrudes into the calyceal lumen
and pelvis. Microscopically, it consists of spindled cells surrounding a par-
tially mineralized osteoid matrix. The tumor is benign, and recurrences have
not been reported (Pediatr Pathol Lab Med. 1995;15:745).
4. Other rare pediatric renal tumors include teratoma U Pediatr Surg.
2010;45:255) and myofibroma (Virchows Arch. 2007;450:231). Renal glo-
mus tumor U Pediatr Surg. 2010;45:e23) and gastrinoma (The Scientific
World I· 2009;9:501) in adolescents have also been described.
IV. PATHOLOGIC STAGING. Most pediatric renal tumors are pathologically staged using
the National Wilms Tumor Study Group (NWTS) staging system. This system
has been recently modified by the COG (Table 20.4). In the current system, any
pretreatment biopsy or intraoperative tumor spillage automatically upstages the
tumor to stage III. Lymph node sampling and triaging of tumor for molecular
studies is required for specific treatment protocols. Pediatric RCCs are best staged
using the tumor, node, metastasis (TNM) system (see Table 20.8).
~ 20 • I
*
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...... 1I It -1ft 70 T..,....nvyCOI'Illlln • YIJ1o1y of hl!tolo!ile footlno,
.,_,,.. ec>llli..,l, 111r01naJ, and - a J
dltloiM!Iotlon
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a 1poc:trum of""""""" and-""
• - be P"""'nl but fal !lla1 d oomplali!-.
o!IIMJIIII'III'IIl'
l.ooo11\on 1;6!1; of111o YloiAe IUmor Oh(ljjld be biosllmol I
1fle IUMOf It >113 ¥\ablo -~
-
lint Jtll••..t 10 Vlll>l&lumor rnusl..,l)lla»113 of tilt tumor mtn
At lllalt213of ll11ulolilo lllnv oonslsb d -ma
011\et 00111~ of IIOI>hrobiMiornoll'lll!' be ltllOOirt In
""llln1 p,po-
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Adult Renal Neoplasms

M•ri~ F. Sem.mo and P<:!ll:r A. Humphrey
L SIOR WMIIImiiN AN~ niiUE SAMPUIIIi. laW 1ilsue II&D1pliag for CWIII>r
iac:lada. porQal &II.C! ndioal acplr!O<tDmiat &ad, ha comm~>llly, .....UC bil>poiat
&lid he .....tic up!,latao.
l. NMdla cara biOPIIJ it ~ io - ~ p~ pri(Jl' to 11.1t
of P"""""""' a&Lui.. lberapi<t .....Z u ..Wair<qii<D.C)' hut ahlatioo or
cryosurgery. It can also be performed if there is a clinical concern for lym-

phoma or metastatic disease. The cores can usually be submitted in 10% for-
malin, with generation of H&E-stained slides. If lymphoma is in the differential
diagnosis, additional core(s) should be submitted fresh for lymphoma workup.
Histopathologic diagnosis of needle core biopsy tissue from renal masses is
highly accurate in establishing a malignant diagnosis (] Urol. 2008;180:2333)
and in renal tumor typing. FISH (B]U Int. 2007;99:290), cytogenetics, compar-
ative genomic hybridization, and/or immunohistochemistry (Arch Pathol Lab
Med. 2011;135:92) rnay aid in typing. RCC Fuhrman grade in needle core tissue
is often lower than grade in the whole tumor (Urology. 2010;76:610).
B. Partial nephrectomy. The surgical resection margin is inked, the specimen is
serially sectioned, and the distance to the resection margin is recorded. Sec-
tions demonstrating the relationship of the mass to the surgical resection mar-
gin and perirenal fat are taken. Intraoperative consultation for gross or frozen
section examination of the margin is often requested (Arch Pathol Lab Med.
2005;129:1505).
C. Radical nephrectomy. A radical nephrectomy includes the kidney; a portion of
the ureter, renal vein, and artery; perinephric fat; and Gerota's fascia. The
adrenal gland may also be present. Intraoperative consultation is most com-
monly requested to grossly confirm presence of a renal or pelvic mass in the
nephrectomy specimen (Arch Pathol Lab Med. 2005;129:1586).
After weighing the entire specimen, the renal hilum is examined to identify
the ureter, renal vein, and artery, and cross sections of these margins are taken
(Fig. 20.1). The renal vein and ureter are opened longitudinally. Areas suspicious
Tumor and IF~-"""""¥:-- Kidney, tumor

radial margin and adrenal gland
Tumor,
cortex and Tumor and renal
perirenal fat ~~-:---=--!=----pelvis ± renal sinus
to include
radial margin
or Gerota's
fascia Transverse
section of
hilar vessels
·- - - Section of distal
l==~ ureter
Normal Kidney and Pelvis

Figure 20.1 Sampling of radical nephrectomy specimen for adult renal tumors. Sections
through renal masses should demonstratt: relationship of mass to capsule, peripheral
fat, renal parenchyma, and renal pelvis. (Modified from: Schmidt WA. Principles and
Techniques of Surgical Pathology. Menlo Park, CA: Addison-Wesley; 1983.)
Chapter 20 • Surgical Diseases of the Kidney I 3! 9
T2b
···r··- -
>10cm
_ _ L_ _ _
Figure 20.2 RCC pathologic primary tumor (pT) stages. (Modified from: Edge SB, Byrd DR,
Compton CC, Fritz AG, Greene FL, Trotti A, eds. AJCC Cancer Staging Manual. 7th ed. New
York, NY: Springer; 2010.)
for tumor involvement of the perirenal soft tissue are inked selectively. Occa-
sionally, lymph nodes are found in the hilar region.
The kidney is then sectioned sagittally. Tumor size, location (upper or lower
pole, cortex, or medulla), involvement of calyceal or pelvic mucosa, invasion
of the capsule or perirenal soft tissue, and involvement of the adrenal gland
are recorded. The uninvolved parenchyma is also examined: color, cortical
thickness, additional focal lesions, and renal pelvis are described. One section
per centimeter of tumor, demonstrating its relationship to adjacent capsule,
renal parenchyma, and pelvis are submitted, as well as sections of any addi-
tional lesions, uninvolved renal parenchyma, and adrenal gland (Fig. 20.2).
Nephroureterectomy for urothelial carcinomas is described in Chapter 21.
II. DIAGNOSTIC FEATURES OF COMMON TUMORS OF THE ADULT KIDNEY
A. Carcinoma. RCC arises from the epithelium of the renal tubules and represents
.....90% of all renal malignancies in adults.
1. Risk factors. The most important risk factor is tobacco smoking. Additional
risk factors include obesity, hypertension, unopposed estrogens, and expo-
sure to arsenic, asbestos, cadmium, organic solvents, pesticides, and fungal
toxins. Patients with tuberous sclerosis, chronic renal failure, and acquired
cystic disease of the kidney have an increased incidence of RCC. About 5% of
patients with acquired cystic disease of the kidney develop renal cell tumors
(see section on Acquired Cystic Disease-associated Renal Cell Carcinoma).
Most RCCs are sporadic, although there are several inherited cancer syn-
dromes that affect the kidney (N Eng J Med. 2005;353:2477), as follows.
a. Yon Hippel4.indau {YHL) disease is inherited in an autosomal dominant
manner and is characterized by hemangioblastomas of the cerebellum and
retina, clear cell RCC, pheochromocytoma, pancreatic cysts, and inner ear
tumors. Typically, renal tumors are multiple and bilateral (e-Fig. 20.20).
Numerous renal cysts, lined by neoplastic clear cells (e-Fig. 20.21), are
also characteristic. This disease is caused by germline mutations of the
VHL tumor suppressor gene on chromosome 3p25.3.
b. Hereditary papillary RCC is also inherited in an autosomal dominant manner

and is characterized by multiple, bilateral papillary RCCs with a type I-
like papillary or tubulopapillary architecture. The disease is caused by
activating mutations of the MET oncogene on chromosome 7q31.
c. Hereditary leiomyomatosis and renal cell cancer is an autosomal dominant
disease caused by mutations in the fumarate hydratase gene. It is character-
ized by benign leiomyomas of the skin and uterus. Patients are predisposed
to RCC and uterine leiomyosarcoma. Tumors have papillary type ll large
eosinophilic cells with large nuclei and nucleoli.
d. Birt-Hogg-Dube syndrome is characterized by cutaneous fibrofolliculo-
mas, trichodiscomas, and acrochordons. Multiple and bilateral diverse
types of renal tumors are present; the chromophobe type is most com-
mon, but oncocytoma, papillary, clear cell, and "hybrid" chromophobe-
oncocytic (e-Fig. 20.22) tumors can also be seen (Arch Pathol Lab Med.
2006;130:1867). The syndrome is autosomal dominant with incomplete
penetrance. The responsible gene, folliculin, is located on chromosome
17p 11.2.
e. Constitutional chromosome 3 translocations are related to an increased risk
of RCC. Tumors are typically of the dear cell type.
f. Succinate dehydrogenase germline mutations in the gene encoding the
B subunit (SDHB) result in a 15% lifetime risk of RCC development.
Several tumor types have been seen, including clear cell carcinoma, pap-
illary carcinoma, and oncocytoma.
2. Clinical diagnosis. Hematuria, pain, and a flank mass is the classical triad
of presenting symptoms, but in North America most renal tumors are now
detected as incidental findings by radiological studies. Other presentations
include weight loss, anorexia, fever, hypercalcemia, erythrocytosis, hyperten-
sion, gynecomastia, anemia, and hepatosplenomegaly. Radiological studies,
especially ultrasound and computed tomography (CT), are useful for detec-
tion and characterization of renal masses, but up to 15% of renal masses
thought to be malignant by radiology are histologically benign, typically
oncocytomas or angiomyolipomas.
3. Histologic typing and diagnosis of renal epithelial malignancies. The 2004
World Health Organization (WHO) classification of neoplasms of the kid-
ney is given in Table 20.5. Recently recognized additional renal carcinoma
types also exist (Mod Pathol. 2009;22:52), including tubulocystic carci-
noma, acquired cystic disease-associated RCC, dear cell papillary RCC, and
thyroid-like follicular carcinoma. Most renal neoplasms can be typed on the
basis of examination of H&E-stained slides; immunohistochemistry is most
often used to confirm a diagnosis of metastatic RCC, although in a minority
of cases, it can be useful in typing (Table 20.6).
a. Clear cell RCC is the most common histologic type. Grossly, these tumors
often have a variegated appearance (e-Fig. 20.23). Bright yellow areas (e-
Fig. 20.24) are due to the high lipid content of the cells, whereas red and
white-yellow regions are due to hemorrhage, fibrosis, and necrosis. Calcifi-
cation and cystic change are fairly common. The cysts can be multilocular
(e-Fig. 20.25) or secondary to necrosis. Histologic sections show round or
polygonal cells with clear or eosinophilic cytoplasm and centrally located
nuclei. The cells are arranged in nests within a fine vascular network (e-
Fig. 20.26). Tumor cells react with antibodies to low-molecular-weight
keratins, RCC marker (RCC Ma), vimentin, epithelial membrane anti-
gen, carbonic anhydrase IX, and CD10. They are usually negative for
expression of CD117, kidney-specific cadherin, and parvalbumin (Arch
Pathol Lab Med. 2011;135:92). Clear cell RCCs are associated with 3p
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has been proposed that multilocular cystic RCC is a subtype of dear cell
RCC (Mod Pathol. 2010;23:931). No recurrences or metastasis have been
reported for this tumor type.
c. Papillary RCC comprises about 10% of RCCs. Grossly, the tumor can be
cystic, hemorrhagic, and necrotic, with a fibrous pseudocapsule (e-Fig.
20.33). Multifocality and bilaterality are more common than for other
RCCs. Microscopically, there is a papillary or tubulopapillary architecture
composed of papillae with thin fibrovascular cores which may harbor
aggregates of foamy macrophages (e-Fig. 20.34) and cholesterol crystals.
There are two types of papillary RCC. Type I tumors have papillae
lined by a single layer of cuboidal cells with scant cytoplasm (e-Fig. 20.34)
that has a basophilic quality. Type II tumors have taller cells with more
abundant eosinophilic cytoplasm, higher nuclear grade, and pseudostrat-
ified nuclei (e-Fig. 20.35). Fuhrman grading may be applied (Am] Clin
Pathol. 2002;118:877), but nucleolar grading may also be useful (Am]
Surg Pathol. 2006;30:1091). Type I papillary RCC is often positive for
vimentin, keratins detected by AE1/AE3 antibodies, CK7, AMACR,
and RCC Ma; immunostains that are usually negative include CD117,
kidney-specific cadherin, and parvalbumin. Type II RCCs have a vari-
able immunophenotype (Arch Pathol Lab Med. 2011;135:92). Geneti-
cally, there is characteristic trisomy of chromosomes 7 and 17, with loss
of chromosome 4, but chromosomal analysis is not used diagnostically.
Papillary RCC, especially type I, has a better outcome than clear cell RCC.
d. Chromophobe RCC accounts for ""5% of RCCs. Grossly, chromophobe
RCCs are well circumscribed, solid, and beige to light brown (e-Fig.
20.36). Tumors are composed of pale polygonal cells with prominent
cell borders, often wrinkled nuclear membranes, and perinuclear halos
(e-Fig. 20.37). The cytoplasm can be finely granular or flocculent (e-Fig.
20.38) to eosinophilic (e-Fig. 20.39). The use of Fuhrman grading is con-
troversial (Am] Surg Pathol. 2007;31:957; Mod Pathol. 2009;22:524).
Diffuse cytoplasmic positivity with Hale's colloidal iron is characteristic,
and these tumors show extensive aneusomy with loss of chromosomes 1,
2, 6, 10, 13, 17, and 21 which can be detected by FISH (Mod Pathol.
2005;18:320).
Hale's colloidal iron and FISH can be used to address the differential
diagnosis of the eosinophilic variant of chromophobe RCC versus onco-
cytoma. For the differential diagnosis with clear cell and papillary RCC,
immunostains can be helpful, although H&E slides are typically diagnos-
tic; immunopositivity for kidney-specific cadherin, parvalbumin, CD117,
EMA, keratins (detected by AE1/AE3 antibodies), and CK7 is typical,
with negative immunostains for vimentin, carbonic anhydrase IX, and
AMACR (Arch Pathol Lab Med. 2011;135:92). Chromophobe RCC has
a much better prognosis than clear cell RCC.
e. Collecting duct carcinoma {CDC) is thought to arise from the collecting
ducts of Bellini. It accounts for < 1% of renal cell tumors. These tumors
are centered in the medulla, have tubular or tubulopapillary architecture,
and have a surrounding desmoplastic reaction (e-Fig. 20.40). The tumor
cells are typically of high nuclear grade and can have hobnail cytology. Use
of immunostains in diagnosis has limitations; when needed, a-methylacyl
coenzyme A racemase (AMACR), CD10, and RCC Ma negativity would
favor CDC over papillary RCC, which figures prominently in the differen-
tial diagnosis, and which would usually be positive for these three markers
(Semin Diag Pathol. 2005;22:51). Immunopositivity for PAX2 and PAX8,
with negative immunostains for p63 and uroplakin Ill, would favor CDC
over urothelial carcinoma. Most tumors present at an advanced stage with

metastatic disease. The prognosis is poor.
f. Renal medullary carcinoma is very rare. Tumors in adults are identical to
those in the pediatric age group as discussed earlier.
g. Renal carcinomas associated with Xp 11.2 trans locations are defined by vari-
ous translocations affecting chromosome Xp11.2, resulting in gene fusions
involving the TFE3 gene. Although rare in adults, they are histologically
identical to those in the pediatric age group as discussed earlier.
h. RCC associated with neuroblastoma is exceedingly rare and occurs in long-
term survivors of childhood neuroblastoma, as discussed earlier.
i. Mucinous tubular and spindle cell carcinoma is an uncommon, pre-
dominantly low-grade renal epithelial neoplasm that is grossly well
circumscribed with gray or light tan, uniform cut surfaces (e-Fig. 20.41).
Microscopically, elongated tubules are separated by a mucinous stroma
(e-Fig. 20.42). The tumor cells are cuboidal or spindle shaped with low-
grade nuclear features. Uncommonly, necrosis, clear cells papillations,
foamy macrophages, and inflammation can be present (Am] Surg Pathol.
2006;30:1554). Sarcomatoid or high-grade epithelial components are also
uncommon. The immunophenotype shows significant overlap with pap-
illary RCC. Loss of chromosomes 1, 4, 6, 8, 13, and 14 and gains of
chromosomes 7, 11, 16, and 17 are characteristic of these tumors, but
molecular analysis is not currently used diagnostically. This tumor type
generally has a good prognosis.
j. Unclassified RCC is a diagnosis that should be reserved for those tumors
that do not fit into other categories. It is a diagnosis rendered in about
5% of RCC cases. Features that place tumors in this group include the
following: a combination of different histologic types, mucin production,
sarcomatoid appearance, presence of epithelial and stromal elements, and
nonidentifiable patterns. This designation is linked to a worse prognosis
than dear cell RCC (B]U Int. 2007;100:802).
Sarcomatoid change (e-Fig. 20.43, e-Fig. 20.44) can be associated with
a specific type of carcinoma or can overgrow the preexisting carcinoma
type and exist in pure form. The percentage of the tumor that is sarco-
matoid should be specified as being less than or greater than 50%. Het-
erologous malignant bone, cartilage, fat, and skeletal muscle or homolo-
gous undifferentiated malignant spindle cells can be seen. Rhabdoid cells
(e-Fig. 20.45) can be found in about 5% of RCC cases, usually dear cell
carcinoma (Am] Surg Pathol. 2000;24:1329-1338). Both sarcomatoid
and rhabdoid features are associated with a poor prognosis.
k. Tubulocystic carcinoma is very uncommon and is of low malignant poten-
tial (Am] Surg Pathol. 2009;33:384 ). Grossly, the tumor is usually solitary
and circumscribed with a spongy "bubblewrap"-like cut surface. Micro-
scopically, sections show tightly packed tubules and cysts up to a few mil-
limeters in diameter (e-Fig. 20.46), separated by fibrous stroma; the lining
epithelium has eosinophilic or amphophilic cytoplasm and large promi-
nent nucleoli, and hobnail type cells may be seen. Most cases are posi-
tive for CK7, CK19, parvalbumin, CD10, AMACR, and kidney-specific
cadherin by immunohistochemistry. Gene expression profiling shows a
distinctive signature similar to but not identical to papillary RCC, with
no overlap with CDC. Most cases are pT1 and only a few patients have
developed metastatic disease.
I. Acquired cystic disease-associated RCC can be seen in patients with end-
stage renal disease, especially after long-term dialysis (Am] Surg Pathol.
2006;30: 141 ). Many RCCs that develop in end-stage renal disease kidneys
are dear cell, papillary, and chromophobe RCCs of usual type. However,
a unique carcinoma, termed acquired cystic disease-associated RCC, is

observed only in end-stage renal disease and acquired cystic disease,
and is characterized by microcystic and cribrifonn/sieve-like architecture
(e-Fig. 20.47), eosinophilic cytoplasm with Fuhrman grade 3 nucleoli,
and frequent intratumoral oxalate crystals. Additional architectural pat-
terns include solid acinar, solid sheet-like, papillary, and macrocystic.
Immunostains show positivity for AMACR, vinculin, and parvalbumin,
with no reactivity for CK7. Another histologic type of carcinoma found in
end-stage kidneys is dear cell papillary RCC (discussed later). Clinically,
acquired cystic disease-associated RCC appears to be more aggressive than
other tumor types in end-stage renal disease.
m. Clear cell papillary RCC can arise in end-stage renal disease but also in
kidneys without end stage features. One group, also called dear cell tubu-
lopapillary RCC (Am J Surg Pathol. 2010;34:1608) or clear cell papillary
and cystic RCC (Mod Pathol. 2009;22:S2), is typically cystic with a fibrous
capsule and is composed of cells with low nuclear grade (Fuhrman 2)
and a cystic, tubuloacinar, and/or papillary architecture (e-Fig. 20.48).
The papillae are prominent and lined by cells with variable amounts of
cleared cytoplasm. A characteristic finding is linear arrangement of the
nuclei in the center or apical portions of the cytoplasm. The immunophe-
notype and molecular genetic profiles are distinct from papillary and clear
cell RCC: CK7 and carbonic anhydrase 9 immunostains are positive, and
immunostains for AMACR, TFE3, and CD10 are negative. Gains of chro-
mosomes 7 and 17 (typical of papillary RCC) and 3p loss (seen in clear
cell RCC) are not present. Most tumors are low stage at presentation and
limited outcome data suggest an extremely favorable prognosis.
A second group of dear cell papillary RCCs, identified on the basis
of CK7 and AMACR positivity, is termed papillary RCC with dear cell
features(] Urol. 2011;185:30). A substantial percentage of these cases has
high nuclear grade, and in this group of tumors the presence of clear cells
is an adverse prognostic indicator compared with conventional clear cell
tubulopapillary RCC.
n. Thyroid-like follicular carcinoma of the kidney is rare, with less than ten
cases reported (Am] Surg Pathol. 2009;33:393). Microscopically, macro-
follicles and microfollides with intraluminal colloid-like secretion are
present (e-Fig. 20.49). A negative TTF-1 immunostain and clinical his-
tory are useful in excluding metastatic thyroid carcinoma.
B. Benign epithelial tumors
1. Papillary adenoma of the kidney is the most common benign epithelial neo-
plasm of renal cells. Grossly, the tumor consists of single or multiple well-
defined white nodules in the renal cortex. The tumors have papillary, tubu-
lar, or tubulopapillary architecture (e-Fig. 20.50), low nuclear grade (e-Fig.
20.51), and measure ~5 mm in diameter. The cytoplasm is scant and non-
cleared, and nuclear grooves can be present. Psammoma bodies and foamy
macrophages are common.
2. Oncocytoma is a benign epithelial neoplasm that comprises ...... 5% of all neo-
plasms of the renal tubular epithelium. The lesion is more frequent in men,
and the peak incidence is during the seventh decade of life. Grossly, the
tumors are well circumscribed and the cut-surface is typically mahogany-
brown (e-Fig. 20.52). A central scar (e-Fig. 20.52 and e-Fig. 20.53) is seen
in up to 33% of cases, generally with larger tumors; such scarring, although
characteristic, is not specific for oncocytoma. Hemorrhage is frequent, but
necrosis is almost always absent.
Microscopically, these tumors are composed of nests and tubules of round
or polygonal cells with granular eosinophilic cytoplasm, round nuclei, and
single central nucleoli (e-Fig. 20.54 ). Scattered larger cells with a higher
nuclear/cytoplasmic ratio and hyperchromatic nuclei can be seen (e-Fig.
20.55). Smaller cells with scant cytoplasm (known as oncoblasts) can also
be noted in some cases (e-Fig. 20.56). A hyalinized or edematous stromal
background is frequent. Microscopic (but not gross) extension into perire-
nal fat and vessels has been described. Rare cases of numerous oncocytic
tumors (oncocytosis) have been described. Chromosomal abnormalities in
oncocytoma include t(5;11) and loss of chromosomes 1 and 14, but assess-
ment for these abnormalities is not usually necessary. No cases of death due
to metastatic disease have been reported.
C. Metanephric tumors (see also the section on pediatric renal neoplasm)
1. Metanephric adenoma occurs in children, and in adults during the fifth and
sixth decades. It is more frequent in women. About 50% of cases are inci-
dental. Tumors are usually well circumscribed, gray to tan to yellow, and
firm or soft. Hemorrhage, necrosis, calcification, and cyst formation are
common. Microscopically, metanephric adenomas are composed of small,
uniform round acini with small lumens (e-Fig. 20.57). The cells are uni-
form with small nuclei and inconspicuous nucleoli. Branching and tubular
configurations are common, as well as a papillary architecture with numer-
ous psammoma bodies. In the differential diagnosis with papillary RCC,
immunostains that are positive for WT1, negative for EMA, and focally pos-
itive for CK7 favor metanephric adenoma.
2. Metanephric adenofibroma is more common in men. Grossly, it is solitary and
partially cystic. Histologically, its cells are similar to those of a metanephric
adenoma but are embedded in a stroma of fibroblast-like spindle cells. Psam-
moma bodies are also common.
D. Mesenchymal tumors
1. Leiomyosarcoma is the most common renal sarcoma; it occurs mainly in adults
and affects women and men equally. They can arise from the renal capsule,
parenchyma, pelvis muscularis, or the renal vein. They are solid, gray-white,
and focally necrotic. Histologically, these tumors are composed of spindle
cells with a fascicular growth pattern (e-Fig. 20.58). Necrosis, nuclear pleo-
morphism, and numerous mitotic figures indicate malignancy. Leiomyosar-
coma is an aggressive tumor with a 5-year survival rate of 29% to 36%, and
most patients die within 1 year of diagnosis. Sites of metastasis include lung,
liver, and bone. Sarcomatoid RCC is in the differential diagnosis and is more
common, and so must be excluded; diffuse desmin and smooth muscle actin
immunoreactivity would favor leiomyosarcoma.
2. Rare primary sarcomas include RMS, angiosarcoma, malignant fibrous histio-
cytoma, chondrosarcoma, low-grade fibromyxoid sarcoma, malignant mes-
enchymoma, and osteosarcoma.
3. Angiomyolipoma is a benign clonal mesenchymal neoplasm tumor com-
posed of thick-walled blood vessels, smooth muscle cells, and adipose tissue
(e-Fig. 20.59). This tumor belongs to the perivascular epithelioid cell tumor
(PEComa) family. The mean age at presentation is 45 to 55 years for patients
without tuberous sclerosis, and 25 to 35 years for patients with tuber-
ous sclerosis. Patients with tuberous sclerosis tend to have multiple, bilat-
eral renal tumors and can have associated pulmonary lymphangioleiomy-
omatosis. Tumors are nonencapsulated, yellow to pink masses (e-Fig. 20.60)
depending on the content of the tissue components. Rarely, angiomyolipo-
mas can extend into the renal vein or the vena cava. Vascular invasion and
lymph node involvement can be present; however, these features are consid-
ered to be evidence of direct extension and multifocality, respectively, rather
than metastatic disease.
Microscopically, the limits between the tumor and the kidney are well
defined. The smooth muscle cells are generally spindled (e-Fig. 20.61) but can
appear round or epithelioid in some cases (e-Fig. 20.62). The smooth muscle
cells often appear to radiate from the outer aspect of the thick-walled, hyalin-
ized blood vessels (e-Fig. 20.63). Nuclear atypia can be present (e-Fig. 20.64).
The amount of fat is variable and can be very focal (e-Fig. 20.62). Uncom-
mon to rare histologic variants include fat-predominant, smooth muscle-
predominant, lymphangioleiomyomatous (e-Fig. 20.65), oncocytoma-like,
sclerosing type, and angiomyolipoma with epithelial cysts (e-Fig. 20.66).
Characteristically, angiomyolipomas coexpress melanocytic markers such
as HMB45 and smooth muscle markers such as smooth muscle actin and
muscle-specific actin. Epithelial markers including cytokeratin are always
negative. Angiomyolipomas can be diagnosed in needle biopsy tissue (e-Fig.
20.67). Classic angiomyolipomas are benign.
4. Epithelioid angiomyolipoma is a potentially malignant mesenchymal neoplasm
that presents more commonly in patients with tuberous sclerosis. It is much
less common than usual angiomyolipoma. Grossly, these tumors are usually
large with tan-gray or hemorrhagic cut surfaces and necrosis. The cells are
epithelioid with abundant granular cytoplasm. Multinucleated cells, nuclear
pleomorphism, mitotic activity, vascular invasion, necrosis (e-Fig. 20.68),
and involvement of perinephric fat can be present. These tumors express
melanocytic markers with variable expression of smooth muscle markers.
These tumors can metastasize to lymph nodes, live.r; lungs, and spine.
5. Leiomyoma is a benign smooth muscle neoplasm that can arise from the
renal capsule, the muscularis of the renal pelvis, or from cortical vascular
smooth muscle. Most are found incidentally. Grossly, they are firm well-
defined masses, although calcification and cysts can be present. Necrosis
should be absent. Leiomyomas are composed of spindled cells arranged in
fascicles, with minimal nuclear pleomorphism and no mitotic activity. They
demonstrate a smooth muscle immunophenotype, with actin and desmin
immunopositivity. So-called capsulomas that look like leiomyomas originat-
ing from the capsule that are HMB-45 positive are thought to be monophasic
leiomyomatous angiomyolipomas.
6. Hemangioma is a benign vascular tumor that presents in young and middle-
aged adults. The tumor is usually unilateral and single, with a red spongy
gross appearance. Microscopically, the lesion is characterized by irregular
blood-filled spaces lined by a single layer of endothelial cells. No mitoses or
nuclear pleomorphism is present.
7. Lymphangiomas are more common in adults and can represent a lymphatic
malformation or can develop secondary to urinary tract infections. Grossly,
they are cystic, encapsulated lesions that can overgrow the entire renal
parenchyma. The cysts are filled with clear fluid and lined by a single layer
of flat endothelium.
8. Juxtaglomerular cell tumors are benign renin-secreting tumors that occur in
younger individuals and are more common in women. Clinically, the tumor
manifests with severe hypertension and hypokalemia. The tumors are solid,
well circumscribed, and composed of sheets of polygonal or spindled cells
with central regular nuclei, well-defined borders, and granular eosinophilic
cytoplasm. Mast cells, hyalinized vessels, and tubular elements are common.
The tumor cells are immunoreactive for renin, actin, vimentin, and CD34.
9. Renomedullary interstitial cell tumors are commonly found during autopsy.
They are present in about 50% of men and women, and frequently they
are multifocal. They are 1 to 5 mm in diamete.r; white or gray, and located
within the renal pyramids. Histologically, they contain small polygonal cells
in a basophilic background. Renal tubules can be entrapped within the tumor

nodules (e-Fig. 20.69).
E. Mixed mesenchymal and epithelial tumors
1. Cystic nephroma is a benign neoplasm that presents after age 30, more com-
monly in women. It is associated with pleuropulmonary blastoma in the
patient or other family members. Grossly, the tumor is well defined, encap-
sulated, and entirely composed of cysts. Solid areas and necrosis are not
present. The cystic change is multiloculated (e-Fig. 20.70), with the fibrous
septa lined by cuboidal epithelium that often assumes a hobnail appearance
(e-Fig. 20.71). The fibrous septa may contain tubules but not blastema or
dear neoplastic cells.
2. Mixed epithelial and stromal tumor predominates in adult perimenopausal
women. A history of estrogen therapy is common. Grossly, the tumor is
composed of solid and cystic areas. Microscopically, the tumor shows tubules
and cysts lined by flattened, to cuboidal, to columnar epithelium. The stro-
mal component is variably cellular and may exhibit myxoid, smooth mus-
cle, ovarian stromal-like, or collagenous features. Fat can be present. The
tumor's behavior is benign. Since it is possible that cystic nephroma and
mixed epithelial and stromal tumor are related, the unifying term renal
epithelial stromal tumor (REST) has been proposed (Am] Surg Pathol. 2007;
31:489).
F. Neuroendocrine tumors
1. Renal carcinoid tumors are very rare and present between the fourth and
seventh decades. There is a tendency for occurrence in horseshoe kidneys,
and there is an association with renal teratomas. Renal carcinoid tumors
are solid, lobulated, and well circumscribed and are histologically similar
to carcinoids in other organs (e-Fig. 20.72) (Am] Surg Pathol. 2007;31:
1539).
2. Neuroendocrine carcinoma, including small cell carcinoma, can rarely arise
within the adult kidney. A primary tumor elsewhere should be clinically and
radiologically excluded. Grossly, the tumor is usually a white, friable, and
necrotic mass. Microscopically, the tumor is composed of small round cells
with hyperchromatic nuclei and inconspicuous nucleoli, arranged in sheets
and trabeculae. Centrally located tumors can be admixed with urothelial
carcinoma. The tumor cells show dotlike cytoplasmic immunostaining with
cytokeratin antibodies and are variably positive for synaptophysin and chro-
mogranin. The prognosis is poor.
3. PN Els are rare primary renal tumors and are discussed in the section on
pediatric renal neoplasm.
4. Primary renal neuroblastoma and paragangliomas/pheochromocytomas are
very rare.
G. Hematopoietic and lymphoid tumors. Primary renal lymphomas usually arise in
transplanted kidneys and are Epstein-Barr virus (EBV)-associated B-celllym-
phoproliferations. Secondary involvement of the kidney by lymphoma is more
common (e-Fig. 20.73 and e-Fig. 20.74). Plasmacytoma can occur as a mani-
festation of disseminated multiple myeloma. Diffuse infiltration of the kidney
secondary to acute leukemias has also been reported.
H. Germ cell tumors include choriocarcinomas and teratomas and are very rare.
I. Metastatic tumors to the kidney include tumors of lung, breast, gastrointesti-
nal tract, pancreas, ovary, and testis and malignant melanoma. Involvement is
usually in the setting of known, widely metastatic disease. Only infrequently
does metastatic tumor mimic a primary renal tumor. Most metastatic masses
are multiple and bilateral.
Ill. HISTOLOGIC GRADING OF RCC should be reported for all RCCs of dear cell and
papillary types, as described earlier and in Table 20.7.
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B. For a core needle biopsy. report the histologic type, histologic grade (Fuhrman
nuclear grade), and any additional pathologic findings (such as inflammation
and glomerular disease).
C. For a nephrectomy, partial or radical, report the tumor site (upper pole, mid-
dle pole, lower pole), focality (unifocal or multifocal), tumor size (largest, if
multiple) in greatest dimension, macroscopic extent of tumor (tumor limited
to kidney, tumor extension into perinephric tissues, tumor extension into renal
sinus, tumor extension beyond Gerota•s fascia, tumor extension into adrenal
gland, tumor extension into major veins [intraluminal, with or without vein
wall invasion], tumor extension into pelvicaliceal system, histologic type, sarco-
matoid features [and percentage if present], histologic grade [Fuhrman nuclear
grade], pathologic TNM stage, and margin status [cannot be assessed, margins
uninvolved by invasive carcinoma, margins involved by invasive carcinoma]).
The site(s) of margin positivity should be specified. For partial nephrectomy
specimens, the renal parenchymal margin, renal capsular margin, and per-
inephric fat margin should be assessed. For radical nephrectomy specimens,
Gerota•s fascial margin, renal vein margin, and ureteral margin should be
addressed. The adrenal gland, if present, should be reported as uninvolved by
tumor, involved by direct invasion, or involved by metastasis. Lymph-vascular
invasion should be designated as absent, present, or indeterminate. Additional
pathologic findings in nonneoplastic tissue 5 mm or greater from the mass
should be recorded, including glomerular disease (type), tubulointerstitial dis-
ease (type), and vascular disease (type). Optional reporting includes cyst(s) and
tubulopapillary adenoma(s).
VI. NONNEOPLASTIC TUMOROUS CONDITIONS include the maldevelopment and nonneo-
plastic cystic diseases discussed earlier. Another important pseudoneoplastic cate-
gory is inflammatory masses such as xanthogranulomatous pyelonephritis (XGP),
renal malakoplakia, and tuberculosis (TB).
A. xanthogranulomatous pyelonephritis (XGP) is a subacute to chronic, unilateral
inflammatory process that can form a mass in the kidney mimicking RCC clini-
cally, radiographically, grossly, and histologically. This disease most commonly
occurs in women from the fourth to the sixth decades of life. XGP typically
presents with fever, flank pain, or a tender flank mass and is frequently compli-
cated by nephrolithiasis. Urine cultures show common urinary tract pathogens,
such as Escherichia coli and Proteus mirabilis. in up to 70% of cases. If the
kidney itself is cultured, an organism can be isolated in 95% of cases.
Macroscopically, XGP can either be confined to the kidney or extend into
the surrounding soft tissue. XGP is typically composed of yellow nodules of
varying size replacing the normal renal parenchyma (e-Fig. 20.75); the nod-
ules can range in size from a few millimeters to several centimeters. Micro-
scopically, the nodules are composed of lipid-laden macrophages (e-Fig. 20.76)
that can mimic low-grade dear cell RCC, especially in small tissue samples
such as needle biopsy specimens. The lesion may also contain reactive and even
multinucleated fibroblasts that can mimic a sarcomatoid component in RCC.
However, XGP lacks the vascularity typically seen in RCC, and moreover, XGP
often exhibits neutrophils, lymphocytes, foreign body giant cells, plasma cells,
cholesterol clefts, and microabscesses. If there is concern for RCC, a panel of
immunohistochemical stains can be helpful: the clear cells in XGP will be pos-
itive for CD68 and vimentin, and negative for epithelial markers such as EMA
and pan-cytokeratins.
B. Renal malakoplakia is uncommon but can form masses simulating a primary
renal neoplasm. Most patients are women who often have extrarenal malako-
plakia in the urinary bladder, ureter, or retroperitoneum. The lesion is bilateral
in about one-quarter of cases. Grossly, there may be diffuse multinodular cor-
tical enlargement or a large yellow mass. Abscesses and cystic spaces may also
Chapter 20 • Surgical Diseases of the Kidney I 371
be seen. Microscopically, the hallmark, as in the urinary bladder, is sheets of

macrophages (von Hansemann histiocytes) with intracellular or extracellular
Michaelis-Gutmann bodies (e-Fig. 20.77). Lymphocytes and plasma cells may
also be present, and with time, fibrosis may develop.
C. Renal tuberculosis. The kidney is the most common site of TB in the genitouri-
nary tract. Grossly, miliary TB in the kidney is remarkable for numerous very
small white nodules, mainly in the cortex. In the ulcerative form of renal TB,
caseating necrosis can involve multiple renal pyramids, with papillary necro-
sis, cavitation, and extension of the necrosis into the renal pelvis. In advanced
disease, necrotic nodules can also efface the renal cortex.
Cytopathology of Renal
Neoplasms
Souzan Sanati
I. INTRODUCTION. The role of fine needle aspiration (FNA) in diagnosis of renal masses
is limited in comparison with many other deep-seated organs, due in part to the suc-
cess of imaging techniques in correct classification of most renal masses as benign
or malignant. FNA is therefore usually considered only when imaging studies show
equivocal results. Since most renal cysts with equivocal radiologic findings usually
require extensive tissue sampling, the role of FNA of renal cysts is limited and often
renders negative results; while FNA can be useful when malignant cellular features
are identified, cytologically negative cyst fluid does not exclude a malignant pro-
cess and thus it has been recommended that these specimens should be reported as
nondiagnostic. FNA sampling of pediatric renal masses has not been advocated.
Despite these limitations, FNA of renal masses, under radiologic guidance, can
be used as an alternative to core biopsy and can be useful in evaluating metastatic
tumors, benign masses (eliminating the need for a surgical procedure), masses diag-
nosed in patients who are poor surgical candidates, and in masses occurring in
candidates for a tumor ablation procedure.
When the renal pelvis is involved by tumo.r; cytologic sampling can be per-
formed endoscopically. Since the endoscopic biopsies are usually small, cell block
processing is advocated, particularly in low-grade lesions where the architectural
features are of diagnostic importance. Cytologic evaluation of ureteral catheteri-
zation samples is also useful for detecting high-grade urotheliallesion of the upper
urinary tract(/ Urol. 2000;164:1901). In the case ofrenal abscesses, FNA can have
a therapeutic as well as diagnostic role.
II. CYTOPATHOLOGY OF RENAL NEOPLASMS
A. Benign neoplasms
1. Angiomyolipoma. Mature adipose tissue, smooth muscle, and blood vessels
are the main components of this tumor (e-Fig. 20.78). Identification of adi-
pose tissue in the FNA biopsy is an important clue for diagnosis; however,
most angiomyolipomas subject to cytological sampling have a low fat con-
tent. The smooth muscle component may have an epithelioid appearance
with significant nuclear atypia, and it is important not to misinterpret this
finding as a malignant process. Immunostains for HMB45 and smooth mus-
cle actin are used to differentiate angiomyolipoma from RCC (Acta Cytol.
2006;50:466).
2. Oncocytoma can be solitary or multifocal and usually has a characteristic
radiologic appearance that includes good demarcation, a central scat; and
a density similar to the uninvolved renal parenchyma. Cytologic samples

are cellular and show numerous singly dispersed large cells and round nests
(usually seen on cell blocks) with abundant granular cytoplasm, round nuclei,
and distinct cell borders. The nucleolus can be prominent (similar to nucleoli
in Fuhrman nuclear grade 2 dear cell RCC), but no necrosis or mitoses
are present (e-Fig. 20.79). Binudeation and/or multinudeation are common
(e-Fig. 20.80). The morphology of this tumor has overlapping features with
chromophobe RCC and granular variant of dear cell RCC; it is therefore
prudent to diagnose these lesions as oncocytic neoplasm on cytologic samples
if the round nested architecture is not seen (Cancer. 2001;93:390).
B. Malignant neoplasms
1. Clear cell RCC is the most common renal malignancy. FNA samples are usually
richly cellular and bloody. The neoplastic cells have abundant vacuolated
cytoplasm that may be more noticeable in air-dried, Diff Quik-stained slides
(e-Fig. 20.81). It is not unusual to see naked nuclei from disrupted tumor
cells in the background. When tumor microfragments are available, a rich
capillary network is seen (e-Fig. 20.82). The degree of nuclear abnormality
and the presence of prominent nucleoli are dependent on the nuclear grade
of the tumor. Application of Fuhrman nuclear grade to cytology samples is
appropriate.
The most common pitfalls which result in a false-positive diagnosis
include XGP, in which macrophages can be mistaken for the malignant cells
of dear cell RCC, and contamination with benign cellular elements such
as benign hepatocytes and adrenal cortical cells. Caution when making a
diagnosis of malignancy on sparsely cellular samples will prevent an over-
diagnosis in these situations.
2. Papillary RCC is characterized by papillary structures, foamy macrophages
which usually distend fibrovascular cores on sections of cell block (e-Fig.
20.83), and psammoma bodies. Some tumors may show higher degree of
cytologic atypia (e-Figs. 20.84 and 20.85). Additionally, nuclear pseudoin-
dusions, nuclear grooves, and cytoplasmic hemosiderin can be identified
(Diagn Cytopathol. 2006;334:797).
3. Chromophobe renal cell carcinoma consists of cells with abundant cytoplasm,
well-defined cell borders, and a perinuclear halo. The nuclei show signif-
icant size variation, irregular contours, and hyperchromasia resulting in a
raisinoid appearance-nuclear features that allow for differentiation from
oncocytoma. Nuclear grooves and/or pseudoindusions and necrotic debris
can also be present. The cell block can be quite helpful when it demonstrates a
trabecular growth pattern; a cell block also provides material for appropriate
stains (such as Hale's colloidal iron).
4. Urothelial carcinoma is the most common tumor arising in the renal pelvis. It
is morphologically similar to urothelial carcinomas arising at other sites. The
tumor consists of large elongated cells with dense cytoplasm and occasionally
"cercariform" cells. The cytologic findings vary according to the grade; cells
from low-grade tumors lack nuclear atypia and may have a spindle shape
(e-Fig. 20.86).
5. Metastatic tumors. The kidney is a common site for metastatic tumors. The
most common origin of metastatic tumors to the kidney is lung. Knowl-
edge of a history of malignancy in another organ and appropriate use of
immunohistochemistry (ideally on sections of cell block) are keys to correct
diagnosis.
Renal Pelvis and Ureter
Souzan Sanati, Frances V. White, and
Peter A. Humphrey
I. NORMAL ANATOMY. The upper tract of the urinary collecting system is composed of
the renal calyces, pelves, and ureters. Renal papillae protrude into the minor calyces,
which expand into two or three major calyces, which in turn are outpouchings of
the renal pelvis, a sac-like expansion of the upper ureter.
The mucosa is normally arranged in folds. The urothelium of the renal pelvis
is three to five cell layers thick, and five to seven cell layers thick in the ureter.
The lamina propria is composed of highly vascularized connective tissue without
a muscularis mucosa; it is absent beneath the urothelium lining the renal papillae
and is thinned along the minor calyces. The thickness and amount of muscularis
propria in the collecting system within the renal sinus fat can be variable; ureteral
muscularis propria is composed of interlacing bundles of smooth muscle, without
inner or outer layers (Mills SE, ed. Histology for Pathologists. 2nd ed. Philadelphia:
Lippincott Williams and Wilkins; 2007:839-907).
II. GROSS EXAMINATION AND TISSUE SAMPLING. Tissue samples include ureteroscopic
biopsies, needle biopsies, pyeloplasty specimens, segmental ureterectomy speci-
mens, radical cystectomy/cystoprostatectomy specimens, and radical nephroureter-
ectomy with urinary bladder cuff resection specimens.
A. Ureteroscopic biopsies are entirely submitted. Because these are often minute in
size, one approach to processing is to submit the biopsy sample for cytology cell
block preparation (Urology. 1997;50:117).
B. Needle core biopsies of renal masses, including urothelial carcinoma involving
the kidney, should be completely submitted. For microscopic examination, it is
recommended that three levels on each of three hematoxylin and eosin (H&E)-
stained slides be produced.
C. Pyeloplasty specimens for ureteropelvic junction obstruction consist of a portion
of distal pelvis with a short segment of ureter attached to it. If the specimen is
intact, it will be funnel-shaped, and narrowing, and/or angulation of the ureter
may be evident. Usually, however, the specimen is longitudinally splayed open
by the surgeon and consists of a flat, fan-shaped pelvis with nub of opened ureter
at the "handle" of the fan. The mucosa should be examined to rule out rare mass
lesions. Measurements include mural thickness of the pelvis and ureter, and both
external and internal diameter of the ureter (the latter at its narrowest point).
If present, narrowing and angulation of the ureter should be noted. Sections
include serial cross sections of ureter and distal pelvis.
D. Segmental ureterectomy is performed for tumors of the proximal or mid-ureter.
The length and diameter of the intact ureter is recorded, with a search for a
mass by palpation and visual inspection. Proximal and distal cross-sectional
margins are taken, and the outer aspect of the ureter is inked. The ureter is then
opened longitudinally and assessed for mucosal abnormalities. After overnight
fixation in 10% formalin, sections are taken to demonstrate the deepest inva-
sion of any lesion(s). At least one section of uninvolved ureter should also be
submitted.
E. Radical cystectomy/cystoprostatectomy with segment Df ureters. Ureteral margin(s)
may be submitted for frozen section for evaluation of carcinoma (and particu-
larly carcinoma in situ). These are shaved margins and should be submitted as
cross sections.
373
F. Radical nephroureterectomy with bladder cuff. Gross examination and sampling

should document the relationship of tumor to adjacent renal parenchyma,
peripelvic fat, nearest soft tissue margin, and ureter. Sections of grossly
unremarkable kidney, pelvis, and ureter should be submitted. The important
urothelial margin is the urinary bladder cuff, which can be sampled as shave
sections.
A. Benign conditions that involve the urothelium in the upper tract have an
appearance similar to those in the urinary bladder. Examples are ureteritis
(e-Fig. 21.1), * pyelitis cystica and glandularis, malakoplakia, nephrogenic ade-
noma, and squamous metaplasia (e-Fig. 21.2).
B. Ureteropelvic junction (UPJ) abnormalities can be seen in biopsies done for UPJ
obstruction (UPJO), which is the most common cause of pediatric hydronephro-
sis and is usually diagnosed by fetal ultrasound. UPJO can also be seen in adults.
Although in a minority of cases ureteropelvic junction obstruction is due to
extrinsic compression by aberrant vessels, most cases are thought to be due to
an "intrinsic" abnormality of the UPJ itself. The pathogenesis of intrinsic UPJO
is not well established. Proposed etiologies include abnormal recanalization of
the proximal ureter during embryogenesis, persistence of fetal mucosal folds,
abnormalities of smooth muscle including disorientation and discontinuity, and
abnormal innervation. UPJO initially results in pelvic and calyceal dilatation,
followed by progressive obstructive renal damage which results in features of
renal dysplasia in patients with end-stage disease. Gross and histopathologic
findings reported in UPJO specimens include increased mural thickness, abnor-
mal mucosal folds/fibroepithelial polyps, smooth muscle hypertrophy and dis-
array (e-Fig. 21.3), attenuation of smooth muscle with increased collagen to
smooth muscle ratio, and chronic inflammation. Prognosis is based on concur-
rent renal biopsy and renal function rather than UPJ findings (Pediatr Nephrol.
2000;14:820, Pediatr Develop Pathol. 2006:9:72, Histopathology 2007;51:
709). Malignancy should also be excluded.
C. Benign epithelial neoplasms are rare and include urothelial papilloma, inverted
papilloma, villous adenoma, and squamous papilloma.
D. Benign nonepithelial neoplasms include the distinctive fibroepithelial polyp,
which is most frequently seen in the proximal ureter of young males. Micro-
scopically, these are exophytic intraluminal projections of a variably inflamed
fibrovascular stroma lined by a normal urothelium. Benign mesenchymal neo-
plasms such as leiomyoma, hemangioma, neurofibroma, and fibrous histiocy-
toma are rare.
E. Urothelial dysplasia in isolated form is rare and displays the same features
as in the urinary bladder, where it is defined as low-grade intraurothelial
neoplasia.
F. Renal pelvic and ureteral cancers are in the vast majority of cases, as in the uri-
nary bladder, of urothelial type (Am J Surg Pathol. 2004;28:1545, Mod Pathol.
2006;19:494, Adv Anat Pathol. 2008;15:127).
Upper tract tumors differ from those in the urinary bladder in the follow-
ing ways. Upper tract tumors are found at a lower frequency; have a stronger
association with long-term analgesic (such as phenacetin) abuse and urinary
tract obstruction; are associated with an increased frequency (in up to 65% of
patients) of synchronous or metachronous urothelial neoplasms elsewhere in the
urinary tract (Murphy WM, Grignon DJ, Perlman EJ. Tumors of the Kidney.
Bladder. and Related Urinary Structures. Washington, DC: American Registry
Chapter 21 • Renal Pelvis and Ureter I 3 75
of Pathology; 2004 ); and tend to present with higher histologic grade and at
higher stage. Also, biopsy of upper tract tumors is more difficult than lower
tract tumors. In about one of four cases, small ureteroscopic biopsies of the
upper tract will be nondiagnostic due to inadequate tissue (Am] Surg Pathol.
2009;33:1540).
1. Risk factors. The main risk factor, as for urinary bladder malignancies, is
smoking. Other risk factors are analgesics (as noted earlier); occupation
in chemical, petrochemical, or plastics industries; exposure to tar, coal,
or asphalt; papillary necrosis; Balkan nephropathy; thorium contrast expo-
sure; hereditary nonpolyposis colorectal cancer (HNPCC) syndrome (Lynch
syndrome II) UUrol. 2011;185:1627); and urinary tract infections or stones
(Eble JN, Sauter G, Epstein ]1, Sesterhenn lA, eds. Tumours of the urinary
system and male genital organs. Lyon: IARC Press; 2004).
2. Clinical features. Most patients are around 70 years of age, and the chief pre-
senting symptoms are hematuria and flank pain. In the majority of patients,
there is a prior history of a bladder cancer, which means that many upper
tract tumors are detected during the course of clinical surveillance after
diagnosis of a bladder tumor.
3. Histologic typing and diagnosis of upper tract tumors are accomplished by
examination of H&E-stained sections. Typing of upper tract urothelial neo-
plasia is the same as that for the urinary bladder as defined in the 2004 World
Health Organization (WHO) classification of neoplasms (see Table 22.1).
4. Urothelial carcinoma is by far the most common type of upper tract
tumor.
a. Gross diagnosis is possible in resection specimens. Patterns of growth
include papillary, polypoid, nodular, ulcerative, and infiltrative. Exo-
phytictumors can fill and distend the pelvis (e-Fig. 21.4), with or without
associated hydronephrosis and stones. High-grade invasive tumors can
grossly involve soft tissue and/or renal parenchyma (e-Fig. 21.5). Exten-
sive renal parenchymal involvement can mimic a primary renal parenchy-
mal neoplasm; in these cases there is often a request for an intraoperative
consultation to determine whether the mass is a urothelial carcinoma or
a renal cell carcinoma (RCC) because the distinction alters the extent of
surgery. The correct diagnosis can usually be determined by straightfor-
ward gross examination alone, but in some cases a frozen section may be
required. The average size of renal pelvic tumors is just under 4 em, with
a range of 0.3 to 9 em (Am] Surg Pathol. 2004;28:1545). Multifocality
in the pelvis and ureter is seen in about one-quarter of cases. In the ureter,
the tumor may be associated with a stricture and hydroureter.
b. Microscopically, the full range of urothelial carcinoma may be seen from
flat intraepithelial neoplasia, including carcinoma in situ (e-Fig. 21.6);
to noninvasive papillary neoplasia, including papillary urothelial neo-
plasm of low malignant potential; to low-grade papillary urothelial carci-
noma (e-Fig. 21.7A and B); to high-grade papillary urothelial carcinoma
(e-Fig. 21.8A and B; e-Fig. 21.9). The full spectrum of invasive urothelial
carcinoma and its variants as found in the urinary bladder can also be
found in the upper tract. Of note, unusual histomorphological variants
seem to be more common in the upper tract (Mod Pathol. 2006;19:494 ),
including carcinomas with micropapillary, lymphoepithelioma-like, sar-
comatoid, squamous, clear cell, glandular, rhabdoid, signet-ring, and
plasmacytoid features or areas.
Primary resections performed for renal pelvic urothelial carcinoma
show high-grade carcinoma in > 70% of cases, with deep invasion
(pT2 or greater) in 45% of cases and with lymph node metastases in
one-quarter of patients. Cancerization of the distal renal collecting ducts

is common in high-grade urothelial carcinoma (e-Fig. 21.10). Levels of
invasion include lamina propria invasion (e-Fig. 21.11); extension into
muscularis propria (e-Fig. 21.12), renal sinus and hilar fat, and peri-
ureteral fat (e-Fig. 21.13); and renal parenchymal infiltration. Metastatic
sites include lymph nodes, peritoneum, and liver. When the differential
diagnosis of a centrally located high-grade carcinoma centers on urothe-
lial carcinoma versus RCC, extensive mucosal sampling is important
because identification of urothelial carcinoma in situ (CIS) is in keeping
with a urothelial primary.
c. Immunohistochemical studies are not usually needed but can be useful in
the differential diagnosis of urothelial carcinoma versus RCC when the
diagnosis is not clear by standard gross and microscopic examination.
A useful marker panel includes cytokeratin CK7, CK20, p63, throm-
bomodulin, uroplakin III, RCC marker (RCC Ma), CD10, PAX2, and
PAX8. Positivity for the first five favors urothelial carcinoma, whereas
immunoreactivity for the last four suggests RCC rather than urothelial
carcinoma (Semin Diagn Pathol. 2005;22:51; Pathol Case Rev. 2010;
15:25).
d. Molecular studies. The most promising test is fluorescence in situ
hybridization (FISH) performed on cells in urine from the upper tract.
In this test (UroVysion/Vysis/Abbott), a mixture of fluorescent probes to
chromosomes 3, 7, 17, and 9p21locus is used to detect numerical chro-
mosomal abnormalities associated with urothelial carcinoma (Expert
Rev Mol Diagn. 2007;7:11). The precise clinical indications for its use
for upper tract tumors are not yet established.
5. Squamous cell carcinoma is rare, is more common in the pelvis, is frequently
associated with nephrolithiasis and/or infection, and often presents with
high-stage and high-grade disease (e-Fig. 21.14A and B). Pure squamous
cell carcinomas should be distinguished from urothelial carcinoma with
squamous differentiation.
6. Adenocarcinoma is rare and may display enteric, mucinous, and/or
signet-ring features. Pure adenocarcinomas should be separated from
urothelial carcinoma with glandular differentiation. Intestinal metaplasia,
nephrolithiasis, and infection are predisposing factors.
7. Small cell carcinomas in very rare cases arise from the renal pelvis. There
may be admixed urothelial carcinoma.
8. Malignant mesenchymal neoplasms are rare. The most common is
leiomyosarcoma. Other exceedingly rare mesenchymal tumor types include
osteosarcoma, rhabdomyosarcoma, fibrosarcoma, angiosarcoma, and
Ewing sarcoma.
9. Hematolymphoid neoplasms in the ureter and renal pelvis typically are due
to secondary involvement.
10. Miscellaneous neoplasms rarely encountered include paraganglioma, carci-
noid tumor, Wilms tumor, malignant melanoma, and choriocarcinoma.
11. Secondary (metastatic} malignancies include carcinomas of the cervix,
prostate, colon, breast, and urinary bladder.
IV. HISTOLOGIC GRADING should be performed for all carcinomas, with urothelial car-
cinoma grade being assigned as low grade or high grade, as for the urinary blad-
der. Pure squamous cell carcinoma and adenocarcinoma may be graded as well-
differentiated, moderately differentiated, or poorly differentiated.
V. PATHOLOGIC STAGING applies only to carcinomas of the ureter and renal pelvis. Stage
is the most important prognostic factor for upper tract carcinoma. The 2010 Tumor,
Node, Metastasis (TNM) American Joint Committee on Cancer/International
Union Against Cancer (AJCCIUICC) staging classification is given in Table 21.1.;
Cltlptet 21 • RaniI Ptll•i:a tftd Utbt I .,,
1'1.1 ]I fJI\, l'IIIMI.!Wt.IWI.ttaliiii(TNIIO .._klllmthwCIIcl- of
' _ R•ll'lllnulld ureter
,,, , . . . . (!)
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l3 Ren.ll llllill lurnm: TUtnOf- beyond """""IIJ1s lntD pelfpelolc: fll
or RN"WI pucr.dlyma
U-.Jtu,.,.: T11110r l'rlldoo l>o!oorld IIIIIOCII\am I!Co peltJ""'IIc flrt
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Subepithelial
connective tissue
Muscularis
propria
\. - I.A. _\ ..... 1-A J

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Peripelvic fat
T= pT
T3
Figure 21.1 Depiction of pT stages pTa, pTl, pT2, and pT3. (Modified
from: Greene FL, Compton CC, Fritz AG, et al., eds. A]CC Cancer Staging
Atlas. New York, NY: Springer; 2006.)
as other mucosal lesions, renal parenchymal lesions, stones, hydronephrosis, and

hydroureter should be documented when present. Microscopically, additional rele-
vant pathologic findings, if present, include lymphovascular space invasion by car-
cinoma, flat urothelial carcinoma in situ {focal vs. multifocal), inflammation, renal
epithelial neoplasia or medical renal disease (glomerulopathy), and metaplasias of
the urothelium, including keratinizing squamous metaplasia and intestinal meta-
plasia. A nomogram predicting cancer-specific survival after nephroureterectomy
for upper tract urothelial carcinoma uses age, tumor grade, pT stage, and pN stage
(Cancer. 2010;116:3774).
The Urinary Bladder
Omar Hameed and Peter A. Humphrey
I. I NORMAL MICROSCOPIC ANATOMY. The wall of the urinary bladder is formed by four
layers (e-Fig. 22.1)."' The thickness of the innermost layer, the urothelium, depends
on the degree of bladder distension, and the shape of its constituent urothelial cells
ranges from smaller cuboidal cells at the base to larger polyhedral cells toward
the surface. Umbrella cells, the most superficial cells, have abundant eosinophilic
cytoplasm and are often binucleated. The underlying lamina propria is separated
from the urothelium by a thin basement membrane and is composed of loose con-
nective tissue with blood vessels, nerves, adipose tissue, and a variable number of
smooth muscle fibers forming a discontinuous muscularis mucosae. Aggregates
of urothelium termed von Brunn nests (e-Fig. 22.2) are often seen as invagina-
tions or separate clusters in the lamina propria. The term cystitis cystica is used
when these nests become prominent and undergo cystic change (e-Fig. 22.3). Cys-
titis glandularis is similar to cystitis cystica except that the cells lining the cysts
are mucin-secreting cuboidal or columnar cells, or true goblet cells, in which case
the term cystitis glandularis with intestinal metaplasia is used (e-Fig. 22.4). These
proliferative lesions, although sometimes seen associated with local inflammation,
represent variants of normal histology. Their main importance lies in the fact that
they can occasionally cause visible lesions simulating a bladder neoplasm. The third
layer, the muscularis propria or detrusor muscle, is composed of large bundles of
muscle fibers and is covered by the outermost adventitial layer, including perivesical
adipose tissue. It is important to note that adipose tissue can also be found in the
lamina propria and wall (e-Fig. 22.5), so identification of fat does not equate to a
specific layer of the bladder wall.
II. GROSS EXAMINATION AND TISSUE SAMPLING OF THE BLADDER. The most common
samples submitted for surgical pathology examination include small biopsies, larger
transurethral resection specimens, and partial and radical (complete) cystectomies.
A. Biopsy specimens. These are usually obtained without cautery ("cold-cup") and
should be immediately immersed in formalin. If multiple biopsies are submitted
separately, as in mapping procedures, they should be processed separately. After
gross examination, bladder biopsies should be marked with ink or hematoxylin,
then placed in a cassette after being put in a fine mesh envelope, wrapped in lens
paper, or sandwiched between sponge pads. After processing, three hematoxylin
and eosin (H&E)-stained slides should be prepared, each with a strip of three
to four levels.
B. Transurethral resection of bladder specimens. These specimens are usually
obtained with the aid of thermal cautery, often for the transurethral resection of
bladder tumors (TURBT). Because of the significant prognostic and therapeu-
tic implications for the presence of muscularis propria invasion by the bladder
neoplasms, it is often necessary to process all of the submitted tissue to ensure
that such foci of invasion are not overlooked.
C. Partial cystectomy specimens. Partial or segmental cystectomy is indicated in
only a minority of bladder cancer patients, typically those who suffer a first
time tumor recurrence with a solitary tumor, and tumor location that allows for
a 1- to 2-cm margin of resection, such as at the dome. Urachal carcinomas at the
379
dome and above, with extension toward the umbilicus, may also be treated by
partial cystectomy, as can carcinoma in a bladder diverticulum. Carcinoma in
situ elsewhere in the bladder (or multifocal tumors) is an absolute contraindica-
tion. The specimens usually consist of a sheet-like portion of tissue that should
be pinned down and fixed overnight. In addition to describing and sampling any
gross tumor(s) as described for cystectomy specimens, the status of the margins
is very important; these can be shaved off or sampled by perpendicular sections,
depending on their relationship and proximity to the tumor(s). Frozen section
of the mucosal margin may be requested.
D. Total cystectomy and cystoprostatectomy specimens. Radical cystoprostatectomy
in men and anterior exenteration in women, along with pelvic lymphadenec-
tomy, are standard surgical approaches for muscle wall-invasive bladder car-
cinoma in the absence of metastatic disease. Cystectomy may be performed in
some cases for nonmuscle wall-invasive bladder carcinoma, if the bladder is non-
functional, or for high-grade pT1 carcinoma that is not responsive to intravesical
therapy. If not sampled separately, a request to perform frozen sections on the
ureteric and urethral margins may be received (Arch Pathol Lab Med. 2005;129:
1585-1601). After orientation and inking, one of two methods can be used to
fix the specimen. The first entails filling the bladder with formalin (through the
urethra, or by using a large bore needle through the dome) and fixing overnight;
the second entails opening the bladder (usually through the urethra extending
upward on the anterior surface) and pinning it flat, then fixing overnight. After
opening the bladder (in the fresh or fixed state), the mucosa is examined for
tumors(s) and, if present, the size, location, pattern of growth (exophytic, endo-
phytic, and/or ulcerated), and depth of invasion are recorded. The mucosa of the
adjacent bladder should also be examined for areas of hemorrhage and discol-
oration that may represent areas of carcinoma in situ. In addition to sampling of
any tumor(s) (three to four sections of each), representative sections need to be
submitted from the different areas of the bladder including the trigone; poste-
rior, lateral, and anterior walls; and the dome (Fig. 22.1). If ureteric and urethral
shave margins were not submitted for frozen section examination, they should
be sampled for permanent sections, as should any possible lymph nodes iden-
tified in the perivesical fat. In cystoprostatectomy specimens, additional blocks
from the prostate and seminal vesicles should be submitted, the extent of which
depends on whether a preoperative diagnosis or suspicion of prostatic carci-
noma exists (see Chap. 29). When the bladder (with or without the prostate) is
removed as part of larger pelvic exenteration specimens (that may include por-
tions of the rectum and/or the gynecological tract in females), it then becomes
imperative to document the presence or absence of involvement of these addi-
tional organs by preferentially sampling suspicious areas, as well as by sampling
the resection margins of these organs.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF THE BLADDER
A. Congenital malformations
1. Urachal abnormalities. The urachus is a vestigial structure that connects the
dome of the bladder to the umbilicus; it normally closes by the fourth month
of fetal life. Persistence or malformations of the urachus can present in child-
hood and occasionally in adulthood; they include urachal remnants (e-Fig.
22.6), patent urachus, urachal cysts, and urachal sinuses, all of which can
result in secondary infection or development of secondary tumors, most fre-
quently adenocarcinoma (e-Fig. 22.7).
2. Exstrophy. This is a rare congenital anomaly characterized by failure of devel-
opment of the anterior wall of the bladder and abdominal wall, usually
resulting in severe secondary infection if left untreated.
B. Inflammatory conditions. Cystitis most frequently has an infectious etiology.
There are, however, specific variants of cystitis that produce somewhat
Chapter 22 • The Urinary Bladder I 3 81
Whole organ bivalve method
Dissection method
Distal
./ureter
)
~Distal
urethra
Bladder
wall and
lesion
Ureteric
orifice
near lesion
B
Figure 22.1 Gross dissection of radical cystectomy specimens by whole organ bivalve
method (A) and by opening along urethra and anrerior wall, with demonstration of sec-
tions to be taken (B). lbree to four sections, or complete embedding, of masses should be
performed. (Modified from Schmidt WA. Principles and Techniques of Surgical Pathology.
Menlo Park, CA: Addison-Wesley Publishing Co.; 1983:525.)
characteristic cystoscopic and/or microscopic appearances. The latter include

hemorrhagic, granulomatous, eosinophilic, and interstitial variants, as well as
malakoplakia.
1. lnfactious cystitis. Acute and chronic cystitis is most frequendy secondary to
bacterial infection (usually by enteric organisms). The incidence is higher in
females, and when intermittent urinary obstruction or stasis is present. Biopsy
during active infection is contraindicated, but biopsy may be performed in

cases of chronic cystitis to rule out neoplasia, especially carcinoma in situ. The
histopathologic features include a nonspecific acute and/or chronic inflam-
matory infiltrate occasionally with lymphoid aggregates/follicles (follicular
cystitis) (e-Fig. 22.8), and a variable degree of lamina propria edema. Of note,
similar findings may be seen in the absence of infection such as following
radiation or cytotoxic chemotherapy. Other infections can produce specific
histologic findings such as viral inclusions (polyoma and herpesviruses) or
granulomas (tuberculosis, fungal infections, and schistosomiasis).
2. Granulomatous cystitis. As noted above, bacterial, fungal, or parasitic infec-
tions can lead to granuloma formation. The most frequent cause, however, is
iatrogenic, either secondary to intravesical Bacille Calmette-Guerin (BCG)
immunotherapy for urothelial CIS and superficially invasive carcinoma
(e-Fig. 22.9), or following TURBT (e-Fig. 22.10).
3. Hemorrhagic cystitis. This is an uncommon side effect of cyclophosphamide
therapy that results in extensive ulceration and hemorrhage that, if severe,
may require cystectomy. Adenovirus infection (in immunocompromised indi-
viduals) may also produce the same pattern.
4. Papillary and polypoid {bullous) cystitis. These related forms of cystitis are
characterized by finger-like (papillary) or broad (polypoid) projections of
edematous, variably inflamed lamina propria covered by reactive nonneo-
plastic urothelium (e-Fig. 22.11). These forms of cystitis are most frequently
seen with prolonged indwelling catheter use. The presence of such an inflam-
matory component in papillary cystitis can help in making the occasionally
difficult distinction from a low-grade papillary urothelial neoplasm.
5. Eosinophilic cystitis. This is characterized by dense infiltration of eosinophils
in the bladder lamina propria (e-Fig. 22.12) and/or wall. Such infiltrates
are most frequently seen adjacent to invasive urothelial carcinoma, but they
also occur in patients with allergic conditions and in patients with parasitic
infections, both often in association with peripheral eosinophilia.
6. Interstitial cystitis. This is an uncommon inflammatory disorder that pre-
dominantly affects middle-aged and elderly women and results in severe
intractable symptoms of culture-negative cystitis. Petechial submucosal hem-
orrhages or ulcers (termed Hunner ulcers) are usually evident cystoscopically.
The microscopic features are nonspecific and include a mixed inflammatory
infiltrate in the lamina propria, often with an increased number of mast cells
that can also involve the muscularis propria (e-Fig. 22.13) and nerves. Clini-
cal correlation is required in these cases because the histological findings are,
at most, consistent with the clinical impression of interstitial cystitis.
7. Malakoplakia. This is an uncommon form of cystitis characterized by the
presence of soft yellowish mucosal plaques composed of inflammatory cells
including abundant epithelioid histiocytes (known as von Hansemann histi-
ocytes) (e-Fig. 22.14) that have granular eosinophilic cytoplasm, and char-
acteristic 3- to tO-micron rounded basophilic intracytoplasmic inclusions
(Michaelis-Gutmann bodies) that contain iron and calcium, best demon-
strated by Prussian blue and von Kossa special stains, respectively. The con-
dition is thought to result from a defect in the ability of histiocytes to degrade
phagocytosized bacteria. Control of urinary tract infection can help control
the disease.
C. Reactive and metaplastic urotheliallesions
1. Squamous metaplasia. This can be of two types, nonkeratinizing and kera-
tinizing. The former is considered a normal finding in the trigone and bladder
neck of females but can rarely be seen in males receiving estrogen treatment
for prostate cancer. In contrast, keratinizing squamous metaplasia is more
common in males in association with chronic irritation and is considered
a significant risk factor for the subsequent development of carcinoma

(Eur Urol. 2002;42:469 and Am J Surg Pathol. 2006;20:883).
2. Intestinal metaplasia. In addition to the intestinal metaplasia occasionally seen
in cystitis glandularis, intestinal metaplasia in the presence of a chronically
irritated bladder can involve the bladder mucosa and lamina propria in a
focal or diffuse manner, resulting in an appearance almost indistinguishable
from colonic mucosa.
3. Nephrogenic metaplasia (adenoma). This is a benign epithelial proliferation
composed of cells resembling renal tubular epithelium (hence the name),
which usually arises in the setting of chronic irritation or injury such as
infection or calculi (Adv Anat Pathol. 2006;13:247). The lesion was believed
to be metaplastic, but more recent evidence has demonstrated that, at least
in renal transplant recipients, nephrogenic metaplasia is derived from shed
renal tubular epithelial cells that may attach to areas of prior injury. Although
most frequently seen in the bladder, it is also quite common in the urethra,
and less so in the ureters and renal pelvis. Nephrogenic metaplasia is usually
an incidental finding, but it may also present as a mass lesion simulating
cancer. Histologically, papillae (e-Fig. 22.15), small tubules (e-Fig. 22.16), or
cystically dilated tubules (e-Fig. 22.17) lined by cuboidal, low-columnar, or
flattened hobnail cells are seen. The importance of nephrogenic metaplasia
lies in the fact that in the bladder it can be confused with adenocarcinoma
(especially clear-cell adenocarcinoma) and glandular variants of urothelial
carcinoma (Mod Pathol. 2009;22:S37-S52); in the urethra it can be con-
fused with prostatic adenocarcinoma. The immunohistochemical reactivity
of nephrogenic metaplasia with cytokeratin 7, a-methylacyl coenzyme A
racemase (AMACR), PAX2, and PAX8 antibodies and the lack of reactivity
with high-molecular-weight cytokeratin (e.g., 34,8E12) and prostate-specific
antigen (PSA) antibodies help in distinguishing it from its mimics.
4. Urothelial hyperplasia. An increase in the thickness of the urothelium
(>?layers) is usually reactive and most frequently seen secondary to chronic
inflammatory conditions. Flat urothelial hyperplasia (e-Fig. 22.18) is more
common than papillary urothelial hyperplasia, which some authors have also
found to be associated with papillary urothelial neoplasms.
5. Pseudocarcinomatous epithelial hyperplasia. This change may be seen after
radiation therapy, chemotherapy, or unassociated with either therapy (Arch
Pathol Lab Med. 2010;134:427). The light microscopic appearance is of
pseudoinfiltrative nests of urothelium in the lamina propria (e-Fig. 22.19),
sometimes with squamous metaplasia. Nuclear atypia may be detected, sec-
ondary to therapy or irritation/ischemia. The irregular nests and aggregates
may appear to wrap around ecstatic vessels with fibrin thrombi, which is a
useful diagnostic finding.
6. Reactive urothelial atypia. Usually seen in a setting of acute and/or chronic
inflammation, reactive atypia may be associated with hyperplastic or thin
urothelium. Nuclear enlargement, often with vesicular chromatin and a sin-
gle prominent nucleolus, is the most prominent finding (e-Fig. 22.20). Mitotic
figures may be increased and cell crowding may be observed; however, polar-
ity, cell uniformity, and maturation are usually well preserved. Acute or
chronic inflammation is often identified.
D. Miscellaneous nonneoplastic conditions
1. Endometriosis. Most frequently seen on the serosal aspect of the bladder in
women with a previous history of pelvic surgery, foci of endometriosis can
also involve the lamina propria or muscularis propria and may be visible
cystoscopically. As elsewhere, at least two of the three histologic features of
endometriosis-endometrial glands, endometrial stroma, and hemosiderin-
laden macrophages-are required for the diagnosis (e-Fig. 22.21).
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epithelium (very similar to so-called "prostatic urethral polyps" of the

urethra).
6. Fibroepithelial polyp. These rare polyps are considered nonneoplastic. They
may be acquired or congenital. About one-half are found in neonates and chil-
dren. Histologically, there is a polypoid configuration, often with club-like
or finger-like projections (e-Fig. 22.22) (Am] Surg Pathol. 2005;29:460).
The surface urothelial lining is normal and there may be tubular or anas-
tomosing urothelium. in the dense fibrous tissue of the polyp fibrovascular
core. Degenerative-type stromal cell atypia, without mitotic activity, may
be present. These polyps have a broader fibrovascular core than urothelial
papillomas and lack the edema and inflammation of polypoid cystitis.
E. Neoplastic urothelial lesions. Urothelial neoplasms are the most common neo-
plasms that involve the bladder:. Histologic typing and diagnosis of urothelial
abnormalities are accomplished by examination of H&:E-stained sections. The
2004 World Health Organization (WHO) classification of tumors of the urinary
tract is given in Table 22.1.
1. Flat urotheliallesions with atypia. In addition to reactive atypia discussed ear-
lier, such lesions include urothelial dysplasia and urothelial carcinoma in situ.
a. Urothelial dysplasia. Dysplasia is an intraepithelial neoplastic urothelial
proliferation characterized by variable degrees of loss of polarity, nuclear
enlargement, and chromatin clumping (e-Fig. 22.23), all of which fall
short of the degree seen in carcinoma in situ. Dysplasia is often identified
in patients with urothelial neoplasms. Occasionally, it may be difficult to
distinguish urothelial dysplasia from reactive atypia; in such situations a
diagnosis of atypia of unknown significance may be warranted. Isolated
urothelial dysplasia progresses to bladder carcinoma in about 15% of
cases (Cancer. 2000;88:625).
b. Urothelial carcinoma in situ. This is a high-grade, often multifocal
intraurothelial neoplastic proliferation characterized by the presence of
unequivocal malignant urothelial cells within the bladder epithelial lining
(Fig. 22.2, e-Fig. 22.24 ). The urothelial proliferation need not involve the
entire thickness of the urothdium, and pagetoid and undermining patterns
of growth are not uncommon. Another common feature is the discohesive
Small Cell
-.-- ---.--
Figure 22.2 Patterns of urinary bladder carcinoma in situ. (From McKenney JK, et al. Am] Surg
Pathol. 2001;25:356, with permission.)
nature of the neoplastic cells that often leads to denudation and a "cling-
ing" pattern of growth in which only scarce malignant cells remain
attached to the bladder wall (e-Fig. 22.25). Surface carcinoma in situ cells
can also extend into and "cancerize" von Brunn nests (e-Fig. 22.26}. In
uncommon cases, glandular differentiation, in the form of more colum-
nar cells with apical cytoplasm, can be noted in urothelial carcinoma
in situ (Am] Surg Pathol. 2009;33:1241). In difficult cases where the
differential diagnosis includes reactive urothelial atypia, immunostains
for cytokeratin 20, p53, CD44, and K.i-67 can be useful {Semin Diagn
Pathol. 2005;22:69). Urothelial carcinoma in situ is often associated with
invasive urothelial carcinoma and carries a significant risk of death from
bladder carcinoma (Cancer. 1999;85:2469}.
2. Papillary urathelial neoplasms
a. Urathelial papilloma. This uncommon benign neoplasm is composed of
delicate papillary urothelial fronds with no or minimal branching or
fusion. The constituent cells are identical to normal urothelial cells, and
no mitoses are present (e-Fig. 22.27). The classic cystoscopic finding is a
solitary lesion in a younger patient with hematuria. Papillomas may recur
(in up to 80% of cases) or progress to higher grade disease (in 2% of cases).
b. Inverted papilloma. Another uncommon neoplasm, an inverted papilloma
has a polypoid or sessile appearance cystoscopically. It is composed of
anastomosing islands and cords of bland urothelial cells that invaginate
and grow downward in the lamina propria with peripheral palisading, no
to rare mitoses, and no to minimal cytological atypia (e-Fig. 22.28) (Am
] Surg Pathol. 2004;28:1615, Cancer. 2006;107:2622). These latter two
features help distinguish this lesion from other papillary neoplasms that
may also occasionally have an inverted growth pattern. Inverted papillo-
mas rarely recur.
c. Papillary urathelial neoplasm of law malignant potential (PUNLMP). This
neoplasm shares the clinical and endoscopic features of papilloma but is
characterized histologically by occasionally fused papillae and ordered, yet
larger, nuclei than are seen in papillomas {e-Fig. 22.29A and B). Mitotic
figures are rare and basal. Compared with papillomas, PUNLMP has
higher recurrence (25% to 35%} and progression rates (up to 4%), and
thus close follow-up is warranted.
d. Noninvasive papillary urathelial carcinoma, law grade. In contrast to
PUNLMP, this urothelial neoplasm shows frequent branching and fusion
of papillae and variations in nuclear size, shape, and contour {e-Fig.
22.30). Mitoses are occasional and may be found at any level. These
tumors tend to be larger than papillomas and PUNLMPs and are more
likely to be multiple. They are also more likely to recur (64% to 71 %)
and progress (2% to 10%).
e. Noninvasive papillary urathelial carcinoma, high grade. These are uncom-
mon noninvasive papillary neoplasms; more frequently there is associated
invasion. They are characterized by frequent branching and fusion with
moderate to marked cytoarchitectural disorder and nuclear pleomorphism
(e-Fig. 22.31). Mitoses are frequent. Similar to low-grade tumors, these
tumors frequently recur (56%) and progress to invasive carcinoma (18%).
3. Invasive urathelial neoplasms. Invasive urothelial carcinomas can have papil-
lary, polypoid, nodular, or ulcerative configurations. Most are cytologically
high-grade tumors. Determination of anatomic depth of invasion by carci-
noma is vital, because pathological stage is the single most important prog-
nostic feature.
Recognition of diagnostic patterns of lamina propria invasion can facili-
tate pT1 stage assignment (Table 22.2). There are several different patterns,
C111ptw 22 • l~t-t Uri11wy B!•erliel I 117
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Figure 22.3 Patrerns of invasion from papillary carcinoma. A: Microinvasive carcinoma at base.
B: Stalk invasion. C: Lamina propria invasion up to muscularis mucosae. D: Lamina propria
invasion beyond muscularis mucosae. (From Amin MB, et al. Am J Surg Pathol. 1997;21:1057,
with permission.)
including those seen with papillary urothelial carcinoma (Fig. 22.3,

e-Figs. 22.32 and 22.33), carcinoma in situ (CIS) (e-Fig. 22.34), nested
carcinoma, and inverted pattern carcinoma (Fig. 22.4) (Am J Surg Pathol.
1997;21:1057). Recently emphasized pitfalls in the diagnosis of lamina pro-
pria invasive urothelial carcinoma include tangential sectioning, thermal arti-
fact, obscuring inflammation, carcinoma in situ involving von Brunn nests,
deceptively bland urothelial carcinoma, invasion into indeterminate type of
muscle, and invasion into adipose tissue within lamina propria (Epstein ]1,
Amin MB, Reuter VE. Bladder Biopsy Interpretation. 2nd ed. Philadelphia:
Lippincott Williams & Wilkins; 2010).
A B c
:·:::::m.:::
...
.,
D E
Figure 22.4 Patrerns of lamina propria invasion. A and B: From carcinoma in situ. C: Nested
variant. D: Endophytic growth and invasion. E: Inverted growth and invasion. (From Amin MB,
et al. Am J Surg Pathol. 1997;21:1057, with permission.)
Chapter 22 • The Urinary Bladder I 389
The level of invasion of carcinoma in pT1 disease is related to patient

outcome, with a worse prognosis for patients with tumors that invade the
muscularis mucosae (pT1b) or deeply into the subepithelial connective tissue,
as quantitated using an ocular micrometer. The WHO 2004 group recom-
mended that some estimate of extent of lamina propria invasion (e.g., pT1a
[invasion above muscularis mucosae] vs. pTl b tumors) be provided, but this
is currently not a formal part of the 2010 TNM system because there is no
established method for estimation that is consistently applicable and repro-
ducible.
Invasion by bladder carcinoma into muscularis propria (muscle wall,
detrusor muscle) (e-Fig. 22.35) is an ominous finding that makes the patient
a candidate for aggressive surgical therapy (radical cystectomy) or radiation
therapy, with or without adjuvant chemotherapy.
Determination of the type of muscle (muscularis mucosae vs. muscu-
laris propria) invaded by carcinoma can occasionally be difficult because of
small tissue sample size, tissue distortion, cautery artifact, poor orientation,
fibrosis, inflammation elicited by destructive growth of invasive tumor, or
even hypertrophy of the normally thin, wispy, and discontinuous muscu-
laris mucosae. The designation of "muscle type indeterminate" is a viable
description in some cases. Smoothelin immunohistochemistry has potential
for distinction of muscularis mucosae from muscularis propria, with the latter
showing strong and diffuse immunostaining, with relatively high sensitivity
and specificity (Am] Surg Pathol. 2010;34:792).
Substaging of pT2 (pT2a, invasion of "superficial" muscle = inner half
vs. pT2b, invasion of "deep" muscle= outer half) and distinction of pT2
versus pT3 can be performed only on radical cystectomy specimens and not
samples from transurethral resections of bladder tumor. Even in cystectomy
specimens, it can be a challenge at times to determine extravesical (pT3)
spread because the boundary between muscularis propria and its fat is not
well demarcated from perivesical fat. Moreover, this boundary can be dis-
torted, obscured, or obliterated by fibrosis and inflammation associated with
infiltrating tumor.
Numerous histologic variants of urothelial carcinoma with divergent dif-
ferentiation have been described (Mod Pathol. 2009;22:596), including the
following.
a. Invasive urothelial carcinoma with squamous differentiation. Focal squa-
mous differentiation, to be distinguished from pure squamous cell carci-
noma, is seen in up to 20% of invasive urothelial carcinomas, with an
increased likelihood of squamous features in high-grade urothelial carci-
nomas. Squamous differentiation may predict a poor response to radio-
therapy and systemic chemotherapy, although this has not been definitely
established.
b. Invasive urothelial carcinoma with glandular differentiation. True glandu-
lar spaces with or without mucin production and/or signet cells is seen
in approximately 6% of invasive urothelial carcinomas (e-Fig. 22.36).
Although this variant needs to be distinguished from adenocarcinoma, it
is not clear whether it behaves any different from classic invasive urothelial
carcinoma.
c. Urothelial carcinoma with trophoblastic differentiation. Trophoblastic dif-
ferentiation can be manifested by the presence of any of the following:
syncytiotrophoblastic giant cells, immunoreactivity for human chorionic
gonadotropin (hCG), or choriocarcinoma. hCG immunoreactivity can be
found in about one-third of high-grade urothelial carcinomas and can be
associated with serum elevations of hCG, but because such immunoreac-
tivity is not of prognostic significance hCG immunostains should not be
performed; a very few cases of urothelial carcinoma with hCG-positive

syncytiotrophoblasts (e-Fig. 22.37) have been reported. True syncytiotro-
phoblasts should be distinguished from the tumoral giant cells of giant
cell carcinoma and from osteoclast-like giant cells, and it is important to
recognize that true syncytiotrophoblasts represent examples of divergent
differentiation and not a primary germ cell tumor of the urinary blad-
der. Nonetheless, this variant should be reported because the prognosis is
extremely poor, with most patients dead of disease within 1 year. Very rare
cases of pure choriocarcinoma of the urinary bladder have been described,
but most likely represent urothelial carcinoma with trophoblasts.
d. Sarcomatoid variant (previously known as carcinosarcoma). This is a
biphasic neoplasm displaying histomorphologic and/or immunopheno-
typic evidence of both epithelial and mesenchymal differentiation (Am
] Surg Pathol. 1994;18:241). Because both components share the same
donal origin, this tumor is another example of a neoplasm with diver-
gent differentiation. This variant accounts for < 1% of bladder malig-
nancies. Previous radiation and cyclophosphamide treatment are pre-
disposing factors. Grossly, the tumor is often exophytic, polypoid, and
deeply invasive into muscularis propria. Microscopically, the growth is
typified by a biphasic population of neoplastic cells with the epithelial
and mesenchymal-like components often in continuity (e-Fig. 22.38A
and B). The carcinomatous component is usually urothelial (85%) but
can be adenocarcinoma, squamous cell carcinoma, or small cell carci-
noma. The amount of the malignant epithelial element varies and in some
cases is only represented by carcinoma in situ; consequently, apparently
pure malignant spindle cell tumors of the bladder should be sampled exten-
sively in an attempt to find epithelial areas. The sarcomatous component is
usually an undifferentiated high-grade spindle cell proliferation arranged
in fascicles, a storiform pattern, or as a patternless confluence. The most
common heterologous element is osteosarcoma, followed by chondrosar-
coma (e-Fig. 22.39), rhabdomyosarcoma, leiomyosarcoma, liposarcoma,
angiosarcoma, or mixtures thereof. The sarcomatous regions are almost
always high-grade. Immunohistochemical staining of the sarcomatoid
variant shows strong, diffuse immunopositivity for pan-cytokeratin (e-Fig.
22.38B ), although epithelial membrane antigen (EMA) immunostaining is
characteristically weak; note, however, that epithelial markers can be neg-
ative and focal immunoreactivity for desmin and smooth muscle actin may
be present. The opposite pattern of staining favors a leiomyosarcoma, in
the correct histopathologic context. Anaplastic lymphoma kinase (ALK)-
1 immunostaining, which typifies inflammatory myofibroblastic tumor, is
negative in sarcomatoid carcinoma. For cases of sarcomatoid carcinoma,
the pathology report should include whether the sarcomatoid carcinoma
is homologous or heterologous, although to date this does not appear to
be of prognostic importance. Surgery and radiation result in 25% survival
at 2 years.
Histologic variants of urothelial carcinoma with unusual growth pat-
terns:
e. Nested variant. This variant is characterized by the presence of infiltrating
small nests and tubules of urothelial carcinoma (e-Fig. 22.40). Despite its
relatively bland low-grade cytological features, this variant is often aggres-
sive. Because of its cytoarchitectural features, florid von Brunn nests, cysti-
tis cystica and glandularis, inverted papilloma, nephrogenic adenoma, and
paraganglioma are all in the differential diagnosis. Clues that are of aid in
establishing the diagnosis are the high density, nearly confluent nests that
can anastomose and invade muscularis propria. Nuclear atypia may be
only focally present and is often found in the deeper aspects of the prolif-
eration. Although the nested variant of urothelial carcinoma has a higher
proliferation index than florid von Brunn nests by MIB-1 immunostaining
(8.8% vs. 2.8%), and a higher p53 immunopositivity (4.2% vs. 1.5%),
the degree of overlap precludes use of these markers as diagnostic tools
(Am I Surg Pathol. 2003;27:1243).
f. Microcystic variant. This variant is also deceptively bland and somewhat
similar to the nested variant, except for characteristic prominent cystic
change (e-Fig. 22.41). It is uncommon, accounting for only about 1% of
bladder carcinomas. Microscopically, there are variable-sized cysts rang-
ing up to 1 to 2 mm in diamete.t The cysts are round to oval and may
contain necrotic material or pink secretions. The layer of lining cells may
be flattened or denuded. The differential diagnosis includes cystitis cys-
tica, cystitis glandularis, and nephrogenic adenoma. Correct diagnosis is
achieved by the detection of an association with usual urothelial carci-
noma, haphazard and infiltrative growth, and variability in cyst size and
shape. In the largest series, 25% of cases had invasion of muscularis pro-
pria and 11 of 12 were high grade (] Urol. 1997;74:722).
g. Micropapillary variant. This rare pattern of urothelial carcinoma resembles
papillary serous carcinoma of the ovary, an important differential diag-
nosis in women. It is characterized by the presence of small nests of cells
and filiform papillae that have retracted from the surrounding stroma
(e-Fig. 22.42). Lymphovascular invasion is common. Admixed invasive
usual urothelial carcinoma is detected in a majority of cases. Because the
proportion of the tumor that is micropapillary seems to be of prognos-
tic significance, the percentage of the invasive tumor that is micropap-
illary should be reported (Adv Anat Pathol. 2010;17:182). This variant
is characteristically aggressive. While muscle wall invasion is commonly
detected, some cases, especially those with a low percentage (<10%) of
micropapillary component and surface micropapillary growth, can be low
stage (pTa or pT1). It has been argued that early cystectomy should be
offered to patients with such low-stage, nonmuscle-invasive micropapil-
lary urothelial carcinoma (Cancer. 2007;110:62). Intravesical therapy is
ineffective (Urol Oncol. 2009;27:3). The 10-year overall survival is 24%.
h. Lymphoepithelioma-like variant. This is characterized by sheets and nests of
poorly differentiated malignant cells that grow in a syncytial pattern with
an admixed dense lymphoplasmacytic infiltrate (e-Fig. 22.43) (Am I Surg
Pathol. 2011 ;35:4 74 ). It may be pure or mixed with usual urothelial carci-
noma. There is a tendency for patients to present with muscularis propria-
invasive disease. The differential diagnosis centers on large cell lymphoma
and severe chronic cystitis, including follicular cystitis; immunostains for
pan-cytokeratin and CD45 are confirmatory, and Epstein-Barr virus is
not present. These are aggressive carcinomas with a 26% mortality at 3
years. These tumors should be treated as other bladder carcinomas-that
is, based on stage-although they do also appear to be chemoresponsive.
Histologic variants of urothelial carcinoma with unusual cytologic fea-
tures:
i. Lymphoma-like and plasmacytoid variants. These variants are exceedingly
rare and are usually admixed with conventional urothelial carcinoma.
However, the diagnosis of urothelial carcinoma in small biopsies com-
posed solely of such variants (e-Fig. 22.44) may be achieved only with
the help of immunohistochemistry (positive reactivity to cytokeratin with
negative reactivity to CD45 and other lymphoid markers).
j. Giant cell variant. This high-grade variant is characterized by numerous
pleomorphic and bizarre tumor giant cells (e-Fig. 22.45), and needs to be
distinguished from urothelial carcinoma with osteoclast-like giant cells,

which represents an unusual stromal response to invasive carcinoma. Out-
comeis poor, with median survival of 11 months (Br I Urol. 1995;75:167).
k. Glycogen-rich (clear-cell) and lipid-rich variants. The main importance
of these rare variants is the fact that they may be confused with
clear-cell adenocarcinoma of the bladder or kidney, and liposarcoma
or signet-ring carcinoma, respectively. The lipid-rich (also known
as lipoid-cell) variant carries a poor prognosis (Br I Urol. 1995;75:167).
I. Urothelial carcinoma with rhabdoid features. Rhabdoid cells may rarely be
observed in urothelial carcinoma; < 10 cases have been reported (Hum
Pathol. 2006;37:16). This variant is a high-grade carcinoma with a poor
clinical outcome. Morphologically, the rhabdoid cells have large, eccentric
nuclei with prominent nucleoli and eosinophilic cytoplasmic inclusions.
In children, primary malignant rhabdoid tumor of the bladder is in the
differential diagnosis, a diagnosis which requires molecular evidence of
mutations of the SMARCB1 gene (Ann Diagn Pathol. 2011; Jul18 [Epub
ahead of print]).
F. Squamous neoplasms
1. Squamous papilloma. This is a very rare papillary lesion that typically presents
in elderly women and, unlike condyloma acuminatum (which can also
involve the bladder), has not been associated with human papilloma virus
infection or with subsequent development of carcinoma (Am I Surg Pathol.
2006;20:883, Cancer. 2000;88:1679). Recurrence is rare.
2. Squamous cell carcinoma. In areas of the world where schistosomiasis is
endemic (parts of Mrica and the Middle East), this type of carcinoma is
the most common primary neoplasm of the bladder. Elsewhere, squamous
cell carcinoma is relatively rare, representing <5% of bladder carcinomas.
Other predisposing conditions include chronic cystitis and chronic irritation
due to vesical lithiasis or long-term indwelling catheters. Smoking is also a
significant risk factor (as for conventional urothelial carcinoma). Squamous
cell carcinomas are represented at a higher percentage in patients with non-
functioning bladders (50% of carcinomas) or diverticula (20% ), and in renal
transplant patients (15%). Keratinizing squamous metaplasia is a potential
precursor and is a risk factor for subsequent detection of carcinoma; 20% to
42% of patients with keratinizing squamous metaplasia are later diagnosed
with carcinoma.
Squamous cell carcinoma typically presents as invasive carcinoma,
although in a few cases pure squamous cell carcinoma in situ may be
detected (Am I Surg Pathol. 2006;20:883). Squamous cell carcinoma in situ
is a strong risk factor for subsequent detection of invasive carcinoma and
approximately 45% of patients with in situ disease are diagnosed with inva-
sive squamous cell or urothelial carcinoma within 12 months (Am I Surg
Pathol. 2006;20:883). Grossly, squamous cell carcinomas are often large,
solid necrotic masses that can fill the entire bladder lumen (e-Fig. 22.46).
Some, howeve.t;, may be flat and infiltrative with ulceration. Microscopically,
the carcinoma should be purely squamous, with keratin production and/or
intercellular bridges (e-Fig. 22.47). Adjacent keratinizing squamous meta-
plasia strongly supports a diagnosis of squamous cell carcinoma; such meta-
plasia is seen in 20% to 60% of cases of invasive squamous cell carcinoma.
Histologic variants of squamous cell carcinoma of the bladder include the
exceptionally rare basaloid variant and the uncommon verrucous variant.
Grading is three tiered (well, moderately, or poorly differentiated), and
histologic grade may correlate with stage and outcome. Howeve.t;, stage is the
most important determinant of outcome. Many patients with squamous cell
carcinoma of the bladder present with muscularis propria-invasive disease,
and this accounts for the poor outcome for most patients. No molecular
genetic abnormalities are currently used for diagnosis or prognosis. Treat-
ment is radical cystectomy, with or without radiation therapy and chemother-
apy.
3. Verrucous squamous cell carcinoma. This variant of squamous cell carcinoma
is seen almost exclusively in patients with schistosomiasis and appears as an
exophytic "warty" mass composed of thickened papillary squamous epithe-
lium with minimal cytoarchitectural atypia and a rounded pushing border.
This tumor is considered to be clinically indolent.
G. Glandular neoplasms
1. Villous adenoma. An uncommon exophytic papillary neoplasm histologically
resembling its colonic counterpart, villous adenoma is usually located in the
trigone and, unless associated with an invasive component, does not recur
following excision.
2. Adenocarcinoma. Primary adenocarcinomas are rare, representing 2% of
malignant bladder neoplasms, and may be of urachal or nonurachal origin.
The latter are more common and usually arise in patients with a nonfunc-
tional bladder or with exstrophy. Urachal adenocarcinomas usually arise
from the dome or anterior wall of the bladder but may also involve urachal
remnants in the anterior abdominal wall. Characteristics of urachal adeno-
carcinomas that are helpful in differentiating them from nonurachal adeno-
carcinomas are that their bulk is in the wall rather than the lumen of the
bladder; they lack an associated in situ component or cystitis glandularis;
and they are sharply demarcated from surface urothelium. Identification
of urachal tumors is important because, unlike nonurachal tumors, surgi-
cal management of urachal adenocarcinomas usually includes excision of
the median umbilical ligament and umbilicus. Bladder adenocarcinomas (of
urachal or nonurachal type) may have different morphologic appearances
including enteric (e-Fig. 22.48A and B), mucinous (e-Fig. 22.7), signet-ring
cell, dear cell, and mixed. The main differential diagnosis for most of these
patterns is the more common metastasis or secondary extension from another
primary tumor site, most notably the prostate and the colon. Immunohisto-
chemistry may be helpful in this situation, especially when the clinical findings
are not helpful or available. Immunoreactivity with PSA and prostatic acid
phosphatase, ,8-catenin (nuclear), or thrombomodulin supports a diagno-
sis of prostate, colon, or bladder adenocarcinoma, respectively; cytokeratin
7 and 20 immunostains are not very useful in this context because of sig-
nificant overlap (Semin Diagn Pathol. 2005;22:69). Adenocarcinoma of the
bladder has a generally poor prognosis with 5-year survival rates ranging
from 18% to 47%.
H. Neuroendocrine neoplasms
1. Paraganglioma. Derived from bladder paraganglia, paragangliomas are typ-
ically found in the muscularis propria and are not infrequently associated
with hypertension and/or headaches, palpitations, and sweating that may be
precipitated by micturition. The tumor is composed of cells with abundant
amphophilic, clear, or eosinophilic cytoplasm arranged in a diffuse or nested
(Zellballen) pattern of growth with an associated thin capillary network
(e-Fig. 22.49). Nuclear atypia can occasionally be prominent. The tumor
is frequently immunopositive for neuroendocrine markers and negative for
cytokeratin (useful in distinguishing the tumor from nested urothelial car-
cinoma), whereas the spindle cells surrounding tumor nests (sustentacular
cells) are positive for 5100 protein. The majority (85% to 90%) of bladder
paragangliomas are benign and do not recur following surgical excision.
2. Small cell carcinoma. This is a rare neoplasm, which is diagnosed even
when mixed with other bladder carcinomas (urothelial, squamous, or
adenocarcinomas) because the presence of any small cell carcinoma com-

ponent has a significant negative impact on prognosis. As with other bladder
carcinomas, hematuria is the most common presentation; however, almost
half of the patients present with metastatic disease, with or without a parane-
oplastic syndrome. Histologically, the tumor cells are characteristically small
with scant cytoplasm, stippled chromatin, and inconspicuous nucleoli with
nuclear molding. Admixture with usual urothelial carcinoma is common and
is present in about one-half of cases (e-Fig. 22.50). The diagnosis can often be
confirmed by immunoreactivity with one or more neuroendocrine markers
such as neuron-specific enolase, chromogranin, synaptophysin, and CD56,
with or without cytokeratin expression (which is usually seen in a dot-like
paranuclear pattern).
3. Rare neuroendocrine tumors. Large cell neuroendocrine carcinomas and car-
cinoid tumors are rare as primary tumors in the urinary bladder.
I. Mesenchymal lesions and neoplasms
1. Myofibroblastic proliferations. These spindle cell proliferations can develop
a few weeks to several months following bladder instrumentation (for
which the term "postoperative spindle cell nodule" has been used), or may
be unrelated to trauma. They can be quite large (up to 9 em in diam-
eter). These neoplasms are composed of myofibroblasts and have been
variously termed inflammatory myofibroblastic tumor (Am I Surg Pathol.
2007;20:592) and pseudosarcomatous myofibroblastic proliferation (Am I
Surg Pathol. 2006;30:787); it remains unsettled as to whether these are
all the same entity or a heterogeneous group of proliferations. Histologi-
cally, these lesions are composed of elongated spindle or stellate cells resem-
bling tissue-culture fibroblasts, embedded in a myxoid matrix that contains
a variable chronic inflammatory infiltrate and extravasated red blood cells
(e-Fig. 22.51). Although enlarged nuclei with prominent nucleoli (as well as
prominent mitotic activity) can be observed, the presence of significant
nuclear atypia and/or abnormal mitotic figures should suggest an alternative
diagnosis such as sarcomatoid carcinoma or leiomyosarcoma. Immunohis-
tochemistry can sometimes be a useful diagnostic tool as the spindle cells
are often positive for cytokeratin, actin, and vimentin. In addition, more
than two-thirds of cases display expression of ALK, often associated with a
translocation involving its locus on the short arm of chromosome 2 (2p23).
Recurrences can be seen and the proliferations can be locally aggressive, with
deep extension into muscularis propria and even into perivesical adipose tis-
sue. In only one case, there has been metastatic spread and in this case overt
sarcomatous features were also present (Am I Surg Pathol. 2006;30:1502).
2. Smooth muscle neoplasms. Leiomyomas and leiomyosarcomas represent the
most common benign and malignant mesenchymal neoplasms of the bladder,
respectively. Leiomyomas usually present as well-circumscribed lamina pro-
pria (two-thirds of cases), intramural, or subserosa! masses and are composed
of intersecting fascicles of spindle cells with abundant eosinophilic cytoplasm
and ovaVelongated nuclei with blunt ends. By definition, there should be no
significant nuclear atypia (hyperchromasia, nuclear membrane irregularities,
or pleomorphism) or evidence of an infiltrative growth pattern. In contrast,
leiomyosarcomas (e-Fig. 22.52) are infiltrative tumors with nuclear atypia,
coagulative tumor cell necrosis, and brisk mitotic activity (usually >5 mitoses
per 10 high-power fields), the latter being a typical feature of high-grade
tumors. In addition to showing immunoreactivity with one or more smooth
muscle markers (actins, desmin, and caldesmon), leiomyosarcomas can also
be at least focally immunopositive for cytokeratin and EMA. Most patients
with leiomyosarcoma develop recurrences and/or metastasis resulting in mor-
tality in almost one-half of cases.
3. Rhabdomyosarcoma. Almost a quarter of all childhood rhabdomyosarcomas

arise in the genitourinary tract, and a significant proportion are of blad-
der origin. Almost all genitourinary tract cases are of the embryonal type.
Grossly, most embryonal rhabdomyosarcomas of the bladder present as poly-
poid intraluminal masses (e-Fig. 22.53) resembling a cluster of grapes (hence
the alternative name botryoid type), with the remainder being deeply invasive
tumors. Histologically, they are mostly composed of small round tumor cells
with a variable admixture of spindle cells embedded in a myxoid stroma,
often with cell condensation under the surface urothelium forming the so-
called "cambium layer" (e-Fig. 22.54). The latter feature, as well as the iden-
tification of rhabdomyoblasts and cross-striations, is particularly useful for
diagnosis. Immunohistochemistry can confirm the diagnosis as these tumors
are usually positive for myogenin and myoDl, among other muscle markers.
The prognosis of embryonal rhabdomyosarcoma has greatly improved with
multimodality treatment, although the alveolar type and the rare rhabdomyo-
sarcomas presenting in adulthood are associated with a worse outcome.
4. Other spindle cell neoplasms. There are other spindle cell neoplasms that
can occasionally involve the bladder including hemangioma, neurofibroma,
solitary fibrous tumor, angiosarcoma, osteosarcoma, and malignant fibrous
histiocytoma. These tumors are histologically identical to their nonbladder
counterparts.
J. Other miscellaneous neoplasms. As discussed earlie.r; the bladder can be involved
by metastatic carcinomas and those extending from adjacent organs, where
clinical data and immunohistochemical findings can help resolve the diagnosis.
Hematolymphoid neoplasms can also involve the bladde.r; either primarily or
secondarily (more common), and may present as mass lesions. Finally, there are
a few reported primary bladder melanomas in the literature.
IV. HISTOLOGIC GRADING OF UROTHELIAL CARCINOMA is indicated as low grade or high
grade, as discussed earlier. Histologic grade of primary adenocarcinoma or squa-
mous cell carcinoma of the urinary bladder can be reported as well-, moderately,
or poorly differentiated.
V. PATHOLOGIC STAGING OF URINARY BLADDER CANCER applies to carcinomas only.
Pathologic primary tumor (pT) 2010 TNM AJCC stage categories are given in
Table 22.2 and illustrated in Figure 22.5. pN and pM groupings are also listed in
Table 22.2.
VI. REPORTING URINARY BLADDER CARCINOMA should follow suggested guidelines (see
the College of American Pathologists urinary bladder cancer protocol and checklists
at http://www.cap.org). For urinary bladder biopsy and transurethral resections, the
histologic type, grade, and depth of invasion are reported. The presence or absence
of muscularis propria should also be noted for each biopsy sample. Carcinoma
in situ, when present in flat urothelium adjacent to papillary tumo.r; should also
be reported. If lymphovascular space invasion by carcinoma is seen, it should be
reported.
For cystectomy (partial or total), radical cystoprostatectomy, and pelvic exen-
teration specimens, the following are reported: tumor location, size in three dimen-
sions, gross growth pattern (papillary, nodular/solid, flat, ulcerated), gross depth of
invasion, gross involvement of adjacent structures (prostate, vagina, uterus, colon),
and relation to surgical margins. Microscopic features that should be reported
include histologic type, grade, site(s) of involvement, growth pattern, extent of
invasion, involvement of other structures, lymphovascular invasion, and margin
status. Important margins include the ureters, distal urethra, perivesical soft tissue
(for cystectomy specimens), and pelvic soft tissue (for exenteration specimens). For
regional lymph nodes, the total number examined, number positive for carcinoma,
presence of extranodal extension, and size of largest metastatic deposit should be
documented.
Subepithelial
connective --+~~~
tissue
-+-- Prostate
-+-------.(__ Urethra
Figure 22.5 Pathologic T staging of carcinoma of the urinary bladder.

(Modified from Greene FL, et al., eds. AJCC Cancer Staging Atlas. New
York, NY: Springer; 2006.)
Cytopathology of the
Urinary Bladder
Rosa M. D~vila
I. TYPES OF SPECIMENS
A. Voided urine normally has a mixture of benign urothelial cells and squamous
cells. Although the squamous cells may be a vaginal or skin contaminant, they
can also be derived from areas of squamous metaplasia that often develop in the
bladder trigone. When urothelial cells are present in papillary-like clusters, the
possibility of a low-grade urothelial neoplasm should be raised. Similar dusters
may also be due to recent instrumentation or lithiasis.
B. Catheterized urine normally has papillary-like dusters of urothelial cells resulting
from the mechanical disruption of the urothelial mucosa. They should not be
confused with low-grade urothelial carcinoma.
C. Bladder washings are obtained by irrigating the bladder with saline instilled
via a catheter, or during cystoscopic evaluation. The cytologic findings in this
specimen type are similar to those seen in catheterized urine.
D. Neobladder or ileal conduit samples have abundant degenerated cells, some of
which are arranged in dusters and have vacuolated cytoplasm. Well-preserved
intestinal-type epithelium is rarely present. A variable number of inflammatory
cells, macrophages, and bacteria are seen (e-Fig. 22.55).
II. INFLAMMATORY/INFECTIOUS PROCESSES

A. Eosinophils. Detection of eosinophils may be difficult in Papanicolaou-stained
slides because the cytoplasmic granules do not stain prominently. Therefore,
identification of eosinophils is based on their bilobed nucleus. Patients with
allergic interstitial nephritis or eosinophilic cystitis, or those who have had recent
bladder biopsies, can display urinary eosinophils.
B. Human polyoma virus (BK virus} is an important cause of morbidity in renal trans-
plant patients. This DNA virus initially infects children and becomes dormant
until the patient becomes immunosuppressed. Because it infects the renal tubular
epithelial cells and urothelial cells, the cellular changes can be detected by urine
cytology. Infected cells are usually arranged singly and have a blackish-blue
inclusion that occupies most of the nucleus. The cellular changes can resemble
those of high-grade urothelial carcinoma, and therefore infected cells are also
known as "decoy cells." In contrast to high-grade urothelial carcinoma, decoy
cells have a round nucleus with smooth contours that has dark, homogenous
staining (Am] Transplant. 2001;1:373).
C. Acute inflammation is common in patients with lower urinary tract infections
that do not require cytologic evaluation. However, urine samples with acute
inflammation may be submitted to cytology when an underlying pathology is
clinically suspected. In addition to the acute inflammation, urothelial cells may
display reactive changes characterized by cellular and nuclear enlargement, a
normal nucleus/cytoplasm ratio, and prominent nucleoli.
D. Trichomonas is commonly identified in cervical-vaginal samples, and occasion-
ally it is identified in urine samples from female and male patients. Because men
are often asymptomatic carriers, the infection is often initially detected by urine
cytology. This protozoan is small (15 to 50 J.{.m) and pear shaped and has a
small nucleus, eosinophilic cytoplasmic granules, and flagella. Flagella are often
difficult to visualize in cytologic samples (e-Fig. 22.56).
Ill. NEOPLASMS
A. Urothelial carcinomas can be divided into the following groups: low-grade neo-
plasms composed of normal-appearing urothelial cells and high-grade neo-
plasms composed of overtly abnormal urothelial cells. Urine cytology has a
poor sensitivity (""30%) in diagnosing low-grade urothelial neoplasms (Acta
Cytol. 1996;40:676). Papillary urothelial dusters in a voided sample, even in
the absence of nuclear atypia, can be a manifestation of low-grade papillary
neoplasm (e-Fig. 22.57). The sensitivity of urinary cytology is approximately
from 40% to 80%, and the specificity >80%, with improved performance for
high-grade urothelial neoplasms (Urol Clin North Am. 2000;27:25). High-grade
urothelial carcinoma cells exhibit nuclear enlargement, a high nucleus/cytoplasm
ratio, and nuclear hyperchromasia with a coarse chromatin pattern (e-Fig.
22.58); in addition, the cells have a tendency to be arranged singly or in small,
poorly cohesive clusters.
One particularly promising method as an adjunct to urinary cytology is flu-
orescence in situ hybridization (FISH) to detect chromosomal abnormalities,
specifically the commercially marketed UroVysion FISH test (Abbott Labora-
tories, Abbott Park, IL). The test uses FISH probes to detect aneuploidy of
chromosomes 3, 4, and 17, and loss of 9p21, in voided urine specimens fixed
on glass slides. The approach has a sensitivity of 72% and specificity of 83%,
and studies with paired data have shown that the UroVysion test has a better
performance than cytology (Urol Oncol. 2008;26:646), although when super-
ficial cancer cases are excluded the differences between UroVysion and routine
cytology almost disappear. Most laboratories employ the FISH-based method in
conjunction with cytology, an approach that can likely be used to lengthen the
interval between surveillance cystoscopy in evaluation of patients with urothe-
lial carcinoma.
B. Squamous cell carcinoma can occur as a primary bladder tumor or can arise
in adjacent organs, such as the uterine cervix, and extend into the bladder.
A more common scenario is the presence of a squamous component within a
conventional urothelial carcinoma. Cytologic features that indicate the presence
of squamous differentiation include neoplastic cells with a variable amount
of dense and eosinophilic cytoplasm, pyknotic nuclei, and bizarre cell shapes
(e-Fig. 22.59). Parakeratotic and/or anucleated cells are often seen in the back-
ground. Poorly differentiated squamous cell carcinoma that is nonkeratinizing
may be confused with high-grade urothelial carcinoma.
C. Adenocarcinoma of the bladder can be primary or metastatic. Although most
(87%) of adenocarcinomas ofthe bladder can be identified as malignant by urine
cytology, only 67% will be additionally classified as adenocarcinoma (Cancer
Cytopathol. 1998;84:335). Columnar or cuboidal cell shapes, with cytoplas-
mic vacuoles within the neoplastic cells, are cytologic features that support the
diagnosis of adenocarcinoma.
Urethra
Souzan Sanati and Peter A. Humphrey
I. NORMAL ANATOMY. The male urethra is divided into three anatomic regions: pro-
static (bladder neck to apex of the prostate), bulbomembranous (apex of the
prostate to inferior surface of urogenital diaphragm), and penile (inferior sur-
face of urogenital diaphragm to the urethral meatus). The prostatic portion is
lined by urothelium, the bulbomembranous portion is lined by pseudostratified or
stratified columnar epithelium, and the penile portion shows a transition from the
stratified columnar epithelium at its origin to squamous epithelium at the meatus.
The female urethra is lined by urothelium in the proximal one-third and squamous
epithelium in the distal two-thirds.
The urethra has associated periurethral glands. Skene glands are present in
females and are concentrated distally. Bulbourethral (Cowper glands) and glands
of Littre, located in the bulbomembranous portion and along the penile urethra,
respectively, are present in males.
A. Urethroscopic biopsy tissue samples should be entirely submitted for histologic
examination, and three levels should be examined.
B. Surgical excision of urethral carcinoma. For men, the type of surgery is depen-
dent on tumor location and extent and includes transurethral resection (TUR),
local segmental excision, partial or radical penectomy, and cystoprostatectomy.
TUR chips should be submitted in their entirety. Segmental excision specimens
should be sampled to include sections of the proximal and distal margins and
area of deepest growth. Urethrectomy (primary or secondary) involves stripping
of all or part of the urethra with preservation of the penis and is performed for
patients with primary urethral carcinoma or secondary involvement by bladder
carcinoma; sampling should include sections of the proximal and distal margins
and area of deepest growth. Gross processing of penectomy and cystoprostate-
ctomy specimens is covered in Chapters 30 and 29, respectively.
For women, local excision of the distal urethra and adjacent vaginal wall
is often sufficient surgical therapy for carcinoma of the urethra; sections of the
mass, and urothelial, radial soft tissue, and vaginal mucosal margins should be
submitted. For proximal urethral cancer in women, cystourethrectomy (ante-
rior exenteration, with excision of part or all of the vagina) is often necessary;
sections of the mass demonstrating relationships with adjacent structures and
depth of invasion, grossly uninvolved urethra and urinary bladder, and ureteral
and radial soft tissue margins should be submitted.
Ill. DIAGNOSTIC FEATURES OF BENIGN DISEASES
A. Congenital anomalies
1. Urethral valves are mucosal folds lined by normal urothelium that project
into the urethral lumen causing obstruction, hematuria, or inflammation.
Posterior urethral valves are usually seen in men and are associated with
bladder neck hypertrophy.
2. Diverticula are invaginations of urethral mucosa usually seen in women as
a result of infection, trauma, or obstruction. They are lined by urothelium
that may undergo squamous or glandular metaplasia (e-Fig. 23.1).*
399
3. Fibroepithelial polyp is a congenital anomaly usually involving the posterior

urethra of male infants and young boys. It consists of a fibrous connective
tissue stalk lined by urothelium.
B. Inflammation and infection
1. Urethritis is an inflammatory response in the urethra that is usually secondary
to sexually transmitted diseases. Diagnosis is made by examination of a ure-
thral smear that shows neutrophils. Polypoid urethritis is usually seen in the
prostatic urethra near the verumontanum and is the result of inflammation
that induces multiple polypoid lesions with edematous stroma, distended
blood vessels, and chronic inflammation (e-Fig. 23.2).
2. Caruncle is a pedunculated or sessile polypoid inflammatory mass in the
distal urethra in postmenopausal women showing a mixed inflammatory
infiltrate with rich vascularity (e-Fig. 23.3A and B).
3. Malakoplakia is a rare urethral granulomatous inflammatory process, show-
ing histiocytes containing characteristic Michaelis-Gutmann bodies, more
commonly seen in women.
4. Condyloma acuminatum of the urethra is caused by human papilloma virus
(HPV), usually serotypes 6, 11, 16, and 18. It can primarily involve the ure-
thra but more commonly arises by direct extension from similar lesions in
adjacent sites including the external genitalia, perineum, and anus. Histolog-
ically, there is a flat or polypoid proliferation of squamous epithelium (e-Fig.
23.4) with koilocytic atypia. Multiplicity and recurrence are common.
c. Metaplasia
1. Squamous metaplasia can occur as a response to chronic inflammatory insults
secondary to infection, diverticula, calculi, or instrumentation.
2. Urethritis cystica and glandularis are small cysts lined by urothelial and glan-
dular cells, respectively.
3. Nephrogenic adenoma (metaplasia) is rare in the urethra. It occurs at the site
of previous damage, often related to a previous surgical procedure. Micro-
scopically, there is a proliferation of tubular and papillary structures, some-
times with cystic change, lined by flattened to cuboidal to hobnail cells with
bland nuclear features. Some cases may be due to implantation and growth
of tubular epithelial cells shed from the kidney.
D. Hyperplasia
1. Urothelial hyperplasia is a reactive thickening of cytologically bland urothe-
lium and can be flat or papillary. In the papillary form the mucosa can be
undulating but still lacks a well-developed fibrovascular core.
2. Prostatic urethral polyp is seen in the verumontanum in young men and con-
sists of benign prostatic tissue arranged in a polypoid or papillary configu-
ration, with projection into the urethral lumen.
IV. DIAGNOSTIC FEATURES OF NEOPLASTIC DISEASES
A. Benign neoplasms are exceedingly rare and have the same appearance as in the
urinary bladder.
1. Benign epithelial neoplasms include villous adenoma, squamous papilloma,
and urothelial (including inverted) papilloma.
2. Benign mesenchymal neoplasms include leiomyoma and hemangioma.
B. Malignant neoplasms originating in the urethra are very rare and are carci-
nomas in the vast majority of cases (Semin Diagn Pathol. 1997;14; Urol-
ogy. 2006;68:1164). Compared with urinary bladder carcinomas, urethral
carcinomas are more often found in women; a much higher percentage are
squamous cell carcinomas; there is a greater percentage of high-grade and
high-stage tumors; and there is a poorer prognosis. The histologic type of pri-
mary urethral carcinomas corresponds to the anatomic site of origin in the
urethra. In general, proximal neoplasms (prostatic urethra in men; proximal
one-third in females) tend to be urothelial carcinomas, whereas distal tumors
Chapter 23 • Urethra I 401
(bulbomembranous or penile in men; distal two-thirds in women) are frequently

squamous cell carcinomas. Often it is difficult to ascertain the precise site of
origin of a urethral carcinoma due to its infiltrative growth and destruction of
normal cells. For primary and secondary urothelial neoplasia, the 2004 World
Health Organization (WHO) urinary bladder classification is used for typing
purposes (Table 22.1).
1. Squamous cell carcinoma is the most common urethral carcinoma in both
sexes. HPV infection has a significant role in its etiology; 30% of squamous
cell carcinomas in men and 60% in women test positive for high-risk HPv,
although molecular HPV typing is not necessary for diagnosis. Grossly, squa-
mous cell carcinoma can be verruciform or grayish-white and scirrhous, with
necrosis (e-Fig. 23.5A and B). Microscopically, most are moderately differ-
entiated and deeply invasive. Squamous cell carcinomas of the distal penile
urethra frequently invade the corpus cavernosum; more proximal tumors
in men may penetrate directly into the urogenital diaphragm, prostate, rec-
tum, and bladder neck. In women, tumors arising in the distal third of the
urethra are commonly low-grade squamous cell carcinoma (e-Fig. 23.6) or
verrucous carcinomas.
2. Urothelial carcinoma rarely primarily involves the urethra but more com-
monly results from secondary involvement by bladder carcinoma; so-called
secondary involvement can represent direct extension, multifocal disease,
and/or lymphovascular invasion. Grossly, carcinoma in situ (CIS) can
be erythematous and/or ulcerative; carcinomas can be papillary, nodular,
ulcerative, and/or infiltrative. Histopathologically, primary or secondary
urothelial tumors can present as pure as, noninvasive papillary neoplasia
(e-Fig. 23.7A and B), or invasive carcinoma, with or without a papillary
component. In men there is a propensity for high-grade carcinoma (CIS or
invasive) to involve the prostate, which can represent in situ duct/acinar
spread and/or stromal-invasive disease. In women, CIS can cancerize subu-
rethral glands, which can mimic invasion.
3. Adenocarcinoma is usually seen in the proximal urethra and can originate in
a diverticulum. Grossly, the tumor is often infiltrative, with or without an
exophytic component, with mucinous, gelatinous, and/or cystic cut surfaces
(e-Fig. 23.8A and B). Microscopically, glandular metaplasia, and urethritis
cystica and glandularis, is frequently seen in adjacent epithelium. In women,
the most common subtype is clear cell adenocarcinoma (40% of cases). In
men, enteric, colloid, or signet-ring histomorphological features are common
but the clear cell type is rare.
a. Clear cell adenocarcinoma is a rare tumor that is almost always found in
women. Histologically, it is similar to clear cell adenocarcinoma of the
vagina or uterus. A distinct feature of this tumor is pattern heterogeneity
within the same neoplasm with solid, tubular, tubulocystic, micropapil-
lary, or papillary structures lined by hobnailed cells (e-Fig. 23.9A and B).
Necrosis and mitoses are frequent. The neoplastic cells are uniform and
ovoid with well-defined borders and amphophilic, acidophilic, or clear
cytoplasm containing glycogen. Nuclei are large and hyperchromatic with
prominent nucleoli. Clear cell adenocarcinoma should be differentiated
from nephrogenic adenoma (metaplasia) which lacks an infiltrative pat-
tern, sheet-like growth, mitotic figures, necrosis, and significant nuclear
atypia (Hum Pathol. 2010;41:594).
b. Non-clear cell adenocarcinoma includes enteric, mucinous (e-Fig. 23.10A
and B), signet-ring cell, and not otherwise specified subtypes. Extension
of adenocarcinoma from adjacent organs should be excluded.
4. Paraurethral gland carcinomas usually develop in paraurethral (Skene
glands in females; glands of Littre in males) and bulbourethral (Cowper)
402 I SECTION v. URINARY TRACl
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Chapter 23 • Urethra I 4 03
glands. Establishing the origin of adenocarcinoma can be difficult because

of mucosal ulceration and obliteration of landmarks by the time of
diagnosis.
5. Other rare carcinomas include small cell carcinoma, adenosquamous carci-
noma, sarcomatoid carcinoma, and lymphoepithelioma-like carcinoma.
6. Melanoma. The urethra is the most common site of primary melanoma of
the genitourinary tract (Am J Surg Pathol. 2000;24:785), but secondary
involvement as a result of spread from the glans penis or vulvar lesions is
more common. Amelanotic melanoma is commonly seen in this location.
Evaluation of margin status and the absence of skip lesions are important
in determining the prognosis (e-Fig. 23.11).
7. Hematopoietic neoplasms. Case reports exist of non-Hodgkin lymphoma,
mucosal associated lymphoid tissue (MALT) lymphoma, and plasmacytoma
in the urethra.
V. HISTOLOGIC GRADING OF URETHRAL CARCINOMA. For squamous cell carcinoma and
adenocarcinoma, well, moderately, or poorly differentiated grades may be applied.
Urothelial carcinomas are graded as low or high grade.
VI. PATHOLOGIC STAGING OF URETHRAL CARCINOMA is presented in Table 23.1. Patho-
logic stage and tumor location are the most significant pathologic prognostic indi-
cators for urethral carcinoma. Proximal tumors have a worse outcome.
VII. REPORTING URETHRAL CARCINOMA. The pathology report of a urethral malignancy
should include the histologic type, histologic grade, anatomic location (proximal
vs. distal), extent of invasion (depth and involvement of adjacent anatomic struc-
tures, if resected), presence or absence of lymphovascular space invasion and per-
ineural invasion, and pathologic tumor, node, metastasis (TNM) staging for resec-
tion specimens (Arch Pathol Lab Med. 2010;134:345). The status of the margins
should also be indicated, if applicable, including identification of the number and
location of positive sites. Any additional findings such as inflammation, metapla-
sia, presence of a diverticulum, or presence of a stricture should also be reported.
SECTION VI
Endocrine System
Thyroid
Changqing Ma, James S. Lewis Jr.,
and Rebecca D. Chernock
I. NORMAL ANATOMY. The thyroid gland is a bilobed organ in the lower neck sur-
rounding, and in intimate contact with, the trachea. It consists of right and left
lobes connected by a small isthmus in the midline (Fig. 24.1). Two functioning
cell types, follicular cells and C (calcitonin) cells, comprise the thyroid follicle. The
follicular component develops from invaginating tissue from the tongue base (fora-
men cecum) at 5 to 6 weeks gestation. Through differential embryonic growth and
migration, the bilobed gland assumes its definitive location in the neck, and the thy-
roglossal duct, the structure along the path of its downward migration, undergoes
atrophy. The C-cells, which comprise 0.1% or less of the total thyroid cell mass,
originate from the ultimobranchial body that develops from the fourth branchial
pouches; they are distributed in a density gradient, being most prominent in the
upper lobes.
The normal gland has a relatively firm consistency, is light brown, and lacks any
obvious nodules. Microscopically, it consists of follicles lined by low cuboidal bland
epithelial cells. This follicular epithelium surrounds a central core of eosinophilic
colloid. In normal glands, the follicles are relatively consistent and regular in size
(e-Fig. 24.1)."' The C-cells lie within the follicular epithelium and are invested by
the basement membrane. They are inconspicuous in normal thyroid.
A. Biopsy. Tissue needle biopsies of the thyroid gland are only very rarely performed
because fine needle aspiration is technically easy to perform, has low morbidity,
and is effective for triaging lesions for further management (see the section on
Cytopathology).
B. Resection. Hemithyroidectomy and total thyroidectomy are common proce-
dures for management of thyroid disease. The specimens from both types of
procedure are handled in a similar manner. The gland is oriented, if possible, on
the basis of the anatomy alone or on markings provided by the surgeon. It is then
measured and weighed. The surface is inspected for any disruptions or attached
soft tissue. Small nodules of tissue situated in the adjacent tissues may repre-
sent lymph nodes, parathyroid glands, or sequestered thyroid in cases of nodu-
lar hyperplasia. The gland should be inked, sectioned in an axial plane (bread
loafed) at intervals of 3 to 5 mm, and described, clearly indicating masses and
their size(s), color, and consistency. For diffuse or inflammatory lesions, three
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Chapter 24 • Thyroid I 405
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lobe).
or four sections from each lobe should be submitted for microscopic exami-
nation, along with one of the isthmus. For a solitary, encapsulated mass, the
entire circumference of the capsule should be sampled, including the surround-
ing thyroid tissue because the distinction of adenoma from carcinoma relies on
features seen in the capsule. For a solitary, nonencapsulated or not completely
encapsulated mass, at least one section per centimeter should be submitted,
and some sections should demonstrate the closest inked margin (Fig. 24.1).
For multinodular glands (nodular hyperplasia), sections of each major nodule
should be submitted, including the edge and/or adjacent soft tissue margin. Any
attached perithyroidal soft tissue should be removed and sampled. Any lymph
nodes or parathyroid glands should be removed and sampled as well, although
oftentimes, the parathyroid gland(s) are first detected by microscopic examina-
tion (the number of parathyroid glands should be recorded in one of the final
diagnostic lines to highlight their presence).
Prophylactic thyroidectomy is currently recommended in infancy, early
childhood, adolescence, or early adulthood for patients with a germline RET
gene mutation. The gross specimen often consists of the thyroid gland with no
grossly identifiable lesions. Each lobe should be entirely submitted sequentially
from superior to inferior to allow thorough examination for C-cell hyperplasia
or medullary microcarcinoma.
A. Developmental. Residual thyroid tissue can be present anywhere along the path
of thyroid migration during development. Carcinomas can rarely develop out-
side of the thyroid gland in these residual tissue sites.
1. Pyramidal lobe. The most common development remnant is a pyramidal
lobe, a linear projection of thyroid tissue from the isthmus pointing cranially
(Fig. 24.1).
2. Thyraalassal duct cyst Failure of closure of the thyroglossal duct commonly
results in a midline cyst known as a thyroglossal duct cyst (TDC). Excision
specimens generally consist of several nondescript fragments of tissue, and a
visible cyst is not always appreciated. Because remnants of the midline tract
pass through the hyoid bone, the latter should be identified. Failure to resect
the bone often results in persistence of the TDC. If a cyst is identified grossly, it
should be sampled in a few sections to document the type of epithelial lining,
which is often obscured by acute inflammation. Because IDCs develop from
a hollow tube, not all of them have thyroid tissue in their walls (only ""5%
406 I SECTION VI: ENDOCRINE SYSTEM
do on routine sections, and only about 40% do even on serial sectioning).

Otherwise, microscopically, these lesions have a squamous or respiratory-
type lining epithelium without a muscular wall (e-Fig. 24.2). TDCs can get
infected and inflamed with associated granulation tissue.
3. Lingual thyroid. Thyroid tissue can be present at the base of the tongue at
the site of developmental invagination, referred to as lingual thyroid. Micro-
scopically, the thyroid tissue is normal.
B. Inflammation (thyroiditis)
1. Acute thyroiditis is caused predominantly by bacteria but rarely can have a
fungal origin. It is usually seen in the setting of neck trauma or as spread from
infection of adjacent structures. Microscopically, the gland is infiltrated by
neutrophils with microabscesses and necrotic foci. Organisms can often be
seen with or without special stains. Patients usually recover with antibiotics.
2. Subacute thyroiditis: de Quervain thyroiditis. Technically referred to as granulo-
matous thyroiditis or subacute thyroiditis, de Quervain thyroiditis is a disease
primarily of women and is thought to be due to systemic viral infection. There
is linkage to HLA type Bw35. Patients present with fevet; malaise, and neck
pain. The disease consists of three phases: hyperthyroidism, hypothyroidism,
and recovery.
Grossly, the gland is usually asymmetrically enlarged and slightly firm.
Microscopically, in the hyperthyroid phase, there is disruption of the follicles,
with depletion of colloid and associated acute inflammation with microab-
scesses. Aggregates of neutrophils in the follicles are a characteristic feature.
Multinucleated giant cells are rare. In the hypothyroid phase, the follicu-
lar epithelium may be very scarce, and there is a florid mixed inflamma-
tory response with lymphocytes, plasma cells, and multinucleated giant cells.
Finally, in recovery, there is regeneration of follicles with fibrosis. Virtually
all patients recover thyroid function after a few months without specific
treatment.
3. Chronic thyroiditis
a. Focal lymphocytic thyroiditis. Also termed nonspecific thyroiditis or focal
autoimmune thyroiditis, this is a common disorder usually discovered
incidentally in surgically excised thyroids, and is found in 25% to 60% of
patients in autopsy studies, and is most common in older women. Patients
are asymptomatic but often have a low level of antithyroid antibodies.
There are no significant gross findings. Microscopically, focal lympho-
cytic thyroiditis consists of aggregates of lymphocytes, occasionally with
germinal centers, between the follicles (e-Fig. 24.3). The lymphocytes can
infiltrate follicles, but there is no significant destruction or damage. The
disease is not progressive.
b. Hashimoto thyroiditis. Hashimoto thyroiditis, or chronic lymphocytic thy-
roiditis, is an autoimmune disease with thyroid enlargement and circulat-
ing antithyroid antibodies. It occurs most often in middle-aged women and
is caused by autoantibodies against thyroglobulin (Tg) and thyroid per-
oxidase (TPO) that lead to inflammation and destruction of the follicles.
There is a familial association, and Hashimoto thyroiditis is often asso-
ciated with other autoimmune diseases. There is also an association with
HLA types D R3 and D R5. Patients can present with a hyperthyroidism or
hypothyroidism and are typically in their sixth decade. Ultrasound shows
an enlarged gland with a hypoechogenic pattern, and anti-Tg and anti-
TPO antibodies are present in approximately 60% and 95% of cases,
respectively.
Grossly, the thyroid is diffusely enlarged and firm with a mildly nodu-
lar surface. On sectioning, lobulation is accentuated due to fibrosis, and
the gland is tan-yellow or off-white rather than the usual brown due to
the abundant lymphoid tissue. Microscopically, there are sheets of lym-

phocytes and plasma cells with abundant germinal centers. Florid inflam-
mation often renders the residual follicles and follicular epithelium incon-
spicuous. The follicles are atrophic with minimal colloid. Characteristic
Hiirthle cell change is present, consisting of large cells with abundant,
granulat; eosinophilic cytoplasm and large, round nuclei with vesicular
chromatin and prominent nucleoli (e-Fig. 24.4). The inflammatory infil-
trate may extend into the perithyroidal soft tissue causing adherence of the
gland at the time of surgery. Finally, there may be fibrous septa between the
lobules, and the follicular epithelium may develop squamous metaplasia.
A number of variants of Hashimoto thyroiditis occur, including fibrous
(sclerosing), fibrous atrophy, juvenile, and cystic forms. Both of the fibrous
variants show the same histologic findings, namely retention of a lobu-
lated pattern with extensive and severe fibrosis. Atrophic follicular cells
are more scattered but show Hiirthle cell change, and there is abundant
chronic inflammation with germinal centers. In the fibrous variant, the
gland is markedly enlarged throughout. In the fibrous atrophy variant, the
gland is small and shrunken. Both of the latter variant forms are associated
with marked hypothyroidism and high titers of antithyroid antibodies.
The differential diagnosis for Hashimoto thyroiditis includes Riedel
thyroiditis and lymphoma. The fibrous variant of Hashimoto thyroiditis
may simulate Riedel thyroiditis with an enlarged fibrotic gland. However,
the fibrosis is typically limited to the gland itself, whereas in Riedel thy-
roiditis there is severe adherence of the fibrotic gland to the neck soft tis-
sues. The dense inflammation in Hashimoto thyroiditis can simulate lym-
phoma, and most thyroid lymphomas do arise in the setting of Hashimoto
thyroiditis. The distinction lies in the absence of sheets of atypical lym-
phocytes, the lack of a strikingly prominent lymphoepithelial pattern, and
the lack of clonality of the lymphocytes by flow cytometry or molecular
studies.
The issue of whether there is an increased risk of papillary carcinoma in
Hashimoto thyroiditis is a controversial topic. Whether this is the case or
not, the nuclear features of the epithelium can simulate those of papillary
carcinoma (including nuclear crowding, clear chromatin, and occasional
nuclear grooves) and develop frequently in the inflamed follicular epithe-
lium. A true carcinoma arising in Hashimoto thyroiditis should stand
out sharply from the neighboring follicular epithelium, as in uninflamed
thyroid.
c. Graves disease. Graves disease is also known as diffuse hyperplasia and is
an autoimmune condition resulting in excess thyroid hormone production.
It causes the majority of cases of spontaneous hyperthyroidism, occurs
most often in the third and fourth decades, and is 5 to 10 times more
common in women. Thyroid-specific autoantibodies, specifically thyroid-
stimulating immunoglobulin (TSI), are present in Graves disease patients.
TSI binds to and stimulates thyrocytes through the thyroid-stimulating
hormone (TSH) receptor, resulting in thyrotoxicosis. Patients also develop
a characteristic infiltrative ophthalmopathy with proptosis.
Grossly, the thyroid gland is diffusely and symmetrically enlarged.
Posttreatment or after long-standing disease, the gland may be some-
what nodular or fibrotic. Microscopically, the gland typically shows a
low-power lobular accentuation due to increased septal fibrous tissue.
Inflammation varies from none, to patchy lymphocytic, to lymphocytic
with formation of germinal centers. The follicular epithelium has increased
amounts of cytoplasm, is convoluted and irregular, and often assumes an
almost papillary appearance. Colloid is decreased or absent and typically
has a "scalloped" appearance from dearing at the interface with the fol-
licular epithelium (e-Figs. 24.5 and 24.6). Treated cases have a variable
morphology and often have cellular nodules mimicking adenomas. After
radioactive iodine treatment, there can be marked nuclear atypia. The dif-
ferential diagnosis includes papillary carcinoma when the stellate outlines
of follicles resemble papillae. The lack of cytologic changes of papillary
carcinoma and the diffuse gland involvement are keys to the correct diag-
nosis of Graves disease.
d. Riedel thyroiditis. A peculiar form of fibrosing disease, Riedel thyroiditis
is a chronic thyroiditis of unknown etiology which is more common in
women, occurs most commonly in the fifth decade, and is commonly
associated with fibrosing disease at other sites such as the mediastinum,
retroperitoneum, and lung. Patients present with firm thyroid enlargement
and local symptoms such as dysphagia, stridor, or dyspnea. Recurrent
laryngeal nerve or sympathetic trunk involvement can lead to hoarseness
or Horner syndrome; compression of the large vessels can lead to superior
vena cava syndrome. Most patients are euthyroid at presentation, but
many subsequently develop hypothyroidism.
Grossly, the thyroid gland is usually received in irregular pieces because
the fibrosis makes it difficult to remove surgically. It is tan to white, firm,
and may have attached muscle. Microscopically, the characteristic find-
ing is dense and hypocellular eosinophilic fibrous tissue with scattered
and patchy aggregates of lymphocytes, plasma cells, neutrophils, and
eosinophils, without germinal centers or granulomas. Rare entrapped and
atrophic thyroid follicles are seen without Hiirthle cell change. A charac-
teristic finding is small veins with infiltrating lymphocytes and myxoid inti-
mal thickening (features of occlusive vasculitis). Marked fibrosis extends
into and involves the surrounding soft tissue as well.
The differential diagnosis includes hypocellular anaplastic thyroid car-
cinoma and fibrous or sclerosing Hashimoto thyroiditis. The lack of necro-
sis or markedly atypical cells rules out anaplastic carcinoma. The lack of
Hiirthle cell change and germinal centers, and the profound degree of
perithyroidal fibrosis, rules out fibrous Hashimoto disease.
C. Nodular hyperplasia. Nodular hyperplasia (clinically termed multinodular goiter)
is an extremely common disorder of the thyroid gland. It is the enlargement of
the gland with varying degrees of nodularity. The pathogenesis is complex but
has been classically related to iodine deficiency with impaired thyroid hormone
synthesis and subsequent TSH stimulation. With supplementation of iodine in
the diet, nodular hyperplasia has also been related either to excess iodine intake
with impaired organification or to genetic factors. It is much more common in
women than men and typically presents in middle age as asymptomatic enlarge-
ment. Large goiters, however, can cause dysphagia, hoarseness, or stridor and
can extend into the upper mediastinum. A small percentage of patients present
with hyperthyroidism (toxic multinodular goiter).
Grossly, the thyroid gland is diffusely but irregularly enlarged. Some glands
can attain a weight of several hundred grams. Parasitic nodules that have only a
tenuous connection to the gland can develop. On sectioning, the nodules may be
semitranslucent, glistening, fleshy, red-brown, tan, and solid or, more commonly,
show varying degrees of degeneration with cystic change, hemorrhage, fibrosis,
and calcification. Heterogeneity of the nodules is typical (e-Fig. 24.7).
Microscopically, nodular hyperplasia is also heterogeneous. The common
appearance is nodules composed of variably sized follicles (e-Fig. 24.8). The
nodules may be quite cellular with tightly packed follicular epithelium and lit-
tle colloid, resembling follicular adenomas or carcinomas. Around cystic areas
there is often fibrosis with variably sized foci of dystrophic calcification and
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Chapter 24 • Thyroid I 41 1
follicular adenomas (microscopic evidence of capsular or vascular invasion

is necessary to render a diagnosis of carcinoma, as discussed below) In con-
trast, widely invasive follicular carcinomas are not encapsulated and show
extensive invasion of the surrounding gland and/or soft tissue beyond the
thyroid (e-Fig. 24.13). Follicular carcinomas are almost always unifocal and
have tan to brown, solid cut surfaces.
Microscopically, the growth pattern ranges from well-formed follicles
throughout, to solid, to trabecular. The nuclei of follicular carcinoma tend
to be round to oval with granular chromatin (e-Fig. 24.14). Mitotic activity
is present, but rarely brisk, and necrosis is lacking. The oncocytic variant
(Hiirthle cell) is the main histopathologic variant and compromises 20%
to 25% of all follicular carcinomas. To be classified as oncocytic variant,
a tumor should be composed of at least 75% oncocytic cells. These cells
have voluminous eosinophilic and granular cytoplasm and nuclei that are
characteristically vesicular with prominent, single nucleoli (e-Fig. 24.15A).
By immunohistochemistry, follicular carcinomas are positive for thy-
roglobulin, thyroid transcription factor-1 (TI'F-1), and low-molecular-
weight cytokeratins, but these are rarely necessary or useful for diagnosis
except occasionally to assess distant metastases as thyroid in origin. In addi-
tion, molecular rearrangements in the PPARy gene, most commonly with
fusion to the PAX8 gene, have been detected in 25% to 50% of cases, but
this finding is not currently useful in diagnosis or management.
The differential diagnosis includes follicular adenoma, medullary thy-
roid carcinoma (MTC), and poorly differentiated (insular) carcinoma. The
distinction of minimally invasive follicular carcinoma from adenoma lies in
capsular and/or vascular penetration alone (Table 24.2). Capsular invasion
is defined by the WHO as tumor penetration through the capsule not caused
by previous fine-needle aspiration. This invasion needs to be definitive and
classically takes the form of "mushrooming" of the tumor outward (e-Fig.
24.14). Vascular invasion is defined by the WHO as the presence of intravas-
cular tumor cells either covered by endothelium or associated with thrombus
(e-Fig. 24.15B). Strict adherence to these criteria is necessary to assure that
the diagnosis of carcinoma is correct.
The distinction of follicular carcinoma from MTC is based on morphol-
ogy (MTCs are rarely follicular in their growth patterns and their nuclei
tend to be more spindle shaped) and immunohistochemical findings (neu-
roendocrine markers, carcinoembryonic antigen [CEA], and calcitonin are
positive in MTC but negative in follicular carcinomas, whereas immunohis-
tochemistry for thyroglobulin is positive in follicular carcinoma and negative
in MTC). Finally, poorly differentiated (insular) carcinomas can be similar to
widely invasive follicular carcinomas, but the former have cells with less cyto-
plasm, minimal follicular architecture, marked mitotic activity, and necrosis.
Unlike PTC, follicular carcinomas only uncommonly involve regional
lymph nodes. Instead, metastasis is via a hematogenous route to lung and
bone. In general, minimally invasive follicular carcinomas have an excellent
prognosis with an approximately 15% long-term mortality. There is evidence
that the presence and extent of vascular invasion may provide further prog-
nostic information for patients with minimally invasive follicular carcino-
mas. Carcinomas that lack vascular invasion have a better outcome ( <5%
long-term mortality), while the presence of four or more foci of vascular
invasion has been shown to correlate with increased tumor-related mortal-
ity. Widely invasive follicular carcinomas have a poorer long-term mortality
approaching 50%.
3. Poorly differentiated {insular} carcinoma. This type of thyroid carcinoma
shows evidence of follicular differentiation but fits morphologically and
biologically between well-differentiated and undifferentiated thyroid carci-

nomas (UTCs). Some prefer to group this variant under the follicular carci-
noma heading, but the WHO recognizes it as a separate entity.
Because the classification of poorly differentiated carcinoma is somewhat
controversial, epidemiologic data are difficult to obtain. In general, however,
it represents much <5% of thyroid carcinomas in the United States, but
between 4% and 7% in Italy and some Latin American countries. It is more
common in women, particularly after age 50. Most cases present as siz-
able asymptomatic masses, with or without pathologically enlarged regional
lymph nodes. There is occasionally a history of a long-standing mass with
recent, rapid growth.
Grossly, most tumors are large with solid gray to white nodules. Necrosis
is common. Extrathyroidal extension is present in some cases but is much less
common than in undifferentiated carcinomas (e-Fig. 24.16). The Turin crite-
ria for the histologic diagnosis of poorly differentiated carcinoma include (i) a
solid, trabecular or insular growth pattern, (ii) lack of well-developed nuclear
features of papillary carcinoma, and (iii) one of the following: convoluted
nuclei, tumor necrosis, or three or more mitoses per high power field (e-Figs.
24.17 to 24.20). Most cells have only a small amount of cytoplasm giving the
tumor a decidedly neuroendocrine look, although the tumor nuclei typically
do not have "salt and pepper" chromatin and often have prominent nucleoli
as well as convoluted nuclei (e-Figs. 24.17to 24.20). Occasional follicles with
colloid can be seen. It is not uncommon for well-formed capsules to be present
around some of the nodules. Vascular invasion is often a prominent feature.
Although rarely useful for routine diagnosis, by immunohistochemistry the
tumors are positive for thyroglobulin and TTF-1, although the thyroglobulin
reactivity may be focal.
The differential diagnosis includes MTC, the solid variant of papillary
carcinoma, and metastasis (particularly from a carcinoid tumor). The mean
5-year survival is approximately 50%.
4. Papillary carcinoma. PTC is defined by the WHO as a carcinoma showing
evidence of follicular differentiation and characterized by distinct nuclear
features. The terminology has become somewhat confusing because papillary
architecture, although very commonly present, is not necessary for diagno-
sis. PTC is the most common malignant thyroid tumor (representing 85% to
90% of differentiated thyroid carcinomas) and has been increasing in inci-
dence, but its prognosis is excellent (discussed later). The commonality of the
tumor relative to its indolent behavior makes the appropriate management
of PTC controversial. PTC occurs across all ages, and women are affected
four times more commonly than men. There is a close link with previous
radiation exposure, particularly in younger patients.
Grossly, most tumors are tan or white with infiltrative borders and a
firm, often gritty cut surface ( e-Fig. 24.21). Cystic change is relatively com-
mon, particularly in lymph node metastases, and the bulk of nodal metastatic
disease may far outstrip the volume of tumor in the thyroid gland. Micro-
scopically, PTC is diagnosed on the basis of a constellation of architectural
and cytologic features (Table 24.3). Architectural patterns include papillary,
trabeculat; micro- and macrofolliculat; solid, and cystic. The stroma around
the tumor is often fibrotic or sclerotic. In tumors with follicles, the colloid
is typically described as "bright" or dark red relative to the normal thyroid.
Another very typical feature is the presence of psammoma bodies (small,
round, laminated calcifications; e-Fig. 24.22). Psammoma bodies are not
pathognomonic of PTC but are an extremely typical feature. Distinct nuclear
features include marked crowding with overlapping of adjacent nuclei, pale
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At surgery, the tumor is usually large, ill-defined, and difficult to excise

due to extensive surrounding soft tissue invasion. Grossly, UTC is usually
fleshy and tan-white with areas of hemorrhage and necrosis. Microscopically,
it is typically composed of a variable admixture of spindle cells, epithelioid
cells, and giant cells. The cells have a moderate amount of eosinophilic cyto-
plasm and almost always show brisk mitotic activity with abundant apoptosis
(e-Fig. 24.27). There is often geographic necrosis, and typically the remain-
ing thyroid gland is obliterated by tumot: On thorough examination, a resid-
ual, well-differentiated carcinoma, either papillary or folliculat; is sometimes
identified. Predominantly spindled tumors often mimic true sarcomas. The
prognosis for UTC is very poor with a median survival of< 6 months despite
aggressive surgery, radiation, and chemotherapy.
Immunohistochemistry for cytokeratin is positive in approximately 80%
of cases, and for epithelial membrane antigen in 30% to 50%. These stains
are useful for diagnosis of tumors in which no obvious carcinomatous dif-
ferentiation is present on H&E in order to confirm that the neoplasm is a
carcinoma rather than a high-grade sarcoma. Howevet; lack of staining with
epithelial markers does not completely exclude the diagnosis of UTC. TfF-1
and thyroglobulin are typically negative in UTC. Nuclear reactivity for PAX8
has recently been reported in approximately 80% ofUTC and may be useful
for diagnosis.
6. Medullary thyroid carcinoma. MTC is a carcinoma with C-cell differentiation
and represents a neuroendocrine neoplasm of the thyroid. C-cells secrete
calcitonin, a peptide that causes increased renal excretion of calcium and
inhibits osteoclasts to prevent calcium liberation from bone.
MTC represents approximately 5% to 10% of thyroid tumors. Although
the majority of cases are sporadic, up to 20% may be familial and there is a
strong association with multiple endocrine neoplasia (MEN) type 2, which
must always be considered in the work-up of patients with MTC. The clinical
presentation of MTC strongly depends on the familial or nonfamilial nature
of the tumor. Sporadic and non-MEN familial MTCs present in fifth or sixth
decades of life as a unilateral thyroid mass. A high percentage of patients
have associated cervical lymphadenopathy. As an index case, familial MTCs
associated with MEN type 2A are frequently detected at a younger age (mean,
third decade) and are multicentric and bilateral. When associated with MEN
type 2B, MTC usually occurs in even younger patients (i.e., in childhood
or adolescence). Virtually all patients with MTC have an elevated serum
calcitonin level.
Grossly, MTC is circumscribed but unencapsulated and is tan-yellow to
white. Sporadic tumors are solitary, whereas familial tumors are often multi-
focal and bilateral. Microscopically, MTC shows great variability. The typical
features are sheets, nests, or trabeculae of polygonal, round to oval cells with
moderate to generous amounts of eosinophilic to amphophilic cytoplasm.
The nuclei are round to oval with a coarse and somewhat granular neuroen-
docrine chromatin. Nucleoli are usually not prominent (e-Fig. 24.28). A great
number of variants have been described including papillary or pseudopap-
illary (e-Fig. 24.29), plasmacytoid, glandulat; giant cell, spindle cell (e-Fig.
24.30), small cell or neuroblastoma-like, paraganglioma-like, and oncocytic.
Another characteristic feature of MTC is the presence of amyloid, which is
formed from the deposition of calcitonin in the peritumoral stroma (e-Fig.
24.28). The amyloid is positive by Congo red staining.
Reactive, or nonneoplastic, C-cell hyperplasia occurs in aging and a
number of other thyroid diseases, most notably hyperparathyroidism and
lymphocytic thyroiditis. Although the histology of C-cell hyperplasia is
somewhat controversial, a few generalities apply. First, reactive C-cell hyper-

plasia is usually unilateral and is not identifiable by H&E examination alone.
Second, if there are aggregates of C cells >50 in number, in nodules, bilater-
ally, or diffusely, neoplastic C-cell hyperplasia is diagnosed.
Neoplastic C-cell hyperplasia, a precursor lesion to MTC that is more
commonly seen in hereditary cases, is identified in the middle third to upper
third of the thyroid lobes where C-cells are preferentially located; it is charac-
terized by groups of enlarged, intrafollicular atypical C-cells that may begin
to obliterate the normal follicular epithelium (e-Figs. 24.31 and 24.32). Neo-
plastic C-cell hyperplasia progresses to medullary carcinoma when C-cells
extend through the follicular basement membrane into the stroma. Distinc-
tion between neoplastic C-cell hyperplasia and medullary microcarcinoma
(a focus of MTC <1 em in greatest dimension) can be difficult. Helpful
histologic features of microcarcinoma include: (i) nuclear pleomorphism,
(ii) expansile growth pattern with C-cell clusters spilling out of follicles,
(iii) sclerotic stroma, and (iv) amyloid deposition.
Immunohistochemistry is useful for diagnosis of MTC. C-cells and MTC
tumor cells are usually strongly positive for calcitonin, synaptophysin,
chromogranin-A, and CEA, but negative for thyroglobulin. Although the dif-
ferential diagnosis of MTC includes thyroid paraganglioma, intrathyroidal
parathyroid adenoma, follicular adenoma, follicular carcinoma, poorly dif-
ferentiated (insular) carcinoma, and oncocytic tumors, all of these tumors
can be differentiated from MTC by histology and immunohistochemistry.
MTC (whether sporadic, non-MEN familial, or in the context of MEN
type 2A or 2B) is strongly associated with activating point mutations of the
RET proto-oncogene. Thirty percent to 66% of sporadic MTCs are associ-
ated with somatic RET gene mutations; 2% to 9% of sporadic MTCs are
associated with de novo RET germline mutations. All cases of hereditary
MTC have germline RET mutations. Recent studies show that each specific
germline mutation of the RET proto-oncogene correlates with/predicts the
age of onset and aggressiveness of the MTC that occurs in that kindred. In
addition, clinical trials of tyrosine kinase inhibitors targeting the RET kinase
have shown promise in reducing tumor burden in metastatic disease.
The behavior of MTC is highly stage dependent. Patients who do not
have metastatic disease are usually cured by total thyroidectomy, and their
10-year survival approaches 100%. Lymph node metastases are very com-
mon, and common distant metastatic sites include the lungs, liver, and bone.
The overall10-year survival for patients with cervical lymph node metastases
is approximately 70% to 80%, and for patients with distant metastases 40%
to 50%. Because C-cell hyperplasia and MTC occur early in life in hered-
itary cases, prophylactic thyroidectomy and central lymph node dissection
should be performed as early as possible in MEN type 2B, although it may be
delayed somewhat in MEN type 2A and familial non-MEN MTC. Among
the different clinical settings in which it arises, the prognosis is best for MTC
in patients with non-MEN familial inheritance, slightly worse for those with
MEN type 2A or sporadic tumors, and worst for those patients with MEN
type 2B.
IV. PATHOLOGIC REPORTING OF THYROID CARCINOMA. American Joint Committee on
Cancer (AJCC) staging of thyroid carcinomas is quite different than for other organ
sites because the histologic type and age of the patient are critical for prognosis
(Table 24.4). Patients with papillary carcinoma that has not metastasized distantly
have an excellent prognosis, particularly those younger than 45 years of age, in
whom survival approaches baseline. Any nonepithelial tumor type is excluded from
staging.
Cytopathology of the Thyroid

Michael E. Hull and Julie Elizabeth Kunkel
I. INTRODUCTION. The advent of widespread use of thyroid FNA has dramatically

decreased surgical excision of abnormal clinical lesions for diagnostic purposes.
The sensitivity for the detection of lesions requiring surgical management is up
to 98%, but specificity lags. The recent introduction of the Bethesda System
for Reporting Thyroid Cytopathology (Am] Clin Pathol. 2009;132:658) has
made possible a more probabilistic approach to the cytology of the thyroid, on
the basis of cellularity, colloidal composition, architectural patterns, and nuclear
morphology.
II. NONDIAGNOSTICIUNSATISFACTORY
A. Six groups of well-visualized follicular cells, with each group containing ten or
more cells, preferably on one slide, constitute the minimal adequacy criteria for
thyroid FNA. Exceptions include paucicellular samples with marked cytologic
atypia; solid nodules with inflammation (i.e., Hashimoto thyroiditis, abscess,
or granulomatous inflammation) in which inflammatory cells predominate
over a very scant epithelial population; and colloid nodule, in which abundant
thick colloid is readily identified in the near-absence of follicular cells. Fail-
ure to meet the above criteria makes an aspirate nondiagnostidunsatisfactory
(e-Fig. 24.33).
B. 11Cyst fluid only" is an important subset of the nondiagnostic category. If only
macrophages and noncolloidal cyst fluid are present, and there is a solid clini-
cal/sonographic lesion, the assumption should be made that the lesion has not
been adequately sampled. In more reassuring clinical/sonographic cases, cyst
fluid only may be consistent with a benign process. It should be clear that "cyst
fluid only" requires careful contextual interpretation.
Ill. BENIGN. This is a category with high negative predictive value (0% to 3% malig-
nancy rate on follow-up). Smears generally show significant amounts of colloid
and variable degrees of cellularity. Benign follicular nodule is the most common
pathologic correlate, but the presence of inflammatory cells or fibrous stroma may
indicate a thyroiditis.
A. Benign follicular nodules, the most common diagnosis in thyroid aspirates, are
characterized by benign follicular cells and colloid in variable proportions,
variably sized follicular groups, Hi.irthle cells, and macrophages. Abundant
colloid and relatively low cellularity are reassuring features of benignity (e-
Figs. 24.34 and 24.35). However, since the pattern is seen in a constellation
of benign proliferations, the underlying process can be diagnosed definitively
only upon resection: nodular goiter, hyperplastic nodules, colloid nodules, and
nodules in the setting of Graves disease.
B. Thyroiditis
1. Acute thyroiditis is rarely sampled given its typical clinical presentation.
The aspirate shows predominantly neutrophils and necrotic debris with
scattered histiocytes and scant reactive follicular cells. Special stains and
culture are required for the identification of microorganisms.
2. Chronic lymphocytic (Hashimoto) thyroiditis. The aspirate is usually cellular
and consists of two major components: (i) a large population of polymor-
phous lymphocytes, plasma cells, tingible body macrophages: and (ii) scat-
tered, two-dimensional clusters of Hi.irthle cells with variable nuclear atypia
(e-Fig. 24.36). Occasional multinucleated giant cells can be identified. Nor-
mal follicular cells and colloid are scant (Diagn Cytopathol. 1994;11:141).
3. Subacute granulomatous (de Quervain) thyroiditis. The aspirate can
be hypocellular due to fibrosis. Granulomas consisting of cohesive
aggregates of epithelioid histiocytes with elongated and kidney-shaped

nuclei, granular chromatin, small nucleoli, and abundant pale cytoplasm
with ill-defined borders (e-Fig. 24.37) are the key diagnostic features.
Multinucleated giant cells and lymphocytes are usually present (Diagn
Cytopathol. 1997;16:214).
4. Riedel thyroiditis. The aspirate is markedly hypocellular, containing
microfragments of stroma with bland spindle-shaped cells. Other cellular
components are absent.
IV. ATYPIA OF UNDETERMINED SIGNIFICANCE/FOLLICULAR LESION OF UNDETERMINED
SIGNIFICANCE (AUSJFLUS}. AUS/FLUS is an equivocal category created in recogni-
tion of the fact that the cytologic pattern does not always allow discrimination
between a benign nodule and a more serious lesion such as a follicular neoplasm
or papillary carcinoma. The features in these aspirates are variable but generally
consist of only rare cytologically or architecturally abnormal follicular groups
in a background of scant colloid (e-Fig. 24.38). The architectural arrangement
of the follicular cells in the lesion is the key to correct classification (Cancer.
1993;71:2598). This designation should generally comprise 7% to 8% of a cytol-
ogy laboratory's thyroid interpretations and carries a 5% to 15% risk of malig-
nancy upon rebiopsy/resection. The recommended follow-up is reaspiration in 6
months; too-early reaspiration may lead to diagnostic difficulty due to resolving
hemorrhage and reparative changes (Cyto]ournal. 2008;5:6).
V. FOLLICULAR NEOPLASM/SUSPICIOUS FOR FOLLICULAR NEOPLASM (FNISFN). FN/SFN
yields richly cellular smears with both architectural and cytologic abnormalities,
consisting of nuclear crowding, overlap, microfollicle formation, and dispersed
single cells (e-Fig. 24.39). However, definitive features of papillary carcinoma are
not seen. Such a pattern brings follicular adenoma, follicular carcinoma, and the
follicular variant of papillary carcinoma into the differential diagnosis, thus
the lesion requires excision; the nonspecificity of the diagnosis underlies use of the
phrase "suspicious for follicular neoplasm" although FN and SFN are equivalent
monikers in the Bethesda system.
Most patients with this diagnosis will undergo lobectomy or complete thy-
roidectomy, although only 20% to 30% of lesions will be malignant upon resec-
tion ( Cyto]ournal. 2008;5:6). This lack of specificity is the impetus for investiga-
tion of ancillary techniques for the prediction of malignancy in these lesions, in
particular determination of the mutational status of the four genetic aberrations
present in the majority of cases of papillary and follicular carcinoma: point muta-
tions of BRAF and RAS, and rearrangements of RET-PTC and PAX8-PPARy
(Mol Cell Endocrinol. 2010;322:29). These investigations may eventually lead to
dual cytologic-genetic triage of patients for surgery and aid in pre- and postop-
erative planning (hemithyroidectomy vs. total thyroidectomy, necessity of node
dissection, radiation treatment, etc.).
VI. FOLLICULAR NEOPLASM, HURTHLE CELL TYPE/SUSPICIOUS FOR FOLLICULAR NEO-
PLASM, HURTHLE CELL TYPE (FNHCT/SFNHCT}. Hiirthle cells are present in both neo-
plastic and nonneoplastic processes (Am] Clin Pathol. 1993;100:231). Hi.irthle
cell neoplasms are cytologically distinct in that they yield hypercellular specimens
composed of an almost pure population of Hi.irthle cells (e-Figs. 24.40 and 24.41 ).
The Hiirthle cells are often dyshesive single cells with prominent nucleoli, and the
background contains an insignificant number of normal follicular cells, lympho-
cytes, and colloid. As is the case in other follicular neoplasms, the diagnosis of
Hi.irthle cell carcinoma is based on assessment of capsular and vascular invasion,
which cannot be evaluated in cytology specimens. It should also be noted, in fur-
ther similarity to FN/SFN, that papillary carcinoma is in the differential diagnosis
of FNHCT/SFNHCT.
VII. SUSPICIOUS FOR MALIGNANCY (SFM}. SFM describes malignant appearing features
similar to those in the "malignant" category (discussed later) but in an insufficient
quantity to allow a confident diagnosis of malignancy. Use of this diagnosis is at

the discretion of the pathologist and will certainly vary between laboratories and
pathologists on the basis of a number of factors including patient population,
cytologic preparations, aspiration technique, and so on. About 50% to 75% of
the lesions in this cytologic category are proven malignant at resection.
VIII. MALIGNANT. This category encompasses cytologic features diagnostic of any of the
malignant primary or metastatic tumors that may be seen in the thyroid. When
the criteria are appropriately applied, there are very few false positives.
A. Papillary thyroid carcinoma (PTC). The specimen is richly cellular, consisting
of large sheets with occasional papillary structures (e-Figs. 24.42 and 24.43).
The follicular cells are slightly enlarged and have crowded nuclei. The char-
acteristic nuclear features are fine open chromatin with nuclear membrane
prominence, small and peripherally located nucleoli, intranuclear cytoplas-
mic inclusions, and longitudinal nuclear grooves (e-Fig. 24.44). Other associ-
ated features include dense cytoplasm, "bubble gum"-like dense colloid, psam-
moma bodies (e-Fig. 24.45), and multinucleated giant cells (Diagn Cytopathol.
1991;7:462).
B. Hyalinizing trabecular adenoma is a controversial entity with great similar-
ity to PTC, including the characteristic nuclear features and RET-PTC rear-
rangements. Definitive diagnosis rests upon histologic examination (Am I Clin
Pathol. 1989;91:115).
C. Poorly differentiated (insular) carcinoma. The aspirate is richly cellular and com-
posed of monomorphic follicular cells with scant colloid. The follicular cells
are present predominantly as single cells, with only occasional crowded dus-
ters and microfollides (e-Fig. 24.46). The necrosis and mitosis seen in tissue
biopsy specimens are rare in smears. There are no distinct cytological features
to render definitive diagnosis (Diagn Cytopathol. 2001;25:325).
D. Anaplastic carcinoma. The aspirate is highly cellular and contains single cells
exhibiting marked nuclear atypia. Necrosis is common. The malignant cells
show diverse morphology, including epithelioid (e-Fig. 24.4 7), spindle-shaped,
and giant pleomorphic cells (Acta Cytol. 1996;40:953).
E. Medullary carcinoma typically generates highly cellular specimens, predomi-
nantly consisting of single cells and loose dusters of cells. The cell morphology
is variable, ranging from monotonous with little atypia, to highly atypical.
The characteristic features are mixtures of plasmacytoid and spindle-shaped
cells with characteristic salt-and-pepper chromatin (e-Fig. 24.48). Amyloid is
usually present, but its distinction from colloid is impossible without a Congo
red stain (Pathologica 1998;90:5).
F. Lymphoma. Most primary thyroid lymphomas develop in the background of
Hashimoto thyroiditis. Diffuse large B-celllymphoma (DLBCL) is the major
type, followed by extranodal marginal zone B-cell lymphoma of mucosa-
associated lymphoid tissue (Am I Surg Pathol. 2000;24:623). DLBCL exhibits
monotonous large atypical lymphoid cells with irregular nuclear contours,
vesicular chromatin, and single prominent nucleoli (immunoblast-like cells)
or multiple nucleoli (centroblast-like cells) (e-Fig 24.49). Marginal zone B-cell
lymphoma shows a heterogeneous population of lymphoid cells with a pre-
dominance of small lymphoid cells, and intermixed plasmacytoid cells and
immunoblasts; monocytoid cells with abundant pale cytoplasm are frequently
seen. Demonstration of light chain restriction by flow cytometry is critical for
diagnosis.
G. Metastatic malignancy. Metastasis to the thyroid is uncommon. The kidney
is the most common primary site of metastatic tumors, followed by lung and
breast. In a patient with a history of malignancy, the differential diagnosis for a
new thyroid nodule should include metastasis. Comparison of the aspirate with
the slides of the primary malignancy, together with immunostains, is critical

for diagnosis.
SUGGESTED READINGS
Adair C. Non-neoplasticlesions of the thyroid gland. In: Thompson LDR, ed. Endocrine Pathology.
New York: Churchill Livingstone; 2006.
DeLellis RA, Nikiforov YE. Thyroid and parathyroid glands. In: Gnepp DR, ed. Diagnostic Surgical
Pathology of the Head and Neck. Philadelphia: W.B. Saunders Elsevier Publishers; 2009.
Lloyd RV, Douglas BR, Young WF. Thyroid gland. In: King DW, ed. Fascicle #1-Endocrine
Diseases, First Series. Armed Forces Institute of Pathology Atlas of Non- Tumor Pathology.
Washington, DC: American Registry of Pathology; 2002.
Nikiforov YE, Biddinger PW, Thompson illR. Diagnostic Pathology and Molecular Genetics of
the Thyroid. Baltimore, MD: Wolters Kluwer Health/Lippincott William & Wilkins; 2009.
Thompson illR. Benign neoplasms of the thyroid gland. In: Thompson illR, ed. Endocrine Pathol-
ogy. New York: Churchill Livingstone; 2006.
Thompson illR. Malignant neoplasms of the thyroid gland. In: Thompson LDR, ed. Endocrine
Pathology. New York: Churchill Livingstone; 2006.
Parathyroid Glands
James S. Lewis Jr.
I. NORMAL ANATOMY. The endodermally derived parathyroid glands develop from

the third (inferior parathyroids) and fourth (superior parathyroids) pharyngeal
pouches. They produce parathyroid hormone (PTH) which acts to increase serum
calcium. They are normally found along the posterior surface of the thyroid gland,
but given their complex embryologic development, normal variations in location
range from within the substance of the thyroid gland, superiorly to the hyoid
bone, inferiorly into the mediastinum, within the thymus gland, or within the
pericardium. Furthermore, 2% to 7% of individuals have more than the usual four
glands.
Most normal parathyroid glands are from 0.3 to 0.6 em in largest dimension,
and the normal aggregate weight of all glands is 120 to 140 mg. They are brown to
yellow-brown, oval, and have a thin capsule. Histologically, they consist of chief (or
principal) cells and oxyphil cells. The former have round nuclei with granular chro-
matin and slightly eosinophilic to clear cytoplasm, and the latter have round nuclei
and more abundant brightly eosinophilic and granular cytoplasm (e-Fig. 25.1).*
Both types of cells are arranged in sheets, nests, and cords. Occasional pseudog-
landular or pseudoacinar foci with central eosinophilic proteinaceous material can
also be seen in the normal parathyroid. Oxyphil cells may also form small nodules
in adults. Intraparenchymal adipose tissue is a normal feature of the parathyroid
glands. It is usually scant in children but progressively increases with age to con-
stitute about 50% of the gland by the fifth decade, and plateaus beyond that time
(e-Fig. 25.2).
Parathyroid cells, both chief and oxyphil, are positive by immunohistochemistry
for PTH, chromogranin-A, and cytokeratins, and this immunoprofile is maintained
in virtually all pathologic processes. Parathyroid cells are negative for thyroglobulin
and thyroid transcription factor-1 (TfF-1 ).
A. Fine needle aspiration. Fine needle aspiration of parathyroid lesions is rarely
performed. The two exceptions are, first, when an adenoma arises within the
substance of the parathyroid gland and thus a biopsy is taken as part of the eval-
uation of a presumed thyroid lesion, and second, when parathyroid carcinoma
presents as a large neck mass.
B. Biopsy. Intraoperative biopsies (frozen sections) are frequently performed to
confirm that the excised tissue is parathyroid because lymph nodes, thymus,
thyroid, and fat may all be mistaken surgically for parathyroid tissue. The tissue
should be weighed, but no other special handling is required.
C. Excision. The initial step in the gross examination of parathyroid glands is
recording their weight and measurements in three dimensions. The glands should
be closely examined and if firm, ragged, or irregulat; should be inked around
their periphery. They should then be sectioned and their color and consistency
described. One section should be taken for histologic examination or, if the
gland is 2 em or larger, two to three sections should be taken.
422
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Chapter 25 • Parathyroid Glands I 4 25
1. Adenoma. Parathyroid adenomas are benign neoplasms composed of chief

cells, oxyphil cells, or a mixture of both. They occur in approximately
0.1% of the population and account for approximately 80% of cases of
hyperparathyroidism. They are more common in women and have a peak
incidence in the sixth and seventh decades of life. Adenomas are sometimes
(albeit rarely) familial. These cases are usually associated with MEN types 1
or 2, or the uncommon hyperparathyroidism-jaw tumor syndrome. Adeno-
mas involve a single gland (it is controversial as to whether patients with more
than one adenoma actually suffer from asymmetrical hyperplasia). Patients
present with signs and symptoms related to hypercalcemia (as detailed in
Section Ill.A.4 on parathyroid hyperplasia). Technetium is concentrated in
parathyroid tissue, so technetium sestamibi scans are often used to detect
and localize the abnormal gland.
Grossly, adenomas are rounded, encapsulated, and tan to reddish-brown,
with an average weight of 1 g (e-Fig. 25.9). On sectioning, they are soft
and homogeneous, although degeneration and cystic change may occur.
Microscopically, adenomas are well-circumscribed, thinly or nonencapsu-
lated masses with little stroma and no, or very minimal, fat, often with a
rim of normal appearing parathyroid gland. The cell population is usually
uniform and predominantly of one cell type, chief or oxyphil (e-Figs. 25.10
and 25.11). The cells are usually arranged in solid sheets, although pseudog-
landular (pseudoacinar or follicular) areas may be seen (e-Fig. 25.12), some-
times containing central eosinophilic material mimicking thyroid follicles.
The nuclei in chief cell adenomas are small, round, regulat; and hyperchro-
matic. The nuclei in oxyphilic adenomas are round and may have prominent
eosinophilic nucleoli. Nuclear pleomorphism can be seen although it is usu-
ally very localized (e-Fig. 25.13). Mitotic figures are scarce (1 or fewer per
10 high-power fields).
The differential diagnosis of an adenoma includes parathyroid hyperpla-
sia. Adenomas simply cannot be distinguished from hyperplasia without sam-
pling more than a single gland because the morphologic features of adeno-
mas overlap those of large hyperplastic glands. If only one gland is provided
for evaluation, it is best to diagnose "hypercellular parathyroid consistent
with adenoma" to reflect the fact that only one gland was excised, implying
that the other glands were not clinically abnormal and that intraoperative
PTI-1 monitoring likely confirmed a sufficient drop after the single gland was
excised. Adenomas can be distinguished from cellular nodules of thyroid
tissue (particularly from patients with nodular hyperplasia of the thyroid)
by finding convincing colloid in thyroid lesions, finding a rim of definite
parathyroid tissue in adenomas, or by immunostaining for thyroglobulin,
PTI-1, and/or TfF-1.
2. Carcinoma. Parathyroid carcinoma is rare and is estimated to be the cause of
<1% of hyperparathyroidism. The average age of patients is between 45 and
55 years, and the sex distribution is roughly equal. The etiology is unknown,
and there is no significant association with any of the familial syndromes that
cause other parathyroid disease. Most patients present with profound hyper-
parathyroidism and hypercalcemia (with serum calcium levels often> 16 mg
per dL with secondary nephrolithiasis, renal insufficiency, and bone involve-
ment with osteopenia and/or "brown tumors"), and most have nonspecific
symptoms such as weakness, fatigue, nausea, and depression.
Grossly, parathyroid carcinomas are usually large, poorly circumscribed,
and adherent to surrounding tissues, particularly the thyroid gland (e-Fig
25.14). They range from 1.5 to 6.0 em or larger, and average 6. 7 g. Their cut
surface is usually gray-white and firm. Microscopically, they usually have a
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Chapter 25 • Parathyroid Glands I 4 27
Cytopathology of the Parathyroid

Glands
Hannah R. Krigman
I. INTRODUCTION. The parathyroid glands are sampled infrequently by fine needle
aspiration (FNA) for diagnostic purposes since adjunct testing (including serum
calcium levels), radiolabeled imaging studies (sestamibi scans), and assessment of
aspirated fluid for parathyroid hormone levels generally obviate presurgical analysis
of parathyroid masses. However, FNA may be used to evaluate the presence of
parathyroid tissue in unusual locations, or to evaluate persistent hypercalcemia
following excision of parathyroid glands. In addition, parathyroid glands may be
incidentally sampled during assessment of thyroid nodules. Rarely, unanticipated
parathyroid adenomas are aspirated as thyroid or neck nodules.
II. CYTOLOGIC FINDINGS. Direct smears show predominantly two dimensional groups of
50 to 100 relatively evenly sized cells with granular cytoplasm. Three dimensional
groups, papillary fragments, and microacinar structures may also be seen (e-Fig
25.18). Oxyphilic epithelial change may be seen rarely. Nuclei are predominantly
round, with finely mottled chromatin (e-Fig 25 .19); sometimes nuclear molding and
overlap are present. Modest variation in nuclear size may be seen, but this feature
has no prognostic importance; mitotic activity is uncommon. The background often
contains stripped nuclei and single cells, macrophages, and colloid-like material.
Diff-Quik stained fresh smears may show fat vacuoles which are a helpful clue
since they are not typically seen in aspirates of thyroid tissue.
Cytologic features do not reliably discriminate among normal parathyroid tis-
sue, hyperplasia, adenoma, or carcinoma.
Differentiation of parathyroid from thyroid in FNA specimens can be problem-
atic. Microacinar groupings of parathyroid cells are an occasional source of error
in that they may be misinterpreted as follicular neoplasm/suspicious for follicular
neoplasm, especially in the setting of parathyroid adenomas. Follicular groups of
parathyroid cells can also be misinterpreted as thyroid follicular cells, although
intervening fat should suggest the correct diagnosis. In problematic cases, parathy-
roid origin can be confirmed with an immunostain for parathyroid hormone.
SUGGESTED READINGS
Absher KJ, Truong LD, Khurana KK, et al. Parathyroid cytology: avoiding diagnostic pitfalls. Head
Neck. 2002;24:157-164.
Agarwal AM, Bentz JS, Hungerford R, et al. Parathyroid fine-needle aspiration cytology in the
evaluation of parathyroid adenoma: cytologic findings from 53 patients. Diagn Cytopathol.
2009;3 7:407-410.
DeLellis RA, Nikiforov YE. Thyroid and parathyroid glands. In: Gnepp DR, ed. Diagnostic Surgical
Pathology of the Head and Neck. 2nd ed. Philadelphia: W.B. Saunders; 2009.
Dimashkieh H., Krishnamurthy S. Ultrasound guided fine needle aspiration biopsy of parathyroid
gland and lesions. Cyto]ournal. 2006;3:6
Lloyd RV, Douglas BR, Young WF. Parathyroid gland. In: King DW, ed. Fascicle #1-Endocrine
Diseases, First Series. Armed Forces Institute of Pathology Atlas of Non-Tumor Pathology.
Washington, DC: American Registry of Pathology; 2002.
Thompson LDR, ed. Endocrine Pathology. China: Churchill Livingstone; 2006.
The Adrenal Gland and
Paraganglia
Louis P. Dehner
I. NORMAL ANATOMY AND HISTOLOGY. The definitive or adult adrenal glands are
located anterior to the upper poles of the kidneys. The glands are pyramidal on
the right and crescentic on the left. In adults the normal combined weight should
not exceed 6 g. Each adrenal gland is divided into head (most medial), body
(middle), and tail (most lateral).
The adrenal gland is composed of an outer cortex and an inner medulla, and
as a compound gland is unique to mammals (Pharmacol Rev. 1971;23:1971).
The gland is derived from two embryologic progenitors: the coelomic epithelium
between the urogenital ridge and dorsal root mesentery as the primordial cor-
tex, and migratory neural crest cells as the future medulla as well as the ganglia
and paraganglia. Migratory neural crest cells are identified as small dark cells
as they pass through the cortex into the center of the gland as individual cells
or small aggregates of neuroblasts (Endocr Pathol. 2009;20:92; Folia Histochem
Cytochem. 2010;48:491); however, a definitive medulla does not become appar-
ent until 1 to 11/2 years of age.
Microscopically, the cortex consists of three zones: the outer zona glomeru-
losa (secreting aldosterone), middle zona fasciculata (secreting mainly cortisol
and minor amounts of sex steroids), and inner zona reticularis (secreting mainly
sex steroids) (e-Fig. 26.1).* The zona glomerulosa is composed of a thin, usually
discontinuous layer of cells with ball-like formations; these cells have less cyto-
plasm than those in other two cortical zones. The zona fasciculata consists of
radial cords or columns of cells with abundant lipid-rich cytoplasm. The cells of
the zona reticularis have compact, finely eosinophilic cytoplasm with or without
lipofuscin pigment. Normally the medulla accounts for 10% of the adrenal vol-
ume and grossly has a gray-white color. The predominant cells in the medulla are
the mature chromaffin cells, the pheochromocytes, organized in nests and cords
(e-Fig. 26.2). The cytoplasm of the chromaffin cells is usually basophilic but may
be amphophilic or even eosinophilic. These cells have indistinct cell borders and
usually a single nucleus which may show variation in size and hyperchromasia.
The chromaffin cells are peripherally surrounded by the sustentacular cells; rare
ganglion cells may be identified in the adrenal unlike their prominence in ganglia
(e-Fig. 26.3). The major function of adrenal medulla is the synthesis and secretion
of catecholamines (epinephrine and norepinephrine).
II. GROSS EXAMINATION OF ADRENAL GLAND SPECIMEN. The adrenal gland is removed
either as part of a radical nephrectomy or for excision of an adrenal tumor. Biop-
sies of the adrenal are generally fine needle aspirations or thin needle biopsies to
evaluate for the presence of metastatic tumor.
A. Needle biopsy. The entire tissue should be submitted for histologic examination.
B. Adrenal gland removed as part of radical nephrectomy. After gross examination
of the kidney, the gland should be measured and weighed. Then the gland
should be serially sectioned at 2- to 3-mm intervals perpendicular to its long
axis; the thickness of the cortex and medulla should be noted. Infrequently,
428
Chapter 26 • The Adrenal Gland and Paraganglia I 429
the adrenal may be invaded directly by renal cell carcinoma or be the site of
discontinuous metastases.
C. Adrenectomy for a primary pathologic process. The first step is to orient the spec-
imen and examine the contour of the adrenal gland. If it is apparent that the
gland has been largely replaced by a mass, the periphery should be inked since
the margins may be important (assuming that the mass has not already invaded
surrounding structures like the liver). If the cortex and medulla maintain their
normal relationship to each other, the thickness of each should be recorded.
Serial sectioning should be done at intervals appropriate for the pathology.
If the disease process is apparently diffuse hyperplasia, the periadrenal soft
tissue should be removed and the gland should be measured and weighed. If
the adrenal contains a solitary mass or multiple nodules, three dimensional
measurements should be obtained. The cut surface of the lesion and its rela-
tionship to any identifiable normal tissue should be described, including color,
consistency, presence of hemorrhage or necrosis, degree of circumscription, and
degree of encapsulation. If any portions of adjacent organs such as liver, kidney,
spleen, or abdominal wall are attached (usually for tumors), their appearance
and relationship to the gland should be noted. A large, en bloc resection will
require numerous sections from the peripheral margins. For diffuse and/or
nodular hyperplasia, representative sections are sufficient.
For a neoplasm, the following sections should be taken: tumor, (including
sections demonstrating the relationship of the tumor to the associated soft tis-
sues and adjacent organs, and relationship of the tumor to uninvolved adrenal
gland); the tumor capsule, if present; margins; periadrenal fatty tissue overlying
a bulging mass; a representative section from uninvolved adrenal, if any; and
regional lymph nodes. The gross description should clearly document the site
of the sections. For large specimens, a gross photograph is extremely helpful
in that it depicts the specimen at a time when landmarks are still maintained
with some anatomic orientation.
D. Neuroblastoma specimens. Neuroblastic tumors present some special issues
regarding acquisition of neoplastic tissue for a variety of special studies to
biologically profile the tumor for children enrolled in a Children's Oncology
Group (COG) protocol. If the specimen is an adrenal-based neuroblastoma
(NB), then generally the amount of available tumor is sufficient for patho-
logic diagnosis as well as for all other ancillary studies. The difficulties arise
in those cases of NB when only biopsies are obtained prior to chemotherapy
in cases of nonresectable tumors. Accuracy of tumor grading is based upon
a thorough microscopic examination which may be limited by the amount of
well-preserved tumor in the specimen.
A resected primary NB of the adrenal, or extraadrenal retroperitoneal mass,
should be examined fresh if at all possible as discussed in section C above. The
COG reference laboratory requests at least 1 g of snap frozen tumor tissue but
will accept any frozen sample of tumor; the snap freezing should occur as soon
as the specimen becomes available following resection. Tumor samples should
be labeled "primary" or "metastatic"; involved bone marrow is required as
well. Storage at -70°C is preferable to -20°C.
Fresh tissue should also be collected for local institutional studies including
conventional cytogenetic studies. Snap-frozen fresh tissue should be saved for
possible molecular studies. Tumor for fluorescent in situ hybridization can be
recovered from formalin-fixed, paraffin-embedded tissue without compromis-
ing the quality of results.
Ill. ADRENAL CORTICAL LESIONS
A. Congenital abnormalities. The most common congenital anomaly of the adrenal
is an incidental finding of heterotopia consisting of microscopic foci of cortical
tissue or nodule(s) along the path of descent of the gonads, although rare cases
of heterotopia at a wide variety of other anatomic sites have been reported.

Heterotopias are identified in 1.5% to 2.7% of groin procedures in males (B]E
Int. 2005;95:407); the spermatic cord, inguinal hernia sac, or paraepididymal
soft tissues are the three most common sites (Int Surg. 2006;91:125). Only
cortical tissue is identified as a rule (e-Fig. 26.4), but cortex and medulla have
been seen in heterotopias in the celiac axis. Other congenital abnormalities
include adrenal union/fusion, adrenal cytomegaly, intraadrenal heterotopic tis-
sue, and congenital adrenal hyperplasia. Adrenal cytomegaly is characterized
by the presence of foci of bizarre cells with eosinophilic granular cytoplasm and
large hyperchromatic nuclei with pseudoinclusions; this finding is detected in
Beckwith-Wiedemann syndrome. Heterotopic tissues within the adrenal gland
include liver, thyroid, and ovarian stroma.
B. Incidental adrenal cortical nodules. Nonfunctional cortical nodules are found in
1% to 10% of autopsies, are usually multiple and bilateral, and can protrude
into adjacent fat. These variably sized nodules have a yellow appearance on
cut surface. Circumscribed but nonencapsulated nodules consist of fasciculata-
type cells with various patterns. Myelolipomatous metaplasia, osseous meta-
plasia, and secondary changes (hyalinization, calcification, or hemorrhage)
may be present. Pigmented nodules are composed of zona reticularis-type cells
with lipofuscin or neuromelanin.
C. Adrenal cortical hypofunction (insufficiency) is divided into primary and sec-
ondary types.
1. Primary adrenal cortical insufficiency (Addison disease) is due to destruc-
tion of the adrenal cortex. In developed countries, over 80% of cases are
due to autoimmune adrenalitis as an isolated manifestation of autoim-
mune polyendocrinopathy (Lancet. 2003;361:1881). This T-cell mediated
immunologic destructive process of the cortex results in small glands which
have a mixed inflammatory infiltrate of lymphocytes, plasma cells, and his-
tiocytes. Lymphoid follicles with germinal centers may be present in a pat-
tern similar to that in Hashimoto thyroiditis (Endocr Dev. 2011;20:161).
Cortical cells may be difficult to identify, but the medulla remains intact.
Various inborn errors of metabolism, hemorrhage, and neoplastic infiltrates
or replacement are other etiologies.
Adrenal cortical insufficiency may occur on the basis of infectious etiolo-
gies including tuberculosis, other bacterial infections (including Meningo-
coccus, Pseudomonas, Streptococcus pneumoniae, and Haemophilus
influenzae), fungal infections, cytomegalovirus, herpes simplex virus, and
toxoplasmosis (in lllV-infected individuals). These infections lead to
destruction of the entire gland. Acute adrenal insufficiency with bilateral
cortical hemorrhage and necrosis are features of the Waterhouse-
Friderichsen syndrome, more commonly seen at autopsy. Adrenoleukodys-
trophy (X-linked peroxisomal disorder) is also manifested by adrenal insuf-
ficiency with cortical nodules composed of enlarged ballooned cells(] Clin
Endocrinol Metab. 2011;96:E925).
Other causes of primary insufficiency include amyloidosis and drugs.
Treatment with mitotane may cause adrenal atrophy with fibrosis.
Congenital adrenal hypoplasia (CAHP) is an uncommon condition that
is more likely to be encountered in a fetopsy or perinatal autopsy. Its esti-
mated incidence is 1:12,500 live births, compared to the incidence of anen-
cephaly of 2.23:10,000 live births in the United States. It should be antic-
ipated that 1% to 2% of fetopsies have CAHP as defined as combined
adrenal weights of <2 g at term, or more accurately an adrenal weight
over body weight of <1:1000. There are three distinct morphologic pat-
terns: cytomegaly type (most common with X-linked inheritance with inac-
tivating deletion mutations at Xp 21.2 in the gene that encodes DAX-1);
anencephalic type (without CNS or pituitary defects but the cortex is atten-
uated and the fetal cortex is inconspicuous); and a miniature type with a
definitive cortex and an attenuated fetal cortex.
2. Secondary and tertiary adrenal cortical insufficiencies. These are due to the
failure of the pituitary gland to secrete ACTH (secondary) or of the hypotha-
lamus to secrete CRH (tertiary). The gland size is decreased. Histologically,
the zona fasciculata is atrophic whereas the zona glomerulosa and medulla
are usually relatively normal.
D. Adrenal cortical hyperplasia can be divided into congenital and acquired types.
1. Congenital adrenal hyperplasia is an autosomal recessive disorder caused
by one of five enzymatic defects that result in a failure in cortisol synthe-
sis; 21-hydroxylase deficiency accounts for 90% to 95% of cases (Lancet.
2005;365:2125; Endocr Dev. 2011;20:80). Marked diffuse hyperplasia
of the zona fasciculata results from ACTH stimulation, whereas cells of
the zona fasciculata are lipid-depleted due to their conversion into zona
reticularis-type cells with compact eosinophilic cytoplasm. Persistent ACTH
stimulation may also give rise to adrenal cortical neoplasms. Nodules resem-
bling hyperplastic adrenal cortical tissue in testis can develop into the so-
called testicular tumors of the adrenogenital syndrome (Am] Surg Pathol.
1988;12:503).
2. Acquired adrenal cortical hyperplasia is a nonneoplastic bilateral process
characterized by a gland weighing in excess of 6 g. The gland has either a
diffuse, nodular, or combined appearance.
a. Diffuse hyperplasia is usually ACTH dependent and caused by a
hyperfunctional pituitary (most often pituitary adenoma, less often
corticotropin-releasing hormone (CRH) from the hypothalamus) or an
ectopic ACTH/CRH-producing tumor. The latter neoplasms include
small cell carcinoma (usually from the lung), low grade neuroendocrine
carcinoma or carcinoid (usually from the lung or thymus), medullary
thyroid carcinoma, pancreatic endocrine neoplasms, and pheochromo-
cytoma (PHEO). Both glands are symmetrically enlarged. The zonae fas-
ciculata and reticularis are expanded with their relative proportions vary-
ing from case to case. The zona fasciculata is lipid depleted but markedly
expanded by a population of cortical cells with abundant eosinophilic
cytoplasm (e-Fig. 26.5).
b. Nodular cortical hyperplasia is ACTH-independent in most cases. The
glands are markedly enlarged, in excess of 15 to 20 gin some cases. The
cortical nodules may constitute a transformation from diffuse hyperpla-
sia in its late stage; these nodules are yellowish and vary from 0.2 to over
4.0 em. Fasciculata-type clear cells, reticularis-type cells, or a mixture of
these two cell types characterize the nodules. Nodular adrenal corti-
cal disease with discrete nodules of hyperplastic zona glomerulosa with
intervening cortical atrophy is seen in children with McCune-Albright
syndrome (Am] Surg Pathol. 2011;35:1311).
c. Primary pigmented nodular adrenocortical disease (PPNAD), an uncommon
but specific form of ACTH-independent nodular hyperplasia, is generally
diagnosed in the second decade of life with or without the other stigmata
of Carney complex (Orphanet] Rare Dis. 2006;1:21; Best Pract Res
Clin Endocrinol Metab. 2010;24:389). A germline heterozygous inacti-
vating mutation of PRKR1A is found in the Carney complex (65% to
70% of cases) and may also have a role in primary PPNAD (Pituitary.
2006;9:211). The glands are of normal size, but the cut surface has scat-
tered pigmented micronodules, or less often macronodules measuring 1
to 4 mm in size. Uniform compact cells with eosinophilic cytoplasm and
some balloon cells are histologic features of the nodules. Pigmentation is
4X2 I SECTION Vlt ENDOCRINE 5YS1EM
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l.Mied""" permllllllltln.
Chapter 26 • The Adrenal Gland and Paraganglia I 4 33
antagonist), characteristic spirolactone bodies are usually present, con-

sisting of small, intracytoplasmic, 2 to 6 ~m round to oval inclusions
with a slightly eosinophilic appearance and two to six concentric rings
within the tumor cells and the adjacent zona glomerulosa (Am] Clin
Pathol. 1970;54:22). A halo separates the spirolactone bodies from the
surrounding cytoplasm.
b. Cushing syndrome is characterized by an adenoma which is sharply
demarcated or even capsulated and has a homogeneous yellow to golden-
yellow appearance or irregular foci of dark discoloration on cut surface.
Solid nests or alveolar-like profiles are present, composed of cells which
are usually larger than the normal cortical cells. The cells of the ade-
noma have features of zona fasciculata-type cells with the occasional
presence of foci of smaller zona reticularis-type cells (e-Fig. 26.9); both
types of tumor cells usually have single, round to oval nuclei with a small
nucleolus. In some cases, there are scattered cells with larger, hyperchro-
matic nuclei with pseudoinclusions; they have no prognostic importance.
Mitotic figures are rare. Fibrosis, organizing thrombi within sinusoids,
and lipomatous and myelolipomatous metaplasia are other features. The
compressed residual cortex is invariably atrophic.
c. Adrenogenital syndrome with sex hormone production is infrequently
caused by an adenoma, but is more often associated with cortical carcino-
mas that produce estrogen. Androgen-producing adenomas are generally
larger than those in Cushing syndrome and are also sharply demarcated
from the adjacent cortex. The tumor cells have the features of zona retic-
ularis cells with compact, eosinophilic cytoplasm.
2. Other adrenocortical neoplasms with special microscopic features beyond
the usual appearance of adenoma and carcinoma include oncocytic tumor,
myxoid adrenal cortical neoplasm, sarcomatoid carcinoma, and the so-
called adrenal blastoma.
a. Black (pigmented) adenoma is associated with Cushing syndrome, and
rarely with Conn syndrome (Int] Surg Pathol. 2004;12:57). In most
cases the tumor is composed predominantly or entirely of cells resembling
zona reticularis cells with a variable amount of intracytoplasmic brown
or golden-brown pigment (lipofuscin) (Endocr Pathol. 1999;10:353).
b. Oncocytic neoplasms include both benign and malignant tumors as in
other organs. A cytoplasm rich in mitochondria accounts for the onco-
cytic appearance of the tumor cells in common with extraadrenal oncocy-
toma. These tumors are often characterized by large, bizarre cells; despite
the presence of these very atypical cells, the histologic criteria to differ-
entiate an adenoma from a carcinoma are applicable (Int] Surg Pathol.
2004;12:231; Ann Diagn Pathol. 2010;14:204). These tumors may have
cytoplasmic globules and inclusions like those seen in pheochromocy-
toma (Virchow Arch. 2008;453:301), which can be problematic since
the latter tumor may also have oncocytic features in rare cases (Am]
Surg Pathol. 2000;24:1552).
c. Myxoid neoplasms are rare morphologic variants of a cortical adenoma or
carcinoma with a focal or near diffuse background that has a myxoid-
mucoid appearance consisting of hyaluronidase digestible Alcian blue
positive stromal mucin. The myxoid stroma separates the tumor cells
into cords, nests, pseudoglands (e-Fig. 26.10), and trabecula (Ann Diagn
Pathol. 2008;12:344; Am] Surg Pathol. 2010;34:973). The individual
tumor cells are spindled to epithelioid. The various microscopic features
for differentiation of a benign from malignant tumor are applicable to
the myxoid variant.
d. Ectopic adrenal cortical adenoma has been reported in the kidney and
within the spinal canal (Mod Pathol. 2009;22:175; Brain Tumor Pathol.
2010;27:121). Ectopic adrenal rest or "tumor" has been described

in congenital adrenal hyperplasia in the testis (Eur ] Endocrinol.
2008;159:489).
e. Sarcomatoid carcinoma, like similar high-grade neoplasms in the lung
and kidney, has a pleomorphic spindle cell pattern in addition to focal
areas of recognizable but poorly differentiated ACC (Pathol Res Pract.
2010;206;59).
f. Adrenal blastoma has been reported in a single case (Hum Pathol.
1992;23:1187). This tumor had a biphasic epithelial and mesenchymal
pattern.
3. Adrenal cortical carcinoma (ACC) is a rare neoplasm with an incidence of
about one per million in the United States. This tumor can occur in any
age, but it appears to have a bimodal age distribution with one peak in chil-
dren <5 years old and another peak in adults in the fourth or fifth decade
of life. However, many of the cases of putative ACC in children, especially
those diagnosed early in the first decade of life, do not behave clinically
as their adult counterparts although the microscopic features associated
with ACC in adults are present (discussed later). ACC is a manifestation
of several hereditary syndromes including Li-Fraumeni syndrome (TP53),
Beckwith-Wiedemann syndrome (CDKN1C/NSD1), MEN1 (MEN1), and
Gardner's syndrome (APC) (] Clin Pathol. 2008;61:787; Adv Anat Pathol.
2011;18:206); together hereditary cases represent 5% or less of all ACCs.
ACC presents as a functioning neoplasm in 50% to 60% of cases, with
Cushing syndrome as the most common presentation (Nat Rev Endocrinol.
2011;7:323). Approximately 40% to 45% of ACCs are confined to the
adrenal and are completely resectable; only 5% to 10% of tumors are
~5 em (T1) whereas 35% to 40% are >5 em (T2); over 50% of cases
have infiltrated beyond the adrenal (T3) or have already metastasized at
presentation in 30% to 35% of cases (T4) (Best Pract Res Clin Endocrinol
Metab. 2009;23:273). The AJCC Staging scheme for adrenal neoplasms is
presented in Table 26.2.
Grossly, hemorrhage and necrosis often provide the background for a
soft yellowish to tan neoplasm with or without cystic areas (e-Fig. 26.11).
However, ACC can have a variety of gross features in terms of coloration
and presence or absence of apparent necrosis. Those ACCs <5 em tend to
have a more uniform gross appearance as do adenomas; these are the cases
which may prove to be problematic in the differential diagnosis between
an ACC and adenoma, a point that is especially pertinent as it relates to
adrenocortical neoplasms in children (Pediatr Dev Pathol. 2009;12:284).
Any number of individual microscopic features have been evaluated in
primary adrenocortical neoplasms in adults for discrimination between an
adenoma and carcinoma. The modified Weiss system requires a score of 3
or more to predict malignant behavior (of the following five microscopic
features): mitotic rate (>5 per 50 hpf), cytoplasm (dense eosinophilic cyto-
plasm in 75% or more of tumor cells since a predominant clear cell pattern
is a feature of adenomas); atypical or bizarre mitotic figures; necrosis; and
capsular invasion (e-Fig. 26.12A-D) (Hum Pathol. 2009;40:757). Once an
adrenal cortical neoplasm has invaded into adjacent soft tissues or organs,
invaded a major vessel, or metastasized to regional lymph nodes or remote
site(s), the diagnosis of ACC is beyond dispute.
Immunohistochemistry to corroborate that a primary neoplasm is cor-
tical in nature is infrequently necessary, with the uncommon exception of
a dilemma between a cortical and medullary neoplasm (which is gener-
ally obvious in most cases). Another example is metastatic renal cell carci-
noma to the adrenal which can mimic a primary cortical neoplasm. A more
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have slightly eosinophilic and finely granular cytoplasm, but they may be
amphophilic to basophilic or even oncocytic (e-Fig. 26.14). A finely vacuo-
lated cytoplasm is present in some cases due to lipid accumulation. Intracy-
toplasmic periodic acid-Schiff positive, diastase-resistant hyaline globules are
yet another finding (e-Fig. 26.15). Melanin-containing PHEOs are rare but
well documented (Hum Pathol. 1993;24:420). The polygonal cells can vary in
size and nuclear detail, with marked nuclear enlargement and hyperchromasia,
although less impressive cytologic atypia is often present. Prominent nucleoli
and nuclear pseudoindusions are additional but inconsistent findings (e-Fig.
26.16). Mitotic figures, if seen, are not an indication of malignancy per se.
The stroma may exhibit extensive hyalinization, fibrosis, or rarely amyloid
deposition, and the vasculature may be prominent. The periadrenal adipose
tissue often has a hibernomatous appearance regardless of the age of the
patient (e-Fig. 26.17). The chromaffin cells are positive for chromogranin
A, synaptophysin, and cytokeratin in 25% of cases (Arch Pathol Lab Med.
1990;114:506); thetumorcellsdonotexpress EMA, melanA, orinhibin. S-100
labels sustentacular cells that are usually located at the periphery of the nests.
Prognostic assessment of an individual PHEO which is confined to the gland
and without metastasis is an exercise in uncertainty. A number of morphologic
features of the tumor have been assessed from the architectural pattern (nested
vs. diffuse), presence or absence of confluent necrosis, mitotic rate, Ki-67 index,
as well as several others including weight (less than or greater than 10 g), but
the conclusion that there are "no absolute histologic criteria for predicting
malignant potential" still seems to be true (Histopathology. 2011;58:155).
Overall, 10% to 25% of PHEOs prove to be malignant as demonstrated by
metastasis to bone, lymph nodes, lungs, and liver (e-Fig. 26.18).
Composite or compound PHEO, a rare variant, is defined in most cases as a
PHEO with a ganglioneuromatous (80%), ganglioneuroblastomatous (20%),
neuroblastomatous (< 1% ), malignant peripheral nerve sheath tumor, or neu-
roendocrine carcinoma (extremely rare) component (e-Fig. 26.19) (Arch Pathol
Lab Med. 1999;123:1274; Am] Clin Pathol. 2009;132:69; ] Clin Pathol.
2009;62:659). This variant accounts for 1% to 3% of all PHEOs, and these
tumors are reported in the clinical settings of neurofibromatosis I and MEN
2A (Am] Surg Pathol. 1993;17:837; Am] Surg Pathol. 1997;21:102). There
is also a case with a cortical adenoma (Ann Diagn Pathol. 2011;15:185).
C. Neuroblastic tumors (also known as peripheral neuroblastic tumors) are embry-
onal tumors arising from the sympathoadrenal neuroendocrine system and are
the most common extracranial solid malignancy of childhood (Nat Rev Can-
cer. 2003;3:203; N Engl] Med. 2010;362:2202). About 600 new cases are
diagnosed each year in the United States. Approximately 98% of neuroblas-
tomas are diagnosed by 10 years of age, and 85% to 90% are detected before
5 years of age. The abdomen and/or retroperitoneum is the site of clinical pre-
sentation in 65% of cases; 50% of NBs arise in the adrenal medulla. Though
most neuroblastomas have a number of overlapping histopathologic features,
they are clinically, pathologically, and molecularly divisible into indolent or
aggressive subtypes (Pediatr Clin North Am. 2008;55:97; Curr Probl Cancer.
2009;33:333; Oncogene. 2010;29:1566).
1. Neuroblastoma (NB} is best characterized as having a variegated appearance
since the gross features are dependent on its morphologic composition and
on secondary changes such as hemorrhage, yellowish foci of necrosis and
calcification, cystic degeneration, and fibrosis; some or all of these find-
ings may be found in any one tumor (e-Fig. 26.20). It is often difficult to
judge whether some of the gross changes are spontaneous since resection
may be preceded by adjuvant therapy. A well-circumscribed, soft, gray-tan
tumm; measuring 2 to 10 em in dimension, with or without hemorrhage and
calcifications, is the common appearance of a poorly differentiated NB
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neuroblastic maturation to ganglionic differentiation in a fibrillary and neu-
romatous or Schwannian background. The macroscopic nodules are com-
posed of neuroblasts in varying phases of differentiation; these are sharply
demarcated from the surrounding stroma that has neuromatous organ-
glioneuromatous features. Nodules of poorly differentiated NB with a high
MKI connote a worse prognosis than those nodules with a low MKI (e-Fig.
26.31); the parameters used to group NBs into favorable and unfavorable
categories also apply to GNB in general (Cancer. 2003;98:2274; Pediatr
Blood Cancer. 2009;53:563). The overall favorable prognosis of GNB is
related to the fact that most tumors are localized and have intermixed rather
than nodular features.
3. Ganglioneuroma (GN) is the most common neoplasm of the sympathetic ner-
vous system in adults and occurs in the posterior mediastinum, retroperi-
toneum, and rarely in the adrenal gland. Most GNs are discovered inci-
dentally, but some are known to present with VIP manifestations (Surgery.
2011;149:99). The tumor is grossly a solid, circumscribed, encapsulated
homogenous gray-white mass, measuring 5 to 10 em (e-Fig. 26.32). Mature
GN is characterized by mature scattered single cells or groups of cells in
a neuromatous stroma (e-Fig. 26.33). The differential diagnosis of matur-
ing GN is intermixed GNB, but the stroma should constitute >50% of the
tumor in the former. There is a definitional fine line between intermixed
GNB and the so-called maturing GN, although in terms of a favorable
prognosis, the distinction is an unimportant one.
V. LESIONS OF THE EXTRAADRENAL PARAGANGLIOMA
A. Normal anatomy and histology. The extraadrenal paraganglia are divided into
two broad categories as they relate to the parasympathetic or sympathetic ner-
vous system. The former are concentrated in the head and neck region with a
close proximity to cranial nerves IX and XII and associated blood vessels and
are further divided into jugulotympanic, vagal, carotid body, laryngeal, and
aorticopulmonary paraganglia (AJR. 2006;187:492). The aorticopulmonary
paraganglia are distributed in parallel with the sympathetic nervous system
along the paravertebral and paraaortic axis and are further divided into cervi-
cal, intrathoracic, and intra-abdominal groups. Paraganglia are also found in
the bladder, prostate, and gallbladdet. All paraganglia have a similar compo-
sition of chief cells arranged in well-defined nests or zellballen surrounded by
peripheral sustentacular cells. Catecholamine concentrations tend to be higher
in the sympathetic paraganglia (including the adrenal medulla). A greater pro-
portion of sympathetic derived paragangliomas are functional and noradren-
ergic (25% to 85% of cases) compared with those of parasympathetic origin
(10% or less) (Histopathology. 2011;58:155).
B. Extraadrenal paraganglioma presents in a sporadic (70% to 75% of cases) or
heredofamilial (25% to 30%) clinical setting, but in children it is almost always
the latter with or without a PHEO (] Pediatr Surg. 2010;48:383). The vari-
ous hereditary paraganglioma syndromes are summarized in Table 26.4. Not
included in this table are the Carney triad (CT) of paragangliomas, gastroin-
testinal stromal tumor (GIST) of the stomach, and pulmonary chondromas
(] Clin Endocrinol Metab. 2009;94:3656;] Int Med. 2009;266;43). The dele-
tion in CT may be located within the 1p or 1q12--q21 region; there are no
mutations in SDHA. SDHB. SDHC. SDHD. KIT. or PDGFRA genes (] Int
Med. 2009;266:93).
1. Parasympathetic paragangliomas have a predilection for the head and neck
with an estimate incidence of 1:30,000 to 100,000; these tumors are found
in association with cranial nerves IX and XII (Head Neck. 2008;31:381).
Though these tumors are rare, the head and neck region is the most common
site (] Clin Endocrinol Metab. 2001;86:5210). Almost 60% of cases are

carotid body tumors at the bifurcation of the internal and external carotid
arteries (Head Neck Pathol. 2009;3:303). Other sites in the head and neck in
approximate descending order of frequency are the glomus jugulare, glomus
tympanicum (both of the latter present in the middle ear), and the glomus
vagale (which presents as a neck mass). The glomus jugulare arises in the
adventitia of the jugular bulb (a dilated portion of the internal jugular vein
at its origin at the jugular foramen) along the auricular branch of cranial
nerve X (also known as Arnold's nerve) or the tympanic nerve (a branch of
cranial nerve IX, also known as the nerve of Jacobson); the glomus tympan-
icum is contiguous with the tympanic nerve in the inferior temporal bone.
Other less common sites in the head and neck include the larynx, paranasal
sinus, salivary gland, orbit, and oral cavity. The thyroid and parathyroid
rarely harbor paragangliomas (Arch Otorhinolaryngol ltal. 2009;29:97;
Head Neck Pathol. 2010;4:37).
The carotid body tumor and glomus vagale are usually resected intact
to reveal a firm to rubbery well-circumscribed mass with a fibrous pseudo-
capsule measuring 1 to 6 em in greatest dimension. A grayish-tan to pale
brownish surface is present on sectioning; focal hemorrhage may be seen.
Groups of uniform pale staining tumor cells with finely granular nuclear
chromatin are surrounded by a prominent and sometimes hemorrhagic
fibrovascular network. Compressed spindle-shaped cells at the periphery
of the cellular nests are S-1 00 protein positive sustentacular cells. There
is typically more cellular pleomorphism, nuclear atypia, and hyperchro-
matism in PHEO in contrast to paragangliomas, although individual case
exceptions are acknowledged. Vascular invasion and infiltrative growth are
unusual findings.
There are some special problems associated with middle ear paragan-
gliomas. These tumors are often submitted in a piecemeal fashion, and
artifacts introduced during excision may yield an initial impression of a
vascular tumor. Immunohistochemistry is helpful in establishing the diagno-
sis and shows a phenotype of reactivity for chromogranin, synaptophysin,
CD56, and focally for cyclin-D1 (Otolaryngol Head Neck Surg. 2010;143:
531).
2. Sympathetic paragangliomas are present in the abdomen-retroperitoneum
in more than 80% of cases, specifically in the superior paraaortic region
(45% of cases), region of the adrenal gland or renal hilum (10%), infe-
rior paraaortic region (30%), and organ of Zuckerkandl or bladder (10%)
(e-Fig. 26.34). The remaining 5% of cases are collectively reported in the
gallbladder, kidney, urethra, prostate, spermatic cord, ovary, and vagina.
Were the adrenal gland included in the discussion, it would be the most com-
mon site of "sympathetic paraganglioma." Unlike the nonfunctional and
infrequently metastasizing parasympathetic paragangliomas, sympathetic
paragangliomas are more often malignant (25% to 65% of cases) with a
metastatic potential in excess of that seen in PHEOs (] Clin Endocrinol
Metab. 2011;96:717). In the absence of metastasis, many of the same
problems exist in the reliable pathologic identification of the potentially
malignant paraganglioma as in the case of a PHEO (e-Fig. 26.35). Size
(> 100 g), confluent necrosis, mitotic and proliferative activity (Ki-67
index), absence of hyaline globules, and small cell morphology are
some of the features that have been correlated with malignant behavior
(Histopathology. 2011;58:155).
3. Gangliocytic paraganglioma usually arises in the periampullary portion of
the duodenum but is also rarely seen in the nasopharynx, lung, esophagus,
mediastinum, pancreas, and appendix (BMC Cancer 20;11:187). It occurs
in patients over a wide age range as a solitary, polypoid mass that projects
into the lumen of the intestine and measures up to 7 em in diameter. It is an
infiltrative neoplasm composed of three cell types in variable proportions,
namely spindle cells, epithelioid cells, and ganglion cells (e-Fig. 26.36). The
spindle cells are elongated and have wavy nuclei resembling Schwanni cells,
with S-100 and neurofilament immunopositivity; these cells can envelope
the ganglion and epithelioid cells, analogous to sustentacular cells. The
larger epithelioid cells are arranged in solid nests, ribbons, or pseudoglan-
dular or papillary structures and have neuroendocrine features including
granular eosinophilic to amphophilic cytoplasm with uniform oval nuclei
and finely stippled chromatin (e-Fig. 26.37); these cells express cytoker-
atin, chromogranin, and synaptophysin. The ganglion cells have atypical
features, and there may be a morphologic continuum with the epithelioid
cell population. Recurrences are rare, but metastatic behavior is restricted
to regional lymph nodes or liver U Gastrointest Surg. 2007;11:1351; Ann
Diagn Pathol. 2011;15:467).
VI. OTHER ADRENAL PARENCHYMAL LESIONS
A. Myelolipoma accounts for <5% of primary adrenal tumors and is usually
detected incidentally on imaging studies of the abdomen, although hemorrhage
with pain is a rare presentation (Asian] Surg. 2009;32:172). Several cases have
been discovered in individuals with congenital adrenal hyperplasia and Cush-
ing syndrome (Exp Clin Endocrinol Diabetes. 2009;117:440; Endocr Pract.
2011;17:441). Whilethetumorcan belargerthan20cm and2 kg, it measures 3
to 5 em in most cases. The tumor has a soft yellow to red surface, depending on
the fat content and presence of hemorrhage. The usual microscopic appearance
includes varying proportions of mature adipose tissue admixed with normal
trilineage hematopoiesis (e-Fig. 26.38). Clonality has been demonstrated (Am
] Surg Pathol. 2006;30:838). Lipoma, angiomyolipoma, and liposarcoma are
in the differential diagnosis.
B. Adenomatoid tumor, like myelolipoma, is usually detected as an incidentaloma
(Am J Surg Pathol. 2003;27:969). The tumor is a well circumscribed, solid
or solid and cystic and measures from 0.5 to 9 em. Microscopically, the
tumor is composed of tubules, cysts, papillary structures, and occasional solid
sheets of low cuboidal cells (e-Fig. 26.39). The tumor cells may have cytoplas-
mic vacuoles with signet ring features (e-Fig. 26.40) but are mucicarmine-
negative for mucin. The tumor cells have the same immunophenotype as
mesothelial cells with reactivity for vimentin, CK7, calretinin, and WT-1
(Pathol Res Int. 2010;2010:702472). Adenomatoid tumor is believed to arise
from mesothelial inclusions within the adrenal gland. The main differen-
tial diagnoses include lymphangioma, metastatic carcinoma, and vascular
tumors, especially epithelioid angiosarcoma (Arch Pathol Lab Med. 2011;135:
268).
C. Mesenchymal tumors include lipoma, schwannoma, hemangioma, leiomyoma,
leiomyosarcoma, angiosarcoma, perivascular epithelioid cell tumor, Ewing
sarcoma-primitive neuroectodermal tumor, solitary fibrous tumor, heman-
gioblastoma, and papillary endothelial hyperplasia.
D. Lymphoma is most commonly seen in previously diagnosed cases of non-
Hodgkin lymphomas (NHL) and occurs in as many as 25% of cases at some
point in the clinical course; bilateral involvement is present in 75% to 80% of
cases, and adrenal insufficiency is a known complication. However, rare exam-
ples of NHL with a primary presentation in the adrenal have been reported
(Exp Clin Endocrinol Diabetes. 2011; 119:20 8). Diffuse large B-celllymphoma
is the most common pathologic type (Mod Pathol. 2009;22:1210), although
Burkitt lymphoma also occurs (Tumori. 2007;93:625). Hodgkin lymphoma of
the adrenal is rare (Br J Haematol. 2010;148:341).
VII. ADRENAL CYSTS are relatively uncommon and are divided into four cate-
gories: parasitic cysts (7%), epithelial cysts (9%), pseudocysts (39%), and
endothelial cysts (45%) (Arch Surg. 1966;92:131; Endocr Pathol. 2008;19:
274).
A. Pseudocysts are discovered as another incidentoma on CT imaging. The usual
appearance is a well-defined cyst with water-like density and a median size of 6
to 10 em in diameter. Pseudocysts are unilocular, are filled with yellow-brown
to bloody amorphous semi-liquid material, and have a wall thickness of 1 to
5 mm. The densely hyalinized connective tissue of the wall may contain focal
calcifications or even metaplastic bone formation, and entrapped cortical tissue
may also be present; the smooth muscle in the wall of the cyst is continuous
with the smooth muscle of the adrenal vein. An identifiable lining is absent. The
differential diagnosis includes a cystic NB or PHEO (Cancer. 2004;101:1537).
B. Vascular cysts are the most common type of adrenal cyst in some series (Arch
Pathol Lab Med. 2006;130:1722). Endothelial cysts are usually well circum-
scribed and multiloculated (e-Fig. 26.41). The endothelial cells are immunopos-
itive for CD31, and if derived from lymphatic endothelium, then also positive
for D2-40 (e-Fig. 26.42A and B).
C. Epithelial cysts are divided into true glandular cysts and embryonal cysts (World
I Surg. 2006;30: 1817), although in some classification schemes cystic adrenal
tumors are also considered in the category of epithelial cysts. Mesothelial cysts
also occur (Endocr Pathol. 2008;19:203).
D. Parasitic cysts are a manifestation of echinococcal infection in the adrenal and
retroperitoneum (Bull Soc Pathol Exot. 201 0; 103:313; Int Surg. 201 0;95:189).
The wall of the cyst is often calcified.
VIII. METASTATIC NEOPLASMS to the adrenal gland are common. Metastatic carcinoma
is present in the adrenals in 25% to 30% of carcinoma-related deaths at autopsy
(Cancer. 1950;3:74). The lung is the most common primary site (adrenal metas-
tases are found in 30% to 35% of cases) (e-Fig. 26.43) followed by the breast;
other malignancies that frequently metastasize to the adrenals include adenocar-
cinomas of the kidney, stomach, and colon (e-Fig. 26.44); melanoma (10% to
15% or greater of cases) (e-Fig. 26.45); hepatocellular carcinoma; and urothe-
lial carcinoma (Clin Endocrinol. 2002;56:95; Am I Surg. 2008;195:363; Semin
Oncol. 2008;35:172). Renal cell carcinoma (RCC) involves the adrenal by either
direct extension from an upper pole tumor or metastasis in 5% to 10% of cases;
metastatic RCC should always be differentiated from a primary adrenal cortical
carcinoma (Am I Surg Pathol. 2011;35 :678; Eur Urol. 2011;60:458) (Table 26.3 ).
Other types of neoplasms arising in the retroperitoneum or as a metastasis to the
adrenal can be differentiated in most cases by immunohistochemistry (see Table
26.3).
IX. ADRENAL GLAND CYTOLOGY
A. Fine needle aspiration (FNA). FNA is frequently used to evaluate adrenal
gland mass lesions and is performed under percutaneous CT and ultrasound
guidance, or endoscopic ultrasound guidance for left adrenal lesions (Diagn
Cytopathol. 2005;33:26). FNA diagnosis of adrenal lesions has an accuracy of
98% and specificity of 100% (Diagn Cytopathol. 1999;21:92). FNA biopsy
of pheochromocytoma is regarded as a relative contradiction due to possible
induction of hypertensive crisis (Radiology. 1986;159:733).
B. Specific Neoplasms
1. Myelolipoma. The aspirate shows a mixture of mature adipose tissue and
hematopoietic elements, including nucleated red blood cells, granulocytes
and precursors, and megakaryocytes (Acta Cytol. 1991;35:353).
2. Adrenal cortical neoplasms. The distinction between adrenal cortical hyper-
plasia, adrenal cortical adenoma, adrenal cortical carcinoma, and normal
adrenal cortical cells is not always possible in cytologic specimens. Radio-

logical correlation is essential.
a. Adrenal cortical adenoma yields moderately cellular smears that con-
tain poorly cohesive sheets of epithelial cells with ill-defined and vac-
uolated cytoplasm, and abundant stripped small round uniform nuclei.
Bubbly and vacuolated lipid background is prominent (e-Fig. 26.46).
Scattered cells show nuclear atypia. Some cells may have cytoplasmic
lipofuscin (Acta Cytol. 1995;39:843; Acta Cytol. 1998;42:1352; Diagn
Cytopathol. 1999;21:92).
b. Adrenal cortical carcinoma yields richly cellular aspirates. It consists of
more frequent single cells that have marked nuclear atypia, intact granu-
lar cytoplasm, eccentric nuclei, and necrosis (e-Fig 26.47). The cytologic
diagnosis of a well-differentiated adrenal cortical carcinoma is difficult
(Acta Cytol. 1997;41:385).
3. Pheochromocytoma. The cytomorphology shows similarity to that of other
neuroendocrine tumors. The aspirate contains abundant isolated and loose
dusters of malignant cells with intervening vasculature (e-Fig. 26.48). The
isolated polygonal or spindle-shaped cells exhibit poorly defined fragile
cytoplasm and fine granular salt-and-pepper chromatin (e-Fig. 26.49). Red
cytoplasmic granules can be seen on Romanowsky-type stains (Acta Cytol.
1999;43:207).
4. Metastatic malignancy. The most common malignancies metastatic to the
adrenal are adenocarcinoma of the lung or breast (e-Fig. 26.50). The con-
firmation of metastasis is straightforward given a known malignant his-
tory. It is important to integrate clinical, laboratory, radiologic, cytologic,
and immunocytochemical findings to differentiate primary neoplasms from
metastasis (Acta Cytol. 1995;39:843).
SUGGESTED READINGS
Chen H, Sippel RS, O'Dorisio SO, et al. The North American Neuroendocrine Tumor Society
consensus guideline for the diagnosis and management of neuroendocrine tumors. Pheochro-
mocytomas, paraganglioma, and medullary thyroid carcinoma. Pancreas. 2010;39:775-783.
Conran RM, Chung E, Dehner LP, et al. The pineal, pituitary, parathyroid, thyroid and adrenal
glands. In: Stocker JT, Dehner LP, Husain AN, eds. Stocker & Dehner's Pediatric Pathology.
3rd ed. Philadelphia, PA: Lippincott Williams & Wilkins; 2011:941-970.
DeLellis RA, Lloyd RV, Heitz PU, et al., eds. World Health Organization Classification of Tumours.
Pathology and Genetics of Tumours of Endocrine Organs. Lyon, France: IARC Press; 2004.
Khan A, ed. Surgical Pathology of Endocrine and Neuroendocrine Tumors. Totowa, NJ: Humana
Press; 2009.
Lloyd RV, ed. Endocrine Pathology. Differential Diagnosis and Molecular Advances. 2nd ed.
New York, NY: Springer; 2010.
McNicol AM. Update of tumours of the adrenal cortex, phaeochromocytoma and extra-adrenal
paraganglioma. Histopathology. 2011;58:155-168.
Raygada M, Pasini B, Stratakis CA. Hereditary paragangliomas. Adv Otorhinolaryngol.
2011;70:99-106.
Thompson LDR, ed. Endocrine Pathology: A Volume on Foundations in Diagnostic Pathology
Series. Philadelphia, PA: Churchill Livingstone; 2006.
Pituitary Gland
I. NORMAL ANATOMY AND HISTOLOGY. The pituitary gland (hypophysis or "under-

growth") is located at the base of the brain, beneath the hypothalamus, within
the sella turcica of the sphenoid bone. The smaller, posterior lobe of the pituitary
contains the neurohypophysis (pars nervosa), which is connected to the hypothala-
mus via the pituitary stalk, or infundibulum ("little funnel"). Associated with the
pituitary stalk is the infundibular portion (pars tuberalis) of the adenohypophysis.
The other portions of the adenohypophysis form the anterior lobe (pars distalis)
and vestigial intermediate lobe (pars intermedia); the latter contains remnants of
Rathke's cleft cyst (glandlike cystic spaces).
Histologically, on H&E-stained sections, normal adenohypophysis (e-Fig.
27.1)* is composed of three different cell types: acidophils (producing growth hor-
mone [GH] or prolactin [PRL]), basophils (producing adrenocorticotrophic hor-
mone [ACTII], thyroid-stimulating hormone [TSH], luteinizing hormone [LH],
and follicle-stimulating hormone [FSH] ), and chromophobes (hypogranulated aci-
dophils and basophils); all are arranged in an acinar pattern. The acini are demar-
cated by a delicate fibrovascular stroma best visualized on reticulin stains. Although
each acinus contains a mixed population of these cell types, the composition varies
regionally. Acidophils dominate the lateral portions, basophils (ACTII and TSH)
dominate the medial portions, and basophilic gonadotrophs (LH and FSH) are
distributed uniformly. Occasionally with aging, basophils extend into the neurohy-
pophysis; this phenomenon, termed "basophilic invasion," should not be misinter-
preted as an infiltrating neoplasm. The posterior pituitary is composed of axons,
axon terminals, and sometimes axonal swellings (spheroids) called Herring bodies
which contain vasopressin and oxytocin. Specialized glial cells (true "pituicytes")
accompany the axons; these are generally considered to be the cells of origin for the
rare and related benign tumors pituicytoma and choristoma (granular cell tumor
[GCT]). Ectopic pituitary tissue can be found in the nasal cavity, sphenoid sinus,
or rarely, within ovarian teratomas.
II. INTRAOPERATIVE EVALUATION AND TISSUE HANDLING. Intraoperative evaluations are
frequently requested for pituitary neoplasms, most often to confirm the clinical
diagnosis of pituitary adenoma. Although frozen sections usually provide ade-
quate information to make an intraoperative diagnosis, a small (1 mm3 ) amount
of tissue should be used for cytologic examination. Intraoperative cytologic smears
or "touch preps" lack freezing artifacts that obscure nuclear details (e-Fig. 27.2)
and thus provide extremely valuable complementary information at the time of
frozen section. Likewise, ample tissue should be reserved for paraffin embedding,
free from freezing artifacts, to preserve antigenicity and morphologic features that
might otherwise be compromised. This point is particularly important in the eval-
uation of a smaller microadenoma, which can easilr be exhausted through cavalier
intraoperative processing. Finally, a small (1 mm3 ) amount of tissue should be
reserved for ultrastructural examination; tissue fixed directly in 3% glutaralde-
hyde and embedded in plastic will retain fine structural features that cannot be
recovered from formalin-fixed, paraffin-embedded tissue.
446
Chapter 27 • Pituitary Gland I 44 7
Ill. NONNEOPLASTIC LESIONS

A. Pituitary hyperplasia. Diffuse expansion of pituitary acini, best evaluated on reti-
culin stained sections, is the histologic hallmark of pituitary hyperplasia. How-
evet; this diagnosis is often difficult to render with certainty. Nodular expansion
of acini with one single hormonal cell type is noted in a variety of clinical scenar-
ios, including pregnancy and estrogen therapy. One must also be cognizant of
the naturally heterogeneous composition of the anterior pituitary, which favors
acidophils laterally and basophils medially.
B. Pituitary apoplexy (e-Fig. 27.3 ). Spontaneous hemorrhage and infarction of non-
neoplastic or neoplastic pituitary may represent a surgical emergency (in the
setting of increased intracranial pressure, subarachnoid hemorrhage, or visual
disturbance). For diagnosis of an underlying tumor in a partially necrotic spec-
imen, a reticulin stain may prove more reliable than immunohistochemistry;
immunohistochemistry should usually be performed but is often difficult to
interpret in this setting.
C. Lymphocytic hypophysitis is a rare autoimmune disorder of the adenohypoph-
ysis that progressively destroys the gland, resulting in panhypopituitarism. The
posterior hypophysis may also be affected, resulting in diabetes insipidus. Lym-
phocytic hypophysitis occurs with greater frequency in pregnant or postpartum
women. Subsets of patients also have other associated autoimmune disorders.
Microscopically, the anterior pituitary demonstrates lymphoplasmacytic infil-
trates without granuloma formation. The resulting glucocorticoid deficiency is
often fatal if untreated.
D. Granulomatous hypophysitis. This rare autoimmune disorder expands and selec-
tively destroys the anterior pituitary with well-formed, noncaseating granulomas
and a mild lymphocytic infiltrate. Radiologically, this condition may mimic ade-
noma, but it also often involves the thyroid, adrenal glands, and testes. Unlike
lymphocytic hypophysitis, it shows no association with pregnancy.
E. Rathke's cleft cyst (e-Fig. 27.4) is a developmental abnormality that arises
between the anterior and the posterior lobes or in the infundibular stalk. Smaller
examples are often detected incidentally. Lesions > 1 em are usually symptomatic
(e.g., cause hypopituitarism or visual disturbances related to compression of the
optic chiasm). The cyst is lined by cuboidal to columnar epithelium that may
or may not be ciliated and may or may not contain goblet cells. Focally, the
epithelium may show flattening and/or squamous metaplasia. The cyst con-
tents are variably serous or mucoid. Xanthogranulomatous inflammation may
be present in cysts that have undergone prior hemorrhage. Surgical resection is
curative.
F. Other. Rarely, in the setting of hypopituitarism, only xanthogranulomatous
inflammation is identified; such a finding may simply represent overwhelm-
ing reactive changes within a craniopharyngioma or Rathke's cleft cyst, but
the entity "xanthogranuloma of the sellar region" that is generally restricted
to the sella has also been described. The pituitary may also become involved
by systemic histiocytic disorders, such as Langerhans cell histiocytosis (LCH),
Rosai-Dorfman disease, Erdheim-Chester disease, and xanthoma dissemina-
tum. LCH in particular may involve the sellar/suprasellar region more often
than is generally considered; diabetes insipidus is a common manifestation.
IV. BENIGN NEOPLASMS
A. Pituitary adenoma is by far the most common sellar neoplasm. Tumors < 1 em
are typically identified as micro adenomas, and those> 1 em are categorized as
macroadenomas. Microadenomas are most often functional (hormone produc-
ing), which explains why they draw clinical attention before reaching larger
sizes. Nonfunctional tumors which account for one-third of all adenomas grow
to a large size before drawing clinical attention. Often, large tumors produce
so-called "stalk effect," in which mild to moderate elevations of PRL hormone
result from compression of the infundibular stalk, which blocks the transport
of dopamine (formerly, PRL inhibitory factor) from the hypothalamus. Some
macroadenomas may compress the normal pituitary tissue and cause panhy-
popituitarism. In many cases, macroadenomas are detected only after they
compress the optic chiasm, producing bitemporal hemianopsia or other vision
disturbance. Approximately half of nonfunctioning adenomas are referred to as
null-cell adenomas. These chromophobic (or less commonly, oncocytic) tumors
have a negative hormonal immunoprofile despite showing reactivity for synap-
tophysin and exhibit few if any secretory granules on ultrastructural analysis
(oncocytic adenomas show abundant mitochondria). Although some null-cell
adenomas appear to express the alpha subunit of the glycoprotein hormones
(LH, FSH, TSH) and thus might be considered "silent" gonadotrophic or thy-
rotrophic adenomas, expression of the alpha subunit has also been documented
in somatotroph adenomas (Neurol Res. 1999;21:247), so this feature may have
limited specificity.
Microscopically, adenomas lack the normal reticulin-rich acinar structure
of the adenohypophysis. Instead, they may appear patternless, pseudorosette-
rich, papillary, endocrine/organoid, or paraganglioma-like and may focally form
glands. In ambiguous cases, a reticulin stain may be useful for accentuating
loss of acinar architecture. Adenoma tumor cells are typically larger than their
nonneoplastic counterparts and have round-to-oval nuclei, delicate stippled
("salt and pepper") chromatin, and small or inconspicuous nucleoli. Occasion-
ally, adenomas exhibit moderate to marked cytologic atypia, but this feature is
not associated with aggressive behavior unless accompanied by elevated mitotic
and/or proliferation indices (see section on "atypical pituitary adenoma"). Cyto-
plasmic quality varies within and among different adenoma (hormonal) sub-
types and may even vary among cells within an individual tumor, particularly in
some tumors that secrete more than one hormone. Cytologic, histochemical, and
ultrastructural features can be used to subtype adenomas. Although immuno-
histochemical profiles of hormone expression and cell proliferation are now in
greater use for this purpose, familiarity with morphologic and histochemical
features remains worthwhile.
Indirect histologic dues toward adenoma biology (hormonal secretion pat-
terns) include the presence of calcium and amyloid bodies (rare) in prolacti-
noma. In prolactinoma treated with dopamine agonists, fibrosis and smaller cells
with high nuclear to cytoplasmic (N/C) ratios are common. Perivascular pseu-
dorosettes usually indicate a gonadotrophic or null-cell adenoma (e-Fig. 27.5).
Deposits of Crooke's hyaline (ringlike cytoplasmic cytokeratin inclusions) most
commonly accumulate in nonneoplastic corticotrophic cells in patients with
hypercortisolism of any cause, including, but not limited to, Cushing disease
(see section on corticotroph adenoma). Strong PAS staining suggests a corti-
cotroph adenoma, whereas weak PAS positivity is seen in glycoprotein (FSH,
LH, TSH expressing) adenomas. Round, weakly eosinophilic, cytokeratin pos-
itive, paranuclear "fibrous bodies" in a relatively chromophobic tumor most
often indicate a GH-producing adenoma (e-Fig. 27.6).
B. Prolactinoma. Also called lactotrophic adenoma, prolactinoma comes to clinical
attention most commonly in women of reproductive age with amenorrhea and
galactorrhea. Men with prolactinoma are usually asymptomatic or present with
decreased libido. Patients with prolactinomas are often treated medically with
a dopamine agonist (e.g., bromocriptine) to impair adenoma cell growth and
inhibit PRL production, but such agents are not cytotoxic. Tumors resected post-
therapy usually show interstitial fibrosis, reduced cell size, and high N/C ratio
and may be reminiscent of small-cell carcinoma (e-Fig. 27.7). However, they are
usually at least focally positive for PRL and display a low mitotidproliferative
index.
Chapter 27 • Pituitary Gland I 44 9
C. Corticotroph (ACTH-producing) adenoma. Corticotroph adenomas most often

present as microadenomas and account for 20% of all pituitary adenomas. These
tumors, which may be as small as 1 or 2 mm, can be difficult to localize radio-
graphically and intraoperatively. In such cases, a surgeon might employ inferior
petrosal sinus sampling to evaluate lateralization and monitor the progress of
resection. Even under these circumstances, exhaustive microscopic evaluation of
the pathology specimen (examining multiple levels using reticulin and immuno-
histochemical stains) may yield no diagnostic findings. Fortunately, a postopera-
tive drop in the patient's serum AC1H level may serve as evidence that the tumor
was removed. Most AC1H adenomas are amphophilic to basophilic, deeply
PAS positive, and arise in the central region of the gland where nonneoplas-
tic corticotrophic cells are concentrated. Crooke's hyaline, appearing as glassy,
eosinophilic cytoplasmic inclusions that ring the nucleus, results from hyper-
cortisolism and represents an accumulation of cytokeratin filaments already
present in corticotrophs (e-Fig. 27.8). Rare AC1H adenomas that themselves
exhibit Crooke's hyaline are called Crooke's adenomas. Other AC1H adenomas
that show immunoreactivity for ACTH but do not secrete biologically signifi-
cant amounts of the hormone and do not induce Cushing syndrome are termed
silent corticotroph adenomas. Such tumors are more common in men, tend
to be invasive, and do not come to clinical attention until they become large
(macroadenomas).
D. Somatotroph (GH cell} adenoma. This subgroup of pituitary adenomas includes
tumors with exclusive GH production (PAS negative), tumors producing GH
and PRL (mammosomatotroph adenomas), and plurihormonal adenomas (most
positive for GH, PRL, and TSH). Somatotroph adenomas (e-Fig. 27.9) usually
appear either intensely eosinophilic or rather chromophobic and show cor-
respondingly strong or weak GH immunoreactivity. Aiding diagnosis, these
chromophobic somatotroph tumor cells often contain weakly eosinophilic,
cytokeratin-reactive paranuclear whorls known as fibrous bodies (e-Fig. 27.6).
The systemic effects of somatotrophic adenomas (acromegaly in adults, gigan-
tism in adolescents) are mediated by insulin-like growth factor-1 (IGF-1). These
tumors may appear in the setting of Carney complex and multiple endocrine
neoplasia I (MEN 1).
E. Gonadotroph adenomas. These adenomas are more common in elderly males,
are often nonfunctional, and grow to a very large size before coming to clinical
attention by compressing adjacent structures; invasion is relatively less common
in this subtype. Histologically, these tumors are usually chromophobic and often
feature pseudorosettes, papillary architecture, or organoid features. Immuno-
histochemically, these tumors exhibit markedly variable reactivity for the beta
subunits of LH and FSH and for the common a subunit.
F. Pituicytoma (infundibular astrocytoma). This rare benign tumor is thought to
arise from the specialized glial cells of the posterior pituitary. Radiographi-
cally, it shows strong postcontrast enhancement. Histologically, it exhibits a
solid growth pattern and is formed by plump spindled tumor cells arranged
in short, interlacing fascicles. Rosenthal fibers and eosinophilic granular bod-
ies are not present. Strong reactivity for vimentin, S100, and TTF-1 (nuclear
pattern), patchy reactivity for GFAP, and no reactivity for cytokeratins, synap-
tophysin and neurofilament have been described (] Neuropathol Exp Neurol.
2009;68:482). Mitotic figures are uncommon.
G. GCT ofthe neurohypophysis (choristoma). Like pituicytoma, this tumor is thought
to be derived from pituicytes; some consider these two tumors to be pheno-
typic variants. Although minute asymptomatic examples are quite common,
larger symptomatic GCTs are relatively rare; these appear in the fifth or sixth
decade with a female predominance and commonly present as an enhancing sel-
lar/suprasellar mass compressing the optic chiasm. Histologically, these tumors
are formed by closely apposed polygonal cells with small nuclei and generous
granular eosinophilic cytoplasm. Spindled/fascicular areas may also be present.
Lymphocytic infiltrates are common and may be robust. Staining with PAS is
resistant to diastase digestion. Immunoreactivity is strong for S100 and TfF-1,
and variable for the lysosomal marker CD68; reactivity for pituitary hor-
mones, synaptophysin, cytokeratins, neurofilament, and chromogranin A is not
observed. Ultrastructurally, these cells show abundant phagolysosomes and no
evidence of neurosecretory granules.
H. Craniopharyngiomas (discussed in Chap. 41).
V. LOW-GRADE NEOPLASMS
A. Atypical pituitary adenomas are histologically similar to benign pituitary ade-
nomas except for elevated mitotic and proliferative indices. For this diagnosis,
current World Health Organization criteria require nuclear immunoreactivity
for the proliferation marker Ki-67/MIB-1 in >3% of tumor cells and extensive
nuclear immunoreactivity for p53; others suggest that Ki-67 labeling > 10%,
regardless of p53 status, correlates with more aggressive behavior and war-
rants an "atypical" designation (Neuroendocrinology. 2006;83:179). Relative
to benign adenomas, these tumors are more likely to invade adjacent structures
and to recur.
VI. HIGH-GRADE NEOPLASMS
A. Pituitary carcinoma is a very rare neoplasm that cannot be diagnosed by its innate
appearance or any known markers; instead, it is diagnosed by the presence
of craniospinal or extracranial metastases. The Ki-67/MIB-1 and p53 labeling
indices are typically high, but cytologic and histologic features are often benign.
These tumors are usually functional, secreting PRL or ACTH. Its designation in
this chapter as a high-grade neoplasm may be inappropriate in the truest sense,
but the term is applied here to reflect its more aggressive clinical behavior.
SECTION VII
Reproductive Tract
Testis and Paratestis

Kiran R. Vij and Peter A. Humphrey
I. NORMAL ANATOMY. The normal adult testis is an ovoid paired organ, measuring
4.5 x 2.5 x 3 cm3 , and weighing approximately 20 g. The testes are suspended
within scrotal sacs by spermatic cords. The testis is covered by a capsule composed
of an outer tunica vaginalis lined by mesothelium, the collagenous tunica albuginea,
and the inner tunica vasculosa. The tunica vaginalis forms a sac filled with serous
fluid. The posterior portion of the testis not covered by a capsule is called the
mediastinum and contains blood vessels, nerves, lymphatics, and the extratesticular
rete testis.
The testicular parenchyma is subdivided into lobules containing seminiferous
tubules separated by fibrous septae. The terminal portions of the seminiferous
tubules drain into the tubuli recti that connect to the tubules of the rete testis at the
mediastinum. The tubules of the rete testis anastomose with the ductuli efferentes,
which form the head of the epididymis and empty into the vas deferens, which
traverses the inguinal canal as a component of the spermatic cord. The testicular
artery arises from the aorta and is the major source of vascular supply to the testes.
The venous drainage occurs through numerous small veins that form a convoluted
mass known as the pampiniform plexus that surrounds the testicular artery. These
small veins anastomose to form the right testicular vein, which drains into the
inferior vena cava, and two left testicular veins, which drain into the left renal
vein.
Histologically, prepubertal and postpubertal seminiferous tubules are quite dif-
ferent. Prepubertal tubules are small, with few or no lumina, and contain mostly
Sertoli cells with a few primordial germ cells (e-Fig. 28.1).* Postpubertal seminif-
erous tubules are larger and harbor Sertoli cells and germ cells at varying stages
of maturation (e-Fig. 28.2). The Sertoli cells abut the basement membrane and
are aligned perpendicular to the membrane; their nuclei are round to oval with
prominent nucleoli, and the cytoplasm has Charcot-Bottcher crystals, which can
occasionally be seen by light microscopy. Within the seminiferous tubules, the least
mature germ cells-spermatogonia-are present along the basement membrane,
with the most mature cells--elongate spermatids-found at the luminal border.
Primary and secondary spermatocytes are found in an adluminal position.
The interstitial tissue between the seminiferous tubules contains Leydig cells,
vessels, and connective tissue. Leydig cells are arranged singly and in clusters
451
452 I SECTION VII: REPRODUCTIVE TRACT
(e-Fig. 28.3), and can be associated with nerves. They are large and irregularly
spherical to polyhedral, with small spherical nuclei and abundant acidophilic cyto-
plasm. The cytoplasm can exhibit lipofuscin and rod-shaped Reinke crystals.
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION. Tis-
sue samples of the testes received for surgical pathologic examination include testic-
ular biopsies, and unilateral and bilateral orchiectomy specimens. Retroperitoneal
lymph node dissection can be performed as part of a staging maneuver for testicular
cancer.
A. Fine needle aspiration biopsy of the testis in infertile patients (with sperm aspi-
ration and cytopathologic examination) is occasionally performed. Cytologic
touch imprints or wet preparations may be made at the time of open testicular
biopsy in infertile patients to rapidly identify the presence of sperm. The role of
cytology in the evaluation of testicular tumors is limited to diagnosis of lymph
node metastases by fine needle aspiration. Although seminomas can usually be
differentiated from nonseminomatous tumors, subtyping of nonseminomatous
tumors cannot be reliably performed by cytology.
B. Testicular biopsies, which can be open or percutaneous, are typically performed
for evaluation of infertility. They are usually contraindicated in the evaluation of
solid testicular masses, with the possible exception of epidermoid cysts, which
can be removed by excisional biopsy. Testicular biopsy specimens are usually
received in Bouin's fixative. An accurate documentation of the number and
size of the biopsy fragments and exact site(s) of the biopsy for each container
should be made during gross dictation. The biopsy fragments should be inked
with hematoxylin to facilitate identification during embedding, wrapped in lens
papet; placed between sponges, and processed entirely. Three hematoxylin and
eosin (H&E)-stained slides, each with three to four serial sections, should be
prepared from each block.
C. Unilateral simple orchiectomy is performed in cases of testicular torsion. Gross
examination of the testis similar to that for tumor cases should be performed.
One section of the testicular parenchyma in relation to the capsule, and a section
each of the epididymis and spermatic cord, should be submitted with description
and sampling of focal lesions.
D. Radical orchiectomy is performed for testicular tumors. The specimen consists of
the testis and paratesticular organs (surrounding tunica vaginalis, epididymis,
soft tissue, and a segment of spermatic cord). In cases of tumor resection, the
specimen should ideally be sent fresh and intact to the surgical pathology labora-
tory for immediate gross examination. Alternatively, when delay is anticipated,
the specimen is placed in 10% buffered formalin and sent intact. In such cases,
tumor morphology is often suboptimal due to poor fixation. The surgeon may
occasionally bisect the specimen to aid fixation; this approach is not recom-
mended as it prevents evaluation of involvement of the tunica by the tumor as
well as procurement of fresh tissue for ancillary studies.
The specimen is weighed, and measurements are recorded in three dimen-
sions. The length and diameter of the resected segment of spermatic cord are
noted separately. The external surfaces of the testis and spermatic cord are
examined for involvement by tumor. The proximal shave resection margin of
the spermatic cord is submitted in a separate cassette, and then the cord is
serially sectioned and inspected for tumor involvement; representative sections
are submitted proximal to distal. The tunica vaginalis is opened anteriorly to
show the tunica albuginea; presence of fluid, if any, within the sac is noted.
The testis is then bisected anteroposteriorly through the epididymis, and serial
sections are made parallel to this plane. Each slice is examined, and the tumor
is described in relation to the epididymis and the tunica albuginea. The size,
colot; and consistency of the tumor should be noted. Foci of hemorrhage, necro-
sis, and variegation, as well as multifocality, if present, should be described.
Chapter 28 • Testis and Paratestis I 4 53
Photographs or digital images should be taken and tissue procured for tumor
bank and ancillary studies such as flow cytometry and karyotyping, if necessary.
The specimen should then be fixed overnight in an adequate amount of 10%
buffered formalin prior to submission of one section per centimeter of tumor.
Representative sections should include heterogenous areas and sections of tumor
in relation to uninvolved parenchyma, epididymis, and tunica albuginea. One
section of grossly normal-appearing parenchyma should be included. If correla-
tion of identified histologic tumor type with serum markers (alpha-fetoprotein
[AFP] and human chorionic gonadotropin [hCG]) is not achieved, additional
sections should be submitted (note that such correlation will not always be per-
fect because metastatic deposits may harbor different elements than the primary
tumor).
E. Retroperitoneal lymph node dissection is performed as a separate procedure. The
specimen is received in 10% buffered formalin. During gross examination, the
tissue fragments should be measured in aggregate and carefully dissected to
harvest as many lymph nodes as possible, and the size of the largest and the
smallest putative nodes should be noted. Possible foci of tumor encountered
during dissection should be measured and sampled. An effort should be made
to sample any area(s) suspicious for viable tumor.
F. Bilateral orchiectomy specimens may be submitted as part of treatment of car-
cinoma of the prostate. Gross examination and sectioning are similar to that
for unilateral simple orchiectomy. Prostate cancer is rarely encountered within
these specimens.
Ill. DIAGNOSTIC FEATURES OF BENIGN DISEASES OF THE TESTIS
A. Congenital abnormalities
1. Cryptorchidism is maldescent of the testis in which the testis is found, after 1
year of age, to be located high in the scrotum, within the inguinal canal,
or in an intra-abdominal location. Grossly, the prepubertal undescended
testis differs little from normal, but after puberty the undescended testis
is smaller. Histologically, there is a progressive loss of germ cells with age,
along with decreased size of seminiferous tubules and increased thickness of
tubular tunica propria. Sertoli cell nodules (e-Fig. 28.4), which are foci of
tubules containing immature Sertoli cells and laminated calcific deposits, are
often seen in cryptorchid testes. These are likely hyperplastic foci, although
they have also been termed tubular adenoma of Pick. Rarely, Sertoli cell
nodules can present as a mass (Am ] Surg Pathol. 2010;23:1874). The
major complications of cryptorchidism are infertility and germ cell neo-
plasia, ranging from intratubular germ cell neoplasia (IGCN) to invasive
germ cell tumors, particularly seminoma, embryonal carcinoma, and embry-
onal carcinoma/teratoma. For patients > 1 year of age, placental-like alkaline
phosphatase (PLAP) and CD117 immunostains can be useful in highlighting
intratubular germ cell neoplastic cells.
2. Anorchism and polyorchidism are absence of testes and more than two testes,
respective!y.
3. In testicular-splenic fusion, encapsulated splenic tissue is found adjacent to
the left testis, which can show germ cell aplasia in the seminiferous tubules.
4. Adrenal conical rests are usually incidental, millimeter-sized nodules of
adrenal cortical tissue that are detected along the pathway of descent of
the testis, including along the spermatic cord and testis.
B. lnfenility. The causes of infertility may be pretesticular, which include endocrine
disorders involving the pituitary and adrenal glands; testicular, including genetic
disorders; or posttesticular, which are mainly obstructive and include varicocele
and cystic fibrosis. The evaluation of the patient includes a detailed clinical
history, physical examination, semen analysis, tests of endocrine function, anal-
ysis of sperm function, and serology for antisperm antibody. Testicular biopsy
..... I SECTION VII· REPRODUCTIVE TRACl
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c. Syphilitic orchitis occurs prior to infection of the epididymis. Histologi-

cally, peritubular lymphocytes and plasma cells can be seen along with
obliterative endarteritis and perivascular plasma cells. Gummas can form
an intratesticular mass with central necrosis.
d. Other types of orchitis include nonspecific granulomatous orchitis and
malakoplakia. Before diagnosing nonspecific granulomatous orchitis, a
specific infectious orchitis, sarcoidosis, lymphoma, and exuberant granu-
lomatous inflammation associated with a germ cell neoplasm should be
excluded.
D. Vascular disorders
1. Systemic vasculitis can affect the testis, but isolated vasculitis involving only
the testis is rare. Testicular vasculitis can cause infarction that clinically sim-
ulates a neoplasm. Most cases of testicular vasculitis are characterized by
polyarteritis nodosa-like changes in small- to medium-sized arteries, with
transmural necrotizing inflammation (Urology. 2011;77:1043).
2. Varicocele is an abnormal dilatation and tortuosity of veins of the pampini-
form plexus of the spermatic cord.
3. Torsion and infarction generally occur in young men and in the setting of
abnormal testicular descent. Initially, there is congestion, edema, and hem-
orrhage, followed by hemorrhagic infarction (e-Fig. 28.7).
E. Atrophy can be caused by many factors, but the morphologic findings are similar.
Grossly, the testis is small. Microscopically, the tubules are decreased in size and
the tunica propria is thickened and hyalinized. End-stage atrophic tubules are
completely hyalinized, without intraluminal cells (e-Fig. 28.8).
IV. TUMORS OF THE TESTIS. These are of germ cell origin in the vast majority of cases,
with sex cord/gonadal stromal tumors occurring in 4% to 6% of cases. The highest
incidence is found in Europe and parts of New Zealand. Populations in Africa
and Asia show a much lower incidence. The 2004 World Health Organization
classification of the neoplasms of the testis is given in Table 28.2.
A. Germ cell tumors. Clinical diagnosis is usually made when the patient presents
with a painless mass in the testis. Associated symptoms such as a dull ache in
the scrotum or lower abdomen may be present. Less commonly, patients present
with gynecomastia and thyrotoxicosis. In 10% of cases, metastatic disease may
produce presenting signs and symptoms. Patients with cryptorchidism are at an
increased risk (about three- to fivefold) of developing germ cell neoplasia both
in the cryptorchid and in the normal contralateral testis. Patients with testicular
microlithiasis, testicular atrophy, infertility, a family history of germ cell neo-
plasia, 46,XY or 45,X/46,XY mixed gonadal dysgenesis, testicular dysgenesis
syndrome (IDS), or a previous history of IGCN are also at an increased risk.
The age of the patient as well as characteristic elevations in the levels of serum
tumor markers in different subtypes of germ cell tumors can be very helpful
in predicting tumor type. Yolk sac tumor and/or teratoma are seen in infants
and children; seminomas and nonseminomatous germ cell neoplasms (includ-
ing embryonal carcinoma, teratoma, yolk sac tumor, and choriocarcinoma) are
found in adolescents and young adults; and spermatocytic seminoma occurs in
patients older than 50 years. Serum AFP levels can be elevated in nonsemino-
matous germ cell tumors, especially those with a yolk sac tumor component,
whereas serum beta-hCG levels are increased in choriocarcinoma and tumors
with syncytiotrophoblasts, which include about 10% of seminomas. The lev-
els of these tumor markers are monitored posttreatment and are indicators of
residual disease.
Imaging studies such as ultrasound are extremely sensitive and inexpensive
in evaluating testicular masses. Localization of the mass, including evaluation
of extratesticular versus intratesticular sites of involvement, and presence of
41& I SECTION VII· REPRODUCTIVE TRACl
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a. lntratubular germ cell neoplasia of unclassified type (IGCNU), a precursor of

invasive germ cell neoplasia, may be found in isolated form or adjacent to
invasive germ cell tumors. Patients with infertility, cryptorchidism, inter-
sex syndrome, gonadal dysgenesis, a history of invasive germ cell tumor
in the contralateral testis, or a retroperitoneal germ cell tumor have an
increased likelihood of development of IGCNU. The high sensitivity of
testicular biopsy in detecting IGCNU makes it a useful screening tool in
these subgroups of patients. IGCNU is not recognizable grossly. Histolog-
ically, the seminiferous tubules show enlarged neoplastic cells with clear
cytoplasm and prominent nucleoli, and often a thickened tunica propria
(e-Fig. 28.9). Mitoses may be seen. Normal spermatogenesis is lacking.
The surrounding stroma may show a prominent lymphocytic response.
The presence of scattered neoplastic germ cells in the surrounding stroma
qualifies as microinvasion (e-Fig. 28.10). Occasionally, the neoplastic cells
spread along the tubules or the rete testis in a pagetoid fashion (e-Fig.
28.11 ). Periodic acid-Schiff histochemical stains highlight glycogen in the
cytoplasm of IGCNU cells; immunohistochemically, PLAP, CD117 (c-kit),
and OCT4 (e-Fig. 28.12) positivity are observed (Semin Diagn Pathol.
2005;22:33). In infants up to 1 year of age, and in patients with delayed
germ cell maturation (cryptorchidism or gonadal dysgenesis), intratubular
germ cells can resemble IGCNU morphologically and by PLAP or OCT4
staining (Am I Surg Pathol. 2006;30:1427), so caution is advised in diag-
nosing IGCNU in these cases. About 90% of pure IGCNU cases progress
to invasive disease within 7 years.
b. Seminoma comprises almost 40% to 50% of all testicular germ cell tumors.
Grossly, it has a homogeneous white to gray cut surface (e-Fig. 28.13).
Necrosis may be seen, but hemorrhage and cystic change are uncom-
mon. Histologically, seminoma is composed of a diffuse sheet of uni-
form cells. The cells may be separated into nests, clusters, or columns by
delicate fibrous septae infiltrated by mature lymphocytes (e-Fig. 28.14).
A parenchymal or intratubular granulomatous reaction may be present
(e-Fig. 28.15). The granulomatous inflammation can be so extensive, both
in the testis and in the draining lymph nodes, that the tumor cells are
nearly obscured, resulting in a misdiagnosis of granulomatous orchitis.
The tumor may entirely replace the normal testicular parenchyma, and
may show an intratubular and less commonly an interstitial growth pat-
tern. Other morphologic patterns include pseudoglandular (e-Fig. 28.16),
tubular, cribriform, and occasionally microcystic appearances (Am I
Surg Pathol. 2005;29:500). In these latter cases, membranous PLAP and
c-kit positivity, together with a negative staining pattern for AFP, inhibin,
pan-cytokeratin, and CD30 can be useful for differentiation from non-
seminomatous germ cell tumors (Table 28.3). An OCT4 immunostain
will mark seminoma and embryonal carcinoma, but not other germ cell
tumor types (Am I Surg Pathol. 2004;28:935). Podoplanin immunore-
activity has been reported in seminomas but not embryonal carcinoma
(Ann Diagn Pathol. 2010;14:331); CD117 (c-kit) (e-Fig. 28.17) is like-
wise immunopositive in seminomas and so can also be useful in sepa-
rating these two entities in difficult cases. Foci of scarring may indicate
a regressed germ cell tumor, and areas of calcification should prompt a
search for possible foci of gonadoblastoma. Although brisk mitotic activ-
ity (>6 mitoses per high-power field) is observed in some tumors, there
is little evidence to support that this denotes a worse prognosis, although
it has been hypothesized that this finding may indicate progression to
embryonal carcinoma. Marked cytologic atypia and mitoses have been
used in the past to define "anaplastic seminoma," but this is not a
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and/or glandular pattern (e-Fig. 28.21). Nuclei are polygonal and vesic-
ular with coarse chromatin. Mitotic activity and necrosis are extensive.
IGCNU can be present at the periphery and frequently displays intratubu-
lar necrosis. Vascular invasion can be seen (e-Fig. 28.22) and should be
differentiated from retraction artifact and artificial implantation during
sampling. Pan-cytokeratin, SALL4, OCT4, and CD30 positivity is seen
in tumor cells; PLAP and AFP are only focally positive in pure tumors.
Epithelial membrane antigen (EMA) is negative, which is important in
the differential diagnosis with somatic carcinomas (Table 28.3). Evalua-
tion of H&E-stained slides is usually sufficient to establish the diagnosis;
in occasional cases, immunohistochemistry is needed to help differentiate
embryonal carcinoma from yolk sac tumor, seminoma, anaplastic large-
cell lymphoma, and/or choriocarcinoma.
e. Yolk sac tumor (endodermal sinus tumor) shows differentiation reminiscent
of embryonic yolk sac, allantois, and extraembryonic mesenchyme. In
infants and young children, it tends to occur in pure form, whereas in
adults it is found as a component of malignant mixed germ cell tumors.
The morphologic appearance is varied and includes reticular (most com-
mon), microcystic, endodermal sinus-like (with Schiller-Duval bodies),
papillary, solid, glandular, alveolar, enteric, polyvesicular vitelline, and
hepatoid patterns (e-Fig. 28.23). Rarely, a neoplastic spindle cell compo-
nent has been observed in association with the myxomatous and reticu-
lar variants. The immunoprofile reveals AFP, glypican 3, SALL4 (e-Fig.
28.24), PLAP, and low-molecular-weight cytokeratin positivity. IGCNU
is commonly seen in adult yolk sac tumors but not as frequently in child-
hood yolk sac tumors. Embryonal carcinoma may, in certain foci, be dif-
ficult to distinguish from yolk sac tumor, and indeed the two tumor types
can appear to merge; however, embryonal carcinoma nuclei are usually
more pleomorphic, and immunostains can help in difficult cases (Table
28.3). Follicle-like areas of granulosa cell tumors in infants may resemble
the solid and cystic pattern of yolk sac tumor, and the enteric pattern may
appear similar to glandular areas in teratoma; usually, the former problem
can be resolved with immunohistochemical stains for AFP and inhibin.
f. Trophoblastic tumors are almost always choriocarcinoma. Pure chorio-
carcinoma is extremely rare; instead, choriocarcinoma most commonly
occurs as a component of mixed germ cell tumors. Grossly, necrotic
and hemorrhagic nodules can be observed. Microscopically, the more
viable peripheral areas show randomly admixed syncytiotrophoblasts,
cytotrophoblasts, and intermediate trophoblasts, although one compo-
nent may predominate giving rise to a monophasic tumor (e-Fig. 28.25);
the tumor has a propensity for vascular invasion (e-Fig. 28.25). Very rare
tumors composed of intermediate trophoblastic cells resembling placen-
tal site trophoblastic tumor have also been reported. The syncytiotro-
phoblasts are positive for hCG, the alpha subunit of inhibin, and EMA,
and the intermediate trophoblasts are reactive for human placental lac-
togen (HPL); all cell types are positive for cytokeratin. The differential
diagnosis includes syncytiotrophoblast-rich seminoma, and isolated syn-
cytiotrophoblasts found in nonseminomatous germ cell tumors.
g. Teratomas include mature and immature teratoma, dermoid cyst, mon-
odermal teratoma, and teratoma with somatic-type malignancies. The
age distribution is bimodal. Teratomas occurring in children are benign,
whereas those in young adults have significant rates of metastases despite
their histologic appearance. Grossly, there are cystic and solid areas,
and cartilage (e-Fig. 28.26) and bone may be evident. Microscopi-
cally, teratomas may show well-differentiated elements derived from one
(monodermal) or all three germ layers (ectoderm, mesoderm, and endo-

derm), or may be immature with fetal-type tissue. Skin and its appendages,
respiratory and intestinal-type epithelium, cartilage, and muscular tissue
are common (e-Fig. 28.27A and B); neural-type tissue is less frequent.
Immature tissues can resemble renal blastema or embryonic neural tube
(Fig. 28.28). Foci with the appearance of a primitive neuroectodermal
tumor (PNET) have been classified as such if present. Pure teratomas are
much less common than the finding of teratoma as an element of a mixed
malignant germ cell tumot:
Dermoid cysts, which harbor keratinizing squamous epithelium and
skin appendages, are very rare and are benign. Epidermoid cysts lack
skin appendages and possibly represent monodermal teratomas; grossly,
epidermoid cysts are distinctive with a ringlike "onion-skin" cut surface
(e-Figs. 28.29 and 28.30).
Several nongerm cell malignant tumors, characterized by expansile
growth of at least a 4x field, including adenocarcinoma, squamous
cell carcinoma, neuroendocrine tumors, sarcomas, and PNET have been
known to arise in primary or metastatic teratomas.
Immunostains are not usually necessary for the diagnosis of ter-
atoma, but it is noteworthy that AFP immunopositivity can be present
in intestinal-type areas of teratomas.
h. Tumors of more than one histologic type (mixed forms), termed "mixed
malignant germ cell tumor," are germ cell neoplasms composed of more
than one type of tumor. They comprise approximately 30% of all germ cell
tumors, and the most frequently encountered components include embry-
onal carcinoma (in about one half of the cases), yolk sac tumor (one
half), and teratoma (about 40% ). About 40% of mixed malignant germ
cell tumors also contain scattered syncytiotrophoblasts. A rare subtype of
mixed germ cell tumor is the polyembryoma with characteristic embry-
oid bodies with central cores of embryonal carcinoma (forming the dor-
sal amnion-like cavity) and a surrounding (ventral) yolk sac component.
Mixed germ cell tumors with an embryonal component are predictive of
a higher stage than tumors with a large seminomatous component, so for
mixed tumors it is very important to quantitate, on a percentage basis,
the amount of embryonal carcinoma (as well as all other components).
Tumors with a yolk sac or teratoma component show a lower incidence
of metastatic disease.
i. Burnt out {regressed) germ cell tumors are germ cell tumors in which the
primary tumor in the testis has undergone necrosis and fibrosis. This most
commonly occurs with seminoma (Am I Surg Pathol. 2006;30:858) and
choriocarcinoma. The most specific histologic finding of a regressed germ
cell tumor is a distinct scar (e-Fig. 28.31) in association with either IGCNU
or coarse intratubular calcifications; however, many cases lack these latter
two features (Am I Surg Pathol. 2006;30:858). Some patients may present
with metastatic disease, the only evidence of the testicular primary being
a scar, with or without IGCNU.
j. New immunophenotypic markers that are transcription factors involved in
the maintenance of stem cell pluripotency have been developed for germ
cell tumors. These markers include SALL4, OCT4, NANOG, SOX2, and
SOX17, and all demonstrate a nuclear pattern of expression. SALL4labels
all IGCNUs, classic seminomas, embryonal carcinomas, and yolk sac
tumors (Am I Surg Pathol. 2009;33:1065). Choriocarcinoma, teratoma,
and spermatocytic seminoma are also variably positive for SALL4. Both
OCT4 and NANOG label IGCNU, classic seminoma, and embryonal car-
cinoma (Am I Surg Pathol. 2004;28:935; Histopathology. 2005;47:48).
SOX2 only labels embryonal carcinoma and teratoma (Am J Surg Pathol.
2007;31: 836; f Pathol. 2008;215: 21) and SOX17 labels IGCNU, classic
seminoma, and yolk sac tumor ( J Pathol. 2008;215: 21; Am J Clin Pathol.
2009;131:131). Among these markers, SALL4 demonstrates the highest
sensitivity and showed particular utility for yolk sac tumors. For yolk sac
tumors, SALL4 is much more sensitive than PLAP, AFP, and glypican-
3 (Am J Surg Pathol. 2009;33:1065). SALL4 is also particularly useful
in differentiating metastatic germ cell tumors from metastatic carcinoma
of nontesticular origin, a setting in which it has been found to be more
sensitive than OCT3/4 (Cancer. 2009; 115:2640).
More recently, an RNA-binding protein UN28 has been discovered as
a novel sensitive and general germ cell tumor marker, which shows sensi-
tivity similar to SALL4 (Hum Pathol. 2010;42:710). A potential diagnos-
tic pitfall that must be kept in mind is that all new markers described above
are also expressed in early fetal germ cells. SALL4 is also expressed in sper-
matogonia in both children and adults (Am JSurg Pathol. 2009;33:1065).
k. Molecular genetics is not currently used in diagnosis. The only exception
is detection of isochromosome 12p by fluorescence in situ hybridization
to identify metastatic germ cell neoplasms, since gain of material in 12p
is the most common structural chromosomal alteration in invasive germ
cell tumors (Mod Pathol. 2004;17:1309).
B. Sex cordlgonadal stromal tumors comprise 4% to 6% of all testicular tumors in
adults and include Leydig cell tumor, Sertoli cell tumor, granulosa cell tumor,
thecoma, and fibroma.
1. Leydig cell tumors are the most common and are known to occur in patients
with gynecomastia, Klinefelter syndrome, and cryptorchidism. Grossly, the
tumor cut surface is homogeneous, solid, and brown to yellow. The tumor
cells are large and polygonal, and have abundant eosinophilic, lipid-laden
cytoplasm (e-Fig. 28.32). The characteristic Reinke crystals are found in only
30% of cases. The tumor cells are positive for inhibin, calretinin, and Melan-
A by immunohistochemistry. The main differential diagnoses are Leydig cell
hyperplasia and syndromic adrenogenital tumors. Leydig cell tumors form a
nodule without seminiferous tubules, whereas in Leydig cell hyperplasia, the
foci are bilateral and multifocal, and wrap around and extend between the
tubules (e-Fig. 28.33); adrenogenital tumors are bilateral and dark brown,
with pigmentation and fibrous stroma. About 90% of Leydig cell tumors are
benign, but constellation of features, including size > 5 em, cytologic atypia,
increased mitotic activity, necrosis, and vascular invasion, favors malignancy.
2. Sertoli cell tumors account for only 1% of all testicular tumors. Grossly,
the mass is usually well circumscribed, with tan-yellow to white, sometimes
hemorrhagic cut surfaces (e-Fig. 28.34). The pathologic subtypes are the
lipid-rich, large cell calcifying (e-Fig. 28.35), and sclerosing variants. The
known clinical associations are with Carney syndrome, Peutz-Jeghers syn-
drome (which can be associated with bilateral tumors), and androgen insen-
sitivity syndrome. Histologically, the cytologically bland cells are arranged
in tubules, potentially with retiform, tubular-glandular, and solid nodular
areas. The tubules can be closed (solid) (e-Fig. 28.36) and are surrounded by a
basement membrane. Intratubular growth can be present. Sertoli cell tumors
should be distinguished from small incidental Sertoli cell nodules (benign
and thought to be nonneoplastic) as can be seen in cryptorchid testes. The
tumor cells are cytokeratin and sometimes inhibin and calretinin positive,
but PLAP and OCT4 negative, by immunohistochemistry. Malignant Sertoli
cell tumors are rare.
3. Granulosa cell tumors and thecoma/fibroma tumors are similar in appearance
to those in the ovary, and are rare in the testis. There are two granulosa cell
tumor variants in the testis, the adult and juvenile types, as in the ovary.
C. Mixed germ cell and sex cord/gonadal stromal tumors include gonadoblastoma,
most commonly seen in the setting of mixed gonadal dysgenesis, ambiguous
genitalia, and 45X/46XY mosaicism. Microscopic examination shows two cell
populations: a germ cell component resembling seminoma and a component
resembling immature Sertoli cells (e-Fig. 28.37). Round deposits of basement
membrane-like material and coarse calcification are common features. A large
number of patients develop invasive germ cell tumors, particularly seminoma,
and so patients are treated with bilateral orchiectomy.
D. Miscellaneous tumors of the testis include carcinoid tumors, tumors of ovarian
epithelial types (serous borderline tumm; serous carcinoma, mucinous cystade-
nomas and cystadenocarcinomas, Brenner tumor, and endometrioid carcinoma),
nephroblastoma, and paraganglioma.
E. Hematolymphoid neoplasms, of which the most common is malignant lymphoma,
comprise 5% of all testicular malignancies. These are the most common bilat-
eral tumors of the testis, and their incidence is higher in elderly men. The most
common subtype is diffuse large B-celllymphoma. The growth pattern is typ-
ically intertubular (e-Fig. 28.38), and the main differential diagnosis is with
seminoma, especially the spermatocytic type. Immunostains can be helpful in
establishing the diagnosis (Table 28.3 ). The prognosis is generally poor. Young
age, low stage, and presence of sclerosis are indicators of a good prognosis.
Isolated plasmacytoma of the testis is rare.
F. Tumors of the collecting ducts and rete. Benign tumors of the rete include
adenoma, cystadenoma, and adenofibroma. Adenocarcinomas of the rete are
rare, and their diagnosis is subject to strict histologic criteria that include
tumor centered on testicular hilum, morphology distinct from any other testic-
ular/paratesticular tumot; solid growth pattern, transition between tumor and
normal tissue, and absence of histologically similar extratesticular malignancy
(especially lung and prostate). The main histologic patterns are tubular, pap-
illary, and solid. The main differential diagnoses are extratesticular adenocar-
cinomas and mesothelioma. The tumor shows extensive regional spread and
distant metastasis, and the overall prognosis is poor.
G. Tumors of the paratesticular organs.
1. Benign. The most common benign neoplasm of the testicular adnexa is adeno-
matoid tumor, representing almost 60% of all cases. These are of mesothelial
origin (Semin Diagn Pathol. 2000;17:294) and arise in the upper or lower
pole of the epididymis as solitary, round to oval nodules invariably <5 em
in size. Histologically, the tumor shows round to oval or slit-like tubules
in a fibrous, hyalinized, and/or muscular stroma (e-Fig. 28.39 and 28.40).
The lining cells are columnar or flat with vacuolated cytoplasm. Adenoma-
toid tumors show immunoreactivity for cytokeratin, EMA, and mesothelial
markers including calretinin and Wf-1. The tumor should be differentiated
from signet-ring-cell carcinoma and mesothelioma; tumors with a more dif-
fuse growth pattern simulate Sertoli or Leydig cell tumors (inhibin positivity,
lipofuscin pigment, and presence of Reinke crystals favor Leydig cell tumor).
Another benign paratesticular tumor is papillary cystadenoma of epi-
didymis (e-Fig. 28.41). About two-thirds of cases are seen in patients with
von Hippel-Lindau disease, and in this setting, the tumor tends to be
bilateral.
A much rarer entity is the retinal anlage tumor or the melanotic neuroec-
todermal tumor, which is composed of two cell populations: larger melanin-
containing cells and smaller neuroblast-like cells.
2. Malignant. Malignant mesothelioma of the testicular adnexa has a micro-
scopic appearance and immunophenotypic profile similar to those of
mesothelioma of the pleura. The main differential diagnoses are with adeno-
matoid tumot; which is better circumscribed, and with carcinoma of the rete
testis.
Primary adenocarcinoma of the epididymis is rare, and may histologically

and cytologically simulate a cystadenoma due to the presence of columnar
cells with clear cytoplasm containing glycogen. Metastatic adenocarcinoma
from other organs should also be excluded.
Desm.oplast:ic small round cell tumor occurs in the epididymis of young
adults. Molecular, genetic, histologic, and immunohistochemical features are
similar to those of the tumor when it occurs at more conventional sites such as
the peritoneum. The tumor should be differentiated from other small blue cell
tumors such as malignant lymphoma and embryonal rhabdomyosarcoma.
Mesenchymal tumors of the scrotum, paratesticular organs, and sper-
matic cord include benign neoplasms such as lipoma, leiomyoma, neurofi-
broma, and granular cell tumor and malignant tumors such as liposarcoma,
leiomyosarcoma (e-Fig. 28.42), malignant fibrous histiocytoma, and rhab-
domyosarcoma (e-Figs. 28.43 and 28.44).
H. Secondary malianancias include metastatic adenocarcinomas from the prostate
(e-Fig. 28.45), lung, and colon, and melanoma.
V. PATHOLOGIC STAGING applies only to germ cell neoplasms of the testis (Fig.
28.1). The 2010 Tumor, Node, Metastasis (TNM) American Joint Committee on
pT1 pT2
= with vascular/
lymphatic invasion
pT3 pT4
Figure 28.1 Pathologic staging of germ cell tumors of the testis. (Modified from:
Greene Fl., Compton CC, Fritz AG, et al., eds. A]CC Cancer Staging Atlas. New York,
NY: Springer; 2006.)
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Prostate
Peter A. Humphrey
I. NORMAL ANATOMY. The normal weight of the prostate is 20 g for ages 20 to 50, and
30 g for ages 60 to 80. Anatomically, the prostate gland comprises three zones: cen-
tral zone, transition zone (where benign prostatic hyperplasia [BPH] occurs), and
peripheral zone (where most carcinomas originate). (e-Fig. 29.1). *Microscopically,
the normal adult prostate is a branching duct-acinar glandular system embedded
in a dense fibromuscular stroma (e-Fig. 29.2). The epithelium has two layers: a
luminal or secretory cell layer and a basal cell layer. Central zone epithelium can
normally have architectural patterns that include cribriform and Roman bridge-like
structures (e-Fig. 29.3 ).
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION.
The most common prostatic parenchymal tissue samples examined in surgical
pathology laboratories in the United States are, in order, 18-gauge needle cores,
transurethral resection of prostate (TURP) chips, radical prostatectomy specimens,
and fine needle aspirates.
A. Needle cores. Needle core biopsy sample handling and processing begins in the
room where the procedure is performed. The needle biopsy tissue should be
immediately placed into a container with fixative, which is usually 10% neutral
buffered formalin, although a few laboratories prefer Bouin's solution, Hol-
lande's solution, or IBF fixative. Bouin and Hollande's solutions are picric acid-
based fixatives that provide superior nuclear detail, but these strong oxidizing
agents can react violently with combustible materials and reducing agents. Fixa-
tion in formalin should be for at least 6 hours. The number of cores received per
container is highly variable, from 1 to >20. If the urologist and treating physi-
cian desire site-specific diagnosis, the core(s) should be placed in separate site-
designated containers. Inking of cores to indicate site, with placement of cores
marked with different colors into the same container, should not be performed
because fragmentation renders site assignment impossible. Gross examination
of prostate needle core tissue is not diagnostic, but is important for correlation
with amount of tissue seen in the histologic sections and so it is vital to record,
for each container, the size and number of tissue cores or fragments. It is recom-
mended that no more than two cores be submitted per cassette for processing
and embedding; some laboratories submit one core per cassette. Prostate cores
can be marked with ink, which facilitates identification during embedding and
the ability to see the cores in the paraffin blocks. Regardless, the cores should be
placed into a cassette after being put into a fine mesh envelope, wrapped in lens
paper, sandwiched between sponge pads, or double-embedded in agar-paraffin
wax. After processing, the cores should be embedded in the same plane, in the
same direction, with even spacing. From each paraffin block, three hematoxylin
and eosin (H&E)-stained slides should be prepared, with three to four serial
sections on each slide. Some laboratories cut interval, unstained sections on
coated slides in case special studies such as immunohistochemistry are needed.
Clinical requests for frozen section diagnosis of prostate needle cores are rare
and should be restricted to patients with clinical evidence of metastatic cancer
who are to undergo immediate treatment (usually orchiectomy) for pain relief.
467
B. TU RP chips. The amount of prostate tissue resected in TURP procedures is vari-

able, ranging from 5 to >75 g oftissue, with a mean of about 25 g. The gross
description should include the aggregate weight of the chips. Recognizable gross
features such as yellow coloration and induration can be recorded, but it has not
been proven that chip color, size, or induration is linked to cancer presence, so
gross selection of specific chips is not required. Although gross TURP chip sam-
pling procedures are not standardized, one initial approach is to submit 12 g of
chips or 6 to 8 blocks of tissue (with 1 to 2 g per cassette). For specimens> 12 g,
the initial 12 g are submitted, with one cassette for every additional 5 g (Arch
Pathol Lab Med. 2009;133:1568). If the patient is younger than 60 years, all tis-
sue should be submitted; all chip tissue should also be submitted if microscopic
examination of partially submitted chips reveals carcinoma in< 5% of tissue, or
if high-grade prostatic intraepithelial neoplasia (PIN) or atypical glands (atypi-
cal small acinar proliferation [ASAP]) is found in sections of partially submitted
chips. One H&E-stained slide, with one or two sections, is typically generated
from each paraffin block of TURP chips.
C. Open suprapubic or retropubic simple prostatectomy {enucleation) tissue. The pro-
static tissue from simple prostatectomies may be submitted to the pathology
laboratory as a single mass or as pieces. The prostatectomy tissue should be
weighed and sectioned at 3- to 5-mm intervals. The gross description for each
piece should include size in three dimensions, weight, firmness, and coloration.
Hard nodules should be sampled, and a total of eight cassettes or one cassette
of tissue for each 5 g of tissue submitted. Additional tissue should be submitted
if carcinoma is histologically detected in initial sections of partially submitted
tissue, although no rules or recommendations exist on how many additional
sections are required. One H&E-stained slide should be made per block.
D. Radical prostatectomy. The entire prostate gland is excised in prostate cancer
surgery using open retropubic or perineal approaches, or using laparoscopic
(including robotic) approaches. The prostate gland is also resected in toto in
radical cystoprostatectomy for bladder cancer.
1. Pelvic lymph nodes. Pelvic lymphadenectomy may be performed as a separate
procedure, often laparoscopic, or during the radical prostatectomy opera-
tion. Sentinel lymph node sampling is not routinely performed.
Frozen section analysis of the sampled lymph nodes may be requested
for patients at risk for nodal metastasis, on the basis of serum prostate-
specific antigen (PSA) level, needle biopsy Gleason score, and clinical stage.
All grossly recognizable lymph nodes should be examined by frozen section;
cytologic touch imprints can be made at the same time. Frozen section diag-
nosis of metastatic carcinoma in lymph nodes is highly specific, but fairly
insensitive. The low sensitivity rate of 58% to 73% is due to sampling error.
Gross sampling of tissue after frozen section should entail submission of
all grossly identifiable lymph node tissue and wide sampling of associated adi-
pose tissue. The gross description of pelvic lymph nodes should include num-
ber, location, and size. One H&E-stained slide is made per paraffin block.
Special studies to detect occult lymph node metastases, such as immuno-
histochemistry for cytokeratins or PSA, or reverse transcription-polymerase
chain reaction (RT-PCR) for PSA RNA, are currently experimental and not
performed in routine practice.
2. Prostate gland and seminal vesicles. The prostate gland and seminal vesicles
from radical prostatectomy procedures may be received fresh or in fixative.
All three dimensions and specimen weight should be recorded. Frozen section
requests on fresh specimens are uncommon, and are usually made to evaluate
margin status; this procedure has a high false-negative rate and is not stan-
dard practice. Fresh specimens are also used for tissue-banking protocols;
after inking the entire outside of the specimen, tissue can be harvested from
Chapter 29 • Prostate I 469
inside the gland while preserving the inked periphery. Alternatively, after ink-
ing, margin sampling, and seminal vesicle amputation (see below), the whole
unfixed gland can be sectioned with a large sharp knife from apex to base at
4-mm intervals perpendicular to the prostatic urethra; areas suspicious for
carcinoma, as judged by palpation or visual inspection, may be sampled by
imprints, scrapes, core biopsy, or small wedge sections.
Fixation of the inked radical prostatectomy specimens (sectioned or
unsectioned) is accomplished by at least overnight (or 24 to 48 hour) room
temperature immersion in 10% neutral buffered formalin at 10 times the
volume of the specimen. For gross examination of unsectioned glands after
inking, the seminal vesicles are amputated (including the soft tissue and pro-
static tissue at the base of the seminal vesicles) and submitted separately as
right and left seminal vesicles. The prostate weight without the seminal vesi-
cles is recorded, and distal apical (urethral) and bladder neck margins are
taken if not already submitted separately by the surgeon. The distal apical
margin is evaluated by amputating the distal5 to 10 mm of the gland, dividing
it into the right and left sides, and submitting radial sections (as for a cervi-
cal cone biopsy). The bladder neck margin can be assessed by a thin 2-mm
shave margin or by conization; the latter is recommended. Ink on tumor cells
is indicative of a positive margin for cone sections and the peripheral margin,
whereas tumor anywhere in shave margin tissue indicates a positive margin.
Vasa deferentia stumps may be sampled using en face sections, but this is
not routine. The prostate gland is serially sectioned in a plane perpendicular
to the urethra at 3- to 5-mm intervals using a long knife. The cut surfaces
should be evaluated for gross evidence of BPH and carcinoma. Photographs
or digital images can be used to document location and gross appearance of
tissue in submitted cassettes. Diagrams (Fig. 29.1) or pictorial maps can also
be used to indicate location of sections and any gross abnormalities.
Both complete and partial embedding methods are acceptable (Mod
Pathol. 2011;24:6). Several protocols for partial submission have been pub-
lished (Scand] Urol Nephrol Suppl. 2005;216:34; Mod Pathol. 2011;24:6).
When there is grossly visible tumor, all lesions grossly suspicious for carci-
noma should be submitted, along with distal apical and bladder neck margins
and seminal vesicles. For cases with no grossly evident tum01; the posterior
aspect of each transverse slice is submitted, as well as a mid-anterior block
from each side, distal apical and bladder neck margins, and the seminal
vesicles. Sections should be submitted as quarters or halves of the prostate,
depending on gland size. Whole-amount sections are rarely made and do not
provide additional morphologic information. If no or minimal tumor is seen
in initial sections of a partially submitted gland, all remaining tissue should
be embedded (including any frozen tissue sent to a tissue bank). If still no
tumor is seen, basal cell and AMACR immunostains of atypical foci should
be performed, and deeper sections should be obtained from block(s) from
the region of the positive needle biopsy and areas of high-grade PIN. If can-
cer remains undetected, the tissue blocks should be flipped, and histologic
sections prepared from the new tissue faces (Mod Pathol. 2011;24:6).
E. Cystoprostatectomy. The prostate gland in radical cystoprostatectomies per-
formed for bladder cancer can be sampled by taking several sections of prostatic
urethra and surrounding prostate tissue, any gross lesions, one block from the
periphery of each side, and both seminal vesicles. The distal urethral shave mar-
gin is important in urothelial carcinoma cases.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF THE PROSTATE
A. Inflammation and infection. Histopathologic identification of inflammatory cells
in the prostate is common, but histologic identification of specific infectious
agents is rare.
An tenor l'linddor neck
Radical Prostatectomy case# _ _ _ _ __
Guide to Taking Sections Name _ _ _ _ __
Weight _ _ _ _ _ grams
Ink right BLACK, Left BLUE o r RED
Guide to block numbers:
Block 1 - Tumor in Methacarn Fixative
Right- odd numbers
Left - even numbers
Posterior
Anterior Anterior Anterior
Righ~
·
3
At>ex
Ovemead
view
'A 2,4
Left~
Posterior Posterior Posterior
Anterior Anterior Anterior
Im
Ri:~no.
Bladder Neck
Ovemead
view
Cl' CilllO.
Posterior Posterior Posterior Left
Figure 29.1 Diagram depicting guide to taking sections from a radical prostarectomy specimen.
(Modified from: True LD. Surgical pathology examination of the prostate gland. Practice survey
by American Society of Clinical Pathologists. Am] Clin Pathol. 1995;103:376.)
1. Asymptomatic inflammatory prostatitis, including acute neutrophilic inflam-

mation and chronic lymphocytic, lymphoplasmacytic, or lymphohistiocyric
inflammation, is common in all prostate tissue samples. Reporting this
inflammation is optional; it may be useful to report if the inflammation is
extensive or persistent in several needle core samples taken over rime, because
prostatic inflammation can raise the serum PSA. Inflammation can be associ-
ated with prostatic glandular atrophy, and reactive nuclear changes including
prominent nucleoli. Inflammation is more commonly associated with benign
epithelial conditions, especially atrophy and BPH, compared with high-grade
PIN and carcinoma where usually only a small percentage of foci (around
10%) are inflamed.
2. Granulomatous prostatitis can clinically elevate the serum PSA and/or present
as a palpable abnormality. The most common type is nonspecific granulo-
matous prostatitis, which is thought to be a response to prostatic secretions
released into stroma by duct-acinar rupture. Microscopically, there is a lobu-
locentric noncaseating granulomatous inflammatory cell in6ltrate with giant
cells (e-Fig. 29.4 ). Variants include xanthogranulomatous prostatitis and pro-
static "xanthoma." Other types of granulomatous prostatitis include infec-
tious and postbiopsy/postresection cases. Infectious granulomatous prostati-
tis is most often Bacille Calmette-Guerin (BCG)-related in patients treated
for bladder urothelial carcinoma (e-Fig. 29.5). Fungal prostatitis is rare and
usually seen in immunosuppressed patients. Postbiopsy/resection granulo-
mas are most often identified in TIJRP chip tissue, and are characterized by
a fibrinoid central zone surrounded by palisading histiocytes.
B. Atrophy of prostatic glands is the benign condition most likely to be misdiagnosed
as prostatic carcinoma by light microscopy. It is a common, age-related pro-
cess that could be related to inflammation, hormones, obstruction, or ischemia.
It can also be treatment related, due to radiotherapy or hormonal therapy.
Histologically, atrophy is defined as cytoplasmic volume loss. It is not neces-

sary to subtype atrophy, but it is important to recognize the existence of dif-
ferent histomorphologic patterns including simple atrophy (with or without
cystic change) (e-Fig. 29.6), sclerotic atrophy, partial atrophy (e-Fig. 29.7), and
postatrophic hyperplasia (or hyperplastic atrophy) (e-Figs. 29.8 and 29.9). Atro-
phy can be confused with carcinoma because it is usually a small gland lesion
that can show a pseudoinfiltrative pattern of growth, stromal sclerosis, nuclear
atypia, and closely packed acini (in postatrophic hyperplasia). Atrophy can also
be noted in cystically dilated peripheral zone glands (e-Fig. 29.10) and in cystic
change in BPH nodules. Another diagnostic pitfall is that atrophic glands can
show a fragmented basal celllayer and even loss of basal cells in a few glands
by immunohistochemical stains (such as 34betaE12 and p63) (Semin Diagn
Pathol. 2005;22:88). In addition, the selective but not specific marker for neo-
plastic epithelial cells alpha-methylacyl coenzyme A racemase (AMACR) can be
immunopositive in atrophy, particularly partial atrophy.
C. Metaplasia or change in cell type in benign prostatic epithelium can be squamous,
transitional cell (urothelial), mucinous, and eosinophilic. These metaplasias are
usually secondary to inflammation, therapy, or injury. They are not preneo-
plastic.
1. Squamous cell metaplasia is most often an incidental finding associated with
inflammation and infarction in BPH nodules. Microscopically, small, solid
nests or partially involved glands with a retained lumen are common (e-
Fig. 29.11). Squamoid cytoplasm and intercellular bridges may be evident,
but keratin pearls are rare. Nuclear atypia, including prominent nuclei and
mitoses, may be present in squamous metaplasia adjacent to infarcts. Squa-
mous metaplasia postradiation or hormonal therapy can be more diffuse and
is frequently immature with less cytoplasm and an absence of keratinization.
2. Transitional cell or urothelial metaplasia should be distinguished from urothe-
lial cells that normally line the prostatic urethra and central ducts of the
prostate. This is usually a focal, incidental finding with small, solid nests
or partial gland involvement by cytologically bland and uniform elongated
cells, with some cells exhibiting nuclear grooves and cytoplasmic dearing
(e-Fig. 29.12).
3. Mucinous metaplasia is replacement of benign luminal epithelium by benign
mucin-secretory cells. This is a focal, incidental microscopic finding in which
the constituent cells have a granular blue cytoplasmic appearance (e-Fig.
29.13 ). Goblet cells and luminal secretion of the mucin are uncommon.
4. Eosinophilic metaplasia is the designation for benign epithelium with large
supranuclear eosinophilic granules, which represent exocrine differentiation
with lysosome-like granules. This uncommon and typically focal change is
more often seen in prostatic ductal epithelium and is associated with vari-
able degrees of chronic inflammation and atrophy. Paneth cell-like change,
which is also characterized by large eosinophilic cytoplasmic granules, is
usually seen in PIN and carcinoma and reflects neuroendocrine differenti-
ation. Paneth cell-like alteration may be found in nonneoplastic prostatic
epithelium after radiation.
D. Hyperplasia. BPH is a clinical diagnosis. Histologically, BPH can be diagnosed
in TURP chips and simple and radical prostatectomy specimens, but it should
not be diagnosed in needle biopsy tissue.
1. Usual nodular epithelial and stromal hyperplasia is the most common mor-
phologic presentation of BPH. Grossly, the nodules, which characteristically
arise in the transition zone and periurethral area, are multiple and vary from
solid white to spongy with cystic change.
a. Pure stromal nodules {nodular stromal hyperplasia) exist. Microscopi-
cally, the nodules can appear myxoid (e-Fig. 29.14), hyalinized, or
leiomyomatous, with spindled, ovoid, or stellate cells. Prominent thick-

walled blood vessels and lymphocytes may be noted.
b. Mixed epithelial and stromal hyperplasia is most common, with variable
admixtures of spindled stromal cells and complex benign glands with com-
plex papillary and branching architecture (e-Fig. 29.15). Cystic change,
inflammation, and basal cell hyperplasia are commonly detected in BPH
nodules. Fibroadenomatoid features (e-Fig. 29.16) can rarely be seen.
c. Epithelial predominant BPH nodules (e-Fig. 29.17) are unusual.
d. Infarcts can be identified in larger BPH nodules and can elevate the serum
PSA.
2. Basal cell hyperplasia is usually discovered in BPH nodules, but can also be
found in peripheral zone needle biopsy tissue, often associated with inflam-
mation. Microscopically, there are two or more layers of basal cells arranged
in acinat; cribriform, and solid growth patterns (e-Fig. 29.18). In usual basal
cell hyperplasia, the basal cells are uniform and cytologically bland, whereas
in so-called atypical basal cell hyperplasia prominent nucleoli are discerned.
The term "atypical" should be avoided because no form of basal cell hyper-
plasia is a known risk factor for neoplasia.
3. Cribriform hyperplasia (e-Fig. 29.19), which is completely benign and not a
risk factor for neoplasia, is an infrequently seen variant of BPH. The luminal
lining cells are cytologically bland and there is a prominent rim of basal cells.
4. Mesonephric remnant hyperplasia is a very rare prostatic proliferation dis-
playing a vaguely lobular or infiltrative pattern of small tubules with
cuboidal epithelium and intraluminal, eosinophilic secretions (Am ] Surg
Pathol. 2011;35:1054). Immunostains for high-molecular-weight cytoker-
atin (34betaE12) and/or p63 can be negative in some cases, and AMACR
can be focally positive. These results may raise concern for prostatic ade-
nocarcinoma, but helpful clues are negative PSA and prostate-specific acid
phosphatase (PSAP) but positive PAX8 immunostains.
5. Verumontanum gland hyperplasia is a benign, small gland proliferation of the
verumontanum and adjacent posterior urethra (e-Fig. 29.20). The closely
packed glands can architecturally be alarming, but the lack of nuclear atypia
and the presence of basal cells rules out carcinoma.
E. Atypical adenomatous hyperplasia (adenosis) is a nodular proliferation of closely
packed small acini (e-Fig. 29.21 ). It is invariably an incidental histologic finding,
most often found in transition zone tissue in TURP chips or prostatectomy
specimens. The densely packed small pale acini are sometimes intermingled with
larger, more complex glands. Nuclear atypia is absent to minimal. The basal cell
layer is fragmented, and, on average, 50% of glands completely lack basal cells.
Of note, AMACR is diffusely positive in about 8% of cases. Adenosis can be
mistaken for well-differentiated Gleason score 2 to 4 adenocarcinoma. It does
not have known premalignant potential.
F. Prostatic intraepithelial neoplasia (PIN) is a proliferation of atypical epithelial
cells in preexisting ducts and acini (synonyms used in the past include atypical
hyperplasia and dysplasia). Currently, PIN is graded as low-grade PIN and high-
grade PIN (HG-PIN), although only HG-PIN has potential clinical significance
and merits reporting. Isolated HG-PIN is diagnosed in about 5% to 10% of nee-
dle biopsies(] Urol. 2006;175:820). It is found in the vast majority of radical
prostatectomy specimens with prostatic carcinoma. Microscopically, there are
four major structural patterns of HG-PIN growth: tufting, micropapillary, crib-
riform, and flat (Mod Pathol. 2004;17:360) (Fig. 29.2 and e-Fig. 29.22). These
patterns are often admixed. At high-power magnification, HG-PIN shows basal
cells (which are typically reduced in number) and atypical luminal cells. Nuclear
abnormalities that should be present to diagnose HG-PIN include increased
nuclear size, increased chromatin clumping and content, and prominent nucleoli.
Figure 29.2 Architectural patterns of h.igh~grade PIN'. (Reproduced from: Humphrey PA. Prostate
Pathology. Chicago, ll.: ASCP Press; 2003. Used with permission.)
The diagnosis can usually be made on H&B-stained sections. Immunostains for

basal cells (34betaE12 and p63) and AMACR can be useful when the differ-
ential diagnosis is HG-PIN' with outpouching versus HG-PIN' with associated
invasive adenocarcinoma (Am] Surg Pathol. 2005;29:529). Isolated HG-PIN in
needle biopsy and 1URP chips has been considered a risk factor for subsequent
detection of carcinoma on rebiopsy, although the level of risk has decreased with
increased 10- to 12-core sampling of the prostate (Am] Surg Pathol. 2005;29:
1201; Urology. 2005;65: 538), such that not all patients necessarily need to
undergo rebiopsy in the first year following diagnosis of isolated HG-PIN
(] Urol. 2006;175: 820). However, patients with two or more cores with
HG-PIN do appear to be at increased risk for subsequent detection of carci-
noma and should be considered candidates for rebiopsy (] Urol. 2009;182:
485).
G. Focal glandular atypia (atypical small acinar proliferation or ASAP) is a descrip-
tive diagnosis for a gland or group of glands with architectural or cytologic
atypia that does not allow for a definitive diagnosis of reactive atypia, atypical
adenomatous hyperplasia, PIN, or carcinoma. If there is significant concern for
malignancy, a diagnosis of focal glandular atypia, suspicious for carcinoma may
be rendered. A diagnosis of atypia is applied in about 3% (range 1% to 9%) of
needle biopsies(] Urol. 2006;175: 820). Distortion artifact, section thickness,
and overstaining can contribute to difficulty in interpretation; immunostains for
basal cells (34betaE12 and p63) and AMACR can be very useful in establishing a
definitive diagnosis when atypia is the initial diagnosis in H&B-stained sections.
Patients with a diagnosis of atypia or ASAP in needle biopsy should be clini-

cally followed and rebiopsied, because about 43% of men are diagnosed with
carcinoma on rebiopsy (/ Urol. 2006;175:820}.
H. Prostate cancer, which is acinar adenocarcinoma in the vast majority of cases,
is a common malignancy in North America, Europe, and Australia, whereas it
is less common in Asia.
1. Risk factors. Proven risk factors for adenocarcinoma include age, family his-
tory, and race. Prostatic adenocarcinoma is uncommonly diagnosed clinically
before the age of 50, whereas a significant minority of men (around 31 %)
in their 30s and 40s have a small adenocarcinoma detectable at autopsy
(In Vivo. 1994;8:1459). Hereditary prostatic adenocarcinoma accounts for
about 10% of prostatic adenocarcinomas. Several candidate genes involved
in hereditary transmission have been identified (Mod Pathol. 2004;17:380},
but currently genetic testing for predisposition to prostatic adenocarcinoma
is not performed. Probable risk factors include dietary fat and androgens;
potential risk factors are cadmium, low vitamin D, low vitamin E, low sele-
nium, herbicides, and sedentary lifestyle.
2. Clinical diagnosis of clinically localized prostate cancer is based on serum
PSA and digital rectal examination (DRE). Serum PSA is widely used for
early detection in the United States, although its use for screening is contro-
versial. Serum PSA level is dearly related to risk for histologic diagnosis of
carcinoma, but risk exists below the most often used 4.0 nglmL prompt for
biopsy (N Engli Med. 2003;347:215). The DREis neither particularly sensi-
tive nor specific for a diagnosis of prostatic carcinoma. Thus, histopathologic
tissue diagnosis is the standard to establish a diagnosis of malignancy in the
prostate. Prostatic carcinoma generally does not cause symptoms until late
in the course of the disease. Local growth into the urethra and bladder neck
can cause increase in frequency and difficulty in urination. Metastatic spread
to bone can produce pain in the lower back, chest, hip, legs, and shoulders.
Response to treatment is followed by serum PSA determinations, and in some
cases, radiologic studies.
3. Histologic typing and diagnosis. The 2004 World Health Organization (WHO)
classification of neoplasms of the prostate is given in Table 29.1.
4. Acinar adenocarcinoma of the prostate is by far the most common type of
prostate cancer.
a. Gross diagnosis of prostatic carcinoma is possible in radical prostatectomy
tissues but not in needle biopsy or TURP chip tissues. Impalpable prostatic
carcinomas detected due to an elevated serum PSA (clinical stage Tlc) are
difficult to impossible to visualize on cut sections of radical prostatectomy
specimens. When visible, the carcinoma can appear nodular and white to
irregular and gray or white-yellow (e-Fig. 29.23).
b. Microscopic diagnosis is based on a synthesis of a constellation of histo-
logic attributes (Table 29.2) (/ Clin Pathol. 2007;60:35). Major criteria
are architecture (pattern of growth), absence of basal cells, and nuclear
atypia.
The architectural patterns of cellular arrangement are well depicted
in the Gleason grading diagram (Fig. 29.3). Well-differentiated prostatic
adenocarcinoma of Gleason patterns 1 and 2 displays abnormal glandular
arrangements in the form of well-circumscribed nodules of closely packed
small acini (e-Fig. 29.24). Gleason pattern 3 usually presents as single,
small, infiltrating glands, with wide stromal separation (e-Fig. 29.25).
High-grade Gleason pattern 4 is cribriform (e-Fig. 29.26}, fused small
acinar, or shows poorly formed glands. High-grade Gleason pattern 5 is
composed of sheets, cords, single cells, or comedocarcinoma (e-Fig. 29.27)
(Am I Surg Pathol. 2005;29:1228; I Urol. 2010;183:433).
Cllojbr 29 • - I ..,.
~ i (J: 1I J!.:1\ I WHO Hlttololfc Cllutflr:l.tbl•t 1-.. of Prottlte
ljtlhlllll ~~
- ......l<tlolftll
Ad fiiiiJP/tllml
(acNI) H11'1111Q ...... - .
cympltoma
........
Alrolllllc
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l'oomy l.eiJ"""'"'
-~~
C.llolil O;'adenOII'oO
SV>ot~ .. ,_pll.,_a (Wimt. tunttlf)
~ Rhobdold tt.mor
i.)m~ Go/In ooll tllmm
Oll<h>lllll 101111 Ol>llldlo eel dill'~ Yo((~ b.lmor
(o.reb:taareoma. u1oon'lil!ltti:t e~momll) Seminoma
P-lnlrlllpllte/lllnooplak (PIN)
Duct.JI ad6nccadll0mt
CJI!rlonn
~
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...
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~tllmMO
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.......,..-...-.
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_.,..... __
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-~-
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&.lcoillli/nfn
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_.
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a..1oo11oormorn•
-11101••-
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Cln:lnoldl!.mor
.......,........
illriiJ1
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u_...
"-"'1om•
,.,.,.blaelomo Malllnont~broua hll1iooyl)mo
- ..
So!Wyftbnlu& tumor
rt.llllllo-lti- Hom.a ....~.
..........
SO"olnol tumot ~ uiiC<lrtoln malllnant IIOleltllol
LAiomyou-
,.,.,..
Lelontyon•
-~~-~~-
~lnan.l
..
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Rhobdomyou,...,...
Chondrou""'""'
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Halllll*m•
---
oti&tl
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LAio~
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HoiTII ....Piflcjbna
Soliuy~broua tum"
TABLE 29.2 Criteria for Diagnosis of Prostatic Adenocarcinoma

MaJor criteria
Architectural: Infiltrative small glands or cribriform glands too large or irregular to represent
high-grade PIN
Single cell layer (absence of basal cells)
Nuclear atypia: Nuclear and nucleolar enlargement
Minor criteria
Intraluminal wispy blue mucin (blue-tinged or basophilic mucinous secretions}
Pink amorphous secretions
Mitotic figures
Intraluminal crystalloids
Adjacent high-grade PIN
Amphophilic cytoplasm
Basal cell absence, the second major criterion, can sometimes be

difficult to evaluate in H&E-stained sections, so in difficult cases and
for small foci of adenocarcinoma (minimal or limited adenocarcinoma),
immunohistochemical staining for basal cells using antibodies against
high-molecular-weight cytokeratins (such as 34betaE12, also known as
CK903) and p63 may be performed (e-Fig. 29.28). While a positive basal
Figure 29.3 Gleason grades 1 to 5.

I •,
I'1.1 I ft! 11 :!;:=:'"""'of
Cllojbr 29. -
I 1'14'1t1Uc YwM Ulo1hlllll (bnllltlonoll C:.ID
bbr
PSA 94-100
(Jill-·
~ c:MtiiiA.. Vre:te'•l CMI'
G f - pulh't)
0
•••
PSAP 89-100 0
P$MA 92 0
NIQCS.I !15 I)
p- 100 6
p63 0-3 8l..Q6
llu..,bomoduln 0 69-&1
UR>jlllkin Ill 0 67
Hlatt-molo:IJior....,qt,t ~n• o-10' 6&-100
and sarcomatoid carcinoma (carcinosarcoma) (Table 29.1). Atrophic pat-

tern adenocarcinoma displays decreased cytoplasm and can thereby mimic
benign atrophy (e-Fig. 29.32). Such cytoplasmic volume loss can be seen with
or without a history of hormonal or radiation therapy. Most cases are Glea-
son pattern 3. Pseudohyperplastic carcinoma is another malignancy that can
resemble benign glands (Am] Surg Pathol. 1998;22:1139). Here, the malig-
nant glands simulate BPH glands (e-Fig. 29.33) in that they are complex with
intraluminal papillary projections, undulating luminal surfaces, branching
patterns, and/or cystic dilatation. Gleason grade assignment is pattern 3.
Foamy gland carcinoma is characterized by xanthomatous cytoplasm and
bland nuclei (e-Fig. 29.34); these carcinomas are usually Gleason score 6 or
7, but can be higher grade. Mucinous carcinoma of the prostate is defined as
adenocarcinoma with at least 25% of the tumor composed of lakes of extra-
cellular mucin (e-Fig. 29.35); this is a rare variant, with a Gleason grade
pattern of 4 (usually) or 3. In the past, this variant was thought to be more
aggressive than usual acinar adenocarcinoma, but recent reports suggest that
it may not be. Signet-ring carcinoma of the prostate is also rare and clini-
cally aggressive; the Gleason grade is 5. Only a few cases of oncocytic and
lymphoepithelioma-like carcinoma have been reported. Sarcomatoid carci-
noma of the prostate is rare (Am] Surg Pathol. 2006;30:1316) and may be a
homologous spindle cell malignancy or heterologous, with an osteosarcoma-
taus (e-Fig. 29.36), chondrosarcomatous, or rhabdomyosarcomatous com-
ponent. In one half of the men there is a history of prostatic adenocarcinoma
treated by hormonal and/or radiation therapy; the outcome is poor. Variants
described since the 2004 WHO classification include microcystic adenocar-
cinoma (Am] Surg Pathol. 2010;34:556), PIN-like adenocarcinoma, and
pleomorphic giant cell carcinoma.
6. Ductal adenocarcinoma is the second most common subtype of prostatic ade-
nocarcinoma (after acinar). Previously known as endometrioid adenocar-
cinoma, in pure form it accounts for about 1% of prostatic cancers and,
when mixed with acinar adenocarcinoma, roughly 5% of prostatic cancers.
Microscopically, these are usually papillary and/or cribriform adenocarcino-
mas (e-Fig. 29.37) that can arise centrally (causing urinary obstruction and
hematuria) or peripherally. Cytologically, tall columnar neoplastic cells with
cleared or amphophilic cytoplasm may be observed. Most patients present
at a more advanced stage, and outcome is worse than that of acinar adeno-
carcinoma. Gleason grade is typically pattern 4.
7. Rare types of prostatic carcinoma include urothelial carcinoma (arising from
central prostatic ducts), squamous and adenosquamous carcinoma (e-Figs.
29.38 and 29.39), basal cell carcinoma, and neuroendocrine carcinoma,
including small cell and large cell neuroendocrine carcinoma (Mod Pathol.
2004;17:316). It is important to exclude primary urethral and urinary
bladder urothelial carcinoma before diagnosing primary prostatic urothelial
carcinoma. An in situ component can be extensive in the prostate, with solid
plugs of cytologically pleomorphic tumor cells, often with comedo necrosis
and stromal inflammation; prostatic stromal invasion is typified by irregular
solid nests and cords. Squamous cell and adenosquamous carcinomas of the
prostate comprise < 1% of all prostatic carcinomas, and in about two thirds
of cases there is a history of hormonal and or radiation treatment (Am] Surg
Pathol. 2004;28:651). Average survival is 2 years. Basal cell carcinoma of
the prostate includes malignant basaloid proliferations (basaloid or basal cell
carcinomas) and also neoplasms that resemble, to a certain degree, adenoid
cystic carcinomas of the salivary glands (Am] Surg Pathol. 2003;27:1523).
Basal cell carcinomas have several growth arrangements, including large
basaloid nests with peripheral palisading and necrosis, a florid basal cell
hyperplasia-like pattern, and an adenoid basal cell hyperplasia-like pattern

(adenoid cystic carcinoma pattern). Small cell carcinoma of the prostate is
quite rare and in one half of cases is admixed with adenocarcinoma; the
histologic appearance is similar to that of small cell carcinoma of the lung.
In one third of cases there is a history of prostatic adenocarcinoma followed
by hormonal therapy (e-Fig. 29.39). A similar history is obtained in most
cases of large cell neuroendocrine carcinoma of the prostate (Am ] Surg
Pathol. 2006;30:684). Outcome is very poor for neuroendocrine carcinomas
of the prostate.
8. Mesenchymal neoplasms are rare. The most common benign mesenchymal
neoplasm is leiomyoma, and the most common malignant mesenchymal neo-
plasms are rhabdomyosarcoma in children and leiomyosarcoma in adults.
Stromal tumors arising from specialized prostatic stroma include stromal
tumors of uncertain malignant potential (STUMPs) (e-Fig. 29.40) and stro-
mal sarcomas.
9. Hematolymphoid neoplasms may involve the prostate, including leukemia,
lymphoma, Hodgkin disease, and multiple myeloma. Leukemic infiltrates
almost always indicate secondary spread, usually of chronic lymphocytic
leukemia. However, about one third of prostatic lymphomas are primary,
most commonly diffuse large B cell lymphoma.
10. Miscellaneous neoplasms rarely encountered in the prostate include cys-
tadenoma, dear cell carcinoma of the utricle/prostate, paraganglioma,
melanocytic neoplasms, and germ cell tumors.
11. Secondary malignancy in the prostate is overall uncommon, but can be seen
in a substantial minority of patients with urothelial carcinoma of the bladder
(e-Fig. 29.41), leukemia, and non-Hodgkin lymphoma.
12. Treatment effects can substantially alter the morphology of prostatic carci-
noma, resulting in difficulty in diagnosis.
a. Hormonal androgen deprivation therapy can cause a decrease in the num-
ber of glands, glandular atrophy, single tumor cells, nuclear pyknosis,
and cytoplasmic vacuolization (e-Fig. 29.42). The carcinoma cells often
resemble lymphocytes or histiocytes. PSA and pan-cytokeratin immunos-
tains can be useful.
b. Radiation therapy can induce striking nuclear atypia in benign glands. Pos-
itive basal cell and negative AMACR immunostains support a diagnosis
of benign atypia. Adenocarcinoma postradiotherapy shows a decrease
in number of neoplastic glands, poorly formed glands and single cells,
cytoplasmic vacuolization, and nuclear pyknosis (e-Fig. 29.43 ). Immunos-
tains for basal cells and AMACR can be useful in diagnosis in this setting
(e-Fig. 29.44).
I. Seminal vesicles are rarely the site of origin of primary disease, whether inflam-
matory or neoplastic. Amyloid can be identified in about 10% of seminal vesi-
cles, as a function of aging; its presence does not indicate systemic amyloidosis
unless there is also co-existing vascular amyloid deposition, which is rare. Semi-
nal vesicles are usually examined for prostatic carcinoma as a part of pathologic
staging.
J. Prostatic urethra urothelium is subject to the same diseases as urothelium in the
urinary bladder, namely inflammation, metaplasia (squamous metaplasia, ure-
thritis cystica and glandularis, and nephrogenic metaplasia [adenoma]), hyper-
plasia, and neoplasms such as papilloma and carcinoma. However, primary iso-
lated malignancies of the prostatic urethra are exceedingly rare, and malignancy
in the prostatic urethra is most often due to secondary synchronous involvement
by urothelial (transitional cell) carcinoma of the urinary bladder.
IV. HISTOLOGIC GRADING OF PROSTATIC ADENOCARCINOMA. Gleason grade is crit-
ical for patient prognosis and management. It is commonly used clinically,
along with serum PSA level and clinical or pathologic stage, in tables (Partin
tables) (http://urology.jhu.edu/Partin_tables/index.html) and nomograms (] Urol.
2001;165:1562; Cancer. 2009;115(Suppl):3107) to predict pathologic stage and
response to treatment and outcome.
Grading should be performed using the modified Gleason system (Fig. 29.3). All
adenocarcinomas of the prostate should be graded, except those that are posthor-
monal therapy or postradiotherapy (when radiation effect is evident). Grading is
based solely on architecture and does not incorporate cytologic atypia or mitotic
counting. At low magnification (40 to 100x), the most common pattern and the
second most common pattern are summed to yield a score (on a scale of 2 to 10).
Recent recommendations in application of the Gleason system include the follow-
ing (Am I Surg Pathol. 2005;29:1228; I Urol. 2010;183:433):
A. Do not (or rarely) assign a well-differentiated Gleason score 2 to 4 to carcinoma
in needle biopsy. This almost always represents undergrading.
B. When three grades are present, in a needle biopsy give most common grade and
worst grade. Thus, if 3 + 4 + 5 are present, the Gleason grade is 3 + 5 = score of
8. In a radical prostatectomy, give the most common and second most common
grades, but if there is a minor tertiary high-grade 4 or 5 pattern, this should be
noted in a comment.
C. If there is 95% high-grade pattern 4 or 5 and 5% or less of 2 or 3, ignore the
lower-grade component. Any high-grade pattern (4 or 5) should be incorporated
into a needle biopsy Gleason score; thus, 98% pattern 3 and 2% pattern 4 in a
needle biopsy is Gleason grade 3 + 4 =score of 7.
D. Variants of prostatic adenocarcinoma can be graded (see above).
E. Cribriform adenocarcinomas are high-grade pattern 4.
F. For needle biopsies, provide grade by clinically submitted container, even if
several cores are within the container. Alternatively, provide the Gleason grade
for the core with highest Gleason score, if different from the overall Gleason
score for cores from that container.
V. PATHOLOGIC STAGING. Staging applies only to adenocarcinomas of the prostate, and
not to sarcomas or prostatic carcinoma variants that are not adenocarcinomas.
The 2010 Tumor, Node, Metastasis (TNM) American Joint Committee on Cancer/
International Union Against Cancer (AJCGUICC) staging classification is given in
Table 29.4. Clinical staging should be distinguished from pathologic staging.
A. Needle biopsy. Pathologic staging is not performed. However, extraprostatic
spread should be diagnosed if carcinoma is seen in fat or in seminal vesicle
tissue.
B. TU RP chips and open prostatectomy. Pathologic staging is not done, but for inci-
dental carcinoma, the amount of carcinoma determined by light microscopic
inspection of tissue involved will place the patient into clinical stage Tla or Tlb.
C. Radical prostatectomy and pelvic lymphadenectomy are used for pathologic
staging (Table 29.4) (Arch Pathol Lab Med. 2009;133:1568).
1. pT2: Organ-confined prostatic carcinoma, subdivided into a, b, and c.
2. pT3: Extraprostatic extension (EPE) by carcinoma, diagnosed if carcinoma
extends into posterolateral periprostatic adipose tissue (e-Fig. 29.45), beyond
the outer boundary of normal prostatic glands at the anterior or apical
prostate, or microscopically into the bladder neck. Note that carcinoma in
skeletal muscle at apex does not always mean EPE. Site(s) and extent ofEPE
should be specified. EPE extent should be given as focal (only a few glands
outside the prostate) or nonfocal. Capsular invasion is not part of the staging
scheme. pT3b is seminal vesicle wall invasion (e-Fig. 29.46).
3. pT4: Gross bladder neck or rectal involvement by carcinoma.
4. pN: Number of involved and total number of examined lymph nodes should
be given.
5. pM: At time of radical prostatectomy, patients are clinical MO (cMO).
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412 I SECTION VII· REPRODUCTIVE TRACl
1' 1,1,! • t ~ tS l l'IIIMII!Wt. ~~e~uta~~~t (1'NIIO . . . _ s.:~~em. hw Prottatfc

' - Cl!el- (t.WIIIIIMl
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T2b NO MC> PSA <20 C' ln ~7
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Tl-2 NO MQ PSA ~ Any\lloooon
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SUGCiESTED READING
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-"""-Ill:,.. c{nc...w ,~ 411- ~DC:..._..., ll.<glotrJ' of
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Penis and Scrotum
Peter A. Humphrey
I. NORMAL ANATOMY. The penis is anatomically composed of three parts: posterior

(root); central body or shaft; and anterior portion composed of glans, coronal sul-
cus, and foreskin (prepuce). In the shaft there are three cylinders of erectile tissues:
a ventral corpus spongiosum surrounding the urethra and two corpora cavernosa.
Histologically, the erectile tissues are characterized by numerous vascular spaces
with surrounding smooth muscle fibers (e-Fig. 30.1).• The tunica albuginea, a
sheath of hyalinized collagen, encases the corpora cavernosa. All three corpora
are surrounded by Buck's fascia, adipose tissue, dartos muscle, dermis, and a thin
epidermis. Distally, the corpus spongiosum forms the conical glans, which is also
composed of a stratified squamous epithelium, lamina propria, tunica albuginea,
and corpora cavernosa. The coronal sulcus is a cul-de-sac just below the glans
corona. The foreskin is a double membrane that has five layers: mucosal epithe-
lium similar to glans epithelium, lamina propria, dartos smooth muscle, dermis,
and epidermis.
The scrotum contains the testes and lower spermatic cords. It consists of skin
that covers the dartos smooth muscle, fibers of the cremasteric muscle, and several
layers of fascia. The skin is pigmented, hair bearing, and loose, with numerous
sebaceous and sweat glands. Lymphatic drainage is to the superficial inguinal lymph
nodes.
II. GROSS EXAMINATION AND TISSUE SAMPLING. Tissue samples include mucosal or
skin biopsies, penile urethral biopsies, foreskin resection specimens, and partial
and total penectomy specimens.
A. Punch and shave biopsies of penile glans and skin should be handled as skin
biopsies from other sites (see Chap. 38).
B. Foreskin resection is indicated for primary carcinomas of this site. The entire
periphery of the mucosal margin should be submitted as a shave resection margin
(usually in three to four sections). The foreskin should then be pinned and fixed
overnight in 10% formalin. Several full-thickness sections should be examined
microscopically to permit evaluation of all five layers.
C. For partial penectomy specimens, the surgical division of the penis is made
2 em proximal to gross tumor extent. Three to four frozen sections are typically
necessary to sample the cut surface of this margin. Permanent sections are taken
as described below.
D. For total penectomy specimens, only proximal urethral and periurethral margin
tissues should be submitted for frozen section, unless the mass is grossly close to
or involves the skin, which should also then be sampled. For permanent sections
of both partial and total penectomy specimens, the foreskin (when present)
should be removed and handled as noted previously. A thin 2-mm shave of all
the structures of the shaft margin should be taken, if not already sampled by
frozen section. One to three additional transverse sections should be taken from
the glans. Any mass(es) should be sampled to demonstrate pattern of growth,
depth of extension, and relationship to normal anatomic structures.
E. Lymph nodes. There may be a clinical request for frozen section(s) of enlarged
inguinal lymph nodes; if positive for carcinoma, a more extended ilioinguinal
483
lymph node dissection may follow. Bilateral inguinal lymphadenectomy speci-

mens may also be received after removal of the primary tumor and a course of
antibiotics, or in patients with T2 tumors, high-grade tumors, or tumors with
vascular invasion (Crit Rev Oncol Hematol. 2005;53:165).
A nomogram has been developed to predict nodal metastases using the pres-
ence of clinically palpable groin lymph nodes and histologic lymphovascular
invasion in the primary tumor(] Urol. 2006;175:1700). The utility of sentinel
lymph node sampling is not yet settled. A prognostic index has also been gener-
ated to predict nodal metastasis (Am J Surg Pathol. 2009;33:1049); this index
incorporates histologic grade, deepest anatomic level involved by cancer, and
the presence of perineural invasion by carcinoma.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF THE PENIS AND SCROTUM
A. Inflammation and infection. Three categories can be defined: inflammatory con-
ditions specific to penis and scrotum, systematic dermatoses (discussed in Chap.
38), and sexually transmitted diseases (BJU Int. 2002;90:498).
1. Phimosis, the clinical condition in which the foreskin cannot be retracted
behind the glans penis, is associated with fibrosis, inflammation, and edema
of the prepuce.
2. Paraphimosis is diagnosed clinically when the foreskin cannot be advanced
back over the glans secondary to fibrosis and inflammation.
3. Balanoposthitis is an inflammation of the glans penis and prepuce, usually in
uncircumcised men with poor hygiene.
4. Balanitis, or inflammation of the glans, occurs in several forms.
a. Plasma cell balanitis (Zoon balanitis) can clinically and grossly mimic car-
cinoma in situ, with presentation as brown or red patches or plaques. The
histologic appearance can vary with time, and the inflammatory cell infil-
trate can vary from patchy and lymphoplasmacytic (early) to dense and
plasmacytic (later) (Am] Dermatopathol. 2002;24:459).
b. Balanitis xerotica obliterans (BXO) is lichen sclerosus of the glans and
prepuce that macroscopically appears as a white patch or plaque. His-
tologically, the squamous epithelium is typically atrophic and hyperkera-
totic, with a band of pale homogenous collagen in the upper dermis and
an underlying lymphocytic infiltrate (e-Fig. 30.2). Complications include
meatal stenosis (urethral stricture), and very uncommonly, squamous cell
carcmoma.
c. Balanitis circinata microscopically resembles pustular psoriasis and is seen
in Reiter syndrome, which includes nongonococcal urethritis, conjunctivi-
tis, and arthritis.
5. Human papillomavirus (HPV) infection can lead to condyloma acuminata. The
growth is typically papillary or warty, and histologically papillomatosis,
acanthosis, parakeratosis, and hyperkeratosis are found; intraepithelial neo-
plasia may also be present. Koilocytes with wrinkled nuclear membranes,
nucleomegaly, cytoplasmic halos, and binucleation may be prominent or
inconspicuous. The causal HPV serotypes are usually 6 or 11, but it is not
necessary to verify the presence of HPV. Bowenoid papulosis is also an HPV-
related proliferation that presents with multiple 2 to 10 mm papules than
can coalesce to form plaques; it is usually caused by serotypes 16, 18, and/or
35. Microscopically, although the appearance is similar to carcinoma in situ,
the clinical course is typically self-limited and benign.
6. Herpes simplex virus (HSV} infection of the male genitalia is usually caused by
subtype 2, which produces multiple vesicles. The diagnosis may be confirmed
by scraping and performance of a Tzanck smear, which reveals multinucle-
ated giant cells with intranuclear inclusions.
7. Scabies is an infestation by a mite that burrows into the keratin layer of the
epidermis with generation of erythematous papules and nodules. Detection
Chapter 30 • Penis and Scrotum I 4 85
of the mites may be accomplished via scrapes or biopsy. In tissue sections,

the 400 ~m mite, eggs, or egg walls, are diagnostic. Dermal eosinophils and
epidermal spongiosis are characteristic responses.
8. Pediculosis pubis is an infection by Pediculus pubis, also known as the crab
louse. Biopsy is not necessary; the lice may be seen by a magnifying lens.
9. Syphilis is caused by Treponema pallidum, a gram-negative spirochete. The
primary lesion, the chancre, is a single, round, craterlike painless ulcer most
often located on the glans or prepuce. Biopsy is not usually necessary, but if
done (e.g., when syphilis is not clinically suspected) it shows a perivascular
lymphoplasmacytic infiltrate. The spirochetes may be identified in the epi-
dermis or in dermal perivascular regions by silver stains (Steiner, Dieterle,
or Warthin-Starry). The secondary and tertiary stages of syphilis are char-
acterized by condyloma latum and gumma, respectively. Smears from the
gray maculopapules of condyloma latum should be examined by dark-field
microscopy for spirochetes since biopsy may yield nonspecific findings. The
gumma is a necrotic mass with surrounding granulomatous inflammation,
and associated obliterative endarteritis with perivascular plasma cells.
10. Gonorrhea is caused by Neisseria gonorrhoeae, a gram-negative diplococcus.
Urethritis with urethral discharge may lead to urethral stricture. Biopsies are
hardly ever performed.
11. Lymphogranuloma venereum is due to Chlamydia trachomatis. Vesicles, then
ulcers, develop in the primary genital phase; biopsies are not useful for diag-
nosis of the primary phase. In the secondary stage, patients develop painful
inguinal lymphadenopathy (bubo). Histologically, the lymph nodes demon-
strate stellate necrosis surrounded by palisaded histiocytes, a nonspecific pic-
ture that can also be seen in cat scratch fever, tularemia, bubonic plague, and
fungal and atypical mycobacterial infections.
12. Granuloma inguinale is caused by Calymmatobacterium granulomatis, a
gram-negative intracellular bacillus. Infection results in ulcers. Smears or
biopsy sections reveal histiocytes with inclusions (Donovan bodies), best
visualized by Warthin-Starry or Giemsa stains.
13. Chancroid is caused by Haemophilus ducreyi, a gram-negative rod, and is
typified by painful nonindurated penile ulcers and lymphadenopathy. The
organisms can be found in smears or histologic sections stained with Giemsa,
Gram, or methylene blue stains.
14. Molluscum contagiosum is caused by a DNA poxvirus and produces multiple
small dome-shaped papules with a central umbilication. Biopsy sections show
a crater with an acanthotic epidermis and the diagnostic intracytoplasmic
viral inclusions (molluscum bodies).
15. Penile infections in acquired immunodeficiency syndrome (AIDS) include almost
all sexually transmitted infections including gonorrhea, syphilis, herpes, can-
didiasis, chancroid, molluscum contagiosum, HPV, scabies, and Reiter syn-
drome.
16. Lipogranulomas in the scrotum or penis are secondary to injections of oil-
based chemicals. Sections show a foreign body, lymphoplasmacytic, and his-
tiocytic reaction to lipid droplets that appear as cleared spaces.
17. Hidradenitis suppurativa, more typical of the sweat glands in the axilla, can
also involve the scrotum. Acute and chronic inflammation, fibrosis, and even
sinus tract formation can occur.
18. Gangrene, as a necroinflammatory process, can involve the scrotum due to a
variety of insults. Fournier gangrene is an extreme fulminant infection of the
genitals, perineum, or abdominal wall (lnt j Urol. 2006;13:960).
19. Idiopathic scrotal calcinosis can occur due to calcification of dermal con-
nective tissue (idiopathic) or in association with keratinous cysts. Multiple
firm nodules are found, measuring several millimeters to several centimeters
in greatest dimension, typically in the scrotum of younger men. Microscopi-

cally, calcific material is present with or without granulomatous inflammation
and/or cyst wall remnants.
20. Elephantiasis or massive scrotal lymphedema is usually secondary to filariasis.
B. Miscellaneous benign nonneoplastic conditions
1. Peyronie disease presents with a painful bending of the erect penis. Histolog-
ically, there is fibrosis or fibromatosis of the tunica albuginea (e-Fig. 30.3).
Rarely, calcification and ossification may occur in these fibrous plaques.
2. Penile cysts
a. Median raphe cyst is most commonly located in the midline on the ventral
aspect of the shaft. It is lined by pseudostratified, columnar, mucinous
epithelial cells.
b. Mucoid cysts are thought to arise from ectopic urethral mucosa, can be seen
on the prepuce or glans, and are lined by stratified columnar epithelium
with mucinous cells.
c. Epidermal inclusion cysts are usually found on the penile shah. They are
also common in scrotal skin. They have the same appearance as they do
elsewhere in the skin.
C. Benign epithelial neoplasms include squamous papilloma, common condyloma,
and the very rare giant condyloma of Buschke-Lowenstein. Most reported
giant condylomas likely represent warty or verrucous carcinomas. Grossly, giant
condylomas are 5 em in average diameter, and microscopically resemble a typi-
cal condyloma except for exuberant surface papillomatosis and pushing bulbous
growth at the base. When carefully defined, they may be locally destructive but
do not show malignant cytologic features or metastasize.
D. Penile intraepithelial neoplasia (PeiN}. Carcinoma in situ (or high-grade squa-
mous intraepitheliallesions) includes erythroplasia of Queyrat (on the glans)
and Bowen disease (on the penile shaft or prepuce). These are HPV-related (usu-
ally serotypes 16 or 18) intraepithelial proliferations. Grossly, erythroplasia of
Queyrat appears as a sharply demarcated patch on the glans, whereas Bowen dis-
ease is a solitary, brownish-red plaque on the shah or foreskin. Microscopically,
the two lesions are similar and are composed of a full-thickness proliferation
of pleomorphic basaloid cells that exhibit loss of polarity (e-Fig. 30.4). Tradi-
tionally, these two high-grade intraepithelial proliferations have been assigned
grade III, while lower grade intraepitheliallesions have been graded as I or II.
Recently, a new PeiN classification scheme has been proposed with cate-
gories of differentiated, undifferentiated, and mixed differentiated and undiffer-
entiated (Am] Surg Pathol. 2010;34:385). Differentiated PeiN is characterized
by hyperkeratosis, parakeratosis, hypergranulosis, acanthosis, elongated rete
ridges, abnormal maturation, intraepithelial pearl formation, prominent inter-
cellular bridges, and atypical basal or prickle layer cells. In the undifferentiated
category are warty, basaloid, and warty-basaloid variants.
E. Penile cancer
1. Risk factors for penile cancer are phimosis, chronic inflammation, lichen scle-
rosus, smoking, ultraviolet (UV) radiation, condyloma, HPV infection (usu-
ally type 18, less commonly type 18), and lack of circumcision(] Am Acad
Dermatol. 2006;54:364). This latter factor is an extremely strong risk factor.
2. Clinical diagnosis. Patients are typically 50 to 70 years of age and present
with an exophytic or flat ulcerative mass of the glans, prepuce, or coronal
sulcus. Patients with advanced disease may present with pubic or scrotal skin
nodules and inguinal lymph node metastasis. Magnetic resonance imaging
of primary penile cancer can help define local tumor extent and any shah
involvement (Radiographies. 2005;25:1629).
3. Histologic typing and diagnosis. The 2004 World Health Organization (WHO)
classification of penile neoplasms is provided in Table 30.1.
Cltlptet 30 • ~it ud Sttebim I 417
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Figure 30.1 Verruciform tumors ofthe penis. A: Verrucous carcinoma with regular papillae, broad
pushing base, and hyperkeratosis. B: Papillary carcinoma with irregular papillae and cores, and
ragged infiltration at base. C: Giant condyloma, with branching cores and koilocytosis. D: Warty
(condylomatous) carcinoma with irregular papillae and koilocytosis. (Modified from: Young RH,
Srigley JR, Amin MB, et al., eds. Tumors of the Prostate Gland, Seminal Vesicles, Male Urethra,
and Penis. Washington DC: Armed Forces Institute of Pathology; 2005:424.)
the distinction of a poorly differentiated squamous cell carcinoma from

another malignant neoplasm such as a sarcoma, melanoma, or lymphoma.
Immunohistochemical detection of carcinoma cells in lymph nodes is of
uncertain significance (BJU Int. 2006;.98:70).
d. Molecular studies are not currently used to diagnose penile carcinoma.
HPV typing is not necessary.
5. Variants of squamous cell carcinoma include basaloid, warty (condyloma-
tous), verrucous, papillary, sarcomatoid, mixed, and adenosquamous (Table
30.1) (Urology. 2010;76(suppl2A):S7). Basaloid carcinoma is an uncom-
mon, aggressive, HPV-related variant that comprises small, uniform, basa-
loid cells with high-nuclear/cytoplasmic ratios and numerous mitoses (e-Fig.
30.9). Warty (condylomatous) carcinoma is one of the verruciform carcino-
mas (the others being verrucous carcinoma and papillary carcinoma) (Fig.
30.1); microscopically, it is hyperkeratotic and papillomatous with cells of
low to intermediate nuclear grade, and koilocytic atypia may be prominent.
Verrucous carcinoma is a very well-differentiated papillary neoplasm with
hyperkeratosis, papillomatosis, and a broad pushing base. Papillary carci-
noma is well differentiated, hyperkeratotic, and has complex papillae and
an irregular infiltrative base; it has a favorable prognosis (Am] Surg Pathol.
2010;34:223). Sarcomatoid (spindle cell) carcinoma is an aggressive, high
Chapter 30 • Penis and Scrotum I 489
grade, deeply invasive spindle cell malignancy with or without heterologous

elements such as muscle, bone, and cartilage; coexisting carcinoma in situ or
invasive carcinoma is usually evident (Am j Surg Pathol. 2005;29:1152). In
adenosquamous carcinoma the glandular component is a minority compo-
nent. In one quarter of cases the carcinoma can be mixed, such as warty-
basaloid, adenocarcinoma-basaloid, and squamous-neuroendocrine.
Rare, recently recognized patterns of squamous cell carcinoma
include pseudohyperplastic squamous cell carcinoma (Am f Surg Pathol.
2004;28:895), carcinoma cuniculatum (Am] Surg Pathol. 2007;31:71), and
pseudoglandular (adenoid, acantholytic) squamous cell carcinoma (Am f
Surg Pathol. 2009;33:551 ). Pseudohyperplastic squamous cell carcinoma can
resemble pseudoepitheliomatous hyperplasia, and carcinoma cuniculatum
is another verruciform squamous cell carcinoma that is low grade and has
deeply penetrating and burrowing patterns of growth. Pseudoglandular squa-
mous cell carcinoma is often deeply infiltrative and of high histologic grade.
6. Rare types of primary penile carcinoma include Merkel cell carcinoma, small
cell carcinoma, sebaceous carcinoma, and clear cell carcinoma.
7. Mesenchymal neoplasms are rare and comprise only 5% of all penile tumors.
The most common benign soft tissue tumors are vascular (hemangioma and
lymphangioma), followed by neural, myxoid, and fibrous tumors. The most
frequent malignant soft tissue tumors are Kaposi sarcoma and leiomyosar-
coma (Anal Quant Cytol Histol. 2005;28:193).
8. Hematolymphoid neoplasms include very rare primary penile lymphomas and
secondary lymphomas.
9. Miscellaneous neoplasms rarely encountered include melanoma (] Urol.
2005;173:1958).
10. Secondary malignancies are rare, with prostatic and urinary bladder carcino-
mas predominating (Int j Surg Pathol. 2010; Jan 14, epub ahead of print).
The corpus cavernosum is the most common site of metastasis, but the spon-
giosum, skin, and glans may also be involved.
F. Scrotal cancer is most commonly squamous cell carcinoma.
1. Squamous cell carcinoma of the scrotum is usually detected as invasive dis-
ease, but some examples of squamous cell carcinoma in situ have been
reported. Associations exist with exposure to soot (in chimney sweeps,
described in 1775 by Sir Percival Pott), machine oil, psoriasis treated with
coal tar, arsenic, psoralens/UV radiation, and HPV infection. Grossly, scrotal
squamous cell carcinomas initially appear as a solitary nodule; later in their
course they show ulceration and induration. Microscopically, most are well
to moderately differentiated (e-Fig. 30.10). The tumor typically invades the
scrotal wall, and larger cancers can involve the testis, spermatic cord, penis,
and perineum. Initial metastatic spread is to ipsilateral inguinal lymph nodes.
Outcome is related to tumor size and pathologic stage.
2. Basal cell carcinomas of scrotal skin are rare and have the same appearance
as they do elsewhere in the skin.
3. Paget disease of the scrotum can be associated with underlying carcinoma
of the urinary bladder, urethra, prostate, or eccrine sweat glands.
4. Sarcomas of the scrotal wall are rare and should be distinguished from parat-
esticular, intrascrotal sarcomas. By far the most common histologic type of
scrotal wall sarcoma is leiomyosarcoma, which likely arises from the dartos
muscle.
IV. HISTOLOGIC GRADING OF PENILE AND SCROTAL SQUAMOUS CELL CARCINOMAS is done
in three tiers as well differentiated, moderately differentiated, or poorly differen-
tiated. Histologic grade of penile squamous cell carcinoma is linked to depth of
infiltration, inguinal lymph metastasis, and survival.
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Chapter 30 • Penis and Scrotum I 4 91
T=pT
I I
Ta: Non-invasive venvcous carcinoma
T1: Tumor invading subepithelial connective tissue
""'
T2: Tumor invading corpus spongiosum or cavemosum
T=pT
T3: Tumor invading urethra or prostate
pT4
T4: Tumor invading other adjacent structures
Figure 30.2 pT staging of penile carcinoma. (Modified

from: Greene FL, Compton CC, Fritz AG, et al., eds. A]CC
Cancer Staging Atlas. New Yor~ NY: Springer; 2006.)
tumor: tumor size, histologic type, histologic grade (Gl, G2, or G3), origin, depth
of invasion (mm), anatomic level of invasion, structures involved, vascular and
perineural invasion (if present), and status of the margins of resection. For regional
lymph nodes, the report should include the number identified and their location,
number involved by tumor, size of metastatic deposit (if present), and extracapsular
extension (if present). Additional pathologic findings that can be noted (if present)
are penile intraepithelial neoplasia and therapy-related changes.
SUGGESTED READING
Epstein JI, Cubilla AL, Humphrey PA. Tumors of the prostate gland, seminal vesicles, penis, and
scrotum. In: Atlas of Tumor Pathology. 4th Series. Washington, DC: American Registry of
Pathology; 2011.
The Ovary
Meredith E. Pittman, John D. Pfeifer,
and Phyllis C. Huettner
I. NORMAL GROSS AND MICROSCOPIC ANATOMY. The ovaries rest on either side of the
uterus and are anchored by the broad ligament. Age and reproductive status greatly
impact size and weight, which range from 3 to 5 em and 5 to 8 g, respectively.
The outer surface of the ovary is white-tan and smooth during early reproductive
years but becomes more bosselated with age due to repeated rupture of ovarian
follicles. The cut surface is vaguely organized into three ill-defined zones: an outer
cortex, underlying medulla, and inner hilum. The cortex and medulla contain cystic
follicles, yellow or orange corpora lutea, and gritty, white corpora albicantia.
Histologic sections of the ovary show a simple cuboidal surface epithelial layer
derived from mesothelium. Within the ovarian cortex and medulla, follicular struc-
tures composed of an inner layer of granulosa cells and an outer layer of theca cells
(e-Fig. 31.1* and e-Fig. 31.2), in varying phases of development, are surrounded
by a stroma comprised closely packed S-shaped spindled cells and collagen (e-Fig.
31.3 ). Centrally hemorrhagic corpora lutea composed of granulosa cells with abun-
dant eosinophilic cytoplasm and smaller theca cells (e-Fig. 31.4 and e-Fig. 31.5),
as well as white acellular corpora albicantia (e-Fig. 31.6), may also be seen. The
hilus, where the ovary connects to the broad ligament, is composed of abundant
blood vessels, nerves, and interspersed eosinophilic hilar cells that are histologically
similar to the Leydig cells of the testis (e-Fig. 31.7). Rete ovarii, the developmental
analogue of the rete testis that are composed of slit-like spaces lined by nonciliated
cuboidal epithelium, are also present in the hilum (e-Fig. 31.8).
II. GROSS EXAMINATION AND TISSUE SAMPLING. The most common ovarian specimens
encountered in surgical pathology are from oophorectomy (with or without hys-
terectomy) or cystectomy procedures. The weight and gross measurements of all
three dimensions are recorded. The capsule is inspected for areas of rupture, adhe-
sions, tumor involvement, or other abnormalities. In most cases, the ovary is then
bivalved along the long axis and any lesions on the cut surface are noted. Solid, cys-
tic, or papillary lesions are thoroughly sampled (one section per em). If no lesions
are identified, one section for every 2 em is adequate. If cysts are present, the color
and consistency of the cyst fluid are noted, and any areas of nodularity or papillary
excrescences are sampled. Prophylactic oophorectomy specimens, performed for a
personal history of cancer or family history of a hereditary cancer syndrome, are
cut perpendicular to the long axis and entirely submitted.
The surgical management of malignant primary ovarian tumors typically
includes a staging procedure, and so an ovary excised for a primary malignancy
will usually be accompanied by multiple abdominal-peritoneal biopsies, an omen-
tectomy specimen, and regional lymph nodes. The small peritoneal biopsies are
submitted entirely. The omentum sample must be serially sectioned. If grossly vis-
ible tumor is present, it should be measured and only one section needs to be
submitted; when no tumor is identified, from 5 to 10 sections are submitted. All
identified lymph nodes are submitted from the lymph node dissections.
Ill. DIAGNOSTIC FEATURES OF COMMON BENIGN DESEASES OF THE OVARY
A. Inflammatory Diseases
1. Infection of the ovary is almost always accompanied by infection of the
fallopian tube (e-Fig. 31.9), systemic infection, or the development of a
493
tubo-ovarian abscess (e-Fig. 31.10). The most common cause of oophori-

tis is ascending pelvic inflammatory disease (PID), which typically causes
intense pelvic pain and is usually polymicrobial. The ovary contains sheets
of neutrophils and often shows paraovarian adhesions or adhesions to an
inflamed fallopian tube. Severe PID may require hospitalization and treat-
ment, while mild PID may resolve on its own. The subsequent fibrosis and
scarring from PID is a leading cause of infertility (e-Fig. 31.11).
2. Autoimmune oophoritis typically presents with oligomenorrhea and infertil-
ity and is often associated with other autoimmune disorders. The ovaries
are normal in size but filled with a lymphoplasmacytic infiltrate in devel-
oping follicles and corpora lutea. Premature ovarian failure and premature
menopause is the final sequelae of the disease.
3. Noninfectious granulomas may also be seen within the ovary as an incidental
finding. These rnay form secondary to systemic diseases such as sarcoidosis,
or represent a foreign body response following a pelvic or abdominal surgery.
B. Cysts
1. Follicular cysts are commonly found in prepubescent and reproductive-aged
women. Normal follicles measure only up to 1 em in greatest dimension,
while follicular cysts measure 2.5 to 10 em in diameter and contain serosan-
guineous fluid. Most cysts are asymptomatic, but women may present with
an abdominal mass or rupture. Follicular cysts are typically unilateral with a
thin, smooth lining comprised an inner layer of granulosa cells and an outer
layer of theca cells similar to normal follicles. Multiple follicular cysts may be
associated with polycystic ovarian syndrome (PCOS) and McCune-Albright
syndrome.
2. Corpus luteum cysts are also common in women of reproductive age. Like
follicular cysts, they may present as a mass or with rupture. On gross exam-
ination, corpus luteum cysts are >3 em, filled with thick hemorrhagic fluid,
and rimmed by a yellow lining. Microscopically, the cyst has an undulating
wall of luteinized granulosa cells (that have abundant, eosinophilic cyto-
plasm) with interspersed peripheral theca cells with an overall architecture
similar to that of a normal corpus luteum.
3. Polycystic ovarian syndrome is an incompletely understood disease character-
ized by anovulation, infertility, hirsutism, and obesity. It is estimated that up
to 10% of American women are affected by this syndrome, which typically
presents between the ages of 20 and 30. In general, the ovaries are enlarged
with a thickened, collagenized cortical surface; multiple, uniform follicles
all in a similar phase of development (typically antral); and an absence of
corpora lutea (indicting infrequent ovulation) (e-Fig. 31.12).
4. Surface epithelial inclusion cysts, thought to be formed by repeated invagi-
nations of ovarian epithelium secondary to surface rupture with ovulation,
are common. By convention, inclusion cysts measure < 1 em in diameter. The
cysts are lined by a single layer of bland flat, cuboidal, or ciliated columnar
cells (e-Fig. 31.13). Psammoma bodies may be seen. It is hypothesized that
benign, borderline, and low-grade serous epithelial neoplasms arise from
these inclusions.
5. Paraovarian or paratubal cysts are found in the hilar region of the ovary
and arise from mesonephric (Wolffian) or paramesonephric (Mullerian) duct
remnants. Mesonephric cysts are lined by simple or stratified epithelium and
have prominent muscular walls (e-Fig. 31.14). Paramesonephric cysts are
lined by columnar epithelium with a mixture of ciliated and nonciliated cells
similar to epithelial inclusion cysts (e-Fig. 31.13).
6. Endometriosis, characterized by endometrial glands and stroma outside the
uterus, is frequently seen within the ovary. Symptomatic patients typically
present during the reproductive years with menstrual-associated pain and
Chapter 31 • The Ovary I 4 95
infertility. Grossly, the ovary shows a thickened cortex, surface adhesions,

and a cyst filled with thick brown fluid resembling chocolate syrup, the
so-called chocolate cyst. Histologically, endometrial epithelium or glands
and stroma with associated hemosiderin-laden macrophages and fibro-
sis are diagnostic (e-Fig. 31.15). Endometriotic cysts frequently contain
areas with atypical cytologic features associated with acute inflammation,
but areas of epithelial tufting, more complex architectural changes, or
increased mitotic activity should raise suspicion for malignancy arising in
endometriosis.
C. Hyperplastic changes
1. Stromal hyperplasia, defined as a proliferation of non-luteinized ovarian
stroma in the cortex and medulla, and hyperthecosis, luteinized stromal cells
in a background of stromal hyperplasia (e-Fig. 31.16), are changes most
commonly seen in the sixth to seventh decades of life and in women with
PCOS. The ovaries are enlarged bilaterally. While most women are asymp-
tomatic, some present with estrogenic symptoms (endometrial hyperplasia)
or androgenic symptoms (acne) due to hormone production by the luteinized
cells.
2. Hilus (Leydig) cell hyperplasia is most common in postmenopausal women
who also have stromal hyperplasia and hyperthecosis. The hilus cell pro-
liferation rarely causes symptoms on its own. Microscopically, eosinophilic
cells with hyperchromatic nuclei are arranged in nests or clusters found most
often in the hilar region of the ovary. Elongated, eosinophilic intracytoplas-
mic inclusions, called Reinke's crystals, may be seen (e-Fig. 31.17).
D. Pregnancy changes
1. Solitary luteinized follicle cysts are benign, unilocular, unilateral cysts that
may grow up to 25 em in diameter. They have the same lining of inner
granulosa cells and outer theca cells as normal follicles.
2. Hyperreactio luteinalis is characterized by symmetric, bilateral enlargement
of the ovaries with multiple luteinized cysts. The condition is associated with
high levels of human chorionic gonadotropin, which also occurs with multi-
ple pregnancies, ovulation induction protocols, and gestational trophoblastic
disease. The cysts usually resolve in the months after completion of preg-
nancy. Microscopically, the follicular cysts are lined by luteinized granulosa
and theca cells with stromal edema.
3. Pregnancy luteoma is seen in the second half of pregnancy and is most com-
mon in multiparous African American women. The lesion consists of hyper-
plastic proliferations of luteinized cells, likely stromal and theca cells. Most
pregnancy luteomas are incidental findings, but 25% of patients will have
symptoms of virilization. Grossly, the nodules are multiple (50%), bilat-
eral (33%), yellow-brown, hemorrhagic, and soft. Microscopically, they are
well-circumscribed collections of large eosinophilic cells that may have many
mitotic figures (e-Fig. 31.18). The nodules regress after pregnancy.
E. Ovarian torsion is frequently accompanied by torsion of the fallopian tube as
well. Torsion is caused by twisting of the adnexa on its fibrovascular pedicle
impeding blood flow into and out of the adnexal structures, ultimately resulting
in infarction. The ovary is often enlarged and hemorrhagic or dusky. Microscop-
ically, there is extensive interstitial hemorrhage, edema, and necrosis of normal
tissue. Frequently there is an associated ovarian mass or neoplasm. Pregnant
women and women with adnexal masses are at increased risk, but torsion can
occur in children, infants, and occasionally in utero. Patients typically present
with acute abdominal pain, nausea, and vomiting, and may have a palpable
adnexal mass.
IV. OVARIAN NEOPLASMS. Table 31.1 presents a simplified version of the World Health
Organization (WHO) histologic classification of tumors of the ovary.
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Chapter 31 • The Ovary I 499
A. Surface epithelial-stromal tumors. Tumors of ovarian surface epithelium rep-

resent approximately two-thirds of all ovarian neoplasms. Ovarian surface
epithelium may differentiate into any Miillerian-type epithelium, accounting
for the various subtypes of ovarian epithelial tumors: serous (resembling fallop-
ian tube), endometrioid (resembling endometrium), and mucinous (resembling
endocervix). Clear cell and transitional tumors have no normal counterpart in
the gynecologic tract. The standard of treatment for these tumors is surgery, with
the goal of staging and optimally debulking the tumot; followed by chemother-
apy for all but certain stage I tumors.
1. Serous tumors are the most common subtype of surface epithelial tumors. The
majority of serous tumors are benign and occur in women aged 30 to 40 years.
About 15% are borderline (also known as low malignant potential [LMP]
or atypical proliferating) tumors. About 25% are malignant. The neoplastic
epithelium, particularly in benign and borderline tumors, resembles fallopian
tube epithelium and contains ciliated and secretory cells; the cystic spaces
contain serous or serosanguineous fluid.
a. Serous cystadenoma/cystadenofibroma. These benign lesions make up
over half of all serous tumors, and they are most commonly found in
reproductive-aged women. Grossly they are uniloculat; smooth-walled
cysts with varying amounts of fibrous stroma, usually ranging from 1 to
10 em. Histologically, the cystadenoma lining is a single layer of colum-
nar and usually ciliated epithelium (e-Fig. 31.19). Cystadenofibromas
show broad stromal papillae lined by simple columnar ciliated epithelium
(e-Fig. 31.20). The lining may be attenuated in longstanding lesions.
Lesions without a cystic component are termed adenofibromas.
b. Borderline serous tumors (serous tumors of low malignant potential) are pre-
dominantly cystic and generally present in perimenopausal women. As
many as one-third are bilateral. These tumors have papillary excrescences
with hierarchically branching stromal cores. The lining alls show mild
to moderate cytologic atypica and demonstrate mildly increased mitotic
activity, stratification, tufting, and detached buds of epithelium (e-Fig.
31.21). Psammoma bodies are frequently seen. By definition these tumors
do not demonstrate stromal invasion.
Although most borderline serous tumors are confined to the ovary,
extraovarian disease such as peritoneal implants may be present at diag-
nosis. The classification of the tumor is then based on the features found
within the primary ovarian tumot; not the extraovarian disease. Peri-
toneal implants associated with borderline tumors are classified as inva-
sive or noninvasive (Cancer. 1988;62:2212). Noninvasive implants have a
smooth interface with the surrounding tissue. By convention, a specimen
that contains only implant with no surrounding normal tissue is assumed
to be a noninvasive implant. Noninvasive implants are subclassified as
epithelial if there is an epithelial proliferation with no stromal response,
and desmoplastic if there is a stromal response that distorts or compresses
the epithelial cells (e-Fig. 31.22). In contrast, invasive implants irregularly
infiltrate and efface the underlying tissue (e-Fig. 31.23); some groups have
proposed that implants with epithelium exhibiting a micropapillary archi-
tecture or solid nests surrounded by a clefted space also represent a form
of invasive implant. The presence of invasive implants is associated with
a poorer prognosis and is generally treated with chemotherapy.
Two additional architectural patterns may be seen in serous border-
line tumors: micropapillary and cribriform. In the micropapillary pattern,
long thin filiform projections (five times as long as they are wide) project
directly from stromal cores without hierarchical branching (e-Fig. 31.24).
The cribriform pattern is often associated with the micropapillary pattern
and shows fusion of the tips of papillae with the formation of cribri-
form structures (e-Fig. 31.25). Both of these patterns are associated with
a poorer prognosis due to their association with invasive implants. Some
groups classify tumors with these patterns as a form of low-grade serous
carcinoma and use the term micropapillary or micro-cribriform serous
carcmoma.
c. Serous borderline tumors with microinvasion. Occasional serous borderline
tumors exhibit microinvasion where individual eosinophilic cells or small
nests of cells, often surrounded by a clefted space, invade the stroma (e-
Fig. 31.26). These microinvasive areas should involve < 3 mm in greatest
linear extent or <10 mm2 in area. The prognostic significance of microin-
vasion is not completely understood, but it may be a risk factor for disease
progression (Am] Surg Pathol. 2006;30:1209).
d. Serous adenocarcinoma most commonly presents as bilateral masses with
widespread peritoneal metastases. Grossly, the tumors are large and fri-
able with multiloculated cysts and polypoid growths. Microscopically, the
tumors may show a wide variety of architectural patterns including papil-
lary, solid, and nested with slit-like spaces. Nuclear atypia is variable but
often marked. By definition, serous carcinomas demonstrate destructive
stromal invasion (e-Fig. 31.27). Psammoma bodies may be present.
Serous carcinomas should be graded into high grade and low grade.
Low-grade tumors (< 12 mitoses/1 0 high power fields and low-grade cyto-
logic atypia) (e-Fig. 31.28) are frequently resistant to chemotherapy and
are more appropriately treated surgically.
A rare form of serous adenocarcinoma known as psammocarcinoma
is characterized by invasive stromal growth, abundant psammoma bodies
in at least 75% of the papillae, and no areas of solid growth > 15 cells
across (Gynecol Pathol. 1990;9:110). This tumor generally has a good
prognosis.
2. Mucinous tumors are the second most common type of surface epithelial
tumor. The neoplastic epithelium resembles intestinal (goblet cells with intra-
cytoplasmic mucin), gastric (foveolar), or endocervical (apical mucin with
no cilia) epithelium. Eighty percent of these tumors are benign, 10% are
borderline, and 10% are malignant. Because mucinous neoplasms are quite
heterogeneous and a malignant component may be very focal, it is imperative
that all mucinous neoplasms be well sampled (one to two sections per em).
a. Mucinous cystadenomas are usually unilateral, multiloculated, and large
(up to 50 em). They have a smooth external surface and are filled with thick
viscous secretions. Microscopically, the cysts are lined by a single layer of
tall columnar cells with bland basal nuclei, most often of intestinal type
(e-Fig. 31.29). If there is a prominent stromal component, the lesion is
termed a mucinous cystadenofibroma.
b. Borderline mucinous tumors (mucinous tumors of low malignant potential)
also present as unilateral, multiloculated masses. They are most common
in perimenopausal women. Grossly, these tumors have thick cyst walls
sometimes with papillary excrescences. Microscopically, mucinous bor-
derline tumors have a stratified, tufted intestinal-type epithelial lining with
mild to moderate nuclear atypia (e-Fig. 31.30). By definition, borderline
mucinous tumors do not demonstrate destructive stromal invasion.
Borderline tumors with endocervical-type epithelium (15% of muci-
nous borderline tumors) have the hierarchical branching pattern of
serous borderline tumors but the papilla are lined by mucinous epithe-
lium; they are often associated with acute inflammation and frequently
with endometriosis and bilaterality (e-Fig. 31.31). Some borderline
tumors show a mixture of endocervical-type mucinous epithelium, serous
epithelium, and even endometrioid epithelium; these tumors also have the
hierarchical branching architecture of serous borderline tumor and are
also associated with endometriosis and bilaterality.
c. Mucinous borderline tumors with intraepithelial carcinoma resemble border-
line tumors but contain focal areas that demonstrate increased cytologic
atypia with marked pleomorphism and prominent nucleoli (e-Fig. 31.32).
No destructive stromal invasion is present, and the intraepithelial carci-
noma does not appear to portend a worse prognosis (Ann Surg Oncol.
2010;18:40).
d. Mucinous borderline tumors with microinvasion. These are mucinous bor-
derline tumors with small foci of invasion either as small nests or as indi-
vidual cells, often with more eosinophilic cytoplasm. These foci should not
exceed 3 mm in greatest linear extent or 10 mm2 in area (e-Fig. 31.33).
e. Mucinous adenocarcinoma also presents in perimenopausal women as uni-
lateral, multiloculated cystic masses. Frank stromal invasion is present
microscopically (e-Fig. 31.34), but mucinous carcinomas frequently
demonstrate areas of benign and borderline mucinous epithelium as well.
Two patterns of invasion are recognized. The expansile pattern shows
crowded glands, little stroma, and, sometimes a cribriform architecture.
The destructive pattern shows single glands or individual cells invading
>3 mm in two linear dimensions or >10 mm2 in area.
Approximately 5% of women with a mucinous ovarian tumor will
present with pseudomyxoma peritonei, a condition in which pools of
mucin, with or without associated neoplastic epithelium, fill the peri-
toneal cavity. Although controversial in the past, there is now general
consensus that most mucinous tumors in the ovary associated with pseu-
domyxoma peritonei are metastatic in origin. The appendix is the most
common primary site (Am ] Surg Pathol. 1995;19:1390). Microscop-
ically, the ovary shows pools of mucin dissecting through the stroma
(pseudomyxoma ovarii) with or without associated mucinous epithelium
(e-Fig. 31.35).
When trying to distinguish a primary from a metastatic ovarian muci-
nous adenocarcinoma, both histologic and immunohistochemical features
are helpful. Primary tumors exhibit benign, borderline, and malignant
epithelium whereas metastatic tumors are more uniformly malignant.
Features seen more commonly in metastatic disease include bilateral-
ity, concomitant extraovarian disease, involvement of the external sur-
face of the ovary, pseudomyxoma ovarii, and extensive lymphovascu-
lar space invasion. Metastatic adenocarcinoma from the colorectum will
show abundant luminal karyorrhectic debris (dirty necrosis), a garland
pattern of glands lining cystic spaces, an abrupt transition between viable
and necrotic epithelium, and immunopositivity for C:I<20 and CEA but
immunonegativity for CK7 and CA125 (e-Fig. 31.36).
3. Endometrioid tumors. The majority of endometrioid tumors of the ovary are
carcinomas. From 10% to 20% are associated with endometriosis, and about
15% have a concomitant endometrioid tumor of the endometrium. Squa-
mous differentiation may be seen in all types of endometrioid tumors.
a. Benign endometrioid tumors are adenofibromas/cystadenofibromas, with
organized endometrial glands arranged in a fibrous stroma. These are quite
rare and can be distinguished from endometriosis by a lack of associated
endometrial-type stroma and hemosiderin-laden macrophages.
b. Borderline endometrioid tumors. Endometrioid tumors that show cytologic
atypia and areas of confluent epithelial proliferation without stromal sup-
port up to 5 mm in maximal dimension are borderline endometrioid
tumors, although there is some controversy as to diagnostic criteria and
IDf I SECTION VII· REPRODUCTIVE TRACl
'' 1.I'! I JJIII

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and cystic on gross examination. Microscopically, they are composed of

papillary cores lined by stratified, cytologically atypical epithelium. By
definition, no benign or borderline Brenner tumor component is present.
6. Carcinosarcoma. Also known as malignant mixed Mi.illerian tumor, carci-
nosarcoma occasionally presents as a primary tumor of the ovary, although
the tumor much more commonly arises in the endometrium. The tumor usu-
ally occurs in postmenopausal women and has a poor prognosis. As with its
endometrial counterparts, ovarian carcinosarcoma contains both malignant
epithelial and malignant mesenchymal elements. Numerous genetic studies
have demonstrated that both elements are derived from the same precursor,
proving that the neoplasm does not represent a collision tumor but rather a
poorly differentiated carcinoma with metaplastic sarcomatous elements. The
epithelial component is most often a high-grade serous carcinoma but may be
endometrioid, mucinous, clear cell, or even squamous carcinoma. Themes-
enchymal component may be homologous to the female genital tract (e.g.,
smooth muscle, endometrial stroma) or may be composed of heterologous
elements not normally found in the female genital tract {e.g., cartilage, fat)
(e-Fig. 31.43).
B. Sex cord-stromal tumors. These tumors represent approximately 8% of ovarian
tumors and comprise the majority of the hormonally active ovarian neoplasms.
1. Granulosa cell tumors
a. Adult granulosa cell tumors {AGCTs) are low-grade neoplasms that occur
most commonly in postmenopausal women but can occur at any age.
AGCTs are the most common ovarian tumors with estrogenic manifesta-
tions {e.g., endometrial hyperplasia or carcinoma). Hormonal manifesta-
tions may alert the clinician to the presence of a granulosa cell tumor, but
patients more commonly present with an adnexal mass. In about 10%
of cases, patients present with hemoperitoneum from tumor rupture. The
tumors are usually large (> 10 em) and unilateral. The cut surface is soft
and yellow-tan with cysts and hemorrhage {e-Fig. 31.44).
These tumors exhibit a variety of histologic patterns including diffuse
{e-Fig. 31.45), trabecular (e-Fig. 31.46), microfollicular, macrofollicular
{e-Fig. 31.47), or gyriform {e-Fig. 31.48), and often more than one pattern
is found within the same tumor. No matter the architecture, the cytology
is usually bland with oval nuclei, longitudinal nuclear grooves, and a
low mitotic rate {e-Fig. 31.49). The microfollicular and diffuse variants
often contain characteristic Cali-Exner bodies consisting of a small col-
lection of eosinophilic material lined by palisaded granulosa cells {e-Fig.
31.50). AGCTs usually exhibit minimal cytologic atypia although areas
of "bizarre" nuclei may be seen {e-Fig. 31.51).
AGCTs are usually confined to the ovary and spread outside the ovary
is a poor prognostic factot: Tumors with high stage, large size, nuclear
atypia, high mitotic activity, and a sarcomatoid pattern have a poorer
prognosis, although tumors with none of these factors may recur. AGCTs
are unusual in that they may recur decades after diagnosis.
b. Juvenile granulosa cell tumors (JGCTs) occur in children and young adults,
typically under the age of 20. They usually present with a palpable mass
and symptoms of hyperestrogenism such as breast development in children
or menstrual irregularities in adolescents. The gross findings are similar
to those of adult granulosa cell tumors with yellow-tan solid areas and
interspersed blood-filled cysts.
Microscopically JGCTs are characterized by solid sheets of cells mixed
with small follicle-like spaces with basophilic or eosinophilic secretions
lined by more mature-appearing granulosa cells. Luteinization is promi-
nent (e-Fig. 31.52). JGCTs exhibit more cytologic atypia and a higher
mitotic rate than adult granulosa cell tumors. The nuclei do not have
grooves (e-Fig. 31.53).
JGCTs are usually confined to the ovary and high stage is a poor
prognostic factor. Unlike their adult counterparts, patients with JGCT
who recur usually do so within 2 years of diagnosis. Cytologic atypia is
not a poor prognostic factor in JGCTs.
2. Fibroma-thecoma
a. Fibromas represent the most common of the sex cord-stromal tumors.
They typically occur in perimenopausal women and are not hormonally
active. Grossly, fibromas are unilateral, solid, and lobulated with a firm,
white-gray cut surface. Cystic degeneration sometimes occurs. Histolog-
ically, they are characterized by interlacing bundles and storiform areas
of spindle cells that show no atypia and few mitoses (e-Fig. 31.54). The
tumor cells stain diffusely positive for vimentin and are usually negative
for inhibin.
Fibromas may be associated with two syndromes. Meig syndrome
(fibroma, ascites, and pleural effusion) and Gorlin syndrome (basal cell
nevus syndrome). In Gorlin syndrome, fibromas occur in younger women
or even children, are often multiple or bilateral, and are calcified.
b. Thecomas are unilateral solid tumors found most often in postmenopausal
women. Hyperestrogenic symptoms are present in 50% to 80% of cases.
Grossly, these tumors have a lobulated, yellow-tan cut surface. Histo-
logically, thecomas comprise round to oval, lipid-laden theca cells in a
fibromatous stroma with little atypia or mitotic activity (e-Fig. 31.55).
Immunohistochemically, the tumor is positive for inhibin. Oil Red 0 fat
stains (which require fresh tissue) highlight the intracellular lipid. When
clusters of lutein cells (eosinophilic cells with large round nuclei) are
present, the tumor is termed a luteinized thecoma (e-Fig. 31.56); this vari-
ant is more common in younger women.
3. Sertoli and Sertoli-Leydig Cell Tumors
a. Sertoli cell tumors are rare, low-grade, nonfunctioning tumors that occur in
women of child-bearing age. Grossly, they are yellow-tan, solid, lobulated
tumors. Microscopically, they are composed of closely packed tubules
separated by fibrous stroma. The tubules are lined by cuboidal to colum-
nar cells with abundant pale eosinophilic cytoplasm with little atypia or
mitotic activity.
b. Sertoli-Leydig cell tumors (SLCT) are rare, unilateral neoplasms that occur
in young women and, in 30% of cases, secrete androgenic hormones.
Grossly, these tumors average 10 em in diameter, are yellow-orange to
red-brown, and frequently have a nodular appearance with a central scar
(e-Fig. 31.57). Well-differentiated SLCTs contain tubules (similar to those
seen in Sertoli cell tumors) and interspersed clusters of Leydig cells that
have abundant eosinophilic cytoplasm (e-Fig. 31.58). SLCT of intermedi-
ate differentiation contain solid cords of Sertoli cells (e-Fig. 31.59), while
Leydig cells may be more difficult to find but are typically located at the
periphery of cellular nodules. The Leydig cells often show lipidization
with foamy rather than eosinophilic cytoplasm (e-Fig. 31.60). Poorly dif-
ferentiated SLCT show densely packed atypical spindled cells resembling
a sarcoma (e-Fig. 31.61).
Two morphologic variants of SLCT occur, usually in association with
tumors of intermediate differentiation or poorly differentiated tumors.
About 20% of SLCf harbor heterologous elements that take the form
of intestinal type mucinous glands (e-Fig. 31.62) or carcinoid tumor,
(which have a good prognosis) or rhabdomyoblastic or cartilaginous dif-
ferentiation (which have a poor prognosis). About 15% of SLCT contain
tubules and slit-like glandular structures, or micropapillary structures

with dense fibrovascular cores (e-Fig. 31.63); these tumors are referred to
as retiform variants. Retiform tumors are less likely to be androgen secret-
ing and more common in younger age groups (average age is 15 years).
4. Steroid cell tumors are uncommon neoplasms composed of large cells with
intracellular lipid that resemble Leydig cells or luteinized stromal cells.
a. Stromal luteomas are benign, hormonally active steroid cell tumors that
usually occur in postmenopausal women. Estrogenic manifestations are
seen in 60% of cases; androgenic in 10% of cases. These well-circum-
scribed tumors grow within the ovarian parenchyma and have a yellow-
brown cut surface. Microscopically, polygonal cells with eosinophilic cyto-
plasm grow in sheets, nests, and cords (e-Fig. 31.64 ). Degenerative changes
may cause slit-like spaces within the tumor. Crystals of Reinke (slender,
eosinophilic, rod-shaped crystals) are absent (e-Fig. 31.17). Stromal hyper-
plasia may occur in the ipsilateral or contralateral ovary.
b. Leydig cell tumors are also benign steroid cell neoplasms found in post-
menopausal women, although these tumors are more often androgenic
or nonfunctioning. The majority of these tumors arise within the ovarian
hilus although they also can occur in the ovarian stroma. The cut surface
is yellow-brown with areas of hemorrhage. Microscopically, large polygo-
nal cells with foamy or granular eosinophilic cytoplasm and round nuclei
grow in cellular clusters separated by pink acellular areas (e-Fig. 31.65).
In order to classify a tumor as a Leydig cell tumor Reinke's crystals must
be identified.
c. Hilus cell tumors are benign tumors composed of the same cell population
but that originate in the hilus of the ovary. Hilus cell tumors are usually
associated with androgenic symptoms.
d. Steroid cell tumor, not otherwise specified (NOS) is a steroid cell tumor that
does not meet the criteria for any of the types mentioned earlier. These are
the most common of the steroid cell tumors and may occur at any age.
They are often hormonally active (50% androgenic, 10% estrogenic), and
their gross morphology is similar to the other types of steroid cell tumor.
Histologically, steroid cell tumor, NOS is composed of large cells with
abundant granular cytoplasm separated by a vascular stroma. Reinke's
crystals are absent. A poorer prognosis is seen in tumors with necrosis,
hemorrhage, an increased mitotic rate, or size > 7 em.
5. Other sex cord-stromal tumors
a. Sclerosing stromal tumors are rare benign neoplasms seen most often in
girls and women <30 years of age. Grossly the tumor is firm to rubbery
and white with areas of cystic degeneration. Histologically, cellular areas,
consisting of both spindled cells and round cells with vacuolated cyto-
plasm, alternate with edematous and collagenized areas giving the tumor
a pseudolobular appearance (e-Fig. 31.66).
b. Sex cord tumors with annular tubules (SCTAT) occur in women of child-
bearing age. In some women, the tumor occurs as a component of Peutz-
Jeghers syndrome, in which case the SCTATs are small and incidental.
Those unassociated with Peutz-Jeghers syndrome form a large, solid, yel-
low mass. Histologically, the tumor is composed of well-circumscribed,
ring-shaped tubules that contain central hyalinized material. The tubules
are lined by cells with pale cytoplasm oriented toward the center of the
tubule, with peripheral elongated nuclei (e-Fig. 31.67).
C. Germ cell tumors
1. Teratomas
a. Mature teratomas are the most common ovarian germ cell tumor. They
occur most often in adult women of reproductive age but may occur at
any age. Grossly, they are usually cystic with a single solid nodule that may
contain fat, teeth, bone, and many other tissue types. The cysts usually con-
tain hair, soft yellow sebaceous debris. Microscopically, mature tissue from
all three germ layers (ectoderm-skin or central nervous system elements;
mesoderm-smooth muscle, teeth, bone; endoderm-respiratory epithe-
lium, GI epithelium, thyroid) may be present (e-Fig. 31.68). The term
dermoid cyst is commonly used to refer to mature cystic teratomas lined
by squamous epithelium that containing skin appendages (e-Fig. 31.69).
Mature cystic teratomas should be thoroughly sampled (one section per
em) in order to exclude an immature component, and to exclude malignant
transformation of one of the mature components (a rare occurrence).
b. Immature teratomas are rapidly growing malignant tumors that occur in
children and young adults. They are unilateral and typically have solid
and cystic components. The solid areas are generally more extensive than
in a mature teratoma. Microscopically, they contain immature or primi-
tive tissue (derived from any or all three germ cell layers) that is usually
mixed with areas of mature tissue. The most common immature element is
neuroectodermal and consists of rosettes, masses, or tubules of primitive
neural cells (e-Fig. 31.70).
Immature teratomas are graded based on the relative amount of imma-
ture tissue present (lnt] Gynecol Pathol. 1994; 13:283 ). Tumors with more
than one low-power field of immature neuroepithelium on any given slide
are considered high grade and require adjuvant chemotherapy.
c. Monodermal teratomas are teratomas in which all the tissue is derived from
one germ cell layer. Struma ovarii is the most common monodermal ter-
atoma and consists of mature thyroid tissue, including follicles and colloid
(e-Fig. 31.71). Secondary changes such as hyperplasia, adenoma, and even
carcinoma may be seen.
2. Carcinoid tumors of the ovary usually arise in a mature cystic teratoma. They
most commonly have an insular or trabecular pattern identical to that seen
in the gastrointestinal tract. The cells are small and uniform with round
nuclei with stippled chromatin. Goblet cell carcinoids are very uncommon.
Metastasis from a primary carcinoid tumor outside the ovary must always be
excluded; metastatic carcinoid tumors are more likely to be bilateral, larger,
unassociated with a teratoma, and more commonly associated with carcinoid
syndrome.
3. Dysgerminoma is the most common malignant germ cell tumor. It occurs as
pure dysgerminoma or as a component of mixed germ cell tumor. Dysger-
minoma develops most commonly in adolescents and young women and is
frequent in patients with ovarian dysgenesis. Patients with dysgerminomas
frequently have elevated LDH levels and occasionally mildly elevated hCG
levels. Pure dysgerminomas have an excellent prognosis when treated by
current therapeutic regimens.
Grossly, dysgerminomas are large and solid with a smooth external sur-
face and a lobulated gray-tan cut surface. The tumor should be thoroughly
sampled (one section per em) for microscopic examination to exclude other
germ cell tumor types, and special attention should be directed to hemor-
rhagic and cystic areas.
Dysgerminomas are analogous to testicular seminomas and have an iden-
tical histologic appearance. They are composed of nests and sheets of uni-
form large round cells with abundant clear cytoplasm, large nuclei, and
prominent nucleoli. Tumor cells are separated by a lymphocyte-rich fibrous
stroma (e-Fig. 31.72). Often a histiocytic or granulomatous infiltrate is
present, and multinucleated syncytiotrophoblastic cells may also be identi-
fied (sometimes the former component can obscure the dysgerminoma cells).
Dysgerminomas are immunopositive for placental alkaline phosphatase

(PLAP), c-kit (CD117), SALL4, and OCT 3/4.
4. Yolk sac tumors, also known as endodermal sinus tumors, occur in young
women, usually with an associated elevated serum alpha fetoprotein (AFP)
level. Yolk sac tumors grow rapidly and have often spread outside the ovary
at the time of diagnosis. They are often a component of mixed germ cell
tumors. Grossly, yolk sac tumors are unilateral and large, with a smooth
external surface and a solid and cystic yellow to tan cut surface. Hemorrhage
and necrosis are often present.
Many different histologic patterns occur, including microcystic, endoder-
mal sinus, macrocystic, solid, polyvesicular-vitelline, papillary, hepatoid, and
glandular. The microcystic pattern is the most common variant and is com-
posed of small cystic spaces lined by cuboidal to columnar cells with clear
cytoplasm and large hyperchromatic nuclei (e-Fig. 31.73). The endodermal
sinus pattern is the second most common pattern. This pattern features char-
acteristic Schiller-Duvall bodies (rounded fibrovascular papillae containing
a single central vessel and lined by columnar tumor cells) (e-Fig. 31.74).
The polyvesicular-vitelline pattern is composed of abundant cystic structures
lined by tumor cells that are embedded in a dense cellular stroma. Yolk
sac tumors are usually immunopositive for expression of keratins, alpha-1-
antitrypsin, glypican 3, and AFP; immunostaining shows a lack of expression
ofEMA.
5. Embryonal carcinoma is a rare neoplasm in the ovary and is often a component
of mixed germ cell tumors. Half of cases occur in prepubertal girls, and
half have elevated beta-HCG levels. Grossly, these tumors are large (""17
em) and solid with areas of hemorrhage and necrosis. Morphologically, the
tumor is identical to embryonal carcinoma of the testis and is composed of
large anaplastic cells with pale eosinophilic vacuolated cytoplasm that grow
in sheets and nests. The nuclei are hyperchromatic with prominent nucleoli
(e-Fig. 31.75). Atypical mitotic figures are common. Embryonal carcinoma
is immunopositive for CD30, cytokeratin, PLAP, SALL4, and OCT 3/4.
6. Polyembryoma is a very rare, highly malignant neoplasm that usually is a
component of a mixed germ cell tumor. Grossly, it presents as a unilateral
solid mass with areas of hemorrhage and necrosis. Microscopically, the tumor
is composed of embryoid bodies (embryonic disks lined by endoderm on one
side, ectoderm on the opposite side, and associated yolk sac and amniotic
cavities).
7. Choriocarcinoma of the ovary is rare in pure form. It is most often seen
as a component of a mixed germ cell tumor, or as a metastasis from gesta-
tional trophoblastic disease. Primary choriocarcinoma presents in children
and adolescents where elevated hCG levels are invariably present. Grossly,
the tumor mass is hemorrhagic, soft, and tan. Microscopically, both cytotro-
phoblast and syncytiotrophoblast are present. Cytotrophoblasts have cen-
trally located hyperchromatic nuclei, prominent nucleoli, clear cytoplasm,
and well-defined cytoplasmic borders. Syncytiotrophoblast are multinucle-
ated and are immunopositive for hCG. While syncytiotrophoblast may be
seen as a component of other germ cell tumors, cytotrophoblast is seen only
in choriocarcinoma.
8. Mixed germ cell tumors make up 10% of germ cell tumors. The most common
combination is dysgerminoma and yolk sac tumor, although any combination
may occur. The relative composition of the various histologic subtypes should
be included in the final report since it can impact therapy and prognosis.
D. Miscellaneous
1. Gonadoblastoma is a rare tumor that contains both germ cell and sex cord-
stromal components. Most patients have gonadal dysgenesis, and over 90%
have a Y chromosome. Approximately half of all cases harbor a malignant

germ cell component, most often dysgerminoma. Gonadoblastoma is benign
unless a malignant germ cell component is present.
Grossly, the tumor is usually small with a yellow to gray cut surface and
areas of calcification. Histologically, the tumor consists of admixed primitive
germ cells and sex cord-stromal derivatives (which resemble immature granu-
losa cells and Sertoli cells) surrounded by abundant basement membrane-like
material, often with calcification.
2. Hypercalcemic small cell carcinoma is a highly malignant tumor with a
poor prognosis that presents in young women. Approximately two-thirds of
patients manifest hypercalcemia. This tumor is generally large and unilateral
with a soft, white-tan, lobulated cut surface that shows areas of hemorrhage
and necrosis. Histologic examination demonstrates sheets of small cells with
scant cytoplasm admixed with follicle-like spaces filled with eosinophilic fluid
(e-Fig. 31.76). The tumor nuclei are round with coarse chromatin and promi-
nent nucleoli (e-Fig. 31.77). Some tumor cells have globular hyaline inclu-
sions producing a vague rhabdoid morphology. The tumor cells are usually
immunopositive for CK, EMA, Wf-1, calretinin, CDlO, and p53 and less
commonly positive for vimentin, NSE, and chromogranin.
3. Primary hematopoietic malignancies of the ovary are extremely rare; how-
ever, secondary ovarian involvement may be seen in up to one-half of all
lymphomas. The tumors are generally bilateral with a fleshy cut surface.
Microscopically, they resemble their nodal or marrow counterparts. The
most common lymphoma with secondary involvement of the ovary is dif-
fuse large B-celllymphoma containing sheets of large noncohesive cells that
have irregular nuclear contours and increased mitotic activity. Immunohis-
tochemical stains such as CD45 (leukocyte common antigen), as well as B
and T cell markers, may be used to demonstrate a hematopoietic origin in
difficult cases.
4. Primary ovarian tumors of mesenchymal origin. A wide variety of these tumors
have been described, although they are quite rare.
a. Vascular tumors, including hemangioma, lymphangioma, and low-grade
angiosarcoma, have all been reported as primary ovarian neoplasms. The
morphologic features are identical to tumors occurring primarily in the
soft tissue.
b. Tumors of striated muscle origin, such as rhabdomyoma and rhab-
domyosarcoma, also rarely occur in the ovary. Primary osteosarcomas and
chondrosarcomas have also been reported. For these neoplasms, generous
sampling of the tumor is required to exclude the presence of an epithe-
lial component which would indicate a carcinosarcoma of heterologous
type.
c. Neural tumors, including neurofibromas, schwannomas, and ganglioneu-
romas, have been reported in the ovary and may present in association
with neurofibromatosis. The morphologic features are identical to tumors
occurring in the peripheral nervous system.
d. Primary ovarian myxomas are rare unilateral tumors composed of stellate
and spindled cells in an abundant myxoid stroma. These tumors must be
distinguished from pseudomyxoma ovarii, which is characterized by pools
of mucin usually with associated strips of mucin-secreting epithelium.
5. Metastases. About 8% of ovarian tumors represent metastases, of which
most are derived from the gastrointestinal tract (especially the large intestine,
stomach, and appendix), breast, uterine corpus, and cervix. In young girls,
ovarian metastases may be from neuroblastoma, rhabdomyosarcoma, Ewing
sarcoma/peripheral neuroectodermal tumor, and malignant rhabdoid tumor
of kidney.
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V. STAGING OF OVARIAN MALIGNANCIES

A. Pathologic staging. Ovarian cancer is staged surgically. The staging procedure
includes bilateral salpingo-oophorectomy, hysterectomy, and omentectomy;
biopsies of multiple pelvic and abdominal peritoneal surfaces; and regional
lymph node dissections (as noted earlier). Cytologic examination of peritoneal
washings is also performed. The 2010 AJCCiillCOFIGO staging classification
B. Items to include in the pathology report. Because tumor histologic subtype and
grade have prognostic and therapeutic significance, these items must always
be included in the final pathology report. For stage I tumors (confined to the
ovary), it is important to note whether tumor involves the surface of the ovary
or whether the ovary was ruptured (preoperatively or during the procedure),
assessments best made at the time of frozen section.
Fallopian Tube
Mitra Mehrad, John D. Pfeifer,
I. NORMAL ANATOMY. The fallopian tubes are formed from the Miillerian
(paramesonephric duct) system and lie within the broad ligament between the ovary
and the uterus. They conduct eggs from the surface of the ovary to the uterine cavity
and are the usual site of fertilization. Each fallopian tube is shaped like an elon-
gated funnel and is divided into four parts from lateral to medial: infundibulum,
ampulla, isthmus, interstitium; the infundibulum contains the finger-like fimbriae
distally.
The fallopian tube mucosa is branched and folded into plicae. The mucosa is
lined by a nonstratified epithelium composed of three cell types: ciliated, secretory,
and intercalated cells. The most common are the ciliated cells, followed by secretory
cells, which together comprise over 90% of the cell population. The intercalated
cells are seen as elongated nuclei sporadically present between the ciliated cells. The
wall of the fallopian tube contains smooth muscle to aid in moving the fertilized
egg into the uterus. The serosa contains abundant blood vessels and is continuous
with the broad ligament.
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION. Fal-
lopian tubes are usually received as a portion of a total abdominal hysterectomy-
bilateral salpingo-oophorectomy specimen. Short cross sections of fallopian tube
are received after tubal ligation procedures. Ectopic pregnancy specimens also usu-
ally contain a portion of fallopian tube. Rarely, specimens are received for primary
fallopian tube tumors.
At the grossing station, the length and diameter of the fallopian tube should be
documented, as well as the presence or absence of a fimbriated end, or evidence
of prior ligation. The serosal surface should also be assessed for the presence of
adhesions, cysts, exudates, rupture, or metastatic tumor. The tube is then serially
sectioned.
A. Benign specimens. Three sections are submitted from a normal fallopian tube,
specifically from the fimbriated end, ampulla, and isthmus end. Additional sec-
tions of any gross lesions are also submitted. Complete cross sections must be
identified from a tubal ligation specimen.
When examining an ectopic pregnancy specimen, an embryo and/or pla-
cental villi will often be grossly evident. Hemorrhagic areas, including blood
dot, along with obvious villous or embryonic tissue, should be submitted for
histologic examination.
B. Neoplastic specimens. Primary tubal carcinoma specimens will show a dilated
lumen filled with a papillary or solid tumor. An ovarian tumor secondarily
involving the fallopian tube is more common than a primary fallopian tube
tumm; and careful sectioning can help distinguish the two. Grossly, papillary and
solid areas should be sampled thoroughly (at least one section per centimeter of
tumor), along with uninvolved areas. If possible, a section showing the tumor's
relationship to the ovary should be submitted.
C. Prophylactic excision specimens. Fallopian tube specimens received as part of
a prophylactic hysterectomy-oophorectomy from patients with hereditary can-
cer syndromes (e.g., BRCA1 syndrome) should be, together with the ovaries,
entirely submitted for histologic examination. For the fallopian tube, the goal
is to ensure sectioning and extensive examination of the fimbria (so-called SEE-
FIM protocol), since the majority of early serous tumors occur in this area.
51 1
Figure 32.1 Approach to sectioning fallopian tube specimens received as part of a prophylactic
hysterectomy-oophorectomy from patients with a hereditary cancer syndrome. Each tube and
ovary should be entirely submitted for histologic examination; note that the fimbriated end of
the tube should be transected, opened longitudinally, serially sectioned longitudinally, and then
entirely submitted for microscopic examination.
The protocol (Am] Surg Pathol. 2006;30:230) specifies that the entire tube is
fixed for at least 4 hours to minimize loss of epithelium during manipulation,
the distal2 em of the fimbriated end is transected, the fimbria! mucosa is secw
tioned longitudinally into four pieces, the remainder of the tube is sectioned
transversely every 2 to 3 mm, and that all the sections are submitted in toto
(Fig. 32.1).
A. lnftammabJry and nonneoplastic lesions of tha fallopian tuba
1. Cystic lesions. Embryologic remnants may be found in the fallopian tube wall,
usually as an incidental finding (e-Fig. 32.1).• Paratubal cysts of Miillerian
origin (ciliated lining) or Wolffian origin (stratified transitional lining) may
be encountered; if the lining is atrophic secondary to compression, the two
can be difficult to differentiate microscopically. Pedunculated paratubal cysts
with a Miillerian epithelium present along the fimbriae are termed hydatid
of Morgagni.
2. lnftammatory lesions
a. Acute salpina;itis usually presents in youngw to middlewaged women, is
generally an ascending infection initiated by Chlamydia or Neisseria
*All ewfigures are available online via the Solution Site Image Bank.
Chapter 32 • Fallopian Tube I 5 13
gonorrhoeae. and is often followed by polymicrobial infection (pelvic

inflammatory disease). The damage to the tube that results from acute
salpingitis may lead to infertility and/or ectopic pregnancy. Grossly, the
tubal lumen may be distended by pus, blood, or secretions. Histologic
sections show marked acute inflammation, with congestion and edema
in the plicae and the tubal wall which often severely distorts the normal
tubal architecture (e-Fig. 32.2).
b. Chronic salpingitis is usually due to resolving acute salpingitis. Grossly,
the tube is often enlarged, fibrotic, distorted, and adherent to the ovary
or adjacent structures. Microscopically, a lymphoplasmacytic infiltrate is
seen in the plicae. Fusion of the tubal plicae after resolution of acute
salpingitis may lead to formation of follicle-like spaces, a histologic pat-
tern known as salpingitis follicularis. Hydrosalpinx may be seen in end-
stage chronic salpingitis, microscopically characterized by a dramatically
thinned wall with few plicae and a lumen filled with clear fluid. Numerous
adhesions are typically present on the serosal surface {e-Fig. 32.3).
c. Granulomatous salpingitis may be caused by tuberculosis, fungal infection,
Crohn disease, or sarcoidosis.
d. Salpingitis isthmica nodosum typically presents in young women. It is asso-
ciated with ectopic pregnancy and infertility, and has an unclear patho-
genesis. Grossly, it presents as 1 to 2 em diameter nodules in the wall of the
fallopian tube isthmus. It is bilateral in 85% of cases. The lesions consist
of outpouchings of tubal epithelium surrounded by a thickened wall of
smooth muscle (e-Fig. 32.4).
3. Other nonneoplastic lesions of the fallopian tube
a. The fallopian tube is a frequent site of involvement of endometriosis. pri-
marily in women of reproductive age, which may be associated with infer-
tility. Grossly, tubal endometriosis consists of dark brown serosal nodules.
The microscopic findings include endometrial glands surrounded by a cuff
of endometrial stroma with associated hemosiderin-laden macrophages
and chronic inflammatory cells (e-Fig. 32.5).
b. Ectopic pregnancy affects 1% to 2% of all conceptions, and the fallop-
ian tube is the most common site; the most commonly involved region
of the tube is the ampulla. Risk factors include prior ectopic pregnancy,
salpingitis, congenital tubal anomalies, salpingitis isthmica nodosum,
and endometriosis. Patients may present with tubal rupture and shock.
Grossly, the fallopian tube is dilated and hemorrhagic with identifiable
chorionic villi, with or without an identifiable embryo. Histologic sec-
tions should show chorionic villi or trophoblast in the tubal mucosa or
wall (e-Fig. 32.6).
B. Neoplastic lesions of the fallopian tube. The World Health Organization
(WHO) histologic classification of tumors of the fallopian tube is presented in
Table 32.1.
1. Benign tumors
a. Adenomatoid tumors are the most common benign tumor of the fallopian
tube. They are usually incidental, unilateral, and present grossly as a well-
circumscribed, white-tan lesion in the tubal wall. They are derived from
mesothelium, and the usual histologic pattern shows slit-like or glandular
spaces lined by a single layer of flattened cuboidal cells (e-Fig. 32.7). The
cells are immunopositive for expression of cytokeratin, calretinin, and
vimentin but are immunonegative for expression of factor VIII-related
antigen and CD31, a profile that indicates a mesothelial rather than a
vascular origin.
b. Other benign tumors include epithelial papillomas, adenomas, cystadeno-
mas, adenofibromas, and leiomyomas. Epithelial papillomas are usually
I
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incidental and show a branched pattern with a central fibrovascular core

lined by a single layer of eosinophilic cells. Adenomas, cystadenomas,
adenofibromas, and leiomyomas all resemble their ovarian and uterine
counterparts.
2. Malignant tumors
a. Primary fallopian tube carcinoma is quite rare. Carcinomas typically occur
in elderly women and are usually widespread at the time of diagnosis with
a correspondingly poor prognosis. Serum CA125 levels may be elevated.
Grossly, the fallopian tube is distended by a papillary or solid tumor.
Serous adenocarcinoma, which microscopically is identical to ovarian
serous adenocarcinoma (e-Fig. 32.8), is the most common subtype of car-
cinoma. Other more uncommon subtypes, such as endometrioid or clear
cell carcinoma, resemble their counterparts in the ovary. The pathologic
staging of primary fallopian tube malignancies is presented in Table 32.2.
Carcinoma in situ, also called serous tubal intraepithelial carcinoma
(STIC), is characterized by cellular stratification, loss of polarity, high-
grade nuclear atypia, increased mitotic activity, and increased apopto-
sis and is distinguished from carcinoma by the lack of stromal invasion
(e-Fig. 32.9).
b. Other primary malignant tumors of the fallopian tube include leiomyosar-
coma, which may affect the fallopian tube or broad ligament, and carci-
nosarcoma (malignant mixed Mullerian tumor). Both tumors microscopi-
cally resemble their ovarian counterparts and are quite rare in the fallopian
tube.
c. Metastatic tumors are the most common category of malignancies involving
the fallopian tube. Consequently, metastasis from a primary tumor at
another site must always be excluded before making the diagnosis of a
primary fallopian tube malignancy. Other primary tumors of the female
reproductive tract, especially the ovary and endometrium, are the most
frequent sources of metastases. It has recently been proposed that small
serous carcinomas of the fallopian tube, particularly in the fimbria in
women with BRCA mutations, are the primary source for peritoneal, and
possibly ovarian, serous carcinomas (Am] Surg Pathol. 2007;31:161).
C. Reporting. The final report in any case of malignancy should include the histo-
logic type and grade of the malignancy; the presence or absence of a precursor
lesion (carcinoma in situ/STIC); tumor size, including depth and width; whether
the malignancy is unifocal or multifocal; and the presence or absence of lym-
phovascular space invasion. In addition, the report should explicitly include all
of the information required for assigning a stage, as well as other information
of clinical interest not required for staging.
Uterus (Corpus)
Jena Beth Hudson, John D. Pfeifer,
I. NORMAL ANATOMY. The uterus is a pear-shaped hollow organ with a normal weight
of between 40 and 80 gin adults. It is divided into the corpus, the lower uterine
segment, and the cervix. The uterine cavity is triangular, measuring on average
6 em in length. It is composed of the inner endometrial lining and the myometrium
or muscular wall, with a serosal covering which extends to the peritoneal reflec-
tion. The peritoneal reflection is shorter anteriorly than posteriorly and so can be
used for orienting hysterectomy specimens.
A. Endometrial biopsy and curettage specimens. The most common endometrial
tissue samplings examined in surgical pathology are endometrial biopsy and
curettage specimens, obtained from cervical dilation and curettage procedures.
Endometrial biopsy samples are obtained from a relatively limited office sam-
pling procedure in which no cervical dilation is required. The dimension (size
range of the largest tissue fragments, or the dimensions of the tissue in aggre-
gate) and/or volume of the specimen should be documented. The entire speci-
men should be submitted for microscopic examination and three H&E stained
levels prepared for microscopic examination.
B. Products of conception specimens are usually obtained by curettage (although
the tissue is often spontaneously passed). The dimension (size range of the
largest tissue fragments, or the dimensions of the tissue in aggregate) and/or
volume of the specimen should be documented. At least three cassettes should
be submitted, focused on any villous tissue that is grossly present, to optimize
microscopic identification chorionic villi both for confirmation of the presence
of an intrauterine pregnancy and to rule out a molar gestation. If the initial
three blocks do not contain villi, the remainder of the specimen should be
submitted; if villi are still not identified, the possibility of an ectopic pregnancy
exists, a result that should be immediately communicated to the clinician.
C. Hysterectomy specimens. The type of hysterectomy (abdominal or vaginal, with
or without salpingo-oophorectomy) should be determined and the size, weight,
and shape of the uterus recorded (the processing of radical hysterectomy speci-
mens, which differs substantially, is discussed in Chap. 34). The uterine serosa
should be carefully examined for any abnormalities, which should be sampled.
The uterus is next bivalved in the coronal plane to show the endometrial cav-
ity and endocervical canal, which are examined and measured. The maximum
thickness of the endometrium and myometrium should also be noted. Both
halves of the uterus are then serially sectioned parallel to the long axis of the
uterus.
1. For specimens excised for benign disease, sections of the anterior
cervix, posterior cervix, anterior endomyometrium, and posterior endomy-
ometrium are submitted. Additional sections of any identified lesions must
also be submitted.
2. For specimens excised for malignancies, contiguous sections of both ante-
rior and posterior endomyometrium, lower uterine segment, and cervix
should be submitted to assess for tumor involvement of the lower uter-
ine segment and cervix. In addition, at least one full thickness section of
the uterine wall containing tumor from both anterior and posterior uterus
(including the serosa from the deepest area of myometrial invasion by the
517
tumor) are submitted to enable calculation of the depth of invasion. Rep-

resentative sections from any other lesions must also be submitted.
Ill. ENDOMETRIUM
A. Dating
1. The endometrial mucosa is composed of glands and stroma. It is divided
into the functional (luminal) layer and the basal (inner) layer. The basal
cell layer acts as a reserve cell layer and is responsible for the regeneration
of the endometrium after menses. The stroma is composed of endometrial
stromal cells and blood vessels.
2. The menstrual cycle is divided into menstrual phase, proliferative phase,
and secretory phase. Menstrual endometrium, present for the first 4 days
of the 28-day cycle, is characterized by glandular (karyorrhectic debris in
glandular cells) and stromal (dense balls of collapsed stroma with surround-
ing neutrophils and reactive epithelium) breakdown, glandular secretory
exhaustion, and background inflammation (e-Figs. 33.1 and 33.2).*
Day 1 of menstrual bleeding is defined as day 1 of the cycle; the menstrual
phase lasts for 3 to 4 days. Proliferative phase begins on day 4 and in an
idealized situation lasts until day 14. Although it usually lasts about 11 days,
it may greatly vary. During the early proliferative phase, the endometrium
is thin and composed of straight, evenly spaced glands in a loose stroma
(e-Fig. 33.3). By day 8 to 10, stromal edema due to estrogen causes increased
endometrial thickening, and the glands become more coiled as the gland-
stroma growth rate increases (e-Fig. 33.4). Throughout the proliferative
phase, the epithelium lining the glands shows nuclear stratification, with a
high mitotic rate in both glands and stroma (e-Fig. 33.5).
The secretory phase begins with ovulation. In an idealized 28-day cycle,
secretory phase begins at day 14 and lasts 14 days, although it may range
from 11 to 18 days. Following an interval phasefrom day 14 to day 15 (dur-
ing which there are no dateable changes), the first dateable feature of early
secretory phase is the appearance of subnuclear vacuoles on day 16, which
appear as a clear zone between the basement membrane and the nucleus
pushing the nucleus toward the glandular lumen. On day 17 the epithe-
lium exhibits uniform subnuclear vacuoles, giving the appearance of "piano
keys" (e-Fig. 33.6). The vacuoles then move to the supranuclear position
(day 18) and eventually are secreted into the glandular lumen (e-Fig. 33.7).
Maximal stromal edema occurs during the mid-secretory phase, around day
22. At day 23, the stroma begins to condense, and the first signs of periarte-
riolar decidualization (where stromal cells acquire abundant, eosinophilic
cytoplasm under the influence of progesterone) become apparent (e-Fig.
33.8). On day 25, this decidualization extends beneath the surface epithe-
lium. Prominent glandular saw-toothing and maximal stromal decidualiza-
tion occur on days 26 to 27 (e-Fig. 33.9). Numerous granular lymphocytes,
marked stromal decidual change, and glandular breakdown are the features
of day 28 of late secretory phases.
B. Pregnancy
1. The earliest gestation-related changes occur following the implantation of
the blastocyst. These changes are characterized by decidualization of the
stroma with edema; the glands exhibit distension with increased secretion
and a serrated architecture (e-Fig. 33.10). By 4 to 8 weeks post implan-
tation, the endometrial epithelium often exhibits a physiologic response
known as the Arias-Stella reaction characterized by glands that have a
Chapter 33 • Uterus (Corpus) I 5 19
hypersecretory pattern and are lined by cells with enlarged, hyperchromatic

nuclei that often jut into the gland lumens (e-Fig. 33.11).
2. The placental implantation site, often seen in curettage specimens obtained
because of a missed abortion, is characterized by decidualized stroma infil-
trated by intermediate trophoblast. Intermediate trophoblast have hyper-
chromatic, angulated nuclei and amphophilic cytoplasm and are often
multinucleated (e-Fig. 33.12). Intermediate trophoblast also normally infil-
trates maternal spiral arteries causing fibrinoid deposition in the vessel wall
which serves to dilate the vessels and increase blood flow to the placenta
(e-Fig. 33.13 ). Intermediate trophoblast also normally infiltrates myometrium
which can occasionally be present in curettage specimens (e-Fig. 33.14).
3. Placental site nodules are incidental, usually microscopic findings that are
characterized by small foci of hyalinized material with entrapped intermedi-
ate trophoblast cells, often with vacuolated cytoplasm (e-Fig. 33.15). They
are thought to arise from the chorionic type intermediate trophoblast of
the fetal membranes which is also vacuolated. Placental site nodules are
occasionally encountered in endometrial biopsies and curettage specimens
but may also be seen in cervical specimens.
4. Abnormalities of implantation. Placenta accreta occurs when a layer of
decidua is not present between the placental villi and the myometrium at
the implantation site (e-Fig. 33.16); fibrin and intermediate trophoblast rnay
be present between villi and myometrium in accreta. Cytokeratin, which
will be positive in trophoblast and negative in decidua, can be used if the
distinction is difficult on H&E. Placenta increta is present when villi invade
into the myometrium, and transmural extension of villi with perforation is
termed placenta percreta.
Risk factors for placenta accreta include prior Cesarean section, placenta
previa, and prior instrumentation, among others. All forms of accreta may
be associated with life-threatening hemorrhage, which may require imme-
diate hysterectomy.
c. Exogenous hormone therapy
1. Estrogen causes proliferation of endometrial glands and stroma. Persistent
exposure to estrogen (exogenous as well as endogenous estrogen, as occurs
with anovulatory cycles, obesity, or an estrogen-secreting tumor) causes
endometrial proliferation with subsequent glandular and stromal break-
down, often clinically interpreted as irregular menstrual bleeding. Micro-
scopically, the findings include stromal condensation with formation of
so-called exodus bodies or stromal blue-balls, glandular degeneration and
apoptosis of the glandular epithelial cells, and fibrin thrombi in stromal
vessels (e-Fig. 33.17).
2. Prolonged exposure to progestogens results in endometrium with a char-
acteristic pattern that includes underdeveloped, inactive glands in a back-
ground of a stroma that shows marked decidual change (e-Fig. 33.18).
3. Tamoxifen is primarily used for the treatment of breast cancer. In the
endometrium, tamoxifen competitively binds to estrogen receptors and acts
as an agonist. It increases the risk of endometrial hyperplasia and adeno-
carcinomas (Ann NY Acad Sci. 2001;949:237), and up to 20% of women
on tamoxifen develop endometrial polyps (Cancer. 2001;92:1151).
IV. COMMON BENIGN DISEASES OF THE ENDOMETRIUM
A. Endometritis
1. Acute endometritis is defined by the presence of acute neutrophilic inflamma-
tion in the stroma of the nonmenstruating endometrium. In severe cases, the
neutrophils are present throughout the stroma, the endometrial epithelium,
and the glandular lumina (e-Fig. 33.19). Acute inflammation present dur-
ing the menstrual phase of the endometrium should not be misdiagnosed as
active infection. Acute endometritis is uncommon and is usually seen only

in postpartum or postabortive endometrium.
2. Chronic endometritis is defined by the presence of plasma cells in the endome-
trial stroma. Associated features include glandular and stromal breakdown,
and dyssynchronous glandular and stromal development (e-Fig. 33.20). The
most common causes of chronic endometritis include Chlamydia trachoma-
tis, Ureaplasma urealyticum, cytomegalovirus, and herpes virus infection.
Infection by Actinomyces israelii or Neisseria gono"heae usually causes a
mixed acute and chronic pattern of inflammation. Granulomatous inflam-
mation is rare; common causes include Mycobacterium tuberculosis infec-
tion, fungal infection, sarcoidosis, and hysteroscopic ablation therapy.
B. Atrophy is most commonly seen in postmenopausal women. Premenopausal
causes include treatment with oral contraceptives or gonadotropin agonists
(Lupron). Patients with premature menopause also show an atrophic pattern.
Microscopically, the endometrium is composed of a thin layer of endometrial
glands lined by an attenuated layer of inactive epithelial cells surrounded by
thin stroma. No mitotic activity is present (e-Fig. 33.21).
C. Metaplasia is the presence of any type of glandular epithelium other than the
normal columnar type. Metaplasia is a common finding in perimenopausal
and postmenopausal women and is often associated with abnormal uterine
bleeding or recent use of exogenous hormonal therapy.
1. Tubal metaplasia consists of foci of normal tubal epithelium within the
endometrial glands, including ciliated, nonciliated secretory, and interca-
lated cells. The ratio of the ciliated to nonciliated cells is cyclical and depends
on hormonal influences.
2. Ciliated cell metaplasia is the most common form of metaplasia. It is com-
posed of a layer of ciliated columnar cells with round to oval nuclei and
abundant pale eosinophilic cytoplasm (e-Fig. 33.22). Ciliated cell meta-
plasia is a normal response of endometrial epithelium to various hormonal
exposures. It is most commonly found in perimenopausal endometrium and
is associated with endometrial polyps, anovulatory cycles, and exogenous
hormonal therapy.
3. Squamous metaplasia (e-Fig. 33.23) is often caused by chronic irritation and
often takes the form of squamous morules or rounded, swirling nests of
squamous cells. Squamous metaplasia must not be confused with endome-
trial hyperplasia or malignancy, although it can occur as a secondary change
in both.
4. Eosinophilic metaplasia or eosinophilic change refers to glandular epithe-
lium with abundant eosinophilic cytoplasm and a central round to oval
nucleus (e-Fig. 33.24). It is often associated with a neutrophilic infiltrate,
the formation of small epithelial papilla, and mild nuclear atypia.
5. Mucinous metaplasia is rare. It is morphologically similar to endocervical
mucinous epithelium in that it consists of columnar epithelium with basally
located oval nuclei and abundant apical mucin (e-Fig. 33.25).
6. Clear cell metaplasia is also rare. It is characterized by columnar cells with
round nuclei and dear cytoplasm.
D. Endometrial polyps are local overgrowths of endometrial glands and stroma that
protrude into the endometrial cavity. Polyps are present in about 20% to 25%
of women and are frequently found in the perimenopausal and postmenopausal
period. Grossly, polyps appear as broad-based to pedunculated lesions; some
pedunculated polyps can extend into the endocervical canal, and even through
the os. Microscopically, polyps are composed of endometrial glands within a
fibrous stroma; the presence of thick-walled blood vessels within the fibrous
stroma is the most common key to the diagnosis (e-Fig. 33.26). Frequently, the
glands are variably shaped and irregularly distributed.
Although endometrial polyps in postmenopausal women usually contain

dilated glands lined by one layer of atrophic epithelium, foci of metaplastic
or hyperplastic epithelium, as well as frank adenocarcinoma, may be present.
Consequently, endometrial polyps should be entirely submitted for microscopic
examination.
1. Polyps with stromal smooth muscle are referred to as adenomyomatous
polyps.
2. Atypical polypoid adenomyoma is a polypoid lesion characterized histologi-
cally by crowded irregular endometrial glands with a complex architecture
and cytologic atypia in stroma that is predominantly composed of smooth
muscle (e-Fig. 33.27). The lesion has a high rate of recurrence after incom-
plete surgical removal and mainly occurs in premenopausal, nulliparous
women. It is associated with a clinical history of infertility.
E. Disordered proliferative endometrium predominantly exhibits a normal prolif-
erative pattern, with mild irregular branching and budding and some cystic
dilation. Howevet; the glands to stroma ratio is not increased-the main fac-
tor that helps to differentiate a disordered proliferative pattern from simple
hyperplasia. The epithelium lining the glands is composed of stratified and
columnar cells with no atypia. Mitotic activity is similar to that of normal
proliferative endometrium (e-Fig. 33.28).
V. ENDOMETRIAL HYPERPLASIA AND ENDOMETRIAL INTRAEPITHELIAL CARCINOMA.
Endometrial hyperplasia is thought to develop as a result of unopposed estro-
genic stimulation. Any disorder that causes an increase in endogenous or exoge-
nous estrogenic stimulation such as polycystic ovarian disease, obesity, or ovar-
ian neoplasms (e.g., thecomas, granulosa cell tumors) can therefore predispose to
endometrial hyperplasia. Abnormal bleeding is the major clinical symptom.
A. Endometrial hyperplasia is subclassified as simple or complex on the basis of the
architectural pattern, and as with or without atypia on the basis of the cytologic
features. Numerous studies have demonstrated that the risk of progression to
adenocarcinoma (specifically, endometrioid adenocarcinoma and its variants)
is more highly correlated with the presence of cytologic atypia than the degree
of glandular crowding.
1. Simple hyperplasia shows a glands to stroma ratio that is slightly increased
(more than 1:1) with prominent variability in size of the glands, glandular
budding, and cystic glandular dilatation (e-Fig. 33.29).
2. Complex hyperplasia is composed of crowded, architecturally complex
glands with little intervening stroma. The glands to stroma ratio is elevated
(at least 3:1) (e-Fig. 33.30).
3. Cytologic atypia, which may be a feature of simple or complex hyperplasia, is
based on the nuclear cytology of the glandular epithelium. The most reliable
indicators of cytologic atypia are an enlarged nucleus that is round rather
than oval, that has coarse clumped chromatin, and that has a prominent
nucleolus (e-Fig. 33.31). A diagnosis of hyperplasia should be made with
extreme caution during the secretory phase of the endometrium because of
the usual crowding of the glands in this phase of the menstrual cycle. The
presence of cytologic atypia must be distinguished from the cellular changes
that accompany metaplasias, and from Arias-Stella reaction.
B. Endometrial intraepithelial neoplasia (EIN) represents an alternative classifica-
tion scheme for premalignant endometrial lesions. EIN is defined as a prolifera-
tion of islands of endometrial glands which have cytological and architectural
abnormalities and are considered to be premalignant. The EIN scheme has
been proposed both as an approach to simplify the diagnosis of premalig-
nant changes and as a classification more closely linked to the genetic changes
in premalignant endometrial epithelium (Gynecol Oncol. 2000;76:287) pre-
ceding transformation into endometrial adenocarcinoma. Studies have shown
that some of these lesions harbor PTEN tumor suppressor gene inactivation,
mutations of k-ras. and microsatellite instability. EIN is considered to be a
monoclonal proliferation composed of cells in early stages of carcinogenesis.
Diagnostic criteria for EIN (http://www.endometrium.org) are based on
size of the lesion, nuclear cytology, and glandular architecture (including glan-
dular crowding with an increased glands to stroma ratio, cytologic differences
between the neoplastic glands and the adjacent normal glands, and an area of
glandular crowding that is > 1 mm in greatest dimension). EIN can have squa-
mous, mucinous, or clear cell differentiation. The treatment of these lesions is
similar to the clinical management of atypical endometrial hyperplasia.
C. Endometrial intraepithelial carcinoma {EIC) is thought to represent the precursor
lesion to serous carcinoma. Microscopically, EIC is composed of glands lined
by cells with the same cytologic abnormalities as serous carcinoma, but without
evidence of myometrial stromal invasion (e-Fig. 33.32). The development of
EIC is independent of prior unopposed estrogenic stimulation and typically
arises in a setting of atrophic endometrium.
VI. EPITHELIAL MALIGNANCIES. Epithelial malignancies are the most common gyne-
cological malignancy in women in developed countries. Endometrial cancer can
be divided into two broad categories which have differences in their clinical and
pathologic features, as well as their underlying genetic abnormalities.
A. Type I tumors consist of endometrioid adenocarcinoma and its variants. They
account for over 80% of endometrial tumors and usually develop in post-
menopausal women in their fifth and sixth decades in the background of long-
term estrogen stimulation. Type I tumors are strongly associated with diabetes
and obesity and have a relatively good prognosis. The endometrial glands and
stroma in Type I tumors are strongly positive for estrogen and progesterone
receptors. In addition, the stromal cells show diffuse strong immunopositivity
for CD10 (CALLA) antigen. Genetically, they show microsatellite instability,
and mutations in the PTEN tumor suppressor gene, k-ras, and CTNNB 1. but
assessment for these genetic abnormalities is not necessary for diagnosis.
B. Type II tumors, for example serous papillary carcinoma, usually occur in women
in their sixth and seventh decades and are not associated with estrogen stimula-
tion, and therefore do not occur in a background of complex atypical hyperpla-
sia but rather in atrophic epithelium. They are more likely than Type I tumors
to be at an advanced stage at the time of presentation and so have a relatively
poor prognosis. Genetically they are characterized by TP53 mutations.
The WHO classification of uterine corpus malignancies is presented in
Table 33.1, and the pathologic staging of uterine corpus carcinomas is shown
in Table 33.2 and Figure 33.1.
1. Endometrioid adenocarcinoma usually arises in the uterine corpus and grossly
usually consists of a raised to exophytic, pink tan, hemorrhagic mass that
projects into the endometrial cavity.
Microscopically, the tumor consists of irregula.t;, confluent, complex
glandular or villoglandular structures lined by pleomorphic stratified
columnar cells with pleomorphic nuclei. The presence of areas with defini-
tive cribriform architecture is a microscopic feature that can be used to
distinguish well-differentiated endometrioid adenocarcinoma from com-
plex hyperplasia with cytologic atypia. Foci of squamous differentiation,
which should not be mistaken as solid component of the tumor, are often
encountered.
Myometrial invasion is recognized by the presence of an irregular
endometrial-myometrial border or by an associated desmoplastic and
inflammatory stromal response (e-Fig. 33.33). The depth of myometrial
invasion compared with the full thickness of the myometrium and the pres-
ence or absence of lymphovascular space invasion should be noted.
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Squt.n'd.i& eel t~~~n::!l"'l:::''''::l Otllor benl!l> """"""hymal1>1mo111
TIINI!Ionllcol c.clrana -ll!ltillllalnl•-•••.,_
Smll coli aol<hlm1 C._,..,me l)n&Ja11ont mblal mLI-
Undll!'--me tlimo<l
otllcn AdOIIOOl!CI)mll
E'lldomdlllll\';l>ell>lool; 0.-bra/M
NonaypUIIM>eiJll&aM Adenollbn>ma
stnplo Ad..,..,..,.
C<m!Mx Alyplcal polyJdd ..... nt
~ hyperplula
stnplo - ..... ,111 tllllt 1 ' -
'J)ot>hobloollc ·~laomo
C<mliox Chod.,..l<hlma
r.,.,.. .._
E'lldomdllll DOtr'l> _ Pl.e<>lrtll sill trat>ho- tlimOf
Ejlllholold trat>hoblullc lumor
.... . ,. . . . .Ill
Molu p""""nclos
Eildomdllll.....,.land "'IIIDd ll.mora IWalldlfllnn mol&
-P-
E'lldomdllll-1 nodulo Oomplelo
E'lldomdllll-1 ..,_,.,..., arade Pal!lal
Ul'ld~~I:I!I/'QOI'I'\1
Stnot:th muatl& turr.:R
Lelomyaun:oma ~~·~. nonmolutrat>ho-losbui
Ejlllll--1 tllo no<lulo and lllaquo
hil:p<>lcl¥1rlont
Sm- muoelo 111m ore of u""""lh
'*"lrtll
"-!lllod olio
- ,....,..,.,.
. . . . . e-n
malpllt -1181
Ser. c:.orll·lllio Iii'"""'
Lelomyano, notdllellolaa apodlled
Nol.rcoctQdon'nl tumn
HIIIDI:!P: ¥1111n11
Mllotlcallf - ..tbrt
...
1\J 11101$ of...,., QlllllYI>O
CdiiAir w.rllnt
OhiO
HomorrhOIIc colkllu \lllllolrt
E'dlll- """"
Jotpolclv.rtlnt ~····---~~~~·--
""fiN nllflilpll"'""
~ ..lllnl
Upololom)'M'II "'lllnl
,......,...,....
uou-
The FIGO srdDig .,....., for aulo.IIXII:iJ>id. ~it .......,_

""tiLe~ of dilfaani&oi.., u cldinocl &y tiLe pm:ao!41ii' of 111andulu
alld aol!d. ~ (&l'COI of llqiWD<I1I.t diH=:allat!Oil uc 110t CCilllld·
ere.! ~of tolid grQWib), 'lll.cioM with 0% to · " «>lid growd> ...
11.ani .. D _ , .
,...,._ m
n•
c.t~p~•
1X Pttndryt.rmor 011nnot be !l!nMMd
TO ~-""'of ptlmll)' 11/mor
1lo Cln:tlom.t In 1111 (pnoln- camom.t)
T1 I TUITIOf con!lnodto""""' Witl
n. lA TuiTIOf llm!lod """cloi1\0ii'llm « lnwdoolim tllln -half
of111e II'(I'Oinolllum
Tlb
T2
Tao
19
II
III A
TUm«ln'OILIM.....,. but-
TUm«ln'OILIM ~Wor """"of ttl& ~urn
not-.! bo)l>nd wru1
TuMOt-MIODOIId/ofa.d.- (dlrecl-nllon«
-)
T.lb 1118 Vqholl-(dl'ect--.,« mldllltalll)
T~ riA TUm«.........,.. bloddor mu .... 01\d/of b<Mol m1100011
Ml Ml Ollourt ...-~o (a<ludlnJI:,_*" ~. pel>lo
...,
r«
.... .,.,.-(II) Mr'tll:t, ,. td l"'leXX)
R.....,.lljmJ:tl """'"....,not be • .,...,~

NO ~ roalonol ~1>11 node - •
Nl RllloneiiJmli!IIOdo m-ol>
N2 R...,lljmJ:tl IIOd• m-.oi&W> ~ ~1>11
no6ol, wlllloriOI!IIout llOIIIot poMc tlfnpll n -
.......,1... (10
Ml(
MO
-.Ia
Ollll.ant
~diiWrt-•
oonnot be • -
Ml Ollourt-llt(eo<ludo&m-.oi&W> penl.llrielymph
node<,~. peMe ....... t l etlMl!!)
lis NO MO
Tlo NO MO
Tlb NO MO
T2 NO MO
T3a NO MO
T3b NO MO
T1-T3 Nl MO
Tl-'D N2 MO
T4 AllYN MO
An/T AnyN Ml
Chapter 33 • Uterus (Corpus} ! 2!
T 1a- Tumor limited to endometrium or invades Tr Tumor invades cervix

less than one-half of the myometrium
T 1b- Tumor invades one-half or more of the
myometrium
T38 - Tumor involves the serosa and/or T4 a- Tumor invades bladder or

adnexa (direct inv or met) rectum
T3 b- Tumor invades vagina or parametrium T4 b- Distant metastasis (excluding
(direct ext or met) adnexa, pelv & abd)
Figure 33.1 pT staging of uterine carcinoma
characterized by glands composed of cells with supra or subnuclear

vacuoles resembling secretory endometrium. The mucinous pattern is
defined by the presence of foci of endometrial glands lined by colum-
nar cells with abundant intracytoplasmic mucin, often with a papillary
architecture. Squamous differentiation (e-Fig. 33.39) is defined as the
presence of sheets of squamous cells which are usually, but not always,
nonkeratinizing.
b. The distinction between endocervical adenocarcinoma and endometrial
adenocarcinoma is often difficult, especially on small biopsy specimens
or when the tumor involves the lower uterine segment. The distinction
has clinical significance, as endometrial adenocarcinomas are treated
with simple hysterectomy and endocervical adenocarcinomas are treated
with radical hysterectomy or radiation therapy. The clinical history can
often be very helpful in the distinction. Woman with endometrial can-
cer are typically obese and often diabetic; their uteri are often enlarged
and they report menometrorrhagia or postmenopausal bleeding. Women
with cervical cancer do not necessarily have a history of obesity but are
often smokers; they have a history of abnormal Pap smears or prior
cervical intraepithelial neoplasia (CIN) or adenocarcinomas in situ (AIS)

and are often symptomatic with postcoital spotting. The presence of
coexisting atypical endometrial hyperplasia, stromal foam cells and
benign morular squamous elements associated with carcinoma favor the
diagnosis of endometrial adenocarcinoma, whereas adjacent cervical AIS
orCIN favor the diagnosis of endocervical adenocarcinoma.
Immunohistochemistry can be useful when histologic features are
ambiguous. The panel of estrogen receptor (ER), vimentin, CEA, and
p16 can usually reliably distinguish between primary endometrial and
endocervical adenocarcinoma, since ER and vimentin will be positive in
endometrial and negative in endocervical adenocarcinoma, while CEA
will only be positive in endocervical adenocarcinomas. Endocervical car-
cinomas are typically diffusely positive for p16 whereas endometrial car-
cinomas show only patchy positivity. In situ hybridization with HPV
may also be helpful; endocervical adenocarcinomas should be positive
and endometrial adenocarcinomas negative.
2. Serous adenocarcinoma is a high-grade tumor characterized by cells with
a high nuclear to cytoplasmic ratio and a high mitotic rate that form a
complex papillary architecture (e-Figs. 33.40 and 33.41). Deep myometrial
and lymphovascular invasion are often present. The distinction between
high-grade serous and grade 3 endometrioid tumors is often difficult and
has high interobserver variability but is clinically meaningful as the survival
is significantly worse for serous adenocarcinoma. Although not useful for
distinguishing serous adenocarcinoma from lower grade endometrioid ade-
nocarcinomas, p16 and PTEN stains are sensitive and specific for high-grade
serous adenocarcinoma when used to distinguish the tumor from grade 3
endometrioid adenocarcinoma.
3. Clear cell adenocarcinoma is a high-grade tumor composed of pleomor-
phic cells with hobnail nuclei (nuclei that jut into the gland lumen), abun-
dant clear cytoplasm, and distinct cell borders arranged in papillary, solid,
and tubular structures, often admixed (e-Fig. 33.42). A characteristic fea-
ture is the presence of hyalinized stromal cores (e-Fig. 33.43). Clear cell
adenocarcinoma is often at an advanced clinical stage at the time of
presentation.
4. Mixed adenocarcinoma is defined as a tumor demonstrating a mixture of
endometrioid adenocarcinoma (or its variants) together with serous, muci-
nous, or clear cell adenocarcinoma. By convention, the minor component
must comprise at least 10% of the tumor.
5. Carcinosarcoma {malignant mixed mullerian tumor or MMMTI comprises
""10% of all uterine malignancies. The diagnostic criteria are based on
the presence of both malignant epithelial and mesenchymal (sarcomatous)
elements. Numerous genetic studies have demonstrated that both elements
are derived from the same precursor, proving that the neoplasm does not
represent a collision tumor. Consequently, the tumor is now considered to
represent a poorly differentiated endometrial carcinoma with metaplastic
differentiation.
Grossly, carcinosarcomas appear to be larger and fleshier than typical
adenocarcinomas and are often described as polypoid. Microscopically, they
consist of areas of adenocarcinoma (typically high-grade serous) intermixed
with a wide range of malignant mesenchymal elements such as smooth
muscle, cartilage, skeletal muscle, or undifferentiated (e-Fig. 33.44 and
33.45). Foci of poorly differentiated cells with marked pleomorphism and
a high mitotic rate with no distinct pattern are not uncommon. Extensive
areas of necrosis are often present. Carcinosarcomas can be divided into
homologous or heterologous tumors depending on whether the malignant
mesenchymal component is normally found in the uterus or not; this dis-

tinction is now recognized to have no clinical significance.
Carcinosarcoma generally has a poor prognosis. Specific adverse prog-
nostic factors include the presence of epithelial component with foci of
serous or clear cell differentiation, deep myometrial invasion, cervical
involvement, and lymphovascular space involvement. The grade of tumor,
type of the mesenchymal element, and mitotic rate have no correlation with
the outcome.
6. Squamous cell carcinoma of the endometrium is rare and usually occurs in
postmenopausal women in association with pyometria and cervical stenosis.
Microscopically, it is identical to squamous cell carcinoma of the cervix, and
so must be distinguished from a cervical primary that has extended into the
endometrium.
7. Other primary malignant tumors. Transitional cell carcinoma (TCC) is
extremely rare and occurs in postmenopausal women. Microscopically,
it consists of sheets of urothelial cells admixed with another type of the
endometrial adenocarcinoma, and must be distinguished from metastatic
TCC from the urinary bladder or ovary. Small cell carcinoma of the
endometrium is extremely rare and comprises < 1% of primary endometrial
malignancies. Microscopically, the tumor has the same cytomorphology as
high-grade neuroendocrine tumors arising at other sites. Undifferentiated
carcinomas do not show differentiation toward any defined tumor pattern.
VII. ENDOMETRIAL STROMAL TUMORS. These are composed of small cells with scant
cytoplasm that morphologically resemble the endometrial stromal cells of prolif-
erative phase endometrium. A subset of tumors exhibit variant morphologic pat-
terns including smooth muscle differentiation, a fibromyxoid component, and sex
cord-like/epithelioid patterns. The t(7;17)(p15;q21) translocation which produces
a JAZF1-JJAZ1 fusion protein is a recurring feature of all classes of endometrial
stromal tumors, but demonstration of its presence is not required for diagnosis.
A. Endometrial stromal nodules are grossly tan to yellow, well-circumscribed
lesions with a smooth border that range from 0.5 to 12 em in greatest dimen-
sion. They are primarily located in the myometrium, and an obvious connection
to the endometrium is not necessary for diagnosis. Histologically, these tumors
are composed of sheets of small cells with scant cytoplasm and an accompany-
ing vascular pattern reminiscent of the spiral arterioles present in the stroma of
proliferative phase endometrium. These tumors stain strongly and are diffusely
positive with CD10 (although a minority of cases may show weak positivity)
and are negative with desmin. Since cellular leiomyomas and leiomyosarcomas
are usually negative for CD10 but positive for desmin, immunohistochemistry
can be helpful in difficult cases. Endometrial stromal nodules are benign and
total abdominal hysterectomy is curative.
B. Endometrial stromal sarcomas predominantly occur in the middle-aged women
and do not share the same risk factors as endometrial carcinoma. On gross
examination, endometrial stromal sarcomas exhibit a tan to yellow cut surface
with an infiltrative border into the surrounding myometrium, often with foci
of hemorrhage and necrosis. Microscopically, the tumor consists of finger-like
projections into the myometrium of cells with similar cytomorphology as those
seen in endometrial stromal nodules, but with a higher rate of mitosis, greater
nuclear pleomorphism, prominent stromal vascularity, and areas of collage-
nized stroma (e-Fig. 33.46). Extensive lymphaticinvasion is the hallmark ofthe
tumor. Hysterectomy with salpingo-oophorectomy is the treatment of choice.
Radiation therapy has been shown to decrease local recurrence rates but does
not have a significant effect on long-term survival. The same chemotherapeutic
agents used to treat soft tissue sarcomas have been used to treat endometrial
stromal sarcoma with widely variable results. Patients with early stage disease
I'1,1,! If!.,
I ~an: FIGO tta;,.., ,.,.__,.IJid Enclomttltll . _ . .
11......D _ , .
,....,._ m
n•
Collj:io1•
,.
....
1X Prtn:tuy tl.l"n« ctntu:t btl • stened
TO No-...:adplt!wyi!Jmor
T1 I Tumllfllmbd to tho ulaM
n. tA Tumor ~5 em In Jff18b:t1t d!mstl.lb'l
Tlb 18 111m« >5""
T2 II 111m« -do lloYond 11\o tnonlo. v.tlilo111o110tlt>
T2o IIA Tumor lnwtte& l!ldl'lllrltf.
T2b liB Tumor I n -oiNt 110M<-
T3
Tao
Tab
...,...1.,.,.-
T~
Ill
lilA
1118
IVA
(II)
-·
Tumor 11'11'11:rtla abdomNI tlaul!lll
More1holl 0110 ello

111m«l•lood• blodder « Rld!.lrn
~ ~nil lymph -annat bt ••IOd

NO No ...,.,.l!rmph , _ .........
Nl R"'"nlllymph ncdo I110butasla
. _.......... 011
Ml( Obart mete!f:Mla tAnrd btl •!tSMt*d
MO NodiAintmlllotal&
Ml OIIUri mlll.tub
abllomnet-
<"""""••-· peM< and
tu,pll'tlllllfqfot•N
fllqo I Tl
fet.,..u••••
NO
••••tutef
MO
cr.eJ.., ••
fllqo lA Tie NO MO
SlqtiB Tlb NO MO
Slqj:lll T2 NO MO
Slqj:IIIIA T3e NO MO
Sla.e>IIIB Tab NO MO
fllqo IIIC TI-T3 Nl MO
fllqo IVA T-' AllyN MO
SIQt Ml Any T AllyN 1.41
women of reproductive age; they can be found in 20% to 30% of women

in their fourth decade, and more than 40% of women in their fifth decade.
They tend to enlarge during pregnancy since they express estrogen and pro-
gesterone receptors. Grossly, leiomyomas are well-circumscribed lesions; they
have a white-tan cut surface and are sharply demarcated from the adjacent
myometrium. Microscopically, leiomyomas are composed of interlacing fas-
cicles of closely packed cells with uniform elongated nuclei and eosinophilic
cytoplasm (e-Figs. 33.47 and 33.48). Degenerative changes including hyaline
change, coagulative necrosis, and hydropic degeneration are often present.
1. Cellular leiomyomas have the same gross features as ordinary leiomyomas
but microscopically demonstrate an increased cellularity with sheets of
spindle cells with hyperchromatic elongated nuclei and a scant amount
of eosinophilic cytoplasm (e-Fig. 33.49). There is no pleomorphism or
increased mitotic activity. These lesions behave as classic leiomyomas. One
important differential diagnosis for this lesion is endometrial stromal sar-
coma. Endometrial stromal sarcomas lack a fascicular growth pattern,
thick-walled vessels, and a cleft-like space between the lesion and the adja-
cent myometrium; in addition, endometrial stromal sarcomas often have
plaques of collagen and foamy cells which are not seen in leiomyomas.
Endometrial stromal sarcomas are typically positive for CD10 and nega-
tive for H-caldesmon and CD44v3. Also, the presence of mast cells is a
sensitive and specific finding, favoring cellular leiomyoma, when there are
usually > 7 mast cells per high power field.
2. Epithelioid leiomyomas are composed of predominantly epithelioid cells with
eosinophilic to clear cytoplasm and fine nuclear chromatin. The cells are
arranged in clusters and as single cells, with no pleomorphism. The mitotic
rate is not elevated, and necrosis is absent. They behave the same as classic
leiomyomas.
3. Symplastic leiomyoma contains scattered enlarged, markedly atypical cells,
often with multiple nuclei. However, the mitotic count is still <10 mitotic
figures per 10 high-power fields (hpf), and no necrosis is present. These
lesions have the same benign behavior as classic leiomyoma.
4. Lipoleiomyoma refers to a classic leiomyoma which contains islands of
mature adipocytes. This variant has no clinical significance.
5. Myxoid leiomyoma consists of fascicles of uniform spindle cells surrounded
by pools of myxoid edematous stroma. Large vessels are not uncommonly
present.
B. Benign metastasizing leiomyoma. Many patients who have benign metastasizing
leiomyoma have a prior history of hysteroscopy with dilatation and curettage,
or other procedures such as myomectomy or hysterectomy. Microscopically,
benign metastasizing leiomyomas have the same histologic features as ordinary
leiomyomas, although they may extend into adjacent vessels. The tumor cells
can migrate to the lung, and lymph node involvement may be present. The dif-
ferential diagnosis includes low grade leiomyosarcoma, and a smooth muscle
tumor of another site (such as gastrointestinal or retroperitoneum) must be
excluded. Some cases may respond to hormonal therapy.
C. Intravascular leiomyomatosis refers to classic leiomyomas that grow into the
lumen of the uterine or pelvic veins. The tumor may migrate or extend into
the inferior vena cava, and even into the right heart. The tumor is composed
of sheets of spindled to round cells with minimal atypia and rare mitoses, for
which the differential diagnosis often includes low grade endometrial stromal
sarcoma. With local control, the tumor has an excellent long-term prognosis.
D. Leiomyosarcoma is the malignant counterpart of leiomyoma. It is the most
common sarcoma of the uterus, with an incidence of 2 to 3 for every 1000
women with leiomyomata. Leiomyosarcoma usually occurs in women over
IJO I SECTION VII· REPRODUCTIVE TRACl
,, /,~,!I J}t§l Stratte for IXIIMCUI IIWIIt SIIWIOIII Mlltde Tu-
..,..""'"
Dope of'***
1 •" ' "nmoto
aiJpla: ' - - mill {i...,lfiQUII) or modaafi>lo....,llt (oi,VIIIclnll
p,...,...,• - lliCTCN
----·-
M~llld<ll
M~ 1.-(MO <20 per 10 hpl, noCTCN, no qplll or no m,.tlhln mild~

alypla -llllomyrma «lllom)'orne \OIIIIina- m-lndM{MI < 5 per 10 hrt-
loa>m)omo; Mil!: 5 por 10 hpf-lrilomyrma \OIIIIIna- m - in<kol)
I~ mllollclndtill. bitt_..,..,
Mil!: 20 Plf hpf, no CTCN, nolfW!eor no""'"' thon mill ~laill::lfW!eololo,..,... wtlll
lmbd
Ml < 10 por hpf, no~ but\01111- m - " ....,. ~ "'YJlle -<typloal
loa>m)orna \01tl Nrtok d """'rr..,..
Ml;. 10 por 10 hpl, no CTCN bulwtlll dll'lllt modln !<>.....,.~ J~Jl!ia
~
Ml < 20 por lOhpf, no CTCN but wtlllfoalll modaofi>lo _,~ <IJplo- otypcoi
lelorr\IO>rno. but~ lmlllld
Ally Ml,- d - mold-'>,_.. ""Yf* and wt111 ~- . . , . , _ , . ,
Ml?; lOper !Ohpt, nofi> rnlde1yJlla blt- ~----•
Ml < 10. noli> mild olyJ:il b<h!lh CTCN --111 rn.-lllmoroftow mdjp\1111 pollntlll,
but..,..,,.,..,lmllod.
IX. OTHER MYOMETRIAL DISEASES

A. Adenomyosis is defined as the presence of benign endometrial glands sur-
rounded by endometrial stroma within the myometrium (conventionally at
least 2.5 mm below the endomyometrial junction). Grossly, the adjacent
myometrium can show a thick trabeculated pattern with punctuate hemor-
rhage. Microscopically, the glands usually show an inactive pattern, or a pat-
tern that is dyssynchronous with the endometrium (e-Fig. 33.52). In some
cases, particularly in postmenopausal women, only the stromal component is
present and the glands are sparse or completely absent. Endometrial adeno-
carcinoma occasionally involves adenomyosis but this occurrence is not con-
sidered myometrial invasion for staging purposes.
B. Postoperative spindle cell nodule is a benign lesion that usually occurs within a
few weeks following endometrial instrumentation or other surgical procedure.
It is grossly pink-tan and friable. Microscopically, it consists of granulation
tissue with surface ulceration, accompanied by a hypercellular proliferation of
spindle cells with elongated nuclei, moderate amounts of pink cytoplasm, and a
high mitotic rate arranged in fascicles. Numerous extravasated red blood cells
are usually present. Postoperative spindle cell nodule must be distinguished
from leiomyosarcoma; the former's distinct fascicular pattern of growth, lack
of pleomorphism, and characteristic clinical presentation are dues to the cor-
rect diagnosis.
C. Adenofibroma is a rare entity that usually occurs in postmenopausal women
who present with abnormal bleeding. Microscopically, it is composed of a
layer of epithelium with bland nuclear cytology that overlies a cellular fibrous
stroma composed of fibroblasts and endometrial stromal cells. Adenofibroma
is a benign entity and hysterectomy is curative.
D. Adenosarcoma is characteristically a tumor of postmenopausal women. It is a
rare neoplasm that arises most commonly from the endometrium and forms
a large, polypoid, lobulated mass that may fill the entire endometrial cavity
and prolapse through the cervical os. Microscopically, adenosarcomas are com-
posed of benign glandular elements in a malignant stroma that shows increased
cellularity, pleomorphism, and a mitotic rate of >2 mitotic figures per 10 hpf
(e-Fig. 33.53). The stroma is often condensed and hypercellular in periglan-
dular areas and often juts into gland lumens in a characteristic pattern (e-Fig.
33.54).
Adenosarcoma is best considered a tumor of low malignant potential and
has a better outcome compared with other uterine sarcomas. However, recur-
rence (which occurs in up to 25% of patients) is associated with very poor prog-
nosis. Treatment includes hysterectomy with bilateral salpingo-oophorectomy.
The pathologic staging of uterine corpus adenosarcoma is shown in Table 33.5.
X. GESTATIONAL TROPHOBLASTIC DISEASE. This group of diseases comprises abnormal
trophoblastic proliferations that arise from gestational trophoblast. In Western
populations, about 0.1% of pregnancies are affected; patients are often at the
extremes of reproductive age, and women with previous gestational trophoblastic
disease are at higher risk.
A. Hydatidiform mole
1. Complete molar pregnancies develop from fertilization of an empty ovum.
Complete moles are diploid but may be either heterozygous (15% of cases
due to fertilization of an empty ovum by two sperm) or homozygous (85%
of cases due to fertilization of an empty ovum by a single sperm with sub-
sequent duplication). Complete moles classically present as a larger uterus
than expected for gestational age that on ultrasound examination shows a
so-called snow storm pattern without fetal parts; the patient's serum hCG
is usually elevated for gestational age. Histopathologically, classic com-
plete moles are composed of markedly enlarged villi with central villous
122 SECTION VII• REPRODUCTIVE TRACl
lltlln
11.ani .. D _ , .
,...,._ m
n•
c.t~p~•
1X Prtn:tuy tl.l"n« ctntu:t btl • stened
TO No-...:adplt!wyi!Jmor
T1 I Tumor a>nflnod to ..,rpu& ut.w1
n. tA Tumllfllmbd to ondomollhmlancloooM<
Tlb 18 Tumor fn'IZidn 11m than OfiOohiltf ~ m~m
Ttc lC 'lllm«lnloodoe on&llohr m010 d ~m
T2 II 1\imor .....,do beyond 11\e """"'· ..tllln 11\e poMI!.
T2o IIA Tumor fnwt.te& adl"'lrrtf.
T2b liB Tumor I n -oiNt poMI:-
T3
Tao
T3b
...,...1.,.,.-
T4
Ill
lilA
1118
IVA
(II)
-·
Tumor fn'Jdws abdomhlf U.va:t
More1holl one ello

Tumor fniJII!Idi!ll bltlddt!r ml.letlllll!lftdlcl bC*I!II MUOOSII
~ RJ;jonll t,mph -annot be ••IOd

NO No ...,.,.l!lmph - moll ......
Nl Re!ilonol t,mph IIOdo , _ _
.... , n•oo
M)( OIIUI'II mauls ..,rd be .....oe.~
MO No dillon! m""'tasl&
<-••"'*""""
............., -·
Ml Obllnl mabnlt8!11s peMI: and
1bllomnat-
._.
fllqo I Tl NO MO
SIQt lA Tla NO MO
Slqj:IIB Tlb NO MO
SIQ:IIC Tic NO MO
Sla.elll T2 NO MO
Sla.e>IIIA T3e NO MO
fllqo 1118 Tab NO MO
SIQt IIIC Tl-l3 Nl MO
SlqtiYA T4 AnyN MO
Slqj:IIYB A1V T Any N Ml
2. Partial molar pregnancies develop from fertilization of a normal ovum by

two sperm, and so have a triploid karyotype. Patients may have a nor-
mal, elevated, or even low serum hCG for gestational age; fetal parts are
sometimes present by ultrasound imaging. Histologically, partial moles are
composed of two populations of villi; one population is essentially normal
or small and fibrotic, and the other exhibits enlargement with at least focal
cavitation, irregular villous outlines, trophoblast inclusions, and subtle cir-
cumferential trophoblast in the form of buds or lacy mounds (e-Figs. 33.59
and 33.60). Partial moles also carry a small but increased risk of subsequent
development of choriocarcinoma.
3. Invasive mole refers to a molar pregnancy (either complete or partial) in
which the villi and associated trophoblast invade the myometrium and
blood vessels or are exported to extrauterine sites (e-Fig. 33.61).
B. Trophoblastic tumors
1. Choriocarcinoma. Although molar pregnancies have a much increased risk
of subsequent development of gestational choriocarcinoma, the tumor can
develop following any type of pregnancy, including normal gestations.
Microscopically, choriocarcinoma is composed of sheets of highly atypical
trophoblast with prominent foci of hemorrhage and necrosis (e-Fig. 33.62).
The pleomorphic trophoblast consists of admixture of cytotrophoblast, syn-
cytiotrophoblast, and intermediate trophoblast and often forms alternating
collections of syncytiotrophoblast and mononucleate trophoblast. Chorio-
carcinoma spreads hematogenously, and the most common metastatic sites
are the lung, pelvis, vagina, liver, and brain. Choriocarcinoma is strongly
positive for cytokeratin, hCG, Mel-CAM, human placental lactogen (hPL),
and placental alkaline phosphatase expression by immunohistochemistry.
2. Placental site trophoblastic tumor is composed of intermediate trophoblast.
Characteristic features include abundant eosinophilic fibrinoid deposition
and dissection of individual smooth muscle cells by the neoplastic cells
(e-Figs. 33.63 to 33.65). This tumor typically presents as a mass which may
be deeply invasive; serum hCG levels are usually only mildly elevated. About
15% of cases exhibit malignant behavior; unlike most forms of gestational
trophoblastic disease, the tumor is not very responsive to chemotherapy.
3. Epithelioid trophoblastic tumor is composed of chorionic-type intermediate
trophoblast. It is a very rare form of trophoblastic disease that has only
recently been recognized as a distinct disease entity. Morphologically, the
tumor is composed of a uniform population of atypical mononucleate cells
arranged in sheets and nests associated with eosinophilic material and sur-
rounded by necrotic debris (e-Fig. 33.66).
XI. SEROSAL TUMORS
A. Endometriosis affects 5% to 10% of women of childbearing age. Patients usu-
ally present with symptoms of pelvic pain, dyspareunia, secondary dysmenor-
rhea and, in some cases, infertility. Many cases remain asymptomatic. The three
most common accepted theories regarding its etiology are retrograde spillage
of menstrual tissue into the pelvic cavity, serosal metaplasia, and a developmen-
tal abnormality. The most commonly involved sites include the ovary, uterine
serosa, fallopian tube, peritoneum, and cul-de-sac. Oral contraceptives have
been shown to have a protective role. Histologically, endometriosis is defined
as the presence of endometrial glands surrounded by endometrial stroma; asso-
ciated hemosiderin deposition and chronic inflammation are usually present.
B. Adenomatoid tumor is a benign peritoneal tumor that originates from mesothe-
lium. It most commonly involves the serosal surfaces of the uterus and fallopian
tubes. Grossly, the tumor usually forms a tan 1- to 2-cm well-circumscribed
nodule. Microscopically, the tumor is composed of tubular and slit-like
spaces lined by a single layer of flattened cuboidal cells with bland cytology
(e-Fig. 33.67). The cells are immunopositive for cytokeratin, calretinin, and
vimentin expression, but immunonegative for factor VIII-related antigen and
CD31 expression-a profile that can be used to distinguish the tumor from
metastatic carcinoma and vascular tumors. Adenomatoid tumor is clinically
asymptomatic and usually found incidentally.
XII. OTHER MISCELLANEOUS NEOPLASMS
A. Lymphoid neoplasms involving the uterine corpus (of which the most frequent
is large B cell lymphoma) most commonly represent a manifestation of dissem-
inated disease. Primary disease of the uterus is extremely rare.
B. Metastatic tumors only rarely involve the uterus. The most common primary
tumors that spread to the uterus are those of breast, lung, stomach, gallblad-
der, thyroid, and melanoma. In most instances, uterine involvement by primary
tumors of the ovary, cervix, bladder, and rectum/colon represents direct exten-
sion rather than hematogenous spread.
SUGGESTED READINGS
Mazur MT, Kurman RJ. Diagnosis of Endometrial Biopsies and Curettings. A Practical Approach.
2nd ed. New York: Springer; 2005.
Tavassoli FA, Devilee P, Eds. Tumours of the Breast and Female Genital Organs. 1st ed. Lyon:
International Agency for Research on Cancer; 2003.
Uterine Cervix
Michael E. Hull
I. NORMAL GROSS AND MICROSCOPIC ANATOMY

A. Gross. The cervix is the tubular distal portion of the uterus, divided into the
ectocervix and endocervix. The smooth ectocervix is covered by a reflection of
the vaginal mucosa, and the anterior and posterior fornices are formed by the
protrusion of the cervix into the vaginal vault. The posterior fornix is deeper
than the anterior fornix. The tan, rugous endocervix is a narrow canal that
begins at the external os. The external os is round and small in the nulliparous
state and becomes slit-like with parity. An internal os, or isthmus, marks the
transition from the endocervix to the endometrium. The parametrial soft tissue,
which attaches to the lateral aspects of the cervix, contains the uterine vessels
and the ureters. The posterior cervix is the anterior border of the pouch of
Douglas, the space between the uterus and the rectum; the anterior cervix is
immediately posterior and inferior to the bladder.
B. Microscopic. The ectocervix is generally covered by squamous epithelium in
continuity with the vaginal epithelium, while the endocervix is lined by colum-
nar mucinous epithelium. The transition from columnar to squamous epithe-
lium through the process of squamous metaplasia occurs over a region of the
cervical epithelium called the transformation zone. In states of low estrogeniza-
tion, the transition occurs approximately at the external os. With higher levels
of estrogen, the transition is observed on the portion of cervix visible in the
vaginal vault. The transformation zone is important diagnostically because it
is the site of the majority of cervical epithelial neoplasms and their precursors
(e-Fig. 34.1)."'
1. Squamous epithelium. The squamous epithelium is composed of a basal layer,
intermediate layer, and superficial layer. The basal layer is one-cell thick and
has a relatively high nuclear/cytoplasmic (N/C) ratio. The N/C ratio decreases
progressively from the basal layer to the superficial layer during normal mat-
uration, and the superficial squamous cells tend to align with their longest
axis parallel to the basement membrane. Directly sampled normal squamous
epithelium in cytologic preparations shows individual and clustered superfi-
cial polygonal squamous cells with pyknotic nuclei, intermediate cells with
somewhat larger nuclei, and more rounded parabasal cells with the highest
N/C ratios. In the estrogenized state, superficial cells predominate.
2. Columnar epithelium. The mucinous columnar epithelium of the endo-
cervix is one cell layer thick, with basal polarization of the cells' nuclei, little,
if any, mitotic activity, and an N/C ratio of about 1:4. Mucinous columnar
epithelium also lines the endocervical glands, which represent infoldings of
the surface epithelium rather than true glands. Directly sampled endocervical
columnar epithelium is seen in cytologic preparations as sheets of uniform
round nuclei in a "honeycomb" arrangement or as single-layered strips of
epithelium with basally oriented nuclei.
2. Squamous metaplastic epithelium. This is an expected finding in the trans-
formation zone of cervical specimens. Histologically, in the immature form,
the squamous epithelium underlies a layer of superficial residual columnar
535
8 7 6 5 4
Figure 34.1 Gross processing of conization specimens. After the endocervical and stromal mar-
gins are inked, the specimen is radially sectioned so that each tissue slice includes the endocervical
margin, the mucosal surface of the endocervical canal, and the ectocervix.
epithelium (e-Fig. 34.1); with full maturation, it may appear very similar to
native squamous epithelium. Metaplastic squamous cells in cytologic smears
occur as either singly dispersed cells or small sheets of cells, and they show
cyanophilic cytoplasm and nuclear sizes and N/C ratios between those of
normal intermediate and basal cells.
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION. Cer~
vical specimens for screening and diagnosis are obtained in several ways.
A. Exfoliative cytolao (Pap test). See the section below on cytology of the uterine
cervix for a discussion of the Pap test.
B. Biopsy. Colposcopic cervical biopsy specimens are small pieces of mucosa and
superficial stroma that are taken, most often, from acetowhite areas identified
visually. Documentation of the number and size of tissue fragments is impor-
tant to ensure that the biopsy tissue fragments are adequately represented on
the slides. If a tissue fragment exceeds 4 mm in maximal dimension, it should
be bisected prior to histologic processing. In general, the biopsy tissue should be
wrapped in lens paper or placed between sponges to avoid loss during process~
ing, and the tissue should be embedded such that the microscopic sections are
perpendicular to the mucosal surface. Three H&E~stained levels are prepared
for microscopic examination.
C. Curettaae. Curettage specimens consist of numerous and often miniscule tissue
fragments in mucus, so it is imperative to both filter the contents of the container
and collect any tissue that may be adherent to the pad or paper submitted
within the specimen container. It is necessary to wrap curettings in lens paper to
avoid loss during processing. The specimens obtained from curettage procedures
should be submitted in their entirety. Three H&E~stained levels are prepared
for microscopic examination.
D. Conization. Ideal cold knife cone biopsy specimens consist of a single tube of
ectocervix and cervical canal surrounded by stroma. Marking sutures attached
by the surgeon enable the sections to be designated using the hours of the clock;
by convention, the mid~anterior location is the 12 o'clock position. The endo~
cervical margin must be identified and inked differentially from the ectocervical
and stromal margins. After fixation, the specimen should be radially sectioned,
with each section encompassing the endocervical margin, the mucosal surface of
the endocervical canal with the transformation zone, and ectocervical margin,
as shown in Fig. 34.1.
E. Loop electrosuraical excision procedure (LEEP). The key to correct processing
of these specimens is identification of the endocervical margin (which may be
Chapter 34 • Uterine Cervix I 53 7
inked by the surgeon to facilitate identification); the ectocervix is smooth and

tan-white, whereas the endocervix is tan and more rugous. The endocervical
margin should be differentially inked from the ectocervical and stromal margins.
Radial sections should be taken perpendicular to the mucosa, encompassing the
endocervical margin, transformation zone, and ectocervical margin in the same
manner as for conization specimens.
F. Radical hysterectomy
1. Uterus. Before opening the uterus, the parametrial soft tissue is inked as it
represents soft tissue margins of interest. The vaginal margins are also inked.
The uterus is then bivalved. If no tumor is visible or the tumor does not
appear to extend into the parametrial soft tissue, the parametrial soft tissue
is removed, sectioned, and completely submitted. If the vagina appears free
of tumor, shave margins are submitted. If tumor appears to extend into the
parametrial soft tissue or vagina, radial sections (that include the tumor's
relationship to the margin) are submitted. If a cervical mass is present, at least
one section per centimeter of tumor, including the deepest extension into the
cervical wall, is submitted. If no tumor is visible, the cervix is amputated and
processed as a conization specimen.
2. Lymph nodes. Separate packets of pelvic and para-aortic lymph nodes are
typically submitted with radical hysterectomy specimens. The lymph nodes
should be separated from the soft tissue and entirely submitted.
Ill. DIAGNOSTIC FEATURES OF COMMON NONNEOPLASTIC DISEASES
A. Inflammation and infection. Acute cervicitis is a pattern of inflammation marked
by a stromal and epithelial neutrophilic infiltrate, with associated stromal
edema, and, often, reactive epithelial atypia. Reactive epithelium shows enlarged
nuclei and prominent nucleoli in a pattern that may be confused with neopla-
sia. Acute cervicitis is usually a nonspecific diagnosis, as the inciting agent can
be any of a wide variety of bacterial, fungal, or protozoan organisms. Chronic
cervicitis consists of a lymphoplasmacytic infiltrate that is also nonspecific. Pap-
illary endocervicitis is a term describing inflamed endocervical mucosa forming
papillary structures.
1. Noninfectious cervicitis. Cervicitis can be due to irritation from chemical
exposure, foreign materials (e.g., pessary, tampons), or surgical trauma. The
cervix may also be a site of involvement in systemic inflammatory conditions
such as collagen vascular disease. The type of inflammatory response may be
neutrophilic, lymphoplasmacytic, or granulomatous.
2. Infectious cervicitis
a. Bacterial cervicitis. Neisseria gonorrhoeae and Chlamydia trachomatis
both produce a mucopurulent cervicitis that requires additional nonhis-
tologic methods for specific diagnosis. With chronicity, C. trachomatis
infection can result in follicular cervicitis, a pattern of intense lymphocytic
infiltration that characteristically includes lymphoid aggregates with ger-
minal centers (e-Fig. 34.2). Although often associated with C. trachomatis,
follicular cervicitis is not specific for that infection.
Actinomyces spp. infection is associated with intrauterine device (IUD)
use and is often asymptomatic. The morphologic pattern is distinctive in
that clusters of purple-red filamentous organisms are seen in curettings
and smears, with associated "sulfur granules" consisting of clusters of
neutrophils with a basophilic center.
Bacterial vaginosis is characterized by "clue cells," squamous cells
coated with bacterial organisms. Gardnerella vaginalis and Mobiluncus
spp. are both implicated in the disease.
b. Viral cervicitis. Herpes simplex virus (HSV; primarily HSV type 2) infec-
tion is characterized by ulceration with enlarged epithelial nuclei, nuclear
molding, multinudeation, and margination of the chromatin in virally
infected cells at the edge of the ulcer (e-Fig. 34.3). Cytomegalovirus is dis-
tinctive for its nuclear and cytoplasmic inclusions. Adenovirus is notable
for its smudged nuclear inclusions. The poxvirus Molluscum contagia-
sum generates large, round, intensely eosinophilic cytoplasmic inclusions,
as it does in its cutaneous sites. Human papillomavirus (HPV) infection is
closely tied to cervical neoplasia; its features are discussed in the sections
on preinvasive and invasive squamous neoplasia.
c. Granulomatous cervicitis. Infectious causes of granulomatous cervicitis
include Mycobacterium tuberculosis and Treponema pallidum infections.
As noted above, noninfectious etiologies are also in the differential diag-
nosis of granulomatous inflammation.
d. Fungal. Candida spp. are commonly encountered in smears and are not
necessarily always pathogenic. They cannot be speciated reliably on their
morphology, in either tissue sections or cervical smears.
e. Parasitic. Trichomonas vagina/is is one of the most common causative
agent of sexually transmitted infections in women. Many infections are
asymptomatic. In Papanicolaou-stained smears or liquid-based prepara-
tions, an ovoid organism with an eccentric nucleus is observed; in liquid-
based preparations, squamous cells may be coated with the organism.
3. Vasculitis. Most cases of vasculitis involving the gynecologic tract are inci-
dental, and the vasculitis is often confined to the cervix. However, some cases
are associated with known collagen vascular disease, and rare cases repre-
sent the first manifestation of a collagen vascular disorder (e-Fig. 34.4) (Int
f Gynecol Pathol. 2000;19:258).
B. Atrophy. Epithelial atrophy is seen in the postmenopausal state, when estro-
gen levels are decreased. As noted in the discussion of normal histology, high
estrogen states are associated with large numbers of superficial squamous cells;
however, when the epithelium is thinned, the histologic picture is dominated by
small cells with increased N/C ratios and nuclei at least as large or larger than
those of normal intermediate cells. The overall appearance of a well-organized
epithelial architecture and a lack of nuclear atypia distinguish atrophy from a
severe squamous dysplasia.
C. Metaplasia. Tubal metaplasia occurs most frequently in the upper endocervix.
Transitional-cell metaplasia is rare and recapitulates urothelium; it is not asso-
ciated with a specific insult and must not be confused with neoplasia. Intestinal
metaplasia is another uncommon metaplasia; it features columnar epithelium
with goblet and Paneth cells.
D. Hyperplasia
1. Squamous hyperplasia consists of thickening of the epithelium with normal
maturation. It occurs in situations of prolapse and chronic irritation.
2. Squamous papilloma is a benign squamous proliferation that covers fibrovas-
cular cores. Squamous papilloma may be associated with HPVinfection but
it does not show classic koilocytic atypia.
3. Microglandular hyperplasia is an increase in glandular elements in the cervical
stroma. Seen in histologic sections, the exuberant proliferation sometimes has
a cribriform architecture that can raise the question of neoplasia. However,
microglandular hyperplasia shows no cytologic atypia, does not infiltrate the
stroma, and is not associated with a desmoplastic reaction.
4. Lobular endocervical glandular hyperplasia is a benign proliferation of bland
glands usually surrounding a central dilated gland and forming a well-
circumscribed lobule.
5. Diffuse laminar endocervical glandular hyperplasia. In this entity, the prolifer-
ation is primarily in the very superficial aspects of the stroma and extends in
a band-like fashion with an intermingled lymphocytic infiltrate that may be
dense.
E. Nabothian cysts are pronounced dilatations of endocervical glands. They are

extremely common (e-Fig. 34.5).
F. Endocervical tunnel clusters are superficial collections of endocervical gland duc-
tal spaces.
G. Mesonephric remnants are developmental remnants of the mesonephric (Wolf-
fian) duct that are occasionally identified in the deep stroma. Mesonephric rem-
nants must not be confused with adenocarcinoma; helpful distinguishing fea-
tures include the bland cytology of the lining epithelium, a lack of atypia in
the overlying endocervical glands, and the absence of a desmoplastic stromal
response (e-Fig. 34.6).
H. Postoperative spindle-cell nodule is a benign proliferation of fibroblasts that
usually occurs following surgical manipulation.
I. Endocervical polyps are often identified colposcopically. They consist of an exo-
phytic configuration of benign glands and stroma usually with thick-walled ves-
sels (e-Fig. 34.7). They must be carefully examined microscopically to exclude
coexisting squamous dysplasia or a glandular neoplasm.
J. Inclusion cysts are benign. Microscopically, they are filled with keratin debris
and are related to surgical manipulation. Similar inclusions occur in the vagina,
following episiotomies.
K. Endometriosis can involve any layer of the cervical stroma, as well as the parame-
triaVparacervical soft tissue.
L. Decidual change occurs in the cervical stroma during pregnancy. The nests of
cells that show abundant amphophilic cytoplasm, a prominent cell borde:r; and
a single, centrally placed nucleus are usually not visible grossly, but sometimes
form polyps (e-Fig. 34.8).
M. The Arias-Stella reaction, characterized by epithelial cells with nuclear enlarge-
ment and clear cytoplasm in response to progesterone, is most commonly seen
in the uterine corpus in pregnancy but may also occur in the cervix. The signif-
icance of the Arias-Stella reaction lies in the fact that it can easily be confused
with a glandular neoplasm.
IV. CERVICAL NEOPLASIA. The WHO classification of cervical tumors is shown in Table
34.1.
A. Benign
1. Submucosal and stromal neoplasms. Leiomyomas identical to those of the
uterine corpus are also seen in the cervix.
2. Blue nevus. This benign melanocytic proliferation is common, noted clinically
as a bluish discoloration of the cervical epithelium. Microscopic examination
shows hyperpigmented spindle cells infiltrating the stroma in a haphazard
pattern (e-Fig. 34.9).
3. Ectopic tissue. While not neoplastic, several types of ectopic tissue may be
seen in the cervix. The most common types of ectopic tissue are cutaneous
adnexal structures and mature cartilage. Prostatic ectopia has also been noted
to occur on occasion (e-Fig. 34.10) and is important to recognize to avoid
overdiagnosis of a glandular malignancy.
B. Malignant and premalignant squamous lesions. Worldwide, cervical cancer is the
third most common malignancy and the fifth most common cause of cancer
mortality in women. Effective screening programs have dramatically reduced
deaths due to cervical cancer in the developed world, but gains have been more
modest elsewhere.
The major risk factor for cervical cancer is sexually transmitted HPV infec-
tion. Although there are >40 different HPV serotypes that infect the female
genital tract, high-risk serotypes (including 16, 18, 35, 39, 45, 51, 56, and
58) are associated with a markedly increased risk of severe squamous dyspla-
sia and subsequent cervical squamous cell carcinoma. Immunodeficiency may
increase the likelihood of persistent infection and may increase the risk of
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squamous intraepitheliallesion [HSIL]) are defined by cytologic abnormali-

ties of the lower two-thirds and full thickness of the epithelium, respectively
(e-Figs. 34.12 and 34.13). Spontaneous regression occurs at lower rates than
in CIN1, ranging from 40% for CIN2 to 33% for CIN3. A more aggressive
approach to the management of these lesions is warranted, because progres-
sion to invasive carcinoma occurs in a higher percentage of these lesions
(Int J Gynecol Pathol. 1993;12:186).
4. Invasive squamous cell carcinoma is recognized by penetration of the epithelial
basement membrane by neoplastic squamous cells with an associated desmo-
plastic stromal response (e-Fig. 34.14). Keratinizing and nonkeratinizing
types are encountered. In contrast to the atypical cells characteristic of high-
grade squamous dysplasia in which the N/C ration is markedly increased,
the invasive tumor cells often exhibit paradoxical maturation evidenced by
abundant eosinophilic cytoplasm.
a. Microinvasive squamous cell carcinoma is defined as a tumor that is not
recognized as such clinically, that invades ::::;5 mm from the basement mem-
brane of the adjacent surface epithelium or endocervical gland from which
it arises, and that extends ::::;7 mm in greatest lateral extent (FIGO Stage
1A). When no lymphatic or vascular involvement is present, when the
entire lesion is excised, and when no dysplasia is present at the margins of
excision, the potential for lymph node metastasis or recurrence is very low.
Long-term follow-up studies have shown that the percentage of patients
harboring residual invasive carcinoma in a hysterectomy specimen after
conization for microinvasive squamous cell carcinoma with maximum
invasion of ::::;1 mm is 0; when invasion is ::::;3 mm, the recurrence and
lymph node metastasis rates are< 1 %. For lesions between 3 and 5 mm in
depth, the recurrence and lymph node metastasis rates increase to around
2% and 4%, respectively (Pathol Ann. 1995;30:103). Most studies find
3 mm of invasion to be a cutoff beyond which recurrence and lymph node
metastasis risk become significant.
b. Squamous cell carcinoma variants
i. Keratinizing tumors contain keratin pearls and nests of tumor cells with
central keratin; cytoplasmic keratinization and keratohyaline granules
are also present. Intercellular bridges can be identified.
ii. Nonkeratinizing tumors show cytoplasmic keratinization of individual
cells and intercellular bridges, but keratin pearls and nests of tumor
cells with central keratin are not present.
iii. Basaloid carcinoma features cells with scanty cytoplasm that resemble
basal type squamous cells. Only rare nests of tumor cells show central
keratin. This variant has an aggressive behavim:.
iv. Verrucous carcinoma is a very well-differentiated squamous cell carci-
noma that betrays its malignant character only in its invasion of the
stroma along broad pushing borders. There is minimal cytologic atypia,
and viral cytopathic effect is absent. Aggressive local invasion and recur-
rence after excision are common. Metastasis is uncommon.
v. Wany carcinoma is a rare squamous malignancy that shows definitive
stromal invasion but also abundant cytologic features of HPV infection.
It may behave less aggressively than other well-differentiated squamous
cell carcinomas.
vi. Papillary squamous cell carcinoma is an exophytic tumor in which
epithelium resembling CIN2 or CIN3 covers fibrovascular cores. Koilo-
cytosis is not characteristic. Although much of the tumor may have the
appearance of a precursor lesion, definitive stromal invasion is present
in the deep aspects of the lesion. Therefore, superficial biopsies of pap-
illary lesions should be interpreted with caution.
Chapter 34 • Uterine Cervix I 543
vii. Lymphoepithelial-like carcinoma resembles the nasopharyngeal tumor of

the same name. The tumor consists of syncytial sheets and islands of
undifferentiated epithelioid cells that have eosinophilic cytoplasm and
large vesicular nuclei with prominent nucleoli in a background that
contains an intense lymphocytic infiltrate.
c. Glandular neoplasms
1. Adenocarcinoma in situ (AIS} is an HPV-associated glandular lesion (most
strongly associated with HPV serotypes 16 and 18) that is a precursor to
invasive adenocarcinoma. Cytologically, AIS is characterized by loss of cyto-
plasmic mucin, cellular stratification, cellular crowding, nuclear enlargement
and atypia, mitotic activity, and epithelial apoptotic debris (e-Fig. 34.15). The
various subtypes can mimic endocervical, endometrial, or intestinal epithe-
lium and can have a papillary or cribriform architectural pattern. AIS is often
seen in conjunction with CIN.
In contrast to in situ squamous epithelial lesions, AIS of the cervix is less
common than its invasive counterpart. Because of this, follow-up studies are
sparse. However, a number of series do show that a positive margin for AIS
in a cone biopsy predicts the presence of either an invasive adenocarcinoma
or the recurrence of AIS (Gynecol Oncol. 2000;79:207), and therefore AIS
requires complete excision.
2. Microinvasive adenocarcinoma. The FIGO definition of microinvasive adeno-
carcinoma is the same as that of microinvasive squamous cell carcinoma. It
is a difficult diagnosis; the subjectivity of histologic assessment and relative
infrequency of in situ adenocarcinoma have presented challenges in the elu-
cidation of the natural behavior of microinvasive adenocarcinoma. What is
known is that it carries a good prognosis. When invasion is :::;2 mm, lymph
node metastases essentially do not occur (Obstet Gynecol. 1985;65:46;
Obstet Gynecol. 1997;89:88; Int] Gynecol Pathol. 2000;19:29).
3. Invasive adenocarcinoma. Infiltrating glands with cribriform and papillary
structures are the architectural characteristics of invasive adenocarcinoma.
A desmoplastic stromal reaction-a feature that helps to distinguish adeno-
carcinoma from both AIS and the hyperplastic entities described earlier-is
present (e-Fig. 34.16).
a. Mucinous adenocarcinoma. Most primary mucinous adenocarcinomas
have cytologic features resembling endocervical glands. The cells are
cuboidal to columnar and show nuclear pleomorphism, nuclear atypia,
and many mitoses. The relatively abundant cytoplasm stains positively for
mucin. Mucinous adenocarcinomas with goblet cells that have an overall
morphology more similar to intestinal epithelium are termed intestinal
variants.
b. Minimal deviation adenocarcinoma (adenoma malignum). This rare entity
comprises only 1% of primary adenocarcinomas of the cervix. The glan-
dular epithelium is so bland in appearance that these lesions may not
be recognized as malignant on biopsy or curettage specimens; increased
mitotic activity and cytologic atypia may be present focally, but these
findings are not prominent. The diagnosis rests on the presence of deep
infiltration, aggregation of glands around vessels or nerves, and a stromal
reaction-features that are easiest to assess on cone biopsy or hysterec-
tomy. The incidence of this lesion is increased in Peutz-Jeghers syndrome.
c. Endometrioid adenocarcinoma. Comprising 30% of cervical adenocarcino-
mas, endometrioid adenocarcinoma of the cervix is identical in appearance
to its counterpart in the endometrium. When well differentiated, the lesion
shows tall columnar cells without mucin. It may be very difficult to distin-
guish a cervical primary lesion from direct extension into the cervix of a
tumor of the uterine corpus. Immunohistochemistry may be of some use
in this situation, as endocervical adenocarcinomas are generally expected

to express CEA and p16, while endometrial adenocarcinomas tend to
be negative for these two markers and positive for vimentin and estro-
gen recepto.t Care should be taken when using immunohistochemistry,
because neither of these profiles has 100% specificity.
d. Well-differentiated villoglandular adenocarcinoma is considered to be a sub-
type of endometrioid adenocarcinoma. It shows an exophytic growth pat-
tern with glandular and villous elements, little nuclear pleomorphism, and
a low mitotic rate, quite similar to an intestinal villous adenoma (e-Fig.
34.17). There is value in the distinction, as this is a tumor of young women
and has a favorable prognosis (Gynecol Oncol. 1997;64:147).
e. Clear cell adenocarcinoma. This tumor's association with in utero diethyl-
stilbestrol (DES) exposure has made it widely recognized. Although rare,
the tumor can occur in patients without a DES-exposure history as well.
Clear cells with hobnail morphology are observed in solid, papillary, and
tubular arrangements. The presence of this tumor in the cervix should
prompt a search for a primary tumor of the ovary, endometrium, or
vagina-sites where this entity is much more common.
f. Serous adenocarcinoma is another adenocarcinoma that only rarely occurs
as a cervical primary tumor.
g. Mesonephric adenocarcinoma. This lesion differs from mesonephric rem-
nants in its cytologic atypia, crowding, and increased mitotic activity. It
arises in the deep lateral cervical walls and a variety of architectural pat-
terns are characteristic, including tubulat; papillary, solid, and retiform.
The behavior of this very rare tumor is generally indolent, if the tumor is
low stage.
D. Other carcinomas
1. Adenosquamous carcinoma. Both squamous and glandular differentiation are
observed in this lesion. To be diagnostic, invasive glandular and squamous
components must be present; that is, adenocarcinoma with adjacent squa-
mous dysplasia is not adenosquamous carcinoma. The epidemiologic profile
is similar to that of squamous cell carcinoma and adenocarcinoma.
2. Glassy cell carcinoma is considered a subtype of adenosquamous carci-
noma that occurs in young women. It carries a poor prognosis because it
is unresponsive to radiotherapy; progression is rapid, and distant metas-
tases are common. Microscopically, the tumor consists of sheets of pleomor-
phic cells with granular eosinophilic cytoplasm, nucleoli, and brisk mitotic
activity. The tumor is usually infiltrated by eosinophils and plasma cells.
An in situ precursor lesion is not typically found in association with this
tumor.
3. Adenoid cystic carcinoma is a rare cervical tumor found mostly in post-
menopausal African American women who present with abnormal bleeding
and a pelvic mass. This tumor shows cystic spaces filled with eosinophilic
hyaline material or basophilic mucin surrounded by palisades of epithelial
cells. The tumor architecture may be tubular, cribriform, or solid. Adenoid
cystic carcinoma of the cervix typically shows more cytologic atypia than is
present in its salivary gland counterpart but displays the same tendency for
perineural invasion and local aggressiveness.
4. Mucoepidermoid carcinoma (MEC) is not currently recognized as a distinct
entity in the WHO classification of cervical tumors. Tumors with strik-
ing similarity to the analogous tumor of the salivary gland do occur in
the cervix, howevet; with epidermoid, intermediate, and mucin-producing
cells. Interestingly, it has been shown that when strict histologic criteria are
applied, cervical tumors with mucoepidermoid morphology harbor the same
t(11;19)( q21;p13) as salivary gland MEC (Am] Surg Pathol. 2009;33: 835).
Chapter 34 • Uterine Cervix I 545
This suggests that MEC may actually be an entity distinct from adenosqua-
mous carcinoma in the cervix.
5. Adenoid basal carcinoma. This tumor occurs in a population similar to that
of adenoid cystic carcinoma. It also shows a nested cribriform architecture.
The epithelium, howevet; is composed of more uniform, round to oval,
basophilic cells, often with squamous differentiation, without significant
atypia or increased mitotic activity. There is often associated CIN. Correct
identification of this lesion is important because it is low grade and does
not have aggressive behavior; in fact, some have suggested that the tumor
is more appropriately termed adenoid basal epithelioma (Am] Surg Pathol.
1998;22:965).
E. Neuroendocrine neoplasms. A variety of neuroendocrine neoplasms may occur
in the cervix, although rarely. The classification of these tumors is similar to that
of neuroendocrine tumors of the lung.
1. Carcinoid. These benign tumors are organoid in their architecture and are
composed of small, oval to spindle cells with granular cytoplasm. Mitoses
are rare in typical carcinoid tumors. Immunoreactivity with neuroendocrine
markers synaptophysin, chromogranin A, and neuron-specific enolase is the
rule.
2. Atypical carcinoid. Moderate cytologic atypia and the presence of 5 to 10
mitotic figures/10 high-power fields (HPFs) are sufficient to classify a carci-
noid as atypical; at least small foci of necrosis are often present (Arch Pathol
Lab Med. 1997;121:34). These tumors generally retain the organoid archi-
tecture of typical carcinoids. Their biologic behavior is difficult to assess sys-
tematically due to the subjectivity involved in separating atypical carcinoids
from typical carcinoids and large cell neuroendocrine carcinomas.
3. Large cell neuroendocrine carcinoma. These tumors show frequent vascular
invasion, have higher mitotic activity than atypical carcinoids (> 10 mitotic
figures/10 HPFs s), and show loss of the organoid architecture seen in less
aggressive neuroendocrine tumors. There may be focal adenocarcinoma-like
areas with abundant cytoplasm and large nucleoli. Necrosis is frequent. The
prognosis is poot; similar to that of small cell carcinoma.
4. Small cell carcinoma. Histologically identical to its counterpart in the lung,
small cell carcinoma (also known as high-grade neuroendocrine carcinoma)
is a tumor of variably sized, round to oval to spindle cells with scanty cyto-
plasm, high mitotic activity, nuclear molding, and frequent crush artifact
(e-Fig. 34.18). Necrosis may be extensive. Clinical series are small due to the
rarity of the tumot; but the prognosis is uniformly poor.
F. Mesenchymal neoplasms. A wide variety of sarcomas may be primary to the
cervix, including leiomyosarcoma, embryonal rhabdomyosarcoma, endometri-
oid stromal sarcoma, alveolar soft part sarcoma, and angiosarcoma (e-Fig.
34.19).
G. Mixed epithelial and mesenchymal neoplasms
1. Adenosarcomas are polypoid lesions that microscopically consist of large
papillae of malignant stroma covered with benign endocervical epithelium.
The stroma can have many different appearances; it can consist of plump,
mitotically active spindle cells or more undifferentiated round cells similar to
those of small cell carcinoma. Heterologous sarcomatous elements may be
present, with skeletal muscle, cartilage, adipose, or osseous differentiation.
Prognosis after excision is apparently good, although only small numbers of
cases have been reported.
2. Malignant mixed Mullerian tumor {MMMT), or carcinosarcoma, also presents
as a polypoid mass. In contrast to its more common counterpart in the uter-
ine corpus, the malignant epithelial component is more often squamous or
basaloid, as opposed to glandular. The sarcomatous component is usually
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homologous, with a spindle-cell morphology similar to that of fibrosarcoma.

In limited series of MMMT of the cervix, it appears that the prognosis is bet-
ter than that of MMMT of the uterine corpus.
H. Hematalymphaid neoplasms. Lymphoma of the cervix is usually a part of systemic
disease.
I. Melanoma. Primary melanoma of the cervix is rare. Vaginal bleeding and a
cervical mass is a common presentation. The tumor is usually low stage at
presentation, but the prognosis is dismal. Morphologically, the tumor is similar
to melanomas of other sites, although cervical melanomas have been noted for
a tendency toward a spindle-cell morphology. Melanin pigment is variable from
tumor to tumor. It is important to identify a junctional component in this lesion
if it is to be classified as primary to the cervix; otherwise, a thorough search for
another primary site is indicated.
J. Secondary malignancies. Most metastases to the cervix arise from tumors at
other sites in the reproductive tract. Aside from the uterus and ovary, the gas-
trointestinal tract and breast are the most common origins of cervical metastases.
V. PATHOLOGIC AND CLINICAL STAGING OF MALIGNANCIES
A. American Joint Committee an Cancer (AJCC) and International Federation of Gyne-
cology and Obstetrics (FIGO} criteria. The AJCC criteria for the staging of cervical
cancer with the corresponding FIGO clinical staging categories are outlined in
Table 34.2.
B. Additional information. The final report in any case of malignancy should explic-
itly include all of the information required for assigning a stage as well as other
information of clinical interest not required for staging; Synoptic reports should
be used to uniformly convey this information. For the cervix, the report should
include: (1) the histologic type and grade; (2) the presence or absence of pre-
cursor lesions (either CIN or AIS); (3) tumor size, including depth and width;
(4) whether the malignancy is unifocal or multifocal; (5) presence or absence
of lymphovascular space invasion; (6) presence or absence of vaginal, paracer-
vicaVparametrial, or uterine extension; (7) margin status; and (8) presence or
absence of lymph node or distant metastases.
Cytopathology of the Uterine

Cervix
Michael E. Hull
I. SPECIMEN TYPES
A. Liquid-based preparations are the most widely used modality for cervical screen-
ing (Pap test). A brush is used to sample the cervix and the sample is placed in
appropriate transport fluid for the proprietary system used in the laboratory, be
it ThinPrep (Hologic, Inc., Marlborough, MA) or SurePath (BD Diagnostics-
TriPath, Burlington, NC). Advantages of this modality include a cleaner back-
ground, a monolayer of cells, increased diagnostic sensitivity, and ease of per-
formance of HPV assays.
B. Conventional smears are made by using a spatula and/or brush to sample the
cervix, smearing the endo- and ectocervical samples on a glass slide, and then
immediately fixing the sample. Papanicolaou staining is performed in the labo-
ratory.
II. EXFOLIATIVE CYTOLOGY OF THE CERVIX
A. Squamous cells. A spectrum of squamous cells, ranging from small parabasal
and metaplastic cells with dense cytoplasm and relatively high N/C ratios
(e-Fig. 34.20), to intermediate cells with medium-sized round nuclei with open
chromatin and polygonal cytoplasmic outlines (e-Fig. 34.21), to superficial cells
with polygonal shapes and small pyknotic nuclei (e-Fig. 34.22) is seen in normal
cervical smears and liquid-based preparations.
B. Glandular cells. The classic appearance of endocervical glandular cells is sheets of
regularly spaced cells forming a so-called "honeycomb" pattern (e-Fig. 34.23);
varying degrees of disruption of this architecture herald reactive and neoplastic
changes. Endometrial cells are smaller and form more three-dimensional aggre-
gates in cytology preparations; they are also typically more degenerated than
endocervical cells. Careful attention to nuclear details such as the chromatin
pattern and nuclear contours is required to separate glandular neoplasia from
reactive atypia.
Ill. THE BETHESDA SYSTEM was developed in 1988 with the objective of standard-
izing terminology to promote better communication between the laboratory and
clinicians. It is in its second edition (2001).
A. Adequacy. A statement of adequacy is required. Cellularity should be 5000 cells
for a liquid-based preparation, and this can be reproducibly and quickly esti-
mated with experience and with the use of reference images. Having 75% of
squamous cells obscured by blood or inflammation renders a specimen unsatis-
factory, as does improper labeling or slide breakage.
B. Negative for intraepithelial lesion or malignancy
1. Microorganisms should be reported. Various causes of vaginitis can be
detected cytologically.
Trichomonas vagina/is is seen as a pear-shaped structure, "'30 ~-tm in
diameter. It has a small pale nucleus and red cytoplasmic granules. The flag-
ella are often difficult to appreciate (e-Fig. 34.24).
Bacterial vaginosis is marked by the presence of "clue cells," which are
coccobacilli-coated squamous cells (e-Fig. 34.25). This is reported as "shift
in flora suggestive of bacterial vaginosis."
Actinomyces infection shows filamentous organisms and the "sulfur gran-
ules" described above. This infection is associated with runs.
Candida albicans is responsible for most cases of vulvovaginal candidi-
asis. Although the fungus can be a commensal microorganism, when it is
accompanied by acute inflammation, the infection is usually symptomatic.
Budding yeast and/or pseudohyphae are seen (e-Fig. 34.26).
Herpes genitalis is usually caused by HSV type 2. Nuclear enlargement,
chromatin margination, multinucleation, nuclear inclusions, and nuclear
molding are evident (e-Fig. 34.27).
2. Other nonneoplastic findings
a. Reactive changes/repair in squamous cells include nuclear enlargement (up
to twice the size of an intermediate cell) with or without multinucleation,
smooth nuclear contours, small nucleoli, and mild nuclear hyperchroma-
sia. In reparative change, there is vacuolization of the cytoplasm and some
of the cells may be elongated, clustering together in a streaming pattern.
Repair implies disruption of the epithelium with subsequent increased pro-
liferation. The relative abundance of cytoplasm and lack of true nuclear
atypia differentiate this from neoplastic squamous proliferations (e-Fig.
34.28). Reactive endocervical cells may show even greater nuclear enlarge-
ment, multinucleation, mild hyperchromasia, and prominent nucleoli
(e-Fig. 34.29).
b. IUD. The recognition of run-related changes is particularly important in
cervical smears, as the singly dispersed cells with vacuolated cytoplasm
and large nuclei seen in this reactive condition can mimic adenocarci-
noma. Despite nuclear enlargement, truly atypical nuclei are not seen.
Correlation with the history is key when run-type changes appear.
c. Normal appearing glandular cells' status posthysterectomy may represent

metaplastic change, adenosis, or misplaced fallopian tube remnants. They
are not considered an epithelial cell abnormality.
C. Epithelial cell abnormality
1. Squamous cell
a. Atypical squamous cells Df undetermined significance (ASCUS) have nuclei
2.5 to 3 times the size of an intermediate cell nucleus. There is minimal
nuclear hyperchromasia and a somewhat increased N/C ratio. Reflex HPV
testing is recommended. While the predictive value of an ASCUSIHPV-
positive sample varies with patient age, in general, the risk of the presence
of CIN2 or greater on follow-up of an ASCUS/HPV-positive Pap test is
similar to that of a low-grade squamous intraepitheliallesion (LSIL) Pap
test, that is, 20% to 30% (N Engl] Med. 2007;357:1579).
b. Atypical squamous cells cannot exclude HSIL are approximately the size of
squamous metaplastic cells, with associated nuclear enlargement result-
ing in an appearance approaching that of a high-grade squamous intraep-
itheliallesion (HSIL). They usually lack the severe hyperchromasia and
abnormal nuclear contours of HSIL, however. Colposcopy is indicated
after this interpretation, since 30% of patients will have CIN2 or greater
on follow-up (Am] Obstet Gynecol. 2003;183:1383).
c. Low-grade squamous intraepithelial lesion {LSIL) is a cytologic lesion with
nuclei greater than three times the size of an intermediate nucleus. Despite
the increased nuclear size, N/C ratios are relatively preserved. Irregular
nuclear contours and hyperchromasia are needed to make this interpreta-
tion. Multinucleation and koilocytic halos are common, but not required
(e-Fig. 34.30).
d. High-grade squamous intraepitheliallesion (HSIL) displays cells with severe
nuclear hyperchromasia and contour irregularities, but the nuclei are gen-
erally smaller than those of LSIL. These severely dysplastic cells are singly
dispersed and in syncytia (e-Fig. 34.31).
e. Squamous cell carcinoma has nuclear abnormalities that are equal to or
more severe than those of HSIL, with the addition of prominent nucleoli
(e-Fig. 34.32). An inflammatory/proteinaceous background (the so-called
"tumor diathesis") favors invasive squamous cell carcinoma, although the
diathesis is seen less prominently in liquid-based preparations.
2. Glandular cell
a. Atypical glandular cells show nuclear enlargement, mild nuclear hyper-
chromasia, variable pleomorphism, and increased N/C ratios. Often in
these cases, only incomplete features of adenocarcinoma are present.
Although not always possible, every attempt should be made to state
whether the abnormal cells are endometrial or endocervical in origin.
b. AIS of the endocervix shows glandular groups with nuclear stratification,
crowding, and sometimes a characteristic "feathering." The nuclei are
generally elongated and lack nucleoli (e-Fig. 34.33).
Adenocarcinoma of the endocervix can be detected by cervical cytol-
ogy in ""80% of cases. However, only 22% of endometrial adenocarci-
noma cases are evident in Pap tests (Acta Cytol. 2007;51:47). The cells
of endocervical carcinoma can be arranged singly or in three-dimensional
clusters, and they show clearly malignant nuclear features including pleo-
morphism, hyperchromasia, irregular nuclear contours, and occasional
large nucleoli. Cytoplasmic vacuoles are often present (e-Fig. 34.34). Cyto-
logically, it can be difficult to distinguish endometrial and endocervical
OriginS.
D. Other. The Bethesda System calls for the use of this category in cases showing
endometrial cells in patients ~40 years of age. Clinical correlation is required to
assess the significance of such cells. The absence of a squamous intraepithelial

lesion must also be documented in the report when "other" is utilized.
IV. ANCILLARY TESTING. HPV testing is recommended in most cases of ASCUS. It is
performed on the unused portion of a liquid-based sample. This testing can be done
using the Digene Hybrid Capture 2 assay (QIAGEN, Inc., Valencia, CA), in situ
hybridization (Ventana Medical Systems, Inc., Tucson, AZ), or polymerase chain
reaction-based assays. These assays test for a number of high-risk HPV subtypes.
Although new assays are available that provide the actual genotype of the infecting
virus, the utility of this more specific information has yet to be determined.
A. Markers of HPV integration such as E6 and E7 mRNA assays and p16 immuno-
histochemistry, among others, are under investigation. Their place in cervical
cancer screening in not well defined, but the hope is that positivity for this type
of marker will be more specific than high-risk HPV infection alone for the even-
tual development of a high-grade squamous intraepitheliallesion or invasive
squamous cell carcinoma.
B. Automated screening of cervical cytology specimens is now in wide use, with FDA
approval. The system used depends upon the proprietary liquid-based Pap test
used in the laboratory.
SUGGESTED READINGS
ACOG Committee on Practice Bulletin. ACOG Practice Bulletin no. 109: Cervical cytology screen-
ing. Obstet Gynecol. 2009;114:1409.
Committee on Adolescent Health Care. ACOG Committee Opinion no. 436: Evaluation and
management of abnormal cervical cytology and histology in the adolescent. Obstet Gynecol.
2009;113:1422.
Ganesan R, Ferryman SR, Meier L, et al. Vasculitis of the female genital tract with clinicopathologic
correlation: A study of 46 cases with follow-up. Int J Gynecol Pathol. 2000;19:258.
Lennerz JK, Perry A, Mills JC, et al. Mucoepidermoid carcinoma of the cervix: another tumor with
the t(11;19)-associated CRTC1-MAMI2 gene fusion. Am] Surg Pathol. 2009;33 :835.
Wright TC Jr, Massad LS, Dunton CJ, et al. 2006 American Society for Colposcopy and Cervical
Pathology-sponsored Consensus Conference. 2006 consensus guidelines for the management
of women with cervical intraepithelial neoplasia or adenocarcinoma in situ. J Low Genit Tract
Dis. 2007;11:223.
Vagina
Rao Watson, John D. Pfeifer,
I. NORMAL STRUCTURE. The vagina is derived from the miillerian ducts and is com-
posed of three layers: mucosa, muscularis propria, and adventitia. The mucosa
is composed of squamous epithelium overlying a lamina propria that contains a
rich vascular and lymphatic network with scattered stromal cells that may show
multinucleation.
II. BENIGN CONDITIONS
A. Infectious diseases
1. Vulvovaginal candidiasis is a common condition that predominantly affects
adult women in their second and third decades. Up to 70% of women
will experience at least one episode in their lifetime. Common predis-
posing factors include antibiotic use, steroid use, oral contraceptive use,
immunosuppression, and uncontrolled diabetes. Pruritus, erythema, and
thick white vaginal discharge are the most common symptoms. Histolog-
ically, squamous epithelial hyperplasia with hyperkeratosis and/or parak-
eratosis is seen. Foci of neutrophilic infiltration of the squamous epithe-
lium are commonly present. Candida can be present in the form of bud-
ding yeasts as well as pseudohyphae, highlighted on GMS or other special
stains.
2. Bacterial vaginosis is most commonly found among adult women. It is caused
by Gardnerella vagina/is, a bacillus which usually grows when the vagi-
nal flora shifts toward a more acidic environment. A watery, malodorous
discharge without significant inflammation is a common symptom. Micro-
scopically, the bacteria overgrow and cover the squamous cells, producing
so-called clue cells.
3. Trichomoniasis, a sexually transmitted disease, is caused by Trichomonas
vagina/is, an oval protozoon with flagella. Microscopically, the organisms
are identified by their bluish-pink body, elongated nuclei, and flagella.
4. Herpes simplex infection is a sexually transmitted disease caused by herpes
simplex virus (HSV). Grossly, the virus causes a mucosal ulceration within a
few days to 2 weeks following the exposure. These lesions are highly infec-
tious until crusting, with final scarring occurring within 2 to 3 weeks of
initial symptoms. The majority of cases are caused by HSV-2, and recurrence
is higher with infection by HSV-2 than by HSV-1.
Microscopically, the ulcerated lesions are characterized by epithelial
necrosis with associated degenerated cells containing viral inclusions, best
identified at the periphery of the ulcer. The cells with viral inclusions have
characteristic features including multinucleation and ground glass nuclei with
a rim of chromatin condensation at the nuclear border surrounded by a cyto-
plasmic halo.
5. Actinomyces-like organisms are most commonly seen in women with noncop-
per intrauterine contraceptive devices.
B. Inflammatory diseases
1. Atrophic vaginitis occurs most commonly in postmenopausal women, but
can also occur during the postpartum period. Grossly, the primary finding is
punctate hemorrhage of the vaginal mucosa. Microscopically, the squamous
cells show decreased glycogen due to lower estrogen levels. Atrophy can be
distinguished from vaginal intraepithelial neoplasia (VAIN) by the monotony
551
of the cell population, uniform chromatin, the lack of cytologic atypia, and
the low mitotic rate.
2. Crohn disease can result in rectovaginal fistula formation and is associated
with fibrosis, chronic inflammation, and granulomas. The differential diag-
nosis includes vaginal fistulas of other etiologies including radiation therapy,
perforated colonic diverticulum, or as a complication of hysterectomy.
3. Stenosis, ulceration, and necrosis are well-described sequelae of radia-
tion therapy. Stenosis can also follow severe bullous erythema multiforme
(Stevens-Johnson syndrome).
C. Cysts
1. Miillerian cysts are the most common type of vaginal cyst and can be lined
by endocervical-, endometrial-, or endosalpingeal-type epithelium (e-Fig.
35.1).*
2. Epithelial inclusion cysts are lined by keratinizing squamous epithelium and
filled with white sebaceous and keratinous debris. They most commonly arise
in areas of previous trauma such as episiotomy sites.
3. Mesonephric cyst. Also known as Gartner duct cysts, they are usually located
along the anterolateral wall of the vagina (along the path of the mesonephric
duct). This type of cyst is lined by low cuboidal, nonmucinous epithelium.
4. Bartholin gland cysts are thought to develop from obstruction of the ducts of
Bartholin glands, which normally open on to the vestibule. The cyst lining
varies from squamous to transitional to mucin secreting (e-Fig. 35.2).
D. Adenosis occurs in about 30% of women who were exposed to diethylstilbe-
strol (DES) in utero and is associated with an increased risk of clear cell adeno-
carcinoma (see section on clear cell adenocarcinoma below). Adenosis usually
involves the upper third of the vagina, but the middle third or lower third are
affected in about 10% of cases. Grossly, adenosis presents as a red erythematous
granular lesion. Microscopically, adenosis is defined by the presence of colum-
nar epithelium of endometrial or endocervical type in the vaginal mucosa or
underlying submucosa (e-Fig. 35.3).
E. Endometriosis of the vagina comprises < 10% of cases of pelvic endometrio-
sis. The diagnosis requires the presence of miillerian-type epithelium (most
commonly endometrioid) and endometrial-type stroma. Hemosiderin-laden
macrophages are often present as well. The presence of endometrial type stroma
can be used to distinguish endometriosis from adenosis (e-Fig. 35.4).
Ill. BENIGN NEOPLASMS. The WHO classification of vaginal tumors is given in Table
35.1.
A. Epithelial
1. Squamous papilloma is usually asymptomatic and can occur at any age.
Grossly, it usually presents as a cluster of papillary lesions. Microscopically,
squamous papillomas have a fibrovascular core and are lined by benign squa-
mous epithelium.
2. Fibroepithelial polyps most commonly occur in adult women during their
reproductive years. They occur in the lower third of the vagina and grossly
have a soft and papillary surface. Microscopically, they are composed of squa-
mous epithelium with underlying hypocellular fibrovascular stroma. Atypi-
cal myofibroblasts are common in the stroma, and scattered multinucleated
cells with bizarre atypical nuclei may also be seen (e-Fig. 35.5). Howevet;
rhabdomyoblasts and a cambium layer are not present and mitotic figures
are rare; these features, together with patient age, distinguish fibroepithelial
polyp from sarcoma botryoides.
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3. Condyloma acuminatum is caused by human papilloma virus (HPV) serotypes

6 and 11. Microscopically, it is composed of papillary fibrovascular cores
lined by squamous epithelium with acanthosis, hyperkeratosis, and parak-
eratosis, with associated viral cytopathic effect or koilocytosis characterized
by nuclear enlargement and irregularity, chromatin clumping and hyperchro-
masia, occasional bi- or multinucleation, and perinuclear clearing.
B. Mesenchymal
1. Leiomyoma is the most common benign mesenchymal tumor of the vagina in
adults, with a mean age at presentation of 40 years. Leiomyomas rarely affect
children. The tumor most commonly develops in the submucosa. Grossly, it
consists of a well-circumscribed, firm mass with a white-tan cut surface.
Microscopically, the tumor is composed of fascicles of spindle cells with
elongated uniform nuclei, fine chromatin, smooth nuclear membranes, and
a moderate amount of eosinophilic cytoplasm. Mitotic figures are rare.
2. Genital rhabdomyoma is a rare tumor of the vagina that shows skeletal muscle
differentiation. It affects middle-aged women, and patients usually present
with vaginal bleeding or dyspareunia. Grossly, it is a solid, polypoid to nodu-
lar lesion that creates a bulging mass under the mucosa. Microscopically,
rhabdomyoma is composed of loosely interweaving bundles of spindle cells
with oval nuclei, abundant eosinophilic cytoplasm, and occasional cross-
striations. Nuclear pleomorphism and mitotic activity are absent. Immuno-
histochemical stains for skeletal muscle markers such as desmin, myogenin,
and myo-D1 are positive. Rhabdomyoma can be distinguished from rhab-
domyosarcoma on the basis of the absence of a dense layer of atypical neo-
plastic cells beneath the epithelium, cytologic atypia, and mitotic activity.
3. Angiomyofibroblastoma is a benign tumor that occurs in the vagina and vulva.
Grossly, it has a well-circumscribed outline with a white-tan cut surface,
and can range from 0.5 to 14 em in maximal dimension. Microscopically,
it is composed of fascicles of spindle cells that have abundant eosinophilic
cytoplasm, elongated nuclei, and minimal to no atypia, although scattered
multinucleated cells may be present. Architecturally, the cells form alternat-
ing hyper- and hypocellular areas, with accentuation of the hypercellular
areas around vessels. The absence of red blood cell extravasation and stro-
mal mucin distinguishes this entity from aggressive angiomyxoma. Surgical
excision is the treatment of choice.
4. Deep ''aggressive" angiomyxoma predominantly affects the sacroiliac soft tis-
sue and perineum of women in their fifth decade. Grossly, it presents as a
large mass with a gelatinous, soft cut surface. Microscopically, it is com-
posed of bland spindle cells with delicate eosinophilic cytoplasmic processes
scattered throughout a hypocellular myxoid stroma. Medium to large thick-
walled hyalinized vessels are commonly present; loose fibrillar arrangements
of collagen fibers (so-called myoid bundles) are typically found around the
thick-walled vessels (e-Fig. 35.6). The stromal cells are usually immunopos-
itive for smooth muscle actin (SMA) and desmin.
5. Postoperative spindle cell nodule is a pseudosarcomatous lesion that most
commonly appears at the site of an excision, a few weeks to months after
the surgery. Grossly, it presents as a small friable reddish mass in the vaginal
vault. Microscopically, it is composed of fascicles of spindle cells with stromal
granulation tissue and extravasated red blood cells. Although atypical mitotic
figures can be seen, atypical nuclear cytology is not present.
The differential diagnosis of postoperative spindle cell nodule includes
vaginal leiomyosarcoma. A clinical history of a recent surgery can aid diag-
nosis.
6. Mullerian papilloma is a benign papillary tumor of childhood. It typically
occurs in the upper vaginal wall of children with a mean age of 5 years.
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grades of dysplasia and invasive carcinoma. Progression into higher grades

rnay take several years to a decade.
Grossly, VAIN appears as an exophytic to verrucopapillary lesion. Micro-
scopically, the squamous epithelium shows nuclear atypia (nuclear enlarge-
ment, hyperchromasia, and irregular nuclear membranes) with koilocytosis
and an increased number of mitotic figures. Grading is based on the extent
to which the thickness of the squamous epithelium shows atypia; VAIN 1,
2, and 3 are defined as the loss of maturation of the lower third (e-Fig.
35.7), lower two-thirds (e-Fig. 35.8), and full thickness (e-Fig. 35.9) of the
squamous epithelium, respectively. Clinical management of VAIN 1 is simple
observation of the patient; the preferred treatment of VAIN 2 or 3 is local
excision or laser ablation.
2. Squamous cell carcinoma of the vagina accounts for 85% of vaginal carcino-
mas and occurs most commonly in women between the ages of 60 and 80
years. In younger women, squamous cell carcinoma is usually associated with
HPV infection. Squamous cell carcinoma often metastasizes to the regional
lymph nodes and has a predilection for distant metastasis to lung and bone.
3. Verrucous carcinoma is a variant of squamous cell carcinoma. It is a slowly
growing, well-differentiated tumor with a warty gross appearance. Micro-
scopically, it demonstrates verruciform architecture with minimal nuclear
epithelial atypia, and a pushing rather than infiltrative margin. Local exci-
sion is the treatment of choice. Local or distant metastasis is extremely rare.
4. Clear cell adenocarcinoma rarely occurs in women who do not have a history
of DES exposure. The lifetime risk of developing clear cell adenocarcinoma
in women exposed to DES is about 0.1 %, with a mean age of 20 years.
In women who have not been exposed to DES, clear cell adenocarcinoma
develops in the postmenopausal years around the age of 60. Patients typ-
ically present with bleeding or a grossly visible mass of the cervix or the
vagma.
Histologically, clear cell carcinoma is composed of cells with pleomor-
phic and hyperchromatic nuclei with abundant clear cytoplasm; hobnailing
is often a prominent feature. The malignant cells may form papillary or tubu-
locystic structures. The most common metastatic sites are the regional lymph
nodes and lung.
5. Other epithelial malignancies. Primary adenocarcinoma of the vagina of non-
clear-cell type is rare; most non-dear-cell adenocarcinomas represent metas-
tasis from the endocervix or endometrium, or other sites such as the ovary,
colon, or breast. The non-dear-cell types of adenocarcinoma that most fre-
quently involve the vagina include mucinous, papillary serous, endometrioid,
and adenosquamous (e-Fig. 35.10).
B. Mesenchymal
1. Sarcoma botryoides (a subtype of embryonal rhabdomyosarcoma) is the
most common malignant vaginal tumor in children, usually affecting girls
<5 years. It is commonly located submucosally and grossly appears as grape-
like clusters of tumor that fill (and in some cases protrude from) the vagina.
Microscopically, the tumor is composed of cells with elongated small nuclei
and a moderate amount of bright eosinophilic cytoplasm. For diagnosis,
at least one microscopic field must show the malignant cells forming a
condensed layer (a so-called cambium layer) beneath an intact epithelium
(Pediatr Dev Pathol. 1998;1:550). The tumor cells are immunopositive for
skeletal muscle markers such as actin, desmin, myo-D1, and myogenin. Sur-
gical excision with radiation and chemotherapy is the treatment of choice,
and the prognosis is usually excellent.
2. Leiomyosarcoma is the most common malignant vaginal sarcoma of adults.
Grossly, the tumor is typically a mass of 3 to 5 em in maximal dimension.
Chapter 35 • Vagina I 55 7
Microscopically, the tumor is identical to its counterparts at other sites in the

female reproductive tract (e-Fig. 35.11).
The criteria for distinguishing leiomyosarcoma from smooth muscle
tumors of uncertain biologic potential are not as well defined for vaginal
tumors as for tumors of the myometrium. Current recommendations are
that tumors >3 em in maximal dimension with an infiltrating margin, mod-
erate to marked cytologic atypia, and ;:::5 mitoses per 10 high-power fields
be diagnosed as leiomyosarcoma (Obstet Gynecol. 1979;53:689).
C. Other tumors
1. Malignant melanoma of the vagina is a rare tumor. It most commonly occurs
in postmenopausal women in the lower third of the vagina. Grossly, it can
present as a bulky palpable mass with or without pigmentation. Microscopi-
cally, the cells have the same cytomorphology as the cells of cutaneous malig-
nant melanoma. Immunohistochemical stains for S-100, HMB45, MelanA,
and vimentin are positive in the malignant cells; immunostains for cytoker-
atin are negative. Vaginal melanomas are treated surgically; radiation and
chemotherapy have not proven effective. The prognosis is very poor and the
recurrence rate is high.
2. Metastasis to the vagina from primary tumors of other sites is uncommon. As
noted above, metastasis usually originates from malignancies of the cervix,
endometrium, ovary, colon, and breast.
Vulva
Danielle H. Carpenter, John D. Pfeifer,
I. NORMAL ANATOMY. The vulva or external female genital region encompasses the
mons pubis, labia majora, labia minora, clitoris, and vestibule. The entire vulva
except for the vestibule is covered by keratinized, stratified squamous epithelium.
The epithelium of the vestibule is glycogenated squamous epithelium. The lateral
aspects of the labia majora and the mons pubis contain hair follicles. Sebaceous
glands are present in the labia majora and the perineum. The clitoris is lined by
keratinizing stratified squamous epithelium overlying paired corpora cavernosa
that contain vascular spaces surrounded by nerves.
The urethral meatus, major vestibular glands (Bartholin glands; e-Fig. 36.1), *
minor vestibular glands, paraurethral glands (Skene glands), and vagina all open
onto the vulva. The Bartholin glands are paired glands that open posterolaterally
on the hymenal ring; they are composed of acini lined by cuboidal mucus-secreting
epithelium that drain into a duct that may be lined by mucus-secreting, transitional,
or squamous epithelium, depending on the location from deep to surface. Skene
glands open on either side of the urethral meatus and are composed of acini lined by
mucus-secreting epithelium that open into ducts lined by transitional epithelium.
A. Vulvar biopsies. Vulvar biopsies should be oriented as for skin biopsies (see Chap.
38) and three H&E stained levels examined.
B. Vulvar resections It is helpful to ask the surgeon to orient the specimen with a
diagram or labeled sutures so that orientation can be maintained during process-
ing. The margins of resection should be inked and, depending on the location of
the resection, the periurethral, vaginal, and perianal margins need to be noted.
In cases with an obvious malignant neoplasm, one section per centimeter of
tumor, including the areas closest to the deep margin; lateral margin, and/or
other margins are recommended. In cases where no tumor is observed grossly,
the entire specimen should be submitted. Because many gynecologic oncologists
consider resection for squamous cancer in this area to be adequate only if tumor
is >8 mm from the margin (Cancer. 2002;95:2331), radial rather than shave
margins should be taken of all but the most obviously negative margins so that
the distance from tumor to margin can be measured.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF THE VULVA. Many inflammatory
and neoplastic conditions that affect the skin will also affect the vulva. These are
discussed in the skin chapters (see Chaps. 38 and 39). This section only covers
those conditions for which the vulva is a common site of disease.
A. Inflammation
1. Bartholin abscess presents as a painful swelling in the area of the Bartholin
gland. Microscopically, there is acute inflammation of the Bartholin duct,
glands, and connective tissue, with purulent luminal contents. The etiology
includes Neisseria gonorrhea, Staphylococcus, or other aerobic or anaerobic
organisms. Treatment includes excision, drainage, and appropriate antibi-
otics.
2. Hidradenitis suppurativa presents as painful subcutaneous nodules in areas
containing apocrine glands, particularly the vulva and axilla. Initial changes
558
Chapter 36 • Vulva I 55 9
include acute and chronic inflammation around hair follicles, which progress
to abscess formation, sinus tractformation, and dermal scarring (e-Fig. 36.2).
Treatment may include laser ablation or total excision of the involved area.
3. Crohn's disease may present as vulvar or perianal erythema, ulceration,
abscesses, or fistulas between bowel and vulva or between two different
areas of vulva. Microscopically, there is acute and chronic inflammation of
the deep dermis, often with associated noncaseating granulomas, fistulas, or
sinus tracts (e-Figs. 36.3 and 36.4).
B. Infection
1. Candida infection is often a chronic inflammatory condition of the vulva that
may be associated with diabetes. It often presents as pruritis and clinically
shows areas of redness with thickened, edematous skin. Microscopically,
there is acanthosis with acute and chronic inflammatory cells in the epithe-
lium, and parakeratosis with neutrophils. Often fungal organisms are visible
on H&E stain in the keratin layer; they are easily identified by silver stains.
2. Syphilis is a sexually transmitted disease caused by the spirochete Treponema
pallidum. The primary lesion of syphilis, the chancre, develops in about
half of the women within 3 weeks of infection and is characterized by one
to sometimes multiple painless, dean-based ulcers. The ulcer heals in 2 to
6 weeks without a scar. Secondary syphilis develops within 6 weeks to
6 months and is characterized by the development of a rash on the palms,
soles, and mucosal surfaces, as well as elevated plaques and papules (termed
condyloma lata) on the vulva and mucosal surfaces. On microscopic sections,
the chancre shows epidermal ulceration, dermal acute and chronic inflam-
mation with numerous plasma cells, and severe arteritis. Condyloma lata are
characterized by marked epidermal acanthosis and hyperkeratosis, dermal
inflammation with numerous plasma cells, and arteritis. The organisms may
be detected on Warthin-Starry, Steine.t;, or Dieterle stains; no organisms are
seen in some cases of active infection.
3. Human papilloma virus {HPV) infection. Condyloma acuminatum, also referred
to as genital warts, is the result of sexually transmitted infection caused by
HPV types 11 (75% of cases) or 6 (25% of cases). They present as asymp-
tomatic, usually multiple or confluent, papillary or papular lesions, and may
occur anywhere on the vulva or perianal region.
Microscopically, condylomata of the vulva typically have a fibrovascu-
lar stalk. The epithelium exhibits acanthosis, papillomatosis, hyperkeratosis,
dyskeratosis, and an accentuated granular cell layer (e-Fig. 36.5). Viral cyto-
pathic effect, termed koilocytosis, takes the form of cytoplasmic clearing
around enlarged nuclei with irregular nuclear outlines and clumped chro-
matin (e-Fig. 36.6). Vulvar condylomata usually follow a protracted course.
They may grow rapidly during pregnancy and then regress after delivery.
Small condylomas may be treated with topical agents while large ones are
excised, or treated with laser ablation or cryotherapy.
4. Herpes simplex virus {HSV). Infection with HSV type 2, or less commonly type
1, is typically heralded by feve.t;, dysuria, and severe pain. Painless vesicles
then appear which progress to an intensely painful ulcer. The ulcer typically
heals in about 2 weeks. Microscopically, epithelial ulceration is surrounded
by virally infected keratinocytes that exhibit multinucleation, "ground glass"
nuclear chromatin, or eosinophilic nuclear inclusions (e-Fig. 36.7).
5. Molluscum contagiosum is a sexually transmitted disease in adults caused by
infection with the Molluscum contagiosum poxvirus. The lesions are small,
3 to 6 mm diameter papules with a characteristic central depression or umbil-
ication, and are usually asymptomatic although perianal lesions may be pru-
ritic. Microscopic features (e-Fig. 36.8) include formation of a cup-shaped
papule with marked epidermal acanthosis, and intracytoplasmic inclusions
that are initially eosinophilic but become more basophilic as the lesion ages.
Most lesions regress spontaneously.
C. Noninfectious squamous lesions
1. Lichen sclerosus presents as symmetric plaque-like areas of white, thinned
epithelium that may be superficially ulcerated. In advanced cases, there may
be scarring of involved areas and stenosis of the introitus.
Microscopically, there is typically a band-like lymphocytic infiltrate in
the upper dermis with spongiotic changes to the basal layer of the epider-
mis and loss of melanocytes from the overlying epidermis. Thinning of the
epidermis with flattening of the rete and a zone of collagenous connective
tissue immediately beneath the epidermis is frequently present (e-Fig. 36.9)
and was the basis for the previous nomenclature "lichen sclerosis et atroph-
icus." However, it is now recognized that a spectrum of histologic changes
can be seen in the disease, ranging from a predominantly lichenoid infiltrate
without dermal collagenization to prominent dermal scarring with minimal
chronic inflammation. Treatment involves high-dose corticosteroids. Post-
menopausal women with lichen sclerosus have a small risk of developing
differentiated VIN (see below) and squamous cell carcinoma.
2. Lichen simplex chronicus (formerly "squamous cell hyperplasia") typically
occurs in adults and presents as a localized area of pruritus (] Reprod Med.
2007;52:3). It is thought to be a nonspecific response triggered by a vari-
ety of irritants. Clinically, the area is white or red, with accentuated skin
markings and sometimes areas of excoriation. The characteristic feature on
microscopy is marked acanthosis without atypia, increased mitotic activ-
ity, inflammation, often with features that overlap other specific dermatoses
(e-Fig. 36.10). Hyperkeratosis may be present. The dermis is normal. Treat-
ment includes limiting exposure to irritants, topical corticosteroids, and
antipruritic agents.
D. Cystic lesions
1. Banholin cyst. Obstruction of the Bartholin duct leads to the accumulation of
secretions and the formation of a cystic dilatation of the duct. The epithelium
lining these cysts may be squamous, transitional, or mucinous. Cysts can be
treated by drainage, marsupialization, or excision of the gland.
2. Keratinous cysts occur at any age and typically affect the labia majora. They
are small, measuring just a few millimeters in maximal dimension, and are
filled with white cheesy material without hair. Microscopically, they are lined
by stratified squamous or flattened epithelium. They can be excised if symp-
tomatic.
3. Mucus cysts occur in the vestibule and are lined by mucinous epithelium with
or without squamous metaplasia. They are probably the result of occlusion
of minor vestibular glands.
IV. TUMORS. The WHO classification of tumors of the vulva is presented in Table 36.1.
A. Benign tumors and tumor-like lesions
1. Fibroepithelial polyps are also known as acrochordons or skin tags. They may
be hyperpigmented, hypopigmented, or flesh-colored, and typically occur on
hair-bearing skin. They usually have a papillomatous or pedunculated growth
pattern and a soft cut surface. Microscopically, the epithelium may be thick-
ened with hyperkeratosis, or may be flattened. The stroma contains loose
bundles of collagen and may be edematous. Fibroepithelial polyps are clini-
cally insignificant but can be excised if they are cosmetically unacceptable.
2. Papillary hidradenoma is a benign tumor that originates from apocrine sweat
glands. It presents as a dome-shaped mass, usually <2 em in diameter, arising
between the labium majus and labium minus. The mass may ulcerate and
bleed but is usually asymptomatic. Microscopically, papillary hidradenoma
forms tubules and acini lined by a luminal layer of epithelial cells and an outer
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+ Anocllloodwlhkhon-.sll
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2. Squamous cell carcinoma may be an incidental finding in a resection for

VIN, may develop in the background of lichen sclerosus or an inflammatory
dermatosis, or may develop in women with no history of VIN. Tumors may
be exophytic, endophytic, or plaque-like and may be located anywhere on
the vulva.
On microscopic examination, nests of invasive carcinoma will exhibit
nuclear atypia, increased mitotic activity, and will be associated with a reac-
tive and desmoplastic stroma. A characteristic feature is keratinization in
nests deep in the stroma (e-Fig. 36.18). Tumors may show warty, keratiniz-
ing, verrucous, basaloid, or mixed features.
In addition to the size of the tumor, the thickness of invasive tumor (as
measured from the top of the granular cell layer to the point of deepest
invasion) as well as the depth of invasion (as measured from the dermal-
epidermal junction at the tip of the nearest normal dermal papilla to the point
of deepest invasion) should be recorded. Lymphovascular space invasion is
also an important prognostic feature and should be noted if found. The
staging scheme for vulvar carcinomas is presented in Table 36.3.
The role of sentinel lymph node biopsy in the management of patients
with vulvar squamous cell carcinoma continues to be evaluated (Curr Opin
Obstet Gynecol. 2004; 16:65 ). The intensity of the histopathologic evaluation
(including immunohistochemistry) determines the frequency at which metas-
tases are identified, which is important since even small metastases/isolated
tumor cells are associated with a small but increased risk for the presence
of more extensive metastatic disease (Curr Opin Oncol. 2010;22:481). The
prognostic importance of different sizes and numbers of nodal metastases is
reflected in the current FIGO staging system (see Table 36.3).
3. Melanoma, though rare, is the second most common malignancy of the vulva
after squamous cell carcinoma. Common presenting symptoms include bleed-
ing, a mass, and pain. Vulvar melanomas may be flat or polypoid and are usu-
ally pigmented, often with satellite lesions. Vulvar melanomas very uncom-
monly arise from a nevus.
The clinical and microscopic features of vulvar melanoma are the same
as those of melanomas arising elsewhere (e-Figs. 36.19 and 36.20) and are
covered in detail in Chapter 40. Important features to note are the thickness,
presence or absence of ulceration, histologic pattern, degree of inflammation,
presence of vascular or perineural invasion, and the presence of satellitosis.
Most vulvar melanoma exhibit an acral-lentiginous pattern; however, those
arising on vulvar skin are more likely to be superficial spreading. Treatment
is wide local excision aiming for 1 to 2 em margins, which can be difficult
to obtain in the vulva without compromising vital structures. The prognosis
for vulvar melanoma is poorer than for melanoma of other skin sites.
4. Paget's disease tends to affect elderly women and presents with patchy, ery-
thematous, excoriated areas of vulvar skin and epithelium. Microscopically,
the squamous epithelium is infiltrated by enlarged cells, either individually
or in clusters, that hug the epidermal-dermal junction (e-Fig. 36.21). These
cells have abundant mucinous cytoplasm, large nuclei with small nucleoli,
and often form small glands within the epithelium. These Paget cells may
also involve the adnexal structures.
Immunohistochemistry is very helpful in distinguishing Paget's disease
from melanoma, which may appear morphologically similar. The neoplastic
cells of Paget's disease will be positive for cytokeratin and CEA (e-Fig. 36.22 ),
but negative for HMB 45; melanoma has the opposite staining pattern.
Cases of Paget's disease associated with underlying carcinoma (usually
vulvar adnexal adenocarcinoma, rectal adenocarcinoma, or bladder carci-
noma) have a significantly worse prognosis, so careful gross and microscopic
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Placenta
Phyllis C. Huettner
I. NORMAL ANATOMY. The normal, unfixed term placenta weighs 350 to 550 g,
trimmed of membranes and cord. The placenta consists of three parts: fetal mem-
branes, umbilical cord, and placental disk.
The fetal membranes insert at the edge of the disk and envelop the fetus and
amniotic fluid. Microscopically, they are composed of a cuboidal amniotic epithe-
lium with underlying connective tissue, a chorionic layer (composed of connective
tissue, intermediate trophoblast, and degenerated villi), and sometimes a layer of
decidua (gestational endometrium) (e-Fig. 37.1).*
The umbilical cord is composed of two umbilical arteries (e-Fig. 37.2) and
one umbilical vein (e-Fig. 37.3) surrounded by Wharton's jelly, a paucicellular
connective tissue matrix. Its outer surface is lined by a layer of cuboidal amniotic
epithelium.
The placental disk is typically oval and microscopically composed of chori-
onic villi surrounded by maternal blood in the intervillous space. The chorionic
villi contain vessels of the fetal circulatory tree embedded in mesenchymal stroma
(e-Fig. 3 7.4). A layer of cytotrophoblast encompasses the villous stroma, and this is
surrounded by a layer of syncytiotrophoblast that is in contact with the intervillous
space. The maternal surface of the placental disk, which is adjacent to the uterine
wall, contains variable amounts of fibrin, intermediate trophoblast, and decidua.
The umbilical cord inserts near the center of the placental disk, and branches of
the umbilical cord vessels arborize over the shiny fetal surface of the disk. Micro-
scopically, the fetal surface of the disk is lined by amnion and chorion.
A. Fetal membranes. The fetal membranes should be assessed for completeness. The
presence of green, blue, or brown staining, indicating meconium or hemosiderin
staining, should be noted. The membranes should be inspected for amniotic
bands, nodules of amnion nodosum or squamous metaplasia, and hemorrhage.
The distance from the insertion to the disk edge should be measured as this gives
a rough estimate of where the placenta was implanted in the uterus. A strip of
membranes should be cut from the rupture site to the disk insertion site, one end
grasped by a forceps, and the strip rolled around the forceps; this membrane
roll should be eased off the forceps into formalin. At least one cross-section of
this membrane roll should be examined.
B. Umbilical card. The length of the cord is measured, including any detached
segments. Note should be made of marginal insertion (at the disk edge), or vela-
mentous/membranous insertion (into the membranes). Abnormalities of cord
color (meconium staining) should be noted. Focal abnormalities such as stric-
ture, hematoma, knots, nodules, plaques, or amniotic bands should be noted
and measured. Cross-sections should be made at regular intervals throughout
the cord length, and the number of vessels and the presence of thrombi should be
noted. At least two cross-sections of cord should be examined microscopically,
avoiding the area just above the insertion site where the two umbilical arteries
fuse.
*All e-figures are available on line via the Solution Site Image Bank.
566
Chapter 37 • Placenta I 56 7
C. Disk. The disk should be assessed for completeness and measured in three dimen-
sions. After examining and removing the membranes and cord, the unfixed disk
should be weighed. The fetal surface of the disk, which is covered by amnion
and chorion, is examined for the same abnormalities as the membranes. The
branches of the umbilical cord vessels are examined for lacerations, calcifica-
tions, and thrombi. The maternal surface of the disk is examined for retropla-
cental hematomas, indentations, or other focal abnormalities. The disk is then
sliced at 1-cm intervals, and each slice is examined and palpated. The color,
location (central vs. peripheral), size, texture (finn vs. spongy), demarcation
(whether well circumscribed or ill defined), and number of all focal lesions are
recorded. An estimate of the percentage of the placental parenchyma involved
by each type of process is noted. Any organized blood clot in the container
is measured. At least two sections of central placenta that include fetal and
maternal surfaces as well as sections of focal lesions should be submitted.
D. Multiple gestation. Placentas from twin gestations may have completely sepa-
rate disks, a fused disk with two gestational sacs, or a fused disk and just one
gestational sac (monoamniotic). If present, the dividing membranes should be
inspected. The percentage of placental parenchyma associated with each twin
should be determined. A roll of the dividing membranes should be made as
for the fetal membranes, and at least one cross-section of the roll should be
examined microscopically to confirm the gross and ultrasound impression of
chorionicity. The chorionic plate vessels should be inspected for anastomoses.
In monochorionic or monoamniotic placentas, the type of anastomoses (artery
to artery, artery to vein, vein to vein) should be investigated by air injection
and recorded, keeping in mind that arteries cross over veins. Note should be
made of unpaired large vessels as these likely represent areas of physiologically
important deep artery-to-vein anastomoses.
Ill. DIAGNOSTIC FEATURES OF COMMON DISORDERS OF THE PLACENTA
A. Fetal membranes
1. Meconium. With recent meconium passage the fetal plate and membranes
will be yellowish-green and slimy. With longstanding meconium passage,
the membranes, fetal plate, and even the umbilical cord will be dull
brown. Microscopically, the amniotic epithelium is stratified and tufted with
pyknotic nuclei. There is marked edema between the amnion and chorion.
Macrophages in this area are filled with yellowish-brown, waxy, meco-
nium pigment (e-Fig. 37.5). Pigmented macrophages may also be seen in
the chorion and decidua.
Meconium passage may be the result of neurologic maturity in the fetal
intestines, but may also be associated with chronic in utero hypoxia, or
stressors closer to delivery. Rarely, meconium induces vascular necrosis in
umbilical vessels.
2. Hemosiderin deposition may stain the fetal plate and membranes brown or
green. Often there is old blood clot where the disk meets the fetal membranes.
Circumvallation (see Section III.D) may also be present. Microscopically, the
membranes do not show the epithelial stratification, tufting, and edema seen
with meconium. Membrane macrophages contain refractile pigment that is
positive with an iron stain. Sometimes a layer of hemosiderin is deposited
in the basement membrane beneath the amniotic epithelium (e-Fig. 37.6).
Diffuse chorioamniotic hemosiderosis is an indication of chronic peripheral
separation and is associated with oligohydramnios in the absence of mem-
brane rupture, preterm delivery, and chronic lung disease.
3. Amnion nodosum forms small, grayish-white, discrete nodules or plaques that
may occur anywhere on the cord or fetal membranes but are most common on
the fetal plate near the cord insertion. These nodules, which represent vernix
caseosa, are easy to remove with a cotton swab. Microscopically, they consist
of fetal squamous cells, amniotic epithelial cells, and sometimes fetal hair
(e-Fig. 37.7). Sometimes nodules of amnion nodosum become re-
epithelialized by contiguous amniotic epithelium.
Amnion nodosum is the result of oligohydramnios. It therefore serves as
a marker of conditions such as renal agenesis that may cause decreased fluid
production, and can also alert to possible complications of oligohydramnios
such as pulmonary hypoplasia and limb positioning abnormalities.
4. Squamous metaplasia. Plaques or nodules of squamous metaplasia are present
in nearly every placenta. They are tan-white and may be seen anywhere on
the cord, membranes, or fetal plate of the disk but are most common on the
fetal plate near the cord insertion. Squamous metaplasia is difficult to remove
with a cotton swab. Microscopically, squamous metaplasia in the placenta is
identical to that present elsewhere in the body. Squamous metaplasia is not
clinically significant.
5. Amniotic bands may appear as shredded amnion on the fetal surface of the
placenta or as thin adhesion-like threads connecting one part ofthe fetal plate
to another, connecting the fetal plate to the umbilical cord, or attached to the
fetal digits or other fetal parts. Microscopically, they are composed of fibrous
tissue often with no attached amnion. Amniotic bands are associated with
a wide variety of abnormalities in the fetus including digital amputations
(e-Fig. 37.8), deft lip and palate, and body-wall defects. Characteristically,
the defects are asymmetric, no two cases are identical, and the spectrum of
defects in any given case does not fit into a recognizable genetic syndrome.
Amniotic band syndrome only extremely rarely recurs in a subsequent ges-
tation.
6. Fetus papyraceous. Occasionally, a mummified remnant of an embryo from
much earlier in gestation will be compressed on the fetal membranes
(e-Fig. 37.9), referred to as fetus papyraceous. It may represent an unrec-
ognized twin gestation or may be the result of selective termination of a
higher order gestation.
B. Umbilical cord
1. Length abnormalities. The normal umbilical cord is 55 to 60 em long. Short
cords (<35 to 40 em) occur in about 5% of cords; they are usually associated
with conditions of decreased fetal movement such as amniotic bands, oligo-
hydramnios, body-wall defects, fetal neuromuscular disorders, and arthro-
gryposis. Long cords (>80 em) occur in about 5% of cords; long cords are
associated with an increased likelihood for encirclement around the fetal
neck or other body part, knots, cord prolapse, and marked cord twisting.
2. Single umbilical anery (SUA}. The incidence of SUA (e-Fig. 37.10) is about
1%, and SUA is about 4 times more common in twins. There is a strong
association between SUA and congenital malformations, mortality, and low
birth weight. SUA is likely an acquired defect as the incidence is lower earlier
in gestation; in fact, the absence of the artery may be the cause of associated
malformations. In some cases of SUA, a small, atrophic remnant of the second
artery can be seen (e-Fig. 37.11). SUA is also strongly associated with other
cord and placental abnormalities such as velamentous insertion, marginal
insertion, extrachorialis, and shape abnormalities.
3. Abnormal cord insenion
a. Marginal insenion. In marginal insertion, the umbilical cord inserts at the
edge of the disk (e-Fig. 37.12). This occurs in 6% to 18% of placentas
and is not clinically significant.
b. Velamentous insenion. In velamentous insertion, the umbilical cord inserts
into the fetal membranes (e-Fig. 37.13). This insertion abnormality is seen
in about 1% of placentas. In about 75% of these cases, the vessels branch
within the membranes before the branches insert into the placental disk;
in 25% of cases, the cord vessels run through the membranes without
branching. Because the branches of the umbilical cord are not protected
by Wharton's jelly, they are at risk for compression, thrombosis, and lac-
eration.
4. Umbilical cord knots. About 1% of umbilical cords have a true knot (e-Fig.
37.14), which may be loose or tight. Differences in the diameter and color
of the cord on either side of the knot should be noted. Cords with size
differences, particularly with a dusky appearance between the knot and the
fetus, are likely to be associated with an adverse outcome. The cord on either
side of the knot should be examined microscopically for thrombi, a feature
that suggests a clinically important knot. The knot should be untied in the
fresh state to look for persistent grooving of Wharton's jelly, a feature that
suggests chronic tightening. The fetal mortality rate for umbilical cord knots
is reported to be between 5% and 11%.
5. Umbilical cord coiling. The normal cord has a left-handed twist, a feature
that is thought to increase turgor, preventing compression of cord vessels. A
coiling index can be determined by counting the number of complete turns
divided by the length of the cord (which can be compared to a standard
reference). About 5% of cords will have no twist, a finding that has been
correlated with increased fetal mortality, operative delivery for fetal distress,
abnormal karyotype, preterm delivery, and fetal heart rate abnormalities.
Hypocoiled cords, with a coiling index below the 1Oth percentile, and hyper-
coiled cords, with a coiling index above than 90th percentile, have also been
associated with a variety of adverse outcomes.
6. Umbilical cord stricture is a focal area of cord that is markedly narrowed
with a depletion of Wharton's jelly, fibrosis, and often thrombosis of the
umbilical cord vessels. The most common location for a stricture is the area
adjacent to the umbilicus. There is a high association between cord stric-
ture and stillbirth, especially early in gestation. Most cases also show excess
twisting.
7. Umbilical cord hematomas occur once in every 5 500 deliveries, although small
hematomas have been documented in 1.5% of cases following ultrasound-
guided cord blood sampling. Hematomas nearly always occur in the portion
of cord closest to the fetus and present as a fusiform swelling with a dark,
hemorrhagic color. It is important not to confuse a true hematoma, which
will be obvious at the time of delivery, with blood accumulation in the cord
as a result of clamping, blood drawing, or other manipulation during or after
delivery. Large hematomas have a perinatal mortality rate of 50%, whereas
small ones have very low fetal morbidity or mortality.
c. Circulatory disorders
1. Infarcts are firm and well circumscribed, with one edge usually abutting the
maternal surface of the disk (e-Fig. 37.15). Early infarcts are red whereas
older infarcts are white. Sometimes there is central hemorrhage. Micro-
scopically, there is collapse of the intervillous space, which crowds the villi
together (e-Fig. 37.16). Depending on the age, the trophoblast may be pale
and degenerative (recent) or show little staining with only ghost outlines of
villi (longstanding).
Infarcts are common, occurring in 10% to 25% of term placentas from
normal pregnancies, typically at the periphery. Extensive infarction, infarcts
>3 em, infarcts that occur in the central placenta, and infarcts in the first or
second trimester of pregnancy are clinically significant and often indicate
significant underlying maternal disease such as pre-eclampsia, collagen vas-
cular disease, or a hereditary thrombophilic condition. Infarcts are caused
by an interruption in the maternal blood supplied by a given spiral artery
to an area of placental tissue.
The normal placenta can lose 15% to 20% of the parenchyma with-
out adversely affecting the fetus. However, in placentas that are chroni-
cally underperfused, such as in pre-eclampsia, a lesser degree of infarction
may be clinically significant. Extensive infarction may cause fetal hypoxia,
intrauterine growth restriction, periventricular leukomalacia in preterm
infants, or fetal death.
2. Massive perivillous fibrin deposition. When large amounts of fibrin are
deposited in the placenta, a firm, white or yellow, slightly gritty, ill-defined
mass is formed (e-Fig. 37.17). Often small pockets of red, villous tissue are
interspersed within strands of white fibrin. Microscopically, there is expan-
sion of the intervillous space by eosinophilic fibrinoid material pushing
the villi away from each other (e-Fig. 37.18); clusters of cytotrophoblast
proliferate in the fibrinoid material, and the villi entrapped in this fibri-
noid material become ischemic. The amount of fibrin deposition needed
for diagnosis of massive perivillous fibrin deposition or to be associated
with an adverse outcome for the fetus is not well established. Some studies
have found that entrapment of 20% of the central-basal terminal villi is
associated with adverse outcomes (Arch Pathol Lab Med. 1994;18:698);
others have defined clinically significant fibrin deposition as fibrin extend-
ing from the fetal to maternal surface and entrapping 50% of villi on at
least one slide (Pediatr Dev Pathol. 2002;5:159). Massive perivillous fibrin
is associated with intrauterine growth restriction, periventricular leuko-
malacia in preterm infants, and fetal death, and may recur in subsequent
pregnancies.
3. Maternal floor infarct. The term maternal floor infarct is a misnomer in that
it is a form of fibrin deposition, not infarction. It is defined as perivillous
fibrin deposition surrounding at least one-third of the villi adjacent to the
basal plate, often with extension of fibrin into the underlying decidua. An
alternative definition is that basal villi of the entire maternal floor be encased
in fibrin at least 3 mm thick on at least one slide. As in massive perivillous
fibrin deposition, the villi in maternal floor infarct that are surrounded by
fibrin undergo ischemic changes.
Maternal floor infarct is quite uncommon, seen in far < 1% of placen-
tas. It is associated with stillbirth, intrauterine growth retardation, preterm
delivery, and neurodevelopmental impairment. It may recur in subsequent
pregnancies.
4. Subchorionic fibrin deposition is common and appears as firm, oval, tan-
white, slightly raised plaques of the fetal surface of the placenta, beneath
the amnion and chorion. On cut section, it is laminated and clearly beneath
the membranes but above the villous tissue (e-Fig. 37.19). Microscopically,
sections show layers of blood and fibrin beneath the chorion. Subchorionic
fibrin plaques are not clinically significant.
5. Retroplacental thrombohematomas occur in about 4.5% of placentas. They
are organized blood clots beneath the maternal surface of the placenta that
indent the placental surface. Recent retroplacental hematomas are soft, red,
easily dislodged, and are often seen in the specimen container rather than
adherent to the placenta by the time the placenta arrives in the labora-
tory. Older hematomas are firm, brown, and densely adherent with def-
inite placental indentation (e-Fig. 37.20). Microscopically, retroplacental
hematomas consist of organized blood clot and fibrin, and the underly-
ing placental parenchyma may be infarcted depending on how long the
hematoma has been present (e-Fig. 37.21). Sometimes the villi immedi-
ately beneath the thrombohematoma exhibit villous stromal hemorrhage
in which the vessels are disrupted and the stroma contains extensive red
cells (e-Fig. 37.22).
Retroplacental hematoma is an important cause of stillbirth. Although

retroplacental hematoma and the clinical syndrome of placental abruption
share many of the same risk factors, in only a third of cases with clinically
identified abruption will a retroplacental hematoma be found on placental
examination (likely associated with rapid delivery before clot can form),
and in only a third of cases in which retroplacental hematoma is identified
will there be a history of placental abruption (likely small and clinically
insignificant).
6. Intervillous thrombohematomas are very common lesions, seen in up to 50%
of normal placentas and 78% of placentas from complicated pregnancies.
They are well-circumscribed, round to oval, very firm lesions with a lam-
inated cut surface (e-Fig. 37.23). Recent intervillous thrombohematomas
are red, whereas older ones are white. They are usually located midway
between the fetal and maternal surfaces. Microscopically, they are com-
posed of layers of red cells and fibrin devoid of villi. A thin rim of infarcted
villous tissue may be present at their periphery.
The blood in intervillous thrombohematomas is of both maternal and
fetal origin, and so they serve as markers of fetomaternal hemorrhage. There
is a very good correlation between the number of intervillous thrombohe-
matomas in the placenta and the degree of fetomaternal hemorrhage as
measured by the Kleihauer-Betke test on maternal blood. Fetomaternal
hemorrhage may be associated with fetal anemia, fetal thrombocytopenia,
fetal death, and maternal sensitization to fetal antigens that are not shared.
7. Subamniotic hematomas are liquid collections of blood that pool between
the amnion and chorion of the fetal plate (e-Fig. 37.24). Microscopically,
they may be difficult to demonstrate as the blood drains out once a cut
is made. Subamniotic hematomas are thought to result from trauma to
chorionic plate vessels when traction is placed on the cord during delivery
of the placenta. It is not usually clinically significant because this occurs
after delivery of the infant. Occasionally, subamniotic hematomas are the
result of injury to vessels during amniocentesis to assess lung maturity or
other procedures.
8. Marginal hematomas occur in about 2% of placentas. They are wedge-
shaped collections of blood at the margin of the placenta where the fetal
membranes meet the placental disk (e-Fig. 37.25). Often there is blood clot
beneath the free membranes in this area. Sometimes a thin layer of blood
forms on the adjacent maternal surface of the placenta, but it does not
indent or infarct the placenta and therefore is not clinically significant.
9. Massive subchorial thrombosis (Breus mole) is a very rare condition, occur-
ring in < 1 in 1000 placentas. It is defined as a red thrombus measuring
at least 1 em in thickness immediately beneath the chorionic plate (e-Fig.
37.26). Microscopically, sections show an organizing blood clot. The patho-
genesis of this rare condition is uncertain.
1D. Fetal thrombotic vasculopathy. Occlusion of vessels in the fetal circulatory
system can involve the large umbilical cord vessels, the chorionic plate ves-
sels, or the smaller fetal vessels within the chorionic villi. Because there is
one continuous circulation between the fetus and the placenta during gesta-
tion, thrombotic lesions in the placenta, particularly when extensive, may
serve as a marker for thrombotic or embolic lesions in the circulation of the
fetus itself. The prevalence of fetal thrombotic vasculopathy is not known.
On gross examination, collections of avascular terminal villi appear as
well-circumscribed pale areas of parenchyma of varying sizes that retain the
same spongy consistency as the surrounding placental tissue (e-Fig. 37.27).
Microscopically, these areas appear as villi with dense, eosinophilic, nearly
acellular stroma with an absence of vessels (e-Fig. 37.28). The villi are
normally spaced without collapse of the intervillous space as is seen in

infarcts, a lesion with which avascular terminal villi may be confused
grossly. A second pattern, formerly termed as hemorrhagic endovasculitis
but now referred to as villous stromal-vascular karyorrhexis, is character-
ized by karyorrhexis of fetal cells such as endothelium, stroma, or blood
cell elements (e-Fig. 37.29). In this pattern, the villi are more cellular than in
avascular terminal villi, with degenerating fetal capillaries and fragmented
red cells. Fetal vascular obstruction in the placenta is usually related to
stasis, hypercoagulability, or vascular damage.
11. Chorangiomas are placental hemangiomas. They are found in about 1%
of placentas and are usually small. Typical chorangiomas are well circum-
scribed, red or gray, and have a firmer consistency than the surrounding
parenchyma (e-Fig. 37.30). Sometimes fibrous septae form lobules within
the chorangioma. Microscopically, chorangiomas are composed of small
capillary-type vessels with a few intermixed larger vessels (e-Fig. 37.31).
Occasional cases may be more cellular, show calcification or degeneration,
or exhibit some cytologic atypia. Chorangiomas do not undergo malig-
nant transformation, and most are incidental findings with no clinical con-
sequences. Large or multiple chorangiomas may cause polyhydramnios,
preterm delivery, antepartum bleeding, hydrops fetalis, fetal anemia or
thrombocytopenia, fetal growth restriction, and cardiomegaly. Infants with
placentas containing chorangiomas have a higher than expected incidence
of hemangiomas elsewhere.
12. Chorangiosis is defined as the presence of~ 10 capillaries per terminal villus
in 10 terminal villi in at least three different regions of the placenta (e-Fig.
37.32). Care should be taken to distinguish chorangiosis from congestion
that makes the vessels appear more prominent. Chorangiosis is found in
about 5% of placentas, typically at term. It is associated with congenital
anomalies, maternal diabetes, maternal anemia, smoking, twin gestation,
and delivery at high altitude.
D. Implantation disorders
1. In placenta accreta, the placenta is abnormally adherent. On gross examina-
tion, the placenta is often severely disrupted or fragmented due to attempts
to remove it manually. Sometimes thick areas of gray myometrial tissue will
be visible on the maternal surface. Microscopically, the key feature is an
absence of decidua between villi and myometrium (e-Fig. 37.33). The pres-
ence of fibrin and trophoblast between villi and myometrium is typical of
accreta. Trophoblast is cytokeratin immunopositive, which is a feature that
can be used to distinguish it from decidual cells.
2. Placenta extrachorialis. Usually the fetal membranes insert at the edge of the
disk. In placenta extrachorialis, the membranes insert away from the disk
edge, leaving a portion of the disk uncovered by fetal membranes. There are
two types of extrachorialis, and they are best distinguished on gross examina-
tion. In circummarginate placentation, the junction between the membranes
and the disk is relatively smooth and flat. In circumvallate placentation,
this junction forms a thick, rolled ridge (e-Fig. 37.34). A given placenta may
exhibit partial or complete extrachorial placentation and may exhibit a com-
bination of circummarginate and circumvallate placentation. The percentage
of the disk circumference involved by each type should be recorded.
Circummarginate placentation is not clinically significant. Complete cir-
cumvallate placentation is thought to be caused by chronic abruption or
peripheral separation at the disk edge and is often associated with diffuse
chorioamniotic hemosiderosis.
3. Shape abnormalities. The placental disk is usually oval or round, but a variety
of shape abnormalities may be seen, the most common or important of which
follow.
a. Succenturiate (accessory) lobe. In about 3% to 5% of placentas, a small

portion of placenta (the succenturiate lobe) is completely separated from
the main disk by membranes devoid of underlying villi. The umbilical cord
almost always inserts into the main disk; the branches of the main vessels
that supply the succenturiate lobe are at an increased risk of thrombotic
events.
b. Bilobed placenta.. Occasionally, the placenta will form two distinct lobes
of approximately equal size, usually connected at one edge by villous
tissue. The umbilical cord usually inserts between the lobes. The clinical
significance of bilobed placenta, if any, has not been established.
E. Maternal disease. Many of the most common and clinically important mater-
nal diseases affecting women during pregnancy have the shared feature of low
uteroplacental blood flow, which results in a characteristic set of changes in the
placenta and a growth-retarded fetus.
1. Pre-eclampsia is the most common maternal disease to occur during preg-
nancy, complicating from 2% to 7% of all pregnancies. It is defined as the
development of hypertension with proteinuria or generalized edema after
20 weeks gestation. Eclampsia is diagnosed when seizures occur in the set-
ting of pre-eclampsia. Pre-eclampsia is a leading cause of maternal and fetal
morbidity and mortality.
On gross examination, the placentas of pre-eclamptic women are often
small. Decidual vasculopathy may be present (in the placentas of normal
women, intermediate trophoblast remodels the intramyometrial segments of
the spiral arteries late in the first trimester or early in the second trimester by
replacing smooth muscle and elastic tissue with fibrinoid material, convert-
ing these vessels into flaccid tubes and thereby dramatically increasing the
blood flow to the placenta; in pre-eclampsia, the remodeling of these intramy-
ometrial segments of spiral arteries does not occur). One form of decidual
vasculopathy is absence of this physiologic transformation; this condition
can only be diagnosed in the decidual tissue adherent to the maternal surface
of the placenta. A second form of decidual vasculopathy is acute athero-
sis; in this condition the spiral arteries exhibit fibrinoid necrosis, infiltra-
tion by lipid-laden macrophages, and often a chronic inflammatory infiltrate
(e-Fig. 37.35). Vessels with acute atherosis, while already narrow, often have
superimposed thrombi further reducing the blood flow through them. Acute
atherosis can be diagnosed in the decidual tissue adherent to the maternal
surface of the placenta but is most frequently seen in decidual spiral arteries
in the membrane roll.
The villi exhibit changes related to low uteroplacental blood flow. They
are small with an increased number of syncytial knots (e-Fig. 37.36) and
exhibit a prominent cytotrophoblast layer, increased villous stroma, and a
thickened trophoblastic membrane (n). Placentas from pre-eclamptic women
are more likely to have infarcts, and the infarcts are more likely to be larger
and/or more numerous. There is also an increased incidence of retroplacental
hematomas in the placentas of pre-eclamptic women.
The hemolysis, elevated liver enzyme levels, low platelet count syndrome
(HELLP) and acute fatty liver of pregnancy complicate a subset of pregnan-
cies with pre-eclampsia. Although there is a much higher rate of preterm
delivery, fetal mortality, maternal complications, and maternal mortality
with these two syndromes compared with pre-eclampsia, the placental find-
ings are not significantly different from those of women with pre-eclampsia
alone.
2. Diabetes. The placental findings in diabetes are variable because the duration
and severity of the disease are highly variable. Women with longstanding
diabetes and significant vascular disease may show placental changes sim-
ilar to those seen in pre-eclampsia. The placentas in the majority of cases,
howevet; are larger and heavier than normal. The microscopic findings are
not specific but are nonetheless characteristic. The villi are often edematous
and immature for gestational age. The cytotrophoblast is prominent. There is
irregular thickening of the trophoblastic basement membrane. Chorangiosis,
avascular terminal villi, and SUA are increased in frequency.
Women with diabetes are more likely to deliver stillborn infants or infants
with malformations and/or macrosomia. There is no relationship between
these adverse outcomes and the severity of the placental findings.
3. Maternal thrombophilic disorders. There is increased interest in the relation-
ship between hereditary thrombophilic disorders (such as protein C and
S deficiency, factor V Leiden, and hyperhomocysteinemia) and pregnancy
complications. Although controversial, it appears that various hereditary
thrombophilic conditions, alone and in combination, are associated with an
increased number and larger infarcts, acute atherosis, spiral artery thrombi,
retroplacental hematomas, and fetal thrombotic vasculopathy.
4. Sickle cell disease. The placentas from women with sickle cell disease may
be small and may have an increased number of infarcts. A characteristic find-
ing is the presence of sickled maternal erythrocytes in the intervillous space
(e-Fig. 37.37). Sickled maternal red cells may also be seen in the placentas of
women with sickle cell trait.
F. Multiple gestations
1. 1Jpes of placentation
a. Diamniotic dichorionic. In this type of placentation, the placental disks may
be completely separate or fused. Each fetus is enveloped by its own ges-
tational sac composed of amnion and chorion. The dividing membranes
are thick. Sections show amniotic epithelium from each twin with fused
chorion from both twins (e-Fig. 37.38).
Dizygous (fraternal) twins exhibit diamniotic dichorionic placenta-
tion, but about 25% of monozygous (identical) twins also exhibit this
type of placentation if the blastocyst splits within the first 3 days post-
fertilization. Because diamniotic dichorionic twins do not share vascu-
lar anastomoses, these twins are the least likely to have complications
such as fetal loss, preterm delivery, and twin-twin transfusion syndrome
(TITS).
b. Diamniotic monochorionic placentation. In this type of placentation, the
placental disks are typically fused. Each fetus is enveloped by its own
gestational sac lined by amnion. A single chorionic layer surrounds both
sacs so that the dividing membranes are composed of only fused amnion
from each sac with no intervening chorion (e-Fig. 37.39).
Twins with monochorionic placentation are monozygous. This type of
placentation is seen in about 75% of monozygous twins and results when
the blastocyst splits between 4 and 7 days after fertilization. Twins with
monochorionic placentation usually share vascular anastomoses and are
therefore at risk for complications such as fetal loss, preterm delivery, and
TITS.
c. Monoamniotic placentation. In monoamniotic placentation, the twins
share a gestational sac, and therefore there are no dividing membranes
(e-Fig. 37.40). Twins with monoamniotic placentation are monozygous,
but only 1% of twins are monoamniotic. This type of placentation results
when the blastocyst splits between 8 and 13 days after fertilization.
Monoamniotic twins have a very high rate of complications, with only
50% surviving to term. They share vascular anastomoses, so are at risk
for the associated complications noted above for monochorionic twins.
In addition, because they share the same gestational sac, these twins have
a high rate of umbilical cord accidents.
2. TTTS complicates about 15% of monochorionic twin gestations. It is the

result of a chronic imbalance of blood flow across the two placental circula-
tions. The vascular anastomoses normally seen in monochorionic placentas
may be artery-to-artery, vein-to-vein, or artery-to-vein. The artery-to-vein
anastomoses are usually at the capillary level and are not visible on gross
examination, but are the most important physiologically as they allow blood
to flow in only one direction and therefore can result in a chronic blood
flow imbalance. Twins with artery-to-artery anastomoses are much less likely
to develop TTTS because these anastomoses tend to cancel any circulatory
imbalances that occur. Hemodynamic imbalance may also be affected by the
type of cord insertion (especially a velamentous cord in the donor twin) and
extensive infarction or other abnormalities of the placenta in one twin (with
resultant increased placental resistance).
Often the twins are discrepant in size. The donor twin supplies blood for
both twins and is hypovolemic, oliguric, and oligohydramnic. The donor twin
may be anemic and hypoglycemic, with small and pale organs. The placental
territory of this twin is usually large, bulky, and pale with edematous villi and
increased nucleated red cells in fetal vessels. The recipient twin experiences
circulatory overload resulting in polyuria, polyhydramnios, and eventually
hydrops fetalis; this twin develops heart failure, hemolytic jaundice, and
kernicterus, with heavy and congested organs. The placental territory of this
twin is small, firm, and congested.
G. Infection. Intrauterine infections can have important consequences for the fetus
including abortion, stillbirth, active infection after birth, and long-term sequelae
such as cerebral palsy, blindness, deafness, and learning disabilities. There are
two patterns of placental infection: ascending and transplacental. Ascending
infections are the most common and are typically caused by bacteria. They result
in inflammation of the fetal membranes (chorioamnionitis) and inflammation of
the umbilical cord (funisitis). Transplacental infections are much less common
and may be caused by viruses, protozoa, and some bacteria. The placenta usually
shows chronic and sometimes acute inflammation within the villi (villitis). Most
cases of villitis, however, do not have an infectious etiology but instead represent
villitis of unknown etiology (VUE).
1. Ascending infections are caused by aerobic or anaerobic organisms that travel
through the cervix or uterine soft tissues to the amniotic cavity. Ascending
infections complicate about 4% of term deliveries but a much higher per-
centage of preterm deliveries. Both the mother and the fetus (after about
20 weeks) respond to the infection. Maternal neutrophils emigrate from
vessels in the decidua through the chorion and eventually into the amnion
(e-Fig. 37.41). They also marginate from the intervillous space to the sub-
chorionic fibrin under the fetal plate of the placenta, and eventually emigrate
through the chorion and amnion of the fetal plate. Fetal neutrophils emi-
grate from chorionic plate vessels toward the amnion. They also emigrate
from the umbilical cord vessels, a process termed funisitis (e-Fig. 37.42).
A staging and grading system has been developed to assess the extent and
severity of both the maternal and fetal inflammatory response (Pediatr Dev
Pathol. 2003;6:435).
In term gestations there is a relationship between the time elapsed since
membrane rupture and the likelihood of developing chorioamnionitis; in
preterm gestations it is thought that the chorioamnionitis precedes and con-
tributes to the development of membrane rupture. There is also a relationship
between acute chorioamnionitis, funisitis, and adverse fetal outcome such as
neonatal sepsis, neonatal pneumonia, cerebral palsy, chronic lung disease,
and necrotizing enterocolitis. Some of these complications may be directly
related to infection and others to the effect of prematurity, but the fetal
response to infection that includes release of cytokines and other molecules

(termed the fetal inflammatory response syndrome) also likely plays a role in
pathogenesis. Adverse outcomes are more tightly linked to funisitis, and are
greater for arteritis than phlebitis; funisitis associated with vascular thrombi
has the highest complication rate of all. These associations have provided
the rationale for staging and grading the inflammatory response in the fetal
membranes and umbilical cord as referenced above.
2. Transplacental infections reach the placenta by hematogenous spread from the
mother. They are usually caused by viruses or protozoa such as those of Tox-
oplasma gondii. rubella, cytomegalovirus (CMV), and herpes simplex virus
(HSV) (TORCH) infections, but some bacteria, most notably Treponema
pallidum and Listeria monocytogenes. may also be spread transplacentally.
The tissue response pattern to infections spread transplacentally is villi-
tis. In cases of villitis, there are usually no findings on gross examination,
although occasionally small yellow nodules may be seen. Microscopically,
the villi contain an inflammatory infiltrate, usually composed of only lym-
phocytes and histiocytes but occasionally containing plasma cells and neu-
trophils (e-Fig. 37.43). Sometimes multinucleated giant cells are seen. Villitis
may be necrotizing or nonnecrotizing; necrotizing villitis, in which there is
destruction of the trophoblastic membranes with fibrin deposition causing
affected villi to agglutinate, is most common. This abnormal agglutination,
rather than the inflammation, is the feature that is most easily recognized on
low-power examination. Usually, villitis is randomly distributed throughout
the placenta, but sometimes villitis only involves the basal villi. There are
subtle features that suggest a specific etiology in some cases; some of the
most common of these are detailed below.
a. CMV. On gross examination, the placenta may be small, normal, or
enlarged and pale. The characteristic microscopic features are necrotiz-
ing lymphoplasmacytic villitis, stromal hemosiderin, necrotizing vasculi-
tis, and areas of villous vessel sclerosis. Cases often show areas of active
villitis as well as areas of scarred villi. In about 20% of cases, viral inclu-
sions are seen involving the fetal capillaries, villous stromal cells, or tro-
phoblast (e-Fig. 37.44). Immunohistochemistry, in situ hybridization, and
polymerase chain reaction (PCR) may all be used to confirm a diagno-
sis of CMV in cases with a clinical suspicion or suggestive microscopic
features.
CMV infections are usually acquired in utero and are more commonly
the result of a primary infection rather than reactivation of latent viral
infection. Infected women are usually asymptomatic.
b. HSV infections are typically acquired during delivery through an infected
birth canal and therefore usually do not cause abnormalities in the pla-
centa. Occasionally, howevet; the virus may be transmitted as an ascending
infection, causing acute necrotizing lymphoplasmacytic chorioamnionitis
with viral inclusions in the amniotic epithelium, or acute funisitis. The
virus may also be transmitted hematogenously, giving rise to necrotizing
or nonnecrotizing villitis. Immunohistochemistry and in situ hybridization
may be helpful in confirming infection. Disseminated HSV infection may
cause severe disease or death of a newborn.
c. Parvovirus B19. On gross examination, the placenta is often large forges-
tational age and pale, as would be expected in any condition causing fetal
anemia. Microscopically, the villi are edematous and there are numerous
nucleated fetal red cells in the villous vessels. Many of the red cell pre-
cursors contain eosinophilic intranuclear glassy inclusions with peripheral
margination of the chromatin (e-Fig. 37.45). Immunohistochemistry or in
situ hybridization may be useful in confirming the diagnosis.
Parvovirus causes a mild disease with a rash in children and is usually

asymptomatic in adults. Pregnant women are more severely affected with
a flu-like syndrome and polyarthralgia; most fetuses are unaffected by
maternal infection. Because red cell precursors, endothelial cells, and car-
diac myocytes are specific targets for parvovirus, fetal anemia and eventual
hydrops fetalis can cause fetal death.
d. Human immunodeficiency virus (HIV} is usually transmitted from mother
to child at delivery or through breastfeeding in the postnatal period.
Transplacental transmission is the least common method of spread. The
role of the placenta in promoting or preventing the spread of mv is
unclear. There are typically no gross abnormalities. The microscopic fea-
tures are not specific and are controversial.
e. Syphilis is caused by the spirochete T. pallidum. Placentas from cases of
syphilis may be normal but are often markedly enlarged, bulky, and ede-
matous. Microscopically, many cases show a classic triad of large, hyper-
cellular, immature villi; villous vascular proliferation with perivascular
fibroblastic proliferation and medial hypertrophy; and villitis that is usu-
ally chronic but may be acute, plasmacytic, or granulomatous. However,
this triad is seen in only 43% of cases, although two of the three fea-
tures are seen in another 4 7% of cases. In addition to the triad, some
cases show necrotizing funisitis or lymphoplasmacytic deciduitis. Special
stains for spirochetes may identify organisms, although often the number
of organisms is very low (e.g., one per slide). PCR identifies cases even
when staining is negative.
f. Toxoplasmosis is caused by the protozoal organism T. gondii. and the
placental findings in congenital toxoplasmosis are highly variable. The
placenta may be normal but is often very large and edematous. Micro-
scopically, the villitis is subtle and nonnecrotizing or results in fibrotic villi.
The inflammatory infiltrate is lymphohistiocytic. Occasionally, true gran-
ulomas are present in the inflammatory infiltrate. In addition to villitis,
some cases show a plasmacytic infiltrate in the decidua, chronic chorioam-
nionitis and funisitis, and thrombosis and calcification of the large vessels
of the chorionic plate. The encysted organisms may be present in the cord,
membranes, decidua, or villi but it are not associated with inflammation
and are very difficult to identify. Tachyzoites released from the cysts cause
marked inflammation and necrosis. Immunohistochemistry, immunofluo-
rescence, and PCR may all aid in the diagnosis.
g. Listeria monocytogenes. The pathologic features of listerial infections
differ in several respects from those of other transplacental infections.
The placenta is typically normal on gross examination but occasionally
small, yellowish-white microabscesses can be seen. Microabscesses, which
feature an abundance of neutrophils between the villous stroma and the
trophoblast as well as extensive necrosis, are present microscopically.
Occasionally, palisaded histiocytes and multinucleated giant cells will be
seen. Usually, acute chorioamnionitis is also present. The organism is a
small rod-shaped or curved gram-positive coccus that can be found in
amniotic epithelial cells, and immunohistochemistry may be more sensi-
tive than routine special stains for its identification.
Listeriosis can have devastating consequences for the fetus. It may
cause spontaneous abortion, prematurity, neonatal sepsis, meningitis, and
death. Infections at birth are typically associated with sepsis and death.
3. VUE. In most cases of villitis, an infectious etiology is not identified; these
cases are referred to as VUE. Serologic studies of both the infant and mother
can be used to exclude many infectious causes if clinically indicated. Most
cases of VUE are seen in the third trimester, and the placenta is normal on
gross examination. Microscopically, about 85% of cases are very mild or

mild; most are necrotizing and have a lymphohistiocytic inflammatory infil-
trate. Sometimes there is vasculitis of the stem villus vessels with associated
downstream a vascular terminal villi. The inflammatory cells in VUE are of
maternal origin.
There are two theories about the pathogenesis of VUE. One theory pro-
poses that VUE is a response to an unrecognized infectious agent, although
many cases have been studied with increasingly sophisticated techniques
and no agent has been identified. The other theory proposes that VUE is
an immunologic phenomenon, specifically a host-versus-graft reaction; the
maternal origin of the inflammatory cells, the tendency of VUE to recur, and
the increased incidence of autoimmune diseases in the mother all support
this theory.
In most cases of VUE, the fetus is unaffected. Adverse fetal outcomes
in the form of intrauterine growth restriction, long-term neurologic deficits,
oligohydramnios, abnormal nonstress tests, abnormal pulsed flow Doppler
studies, abnormal biophysical profiles, and perinatal mortality are related to
the severity of the villitis.
SECTION VIII
Skin
Skin: Nonneoplastic
Dermatopathology
Samuel J. Pruden II, Kimberley G. Crone, and
Anne C. Lind
I. NORMAL MICROANATOMY. Microscopically, the skin is composed of three compart-

ments: the epidermis (a keratinizing epithelium), the dermis (a connective tissue
matrix), and the subcutis (a layer of adipose tissue). Specific features and the rel-
ative size of each of these compartments vary with body site and age and reflect
the many functions of the skin. Familiarity with regional anatomical differences
helps recognition of subtle abnormalities, provides a clue to the site of a biopsy
if that information is not provided, and aids in the formulation of a differential
diagnosis that is appropriate to specific anatomic locations.
The epidermis, a stratified squamous epithelium, rests on a normally invisi-
ble basement membrane. The keratinocytes of the basal layer (stratum basale)
have a generative function and are anchored to the basement membrane by
hemidesmosomes. Immediately above the basal layer is the variably thick
"prickle" or "spinous" cell layer (stratum spinosum). This name refers to the
slender eosinophilic processes that extend between adjacent keratinocytes as seen
by light microscopy. These processes correspond to the desmosomes or cytoplas-
mic attachment plaques. Above the spinous layer is the granular layer (stratum
granulosum), which features fine intracytoplasmic basophilic keratohyaline gran-
ules. The granular cell layer is 1 to 3 cells thick and forms a water-tight barrier. The
most mature and outermost cell layer of the epidermis, the cornified layer (stra-
tum corneum), is composed of flat keratinocytes without nuclei. Keratinocytes are
derived from ectoderm and produce type I small acidic (K9-20) and type ll large
neutral-to-basic (K1-8) keratins. Transit time through all layers of the epidermis
is "'28 days.
Cells other than keratinocytes are also present in the epidermis. Melanocytes
originate in the neural crest and are normally located in the basal layer slightly
beneath the basal keratinocytes. They are histologically distinct, having a rounded,
hyperchromatic nucleus as compared with the more elongated nucleus of the basal
keratinocyte. They sometimes show a pericytoplasmic clearing/vacuole. This is an
artifact of fixation and is attributable to their lack of desmosomes. The melanocyte
to basal keratinocyte ratio varies from 1:10 on truncal skin to 1:3 on facial skin,
and the ratio is affected by solar damage/sun exposure as well as anatomic site.
Melanocytes produce melanin, which has an ultraviolet light-protective function.
Melanin is packaged in melanosomes that are exported via slendet; elongated,
dendritic processes that extend between keratinocytes.
Langerhans cells are usually histologically invisible and function as antigen-
presenting cells. They are suprabasal dendritic cells that, when seen in aggregates,
579
580 I SECTION VIII: SKIN
have a coffee-bean shaped nucleus. Langerhans cells are visible only via spe-
cial stains or in aggregates in chronic inflammatory disorders and in Langerhans
cell histiocytosis. Merkel cells are located along the stratum basale and are also
histologically invisible. They have recently been determined to be derived from
progenitor keratinocytes and are presumed to serve in tactile perception.
Small, slender, regularly spaced downward extensions of the epidermis (rete)
divide the superficial dermis into papillae. The papillary dermis, located immedi-
ately beneath the basement membrane, is composed of fine collagen and elastic
tissue fibers and contains the capillary loops of the vascular plexus. The reticular
dermis, with its haphazardly arranged thick collagen bundles and elastic tissue
fibers, is separated from the papillary dermis by the superficial vascular plexus.
Elastic fibers, a component of both the papillary and the reticular dermis, are
usually visible only with the aid of special stains. The dermis provides structural
support and flexibility to the skin.
The dermis also contains adnexal structures, arrector pili muscles, nerves,
and blood vessels. Adnexal structures in the skin include hair follicles and the
eccrine, apocrine, and sebaceous glands. Eccrine glands develop as downgrowths
of the epidermis. They are present as secretory coils in the deep dermis and have
a vertically oriented duct that communicates directly through the epidermis by
way of a pore called the acrosyringium. Alternatively, apocrine glands develop
from the follicular unit and communicate to the surface via a connection through
the follicular infundibulum. Like the eccrine gland they have a deep coil and
a vertically oriented duct, although the apocrine coil has a larger central space
than the eccrine coil and has the classic eosinophilic cytoplasmic apical bleb. Like
the apocrine gland, sebaceous glands are outgrowths of the follicular infundibula.
They form lobules that connect to the follicle via a small duct and are composed of
peripheral basaloid cells and central mature sebocytes with vacuolated cytoplasm.
Hair follicles have varied features depending on the type of hair they produce
and, if they are a terminal hair, whether they are actively growing (anagen), rest-
ing (telogen) or involuting (catagen). The infundibular portion of the follicle is
histologically identical to the epidermis. This portion extends from the epidermal
surface to the sebaceous duct. The isthmus has trichilemmal keratinization and
extends from the sebaceous duct to the insertion point of the arrector pili muscle
(the bulge). The lower segment of the follicle extends from the bulge to the bulb,
the classic ball-and-claw that is seen at the base of every hair follicle.
The subcutis is composed of mature adipose tissue separated into lobules by
fibrous septae. Fully lipidized adipocytes have a slender, crescentic, barely visible
nucleus that has been displaced to the periphery of the cell by the accumulated
lipid. The deep vascular plexus separates the subcutis from the reticular dermis.
II. COMMON DESCRIPTIVE TERMS
A. Acantholysis: Loss of attachment(s) between keratinocytes (e-Fig. 38.1).*
B. Acanthosis: Thickening of the epidermis (e-Fig. 38.2).
C. Bulla: Fluid containing space in the epidermis, > lcm in size.
D. Dyskeratosis: Abnormal keratinization that results in altered eosinophilic cyto-
plasm; individual dyskeratotic cells may be referred to as Civatte or colloid
bodies (e-Fig. 38.3).
E. Epidermotropism: Migration of malignant cells into the epidermis (e-Fig. 38.4).
F. Exocytosis: Migration of benign, nonepithelial cells into the epidermis, com-
monly seen in association with spongiosis (e-Fig. 38.5).
G. Hypergranulosis: Thickening (increased number of layers) of the granular layer
(e-Fig. 38.6).
H. Hyperkeratosis: Thickening of the stratum corneum.
Chapter 38 • Skin: Nonneoplastic Dermatopathology I 581
I. Orthokeratosis: Appropriately mature stratum corneum composed of superfi-

cial keratinocytes without nuclei. Seen in a characteristic loose "woven,. (bas-
ketweave) pattern on nonacral skin, and densely compact on the acral skin of
the palms and soles (e-Fig. 38.7).
J. Parakeratosis: Abnormally retained keratinocyte nuclei in the stratum corneum
(e-Fig. 38.8).
K. Spongiosis: Fluid/edema creating a space between adjacent cells in the stratum
spinosum, which makes the desmosomes appear prominent (e-Fig. 38.9).
L. Vacuolar change: Clearing of basal keratinocyte cytoplasm secondary to
inflammation at the epidermal-dermal junction (e-Fig. 38.10).
Ill. GROSS EXAMINATION AND TISSUE SAMPLING. The skin biopsy/excision is generally
received in the laboratory in a fixative such as 10% formalin. If special studies
are required, a nonfixative preservative (e.g., Michel's medium) is required. The
gross description should include all pertinent information, including tissue size
in centimeters (length x width x thickness), presence or absence of epidermis,
color, presence or absence of hair (especially if from the scalp), and alterations to
the epidermal surface (including documentation of the dimensions, color, and dis-
tance to the nearest margin of discrete lesions). All surfaces, except the epidermis,
must be inked before sectioning. Avoiding the use of black ink facilitates inter-
pretation of commonly used special stains, immunostains, natural pigments, and
some exogenous pigments. If the clinician has provided orientation for specific
margin identification, inking with two or more colors is required.
Shave or punch biopsies with a greatest epidermal dimension of <0.3 em
are submitted for processing without sectioning. Specimens with a greatest epi-
dermal measurement of at least 0.4 em are sectioned vertically through the epi-
dermis resulting in pieces of relatively uniform thickness (""0.2 to 0.3 em thick)
(Figs. 38.1 and 38.2). As seen in Fig. 38.1, if an epidermal lesion is present, sec-
tioning that will best represent the lesion and its relationship to the nearest margin
is optimal. Biopsy tissue is otherwise sectioned along the longest epidermal axis,
Punch Biopsy
J lll
11111
,, ,cut
,
,,
Demonstrate
nearest margin
Figure 38.1 Gross processing
of a punch biopsy.
Shave Biopsy
Figure 38.2 Gross processing of a shave biopsy.
thus maximizing microscopic visualization (which is particularly important when

incisionaVwedge biopsies are performed).
Punch biopsies of the s~p for evaluation of alopecia are sectioned horizon-
tally (every 0.2 to 0.3 em) to permit evaluation of follicular density and architec-
ture at various tissue levels (Fig. 38.3). One plane of section should separate the
tissue at the level of the deep reticular dermis. The two pieces of tissue are placed
with each superficial surface down in the tissue cassette and are subsequently
embedded with this same orientation. This allows sections that include complete
sequential discs that include en face sections of all of the follicles in the specimen.
If embedded and sectioned appropriately, the superficial sections will also include
a peripheral rim of epidermis and basement membrane zone. The deeper levels
will highlight the infundibulum and isthmus; the lower segment will highlight the
hair bulb.
Elliptical biopsies and excisions are approached, in genera~ in a similar fash-
ion. If an ellipse is oriented by a suture or any other means, inks of different
colors are applied to the two long margins to allow specific margin identification
under the microscope. Sections of an ellipse should be taken at regular intervals
of 2 to 3 mm to allow a reasonable assessment of the true margin. Laboratories
vary in their handling of the tip ends of ellipses; the recommended method is
Punch Bio~psy
for Alopecia
cut
cut
Figure 38.3 Gross processing of

an alopecia biopsy.
Chapter 38 • Skin: Nonneoplastic Dermatopathology I 58 3
Unoriented Ellipse Oriented Ellipse
A4 A3 A2 A4 A3 A2
cut cut cut cut
cut cut cut cut
Figure 38.4 Gross processing of an elliptical excision specimen.
illustrated in Figure 38.4. To prevent embedding errors, cassettes should never be

overcrowded. Sponges can help prevent distortion of the tissue during process-
ing; for example, submit punch biopsies on one sponge, and lay the pieces of a
shave biopsy flat between two sponges. If alternative embedding is required, as
for alopecia biopsies, dear instructions to the histology technicians are helpful in
ensuring appropriate sections.
Frozen sections may be helpful in evaluating margins of cutaneous carcino-
mas and may (rarely) be requested if a life-threatening condition (such as toxic
epidermal necrolysis [TEN]) requires tissue diagnosis prior to treatment. Frozen
sections for diagnosis or margin examination of melanocytic neoplasms are never
indicated and may compromise diagnosis on the basis of subsequent permanent
sections. The majority of dermatopathology diagnoses are best made on ade-
quately processed, fixed tissue.
IV. INFLAMMATORY DERMATOSES
A. Lichanoidiintarfaca: Characterized by basal keratinocyte damage; may have a
band of lymphocytes in the upper dermis (lichenoid) or show only vacuolar
alteration of basal keratinocytes and dyskeratosis (interface).
1. Lichen planus
a. Clinical: Multiple, flat topped papules and plaques, pruritic, sometimes
featuring superficial white lines (Wickham's striae), most common in
adults, can involve skin, hair, nails, and mucous membranes.
b. Microscopic: Compact hyperkeratosis, acanthosis, band of lymphocytes
at the epidermal-dermal junction, dyskeratotic keratinocytes in (Civatte
bodies) and under (colloid bodies) the epidermis, rete with a "saw-tooth"
pattern, melanophages {e-Fig. 38.11).
2. Lichen planus-like keratosis
a. Clinical: A single red, scaly plaque on sun-damaged skin (arms and
chest/shoulders); the clinical impression is frequently that of a basal cell
carcinoma or other nonmelanoma skin cancet.
b. Microscopic: Basketweave orthokeratosis with patchy parakeratosis,
obscuring band of lymphocytes at the epidermal-dermal junction, flat-
tened/atrophic epidermis sometimes, vacuolar alteration, dyskeratosis,
melanophages (e-Fig. 38.12A and B).
3. Erythema multiform• {EM)IStavans-Johnson syndrome (SJS)nEN
a. Clinical: EM is a clinical reaction pattern characterized by symmetric,
targetoid lesions seen best on the palms and soles; it may involve the
mucosa, and it is associated with multiple "triggers, (most commonly
HSV). SJS and TFN always have mucosal ulcerations with cutaneous
sloughing. SJS involves <10% of body surface area (BSA) whereas TFN
>30% BSA.
b. Microscopic: Vacuolar alteration of basal keratinocytes, aggregates of

dyskeratotic keratinocytes throughout the epidermis, possible subepi-
dermal bulla with full-thickness epidermal necrosis, lymphocytes in the
epidermis and around the superficial vessels (e-Fig. 38.13).
4. Lupus erythematosus
a. Clinical: Lupus can involve the skin only or involve multiple organ
systems. Skin findings, which vary with the clinical disease, include a
"butterfly" -shaped erythematous rash on the cheeks and nose, scaly ery-
thematous lesions on sun-exposed skin and/or scarring, and alopecic
plaques.
b. Microscopic: The spectrum of changes includes epidermal atrophy, fol-
licular plugging, vacuolar alteration of basal keratinocytes, scattered
dyskeratotic keratinocytes, thickened basement membrane, superficial
and deep perivascular and periadnexal infiltrate of lymphocytes, some
extravasated erythrocytes, and interstitial dermal mucin (e-Fig. 38.14).
Direct immunofluorescence of a well-established lesion usually shows
linear-granular immunoglobulin G (IgG), IgM, and/or complement
deposition at the basement membrane.
5. Graft versus host disease (GVHD}
a. Clinical: GVHD may be hyperacute (rarely, <14 days from transplant),
acute, or chronic. Acute cutaneous GVHD is often a morbilliform erup-
tion with sudden onset that favors acral sites (palms, soles, head, and
neck) and may occur before gastrointestinal or hepatic manifestations.
Chronic GVHD may clinically resemble lichen planus (lichenoid GVHD)
or morphea (sclerodermoid GVHD).
b. Microscopic: Acute GVHD shows epidermal atrophy with a spectrum
of changes at the basement membrane zone (see the section on grad-
ing). Because the process is mediated by lymphocytes, lymphocytes in
the epidermis and superficial dermis are required for the diagnosis. The
histologic changes of acute GVHD are indistinguishable from those seen
in response to marrow engraftment (cutaneous eruption of lymphocyte
recovery). Grading of acute GVHD:
Grade 0: No histologic alteration by skin biopsy.
Grade 1: Vacuolar alteration.
Grade 2: Dyskeratotic keratinocytes in the epidermis and/or adnexal
epithelium (e-Fig. 38.15).
Grade 3: Clefting of the epidermis from the dermis.
Grade 4: Complete separation of the epidermis from the dermis.
Grades 1 to 4 require the presence of lymphocytes in the dermis
and epidermis (the number of lymphocytes may vary); eosinophils
may also be present and do not necessarily support the diagnosis
of a reaction to a medication.
B. Psoriasiform: Characterized by regular elongation of the rete.
1. Psoriasis
a. Clinical: It has diverse clinical forms (vulgaris, pustular, erythrodermic,
guttate), the most common being psoriasis vulgaris, which presents as
erythematous plaques with a thick scale on the extensor surfaces (knees,
elbows). Guttate psoriasis is often more acute and has many raindrop-
like, mildly scaly papules.
b. Microscopic: Confluent parakeratosis, absent granular cell layer, regular
elongation of the rete, neutrophils in the stratum corneum (Munro's
microabscess) and stratum spinosum (spongiform pustule of Kogoj),
widely dilated papillary dermal vessels, increased mitoses, and a
lymphocytic infiltrate in the dermis characterize psoriasis vulgaris

(e-Fig. 38.16). Guttate psoriasis has mounds of parakeratosis, but lacks
the regular acanthosis of psoriasis vulgaris.
2. Lichen simplex chronicus
a. Clinical: Thick, hyperkeratotic (and sometimes hyperpigmented)
plaques, commonly on the back of the neck, extensor forearms, and
legs, or genital region, caused by chronic rubbing or scratching.
b. Microscopic: Compact orthokeratosis, a thickened stratum granulosum,
thick and elongated rete. The papillary dermis characteristically has col-
lagen fibers that run toward the skin surface (so-called vertical streaks).
The low-power impression may be that of acral-type skin. Acral-type
epidermis when paired with dermal hair follicles points to a diagnosis of
lichen simplex chronicus (e-Fig. 38.17).
3. Other. Other entities with a psoriasiform pattern include dermatophytosis,
pityriasis rubra pilaris, mycosis fungoides, and chronic spongiotic derma-
titis.
C. Spongiotic: Characterized by expanded space between adjacent keratinocytes
of the spinous layer. Acute spongiosis has spongiosis and vase-shaped vesicles
but no or minimal parakeratosis; subacute spongiosis has definitive paraker-
atosis; chronic spongiosis has psoriasiform hyperplasia and areas of compact
orthokeratosis mixed with parakeratosis.
1. Eczema/atopic dermatitis/allergic or irritant contact dermatitis
a. Clinical: Erythematous scaly patches or plaques, variable distribution,
any age/gender; age/gender and distribution vary with the inciting agent;
atopic dermatitis in children is classically periflexural.
b. Microscopic: Parakeratosis, spongiosis, variable acanthosis, and a super-
ficial perivascular inflammatory infiltrate that may be purely lymphocytic
or may have lymphocytes mixed with other inflammatory cells such as
eosinophils; the degree of change depends on the chronicity of the lesion.
2. Pityriasis rosea
a. Clinical: Scaly erythematous patches/plaques; classically distributed in a
"Christmas tree-like" pattern on the back; less common patterns exist,
including lesions on the extremities (inverse pityriasis rosea).
b. Microscopic: Mounds of parakeratosis, irregular acanthosis, patchy
spongiosis, and a mild superficial perivascular lymphocytic infiltrate
with extravasated erythrocytes. Prominent exocytosis of lymphocytes
is occasionally present and erythrocytes may extend into the epidermis.
(e-Fig. 38.18).
3. Eosinophilic spongiosis
a. Clinical: Varied, depending on etiology. Etiologies include allergic con-
tact dermatitis, arthropod bite reactions, drug eruptions, the urticarial
phase of pemphigus or pemphigoid, and the first stage of incontinentia
pigmenti.
b. Microscopic: Spongiosis with exocytosis of eosinophils; the degree of
either may vary, and the epidermis may be acanthotic (e-Fig. 38.19).
4. Other. Other entities that have a spongiotic tissue pattern include early and
partially treated psoriasis and mycosis fungoides.
D. Vesiculobullous: Characterized by a vesicle or bullae.
1. Bullous pemphigoid
a. Clinical: Tense blisters on an erythematous base, may be pruritic, may
be generalized, usually in the elderly. Variants include pemphigoid ges-
tationis, which occurs in pregnancy, and cicatricial pemphigoid, which
has mucosal involvement and resolves with mucosal scarring.
b. Microscopic: Subepidermal vesicle/bulla with a dermal infiltrate of
eosinophils. A variant without significant inflammation also occurs
(e-Fig. 38.20). Direct immunofluorescence shows linear deposition of

IgG and C3 at the basement membrane zone. Collagen IV can be demon-
strated at the floor of the blister by immunoperoxidase or immunofluo-
rescence stains.
2. Pemphigus vulgaris
a. Clinical: Painful oral erosions and flaccid cutaneous blisters that extend
easily with lateral pressure (Nikolsky's sign).
b. Microscopic: Suprabasilar acantholysis without dyskeratosis (e-Fig.
38.21A and B). Direct immunofluorescence shows intercellular IgG and
complement. Indirect immunofluorescence may also be useful for diag-
nosis and to follow response to therapy.
3. Porphyria cutanea tarda
a. Clinical: Blistering of sun-exposed skin, especially hands and face; milia
(small subepidermal cysts) may develop. Patients may have hypertri-
chosis.
b. Microscopic: Subepidermal blisters with minimal inflammation. The der-
mal papillae extend into the blister floor (festooning) (e-Fig. 38.22). The
thickened vascular basement membrane(s) can be highlighted by periodic
acid-Schiff (PAS) stain.
4. Epidermolysis bullosa acquisita
a. Clinical: Blister formation at sites of trauma, generally in adults.
b. Microscopic: Subepidermal bulla with a sparse inflammatory infiltrate.
Direct immunofluorescence shows basement membrane linear deposi-
tion of IgG and complement as in bullous pemphigoid; however, colla-
gen IV will be seen at the roof of the blister by immunoperoxidase or
immunofluorescence stains.
5. Dermatitis herpetiformis
a. Clinical: Highly pruritic, small vesicles and/or excoriations on the exten-
sor surfaces (elbows, knees, and buttocks/sacrum) associated with gluten
sensitivity.
b. Microscopic: Neutrophils in the dermal papillae with a small subepider-
mal deft (e-Fig. 38.23). Direct immunofluorescence shows granular IgA
at the dermal papillae.
E. Granulomatous: Characterized by a variable number and arrangement of epithe-
lioid histiocytes.
1. Necrobiotic granuloma
a. Granuloma annulare
i. Clinical: Clinically variable; lesions may be localized or generalized,
annular and erythematous, papules, plaques or patches, subcutaneous
nodules or perforating.
ii. Microscopic: Epithelioid histiocytes palisade around dermal collagen
with nuclear dropout; there may be an associated lymphocytic infil-
trate, and interstitial mucin may be increased in the center. The inter-
stitial variant has less definitive architectural features and may appear
at scanning power as a vaguely patterned, hypercellular dermis. Sub-
cutaneous granuloma annulare has a tumor-like presentation with
central necrosis with mucin, palisaded histiocytes, and a hypervascu-
lar rim containing chronic inflammatory cells (e-Fig. 38.24).
b. Necrobiosis lipoidica
i. Clinical: A chronic plaque with a depressed yellow center and an ery-
thematous, raised rim/border, usually pretibial; female predilection;
may be associated with diabetes.
ii. Microscopic: Dermal collagen is fibrotic; layers of fibrosis alternate
with zones of histiocytes and plasma cells. The abnormalities fill the
dermis and may extend into the septae of the subcutis (e-Fig. 38.25).
2. Sarcoidal granuloma (sarcoidosis)

a. Clinical: Highly variable, but with a predilection to areas of cutaneous
injury/scars; up to one-third of patients with sarcoidosis will have skin
lesions. Presentations include annular lesions, areas of hypopigmenta-
tion, ichthyosis-like changes, alopecia, subcutaneous nodules, and ery-
thema nodosum.
b. Microscopic: Naked, noncaseating granulomata are classic. The type of
dermal granulomatous inflammation is highly variable in patients with
documented systemic sarcoidosis (e-Fig. 38.26).
F. Vasculopathic: Characterized by damaged/incompetent endothelial cells. This
broad category includes vascular damage mediated by neutrophils, lympho-
cytes, eosinophils, or histiocytes; vascular occlusive disease related to thrombi,
emboli, or vascular deposits of calcium; noninflammatory vascular damage;
urticaria; and neutrophilic dermatoses.
1. Acute vasculitis (leukocytoclastic vasculitis)
a. Clinical: Palpable purpura, generally on the extremities. Clinical variants
include Henoch-Schonlein purpura, hemorrhagic edema of childhood,
urticarial vasculitis, and mixed cryoglobulinemia.
b. Microscopic: Involves the vessels at the interface of the papillary
and reticular dermis (the superficial vascular plexus) with perivascu-
lar neutrophils, nuclear debris, fibrinoid necrosis of the vessel walls,
and extravasation of erythrocytes. Direct immunofluorescence may be
positive (e.g., Henoch-Schonlein purpura) for IgA and complement
(e-Fig. 38.27).
2. Granuloma faciale
a. Clinical: Reddish brown plaque(s) on the face.
b. Microscopic: A dense, mixed inflammatory infiltrate separated from the
epidermis by a grenz (uninvolved) zone; eosinophils, neutrophils, lym-
phocytes, mast cells, and plasma cells fill the superficial to mid papillary
dermis; there are associated classic dilated, thin-walled blood vessels
(e-Fig. 38.28).
3. Lymphocytic vasculitis
a. Clinical: Highly varied; diverse clinical conditions ranging from pri-
mary dermatoses, morbilliform viral exanthema, to connective tissue
diseases; all have a lymphocytic vasculitis as all or part of their histo-
logy.
b. Microscopic: Superficial perivascular lymphocytes arranged tightly
around the vessels with plump endothelial cells and with variable num-
bers of extravasated erythrocytes; there is no fibrinoid necrosis or leuko-
cytoclasis.
4. Vasa-occlusive vasculopathies
a. Clinical: Varied presentations include purpura, livedo reticularis and
ulcer/infarct.
b. Microscopic: Occlusion of any of the vessels in the cutaneous vasculature
with cellular thrombi (in Coumadin necrosis, heparin necrosis, dissem-
inated intravascular coagulation, TIP, heritable disorders of coagula-
tion) or acellular/hyaline thrombi (essential cryoglobulinemia). Choles-
terol emboli may be seen after instrumentation; marrow emboli may be
seen after trauma; vascular calcification can lead to vascular occlusion if
extreme (calciphylaxis).
5. Neutrophilic dermatosis (Sweet's syndrome)
a. Clinical: Erythematous plaques, abrupt onset, tender and nonpruritic;
most common on the head, neck and upper extremities; accompanied by
fever/constitutional symptoms and leukocytosis; up to 20% associated
with malignancy.
b. Microscopic: Diffuse superficial dermal infiltrate of neutrophils; presence

or absence of vasculitis and/or folliculitis is controversial. There may be
associated marked papillary dermal edema leading to pseudobullae.
6. Urticaria
a. Clinical: Wheals that are pruritic and generally last <24 hours, at any
body site. Persistence for >24 hours, burning rather than itching, and
resolution of a lesion with a purpuric macule suggest a diagnosis of
urticarial vasculitis.
b. Microscopic: At low power may appear histologically normal; sparse
perivascular and interstitial infiltrate of neutrophils, eosinophils, lym-
phocytes, mast cells, and plasma cells without extravasation of erythro-
cytes or necrosis. Dermal edema may be noted, and there may be neu-
trophils in the vascular lumina (e-Fig. 38.29).
7. Pigmented purpuric dermatosislcapillaritis
a. Clinical: Multiple clinical variants.
i. Schamberg disease -multiple, small, "cayenne-pepper" -like macules
sprinkled on the lower extremities, usually transient and self-limited.
Most common in young adults.
ii. Lichen aureus- solitary or multiple, golden-brown plaques on trunk
or extremities; most common in young adults; may persist for
years.
b. Microscopic: Lymphocytes surround the capillaries of the papillary der-
mis, with scattered extravasated erythrocytes; an iron stain will highlight
siderophages. Some vacuolar alteration, as seen in lichenoid dermatitis,
may be present. All variants have similar histology and are best distin-
guished by their clinical presentation (e-Fig. 38.30).
8. Noninflammatory purpura
a. Clinical: Purplish/erythematous discoloration of a small (petechia) or
large (purpura/ecchymosis) area of skin at any body site. No age/gender
predilection.
b. Microscopic: Erythrocytes in the dermis without associated inflamma-
tion; amount/degree varies. In senile purpura, there are telangiectatic
blood vessels and there is usually severe solar elastosis (e-Fig. 38.31).
G. Panniculitis: Characterized by inflammatory changes to the septa or lobules of
the subcutaneous adipose tissue.
1. Erythema nodosum (Septal)
a. Clinical: Single or multiple, painful, erythematous nodules on the ante-
rior lower leg/shin, although other sites can be involved; predilec-
tion for young adults. This is a cutaneous reaction that is associated
with multiple systemic diseases and medications, including inflamma-
tory bowel disease, sarcoidosis, infections (viral, bacterial, and rick-
ettsial), leukemia/lymphoma, antibiotics, salicylates, and oral contra-
ceptives.
b. Microscopic: Adipose tissue septa are affected, and the lobules are rela-
tively spared. Early lesions have neutrophils in the septa, and late lesions
have variable numbers of lymphocytes, multinucleate giant cells, histio-
cytes, and eosinophils; septa become fibrotic over time (e-Fig. 38.32).
2. Erythema induratumlnodular vasculitis (Lobular)
a. Clinical: Single or multiple erythematous nodules on the posterior lower
leg/calf; any age/gender may be affected.
b. Microscopic: Adipose tissue lobules are infiltrated by histiocytes in well-
to poorly formed granulomata; lymphocytes, neutrophils, and plasma
cells may be present. A mixed cellularity vasculitis is usually seen,
and some vessels of the subcutis can have the classic alterations of an
acute/necrotizing vasculitis (e-Fig. 38.33A and B).
3. Subcutaneous fat necrosis of the newborn (Lobular}

a. Clinical: Firm, sometimes nodular areas in body sites associated with
increased adipose tissue (shoulders/buttocks/cheeks) of an otherwise
healthy newborn infant. Some infants develop acute or delayed hyper-
calcemia.
b. Microscopic: Adipose tissue lobules with numerous multinucleate histio-
cytes that contain classic, radiating, needle-like inclusions; lymphocytes
and histiocytes may also be present (e-Fig. 38.34).
4. Calcifying panniculitis (Lobular)
a. Clinical: Highly varied depending on the underlying systemic pathology
including disorders of coagulation, peripheral vascular disease, localized
trauma/inflammation, and calciphylaxis of renal failure.
b. Microscopic: Stippled basophilic calcium deposits in and around vessels
that range in size from capillaries to arterioles, with lipophages, variable
inflammation, and variable erythrocyte extravasation.
5. Membranous lipodystrophy (Lobular)
a. Clinical: Varied disease processes are associated with this histologic alter-
ation. It is most commonly seen in lipodermatosclerosis (sclerosing pan-
niculitis) which manifests as indurated plaques of the inferior lower
extremities resulting in an "inverted champagne bottle" clinical appear-
ance.
b. Microscopic: Small cysts in the lobules of the subcutis that have a
complex eosinophilic cuticle with frond-like excrescences (arabesque);
minimaVno inflammatory response (e-Fig. 38.35).
6. Other. Inflammatory and/or structural alterations with varied clinical and
histologic presentations can also be seen in the subcutaneous adipose tissue
as a result of primary inflammatory dermatoses such as lupus (profun-
dus), human immunodeficiency virus (IllY) infection, pancreatitis, alpha
1-antitrypsin deficiency, or in response to venous insufficiency, trauma, or
infection.
V. COMMON CUTANEOUS INFECTIONS
A. Viral
1. Molluscum contagiosum
a. Clinical: Single to multiple, small (1 to 2 mm), umbilicated papules,
most common in children and young adults, no site or gender predilec-
tion. In immunosuppressed persons, lesions may be numerous and large
(>1 em).
b. Microscopic: Exo-/endophytic, acanthotic, and papillomatous epider-
mis, with classic homogeneous eosinophilic cytoplasmic inclusions
(Henderson-Patterson bodies) that displace and compress keratinocyte
nuclei into small peripheral crescents; keratinocyte cytoplasm has a vio-
laceous hue (e-Fig. 38.36).
2. Verruca vulgaris
a. Clinical: Single to multiple, hyperkeratotic (verrucous) lesions, no age or
gender predilection, occur(s) most commonly on exposed skin.
b. Microscopic: Compact orthokeratosis with parakeratosis at the peaks
of a papillomatous epidermis, hypergranulosis due to increased num-
ber and size of irregular keratohyaline granules (viral keratohyaline),
keratinocytes with large irregular hyperchromatic nuclei with a conspic-
uous perinuclear halo (koilocytes), and elongated rete that bow inward
toward the center of the lesion. Any of these alterations can be lost over
time (e-Fig. 38.37).
3. Verruca plana
a. Clinical: Usually multiple, small, flat-topped, flesh-colored to pink
papules; most common on the face and hands; occur at any age in both
genders; may spread to adjacent skin by trauma such as scratching or

shaving.
b. Microscopic: Hyperorthokeratosis (basketweave), hypergranulosis/viral
keratohyaline, some scattered koilocytes, epidermal acanthosis with a
relatively flat surface and base (e-Fig. 38.38).
4. Condyloma acuminatum
a. Clinical: Single or multiple, flesh-colored, papillomatous papules/
plaques; external genitalia, perineum, and anus.
b. Microscopic: Variable parakeratosis with compact orthokeratosis, acan-
thosis with mild papillomatosus, viral keratohyaline, and koilocytes
in varying amounts. If dysplasialdysmaturation of the keratinocytes is
noted, testing for high-risk human papilloma virus (HPV) serotypes (e.g.,
serotypes 16 and 18) can be performed. Striking cytologic atypia can
also be incited by topical treatment with agents such as podophyllin
(e-Fig. 38.39).
5. Myrmecialdeep palmoplantar wart
a. Clinical: Single to multiple, small, hyperkeratotic, variably papilloma-
tous; restricted to acral or periungual skin (hands/feet).
b. Microscopic: Exo-/endophytic, compact hyperortho- and parakeratosis,
viral keratohyaline, prominent basophilic nuclei, koilocytes, multiple
large irregular eosinophilic cytoplasmic inclusions. Nuclear size is not
affected by the viral inclusions, and the nucleus is not displaced by the
inclusions (e-Fig. 38.40).
6. Herpes (HSV/varicella zoster virus)
a. Clinical: Variable.
i. Primary herpes simplex - Painful grouped vesicles which ulcerate;
ulcer frequently has a scalloped border.
ii. Primary varicella- Vesicles on an erythematous base following a one
to two day prodrome; vesicles are highly pruritic and accompanied
by fever; trunk is a favored site; lesions appear in crops, crust in
<24 hours; most commonly seen in children.
iii. Recurrent varicella (zoster)- Commonly restricted to a dermatome
(an area innervated by one nerve); may otherwise appear similar to
HSV infection.
iv. Herpes folliculitis- Painful, erythematous nodules.
b. Microscopic: Acantholysis, dyskeratosis, multinucleate keratinocytes
with irregularly shaped nuclei, chromatin displaced to the nuclear mem-
brane by central homogeneous basophilic material that corresponds to
the viral protein. Neutrophils, hemorrhage, and necrosis are invariably
present, but variable in amount. Herpetic lesions commonly display peri-
lesional interface dermatitis and sebocytis. In herpes folliculitis, similar
changes are restricted to some portion of the hair follicle epithelium and
the epidermis is usually spared (e-Fig. 38.41).
B. Fungal. The most common fungi present on the surface of the skin is the yeast
form of Malassezia sp. (Pityrosporum). These small, ovoid forms in the stratum
corneum are incidental findings and, unless seen deep to the infundibulum of
the hair follicle and associated with inflammation, are usually not reported.
1. Dermatophytosis
a. Clinical: Variable; pustules at the advancing border suggest tinea.
i. Tinea corporis - Annulat; erythematous plaques with scale; any
site/age/gender.
ii. Tinea capitis/tinea barbae- Folliculitis of scalp or face.
iii. Tinea faciei- Facial erythema and scaling.
iv. Tinea pedis/athlete's foot- Itchy, scaly rash on feet.
v. Onychomycosis - Thickened and/or discolored nails.
b. Microscopic: Fungal hyphae are present in the stratum corneum, usually/

often visible without the use of special stains such as Gomori
methenamine silver (GMS)/PAS. All other changes are highly variable,
including parakeratosis, intracorneal neutrophils, and dermal inflam-
mation. Clippings of nails for the evaluation of fungal nail infections/
onychomycosis are processed after softening in a phenol solution
(such as a depilatory) and should always be stained for organisms
because the hyphae are usually invisible without special stains (e-Fig.
38.42).
2. Angioinvasive fungus
a. Clinical: Necrotic papules/plaques in severely immunosuppressed per-
sons; frequent, but not invariable, systemic symptoms.
b. Microscopic: Ischemic necrosis of the epidermis and superficial dermis;
extravasation of erythrocytes; fungal hyphae present in mid to deep
dermal blood vessels of all sizes with extension through the vascular
wall; inflammation type and amount highly variable and may be absent
(e-Fig. 38.43).
3. Traumatically implanted fungus/dematiaceous fungus
a. Clinical: Variable from scaly and erythematous to nodular and verrucous;
often at sites of trauma or implantation of vegetable matter; clinician
may see foreign material such as a wood splinter. Verrucous lesions are
common in chromomycosis; draining sinuses in mycetoma.
b. Microscopic: Pseudocarcinomatous epidermal hyperplasia and/or ulcer-
ation; mixed acute, chronic, and granulomatous inflammation; brown/
golden hyphae and spore forms (Medlar bodies/"copper pennies") can
be seen depending on the organism (e-Fig. 38.44).
VI. DERMAL DEPOSITS
A. Myxoid
1. Myxoma
a. Clinical: Usually solitary, flesh-colored, papule or nodule; most common
in adults, with no site predilection.
b. Microscopic: Pools of acellular, hypovascular, wispy blue material in
the papillary and/or reticular dermis; usually moderately well circum-
scribed but not encapsulated. Stains such as colloidal iron or Alcian
blue at low pH can be used to confirm the presence of interstitial mucin
(e-Fig. 38.45).
2. Digital mucous cyst
a. Clinical: Solitary dome-shaped papule or nodule that may appear cystic,
on the distal finger/toe in proximity to joint or nail; middle-aged to elderly
adults, slightly more common in women.
b. Microscopic: Compact hyperorthokeratosis with variable other epider-
mal change(s); acellular, wispy blue material in the papillary dermis; there
may be cystic degeneration.
3. Calcinosis cutis
a. Clinical: Varied, ranging from small firm papules, to plaques, to tumor
nodules.
b. Microscopic: Basophilic deposits usually in the dermis, varying from
small areas with a cracked geographic appearance, to large well-
circumscribed pools with less intense basophilia and a homogeneous
appearance (e-Fig. 38.46).
4. Osteoma cutis
a. Clinical: Highly varied. One common variant presents as multiple firm
papules, most common on the face at the sites of acne scars; may be
seen as a secondary change in cysts, benign cutaneous neoplasms, or
malignant cutaneous neoplasms.
b. Microscopic: Trabecular bone with fatty replacement of the marrow cav-

ity and/or hematopoiesis; osteoblasts in Haversian canals; sometimes
osteoclasts (e-Fig. 38.47).
5. Gout
a. Clinical: Nodules/dermal deposits; ear or periarticular sites.
b. Microscopic: Dermal/subcutaneous nodular amorphous pink deposits
with variable surrounding foreign body giant cell reaction. After pro-
cessing, birefringent needles are not detectable (e-Fig. 38.48).
6. Amyloid
a. Systemic
i. Clinical: Papular or nodular lesions; purpura.
ii. Microscopic: Hyaline deposits in the dermis or amyloid rings around
blood vessels and adipocytes.
b. Primary cutaneous
i. Clinical: Macular and lichen amyloidosis are seen as pebbled/
pigmented areas on the upper back and shins, respectively; nodu-
lar amyloid presents as solitary or multiple nodules in varied
locations.
ii. Microscopic: Macular amyloidosis appears histologically normal at
scanning power; classic amyloid deposits are seen in the dermal papil-
lae; lichen amyloidosis is similar to macular with the added abnormal-
ity of lichen simplex chronicus; nodular amyloidosis has large dermal
aggregates of amyloid and plasma cells.
VII. KERATINOUS CYSTS. These are epithelial-lined cystic structures in the dermis
and/or subcutis. They vary in manner of keratinization and/or cyst contents.
A. Infundibular type
1. Clinical: Solitary, slow growing, dome-shaped, mobile nodule with a small
opening to the skin (punctum); most common on the face, neck, and
trunk.
2. Microscopic: Lined by stratified squamous epithelium with orthokeratotic
(basketweave) keratin and an intact granular layer, but devoid of rete. The
cyst contains laminated loose keratin (e-Fig. 38.49).
B. Trichilemmal type (pilar, or isthmuslcatagen)
1. Clinical: Solitary or multiple, smooth, mobile, firm nodule(s); most common
on the scalp of females.
2. Microscopic: Lined by stratified squamous epithelium with an abrupt tran-
sition to compact orthokeratotic keratin; granular layer is absent or min-
imally present; keratinocytes grow larger toward the lumen. The cyst
contains compact eosinophilic keratin, which may be focally calcified
(e-Fig. 38.50).
C. Steatocystoma
1. Clinical: Uncommon, solitary (simplex) or multiple (multiplex), smooth,
firm, yellow or skin-colored cystic papules and nodules; most common on
the trunk and proximal extremities.
2. Microscopic: An undulating cyst lined by a thin, stratified squamous epithe-
lium that is covered by a homogeneous eosinophilic cuticle; sebaceous
glands may communicate with the cyst cavity, which is usually devoid of
contents (e-Fig. 38.51).
D. Vellus hair cyst
1. Clinical: Multiple, small, asymptomatic skin-colored papules; most com-
mon on the chest and axillae of children or young adults.
2. Microscopic: Small cyst lined by stratified squamous epithelium showing
either epidermal or trichilemmal keratinization; contains multiple small vel-
Ius hairs intermixed with keratin (e-Fig. 38.52).
VIII. DISORDERS OF COLLAGEN AND ELASTIN

A. Morphea/localized scleroderma
1. Clinical: One or more depressed, firm, bound-down plaques; they are
slightly erythematous to violaceous during the inflammatory stage; they
develop an ivory white appearance with chronicity; most common on the
trunk or extremities in children or young adults, with a slight female pre-
dominance.
2. Microscopic: Thick hypocellular collagen bundles replace the usual hap-
hazardly arranged reticular dermal collagen; adipose tissue around eccrine
coils may be absent; arrector pili muscles may show hypertrophy; dermal-
subcutaneous interface is flattened; plasma cells may be present and, in
early-stage/inflammatory-stage morphea, deep interstitial plasma cells may
be the only alteration (e-Fig. 38.53).
B. Lichen sclerosus et atrophicus
1. Clinical: Slightly depressed, pale patches/plaques with a tissue paper-like
surface; preferentially involves the female genitalia; more common past
middle age; extragenital lesions most common on hair-bearing skin; called
balanitis xerotica obliterans when the glans penis/prepuce is affected.
Genital lesions give rise to squamous cell carcinoma in a minority of
cases.
2. Microscopic: The unusual combination of compact hyperkeratosis and epi-
dermal atrophy is a clue to the diagnosis at scanning magnification. Early
lesions are lichenoid, mimicking lichen planus; lesions eventually develop
homogenization/pallor of the papillary dermal collagen, which is acellular
but contains prominent dilated vessels. Evidence of a preexisting lichenoid
infiltrate may be seen as a band of lymphocytes with melanophages beneath
the altered collagen (e-Fig. 38.54).
C. Pseudoxanthoma elasticum
1. Clinical: Small, flesh-colored papules with the appearance of plucked
chicken skin, most common on the neck/axillae; may be associated with
retinal changes (angioid streaks) and incompetence of blood vessels man-
ifested by hypertension, cerebrovascular accidents, and sudden cardiac
death. Involvement of the vessels of the gastrointestinal system may result
in bleeding/hemorrhage. Hereditary, with both autosomal dominant and
recessive forms that map to chromosome 16. A nonhereditary, acquired
variant affects only the skin and is most common in the periumbilical region
of obese multigravid women.
2. Microscopic: Short, coiled, beaded basophilidamphophilic elastic tissue
fibers are present in the superficial reticular dermis (e-Fig. 38.55). Calcium
is deposited on these abnormal elastic fibers and can be demonstrated by a
calcium stain such as von Kossa.
D. Solar elastosis
1. Clinical: Wrinkled skin at sites of extensive sun exposure, usually seen in
the elderly or in persons with excess exposure to ultraviolet light.
2. Microscopic: Amorphous, acellular basophilic material replaces the papil-
lary and superficial reticular dermis (e-Fig. 38.56).
IX. DISORDERS OF EPIDERMAL MATURATION AND KERATINIZATION
A. Acantholytic dyskeratosis
1. Clinical: Varied, including malodorous, erythematous lesions in seborrheic
areas; solitary pruritic lesions on the trunk; solitary warty lesions most
commonly on the head and neck.
2. Microscopic: Suprabasilar deft, acantholysis, dyskeratosis, elongated rete
(villi), and distinctive granular keratinocytes that are perfectly round (corps
ronds), or parakeratotic and acantholytic (corps grains) (e-Fig. 38.57).
B. Granular parakeratosis
1. Clinical: Brownish keratotic plaques at sites where skin rubs against skin;
most common in the axillae; may be pruritic.
2. Microscopic: Keratohyaline granules are present in a confluent, thick layer
of parakeratotic keratin (e-Fig. 38.58).
c. Parakeratosis
1. Clinical: Multiple clinical types; most commonly multiple erythematous
macules with a thread-like palpable scale at their border on sun-damaged
skin (disseminated superficial actinic porokeratosis [DSAP]).
2. Microscopic: All lesions of porokeratosis have one or more cornoid lamel-
lae consisting of a thin spire of parakeratotic keratin arising from an area
of the epidermis without a granular layer and usually associated with iso-
lated, scattered, dyskeratotic keratinocytes. If the entire lesion is sampled, a
cornoid lamella can be seen at both edges/sides of the lesion. DSAP has an
atrophic epidermis between the cornoid lamella and a patchy/discontinuous
band of lymphocytes in the superficial dermis (e-Fig. 38.59).
Nonmelanocytic Tumors
of the Skin
Nathan C. Walk, Yumei Chen, Anne C. Lind,
Friederike Kreisel, and Dongsi Lu
I. BENIGN TUMORS OF THE EPIDERMIS

A. Seborrheic keratosis
1. Clinical: Single or multiple discrete papules or plaques, typically measuring
0.5 to 1.0 em, variably pigmented and with a "stuck-on" appearance, only
on hair-bearing skin; most commonly found in people >30 years of age.
2. Microscopic: Exophytic or endophytic lesion sharply demarcated from adja-
cent epidermis, composed of basaloid and often homogeneous cells mixed
with squamoid cells. Intraepidermal pseudocysts filled with loose orthok-
eratotic keratin are usually present. There are many microscopic patterns
of seborrheic keratoses, the most common being hyperkeratotic, acan-
thotic, reticulated, and clonal. Features of irritation are commonly seen
(e-Fig. 39.1).
B. Clear cell acanthoma
1. Clinical: Uncommon, slow growing, pink to brown, dome-shaped nodule or
small plaque, most frequently on the leg of middle-aged and elderly individ-
uals.
2. Microscopic: Acanthotic epidermis with a sharply demarcated proliferation
of keratinocytes with pale/dear cytoplasm with associated parakeratosis,
agranulosis, and neutrophils in the stratum corneum and spinosum. Periodic
acid-Schiff (PAS) stain highlights cytoplasmic glycogen (e-Fig. 39.2 ). *
C. Large cell acanthoma
1. Clinical: A sharply demarcated, scaly patch on the sun-exposed skin of
middle-aged and elderly individuals.
2. Microscopic: Sharply demarcated acanthosis, keratinocyte nuclear and cyto-
plasmic enlargement (about 2x normal), hypergranulosis and hyperorthok-
eratosis (e-Fig. 39.3).
II. PREMALIGNANT AND MALIGNANT TUMORS OF THE EPIDERMIS
A. Actinic keratosis
1. Clinical: Scaly, erythematous papules or nodules on sun-exposed skin of the
head and neck, upper and lower extremities; more common in fair-skinned
individuals.
2. Microscopic: Patchy parakeratosis and agranulosis that often spare adnexal
ostia, irregular downward buds of atypical (dysplastic) basal keratinocytes.
Dyskeratotic cells are sometimes present. Dysplasia does not involve the full
thickness of the epidermis, and solar elastosis is frequently seen (e-Fig. 39.4).
3. Genetics: Approximately 50% of actinic keratoses show TP 53 mutations and
overexpression of cyclin D1, whereas 16% have independent activation of
Ras. Loss of heterozygosity is commonly seen on chromosome 3p (31 %), 9p
(39%), 9q (22%), 13q (52%), 17p (64%), and 17q (46%); human papilloma
virus (HPV) is detected in 41% of cases.
595
B. Squamous cell carcinoma in situ (Bowen disease}

1. Clinical: Sharply demarcated, scaly, often hyperkeratotic macule, papule, or
plaque; most common in sun-exposed areas (particularly the face and legs);
more common in fair-skinned, older individuals.
2. Microscopic: Keratinocytes with enlarged, hyperchromatic nuclei and mini-
mal cytoplasm occupy the full thickness of the epidermis; variable acantho-
sis, hyperparakeratosis, agranulosis, dyskeratosis, mitoses above the basal
layer and loss of maturation; the neoplastic cells populate adnexal structures
(e-Fig. 39.5).
3. Genetics: Increased expression and mutation of TP53 have been observed.
Allelic deletion of one or more chromosome 9q markers has also been
detected in occasional lesions.
C. Invasive squamous cell carcinoma (The American Joint Committee on Cancer
[AJCC] Tumor, Node, Metastasis [TNM] staging scheme for cutaneous squa-
mous cell carcinoma and other cutaneous carcinomas is given in Table 39.1.)
1. Clinical: Early lesions are finn, skin-colored, or erythematous nodules; later
lesions are shallow ulcers with firm, elevated, or indurated surroundings,
sometimes with crust or scale; most common in fair-skinned, older individ-
uals on sun-exposed areas and in immunocompromised patients. Metastatic
potential depends on location, histology, and precursor lesions/etiology.
2. Microscopic: Acanthosis, invasion of the dermis by individual or nested
keratinocytes with variable amounts of cytoplasm and nuclear pleomor-
phism, dyskeratosis, and swirls of parakeratotic keratin (keratin pearls); there
may be dermal desmoplasia and variable inflammation. Acantholysis, per-
ineural invasion, and sarcomatoid change predict more aggressive behavior
(e-Fig. 39.6).
D. Basal cell carcinoma
1. Clinical: Variable presentation; most commonly a dome-shaped, firm papule
with a pearly, telangiectatic surface; slowly growing; primarily on the head,
neck, and trunk. The most common skin cancer in Caucasians; rarely metas-
tasizes unless neglected.
2. Microscopic: Variable histology; Common features include basaloid cells with
hyperchromatic nuclei and scant cytoplasm, usually connected to an unre-
markable or ulcerated epidermis. Basaloid islands show peripheral palisad-
ing, mitoses, and apoptosis; a retraction artifact separates the basaloid cells
from the surrounding basophilic, variably mucinous stroma (e-Fig. 39.7).
3. Genetics: Mutations of genes PTCH1 on chromosome 9q22.3 and SMD on
chromosome 7q31-32 involved in activating hedgehog signaling pathway
have been identified in both sporadic basal cell carcinomas and basal cell
nevus syndrome, a rare autosomal dominant (AD) disorder.
Ill. TUMORS OF THE CUTANEOUS APPENDAGES. Tumors of the cutaneous appendages
have a somewhat nebulous nomenclature. A traditional system organizes the
tumors on the basis of their origin from one of the normal cutaneous appendages:
hair follicle, sebaceous gland, eccrine gland, or apocrine gland. However, complex
adnexal tumors also occur that have abnormalities of two or more of the normal
skin constituents, including the epidermis itself, and there is sometimes controversy
as to a tumor's precise histogenesis (i.e., eccrine vs. apocrine, and so on). The fol-
lowing is a succinct review of the major tumors derived from the skin appendages.
A. Benign appendage tumors. Benign adnexal tumors have architectural symmetry
when viewed at low power and, usually, a distinctive cleft between the stroma
of the tumor and the native dermal collagen.
1. Hair follicle tumors
a. Trichofolliculoma
i. Clinical: Rare, solitary, small (< 1 em), skin-colored papule on the face.
A tuft of fine hairs protrudes from the center.
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ii. Microscopic: One or more cystically dilated hair follicles with radiating
follicles that project into a relatively cellular stroma. The secondary hair
follicles are of variable maturity and may give rise to more hair follicles
(e-Fig. 39.8).
b. Trichoepithelioma
i. Clinical: Usually solitary, skin-colored papule, most common on the
central face; if multiple, associated with AD inheritance. The desmo-
plastic variant is a solitary, firm annular lesion with a raised border
and central depression that occurs exclusively on the face.
ii. Microscopic: A relatively well-circumscribed dermal tumor composed
of islands of basaloid cells with pilar differentiation in a cellular stroma;
papillary mesenchymal bodies are characteristic. Small keratinous cysts
and foci of calcification are often present. The tumor epithelium may
mimic basal cell carcinoma, but there is no retraction artifact or muci-
nous stroma (e-Fig. 39.9). The desmoplastic variant shows a well-
circumscribed lesion in the upper and mid-dermis composed of cords or
small nests of basaloid cells in a sclerotic stroma (e-Fig. 39.10). It may
be confused with a syringoma or a morphea-form basal cell carcinoma.
c. Trichoadenoma (of Nikolowski)
i. Clinical: Rare nodule on the face or buttocks.
ii. Microscopic: A well-demarcated dermal tumor composed of multiple
cyst-like structures lined by multilayered keratinizing pilar-type squa-
mous epithelium. The cysts contain keratinous debris and no hair shafts
(e-Fig. 39.11).
d. Dilated pore of Winer
i. Clinical: Common, usually solitary, comedo-like lesion on the head and
neck, or trunk.
ii. Microscopic: A cystically dilated hair follicle, filled with loose keratin
and lined by acanthotic stratified squamous epithelium with irregular
budding (e-Fig. 39.12).
e. Trichilemmoma
i. Clinical: A solitary, small, skin-colored/pink or brown papule on the
face and neck; multiple lesions are associated with Cowden syndrome.
ii. Microscopic: A well-circumscribed, endo-exophytic tumor composed
of clear squamoid cells with glycogenated cytoplasm. The tumor
extends from the epidermis with a lobular configuration. There is a
thin peripheral rim of palisading columnar cells and variable, thick,
eosinophilic basement membrane (e-Fig. 39.13).
f. Pilomatrixoma (Calcifying epithelioma of Malherbe)
i. Clinical: Firm, deeply located nodule, most common on the face and
upper extremities; onset frequently in childhood.
ii. Microscopic: A sharply demarcated tumor in the lower dermis and,
often, the subcutis. The tumor is composed of large, irregularly shaped
tumor islands and intervening stroma. Two basic cell types are present:
basaloid cells and "shadow" or "ghost" cells. The basaloid cells resem-
ble the cells in basal cell carcinoma and are present at the periphery
of the tumor islands. The shadow cells, which have eosinophilic cyto-
plasm, distinct cell borders, and no nuclear staining, occupy the center
of tumor islands. Several layers of transitional cells with intermediate
features may be present. In addition, dystrophic calcification, a mixed
inflammatory infiltrate, hemosiderin, melanin, bone, and foreign body
giant cells can be seen (e-Fig. 3 9.14).
g. Fibrofolliculomaltrichodiscoma
i. Clinical: Skin-colored papules, most common on the face; may be
solitary or multiple. Multiple lesions are seen in Birt-Hogg-Dube
Chapter 39 • Nonmelanocytic Tumors of the Skin I 59 9
syndrome, which has an AD inheritance and carries a high risk for

developing renal tumors, pulmonary cysts, and pneumothorax.
ii. Microscopic: Spectrum of changes with fibrofolliculoma at one end
and trichodiscoma at the other. Fibrofolliculoma is composed of thin
strands of follicular epithelium extending from a hair follicle structure
into a stroma that is well defined and composed of loose connective
tissue with fine fibrosis. A trichodiscoma has a dominant stromal com-
ponent that is characterized by strands of loose connective tissue with
intermixed fibroblasts resembling an angiofibroma (e-Fig. 39.15).
2. Eccrine tumors
a. Syringoma
i. Clinical: Usually multiple, skin-colored, small, firm papules on the
lower eyelids and cheeks, more common in women; onset is often at
puberty.
ii. Microscopic: Multiple small ducts in a dense fibrous stroma in the der-
mis. The ducts are lined by two layers of cuboidal epithelium, and
sometimes are "tadpole" or "comma-like." Solid nests and strands of
tumor cells can be present (e-Fig. 39.16).
b. Eccrine {or apocrine} mixed tumor {chondroid syringoma}
i. Clinical: Solitary, firm, dermal or subcutaneous nodule on the head and
neck.
ii. Microscopic: Well-circumscribed dermal tumor of small epithelial cells
in solid nests, small clusters, and ducts in a prominent myxoid, carti-
laginous, and/or fibrous stroma (e-Fig. 39.17).
c. Cylindroma
i. Clinical: Solitary nodule on the head and neck; predominantly in
middle-aged to elderly women; rarely undergoes malignant transfor-
mation; if multiple, associated with AD inheritance and an occurrence
as "turban tumors"
ii. Microscopic: A poorly circumscribed dermal tumor composed of irreg-
ularly shaped nests of basaloid cells, surrounded by a thick eosinophilic
basement membrane. The cellular nests fit together in a "jigsaw" pat-
tern (e-Fig. 39.18).
d. Spiradenoma
i. Clinical: Usually solitary, firm, tender or painful dermal nodule. There
is no characteristic distribution. They occur primarily in young adults.
ii. Microscopic: One or more sharply demarcated dermal nodule(s) of
basaloid cells. Two types of cells are present: small lymphocyte-like
cells with hyperchromatic nuclei and larger epitheliod cells with more
open chromatin. There may be a thin pseudocapsule; a few duct-like
structures are present (e-Fig. 39.19).
e. Poroma group {including eccrine poroma, dermal duct tumor, and hidroacan-
thoma simplex}
i. Clinical:
{a} Eccrine poroma---solitary, sessile or slightly pedunculated, pink
nodule often on the plantar or palmar skin, or other locations with
sweat glands.
{b) Dermal duct tumor-a solitary, firm nodule on the head, neck, and
extremities.
{c) Hidroacanthoma simplex-a solitary plaque or nodule on the
extremities and trunk; clinically resembles seborrheic keratosis or
basal cell carcinoma.
ii. Microscopic:
(a} Eccrine poroma-a circumscribed tumor composed of columns of
basaloid cells extending from the lower epidermis into the dermis
within a loose, vascular stroma. Ducts and, rarely, small cysts may
be seen in the tumor columns. There is a sharp demarcation from
the epidermis (e-Fig. 39.20).
{b) Dermal duct tumor-islands of basaloid cells similar to those of
eccrine poroma, located entirely in the dermis. An epidermal con-
nection may be found if multiple sections are examined.
{c) Hidroacanthoma simplex-islands of basaloid cells similar to those
of poroma, confined to the epidermis.
f. Acrospiroma {nodular hidradenomai solid-cystic hidradenomai apocrine/
eccrine hidradenomai clear cell hidradenoma)
i. Clinical: Solitary nodule, 0.5 to 2.0 em or more, no site predilection,
disputed histogenesis. Nomenclature for this entity is confusing.
ii. Microscopic: Variable histologic appearance, as reflected by the nosol-
ogy; usually a circumscribed, nonencapsulated, multilobula.r; cen-
tral dermal tumor with variable proportions of cystic and solid
areas; tumor cells have variable clear or eosinophilic cytoplasm and
are round, fusiform, or polygonal. Duct-like structures are typi-
cally present and may have a squamous appearance. The stroma
varies from rather fine fibrous tissue to dense hyalinized collagen
(e-Fig. 39.21).
3. Sebaceous hyperplasia and tumors
a. Sebaceous hyperplasia
i. Clinical: Yellow to whitish papules with central umbilication, typically
on the face of older individuals. It may mimic a basal cell carcinoma.
ii. Microscopic: Multiple large, but otherwise normal, sebaceous lobules
centered on a large central orifice; solar elastosis is frequently present
(e-Fig. 39.22).
b. Sebaceous adenoma
i. Clinical: Rare, solitary or multiple, pink or flesh-colored, usually
< 1 em nodule(s) on the face or scalp of adults; may be associated
with Muir-Torre syndrome (visceral carcinoma).
ii. Microscopic: Multiple circumscribed sebaceous lobules usually cen-
tered in the superficial to mid-dennis; composed of peripheral basaloid
germinative cells, central mature sebaceous cells, and a variable zone
of transitional forms. Mature cells usually outnumber the basaloid
cells (e-Fig. 39.23 ).
iii. Genetics: Loss of protein expression of DNA mismatch repair genes
(MSH-2, MSH-6, MLH-1, and PMS-2) has been found in a number of
patients with sebaceous adenoma. There is a high correlation between
lost expression of mismatch repair (MMR) proteins and microsatellite
instability (MSI).
4. Apocrine tumors
a. Apocrine hidrocystoma {apocrine cystadenoma)
i. Clinical: Solitary, translucent to bluish nodule, predominantly occur-
ring on the face.
ii. Microscopic: Several large cysts in the dermis; the cyst wall is com-
posed of an outer myoepithelial layer and an inner layer of cuboidal to
columnar cells with apocrine decapitation secretion; pseudopapillary
projections may be present (e-Fig. 39.24).
b. Syringocystadenoma papilliferum
i. Clinical: Varied; most commonly a raised, warty plaque on the scalp;
associated with nevus sebaceous in approximately one-third of the
cases.
ii. Microscopic: Deep invaginations of duct-like structures extend from an
acanthotic, variably papillomatous epidermis into the dermis; they are
Chapter 39 • Nonmelanocytic Tumors of the Skin I 601
lined by squamous epithelium in the upper portion and by two-layer,

sweat duct-like epithelium in the lower portion; numerous plasma
cells are present in the connective tissue stroma of the papillae (e-Fig.
39.25).
iii. Genetics: Allelic deletions of the patched gene on chromosome 9q22
and loss of heterozygosity of chromosome 9p21 have been reported
in syringocystadenoma papilliferum.
5. Complex adnexal tumors: Nevus sebaceous of Jadassohn {organoid nevus)
a. Clinical: Solitary, yellow or waxy, patch or plaque with alopecia on the
scalp (most common), face, neck, or trunk; present at birth or first noticed
in early childhood.
b. Microscopic: A complex hamartoma involving the epidermis, piloseba-
ceous unit, and ducts/glands. There is variable epidermal acanthosis, papil-
lomatosis, and abnormally formed/abortive hair follicles with bulbs that
do not extend into the subcutis; increased numbers of large sebaceous
glands are present around and after puberty; dilated apocrine glands
are found in up to 50% of cases (e-Fig. 39.26). Secondary tumors may
develop, such as syringocystadenoma papilliferum.
B. Malignant tumors of the cutaneous appendages are rare and far less common
than their benign counterparts. As with the benign tumors discussed earlier, a
malignant neoplasm can arise from any of the normal skin appendages, and there
may be more than one component to a particular tumor. Only extramammary
Paget disease will be discussed.
1. Clinical: A red, scaly plaque, typically in areas rich in apocrine glands such
as the anogenital region and, less commonly, the axilla; usually pruritic and
slowly spreading. An underlying adnexal carcinoma is present in approxi-
mately 20% to 25% of cases; visceral carcinoma (rectal, prostate, bladder,
cervix, or urethra) is present in 10% to 15% of cases.
2. Microscopic: Similar to mammary Paget disease, there are large, pre-
dominantly intraepidermal, epithelioid tumor cells with large, pleomor-
phic nuclei and abundant pale cytoplasm that may or may not con-
tain mucin. The tumor cells are concentrated in the lower epidermis and
randomly dispersed throughout the epidermis; they may also be seen in
the superficial dermis. Special stains may be needed to distinguish extra-
mammary Paget disease from melanoma in situ and squamous cell car-
cinoma in situ with pagetoid features. The tumor cells are positive by
mucicarmine and PAS stain; carcinoembryonic antigen (CEA), epithelial
membrane antigen (EMA), and low-molecular-weight keratin immunos-
tains are positive; Melan-A/Mart-1 and S-100 immunostains are negative
(e-Fig. 39.27).
IV. BENIGN AND MALIGNANT TUMORS OF MESENCHYMAL ORIGIN. Classification of mes-
enchymal neoplasms is based on the mature, nonneoplastic tissue from which they
are believed to be derived: blood vessel, nerve, smooth muscle, skeletal muscle,
bone, cartilage, fibrous tissue, neuroendocrine tissue, or hematopoietic elements.
Many of these tumors are discussed elsewhere in this text (see Chaps. 43 to 44 and
46 covering hematolymphoid and soft tissue malignancies, respectively); only the
most common entities will be discussed here.
A. Vascular tumors
1. Lobular capillary hemangioma {pyogenic granuloma)
a. Clinical: Common, rapidly developing polypoid or pedunculated, red,
eroded papules or nodules that bleed easily; seen on mucous membranes
and skin.
b. Microscopic: An exophytic, frequently ulcerated, proliferation of capillar-
ies divided into lobules by fibrous tissue septae; variably cellular, often
surrounded by a collarette of epidermis (e-Fig. 39.28).
2. Arteriovenous hemangioma (acral arteriovenous tumor}

a. Clinical: A solitary, usually asymptomatic, red or purple, enlarging or
bleeding papule, averaging 4 mm in diameter; most common on the lips,
perioral skin, nose, and eyelids of middle-aged to elderly men.
b. Microscopic: Well-circumscribed, nonencapsulated vascular tumor in the
upper to mid-dermis, composed of closely packed, large caliber, thick-
walled and thin-walled vessels (e-Fig. 39.29).
3. 11Cherry" angioma (senile angioma, Campbell de Morgan spot}
a. Clinical: Solitary or multiple tiny, bright red papules, predominantly on
the trunk and proximal extremities; common in adults, almost universal
in the elderly.
b. Microscopic: Early lesions with one or more dilated interconnecting thin-
walled vessels in the papillary dermis. Established lesions are polypoid
with an epidermal collarette and are composed of dilated and congested
vascular channels in the papillary dermis (e-Fig. 39.30).
4. Angiokeratoma
a. Clinical: Single to multiple red to black papule(s) or plaque(s) that may be
warty or hyperkeratotic; multiple variants exist with characteristic clinical
presentations and carrying various eponyms.
b. Microscopic: Markedly dilated and congested papillary dermal vessels
form cavernous channels that have an intimate association with the epi-
dermis; irregular acanthosis and variable hyperkeratosis (e-Fig. 39.31).
5. Lymphangioma {cystic lymphatic malformation)
a. Superficial lymphangioma (lymphangioma circumscriptum)
i. Clinical: Multiple, localized, scattered, or grouped translucent vesicles
or papulovesicles; may be red or purple due to intralesional hemorrhage
and thrombus formation; usually congenital, sometimes presents later
in childhood and is rare in adults. There may be a deeper component.
ii. Microscopic: Multiple dilated, thin-walled lymphatic channels in the
papillary and upper reticular dermis; epidermis is sometimes acanthotic
and forms a collarette (e-Fig. 39.32).
b. Deep lymphangioma (including cavernous lymphangioma and cystic hygroma)
i. Clinical: Usually solitary, rubbery, skin-colored nodules that may result
in swelling of the soft tissue; most common on the face, trunk, and
extremities; varies from spongy to a large cystic tumor. Most are present
at birth or the first few years of life.
ii. Microscopic: Variable; in general, there are irregularly dilated vascu-
lar channels of variable size in the dermis, the subcutis, and deeper
tissue. The vessels are thin- or thick-walled, and contain luminal pro-
teinaceous fluid or lymphocytes. There is no endothelial atypia or
mitotic activity. The intervening stroma is unremarkable or fibrotic
(e-Fig. 39.33).
6. Glomus tumor and glomangioma
a. Clinical: Painful, solitary or multiple, red to purple subungual macule(s);
may be a nodule at other sites. Glomangioma is less painful.
b. Microscopic: Well-circumscribed or encapsulated dermal tumor composed
of sheets of glomus cells surrounding blood vessels. The glomus cells are
homogeneous with eosinophilic cytoplasm and dense, round nuclei. The
tumor stroma is fibrous and often pale-staining (e-Fig. 39.34). A glo-
mangioma is predominantly vascular and has fewer glomus cells (e-Fig.
39.35). The glomus cells are positive for smooth muscle actin and negative
for endothelial markers such as CD31.
7. Kaposi sarcoma
a. Clinical: Early lesions are ecchymotic macules or patches; later lesions
are bluish or purple papules, nodules, plaques, and tumors; all are pal-
pable. There are four clinical subtypes with similar cutaneous findings:
Classic, African (endemic), Epidemic (human immunodeficiency virus

[HIV]-associated), and Iatrogenic (immunosuppressive therapy related).
Regardless of type, it is a low-grade malignancy that is slowly progressive
but may involve internal organs. The etiologic agent is human herpes virus
type 8 (HHV-8) in all types.
b. Microscopic: Early lesions show a dermal proliferation of irregular slit-
like vascular channels with extravasated erythrocytes, hemosiderin, and
plasma cells. The endothelial cells are plump or inconspicuous; there is no
significant cytologic atypia. The vascular channels infiltrate between col-
lagen bundles and surround existing blood vessels (promontory sign) and
appendages (e-Fig. 39.36). As the lesions evolve into papules, plaques, and
nodules, there are increased spindle cells between the poorly defined slit-
like vessels. Intracytoplasmic eosinophilic hyaline globules (PAS positive)
can be identified within the tumor cells (e-Fig. 39.37).
8. Epithelioid hemangioendothelioma
a. Clinical: Firm, tan-pink subcutaneous nodules and plaques, uncommonly
involving the dermis, measuring several centimeters in maximal diameter;
common sites are the trunk and extremities. The lesion is seen primarily
in adults and has a slight female predilection; 40% of tumors recur, and
""'15% develop distant metastasis.
b. Microscopic: Sheets and cords of large polyhedral tumor cells in the sub-
cutis/dermis. The tumor cells have amphophilic cytoplasm, prominent
cytoplasmic vacuoles, and round nuclei often with small nucleoli; the cells
have a tendency to grow around preexisting large vessels. The stroma is
variably fibrous or myxoid. Mitotic figures and necrosis may be seen and
may indicate worse clinical behavior. The tumor cells are immunoreac-
tive for endothelial cell markers CD31, CD34, von Willebrand factor, and
thrombomodulin (e-Fig. 39.38).
9. Angiosarcoma
a. Clinical: Single or multifocal, purpuric or black plaque(s) on the head and
neck of elderly patients, or purplish-red papules or polypoid tumors in
the chronically edematous skin associated with lymphedema and/or prior
radiation.
b. Microscopic: Poorly circumscribed and often multifocal proliferation
of anastomosing, infiltrative vascular channels in the dermis and sub-
cutis with prominent extravasation of erythrocytes and hemosiderin.
The vascular channels are lined by crowded, variably plump, and atyp-
ical endothelial cells; papillary processes sometimes extend into the
lumens of the vessels. Poorly differentiated tumors with an epithe-
lioid cellular morphology may resemble carcinoma or melanoma. The
tumor cells are immunopositive for endothelial cell markers such
as CD31, CD34, factor VIII-related antigen, and Ulex europaeus
(e-Fig. 39.39).
B. Neural and neuroendocrine tumors
1. Traumatic neuroma
a. Clinical: Usually firm, pea-sized nodule in the subcutis and deep soft tissue
at sites of previous injury; may be painful.
b. Microscopic: Irregularly arranged nerve fascicles embedded in fibrous scar
tissue. Each fascicle is surrounded by fibrous tissue and perineural cells.
There are scattered mast cells (e-Fig. 39.40).
2. Solitary circumscribed neuroma (palisaded and encapsulated neuroma)
a. Clinical: Uncommon, usually solitary, skin-colored or pink papule, most
common on the face of middle-aged adults; slow-growing, painless, and
< 6 mm in diameter.
b. Microscopic: Well-circumscribed and partially encapsulated dermal nod-
ule composed of fascicles of bland spindle cells with amphophilic
cytoplasm and serpiginous nuclei that appear parallel to each other within
a single fascicle. No intervening fibrous tissue is present (e-Fig. 39.41).
3. Neurofibroma
a. Clinical: Soft, skin-colored, pedunculated papules or nodules. Multiple
lesions in a segmental or widespread distribution are related to neurofi-
bromatosis.
b. Microscopic: A nonencapsulated dermal or subcutaneous tumor charac-
terized by loosely arranged, wavy spindle cells in a pale-staining stroma.
There are increased small caliber vessels and mast cells in the stroma. The
spindle cells tend to surround adnexal structures rather than displace them
(e-Fig. 39.42).
4. Cutaneous schwannoma {neurilemmoma}
a. Clinical: Uncommon, slowly growing, usually solitary tumor/nodule with
a predilection for the limbs of adults. The neoplasm can be either sporadic
or associated with neurofibromatosis 2 (NF-2). Schwannomas are more
commonly seen in the deep soft tissue, intracranially, or intraspinally.
b. Microscopic: Similar to their soft tissue counterpart; form a circumscribed
and encapsulated subcutaneous/dermal nodule. The tumor cells are spin-
dled Schwann cells with indistinct cytoplasmic borders arranged in inter-
lacing fascicles. There are hypercellular areas (Antoni A) containing rows
or palisades of nuclei aligned around eosinophilic cellular processes (Vero-
cay bodies) and hypocellular (Antoni B) areas. No axons are present
(e-Fig. 39.43).
5. Merkel cell carcinoma
a. Clinical: A rapidly growing, often ulcerated, red nodule or plaque usually
arising on the sun-exposed skin of the elderly, particularly on the head,
neck, and extremities. An association with polyoma virus infection (specif-
ically, Merkel cell polyoma virus [MCPyV]) has been demonstrated in the
majority of cases. AJCC TNM staging scheme for Merkel cell carcinoma
b. Microscopic: Trabeculae, nests, and sheets of small cells with scant cyto-
plasm, indistinct cytoplasmic borders, vesicular nuclei with nuclear mold-
ing, and multiple small nucleoli that infiltrate the entire dermis and some-
times the subcutis. There are numerous apoptotic forms and mitoses. Local
intralymphatic spread is commonly seen. The tumor cells are positive for
neuron-specific enolase, chromogranin, and synaptophysin and have a
characteristic "paranuclear dot-like" staining pattern for low-molecular-
weight keratin such as CK20. The tumor cells are negative for CD45,
S-100, and TTF-1, allowing distinction from hematopoietic and
melanocytic neoplasms, as well as metastatic small cell carcinoma of the
lung (e-Fig. 39.44).
c. Genetics: Deletion of chromosome lp36 is commonly seen; numerous
other chromosomal abnormalities have been described, of which trisomy
6 is the most common.
6. Granular cell tumor
a. Clinical: Asymptomatic, solitary, skin-colored nodule <3 em in diameter;
multiple lesions in ""10% of cases; most common in Mrican American
adults and women. Malignant counterpart is exceedingly rare.
b. Microscopic: A nonencapsulated dermal-based tumor composed exclu-
sively of large polyhedral cells with abundant fine to coarsely granular
eosinophilic cytoplasm, and small oval centrally located nuclei. The gran-
ular cells infiltrate between collagen bundles and surround adnexa. The
epidermis may be markedly acanthotic as in pseudocarcinomatous hyper-
plasia (e-Fig. 39.45). Tumor cells are usually positive for S-100, except
in a subset of congenital lesions reported as CD34-positive granular cell
dendrocytosis.
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2. Dermatofibroma
a. Clinical: Brownish, round, firm dermal nodules, usually < 1 em in diame-
ter; most common on the legs of young adults.
b. Microscopic: Poorly circumscribed dermal proliferation of spindled fibrob-
lasts, histiocytes, and blood vessels. Fibroblasts at the periphery surround
the collagen bundles ("collagen-trapping"). Dermatofibromas are histo-
logically varied, and there are many corresponding named variants such
as cellulat; aneurysmal, "Monster" cell, and so on. The epidermis typically
shows acanthosis, basal keratinocyte hyperpigmentation, and a broad flat-
tened rete or a basaloid proliferation which can be mistaken for basal cell
carcinoma in a superficial shave biopsy (e-Fig. 39.47).
3. Dermatofibrosarcoma protuberans
a. Clinical: Solitary or multiple polypoid nodules arising in an indurated
plaque on the trunk or extremities of adults; slowly growing locally aggres-
sive tumor; rarely metastasizes.
b. Microscopic: A cellular dermal tumor composed of homogeneous spin-
dle cells arranged in a storiform or cartwheel pattern. The tumor cells
infiltrate between adnexa, and there is characteristically extension into
the subcutis with fat trapping. Occasional mitotic figures can be found,
and atypia is mild. The epidermis is normal, atrophic, or ulcerated (e-Fig.
39.48).
c. Genetics: The tumor characteristically exhibits the translocation
t(17;22)(q22;q13), which results in production of a COLIA1-PDGFB
fusion protein.
4. Giant cell fibroblastoma: A histologic variant of dermatofibrosarcoma protu-
berans, primarily affecting children; it has a strong male predominance and a
similar anatomic distribution; harbors the same translocation as dermatofi-
brosarcoma protuberans.
a. Clinical: Grossly, it is a firm yellow or gray tumor with a gelatinous or
rubbery consistency without hemorrhage or necrosis.
b. Microscopic: The tumor is usually hypocellular and composed of wavy
spindle cells and scattered giant cells with hyperchromatic and angu-
lated nuclei. The stroma is variable from myxoid, to collagenous, to scle-
rotic. Scattered mast cells are seen within the stroma. Irregular branch-
ing "angiectoid" spaces resembling dilated lymphatics may be seen, lined
by spindled or multinucleated cells morphologically identical to those of
the surrounding stroma (e-Fig. 39.49). The lining and stromal cells are
immunopositive for CD34 and immunonegative for CD31, S-100, actin,
desmin, and EMA.
5. Angiofibroma
a. Clinical (major types}
i. Fibrous papule of the face: A solitary, firm, dome-shaped, often flesh-
colored lesion on the nose or central face.
ii. Pearly penile papules: Tiny white papules, 1 to 3 mm in diametet;
arranged in groups or rows on the coronal margin of the penis.
iii. Adenoma sebaceum (tuberous sclerosis): Multiple papules or nodules
with a predilection for the butterfly area of the face.
iv. Digital fibrokeratoma: A solitary, thin, tall horn on a digit.
b. Microscopic: Dermal fibrosis; dilated small vessels; and variably enlarged,
angulated, or stellate fibroblasts. Concentric fibrosis around the blood
vessels is more prominent in adenoma sebaceum than in fibrous papule
of the face (e-Fig. 39.50). The epidermis of digital fibrokeratomas shows
hyperkeratosis and acanthosis of acral skin (e-Fig. 39.51).
c. Genetics: Mutations of two genes, TSC1 on chromosome 9 and TSC2 on
chromosome 16, are identified in patients with tuberous sclerosis.
6. Acrochordon (skin tag, soft fibroma, fibroepithelial polyp, and so on)

a. Clinical: Flesh-colored, pedunculated papules or nodules with irregular or
smooth surfaces. Most common on the axilla, neck, groin, and eyelids;
incidence increases with age; more common in obese women.
b. Microscopic: Polypoid, variable epidermal change, well vascularized, loose
dermal connective tissue. A variable amount of fat can be seen in the
dermis of larger lesions (soft fibroma). No appendages are present (e-Fig.
39.52).
7. Epithelioid sarcoma
a. Clinical: Epithelioid sarcoma has recently been divided into two distinct
subtypes.
The distal type presents as one or more slowly growing, firm, painless,
tan-white subcutaneous nodules with an indistinct infiltrating margin. The
tumor occurs most commonly on the distal extremities, particularly the
hand and wrists. Ulceration and sinus formation may be present weeks
or months after the lesion is first noted. The local recurrence rate is near
80%; distant metastasis occurs in 30% to 45% of cases, most commonly
to the lymph nodes and lung.
The proximal type is not characteristically a tumor of the skin or sub-
cutaneous tissue, but instead involves the pelvis, perineum, and/or genital
tract. It is discussed in more detail in Chapter 47.
b. Microscopic: The proximal type arises in the subcutaneous or soft tissue (it
is rarely dermal) and has a nodular arrangement of tumor cells that tend
to palisade around geographic central degeneration/necrosis. A lympho-
histiocytic infiltrate often surrounds the tumor nodules. The tumor cells
are large and polygonal with abundant deeply eosinophilic cytoplasm,
round to oval nuclei, and prominent nucleoli. Plump spindled tumor cells
are sometimes present. Mitoses are frequent. The tumor cells are positive
by immunohistochemistry for low- and high-molecular-weight keratins,
EMA, and vimentin and are also positive for CD34 in 60% to 70% of
cases. The architectural features are similar to those seen in deep granu-
lomatous processes such as deep granuloma annulare.
D. Fatty and muscular tumors
1. Lipoma
a. Clinical: Common, asymptomatic, soft, subcutaneous nodule.
b. Microscopic: Unremarkable mature adipocytes are surrounded by a thin
fibrous capsule; there is a paucity of fibrous septae.
2. Angiolipoma
a. Clinical: Often multiple, painful, subcutaneous nodules of the extremities
or trunk; usually appears after puberty.
b. Microscopic: Varying proportions of mature adipose tissue and small-
caliber blood vessels with fibrin thrombi, surrounded by a thin fibrous
capsule, and arranged vaguely into lobules by fine incomplete fibrous sep-
tae (e-Fig. 39.53).
3. Leiomyoma
a. Clinical: Solitary or multiple red nodules, often painful.
b. Microscopic: Pilar leiomyoma is a circumscribed, nonencapsulated tumor
composed of interlacing smooth muscle bundles (e-Fig. 39.54). Scro-
tal leiomyomas are similar but often have ill-defined or focally
infiltrative margins. Angioleiomyoma is a deep dermaVsubcutaneous,
well-circumscribed nodule of interlacing smooth muscle bundles between
multiple thick walled vessels (e-Fig. 39.55).
4. Leiomyosarcoma
a. Clinical: Rare, dermal or subcutaneous nodule or plaque, most common
on the extremities. Dermal leiomyosarcomas frequently extend into the
subcutaneous tissue; there are no confirmed cases of metastases from

dermal tumors. Subcutaneous leiomyosarcomas have greater tendency for
local recurrence and metastasis.
b. Microscopic: Interlacing, hypercellular smooth muscle bundles with pleo-
morphic and hyperchromatic nuclei, and occasional mitoses (at least one
per 10 high-power fields).
V. CUTANEOUS LYMPHOID INFILTRATES. These infiltrates are highly varied and encom-
pass reactive lymphoid infiltrates, primary cutaneous lymphoma, and cutaneous
involvement by systemic disease/nodal lymphoma. With the exception of mycosis
fungoides (MF), determination of whether the skin is the primary or a secondary
site of a lymphoma requires a complete clinical examination for appearance and
distribution of cutaneous lesions, presence or absence of lymphadenopathy and/or
organomegaly, systemic symptoms, and peripheral blood smear abnormalities. In
contrast to nodal lymphomas, of which B-celllymphomas comprise the vast major-
ity, ""70% of cutaneous lymphomas are of T-cell origin, of which MF is the most
common (accounting for about 50% of all cutaneous lymphomas overall). The
World Health Organization/European Organization for Research and Treatment
of Cancer (WHO/EORTC) classification and the current WHO classification are the
most current classifications on cutaneous lymphomas and are uniformly accepted
by pathologists, dermatopathologists, dermatologists, and oncologists worldwide.
Table 39.3 summarizes the WHO/EORTC classification of cutaneous lymphomas.
Since MF and primary cutaneous CD30+ lymphoproliferative disorders are the most
common cutaneous lymphomas, only these will be discussed in more detail below.
A. Mycosis fungo ides
1. Clinical: Patches, plaques, and/or tumors with a wide anatomic distribution;
most common in middle-aged to elderly persons, but documented in children.
Usually pruritic, may present with or develop erythroderma; clinical course
is usually indolent. MF is, as a rule, limited to the skin, with widespread
distribution and a protracted disease course. Extracutaneous spread may
occur in advanced stages, mainly to lymph nodes, liver, spleen, lungs, and
blood. Malignant cells of MF involving the blood in advanced stages are cere-
briform in appearance and called Sezary cells. Disease progression involving
peripheral blood must be distinguished from primary Sezary syndrome, a rare
T-celllymphoma that presents as a triad of erythroderma, generalized lym-
phadenopathy, and malignant T-cells (Sezary cells) in the peripheral blood.
The staging scheme for MF and Sezary syndrome is outlined in Table 39.4,
and the histopathologic staging of lymph nodes in MF and Sezary syndrome
are outlined in Table 39.5.
2. Microscopic: Histologic features vary with clinical presentation. Epidermal
changes range from atrophic to psoriasiform. Spongiosis may be dispropor-
tionate to the intraepidermallymphocytes.
a. Patch stage/early MF is histologically subtle. A patchy, paucicellular
band of lymphocytes is present in a fibrotic papillary dermis; lympho-
cytes extend into the epidermis (epidermotropism); intraepidermallym-
phocytes are larger than the dermal lymphocytes and have hyperchro-
matic, cerebriform nuclei; lymphocytes are separated from keratinocytes
by a halo and file along the basal layer of the epidermis (so-called
"string of beads" pattern). Aggregates of atypical intraepidermallympho-
cytes (Pautrier microabscesses) are rare. Dermal lymphocytes align along
collagen bundles (e-Fig. 39.56).
b. Plaque stage has easily identifiable Pautrier microabscesses in the epi-
dermis and a more prominent band of lymphocytes in the upper dermis
(e-Fig. 39.57).
c. Tumor stage shows a dense nodular or diffuse infiltrate filling the der-
mis and extending into the subcutis. Epidermotropism may be lost.
There may be transformation to a large cell phenotype. The infiltrate is
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diagnostic or surgical procedure. Metastases tend to occur on the skin near the
primary malignancy; metastasis to sites distant from the primary tumor is more
common in tumors that demonstrate angioinvasion (e.g., primary tumors of the
kidney or lung).
Rarely, a cutaneous metastasis is the first indication of a visceral malignancy; the
umbilicus, and less frequently the scalp, is a particularly common site of involve-
ment in this setting. Adenocarcinoma is more frequently observed as a cutaneous
metastasis than is squamous cell carcinoma or urothelial carcinoma. Cutaneous
metastases are variable in clinical appearance and can occur either as solitary or
multiple papules/nodules, or as mimicks of inflammatory/infectious conditions.
A. Sister Mary Joseph nodule
1. Clinical: This condition was named after Sister Mary Joseph (1856-1939), a
surgical assistant for Dr. William Mayo, who noted the association between
paraumbilical nodules observed during skin preparation for surgery and
metastatic intra-abdominal cancer confirmed at surgery. It is usually a soli-
tary, firm, indurated nodule, sometimes with surface fissuring or ulceration;
it is variable in size and can be painful. Common underlying tumors include
gastrointestinal malignancy (>55%, male predominance) and ovarian malig-
nancies (34%).
Sister Mary Joseph nodules account for 60% of all malignant umbilical
tumors (primary or secondary). Most patients die within months after the
appearance of the umbilical tumor.
2. Microscopic: Most commonly, a dermal-based adenocarcinoma with histol-
ogy resembling the primary malignancy. Signet-ring cell morphology may be
present, especially from a gastric primary (e-Fig. 39.60).
B. Metastatic breast carcinoma
1. Clinical: Typically small papules, ranging from 1 to 2 mm in diameter to large
tumor masses, on the anterior chest wall. Intralymphatic spread of tumor
cells (inflammatory carcinoma) can manifest as a diffuse, warm, indurated
plaque (carcinoma erysipeloides). Scalp metastasis can present as alopecia
(alopecia neoplastica), which is also seen in metastases from other visceral
sites, especially from a lung primary.
2. Microscopic: Usually a poorly differentiated adenocarcinoma. Ductal and
sometimes lobular arrangements are present. The proportion of intravascular
tumor varies (e-Fig. 39.61).
C. Metastatic renal cell carcinoma
1. Clinical: Typically erythematous, vascular papule or nodule, may be misdiag-
nosed as lobular capillary hemangioma or Kaposi sarcoma; solitary in 15%
to 20% of cases. The scalp is a common site of involvement.
2. Microscopic: Usually well-circumscribed, dermal nodule(s) composed of
sheets and nests of polygonal tumor cells with clear cytoplasm, distinct cyto-
plasmic borders, and enlarged hyperchromatic nuclei; associated with a del-
icate rich vascular stroma, red blood cell extravasation, and hemosiderin
deposition. The histologic features can be bland, and the differential diagno-
sis includes other primary and metastatic tumors with clear cell morphology
(e-Fig. 39.62).
Skin: Melanocytic
Lesions
Anne C. Lind, Nils Becker, Emily A. Bantle,
and Louis P. Dehner
I. BACKGROUND. Melanocytes are melanin-synthesizing cells derived from the neural

crest. During the first 3 months of gestation, they migrate into the ectoderm where
they normally occupy a space slightly beneath the basal keratinocytes. They can be
differentiated from adjacent keratinocytes by their rounded and slightly hyperchro-
matic nucleus as compared with the more elongated or ovoid nucleus with evenly
dispersed chromatin of the basal keratinocyte. In addition, melanocytes usually
exhibit a pale often eccentric rim of eosinophilic cytoplasm and are separated from
the neighboring basal keratinocytes and/or basement membrane by a clear space
or halo (e-Fig. 40.1).* Occasionally, dendritic processes can be seen as they extend
between the keratinocytes. The ratio of melanocytes to basal keratinocytes depends
on the body site. On the trunk and extremities it is approximately 1:7 to 10, while
the ratio on the face and external genitalia is about 1:3.
The main function of melanocytes is the production of melanin, a tyrosine-
derived photoprotectant which converts UV radiation into heat, thus prevent-
ing UV-related DNA damage. Melanin is exported via melanosomes, which are
membrane-bound and have an internal lattice-like structure. Melanin granules are
deposited on the lattice, eventually obscuring this architectural feature. Melanin
pigment found in the skin is classified as eumelanin (brown or black pigment) or
pheomelanin (yellow-red pigment), the latter of which is rich in sulfur.
In addition to the general considerations for gross examination of skin spec-
imens (Chap. 38), a complete description of the background skin color and the
clinical "ABCDs" (see section on melanoma ) is required for biopsies or excisions
of a pigmented lesion. Therefore, the size of the surface lesion, distance to or
presence at the margin, color regularity or irregularity, and border, should all be
included in the gross description.
Diagnosis of a melanocytic lesion is based on multiple architectural and
cytologic criteria. In most cases, these microscopic features result in a specific
diagnosis that reliably correlates with the anticipated biologic behavior of the
melanocytic proliferation. Features associated with benignancy include symme-
try, circumscription, maturation, predominance of nested melanocytes, cohesive
nests of melanocytes, and melanocytes with regular nuclear borders and without
nucleoli or atypical mitoses. A comparison of the histologic criteria for benign and
malignant melanocytic proliferations can be found in Table 40.1. The status of
the margins should be included routinely on all pathology reports of melanocytic
proliferations, even in punch biopsy specimens. Although there admittedly is no
consensus on this point, the margin status provides additional information that is
often clinically useful.
Immunohistochemical stains can be helpful in distinguishing melanocytic from
nonmelanocytic lesions, confirming nodal micrometastases, or demonstrating con-
fluence or nonconfluence of melanocytes. Although several stains are available,
including S100, Melan-A (MART-1), HMB-45, MITF, and NKI-C3, no immunos-
tain is capable of reliably differentiating malignant from benign melanocytic
lesions.
613
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Chapter 40 • Skin: Melanocytic Lesions I 61 7
regardless of age or site. Pagetoid spread may be seen in an otherwise typical

Spitz nevus. Caution is required when considering the diagnosis of an epithe-
lioid Spitz nevus in a person past middle age and/or on sun-damaged skin.
The designation of "Spitz tumor" has been suggested for the Spitz nevus with
atypical features (e-Fig. 40.7), if not all Spitz nevi. Although the diagnostic
challenge presented by a melanocytic lesion with Spitz features is admittedly
difficult, the overwhelming majority of Spitz nevi occurring in children and
adolescents have benign behavior.
F. Pigmented spindle cell nevus of Reed
1. Clinical: A pigmented spindle-cell nevus of Reed is a heavily but evenly pig-
mented, small, acquired lesion in the shoulder/pelvic girdle region. It is most
common in women in their second and third decades.
2. Microscopic: It is a symmetric and well-circumscribed proliferation of spin-
dled melanocytes in the epidermis and superficial dermis with a pattern rem-
iniscent of a woven basket. Fascicles of spindled cells are oriented vertically,
horizontally, and tangentially. A broad zone of melanophages in the superfi-
cial dermis is invariably present beneath the melanocytes (e-Fig. 40.8). The
relationship of the Reed nevus to the Spitz nevus is uncertain.
G. Blue nevus
1. Clinical: It is a congenital or acquired, small lesion that is flat to slightly
raised. Unlike most other melanocytic lesions, it is blue rather than black or
brown. They occur most commonly on the head, neck, and extremities.
2. Microscopic: A common blue nevus is a superficial, dermal, lentil-shaped,
variably cellular proliferation of spindled and dendritic melanocytes
that interdigitate between collagen bundles. Melanophages are frequently
admixed. The epithelioid variant has, as the name implies, melanocytes with
a more epithelioid appearance (e-Fig. 40.9). The differential diagnosis of a
common blue nevus includes dermal melanocytosis.
H. Cellular blue nevus
1. Clinical: It is a large (1 to 2 em) heavily pigmented, blue to blue-black, raised
lesion that is most common on the scalp or buttocks.
2. Microscopic: This lesion is characterized as a mid- to deep-dermal multinodu-
lar biphasic proliferation. The nodular centers are composed of small epithe-
lioid melanocytes, with pale to amphophilic cytoplasm. The nodules are sur-
rounded by a rim of pigmented spindled melanocytes and melanophages.
There may be extension of the nodules into the contiguous subcutis in some
cases. Despite the striking tumor-like appearance, a consideration of malig-
nancy should not be entertained in the absence of necrosis, atypical mitosis,
and prominent nucleoli (e-Fig. 40.10).
I. Dermal melanocytosis {Nevus of Ota, Nevus of Ito, Mongolian spot, Hori nevus)
1. Clinical: These nevi are characteristically flat, slate-gray to blue to dark brown
patches on the face, shoulder, sacrum, or bilateral temples. They predomi-
nantly occur in people of Asian descent and are more common in females.
2. Microscopic: These lesions have identical features and some overlap with
those of a blue nevus. Instead of a superficial lentil-shaped lesion they are
present in the mid- to deep-dermis as ill-defined, paucicellular infiltrates of
pigmented, spindled, and/or dendritic melanocytes (e-Fig. 40.11).
J. Congenital and congenital-paHern nevi
1. Clinical: Congenital or congenital-pattern nevi are pigmented lesions that
are usually present at birth or that appear during infancy. They are classified
as small (<2 em), intermediate (2 to 20 em), or large (>20 em). They are
frequently varied in pigmentation and have irregular borders. These nevi
may be hairy and/or become progressively more hairy. Involvement of the
leptomeninges by benign or malignant melanocytes in a child with a large or
giant congenital nevus is known as neurocutaneous melanosis.
2. Microscopic: A junctional component exists in these lesions in early infancy,

but the melanocytes are predominantly dermal with time. Individual nevoid
to small epithelioid melanocytes in the dermis extend along or into dermal
structures (such as the adnexa, arrector pili muscles, and small peripheral
nerves) and into lymphovascular spaces. The subcutis and deeper structures
may also be infiltrated by nevus cells. The epidermal melanocytic pattern
is highly variable, ranging from no significant melanocytic proliferation, to
nested melanocytes, to confluent lentiginous hyperplasia. Mitotic figures,
but not atypical forms, may be present at any level. Proliferative nodules
composed of monomorphous melanocytes and with brisk mitotic activity
may be present and might lead to an erroneous interpretation of melanoma.
An apparent congenital nevus, especially from the scalp, with a more
complex pattern of spindle cells, perivascular pseudorosettes, and tactoid
bodies likely represents a so-called neurocristic hamartoma.
A congenital-pattern nevus is regarded as an acquired lesion that has
a dermal growth pattern similar to that of a congenital nevus, including a
predominance of individual cells rather than nests, and preferential growth
in or along adnexal structures (e-Fig. 40.12).
K. Deep penetrating nevus
1. Clinical: It is a pigmented lesion that is usually acquired during early adult-
hood. It is of small size and found most commonly on the upper half of the
body. Occasionally, it may resemble a blue nevus.
2. Microscopic: Deep penetrating nevi show dermal fascicles and nodules of
epithelioid to spindled melanocytes with conspicuous granulat; gray-brown
cytoplasm admixed with melanophages and some nevoid melanocytes. They
may extend to the subcutis in a mixed pushing and infiltrative pattern.
Nuclear pleomorphism, mitotic figures, and nucleoli may be noted. This
lesion may be seen as one component of a combined nevus. The combina-
tion of the cytologic and architectural features may lead to a concern for
melanoma (e-Fig. 40.13).
L. Halo nevus
1. Clinical: A halo nevus shows a characteristic central zone of pigment with
a circumferential rim of hypo-/depigmentation. It may be raised or flat and
sometimes is noted as a recent change in a preexisting pigmented lesion.
2. Microscopic: Sections show a junctional, compound, or intradermal
melanocytic nevus which has been obscured or nearly obscured by a dense
band of lymphocytes. Regressive changes (immature fibroplasia, neovascu-
larization, paucicellular lymphocytic infiltrate, melanophages) may be seen at
the perimeter. The lymphocytic infiltrate should not impart a sense of asym-
metry, and the epidermal melanocytes, if present, should be predominantly
nested (e-Fig. 40.14).
M. Nevus with architectural disorder/Clark nevus/dysplastic nevus
1. Clinical: Nevi with architectural disorder are highly variable in appearance,
ranging from small, symmetric, and evenly pigmented to large (>6 mm),
irregularly shaped, and irregularly pigmented. Nevi with architectural disor-
der are a cutaneous marker for the dysplastic nevus syndrome where multiple
atypical nevi (> 100) can be present. Nevi with architectural disorder are a
controversial entity of uncertain significance outside of this syndrome. The
lack of uniformity in diagnostic terms is one sign of this controversy.
2. Microscopic: This junctional or compound melanocytic proliferation has its
diagnostic features centered on the epidermal-dermal junction. The nested
melanocytes bridge between adjacent rete, nested melanocytes arise between
or from the sides of rete (rather than the tips), eosinophilic collagenous tis-
sue drapes beneath the epidermis in a festoon-like pattern (lamellar fibro-
plasia), and there is nonconfluent lentiginous melanocytic hyperplasia. The
Chapter 40 • Skin: Melanocytic Lesions I 61 9
melanocytes may have a bland, nevoid appearance; have a size greater than
that of the basal keratinocytes (mild cytologic atypia); have increased size and
variable chromatin patterns (moderate cytologic atypia); or have increased
size, variable chromatin patterns, pleomorphism, and nucleoli (severe cyto-
logic atypia). A nevus with architectural disorder and severe cytologic atypia
should be considered a borderline melanocytic proliferation; a conservative
but complete excision should be encouraged (e-Fig. 40.15).
N. Nevi on special sites
1. Clinical: The clinical appearance varies with site. Special sites include umbili-
cus, acral skin (palms/soles), areola, and mucocutaneous sites such as the
conjunctiva, anus, and external genitalia.
2. Microscopic: In addition to the features associated with junctional and/or
compound nevi, the junctional melanocytic proliferations at these sites may
have an increase in the number of individual melanocytes with either a lentig-
inous or pagetoid pattern, loss of cohesion, and asymmetry.
0. Combined melanocytic nevus
1. Clinical: They vary in appearance. They may be flat, raised, or both and
may be variably pigmented. Although they can present at any age, they are
most common in the first three decades. The combination of features in these
lesions (asymmetry, color variations, and possible border irregularities) often
raises concern for melanoma.
2. Microscopic: Combined melanocytic nevi may have any of the features
of the previously described nevi in a side-by-side arrangement or top-to-
bottom arrangement. One of the more common combinations is a com-
pound melanocytic nevus and a blue nevus; the most worrisome combina-
tions include a compound melanocytic nevus with a deep penetrating nevus,
or a side-by-side Spitz nevus and compound nevus.
P. Recurrent melanocytic proliferations
1. Clinical: These are characterized by a sudden appearance or reappearance
of pigment in a scar from a previously biopsied or excised nevus and/or
melanoma. In some instances, the patient may not be able to provide reliable
information regarding the biologic nature and/or diagnosis of the previous
lesion.
2. Microscopic: Lentiginous melanocytic hyperplasia and randomly scattered,
irregular nests of melanocytes are present above an immature or mature
dermal scat. The features of a recurrent nevus may be indistinguishable from
a recurrent melanoma and have been referred to as "pseudomelanoma."
Adjacent lesional tissue outside of the scar and/or knowledge of the initial
biopsy or excision findings is imperative for arriving at the correct diagnosis
(e-Fig. 40.16).
Q. Nevus ambiguous. This category could include any/all melanocytic lesions that
have some of the classic features of a distinctive subtype (such as a Spitz nevus)
but are variant in others.
Ill. MELANOMA is a malignant melanocytic neoplasm that can occur in any tissue on any
body site. Ninety percent of all melanomas arise in the skin, and only cutaneous
melanoma will be considered in this section.
Melanoma accounts for only a small percentage (3% to 5%) of primary cuta-
neous malignancies; basal cell carcinoma and squamous cell carcinoma are far more
prevalent (95% to 97%). However, melanoma accounts for >50% of cancer deaths
from a primary cutaneous malignancy. The incidence of melanoma has increased
over the past 25 years, but the recognition and diagnosis of lower stage lesions
account for a substantial proportion of this increased incidence. As expected, the
overall survival has shown improvement as pathologically lower stage lesions are
diagnosed. Melanoma is most common in individuals >50 years of age but affects
persons of all ages, including infants. It is slightly more common in males, in whom
the pattern of distribution is slightly different. In males, melanoma is more com-

mon on the trunk/head and neck; in females, melanomas on the lower extremities
are more common. There are numerous histologic types (see Table 40.3), and early
study results suggest that the genetic signature of melanoma is different depend-
ing on its association with sun exposure (chronic, intermittent, or none; N Engl
J Med. 2005;353:2135). While the majority of melanomas have easily recognized
histologic patterns, some subtypes, such as nevoid and spitzoid, are unsettlingly
similar to benign melanocytic lesions. In addition, amelanotic melanomas have a
clinically more challenging appearance due to their lack of pigmentation. They are
considered to be poorly differentiated melanomas.
Clinical features may vary slightly with anatomic site and/or histologic subtype;
howevet; in general, a melanoma is Asymmetric, has an irregular Border, its Color
is uneven, and it has a Diameter that is >6 mm. These clinical features have been
described as the ABCDs of pigmented lesions. The patient may report a change in
a preexisting pigmented lesion and/or complain of pruritus. A clinical description
of a worrisome pigmented lesion can be helpful, especially in the case of a biopsy
with borderline atypical histologic features. It should always be considered that a
partial biopsy may not be representative of the overall pathology.
A. Melanoma in situ
1. Microscopic: There is an increase in the number of atypical, enlarged
melanocytes found only in the epidermis. These cells show a spectrum of
nuclear enlargement and hyperchromatism. There may be prominent page-
toid spread (i.e., melanocytes are present above the suprapapillary plate) in a
superficial spreading pattern, or confluent spread of melanocytes at the level
of the basal keratinocytes in a lentiginous pattern. Nests may or may not be
present, but if present are randomly distributed and vary in size and shape.
Melanocytes are not seen in the dermis, although features of regression may
be seen and warrant a comment because invasion may have been present
earlier (e-Fig. 40.17).
2. Differential diagnosis: Sun-induced melanocytic hyperplasia, recurrent
melanocytic proliferations, lentiginous dysplastic melanocytic nevus, page-
toid Spitz nevus, and psoralen and ultraviolet A light (PUVA) lentigo.
B. Malignant melanoma-superficial spreading type
1. Clinical: Superficial spreading melanoma often occurs on the trunk of men
and lower extremities of women; however, it can be found on any body site.
2. Microscopic: Increased numbers of melanocytes with enlarged nuclei and
increased amounts of cytoplasm are present at all levels of the epidermis,
pagetoid spread is easily identified, and melanocytes with similar cytologic
features are present in the dermis. In the dermis, melanocytes may be seen as
individual cells, in small dusters, and/or in sheets (e-Fig. 40.18).
3. Differential diagnosis: Compound melanocytic nevus.
C. Malignant melanoma-lentiginous type (lentigo maligna melanoma)
1. Clinical: This type usually develops on the face or sun-exposed areas of the
upper extremities of elderly patients.
2. Microscopic: Increased numbers of melanocytes with enlarged nuclei and a
mild increase in cytoplasm are present in a confluent pattern at the level of
the basal keratinocytes without sparing of the suprapapillary plates. Poorly
formed and randomly scattered nests of melanocytes may be present; page-
toid cells are not seen. An artifactual subepidermal cleft might be noted, and
melanocytes might extend along the adnexal epithelium. In this variant, cyto-
logic atypia might not be prominent. Melanocytes are present in the dermis
(e-Fig. 40.19).
3. Differential diagnosis: Sun-induced melanocytic hyperplasia, recurrent
melanocytic proliferations (benign or malignant), and lentiginous dysplas-
tic melanocytic nevus.
Chapter 40 • Skin: Melanocytic Lesions I 621
D. Malignant melanoma-nodular type

1. Clinical: These can be found on any body site. Characteristically, they lack
an initial radial growth phase, leading to a nodular or polypoid appearance.
2. Microscopic: Melanocytes with atypical, but often monotonous, nuclear and
cytoplasmic features including enlarged nuclei, irregular nuclear borders,
increased amounts of cytoplasm, and abundant mitotic activity (including
atypical forms) are present in the dermis in a large nodule or sheet. There may
be no apparent intraepidermal melanocytic proliferation, and if malignant
melanocytes are present in the epidermis, they do not span the entire dermal
tumor. Ulceration is frequently noted (e-Fig. 40.20).
E. Malignant melanoma-desmoplastic-neurotropic type
1. Clinical: It has a predilection for the head and neck region and is often non-
pigmented.
2. Microscopic: There is a predominantly dermal proliferation of individ-
ual, large, spindled melanocytes with prominent nuclear pleomorphism.
Melanocytes are noted between collagen bundles and along nerves. The lesion
may be poorly defined and variably cellular. Unlike most types of cutaneous
melanoma, the desmoplastic melanoma may be S-1 00 negative and gener-
ally is nonreactive with other markers of melanocytic differentiation such as
Melan-NMART-1 and/or HMB-45. Apparent expression of smooth muscle
actin (SMA) has been noted (e-Fig. 40.21).
3. Differential diagnosis: Malignant fibrous histiocytoma, scar, dermatofibroma,
sclerotic dermatofibrosarcoma protuberans, and leiomyosarcoma.
F. Malignant melanoma-nevoid type
1. Microscopic: Sections show a symmetric, well-circumscribed, nested pro-
liferation of small, nevoid melanocytes with dermal maturation. At low
power, it may be indistinguishable from a compound melanocytic nevus. At
higher power, deep dermal melanocytes have prominent nucleoli and mitoses
(e-Fig. 40.22).
2. Differential diagnosis: Melanocytic nevus.
G. Malignant melanoma-spitzoid type
1. Microscopic: Epithelioid and spindled melanocytes in a predominantly nested
pattern are seen in an acanthotic and hyperkeratotic epidermis. The epider-
mal architectural features are strikingly similar to a Spitz nevus. Dermal
melanocytes may be preferentially present in sheets or in large confluent
nests, and lack maturation. There is a degree of nuclear pleomorphism that
exceeds the atypia seen in the usual Spitz nevus. Mitotic figures, including
atypical forms, may be present at all levels of the dermis and may be asym-
metrically distributed. A lymphocytic response, if present, may abut the deep
aspect of the lesion and provide a suggestion of asymmetry. Frequently the
nuclear to cytoplasmic ratio is greater than that of a Spitz nevus, and there is
no apparent decrease in nuclear size between the superficial and deep dermal
melanocytes (e-Fig. 40.23).
2. Differential diagnosis: Spitz nevus.
IV. MICROSCOPIC STAGING OF MELANOMA. The Tumor, Node, Metastasis (TNM) clas-
sification of malignant melanoma as published in the American Joint Committee
on Cancer (AJCC) guidelines from 2010 is provided in Table 40.4. The following
changes have been made from the 2002 guidelines: The use of the mitotic rate
per mm2 instead of Clark level for categorizing T1 melanomas, the inclusion of at
least one melanoma-associated immunohistochemical marker in the detection of
nodal metastases (unless a diagnosis can be made by cellular morphology), and the
removal of the prior existing 0.2 mm threshold for positive lymph nodes.
A. Breslow thickness is the single most important prognostic parameter associ-
ated with an invasive melanoma. An optical micrometer, calibrated to the
microscope, is aligned perpendicularly to the epidermis, and the melanoma is
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V. NONMELANOCYTIC PIGMENTED LESIONS. The pathologist is sometimes presented

with a biopsy or excision of skin for a clinically pigmented lesion, often with the
clinical description of "atypical pigmented lesion; rule out melanoma." Lesions
that are not composed of melanocytes microscopically, but can mimic a nevus
or melanoma clinically, include: pigmented seborrheic keratosis (e-Fig. 40.24),
pigmented basal cell carcinoma (e-Fig. 40.25), pigmented actinic keratosis, solar
lentigo (e-Fig. 40.26), dermatofibroma (e-Fig. 40.27), postinflammatory pig-
mentary alteration (e-Fig. 40.28), intracorneal hemorrhage (calcaneal petechiae)
(e-Fig. 40.29), tinea nigra (e-Fig. 40.30) and Monsel's solution in a biopsy site
(e-Fig. 40.31).
SUGGESTED READINGS
Banerjee SS, Harris M. Morphological and immunophenotypic variations in malignant melanoma.
Histopathology. 2000;36:387-402.
Barnhill RL, Piepkorn M, Busam KJ. Pathology of Melanocytic Nevi and Malignant Melanoma.
2nd ed. New York, NY: Springer; 2004.
Crowson AN, Magro CM, Mihm MC. The Melanocytic Proliferations. A Comprehensive Textbook
of Pigmented Lesions. New York, NY: Wiley-Liss; 2001.
Culpepper KS, Granter SR, McKee PH. My approach to atypical melanocytic lesions. JClin Pathol.
2004;57:1121-1131.
Elder D, van den Oord ]. Pathology and pathophysiology of melanocytic disorder. Histopathology.
2002;41(Suppl. 2):120-146.
Massi G, Leboit PE. Histological Diagnosis of Nevi and Melanoma. Berlin, MA: Steinkopff Darm-
stadt/Springer; 2004.
Thompson JF, Morton DL, Kroon BBR. Textbook of Melanoma. New York, NY: Martin Dunitz;
2004.
SECTION IX
Nervous System
Central Nervous
System: Brain, Spinal
Cord, and Meninges
I. INTRODUCTION. The task of evaluating a neuropathologic specimen often seems

daunting, given the complexity of this organ system and its ever-enlarging list of
diseases. However, when a methodical approach is applied using clinical, radio-
logic, histological and, increasingly, molecular information, the chances of error
can be significantly reduced.
II. ANATOMY AND HISTOLOGY. The central nervous system (CNS) consists of cere-
brum, cerebellum, brain stem, spinal cord, meninges, 12 paired cranial nerves,
and the blood vessels supplying these structures. The brain and spinal cord are
enclosed within the skeletal confines of the cranium and vertebral canal. The
mature (adult) brain weighs around 1200 to 1400 g. The meninges covering the
brain and spinal cord are of two principal types: (1) the dense fibrous dura mater
and (2) the more delicate leptomeninges (pia and arachnoid mater). The cerebrum
is divided into right and left hemispheres by a thick dural fold, the falx cerebri. A
second dural fold between the cerebrum and the cerebellum (tentorium cerebelli)
divides the brain into infra- and supratentorial compartments. Infratentorially,
the midbrain, pons, and medulla oblongata (cranial to caudal) form the brain
stem, which is connected to the cerebellum by means of three (superior, mid-
dle, and inferior) cerebellar peduncles. The supratentorial compartment contains
cerebral cortex (frontal, temporal, parietal, and occipital lobes), white matter, and
deep gray nuclei, such as basal ganglia, thalamus, and hypothalamus. The tenn
"neuraxis" is sometimes used to refer to brain and spinal cord; thus, lesions that
involve brain parenchyma are said to be "intra-axial" (e.g., astrocytoma, central
neurocytoma, ependymoma), and those located outside of the parenchyma are
referred to as "extra-axial" (e.g., meningioma, hemangiopericytoma [HPC], soli-
tary fibrous tumor [SFf], and schwannoma of cranial nerve VIII). Similarly, in
the spine, the terms "intramedullary" and "extramedullary" are used to denote
lesions within or adjacent to the spinal cord parenchyma, respectively.
The CNS is composed of two tissue types, namely gray and white matter, that
differ qualitatively on gross and microscopic examination. Neuronal cell bodies
and dendrites reside mostly in the gray matter (cortex and deep gray nuclei), and
axons create the framework of the white matter. Glial cells (astrocytes, oligoden-
droglia! cells, and microglia) are present in different proportions in these tissues.
Oligodendrocytes are more populous in the white matter; their processes fonn
the sheaths of myelin that insulate CNS axons. The eosinophilic, finely granu-
lar to fibrillary material between cell bodies is often referred to as "neuropil"
625
626 I SECTION IX: NERVOUS SYSTEM
and is formed by the processes of neurons (axons and dendrites) and glial
cells. Neuronal morphology varies significantly, with cell body size ranging from
< 15 f.'ID (e.g., small neocortical granular stellate neurons) to 100 /A-ID (Betz cells of
the primary motor cortex). For descriptive purposes, neocortical pyramidal neu-
rons are often considered the morphologic prototype. These cells contain abun-
dant amphophilic cytoplasm, clumpy basophilic Nissl substance, a large round
central nucleus, a prominent nucleolus, coarse proximal cytoplasmic processes,
and a prominent apical dendrite oriented perpendicular to the cortical surface.
Ependymal glial cells form a ciliated cuboidal epithelium that lines the ventricles
and central canal and focally transitions with epithelium of the choroid plexus.
The choroid plexus, which produces cerebrospinal fluid (CSF) within the ven-
tricles, is papillary; its branching fibrovascular cores are lined by a specialized
epithelium with a hobnailed apical surface.
Ill. INTRAOPERATIVE EVALUATION, GROSS EXAMINATION, AND TISSUE SAMPLING. Eval-
uation of a surgical neuropathology specimen often begins with an intraoperative
consultation, which may be requested by the surgeon (1) to confirm the presence of
lesional tissue, (2) to provide a preliminary diagnosis that will guide surgical man-
agement (e.g., aggressive surgery for ependymoma, limited biopsy for lymphoma,
culture sample to microbiology for abscess), and (3) to sample fresh or frozen tis-
sue for ancillary studies (e.g., Western blot for Creutzfeldt-Jakob disease [CJD],
molecular pathology, tumor banking, karyotyping). For optimal evaluation, spec-
imens should ideally be submitted on Telfa nonstick gauze pads saturated with
normal saline. Tissue that has been soaked in saline is certainly acceptable, but is
more likely to fragment during transport and may demonstrate more severe freez-
ing artifacts. Fresh brain tissue, especially small biopsy specimens, should never
be placed on dry gauze or tissue paper, because subsequent tissue retrieval from
these materials is almost impossible. Water content may be reduced through very
gentle blotting on a clean dry plastic surface, but fresh brain tissue is very fragile,
and improper handling can introduce cellular "touch" and "crush" artifacts.
For intraoperative diagnosis, small portions of the fresh specimen should be
chosen for freezing, for cytologic "smear" or "touch" preparations, and for possi-
ble ultrastructural examination. Freezing or otherwise exhausting the entire spec-
imen for intraoperative diagnosis should be avoided, for several reasons. First,
the techniques available during intraoperative examination seldom yield suffi-
cient information for a definitive final diagnosis; most diagnoses require the fine
histologic detail afforded by paraffin sections and information from immunohis-
tochemical stains and other ancillary molecular tests. Second, freezing introduces
artifacts (e.g., ice crystals, clumping of nuclear chromatin). Third, on some occa-
sions, surgical attempts to obtain additional diagnostic material from the patient
cause bleeding or other complications that prevent further tissue acquisition.
The manner in which intraoperative specimens are processed for diagnosis is
important. Before any tissue is frozen, a block of embedding compound (com-
monly referred to as "OCT"), formed within an empty tissue well within the
cryostat, should be frozen fast to a cryostat "chuck." After freezing, the chuck
and OCT block are removed sharply and inverted; the tissue should be placed
centrally on the flat surface of the frozen OCT block, immediately covered with
a minimal amount of liquid OCT, and frozen from above by a flat, prechilled
metallic weight (e-Fig. 41.1)."' This process maximizes rate of freezing and min-
imizes (but does not eliminate) ice crystal formation. Cytologic evaluation by
smear preparation is extremely helpful because it lacks freezing artifacts and
preserves nuclear details. Smears can be prepared by gently compressing a very
small amount of representative tissue between two glass slides, gently sliding them
apart, and immediately fixing both smeared slides in 95% alcohol (e-Fig. 41.2).
Lastly, a small tissue fragment (1 mm3 ) should also be fixed in glutaraldehyde and
Chapter 41 • Central Nervous System: Brain, Spinal Cord, and Meninges I 6 27
processed for potential electron microscopic studies, particularly if the intraoper-

ative diagnosis is unclear.
Gross examination and tissue sampling for permanent sections is less com-
plicated. Most neuropathologic specimens are small and/or fragmented, limiting
full appreciation of meaningful gross features. Even when large resection spec-
imens are submitted intact, gross abnormalities are often absent or subtle. In
fact, radiographs are commonly considered the "gross pathology" for CNS biop-
sies. In either case, specimens are usually entirely submitted for histologic analysis
after adequate formalin fixation (a few hours for smaller specimens and overnight
fixation for large specimens). Because most neuropathologic processes are hetero-
geneous, even when the diagnosis seems clear, if a resection specimen is too large
for complete processing, it should still be extensively sampled. For similar rea-
sons, Cavitronic UltraSound Aspirator (CUSA) material should not be dismissed
as worthless; such material may be somewhat less preserved than resected tissue
due to partial autolysis and other artifacts, but occasionally it provides essential
clues to the final diagnosis.
A. CNS biopsy for special circumstances. Brain and sometimes meningeal biop-
sies for nonneoplastic indications are occasionally performed (e.g., nonresolv-
ing chronic meningitis, neurosarcoidosis). Similarly, a "blind" frontal lobe
biopsy is sometimes obtained for neurodegenerative disorders that do not
have a clearly defined etiology, particularly in younger patients. When a prion
protein disease like CJD is suspected, the neuropathologist should be given
advance notice and should be involved from the outset. One piece of cortex
from the biopsy should be snap frozen for Western blot analysis by a reference
laboratory such as the National Prion Disease Pathology Surveillance Cen-
ter (NPDPSC) (special shipping containers must be used, and specific proce-
dures must be followed; see http://www.cjdsurveillance.com/). The remaining
tissue should be fixed in 10% neutral buffered formalin for 24 hours, fol-
lowed by immersion in 88% to 98% formic acid (undiluted stock solution)
for 1 hour, prior to routine processing. If initial histologic examination reveals
pathologic features consistent with CJD or fails to suggest an alternative diag-
nosis to explain the clinical findings, paraffin-embedded material (blocks or
unstained slides) must accompany the frozen specimen to the reference labora-
tory to allow immunohistochemistry to be performed. C]D pathology can be
patchy within the brain (even when the abnormal protein is widespread), and
rare cases of C]D are caused by a protease-sensitive prion requiring immuno-
histochemical rather than immunoblot analysis for definitive diagnosis (Ann
Neurol. 2010;68:162). Lab equipment (gloves, instruments, etc.) are decon-
taminated with 1N sodium hydroxide solution for a minimum of 1 hour or,
alternatively, in 10% or 20% bleach solution for 1 hour, followed by auto-
claving. Many disinfection protocols that may be more or less appropriate for
particular circumstances have been reported (Infect Control Hosp Epidemiol.
2010;31:107). Frozen tissue diagnosis should not be attempted on tissues sus-
pected of prion protein diseases.
B. Ancillary studies
1. Electron microscopy {EM). The utilization of EM for diagnosis of CNS lesions
is labor intensive, time-consuming, and expensive, and it is mainly used
to evaluate nerve and muscle biopsies. However, ultrastructural evalua-
tion remains invaluable for many other neuropathologic diagnoses, includ-
ing CADASIL (see Section VIII.D), neuronal ceroid lipofuscinoses, and for
distinguishing ambiguous brain tumor cases, particularly meningioma and
ependymoma.
2. Immunohistochemistry is now routinely used in evaluation of complex surgi-
cal neuropathology cases, especially in the area of tumor neuropathology.
Frequently utilized antibodies and their immunoreactivity for common
tumor types are summarized in Table 41.1.
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Chapter 41 • Central Nervous System: Brain, Spinal Cord, and Meninges I 629
a. Glial markers. The most commonly used glial marker in neuropathology

practice is glial fibrillary acid protein (GFAP). This intermediate filament
protein is fairly (but not completely) specific for glial lineage. However,
it does not reliably distinguish astrocytic, oligodendroglia!, and ependy-
mal tumors from one another, and may also be encountered in other
tumors with glial differentiation such as choroid plexus tumors, medul-
loblastomas and/or primitive neuroectodermal tumors (PNETs), gangli-
ogliomas (GGs). GFAP can even be detected to some extent in nonglial
neoplasms, such as nerve sheath and cartilaginous tumors.
b. Neuronal markers. The most commonly used neuronal markers include
neurofilament (NF) protein, synaptophysin (SYN), chromogranin, and
Neu-N.
SYN is one of the more sensitive markers of neuronal differentiation.
It is typically found even in the most primitive neuronal tumors (medul-
loblastomas and PNETs) and is very useful for highlighting neoplastic
ganglion cells, pituitary adenomas and carcinomas, carcinoid tumors,
neurocytomas, and paragangliomas. One major disadvantage of SYN
is that it fails to differentiate native (entrapped) neuropil from tumor
neuropil. Additionally SYN stains a proportion of tumors (e.g., pilocytic
astrocytomas [PAs]) that are generally not considered to have neuronal
differentiation. In some cases, however, this feature is actually somewhat
useful; classic oligodendrogliomas show dot-like paranuclear reactivity
for SYN.
NF is a heteropolymer unique to neurons and axons. Mature neu-
ronal tumors, such as GGs, often stain for NF; however, more primitive
neuronal tumors such as medulloblastomas are often negative. Addition-
ally, NF also stains normal axons, a property that is of great utility for
demonstrating an infiltrative growth pattern by highlighting entrapped
axons (e-Fig. 41.3).
Neu-N is a marker of advanced neuronal differentiation; it has the
advantage of clearly marking neuronal nuclei and cell bodies rather than
surrounding neuropil. As a result, it is particularly useful for identifying
architectural abnormalities in cortical dysplasia, and for highlighting
neuronal loss (e.g., in mesial temporal sclerosis) and neurons entrapped
within invasive tumors. Surprisingly, most neoplastic ganglion cells in
GGs are negative for Neu-N; this feature can be useful because entrapped
cortical neurons are virtually always strongly positive.
c. Epithelial markers. The commonly used epithelial markers include cytok-
eratin (CK; AE1/AE3), epithelial membrane antigen (EMA), and CAM
5 .2. CKs are used predominantly in the diagnosis of metastatic carci-
nomas, but are also used to identify craniopharyngiomas, chordomas,
and choroid plexus tumors. Due to cross-reactivity with GFAP, gliomas
(and reactive astrocytes) may show CK reactivity, a major pitfall in the
differential between glioblastoma (GBM) and metastatic carcinoma. In
such instances, CAM 5.2 is recommended, because gliomas (and reactive
astrocytes) are virtually always negative. EMA is frequently used in the
identification of meningiomas, which, unlike true epithelial tumors, usu-
ally display minimal to no CK expression (secretory meningioma is an
exception). EMA is also useful in the diagnosis of ependymomas, along
with CD99 and D2-40 antibody (podoplanin).
d. S-1 DO protein is a marker of neuroectodermal cells, including
melanocytes, glia, Schwarm cells, chondrocytes, and the sustentacu-
lar cells in tumors such as paraganglioma, pheochromocytoma, and
olfactory neuroblastoma. In conjunction with collagen IV, which stains
basement membranes, S-100 is particularly helpful for demonstrating
Schwann cell differentiation in benign and malignant peripheral nerve

sheath tumors (MPNSTs).
e. Proliferation markers are used in conjunction with mitotic counts in brain
tumors to guide determination of tumor grade and prognosis. The most
widely used marker is Ki-67, which labels nuclei that are not in the Go
phase of the cell cycle.
f. "Molecular test" markers. Recently, several antibodies have been devel-
oped that can be used in lieu of more complicated molecular tests for
diagnosis and prognosis of various neoplasms.
i. INI1/BAF47 deletions are detected in "'70% of the atypical tera-
toid/rhabdoid tumors (ATIRTs); however, loss of the corresponding
INil protein is even more common than the genetic alteration. Loss
of INI1 nuclear immunoreactivity in tumor cells is used as a surro-
gate for genetic testing to demonstrate biallelic inactivation of the
gene.
ii. lsocitrate dehydrogenase (IDH 1/IDH2} mutations have been detected in
the majority of diffuse gliomas, with the notable exception of pri-
mary (de novo) GBM, and appear to play a fundamental and early
role in oncogenesis (N Eng] Med. 2009;360:765). The most com-
mon mutation is in the IDH1 gene (R132H), and is recognized by
monoclonal antibody IDH-1. Diagnostically, this immunohistochem-
ical stain shows greatest promise for its potential to distinguish low-
grade diffuse glioma from gliosis (Am J Surg Pathol. 2010;34:1199;
Acta Neuropathol. 2010;119:509). Prognostically, the presence of
this mutation is favorable, as such tumors appear to show greater
response to therapy.
3. Molecular diagnostics involves the measurement of diagnostically or prog-
nostically relevant pathologic features at the DNA (epigenetic, genomic
[nuclear or mitochondrial], mRNA, or protein levels.
The most common and practical approaches to detect changes in DNA
(deletions of chromosomal regions, amplifications of oncogenes, or loss of
specific tumor suppressor genes) include fluorescence in situ hybridization
(FISH) and quantitative polymerase chain reaction (PCR) techniques. FISH
has the advantages of simplicity, morphologic preservation, and minimal
tissue and purity requirements. However, FISH is insensitive to very small
deletions/amplifications, substitution mutations, or epigenetic modification
(e.g., methylation).
The most notable use of FISH in surgical neuropathology is to detect
losses of chromosomal arms 1p and 19q testing as a prognostic and/or
management tool for adult patients with oligodendroglia! tumors (Adv
Anat Pathol. 2005;12:180). Other clinical applications of FISH include
detection of EGFR amplification and/or 1Oq deletions to distinguish the
small cell variant of GBM from anaplastic oligodendroglioma; 22q11.2
deletion (INI1 locus) to distinguish ATIRT from variants of medulloblas-
toma (Hum Pathol. 2001;32:156); isochromosome 17q (i17q) and NMYC
or CMYC amplifications to diagnose and predict outcome for medulloblas-
tomas, large cell/anaplastic medulloblastomas and other aggressive forms
of CNS-PNET; meningioma-associated deletions (NF2, DAL1, 1p, 14q)
to distinguish anaplastic meningiomas from other malignancies or benign
meningiomas from foci of meningothelial hyperplasia; and (9p21) to pro-
vide prognostic information about higher-grade meningiomas.
A common use ofPCR-based techniques is to detect 06-methylguanine-
DNA methyltransferase (MGMT) gene methylation, which blocks MGMT
transcription. As the MGMT gene product plays an important role in DNA
repair, GBMs with this epigenetic modification show greater sensitivity to
alkylating agents (like temozolomide) and radiation therapy (N Eng] Med.

2005;352:987).
IV. BASIC ELEMENTS OF CNS PATHOLOGY. The cells and tissues of the CNS are capable
of displaying diverse histologic abnormalities. A few of these are pathognomonic
for a given disease, but most diseases require a constellation of findings to suggest
a diagnosis.
A. Neurons.
1. Axonal injury. When an axon is damaged and the associated neuron survives,
the axon itself may form a swelling, or axonal spheroid, at the site of injury.
If the axon is severed, in most cases the distal portion will disintegrate
in an active cellular process called Wallerian degeneration. In some cases,
the axotomized neuron will undergo "chromatolysis," in which the cell
body appears mildly swollen and achromatic; this change reflects a loss of
Nissl substance that occurs as the cell alters its metabolism to allow repair
of its damaged axon.
2. Apoptosis. If neuronal injury is severe enough to cause cell death, neu-
rons may undergo apoptosis (as occurs in the basis pontis and subiculum
from hypoxic ischemic injury late in gestation, a pattern labeled with the
misnomer pontosubicular necrosis). Alternatively, neurons may undergo
necrOSIS.
3. Necrosis. The classic histologic appearance of acute neuronal necrosis
includes (1) variably intense cytoplasmic eosinophilia (accounting for the
name "red neurons") and (2) shrunken pyknotic nuclei (e-Fig. 41.4). Red
neurons require 12 to 24 hours to develop within a living brain; individ-
uals who die within minutes or a few hours of an ischemic stroke do not
show red neurons in affected region(s). Although red neurons are commonly
caused by ischemia, many insults (e.g., hypoxia, hypoglycemia, epilepsy,
herpes simplex virus [HSV] infection) can also cause neuronal necrosis. It
is important to note that a common artifact caused by overmanipulation
of fresh brain tissue can cause normal healthy neurons to (superficially)
resemble red neurons. Neurons affected by this "dark cell change" usually
show more basophilia than red neurons, a nucleus that is less distinct within
the cell body, and an apical dendrite that resembles a spiral or corkscrew
(e-Fig. 41.5).
4. Ferrugination. Occasionally, damaged neurons around the edge of a remote
infarct or traumatic injury become encrusted with basophilic iron and cal-
cium salts. This condition is often referred to as mineralization or ferrugi-
nation.
5. Binucleation of neurons, rare in normal brains, is infrequently noted in
dysplastidmalformative processes (e.g., tuberous sclerosis, TS), in certain
neoplasms (e.g., GG), and in Alzheimer disease (AD) (Neuropathol Appl
Neurobiol. 2008;34:457) (e-Fig. 41.6).
6. lntraneuronal inclusion bodies form within neurons under many different
circumstances. Some are pathognomonic, some are associated with one or
more diseases, and others appear to have no pathologic significance.
a. Pick bodies are round, tau-positive intracytoplasmic neuronal inclusions.
In Pick disease (a form of frontotemporal dementia), these are argy-
rophilic by Bielschowsky and Bodian (but not Gallyas) silver stains and
are abundant in neurons of the cortex, hippocampus, and dentate gyrus.
A similar (but less abundant, Gallyas positive) inclusion is seen in corti-
cobasal degeneration.
b. Lewy bodies (LBs). Classic LBs, which occur within pigmented neurons of
the brainstem, are spherical cytoplasmic inclusions with an eosinophilic
core and a pale halo (e-Fig. 41.7). By contrast, cortical LBs appear as sub-
tle spheres of homogeneous eosinophilia. Making detection even more
difficult, cortical LBs are not argyrophilic. Fortunately, LBs all show
immunoreactivity for ubiquitin and a-synuclein (e-Fig. 41.8). These
lesions (along with similar inclusions [Lewy neurites] that appear within
cell processes) are seen in Lewy body disorders (e.g., Parkinson disease
[PD], and dementia with Lewy bodies [DLBs]).
c. Marinesco bodies are small, eosinophilic, strongly ubiquitin-positive
intranuclear inclusions located chiefly in pigmented brain stem neurons
(e-Fig. 41.9). These have no known pathologic significance.
d. Neurofibrillary tangles (NFTs} (e-Fig. 41.10) are argyrophilic intracyto-
plasmic filamentous aggregates of hyperphosphorylated tau protein.
Although characteristic of AD, they also appear in many other neurode-
generative disorders, and in rare GGs.
e. Hirano bodies are brightly eosinophilic rod-shaped or elliptical cytoplas-
mic inclusions that occur within the proximal dendrites of neurons, par-
ticularly in the hippocampus. Although not specific, they are particularly
numerous in brains with AD pathology (e-Fig. 41.11).
f. Granulovacuolar degeneration (GVD} is common in hippocampal pyrami-
dal neurons in AD, and less common in older brains without AD. GVD
resembles many small bubbles, each with a small basophilic granule.
g. TDP-43 neuronal cytoplasmic inclusions (NCis}, as the name suggests, are
immunoreactive for TDP-43. Although NCis are characteristic of a sub-
set of frontotemporal dementias, they are not specific, and may be seen
in the temporal lobe in other neurodegenerative diseases (e.g., AD).
h. Bunina bodies are eosinophilic ubiquitin and TDP-43 immunoreactive
intracytoplasmic inclusions that form in motor neurons in cases of famil-
ial and sporadic amyotrophic lateral sclerosis (ALS).
B. Astrocytes
1. Reactive astrocytosis. Normally, astrocytes are evenly dispersed (albeit with
regional variation), mitotically silent, and GFAP positive. In most forms
of brain injury, astrocytes become hypertrophic, increase their GFAP con-
tent, and may proliferate. Reactive astrocytes have prominent stellate pro-
cesses and, often, abundant eccentrically distributed glassy cytoplasm that
inspires the moniker "gemistocyte." Nevertheless, reactive astrocytes gen-
erally maintain an even distribution, and do not exhibit nuclear atypia
(radiation exposure is an exception). Grossly, tissues affected by chronic
astrocytosis are usually firm; thus, the term "gliotic" is used to describe
brain tissues that appear unusually firm or rubbery.
2. Bergmann gliosis refers to an accumulation of astrocytic nuclei (usually in
association with neuron loss) within the Purkinje cell layer of the cerebel-
lum.
3. Alzheimer type II astrocytes, with swollen pale nuclei and minimal visible
cytoplasm, appear in hyperammonemic states (e.g., liver failure, Wilson
disease). In the cortex and striatum, these nuclei are round with a prominent
nucleolus (e-Fig. 41.12); in the pallidum, dentate nucleus, and brainstem,
they are irregular and lobated.
4. Corpora amylacea are basophilic, round, concentrically lamellated aggre-
gates of polyglucosan (polyglucosan bodies) that develop within astrocytic
processes. Common in normal brains, particularly near ventricular and pial
surfaces, these become more numerous with age. Similar structures (Lafora
bodies) form in far greater numbers in astrocytes and neurons (and in eccrine
sweat glands) in Lafora body disease.
5. Rosenthal fibers (Rfs} are brightly eosinophilic, somewhat refractile, irregu-
lar/beaded structures that range from "'1 0 to 40 ~-tm in diameter. EM reveals
them as swollen astrocytic processes filled with electron-dense amorphous
granular material and glial filaments. RFs are commonly observed in PA, but
are also commonly seen in nonneoplastic tissues adjacent to slowly growing

neoplasms (e.g., craniopharyngioma, ependymoma), cysts, and syrinx; such
RF-abundant tissue reaction is called piloid gliosis (e-Fig. 41.13).
6. Eosinophilic granular bodies (EGBs}, although not present in nonneoplastic
astrocytes, are found in slowly growing astrocytic and glioneuronal tumors
(PA, GG, and pleomorphic xanthoastrocytoma [PXA]). EGBs appear on
H&E sections (or cytologic smear preparations) as refractile clusters of
small, round hyaline droplets, and are PAS-positive (e-Fig. 41.14).
C. Microglia. Normal residents of the brain, microglial cells are of monocytic
lineage (and are consequently immunoreactive for leukocyte common anti-
gen [LCA/CD45] as well as CD68 and CD163). In response to various sig-
nals, these cells undergo activation, whereupon they change their morphology
(appearing as irregular elongated "rod cells"), become motile, and intensify
their communication with other cells via secreted factors (e.g., cytokines and
interleukins). They may also participate in limited phagocytosis. Small clus-
ters of activated microglial cells (microglial nodules) are characteristic of viral
encephalitis and may be seen decorating dying neurons, in a process called neu-
ronophagia. Capacity for phagocytosis increases in the setting of injury, infec-
tion, or demyelinating disease when microglia differentiates into macrophages.
Monocytes and macrophages may also be recruited into the CNS from the sys-
temic circulation. Occasionally, brisk mitotic activity among macrophages in
these settings (particularly in tumefactive multiple sclerosis (MS), which radio-
graphically resembles GBM) can cause diagnostic confusion with gliomas.
Gliomas themselves often contain a large number of microglial cells, which
appear to play a role in tumorigenesis (Cancer Res. 2008;68:10358).
D. Cerebral edema is an increase in brain volume due to increased water content.
Depending on its pathogenesis, cerebral edema can be classified as vasogenic,
cytotoxic, osmotic, or interstitial (resulting from obstructive hydrocephalus).
However, combinations of different edema types often coexist. In vasogenic
edema, the fluid collection is predominantly extracellular and results from
breakdown of the blood-brain barrier. In cytotoxic edema, the fluid accumu-
lation is intracellular, the result of impaired Na/K-ATPase function in glial
cells caused by toxins, ischemia, or various other conditions. Osmotic edema
results when osmolality of the brain interstitial fluid exceeds that of the plasma.
Interstitial edema results from transependymal flow from the ventricles in the
setting of obstructive hydrocephalus.
E. Hydrocephalus, an abnormal increase in the intracranial volume of CSF asso-
ciated with dilatation of all or part of the ventricular system, may be classi-
fied as communicating, noncommunicating (obstructive), normal pressure, or
ex vacuo.
1. Noncommunicating hydrocephalus results from physical blockage of CSF
flow, usually by tumor compressing a narrow channel, such as the foramen
of Monro or the cerebral aqueduct.
2. Communicating hydrocephalus results from impaired resorption of CSF at
the arachnoid granulations, as may occur in the setting of subarachnoid
hemorrhage or meningitis; less commonly, it may result from increased CSF
production, for example, by a choroid plexus papilloma.
3. Normal pressure hydrocephalus (NPH}, characterized classically by the clini-
cal triad of dementia, ataxia, and incontinence, is poorly understood. NPH
may represent a circumstance in which production and resorption of CSF
reach a new equilibrium after a prolonged period of impaired resorption.
4. Hydrocephalus ex vacuo describes a state of ventricle expansion due to loss
of adjacent parenchyma or generalized cerebral atrophy (as occurs in AD).
F. Intracranial pressure (ICP) and brain herniation. Normally, the cranial cav-
ity contents (blood, brain, and CSF) are maintained in volumetric balance
within the rigid skull and dura. ICP increases when this balance is strained,
as may result from diffuse brain edema, increased cerebral blood flow and
blood volume, or development of space-occupying lesions (e.g., tumor, abscess,
hematoma, or large edematous infarct). Elevated ICP, if not treated, can
cause herniation; herniation syndromes include subfalcine (cingulate gyrus),
transtentorial (uncal), and cerebellar tonsillar/brainstem herniations.
G. Duret (secondary brain stem) hemorrhage occurs when penetrating pontine arter-
ies (which arise perpendicularly from the basilar artery) become kinked in asso-
ciation with brainstem herniation; the resulting acute hemorrhagic infarction
of the pons is often fatal.
V. NEOPLASMS OF THE CNS. For most CNS tumors, incidence varies greatly with age
and gender. Among adults, metastases, GBM, and meningioma are the most com-
mon CNS neoplasms; among children, PA, medulloblastoma, and ependymoma
are far more common. Likewise, tumors often differ in their radiographic features
and propensity for certain anatomic sites. For this reason, microscopic evaluation
of a CNS biopsy specimen is incomplete without considering the neuroradiologic
findings that describe the targeted lesion in situ. Indeed, neuroradiologic assess-
ment can be considered to be the neuropathology surrogate for gross examination.
Imaging studies are particularly helpful when evaluating small biopsy samples.
The World Health Organization (WHO) currently lists more than 100 types of
CNS tumors and their variants (Table 41.2). Table 41.3 organizes common CNS
tumor diagnoses on the basis of location, patient age, and imaging characteristics.
Histologic features are also critical for diagnosis. Because a final diagnosis may
not be obvious from an initial histomorphologic examination, it is worthwhile
to begin with a broad differential based on a specimen•s general histopathologic
pattern. Table 41.4lists eight major histopathologic patterns that may be encoun-
tered, along with their most commonly associated diagnostic entities. Once a
differential diagnosis has been formulated on the basis of these data, closer exam-
ination of microscopic details can refine the differential further and suggest what
ancillary tests (if any) are required to arrive at a final diagnosis.
A. Gliomas
1. Diffuse (infiltrating) astrocytomas (DAs), WHO grade II. Diffuse gliomas are
the most frequent primary CNS neoplasms. Because they are diffusely infil-
trative, complete resection is nearly impossible. On MRI, DAs are nonen-
hancing, T1 hypointense, T2/FLAIR hyperintense, ill-defined intra-axial
masses that can occur throughout the neuraxis, but commonly involve
the cerebral hemispheres. Clinically, they present with new-onset seizures
(the most common symptom), headaches, or functional neurologic deficits.
Grossly, these lesions may appear gray-tan to gelatinous and obscure the
native gray-white junction. Microscopically, tumor cells invade adjacent
cortex along white matter tracts. They aggregate around neurons and
blood vessels and beneath pial and ependymal surfaces to produce the
so-called secondary structures of Scherer. The cytologic features of astro-
cytic tumor cells can vary widely from uniform and minimally atypical,
to highly pleomorphic both in cytoplasmic and nuclear features (e-Fig.
41.15). Cells with elongate, irregular, hyperchromatic nuclei with minimal
cytoplasm are seen in fibrillary astrocytomas, whereas cells with eccentri-
cally located nuclei and abundant eosinophilic cytoplasm characterize the
gemistocytic variant. Tumor cells are often, but not invariably, immunore-
active to GFAP. A recently identified immunohistochemical marker,
antibody IDH-1, is proving invaluable for distinguishing the majority of
DAs (but not primary GBMs) from gliosis. Mitotic activity is very low or
nonexistent.
2. Anaplastic astrocytoma (AA}, WHO grade Ill, is a diffusely infiltrating glioma
with a mean age of presentation in the fifth decade. AA may appear
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hypercellular sheets of scGBM appear bland. At higher power, scGBM

shows slightly oval or elongate nuclei with mild hyperchromasia
and delicate chromatin, perinuclear halos, microcalcifications, and
"chicken-wire"-like branching capillaries. However, unlike most oligo-
dendrogliomas, its mitotic rate is remarkably high (e-Fig. 41.17). This
dissonance (bland histology and abundant mitotic figures) is often the
first clue to the diagnosis (Adv Anat Pathol. 2005;12:180). Other help-
ful clues include radiographic ring-enhancement and, when present,
focal EH and pseudopalisading necrosis. However, a subset of anaplas-
tic oligodendrogliomas can show these features, and one-third of
scGBMs do not.
Immunohistochemical and molecular characteristics can be used
to distinguish scGBM and oligodendrogliomas. The tumor cells in
scGBM often contain thin, GFAP-positive cytoplasmic processes; GFAP
reactivity in oligodendrogliomas is usually restricted to minigemis-
tocytes, gliofibrillary oligodendrocytes, and entrapped reactive astro-
cytes. The MIB-1 (Ki-67) labeling index is typically much higher in
scGBM than in most anaplastic oligodendrogliomas, although the
ranges of these indices overlap. More importantly, reactivity with anti-
body IDH-1 is uniformly absent in scGBM (a primary GBM) and
present in the majority of oligodendrogliomas. Additionally, FISH
studies have shown EGFR amplifications in ...... 70% and deletions of
chromosome 10q in >90% of the scGBMs (e-Fig. 41.18); these are
rare in oligodendroglioma. In contrast, codeletion of chromosomal
arms 1p and 19q, common in oligodendrogliomas, is not observed in
scGBM.
b. Gliosarcoma {GS) WHO grade IV, a variant of GBM, is often super-
ficially located and, radiographically, deceptively circumscribed. GS
exhibits astrocytic and sarcomatous elements, the latter usually resem-
bling fibrosarcoma or malignant fibrous histiocytoma. However, GS
may also exhibit bone, cartilage, muscle, and even epithelial lines of
differentiation; when the latter is present, the tumor may be called
adenoid GBM. Molecular studies have demonstrated similar genetic
abnormalities within all histologic elements of the tumor, suggesting a
clonal origin. The sarcomatous elements in GS are reticulin rich, typi-
cally negative for GFAP, and positive for vimentin, smooth muscle actin,
muscle-specific action (MSA), etc., based on the sarcomatous differenti-
ation pattern. In contrast, the glial regions are reticulin poor and GFAP
positive.
4. Gliomatosis cerebri is an extensively infiltrative glioma, often of astrocytic
type, that involves three or more lobes of the cerebrum and often extends
into brain stem, cerebellum, and/or spinal cord as well. Microscopically,
gliomatosis most often has features of AA but is associated with a poor
prognosis regardless of the grade. Foci of progression to GBM are not
uncommon.
5. Circumscribed astrocytomas
a. Pilocytic astrocytoma {PA), WHO grade I (e-Fig. 41.19), is a slowly grow-
ing, rather circumscribed neoplasm frequently occurring in children
and young adults, with a predilection for cerebellum. It also occurs in
the optic pathway, hypothalamus, dorsal brain stem, spinal cord, and
rarely, cerebral hemispheres. Classically, these tumors appear in imag-
ing studies as cystic lesions with an enhancing mural nodule. In the
brain stem, they may occur as exophytic lesions. Histologically, these
tumors have a biphasic pattern in which compact pilocytic areas are in-
terspersed with microcystic loose and/or spongy areas. PAis variably
cellular and populated by bipolar cells with long hair-like (piloid) pro-
cesses. RFs and EGBs commonly occur within the compact (dense)
regions and, although not specific for PA, provide very helpful diag-
nostic clues. Mitotic activity is low. The blood vessels within PAs are
commonly hyalinized, and often exhibit linear glomeruloid tufts of EH.
This feature, particularly when coupled with the marked nuclear atypia
that commonly occurs in PA, can lead to confusion with high-grade
glioma. However, the EH and nuclear atypia of PA have no prognostic
significance. PAs usually follow a benign clinical course.
b. Pilomyxoid astrocytoma (PMA}, WHO grade II, is a piloid neoplasm closely
related to PA that presents in infants at a median age of 10 months, and
favors the hypothalamus and optic chiasm. On MRI, PMA appears cir-
cumscribed and solid, hypointense on T1 and hyperintense on 1'2, and
shows homogenous contrast enhancement. The histologic hallmark of
PMA is the presence of monomorphous bipolar cells in a markedly
mucoid matrix, with a prominent angiocentric arrangement that resem-
bles the perivascular pseudorosettes of ependymoma (e-Fig. 41.20). As
defined, PMA does not show RFs or EGBs. Nuclear atypia is uncom-
mon. Mitoses can be present. Vascular proliferation resembling that in
PAis characteristic. Necrosis may be seen. The tumor cells show strong
and diffuse immunoreactivity for GFAP, S-100, and vimentin. A subset
of tumor cells is also positive for SYN. Clinically, PMAs behave more
aggressively than PAs, with frequent local recurrences and, often, CSF
seeding at the time of diagnosis.
6. Pleomorphic xanthoastrocytoma (PXA}, WHO grade II, is an epileptogenic
neoplasm commonly located in the superficial cortical regions of the tem-
poral lobe, often with meningeal attachment. Histologically, this tumor is
composed of large pleomorphic, variably GFAP-positive astrocytes inter-
mingled with less conspicuous spindled cells (e-Fig. 41.21). Some of the
tumor cells have bizarre nuclei and nuclear pseudoinclusions, and multi-
nucleated giant cells are common. EGBs are almost always evident. Xan-
thomatous tumor cells, although diagnostically helpful, only occur in 25%
of the cases. Other features include perivascular plasmalymphocytic cuff-
ing and scant mitoses. Reticulin silver stain highlights a network of basal
laminae that surrounds individual tumor cells. Neuronal differentiation
is often evidenced by reactivity for SYN and NF protein; reactivity for
CD34 is frequently observed. Grade II PXAs have a relatively favorable
prognosis (81% 5-year survival). Anaplastic (WHO grade III) transforma-
tion, characterized by less pleomorphism, increased proliferative activity,
necrosis, and/or microvascular proliferation, occurs in 15% of the PXAs;
reticulin staining is often diminished in these cases.
7. Subependymal giant cell astrocytoma (SEGA}, WHO grade I, is almost exclu-
sively found in children and young adults with tuberous sclerosis (TS).
These well-circumscribed, intraventricular tumors usually occur near the
foramen of Monro and cause symptoms associated with obstructive hydro-
cephalus. Imaging studies reveal contrast enhancement and calcification.
Histologically, the tumor cells may be spindled, epithelioid, or gemistocyte-
like, and may appear in sweeping fascicles, large clusters, or perivas-
cular pseudorosettes. The gemistocyte-like cells have abundant glassy
eosinophilic cytoplasm and nuclei resembling those of ganglion cells,
with fine granular chromatin and prominent nucleoli (e-Fig. 41.22). The
hybrid astrocytic and neuronal features of SEGAs are also reflected in their
immunohistochemical profile, with focal reactivity for both GFAP and one
or more neuronal markers (e.g., SYN). Mitoses are rare. Microvascular
proliferation and necrosis are typically absent.
8. Oligodendrogliomas, WHO grade II, comprise 10% to 25% of all the adult
gliomas, behave less aggressively than astrocytomas, and show slower pro-
gression and longer patient survival. The most useful histologic feature
for distinguishing oligodendroglia! neoplasms from astrocytic tumors is
nuclear morphology; oligodendroglioma nuclei are nearly spherical, with
delicate chromatin, crisp envelopes, and inconspicuous nucleoli (e-Fig.
41.23). Other helpful features include dear perinuclear haloes that often
result as an artifact of routine histologic processing; a delicate hexag-
onal array of capillaries that resembles chicken wire; and mucin-rich
microcystic spaces, microcalcifications, predominantly cortical involve-
ment, and perineuronal satellitosis. Immunohistochemically, classic oligo-
dendroglioma cells show no reactivity for GFAP. However, oligoden-
drogliomas often contain three cell types that do show such reactivity,
specifically mini-gemistocytes, gliofibrillary oligodendrocytes, and reactive
astrocytes. The mini-gemistocyte, named for its superficial resemblance
to the larger astrocytic gemistocyte, has a belly of glassy, eosinophilic
cytoplasm and an eccentrically placed oligodendroglia! nucleus. Gliofibril-
lary oligodendrocytes are indistinguishable from classic oligodendroglia!
tumor cells on H&E stained sections, but show a GFAP-positive rim of
cytoplasm and a tadpole-like tail. Entrapped, nonneoplastic astrocytes
generally retain their characteristic stellate morphology.
In addition to oligodendrogliomas, several other tumors exhibit the
so-called fried egg appearance with rounded nuclei and dear perinuclear
haloes; most notably dysembryoplastic neuroepithelial tumor (DNT),
PA, central neurocytoma, dear cell ependymoma, and mixed oligoas-
trocytoma (MOA). Nevertheless, this differential diagnosis can usu-
ally be resolved using clinical information, other histologic clues (e.g.,
RFs, EGBs, atypical astrocytic nuclei), immunohistochemical stains (e.g.,
GFAP, Neu-N, SYN, NF, IDH-1, EMA, CD99, D2-40, MIB-1), and FISH
studies.
Ancillary genetic testing of oligodendrogliomas (Adv Anat Pathol.
2005;12:180) is now routinely performed. Several studies utilizing FISH
techniques have established the presence of 1p and 19q codeletions in
60% to 90% of the oligodendrogliomas (e-Fig. 41.24); these tumors
behave less aggressively, are more sensitive to chemotherapy and radia-
tion, and are thus considered genetically favorable. Apart from the obvi-
ous prognostic implications, identification of 1 p and 19q codeletions also
helps to differentiate oligodendroglioma-like mimics from true oligoden-
drogliomas. Unfortunately, pediatric oligodendrogliomas generally do not
show codeletion; those that do harbor the codeletion frequently occur in
teenagers and older children and likely represent the adult type oligoden-
droglioma(] Neuropathol Exp Neurol. 2003;62:53).
9. Anaplastic oligodendroglioma, WHO grade Ill. On the basis of WHO criteria,
anaplastic oligodendroglioma must have "significant mitotic activity, EH,
or necrosis" along with increased cellularity and marked atypia (WHO
Classification of Tumors of the Central Nervous System. Lyon, France:
IARC Press; 2007). High-grade oligodendrogliomas additionally often
show greater cytologic pleomorphism, epithelioid morphology, prominent
nucleoli, and sharper cell borders. Necrosis is more often infarct-like rather
than pseudopalisading. The exact number of mitoses required for diag-
nosis has not been established, but a mitotic index of six or more per
ten high-power fields (40x) is often used to assign anaplastic grade Ill
(based on J Neuropathol Exp Neurol. 2001 ;60:248) in the absence of EH
or necrosis. Anaplastic transformation can be focal or widespread. Some
oligodendrogliomas appear otherwise low grade, but contain hypercellu-
lar nodules with increased mitotic activity; such cases may be designated
oligodendroglioma with focal anaplasia, WHO grade III. Neuroimag-

ing studies of anaplastic oligodendroglioma often, but not invariably,
show patchy or homogeneous contrast enhancement. Ring enhancement
is uncommon and predicts poor prognosis. scGBM is usually considered
in the differential diagnosis.
10. Mixed oligoastrocytoma {MOA}. Although the majority of gliomas, when
adequately sampled and well fixed, show classical features of either oligo-
dendroglioma or astrocytoma, many cases show ambiguous or intermixed
features and are therefore included in the category of MOA. MOAs exhibit
low diagnostic concordance rates even among expert neuropathologists
(J Neuropathol Exp Neurol. 2003;62:1118), largely due to the absence of
specific markers for either astrocytic or oligodendroglia! differentiation.
Histologically, the oligodendroglia! and astrocytic components manifest
either as geographically separate or, more often, as intermingled forms.
Unlike the classical oligodendroglioma, only a small subset of MOAs
exhibits 1p and 19q codeletions. When MOAs exhibit an increased mitotic
index, EH, or palisading necrosis, they are designated as anaplastic (WHO
grade III); in the absence of EH or necrosis, the threshold for mitotic
index is not well established, but the threshold of six or more mitotic fig-
ures per ten high-power fields (40x) is often applied (] Neuropathol Exp
Neurol. 2001;60:248). MOAs with necrosis are sometimes referred to as
GBM with oligodendroglia! component, WHO grade IV U Clin Oncol.
2006;24:5419); they have a prognosis better than that of classic GBMs,
but worse than that of anaplastic MOAs lacking necrosis.
B. Ependymal neoplasms
1. Ependymoma, WHO grade II (e-Fig. 41.25). Ependymomas occur as discrete
enhancing masses. In children, they favor a fourth ventricular location; in
young adults, the spinal cord. However, these associations are not abso-
lute, and supratentorial tumors, often unassociated with the ventricles,
can present in either age group. CSF dissemination occurs in <5% of the
cases. Calcification in ependymoma is common when the lesion is located
intracranially, whereas cyst formation is common in supratentorial cases.
Smear preparations of ependymoma show uniform round to oval nuclei,
distinct nucleoli, and spindled to epithelioid morphology; histologically,
the tumor cells form perivascular pseudorosettes and, in 5% to 10% ofthe
cases, true ependymal rosettes. True rosettes (or ependymal canals) have
central lumens reminiscent of the young central canal of the spinal cord,
whereas perivascular pseudorosettes have vessels at the center, surrounded
by a nucleus-free zone of radially oriented tumor cell processes. Ependy-
mal canals, elongate versions of true rosettes, are typically seen in only the
most differentiated examples. lmmunohistochemically, most ependymomas
exhibit reactivity for GFAP. Many cases also show reactivity for EMA,
CD99, and/or podoplanin (antibody D2-40) along the luminal surface of
true rosettes/ependymal canals or within paranuclear intracytoplasmic dot-
like structures, thought to represent intracytoplasmic lumina. In difficult
cases, EM studies can confirm ependymal differentiation by demonstrat-
ing long zipper-like intercellular junctions, microvilli, cilia, and/or intracy-
toplasmic lumina. Variants of ependymoma include the clear cell (mimics
oligodendroglioma), cellular (mimics medulloblastoma or PNET), tanycytic
(mimics schwannoma or PA), and papillary (mimics choroid plexus tumors)
subtypes. Rarely, ependymomas may contain cells with melanin pigment, or
xanthomatous, signet ring, giant cell, or neuronal features. Poor prognostic
indicators include age < 3 years, posterior fossa location, and anaplastic
tumor grade. Grade ll ependymomas have a 5-year progression-free sur-
vival rate of 60% to 80% as compared with 25% to 50% in anaplastic
ependymomas.
2. Anaplastic ependymoma, WHO grade Ill. Several grading systems have been
proposed for ependymomas, but no consensus has been achieved. According
to the WHO 2007 criteria, anaplastic ependymomas are characterized by
increased cellularity, brisk mitotic activity, pseudopalisading necrosis, and
microvascular proliferation. Necrosis by itself in the absence of pseudopal-
isading does not warrant the diagnosis of anaplasia, because lower-grade
ependymomas also can show degenerative changes.
3. Myxopapillary ependymoma, WHO grade I, is a slowly growing tumor of young
adults that occurs almost exclusively in the cauda equina region, where it
arises from the filum terminale. Grossly, these tumors are encapsulated,
intradural, sausage-shaped masses with a gelatinous interior. Histologi-
cally, the tumors show variable proportions of ependymal and papillary
patterns. In papillary areas, the tumor cells form irregularly ovoid rings
of cuboidal-to-columnar epithelium, each ring formed around a hyalinized
central vessel, some fibrous stroma, and a rim of pale basophilic mucin.
Pale basophilic often bubbly mucinous (myxoid) material also separates the
papillae. In ependymal areas, the tumor cells are spindled and slendet; and
are arranged in loose fascicles and rather extravagant perivascular pseu-
dorosettes (e-Fig. 41.26). The tumor cells are immunoreactive for GFAP,
S-100 protein, and vimentin, with variable staining for EMA, CD99, and
podoplanin (antibody D2-40). These tumors have a favorable prognosis fol-
lowing complete resection. If untreated or incompletely resected, rare cases
can progress and invade perispinal tissues. Occasional ependymal tumors
in this location show a mixture of myxopapillary and typical ependymoma
features; the clinical behavior of such tumors is incompletely understood.
4. Subependymoma, WHO grade I, is a benign, slowly growing, solid tumor
related to ependymoma. They are often asymptomatic, discovered inciden-
tally on CT and MRI studies performed for other reasons. Occasionally,
they undergo intratumoral hemorrhage and become life threatening by
exerting mass effect. Prognosis is excellent following resection. Grossly,
they are sessile or pedunculated masses within the ventricles (lateral >
fourth > third). Histologically, they are lobulated and well demarcated.
The tumor nuclei resemble those of ependymoma and appear in irregu-
lar clusters within a dense fibrillar background formed by tumor cell pro-
cesses (e-Fig. 41.27). Mitotic figures are rare. Occasional pseudorosettes
are not unusual. Secondary degenerative changes include microcyst forma-
tion, hemosiderin deposition, vascular hyalinization, myxoid change, and
calcification. Nuclear pleomorphism and microvascular proliferation are
occasionally seen; these do not warrant a higher-grade diagnosis. It is worth
noting that subependymoma may appear as part of a compound ependymal
tumor; in such cases, tumor grading should be assigned according to the
ependymal component.
C. Choroid plexus tumors
1. Choroid plexus papilloma, WHO grade I (e-Fig. 41.28), is a benign lesion com-
monly located in the lateral ventricles in children, and in the fourth ventricle
in adults. It typically presents with symptoms associated with obstructive
hydrocephalus. Histologically, choroid plexus papilloma resembles normal
choroid plexus. Howevet; its fibrovascular cores are lined by a cuboidal-
to-columnar epithelium that lacks the superficial intercellular spaces that
impart a cobblestone appearance to normal choroid plexus. Mitotic activity
is low. Clear cytoplasmic vacuoles are noted in some cases. Occasionally,
infarct-like necrosis may be present, but has no prognostic significance.
Focal ependymal differentiation is common. Tumor cells show immunore-
activity for S-100, CAM 5.2, transthyretin, and GFAP (focally) but not for
EMA and carcinoembryonic antigen (CEA).
2. Choroid plexus carcinoma, WHO grade Ill (e-Fig. 41.29), usually presents in
children <3 years of age as an enhancing, intraventricular mass. These
tumors are highly aggressive, often metastasize via the CSF, and are nearly
uniformly fatal. Histologically, the tumors are more solid and complex
than papillomas. High-grade cytology and frequent mitoses are the rule.
Foci of necrosis are characteristic, and microvascular proliferation may be
seen. Focal small cell features may mimic embryonal tumors (e.g., PNET).
Immunoreactivity for S-100, CAM 5.2, and transthyretin is characteristic; is
variable for GFAP; and is not typically observed for EMA and CEA. The dif-
ferential diagnosis includes anaplastic ependymoma (GFAP+, CAM 5.2-,
EMA+/-), GBM (GFAP+, CAM 5.2-), medulloblastoma/PNET (SYN+,
CAM 5.2- ), and metastatic carcinoma (in older patients, EMA+, CEA+/-,
S-100 +1-, CAM 5.2+).
D. Other glial neoplasms.
1. Angiocentric glioma {AG}, WHO grade I (e-Fig. 41.30). First reported in 2005,
AG is a rare supratentorial tumor associated with epilepsy in children and
young adults. Radiographically, AG appears as a T2/FLAIR hyperintense,
nonenhancing mass that expands the cortex with minimal mass effect. In
some cases, a projection of the tumor extends to the ventricle. Histologically,
AG shows uniform, slender spindled glial cells, often arranged either radi-
ally from or parallel to vessels, and, occasionally, perpendicular to the pia.
AG shows some ependymal features, including dot-like immunoreactivity
for EMA, and microvilli and zipper-like junctions by EM. The proliferation
index is low.
2. Astroblastoma {AB}, not yet graded {e-Fig. 41.31 ). As with AG, AB is asso-
ciated with epilepsy in children and young adults, but AB is also associ-
ated with headaches and vomiting, and shows a female predominance. On
MRI, AB is usually large, lobulated, well demarcated, solid with an occa-
sional cystic component, T2-isointense with gray matte.t; and enhancing.
Histologically, AB bears some resemblance to ependymoma, but has stout
(rather than fibrillar) cell processes that form the defining astroblastic pseu-
dorosettes. Vascular hyalinization is usually robust; in some tumors, this
architectural pattern assumes a papillary appearance. Like ependymoma
and AG, AB shows dot-like immunoreactivity for EMA; reactivity for GFAP
highlights the abbreviated cell processes within pseudorosettes. ABs show-
ing >5 mitoses per ten 40x fields, anaplastic nuclear features, increased cel-
lularity, microvascular proliferation, and pseudopalisading necrosis are con-
sidered anaplastidmalignant, but may still be resectable due to this tumor's
noninfiltrating growth pattern.
3. Chordoid glioma of the third ventricle {CG}, WHO grade II, is a rare tumor of
adults that occurs almost exclusively in the vicinity of the anterior third ven-
tricle and hypothalamus. Radiographically, CG is solid, well demarcated,
T2 intense, and strongly contrast enhancing. Histologically, as its name
suggests, CG focally resembles chordoma, showing cords and dusters of
eosinophilic epithelioid cells within a bubbly basophilic mucinous matrix;
where mucin is lacking, the cells (strongly GFAP positive, modestly reac-
tive for EMA) appear in sheets. Lymphoplasmacytic infiltrates are typically
seen. Immunoreactivity for GFAP (and absence of whorls, psammoma bod-
ies, and physaliferous cells) distinguishes CG from chordoid meningioma
and chordoma.
E. Neuronal and glioneuronal neoplasms
1. Ganglion cell tumor {GG and gangliocytoma}, WHO grade I. Ganglion cell
tumors are epileptogenic and appear most often in the temporal lobes. On
imaging, they commonly enhance and appear solid, cystic, or both. A cyst
with an enhancing mural nodule is a common (but not specific) pattern.
Microscopically, they are usually cortically based, microcystic, variably
fibrotic, and calcified, and they often show perivascular lymphocytic cuffing
(e-Fig. 41.32). Dysmorphic neurons (that may exhibit cytomegaly, vac-
uolated cytoplasm, coarse Nissl substance, irregular multipolar processes,
and/or nuclear abnormalities including binucleation) are the defining
feature. Architecturally, the ganglion cells are clumped or haphazardly
arranged in comparison with the laminar well-ordered arrangement of the
normal cortex. GGs have a variable glial component, typically astrocytic;
glial predominant GG may resemble DA or PA. Other features commonly
noted include EGBs and RFs (although the latter is more common at the
tumor periphery, and may result from piloid gliosis). Cortical dysplasia may
be seen adjacent to GG. High-grade glial transformation is exceedingly rare
and difficult to define. The glial component is immunoreactive for GFAP;
the neuronal component is variably immunoreactive for SYN, NF, and chro-
mogranin, but is usually immunonegative or minimally positive for Neu-N.
A subset of cells both within and adjacent to the tumor often shows
immunoreactivity for CD34; these CD34+ cells are "spider-like,, charac-
terized by long, stellate, ramified processes, and are thought to represent
progenitor cells. GGs have a favorable prognosis after surgical resection.
2. Desmoplastic infantile astrocytoma and desmoplastic infantile ganglioglioma
(DIAIDIG), WHO grade I (e-Fig. 41.33), typically occur in children younger
than 2 years, and are located superficially within the frontoparietal region.
Radiologically, they are very large, attached to the dura, brightly enhanc-
ing, and are associated with a very large, occasionally multiloculated cyst.
Histologically, the tumors are biphasic. In the collagen N- and reticulin-rich
phase, the tumor cells are arranged in a storiform or fascicular pattern. The
astrocytic cells, spindled and gemistocytic, are often inconspicuous within
the desmoplastic background, but can be identified by GFAP immunos-
taining. The neuronal component (present in DIG, absent inDIA) is also
subtle because the polygonal neuronal tumor cells are often considerably
smaller than those of conventional GG. The other component of the tumor
resembles PNET, with abundant small cells and many mitotic figures, and
lacks reticulin and collagen. EH and necrosis may be observed but are not
associated with an unfavorable prognosis.
3. Dysplastic cerebellar gangliocytoma (DCG} (Lhermitte-Duclos disease}, WHO
grade I. This unique cerebellar neoplasm is often associated with Cowden
syndrome (autosomal dominant PTEN/MMAC-1 mutation). Patients with
DCG often present with cerebellar dysfunction and/or obstructive hydro-
cephalus. DCG, which is Tl hypointense and T2 hyperintense, has a char-
acteristic striped appearance with both modalities. Microscopically, DCG
presents as a unilateral expansion of cerebellar folia wherein the internal
granular layer is replaced by dysmorphic ganglion cells; there is no glial
component. Abnormal vascular proliferation is sometimes noted, and white
matter is occasionally vacuolated. Evaluation for other features of Cowden
syndrome (breast and gastrointestinal lesions) is warranted in patients with
DCG.
4. Central neurocytoma, WHO grade II (e-Fig. 41.34), occurs in the lateral ven-
tricles near the foramen of Monro in young or middle-aged patients with
obstructive hydrocephalus. On imaging studies, these masses are large, glob-
ular, enhancing, and often calcified. Histologically, the tumor cells have a
uniform appearance with round nuclei, finely granular chromatin, incon-
spicuous nucleoli, sparse eosinophilic or clear cytoplasm, and delicate fib-
rillar cell processes (in contrast to the coarse processes of glial tumors).
The cells often appear in solid monotonous sheets that may also exhibit
prominent chicken-wire vasculature, and/or neurocytic (Homer-Wright)
rosettes, and/or perivascular pseudorosettes, and/or calcifications. Thus, the
differential diagnosis often includes oligodendroglioma and ependymoma.

In most cases, the brain/tumor interface of central neurocytoma is
solid/pushing rather than infiltrative. Immunohistochemistry shows diffuse
and strong immunoreactivity with SYN, variable reactivity for Neu-N and
NF, and focal reactivity for GFAP. Occasionally, neurocytomas occur within
the parenchyma of the cerebral hemispheres, cerebellum, or spinal cord;
these extraventricular neurocytomas often exhibit more pronounced gan-
glion cell and astrocytic differentiation. A small subset of neurocytomas
(central and extraventricular) with microvascular proliferation, necrosis,
and elevated mitotic/proliferative index (MIB-1 labeling index of 2% or
more) show more rapid recurrence, and are identified as atypical, although
they are still considered WH 0 grade IT.
5. Dysembryoplastic neuroepithelial tumor (DNT}, WHO grade I, is a benign,
slowly growing tumor of adolescents and young adults who have a history of
longstanding, drug-resistant partial seizures. On imaging, DNT appears in
an area of expanded cerebral cortex, with a predilection for the medial tem-
poral lobe. DNT is Tt hypointense, T2JFLAIR hyperintense, and nodular;
some complex varieties show enhancement (see below). Microscopically,
simple DNT demonstrates only the specific glioneuronal element; patterned
intracortical mucin-rich nodules each exhibit a network of bundled axons
and delicate capillaries that divide the mucin into microcystic pools. This
network is decorated by oligodendroglioma-like cells and occasional stellate
astrocytes, and some of the microcytic pools show morphologically normal
"floating" neurons (e-Fig. 41.35). Mitotic figures are rare to absent. Com-
plex DNT consists of the specific glioneuronal element with an additional
component referred to as glial nodules which are histologically identical to
PA, GG, diffuse glioma, or another glial neoplasm. Cortical dysplasia may
be seen adjacent to DNTs. Simple DNTs are commonly misdiagnosed as
oligodendrogliomas and, if sampling is incomplete, complex DNTs may be
misdiagnosed according to the glial nodule subtype that is present. There-
fore, review of clinical history and radiographic imaging is essential in eval-
uating specimens from temporal lobe epilepsy surgical resections. DNT has
a benign course and postsurgical seizure resolution is common.
6. Papillary glioneuronal tumor (PGNT}, WHO grade I, is a contrast-enhancing,
well-circumscribed, solid or cystic lesion that arises most commonly in
the periventricular white mattet; usually in the temporal lobe (Am I Surg
Pathol. 199 8;22: 1171 ). Patients, most commonly young adults, present
with seizures, headaches, and vision disturbance. Histologically, as the
name suggests, these tumors have glial and neuronal elements, and a pap-
illary appearance. The glial element (GFAP-positive) forms a loose layer
of spindled-to-cuboidal cells closely apposed to the surfaces of hyalinized
fibrovascular cores; between these pseudopapillary structures are the neu-
ronal tumor cells (SYN positive), which can range morphologically from
neurocytic to ganglion cell-like. Mitotic activity, atypia, vascular prolifera-
tion, and necrosis are rare to absent (e-Fig. 41.36).
7. Rosette-forming Glioneuronal Tumor of the Fourth Ventricle (RGNT}, WHO
grade I. This tumot; most common in young adults, always involves the
ventricular system of the posterior fossa, and in unusual cases may extend
upwards into the proximal supratentorial ventricles (Am I Surg Pathol.
2002;26:582). Consequently, RGNT usually presents with sequelae of
obstructive hydrocephalus. MRI shows RGNT to be circumscribed and
solid, T2 hyperintense, T1 hypointense and, in some cases, heterogeneously
enhancing. Histologically (e-Fig. 41.37), RGNT is biphasic, characterized
by a uniform population of small neurocytic cells and a second glial ele-
ment that resembles PA. The neurocytic cells have hyperchromatic round
nuclei and a small amount of cleared cytoplasm; classically, they form sin-
gle or clustered rosettes and pseudorosettes within a relatively hypocellular
eosinophilic matrix of astrocytic tumor cell processes. The centers of these
rosettes and pseudorosettes are more intensely eosinophilic and contain
fine, SYN -immunoreactive cell processes. The surrounding matrix shows
immunoreactivity for GFAP, and may exhibit EGBs and RFs. Surgical resec-
tion is curative.
F. Pineal parenchymal tumors
1. Pineocytoma, WHO grade I, is more common in adults. On MRI, pineo-
cytoma appears as a Tt hypo- or isointense, T2 hyperintense, uniformly
enhancing, often calcified, discrete mass in the region of the pineal gland.
Through mass effect, pineocytoma often causes increased ICP and upward
gaze palsy (Parinaud syndrome). Histologically, pineocytoma tumor cells
(SYN+, Neu-N+) have round-to-oval nuclei, salt and pepper chromatin,
inconspicuous nucleoli, and poorly demarcated cell borders, and often
form pineocytic rosettes (large, exaggerated Homer-Wright rosettes) (e-Fig.
41.38). Mitotic activity is very low. Pineocytomas may be difficult to dis-
tinguish from normal pineal tissue in a small biopsy, although a lobular
pattern with gliovascular septae favors the latter.
2. Pineoblastoma, WHO grade IV, is a rapidly growing malignant tumor that
predominantly occurs in children and very young adults. Occasionally,
pineoblastomas occur in association with bilateral retinoblastoma (trilat-
eral retinoblastoma). On MRI, pineoblastomas are inconsistent: hypo- to
isointense on Tt, hypo- to hyperintense on T2, with homogeneous or het-
erogeneous enhancement. Their borders appear more infiltrative than those
of pineocytoma, and leptomeningeal metastasis (through CSF dissemina-
tion) is common at presentation. Grossly, they are soft, friable, and poorly
demarcated. Hemorrhage and necrosis may be present. Calcification is rare.
Histologically, they resemble other small blue cell tumors (e.g., medul-
loblastoma, PNET). The primitive-appearing cells have little cytoplasm,
hyperchromatic molded nuclei, and are densely packed in sheets occasion-
ally interrupted by Homer-Wright rosettes, Flexner-Wintersteiner rosettes
(which have a central lumen), and zones of necrosis. Mitotic activity is
generally high. Invasion of the adjacent pineal gland and leptomeninges is
common. Immunoreactivity for neuronal markers (e.g., NF, SYN, chromo-
granin, and neuron-specific enolase) is weak and focal relative to that seen
in pineocytoma. Occasionally, these tumor cells may additionally express
antigens related to photoreceptor or pineal differentiation, including retinal
S-antigen, rhodopsin, and melatonin.
3. Pineal parenchymal tumor of intermediate differentiation (PPTID), WHO grade
11-111, is a recently recognized entity that occurs more commonly in adults.
Histologically, PPTID is rather cellulat; lacks pineocytic rosettes, and has
slightly more pronounced cytologic atypia and a higher mitotidproliferative
rate than pineocytoma (MIB-1 indices from 10% to 16% for PPTID vs. 1%
to 2% for pineocytoma). PPTID appears in two main patterns: diffuse, with
mildly irregular round-to-oval nuclei arranged in a sheet-like pattern, and
pseudolobulated, wherein tumor cells are divided into lobules delineated
by vessels. A higher grade (Ill) is assigned to PPTID when the mitotic index
exceeds six mitoses per ten 40 x fields or when immunoreactivity for NF
protein is nearly absent; for this reason, definitive grading is best reserved
for resection rather than limited biopsy specimens. Recurrence and survival
rates are intermediate between those of pineoblastoma and pineocytoma.
4. Papillary tumor of the pineal region (PTPR), WHO grade II or Ill, has been
reported in children and adults, with a peak in the third decade. MRI shows
a large, well-circumscribed, T2-intense, enhancing (occasionally cystic)
mass in the vicinity of the pineal gland. Histologically, PTPR exhibits papil-
lary features; large, pale-to-eosinophilic tumor cells form columnar epithelia
around hyalinized vessels (e-Fig. 41.39). In other, more solid areas, tumor
cells with round-to-oval nuclei and clear or vacuolated cytoplasm may form
true ependymal rosettes. Mitotic activity is variable (0 to 10 per 10 high-
power fields), and necrosis may be seen; EH is usually not observed. PTPR
does not infiltrate the pineal gland.
This histologic appearance resembles that of ependymoma. Further,
both tumors can show focal membranous and dot-like reactivity for EMA
(consistent with the fact that PTPR is thought to arise from the specialized
ependymal cells of the subcommissural organ). The key features that dis-
tinguish PTPR are minimal reactivity for GFAP and strong reactivity for
CK18. Because PTPR has only recently been described, official grading cri-
teria have not yet been defined; howeve.t; a mitotic index of five or more per
ten 40x fields has been associated with poorer prognosis.
G. Embryonal tumors, most common in children, are so-named because their "small
blue cell" histology is reminiscent of elements of the embryonic nervous sys-
tem. These tumors are bulky, grow rapidly, and seed the CSF pathways. All
embryonal tumors are inherently malignant (WHO grade N); howeve.t; the
prognoses for these tumors vary widely, as some respond very well to current
treatments.
1. Medulloblastoma, WHO grade IV, the most common embryonal CNS tumo.t;
is thought to originate either from remnants of the external granular layer
or from the fourth ventricular germinal matrix. On imaging, they are often
T1 hypointense, T2 hyperintense, noncalcified, homogeneously contrast-
enhancing cerebellar masses with restricted diffusion. In approximately
one-third of the classic cases, the tumor has already seeded the CSF path-
ways focally (with so-called drop metastases in the lumbosacral spinal cord)
or diffusely (with so-called icing of the subarachnoid space) at the time
of presentation. Distant metastases, most often involving bone and lymph
nodes, are rare. Several histologic subtypes of medulloblastoma have been
described including classic/undifferentiated, desmoplastic/nodula.t; "with
extensive nodularity," large cell, and anaplastic.
Classidundifferentiated medulloblastoma (e-Fig. 41.40), the most com-
mon subtype, appears as a patternless sheet of abundant small tumor cells
with hyperchromatic nuclei and minimal apparent cytoplasm. The nuclei
show a degree of molding and are often described as round-to-oval or
carrot-shaped. Nucleoli are inconspicuous. Mitotic figures and apoptotic
bodies are often abundant. Microvascular proliferation and necrosis may
be present, but are not prominent. About 40% of the cases exhibit Homer-
Wright rosettes. In addition to seeding the CSF, this tumor often reinvades
the cerebellar parenchyma from the subarachnoid space via Vrrchow-Robin
spaces.
The desmoplastidnodular variant (e-Fig. 41.41) has a characteristic low-
power appearance of rounded, pale islands separated by dark internodu-
lar tissue. The pale islands (reticulin free) contain cells with a relatively
neurocytic phenotype that features round to oval nuclei, open chromatin,
moderate cytoplasm, cell processes that form a neuropil-like stroma, and
little mitotic activity. The internodular tissue (reticulin rich) is formed by
densely packed, primitive-appearing cells with hyperchromatic malleable
nuclei, smudged chromatin, little cytoplasm, and high proliferative activ-
ity. This variant is genetically distinct and has a slightly more favorable
prognosis than other subtypes (Acta Neuropathol. 2006;112:5).
Another prognostically favorable variant is medulloblastoma with
extensive nodularity. This rare tumo.t; found almost exclusively in infants,
radiographically may resemble a duster of grapes (T1 imaging, postcon-

trast). Histologically, it resembles the desmoplastic variant, but is dominated
by pale, reticulin-free nodules with neuronal differentiation; primitive areas
are diminished. Mter treatment, this tumor has been reported to undergo
complete gangliocytic differentiation.
In contrast, the rare large cell variant of medulloblastoma behaves rel-
atively aggressively and is less responsive to therapy. The cells of large
cell medulloblastoma have relatively more cytoplasm than those of classic
medulloblastoma, as well as larger nuclei, vesicular chromatin, and promi-
nent nucleoli. It is worth noting, however, that this large cell histologic
pattern may not be uniform within a tumor, and intermixed anaplastic
tissue is often associated.
Anaplastic medulloblastomas feature cellular pleomorphism, cyto-
megaly, hyperchromasia, a markedly elevated mitotic index, atypical
mitoses, abundant apoptotic bodies, apoptotic lakes, and cell wrapping
(e-Fig. 41.42). Although the common cooccurrence of large-cell and
anaplastic features has led some to advocate a combined large-celV
anaplastic medulloblastoma category, anaplasia can be observed in clas-
sic medulloblastomas that lack large-cell features.
Medulloblastomas are generally immunopositive for SYN and vari-
ably positive for GFAP, and have high MIB-1 (Ki-67) labeling indices.
Cytogenetic and FISH studies of medulloblastomas often show loss of
chromosome 17p and, when coupled with duplication of its long arm,
formation of isochromosome 17q (i17q). i17q is encountered in about
30% of medulloblastomas and is considered a relatively specific diagnostic
marker; whether it is associated with poorer prognosis is less clear. Other
genetic aberrations that clearly are associated with aggressive behavior and
shortened survival times are amplification of C-MYC, and less commonly
MYCN, oncogenes. These amplifications are observed in 4% to 17% of
medulloblastomas and are observed in a greater percentage of anaplas-
tic/large cell medulloblastomas. In contrast, immunoreactivity for nuclear
beta-catenin, which is observed in 15% to 20% of sporadic cases, is asso-
ciated with favorable prognosis.
The 5-year survival rate for medulloblastoma overall is 60%, following
current treatment protocols (gross total resection, craniospinal radiation
therapy, and adjuvant chemotherapy). Patients with the extensively nodu-
lar variant fare relatively better; other patients, particularly when younger
than 3 years and/or with limited surgical resections and/or with CSF dis-
semination, have relatively unfavorable prognoses.
2. CNS-PNET, WHO grade IV, is often histomorphologically identical to classic
medulloblastoma but occurs outside the cerebellum. On imaging, CNS-
PNET is contrast enhancing and may exhibit calcification, hemorrhage,
and/or necrosis. Some features of neuronal differentiation (e.g., diffuse
and widespread immunoreactivity for SYN, NF proteins, NSE) are com-
mon; reactivity for GFAP is usually strong within only a subset of cells. As
with medulloblastoma, several variants of CNS-PNET have been described.
Tumors with extensive neuronal differentiation have been alternatively
termed "cerebral neuroblastomas" or, when ganglion cells are observed,
ganglioneuroblastoma. Other rare cases showing multilayered true rosettes
(with central lumens) that blend into a surrounding sheet of dense tumor
cells are called ependymoblastoma (e-Fig. 41.43). A newly recognized tumor
has several of these features (Pediatr Develop Pathol. 2000;3:346) and is
aptly named "ETANTR" (embryonal tumor with abundant neuropil and
true rosettes) (e-Fig. 41.44 ); whether this aggressive tumor is distinct or rep-
resents a variant of CNS/PNET is not yet clear. A final variant of CNS/PNET
is medulloepithelioma, defined by arrangements of neoplastic neuroepithe-

lium reminiscent of the developing neural tube that appear among other
areas with PNET-like histology.
3. ATJRT, WHO grade IV (e-Fig. 41.45), is a densely cellular tumor of infants and
young children (most are <3 years old, with a male predominance) that
can occur anywhere in the neuraxis. Radiographically, AT/RT is usually
large, cystic, hemorrhagic, focally necrotic, and heterogeneously enhanc-
ing; not uncommonly, nodular leptomeningeal spread is found at presenta-
tion. Histologically, AT/RT is defined by rhabdoid cells with eccentrically
placed vesicular nuclei, prominent nucleoli, and large eosinophilic paranu-
clear whorls of intermediate filaments. The rhabdoid cells may appear only
focally, or not at all, in a biopsy specimen; tissue heterogeneity is com-
mon in AT/RT. A small blue cell component is seen in 65% of the cases
and may predominate. For this reason, AT/RT can easily be confused with
medulloblastoma (particularly the large celUanaplastic variant) or CNS-
PNET. Other areas of AT/RT may show glial, mesenchymal, epithelial, or
papillary features, potentially conjuring an even broader differential diag-
nosis. AT/RT typically shows immunoreactivity for vimentin, EMA, CK,
and smooth muscle actin, and often, for SYN and GFAP as well. Because
AT/RT can mimic other entities and is relatively common among infantile
CNS tumors, a very low threshold should be maintained to rule out this
diagnosis when confronted with a tumor in a child under 3 years of age. A
highly sensitive and specific immunohistochemical stain that differentiates
AT/RT from other embryonal tumors targets the BAF47/INI1 protein, the
product of the INI1/BAF47/hSNF5 tumor suppressor gene on chromosome
22q11. AT/RT results from biallelic inactivation of this gene, and hence
virtually all AT/RTs show loss of intranuclear reactivity for BAF47/INI1.
These are highly aggressive tumors; mean survival is 17 months.
H. Tumors of the meninges
1. Meningiomas, which bear resemblance to the arachnoidal cells that nor-
mally inhabit the inner surface of the dura, are most often intracranial and
extra-axial, appearing over the cerebral convexities, parasagittally along
the falx cerebri, along the skull base or tentorium, or in the optic nerve
sheath. Less commonly, meningiomas appear within the spine, where tho-
racic segments are favored. Rarely, meningiomas occur within a ventricle,
presumably arising from the tela choroidea, a leptomeningeal invagina-
tion at the base of the choroid plexus. Most meningiomas occur in adults
between 20 and 60 years of age, with a peak incidence around 45 years
and a slight female preponderance (female to male ratio of 3:2). Spinal
meningiomas are particularly more common in women (female to male
ratio of 9:1). Radiation-induced meningiomas (which can appear two or
more decades after radiotherapy for other brain tumors or for Tinea capitis)
are well recognized but rare. Many benign meningiomas (WHO grade I)
grow slowly and come to clinical attention incidentally or only after they
have grown to very large size and have begun to cause headaches, seizures,
or focal neurologic deficits by compressing adjacent structures. In contrast,
atypical (WHO grade II) and anaplastic (WHO grade III) forms can be
aggressive, show more rapid growth, a greater propensity to recur follow-
ing resection, and, in some cases, invasion of the CNS. It is important to
note, however, that even a benign meningioma can show invasion of soft
tissue and bone, and may come to clinical attention after invading the orbits
or sinuses; invasion of soft tissue and bone is not considered a criterion for
WHO grading. On imaging, meningiomas are extra-axial, homogeneously
enhancing lesions. Trailing of the enhancement into adjacent dura is a useful
radiologic sign (the so-called dural tail) but can also be observed in other
dura-based tumors. Hyperostosis of the adjacent skull is a suggestive radio-

graphic finding that is somewhat more specific, and is often associated with
bone invasion.
Grossly, meningiomas are spherical to lobulated, firm or rubbery, usually
well circumscribed, and firmly attached to the inner surface of the dura.
Meningiomas that occur along the sphenoid wing may grow "en plaque"
as flat, carpet-like masses. Most invade the underlying dura or dural sinuses
but do not involve the pia or the underlying CNS.
The microscopic features of meningioma are highly variable, and
13 histologic variants are recognized by the WHO classification system
(e-Fig. 41.46). However, the patterns that characterize these variants often
appear together within a single tumor. Although cytologic smear prepa-
rations of meningioma are occasionally nondiagnostic, they usually show
three-dimensional clusters (sometimes exhibiting whorls) of epithelioid to
spindled cells, reflecting the cells' rather cohesive nature. Tumor cell nuclei
are generally round to oval and often contain intranuclear pseudoinclu-
sions (cytoplasmic invaginations) and nuclear clearings; these latter features
are characteristic, but not specific. Likewise, concentric microcalcifications
(psammoma bodies) provide some diagnostic reassurance. Histologically,
the most common cytoarchitectural patterns in meningioma are fibrous
(fascicles of spindled cells) and meningothelial (dominated by whorls and
fascicles); identification of these patterns-even focally-may provide the
strongest initial clue to diagnosis, particularly when an uncommon histo-
logic pattern dominates the specimen. Nevertheless, familiarity with the
variants of meningioma is critically important to avoid misdiagnosis, par-
ticularly when specimens are small. In such cases, immunohistochemistry is
indispensible for confirming the diagnosis. Some of the common differen-
tial diagnoses for meningioma subtypes are provided in Table 41.5. Most
meningiomas display patchy or weak membranous reactivity for EMA and
reactivity within nuclei for progesterone receptor (PR) (although reactivity
for PR is less common among atypical and anaplastic tumors). Relevant for
the often-encountered differential diagnosis of schwannoma versus menin-
gioma for a spindled CNS neoplasm, collagen IV reactivity is not observed
in meningioma, and S100 reactivity is typically patchy rather than diffuse. In
unusual circumstances, when immunohistochemistry fails to provide diag-
nostic clarity, EM may be helpful; meningioma cells have interdigitating cell
processes with scattered desmosomes, and do not secrete a basement mem-
brane. FISH for common genetic changes may also be useful for guiding
diagnosis (see below).
a. Histologic variants, grading, and prognosis. Roughly 80% of meningiomas
are considered histologically benign (WHO grade I) and have a low
risk of recurrence after gross total resection (rv5% at 5 years, rv20%
at 20 years). Atypical (WHO grade IT) meningiomas constitute 15% to
20% of cases and have a higher risk of recurrence (""30% or more at
5 years). Anaplastic (WHO grade III) meningiomas account for 1% to
2% of cases, are associated with even higher recurrence rates, and often
prove fatal (median survival <2 years). After histologic grade, the most
influential prognostic factor for recurrence is extent of surgical resection;
subtotally resected benign meningiomas have a recurrence rate of rv30%
to 40% at 5 years. Other prognostic factors include male gender and
young age; each is unfavorable. Genetic characteristics of a meningioma
also influence prognosis, as discussed below.
One determinant of WHO grade is the histologic pattern.
Meningothelial, fibrous, transitional, psammomatous, angiomatous,
microcystic, secretory, lymphoplasmacyte-rich, and metaplastic variants
are considered WHO grade I unless they have additional superimposed
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Consistent with increased mitotic activity, atypical meningiomas also

generally exhibit moderately elevated Ki-67 (MIB-1 antibody) labeling
indices. Because measurements of Ki-67labeling index can show interin-
stitutional variability, this feature is not officially considered in the WHO
grading system. However, it does represent a reasonable surrogate for
mitotic activity when cytologic preservation is poor or when a specimen
is particularly small.
c. Brain invasion. Brain invasion by meningioma is characterized by
entrapped islands of brain parenchyma at the periphery of the tumor
and/or irregular tongue-like protrusions of tumor tissue in attached brain
parenchyma. On H&E sections, this phenomenon can be quite subtle,
so immunohistochemistry for GFAP is often warranted for confirmation
(e-Fig. 41.48). Brain invasion may occur in tumors that are otherwise his-
tologically benign, atypical, or anaplastic. Although brain invasion has
historically been assumed to be a malignant feature, clinicopathologic
correlation studies have revealed statistical outcomes more consistent
with atypical meningiomas. Therefore, brain-invasive meningiomas are
considered WHO grade II even when their histologic patterns are other-
wise benign.
d. Anaplastic (malignant) meningioma, WHO grade Ill. Anaplastic menin-
giomas that are not overtly papillary or rhabdoid must exhibit a
markedly elevated density of 20 or more mitotic figures per ten 40x
fields or show histologic features of frank malignancy reminiscent of
carcinoma, melanoma, or high-grade sarcoma (Cancer 1999;85:2046)
(e-Fig. 41.49). Consequently, the differential diagnosis for these anaplas-
tic meningiomas often includes one or more of these malignant tumors.
Complicating this situation somewhat is the tendency for meningioma
to harbor metastatic carcinoma from outside the CNS (most commonly
breast and lung). In such cases, when immunohistochemical data are
insufficient to support a diagnosis with confidence, ultrastructural and
FISH studies probing for genetic changes common to meningioma may
be useful.
e. Genetic changes in meningioma. Most meningiomas of all grades show
loss of chromosome 22q, site of the NF2 gene, thought to be important
for tumor development. A genetic alteration with prognostic implications
is loss of the CDKN2A/p16 region on 9p21. Characteristic of the major-
ity of anaplastic meningiomas, loss of 9p21 is associated with decreased
survival; tumors that lack this alteration show survival patterns more
consistent with atypical rather than anaplastic grade.
2. Hemangiopericytoma {HPC), WHO grade 11-111, and Solitary Fibrous Tumor
{SFT), ungraded, but benign (e-Fig. 41.50). Historically, these two uncommon
tumors have been regarded as distinct entities, but their overlapping clinical
and histologic features suggest that they may represent ends of a continuum.
Radiologically, they resemble meningioma; they are dura-associated, solid,
uniformly enhancing, and appear in a similar anatomical distribution. How-
ever, unlike meningioma, HPC does not show intratumoral calcification
and can be associated with bone lysis rather than hyperostosis. Addition-
ally, HPC appears hypervascular by angiography, and may receive a dual
blood supply from the meninges and brain. Microscopically, this hyper-
vascularity is contributed by slit-like vascular channels lined by flattened
endothelial cells and frequent ectatic, thin-walled and branching "staghorn
vessels." Although such vessels are not specific, they provide a very helpful
diagnostic clue.
The parenchyma of these tumors is usually biphasic. Hypocellular areas
of SFT are strongly collagenous and hyalinized; cellular areas of SFT are
composed of a patternless or mildly fascicular arrangement of elongated

spindled-to-oval cells that are intermixed with brightly eosinophilic, inter-
laced collagen bundles. Mitotic figures are sparse, and other classically
anaplastic features are not observed. When the cellular areas of SFT become
particularly dense, they may resemble HPC. Cellular areas of HPC contain
oval-to-epithelioid cells with round nuclei and scant cytoplasm that are
densely arranged in a random or vaguely nested pattern with little inter-
vening stroma; collagen is not prominent. Within this dense background,
paucicellular areas of HPC appear as pale islands. A diagnosis of anaplasia
for HPC requires either >5 mitotic figures per ten 40x fields or the presence
of necrosis, plus two or more of the following: hypercellularity, moderate or
severe nuclear atypia, and hemorrhage. In ambiguous cases, special stains
can guide diagnosis; HPC shows at best only focal immunoreactivity for
CD34 but has abundant stromal reticulin fibers that surround cells individu-
ally and in small groups; SFT stains strongly for CD34 and is reticulin-poor.
Both neoplasms show immunoreactivity for vimentin, bcl-2, and CD99, and
no reactivity for S-1 00, CD 31, EMA, and PR. FISH studies can also be help-
ful, as loss of chromosome 22q has not been reported in HPC. It is worth
noting, however, that loss of 9p21, a poor prognostic marker in high-grade
meningiomas, has been reported in about 25% of HPCs.
I. Hemangioblastoma {HB), WHO grade I. Sporadic cases of this benign, slowly
growing, highly vascular neoplasm occur most often in the posterior fossa
of adults, favoring the cerebellum. HBs located elsewhere (e.g., brain stem,
spinal cord, retina, cerebrum), particularly when multiple and/or occurring
in younger patients, are more commonly associated with von Hippel-Lindau
disease (VHL); therefore, discovery of an HB with any of these unusual char-
acteristics warrants thorough screening for other manifestations of VHL (e.g.,
endolymphatic sac tumor, pheochromocytoma, pancreatic cyst or islet tumor,
renal cyst or clear cell carcinoma, and papillary cystadenoma of the epi-
didymis). On imaging studies, the classic HB is well-circumscribed, intra-axial,
and cystic, with an enhancing mural nodule that is hypervascular on angiogra-
phy. Microscopically, hemangioblastomas have two main components: stromal
and vascular. Stromal cells show variable morphology, but typically are large
and polygonal with vacuolated lipid-laden cytoplasm (e-Fig. 41.51). Kary-
omegaly and nuclear degenerative atypia among these stromal cells are com-
mon and without significance. The vascular component is characterized by
thin-walled channels lined by flattened endothelial cells. On the basis of the
relative prominence of these two components, two histologic variants have
been described: cellular and reticular. The less common stroma-rich cellular
variant may show greater Ki-67labeling index and propensity to recur than the
more common reticular variant. Mast cells are common within HBs and foci of
extramedullary erythropoiesis are present in a minority of cases. Occasionally,
these tumors produce brisk piloid gliosis in the adjacent brain parenchyma.
On the basis of radiographic and histologic patterns, the differential diagno-
sis often includes PA and metastatic renal cell carcinoma (RCC), requiring
immunohistochemical stains such as D2-40/podoplanin or inhibin (positive in
stromal cells), as well as EMA, CD10, and RCC (all variably positive in RCC)
and GFAP (positive in PA and most often negative in stromal cells).
J. Tumors of the sella and/or suprasellar region {other than pituitary adenomas).
1. Craniopharyngiomas, WHO grade I, due to their location, may present clin-
ically with hormonal abnormalities, vision disturbance, and obstructive
hydrocephalus. These partly cystic, histologically benign epithelial tumors
are thought to originate from Rathke's pouch epithelium. On MRI, these
tumors often show T1 bright cyst contents and postcontrast enhance-
ment of solid areas and any cyst capsule(s). Two morphologically distinct
subtypes have been described, adamantinomatous and papillary; the for-

mer shows heterogeneous calcifications on CT, whereas the less common
papillary variant does not.
a. Adamantinomatous craniopharyngiomas occur more commonly in chil-
dren aged 5 to 15 years, but show a second peak in the fifth and sixth
decades. Grossly, cyst contents have the appearance of dark green-brown
machine oil. Calcifications are frequently noted. Microscopically, this
variant resembles adamantinoma of the bone (tibia) and ameloblastoma
of the jaw. The adamantinomatous variant is formed by a complex squa-
mous epithelium that shows: basal palisading (a columnar orientation of
the most peripheral layer of nuclei); an overlying zone of moderate cellu-
larity in which focal areas resemble a network of processes over a cleared
background (stellate reticulum); and central keratinization, which pro-
duces nodules of pale, anucleate, cohesive cell remnants (wet keratin)
(e-Fig. 41.52). Areas of xanthogranulomatous inflammation and abun-
dant cholesterol clefts are common. The border of this tumor is usu-
ally irregular and adherent to surrounding structures, and the adjacent
parenchyma typically shows robust piloid gliosis. Although histologi-
cally benign, its adherent somewhat invasive nature leads to frequent
recurrence even after gross total resection. Consequently, adamantino-
matous craniopharyngioma can cause considerable morbidity, particu-
larly given its close association with the optic chiasm and pituitary.
b. The papillary variant (e-Fig. 41.53) is less common, is not typically seen in
children, and most commonly occurs in the third ventricle or suprasel-
lar region rather than in the sella itself. Grossly, it is seldom cystic; any
cysts observed contain clear fluid rather than "machine oil." Calcifi-
cations are absent, and the interface with the brain is smoother and
less adherent. Histologically, the papillary variant is formed by a well-
differentiated, nonkeratinizing squamous epithelium that overlies broad,
loose fibrovascular cores. The basal layer shows increased density, but
without strong palisading. Beyond the apical surface, the tissue degrades
(undergoes dehiscence), effectively leaving crude pseudopapillae of viable
tissue around stromal cores. Occasionally, the epithelium may show focal
cilia or goblet cells. In spite of clearer demarcation from the adjacent
brain tissue in the papillary variant, the tumor has a similarly high rate
of recurrence as the adamantinomatous variant.
K. Germ cell tumors (GCTs). Included in this category are germinoma, teratoma,
yolk sac tumor, embryonal carcinoma, and choriocarcinoma. Typically, these
brain tumors occur in children or young adults. A pineal location, classi-
cally associated with increased intracranial pressure and Parinaud syndrome,
is more common in males; a suprasellar location, classically associated with
diabetes insipidus, vision changes and hypopituitarism, is slightly more com-
mon in females. In some cases, both areas are involved. Less common sites
include the basal ganglia and thalamus. On MRI, most GCTs are heteroge-
neously T2 hyperintense and show heterogeneous enhancement. Calcification
and complexity are common in teratomas, and hemorrhage is strongly asso-
ciated with choriocarcinoma. Histologically, these tumors are homologous to
their gonadal and mediastinal counterparts. Occasionally, the marked lym-
phocytic infiltrate and/or granulomatous response seen in germinomas can
mask the underlying large, clear cells with central round nuclei and large
nucleoli that define the neoplasm. In such cases, immunostaining for c-kit
(membranous pattern), OCT4 (nuclear pattern), and placental alkaline phos-
phatase (PLAP) (cytoplasmic and membranous pattern) may be applied to aid
diagnosis (Table 41.1, e-Fig. 41.54). Rare germinomas contain syncytiotro-
phoblastic elements and should not be confused with choriocarcinoma. Pure
germinomas are exquisitely radiosensitive and have an excellent prognosis;

those with syncytiotrophoblastic cells show a greater tendency to recur, and
modestly less favorable survival. Mature teratomas follow a benign clinical
course and may be cured by gross total resection. Yolk sac tumors, embryonal
carcinoma, and choriocarcinoma (and mixed tumors composed of more than
one of these elements) confer less favorable prognoses.
L. Lymphomas and histiocytic tumors.
1. Primary CNS lymphomas (PCNSLs) are malignant lymphomas that occur in
the absence of systemic lymphoma. PCNSL is more common among but not
exclusive to elderly or immunosuppressed patients. On imaging, PCNSLs
occur as single or multiple homogeneously enhancing lesions; some appear
well circumscribed (resembling, e.g., a metastasis or tumefactive MS) and
others appear somewhat diffuse (resembling a diffuse glioma). A periven-
tricular location is common. These lesions often respond dramatically to
steroid therapy, at least initially; although this radiographic response may
provide a diagnostic clue, steroid treatment is usually withheld when pos-
sible until a tissue diagnosis is secured.
Microscopically, PCNSLs appear as parenchymal and angiocentric
infiltrates of highly atypical lymphocytes (e-Fig. 41.55) accompanied by
nonneoplastic CD3+ T cells. The neoplastic cells usually have centroblast-
like or immunoblast-like morphology and are immunoreactive for B-
lymphocyte markers (e.g., CD20, CD79a); most PCNSLs are considered
diffuse large B-cell lymphomas (DLBCLs). Mfected vessels show mural
expansion by this mixed population, a feature that can be highlighted by
reticulin stain. In some cases, nonneoplastic foamy histiocytes and reactive
astrocytes may also be abundant in the parenchyma. Necrosis and evidence
of Epstein-Barr virus (EBV) (by immunostain or in situ hybridization for
EBV-encoded RNA [EBER]) are features associated with acquired immun-
odeficiency syndrome (AIDS) or immunosuppression-associated PCNSL.
Uncommon variants of lymphoma that affect the CNS include: lym-
phomatoid granulomatosis, an angiodestructive EBV-driven high-grade
B-celllymphoma that affects the brain in 25% of cases (more commonly, the
lung and skin) and features fibrinoid angionecrosis and poorly formed gran-
ulomas (lymphohistiocytic nodules); and intravascular B-celllymphoma, a
high-grade extranodal B-celllymphoma characterized by its restriction to
vessel lumina. Both of these variants lead to vascular occlusion and infarcts.
T-celllymphomas are encountered only rarely as PCNSLs. When lymphoma
metastasizes to the CNS (secondary CNS lymphoma) it typically appears in
epidural, dural, or leptomeningeal locations and is accompanied by systemic
disease.
2. Histiocytic Disorders
a. Langerhans cell histiocytosis (LCH) is the term officially endorsed by
the Histiocyte Society to replace previous terms (e.g., Hand-Schiiller-
Christian disease [HSCD], eosinophilic granuloma, histiocytosis X, etc.)
LCH primarily affects children, with a median age of 12 years. The
most common manifestation involves an isolated osteolytic lesion of
the skull (eosinophilic granuloma) that may also involve the underly-
ing meninges and cortex. When multiple, such lesions may be accom-
panied by hypothalamic involvement (consistent with HSCD). Although
it is rare to encounter isolated involvement of the hypothalamus, pitu-
itary, and optic chiasm (e-Fig. 41.56), this region shows radiographic
changes in"' 50% of cases with disseminated LCH, and 25% of children
with multifocal disease present with diabetes insipidus. Histologically
(e-Fig. 41.57), LCH is characterized by Langerhans cells accompanied
by variable amounts of nonneoplastic reactive elements that may include
eosinophils, neutrophils, macrophages, lymphocytes, plasma cells, and

multinucleated giant cells. Langerhans cells are recognizable morpholog-
ically by their grooved "coffee bean" nuclei; immunohistochemically by
reactivity for S100, CDla, and langerin (CD207); and ultrastructurally
by the presence of Birbeck granules.
b. Non-LC Histiocytic disorders are far less common in the CNS. Intracra-
nial Rosai-Dorfman disease (e-Fig. 41.5 8) affects children and adults and
can mimic meningioma; it most often affects the meninges as a contrast-
enhancing mass, and may induce a nonneoplastic hyperplastic response
in the associated arachnoidal cells. However, the other constituents of the
tumor include plasma cells, lymphocytes, and, most importantly, scat-
tered large pale histiocytes (S-100+, CD68+, CDla-) with prominent
nucleoli. In a majority of cases, these large histiocytes exhibit the diagnos-
tic feature of emperipolesis (engulfment of lymphocytes or plasma cells).
Erdheim-Chester disease affects adults and may involve any part of the
CNS or its coverings. Meningeal and perivascular lesions feature foamy
histiocytes (S100 variable, CD68+, CD1a-, factor Xilla+) but cells in
the parenchyma may resemble activated microglia or neoplastic astro-
cytes; sparse lymphocytes, plasma cells, eosinophils, and multinucleated
histiocytes may also be seen. Definitive diagnosis usually requires corre-
lation with clinical and radiologic evidence of systemic involvement, par-
ticularly bone pain and sclerosis of the long bones. Juvenile xanthogran-
uloma (JXG) (e-Fig. 41.59) usually appears as an isolated cutaneous
nodule in children, but can also occur as an isolated lesion in the brain
or meninges. The histiocytes of this lesion are S-100-, CD68+, CD1a-,
Factor X111a+, CD11c+, and lysozyme negative, an immunopheno-
type similar to that of plasmacytoid monocytes. Scattered Touton giant
cells, lymphocytes, and eosinophils are commonly seen. JXG is benign,
but may cause seizures. Histiocytic sarcoma is a very rare tumor that
shows cytologic atypia, necrosis, and a high proliferative index; stains
only with macrophage markers CD68 and CD 163; and is associated with
poor prognosis.
M. Metastatic tumors to the CNS are the most common CNS neoplasms, and occur
in up to 30% of adult and 6% to 10% of pediatric cancer patients. Common
sources in adults include carcinomas of lung, breast, kidney, and colon, as well
as melanomas. In children, leukemia, lymphoma, osteogenic sarcoma, rhab-
domyosarcoma, and Ewing sarcoma are the most common primaries. Radio-
logically, the majority of CNS metastases implant in the cerebral hemispheres at
the junction of cortex and white matter, appear well circumscribed, and show
ring-like or diffuse contrast enhancement. Histologically and immunopheno-
typically, metastases usually resemble their primary tumors at least focally; it
is therefore always worthwhile to review slides from a putative primary tumor
whenever possible.
VI. INFLAMMATORY AND INFECTIOUS DISORDERS
A. Inflammatory demyelinating disorders are characterized by myelin loss with rel-
ative preservation of axons; this finding is often accompanied by abundant
foamy macrophages and perivascular lymphocytes. Classic radiographic exam-
ples of myelin disorders are not generally biopsied, but when clinical and/or
radiologic data are unusual or ambiguous, neurosurgery may be recommended
to obtain a tissue sample.
1. Multiple sclerosis (MS) is an idiopathic demyelinating disorder with genetic,
environmental, and infectious influences. Historically, several subtypes have
been described, including relapsing-remitting, secondary progressive, pri-
mary progressive, acute monophasic (Marburg disease), and acute tumefac-
tive. The relapsing-remitting form (RRMS), characterized by intermittent
attacks and partial recovery from neurologic deficits during periods of
remission, is more common in young adults (women > men with a ratio of
2:1) and is themostcommonformin the United States (""80% of cases). Less
common forms involve rapid, unremitting clinical deterioration-either de
novo (primary progressive MS [PPMS]) or in the setting of RRMS (sec-
ondary progressive MS [SPMS]). Demyelinative MS plaques in these sub-
types most often involve white matter periventricularly, and in the optic
pathway, brain stem, and/or spinal cord. Radiographically, chronic inac-
tive plaques are hypointense on T1 and diffusion-weighted images; plaques
with active inflammation are hyperintense on T2 and show postcontrast
enhancement. This latter feature can be responsible for some radiolog-
ical diagnostic uncertainty in cases of Marburg disease or acute tume-
factive MS. In acute tumefactive MS, a large isolated plaque appears as
a T2-hyperintense, peripherally enhancing lesion that resembles a high-
grade glioma or lymphoma. Although the rim of enhancement around
the demyelinative lesion may be incomplete at the cortical side (forming
a horseshoe-shaped profile that would be highly unusual in a neoplasm),
this feature is not robust enough for conclusive diagnosis; consequently,
such lesions, although uncommon, are often biopsied. At frozen section,
tumefactive MS can represent a great diagnostic challenge; in such cases,
examination of a cytologic smear preparation is often very helpful as it
preserves important cellular details that can be obscured by freezing arti-
fact. Microscopically, an active demyelinative MS plaque shows (1) loss
of myelin, (2) relative preservation of axons, (3) perivascular nonneoplas-
tic lymphocytic infiltrates, (4) numerous foamy macrophages, (5) reactive
astrocytosis, and (6) cerebral edema. Mitotic activity among astrocytes and
macrophages may be brisk, but nuclear atypia should be minimal. Gran-
ular mitoses and Creutzfeldt cells (which contain multiple micronuclei),
although not specific for MS lesions, are characteristic. Myelin loss with
axonal sparing (which distinguishes this lesion from an axon-destroying
infarct) is best visualized by comparing Luxol fast blue-PAS (LFB-PAS)
stained sections to others stained with Bielschowsky silver or NF protein
immunohistochemistry (e-Fig. 41.60). On LFB-PAS sections, the border of
demyelination is usually abrupt, and blue-green debris (representing par-
tially metabolized myelin) is visible within macrophages. In rare cases that
cannot be satisfactorily diagnosed with special stains, FISH studies can be
applied to evaluate for glioma-associated chromosomal changes.
2. Acute disseminated encephalomyelitis (ADEM}Iperivenous encephalomye-
litis represents an unusual autoimmune response among children and young
adults that is induced by recent infection by measles (most commonly,
but also), mumps, varicella, influenza, rubella, Campylobacter jejuni, or
Mycoplasma pneumoniae or vaccination (smallpox or rabies). Symptoms
include headache, fever and vomiting, followed by weakness, ataxia, and
visual and sensory loss with progression to stupor and seizures. Radio-
logically, ADEM usually appears as many small T2-weighted and FLAIR
hyperintensities in subcortical and periventricular white matter and the
spinal cord; the deep gray matter and cortex may also be affected. This
clinicoradiographic pattern is characteristic enough that biopsy is not
always performed. Nevertheless, when obtained, biopsy material shows
perivenous demyelination with axonal sparing, mononuclear infiltrates,
macrophages, activated microglia, and occasional petechial hemorrhages.
ADEM is thought to result from a myelin-directed, cross-reactive immune
response to a foreign antigen.
3. Demyelinating viral infections. Viral infections that commonly cause CNS
demyelination include human immunodeficiency virus (HIV leukoen-
cephalopathy and vacuolar myelopathy [HIVL, HIVVM]), human
T-celllymphotropic virus-1 (HTLV-associated myelopathy/tropical spastic
paraparesis [HAM!fSP]), JC virus (progressive multifocal leukoen-

cephalopathy [PML]), and measles virus (subacute sclerosing panencephali-
tis [SSPE]).
a. HIVLJHIVVM. White matter damage by HIV is thought to be mediated
by virally infected macrophages and giant cells; oligodendrocytes are
not directly infected. HIVL is often associated with HIV-associated
encephalitis (HAVE) but begins earlier, often introducing cognitive
impairment (up to 50% of AIDS patients develop dementia ascribed
to HIVL and 1-llVE). Histologically, HIVL may show subtle white mat-
ter pallor, with variable numbers of microglial nodules, macrophages,
and multinucleated giant cells. HIVVM targets the spinal cord in ""25%
of AIDS cases, producing vacuolar myelopathy in the dorsal and lateral
columns that resembles subacute combined degeneration.
b. HAMITSP, characterized by paraparesis, spasticity, and hyperreflexia of
the lower extremities and positive Babinski sign, affects 1% to 5% of
individuals who are seropositive for HTLV-1. As the clinical presenta-
tion suggests, HAM!fSP preferentially affects the lateral columns (cor-
ticospinal tracts), most severely in the thoracic spinal cord; correspond-
ing changes (T2 hyperintensity, thoracic spinal cord atrophy) are visible
radiologically. Although biopsy is uncommonly undertaken, histology of
the cord shows leptomeningeal and perivascular lymphocytic infiltrates,
foamy macrophages, widespread myelin loss, and axonal dystrophy of
the lateral columns.
c. PML is caused by reactivation of a latent JC papovavirus (much of the
general population is seropositive) in the setting of immunosuppression
(e.g., lymphoproliferative disorders, AIDS, monoclonal antibody treat-
ments for autoimmune diseases). Patients experience 3 to 6 months of
progressive neurologic symptoms that reflect the anatomic distribution
of the lesions, which favor the subcortical and deep cerebral white matter,
cerebellum, brainstem and, rarely, the spinal cord. Personality changes
are often prominent, followed by dementia and death. In some cases,
restoration of immunocompetency may halt the disease; in other cases,
the restored inflammatory response hastens clinical decline (the so-called
immune reconstitution inflammatory syndrome [IRIS]). On MRI, the
lesions of PML are T2-hyperintense, T1-hypointense, and nonenhanc-
ing; however, enhancement is common in the setting of IRIS. Histologi-
cally, PML shows foci with myelin loss, a variably modest lymphocytic
infiltrate (more pronounced in IRIS), and, within the center of larger
lesions, few oligodendrocytes. Some oligodendrocytes show the hallmark
lesion of PML, a large nucleus with marginated chromatin and a glassy
inclusion that appears plum colored on H&E sections (e-Fig. 41.61).
Ultrastructurally, the papovavirus particles exhibit a "stick and ball" or
"spaghetti and meatballs" pattern. Also noted within and around these
lesions are pseudoneoplastic astrocytes with atypical nuclei; these cells
do not appear to produce or contain virus particles, and should not be
mistaken for astrocytoma. Immunohistochemistry for JC virus proteins,
EM, and in situ hybridization studies may be used to confirm the presence
of the virus.
d. SSPE (e-Fig. 41.62) is a rare sequela of measles in which a defective
measles paramyxovirus targets oligodendrocytes and neurons resulting
in a slowly progressive encephalitis that results in coma and death. SSPE
may present as many as 3 to 10 years after initial infection. Early in
the course, radiographic findings are minimal; periventricular T2- and
FLAIR hyperintensities appear late. Antimeasles IgG titers in CSF may
be elevated. Histologically, SSPE shows large areas of myelin loss and
neuron loss, leptomeningeal and perivascular lymphocytic infiltrates,

and, most importantly, eosinophilic viral inclusions in the nuclei of oligo-
dendrocytes and neurons. Diagnosis may be confirmed through immuno-
histochemistry for measles virus associated proteins.
B. CNS Infections may be broadly classified into meningitides, encephalitides, and
abscesses.
1. Meningitis may be caused by bacteria, mycobacteria, viruses, fungi, and
parasites. Although CSF analysis is often sufficient for diagnosis, biopsy is
occasionally necessary.
a. Bacterial meningitis is characterized in its acute stage by abundant neu-
trophils within the subarachnoid space, and subpial reactive gliosis.
Organisms are usually difficult to identify. In the chronic stage, neu-
trophils are supplanted by mononuclear cells, and granulation tissue and
fibrosis may appeal:.
b. Tuberculous meningitis can mimic bacterial meningitis, but more com-
monly exhibits a patchy granulomatous infiltrate of epithelioid histio-
cytes, multinucleated giant cells, and mononuclear cells. As with bac-
terial meningitis, organisms are difficult to identify, even with acid-fast
stains; ancillary testing (PCR, culture) may aid diagnosis.
c. Viral meningitis often shows meningeal and perivascular lymphocytic
infiltrates that may extend into Virchow-Robin spaces. Because these
infiltrates can be minimal and patchy, absence of inflammation on biopsy
does not rule out the diagnosis. Identification of microglial nodules in
the parenchyma supports an additional diagnosis of viral encephalitis.
d. Fungal meningitis provokes a mononuclear/granulomatous inflammatory
response similar to that of tuberculous meningitis, but the organisms are
usually easily identified by histochemical stains. Nevertheless, culture
is required for definitive speciation. In contrast to yeast forms (Histo-
plasma, Blastomyces, Cryptococcus), pseudohyphal and hypha! organ-
isms (Candida, Aspergillus, Zygomycetes, Fusarium, Coccidioides) can
be angioinvasive and compromise blood supply leading to infarction.
2. Encephalitis is most often caused by viruses, and less commonly by parasites.
a. Viral encephalitis is recognized histologically by the presence of
meningeal and perivascular lymphocytes accompanied by parenchymal
microglial nodules. Occasionally, a microglial nodule may be observed
around a dying neuron (termed neuronophagia). Hundreds of viruses can
cause encephalitis, and most do so without forming distinctive inclu-
sions, so serologidlaboratory tests and clinical observations are usu-
ally required for diagnosis. Nevertheless, a small subset of pathogens is
responsible for most clinically significant cases.
i. West Nile Virus (WNV) has become the leading cause of epidemic viral
encephalitis in the United States within the last decade, and now
spans the continent. This arbovirus (arthropod-borne) infection has
a peak incidence in the summer or early autumn. In the acute phase,
WNV can induce a neutrophil response that is visible in the meninges
and CSF. In the chronic phase, microglial nodules are most abundant
in the spinal cord, thalamus, and substantia nigra pars compacta.
WNV encephalitis is best diagnosed by detection of specific IgM in
the CSF rather than by PCR.
ii. Herpes encephalitis is the most common cause of sporadic viral
encephalitis is the United States. Asymmetric, bilateral involvement
of the temporal lobes is characteristic, and when severe involves
hemorrhage and necrosis. Clinically, involvement of the temporal
lobes may cause hallucinations, agitation, personality changes, and
psychosis. PCR testing of CSF for HSV shows high sensitivity and
specificity, but sensitivity might be lower early in the course of

infection. Histologically, in addition to perivascular lymphocytes
and microglial nodules, biopsy material may show areas of necro-
sis with foamy macrophages and hemorrhage. Intranuclear Cowdry
A ("owl's eye") and Cowdry B (eosinophilic translucent) inclusions
may also be visible in some cases. Immunohistochemistry or in situ
hybridization for HSV-1 and HSV-2 may be applied to confirm the
diagnosis.
iii. Varicella zoster virus (VZV) encephalitis is more common with
immunosuppression, and targets several structures leading to dif-
ferent injuries. By infecting and damaging large- and medium-sized
vessels, VZV causes bland and hemorrhagic infarcts. By damaging
small vessels, VZV causes small, deep, ovoid ischemic lesions, and
then infects the newly exposed oligodendrocytes, creating areas of
demyelination; intranuclear Cowdry A inclusions (e-Fig. 41.63) are
often visible within glial cells at the lesion's edges. Less commonly,
VZV infects ependymal cells causing ventriculitis.
iv. HIV encephalitis (microglial nodule encephalitis), in conjunction with
HIV leukoencephalopathy, is associated with the dementia that
complicates AIDS. The histopathology features widespread, robust
microglial nodules with associated lymphocytes, reactive astrocytes,
and occasional multinucleated giant cells that harbor the virus. Neu-
rotoxic cytokines and other damaging chemical agents (e.g., reactive
oxygen species) may also play a role in this poorly understood demen-
tia, which is also associated with cortical and subcortical atrophy.
v. Rocky Mountain spotted fever is caused by rickettsia, but can mimic
viral encephalitis and is therefore mentioned here. The organism tar-
gets vascular endothelium and smooth muscle cells, and thus pro-
duces vasculitis, thrombosis, microinfarcts, and petechial hemor-
rhage without fibrinoid necrosis. Microglial nodules and mononu-
clear infiltrates with a leptomeningeal and perivascular distribution
are commonly seen.
b. Parasitic encephalitis
i. Neurocysticercosis (e-Fig. 41.64 ), caused by the pork tapeworm, Tae-
nia solium, is the most common CNS parasitic infection worldwide
and a common cause of seizures. The larvae form thin-walled cysts
in muscle, brain, eyes, liver, and lung. Each cyst contains a scolex
that can be detected radiographically or histologically and serves as
the pathognomonic feature of the lesion. Death of the organism trig-
gers a brisk mixed and granulomatous inflammatory response. Over
months to years, inflammation subsides; the cyst becomes fibrotic
and, eventually, a small calcified nodule.
ii. Cerebral malaria affects fewer than 10% of infected individuals, favor-
ing those without immunity (<4 years of age, foreign visitors). Pre-
senting symptoms include fever, headache, backache, photophobia,
vomiting, variable neurologic deficits, and seizures. The disorder is
caused by the deposition/lodging of parasitized erythrocytes within
the microvasculature of the brain, which is accompanied by petechial
hemorrhages in the white matter around necrotic vessels. Over time,
the ring hemorrhages are resorbed by microglia, macrophages, and
astrocytes, forming Di.irck granulomas. Prompt treatment with cor-
ticosteroids and antimalarial drugs reduces mortality.
iii. Amebic encephalitis (AE) is caused by several organisms, includ-
ing Naegleria fowleri, Balamuthia mandrillaris, and Acanthamoeba
species; Entamoeba histolytica tends to form abscesses. N. fowleri-
usually encountered during fresh water swimming-enters the CNS
through the cribriform plate; the other parasites enter hematoge-

nously from other infected organs. Clinically, AE with N. fowleri
presents in previously healthy children and young adults as acute
meningitis and progresses to coma and death in 2 to 3 days. Histo-
logically, mononuclear inflammation of the meninges is scanty, but
the adjacent brain shows extensive hemorrhagic necrosis. Organ-
isms are visible in the subarachnoid space and around vessels. These
trophozoites bear a striking resemblance to foamy macrophages but
have a prominent central nucleolus. AE with B. mandrillaris and
Acanthamoeba has a similar dismal prognosis, but a more chronic
course.
iv. Toxoplasmosis is seen in neonates as a congenital infection (one of the
TORCH infections) that leads to microcephaly with dystrophic cal-
cifications. Later in life, infection may cause feve.r; a maculopapular
rash, and malaise or may be asymptomatic. In the setting of sub-
sequent immunosuppression (particularly AIDS) a dormant infec-
tion can undergo reactivation and present with multiple deep-seated
and/or cortical enhancing lesions. Grossly, the focal lesions of cere-
bral toxoplasmosis are multiple, discrete, and necrotic; less com-
monly, toxoplasmosis may appear as a more diffuse encephalitis. His-
tologically (e-Fig. 41.65), the necrotic lesions show peripheral mixed
inflammatory infiltrates, neovascularization, and gliosis. Perivascu-
lar inflammation and fibrinoid necrosis of vessels may also be seen.
Although H&E stains are usually adequate for identifying Toxo-
plasma gondii. immunostains for the protozoan offer greater sensi-
tivity and specificity. In unusual cases when cerebral toxoplasmosis
manifests diffusely without necrotic foci, histologic changes include
microglial nodules and reactive astrocytosis.
3. Brain abscess may be caused by bacteria, protozoa, or fungi. Most cases
result from direct spread from the paranasal sinuses, middle ea.r; or dental
root; in other cases, the organism travels hematogenously from another site.
In children, congenital heart defects may allow septic emboli to bypass the
pulmonary circulation; in adults, lung infections and endocarditis are the
most common sources of hematogenous inoculation. Damaged brain tissue
and immunosuppression contribute in some cases. Abscesses mature over
a 2-week period. Days 1 to 2 involve endothelial swelling and neutrophil
invasion; over days 3 to 4, necrosis and macrophages are prominent, and
lymphocytes and plasma cells join the infiltrate; over days 5 to 7, granu-
lation tissue forms around the necrotic center; and over days 8 to 14, the
capsule becomes strengthened by collagen and fibrosis, traversed by radially
oriented capillaries and surrounded by gliosis.
C. Granulomatous inflammation
1. Sarcoidosis affects the CNS in 5% of the cases, most often involving the
basal meninges, cranial nerves, optic tracts, and/or hypothalamus. Facial
nerve palsy is the most common presenting symptom, but other focal deficits
also occu.r; reflecting loss of function of involved structures. CNS-only sar-
coidosis is quite rare, but cases of systemic sarcoidosis that also affect
the CNS often receive neurosurgicaVneuropathologic attention because the
neurologic symptoms are so worrisome. Radiographically, the involved
structures show postcontrast enhancement. Grossly, the meninges and cra-
nial nerves may appear nodular and thickened. Microscopically, affected
structures show epithelioid granulomas with giant cells (e-Fig. 41.66).
Necrosis, although it may be seen focally, is not a usual feature of neurosar-
coidosis, and should encourage consideration of other diagnoses. Because
sarcoidosis is a diagnosis of exclusion, reasonable efforts to exclude other
etiologies for granulomatous inflammation (e.g., fungal, tuberculous, and
rheumatologic) must be made. Ultimately, a pathologic diagnosis no more

specific than granulomatous inflammation (accompanied by an appropriate
written comment) is usually warranted.
VII. SEIZURE DISORDERS. Surgically curable seizure disorders are broadly classified
into neoplastic and nonneoplastic lesions. Distinct neoplastic entities that generate
chronic seizures include DNT, GGs, and PXAs, and are discussed above. Common
nonneoplastic entities are discussed below.
A. Hippocampal (mesial temporal) sclerosis (HS) is a disorder characterized by
neuronal loss and gliosis of the hippocampus and, in some cases, adjacent
mesial temporal structures such as the amygdala and entorhinal cortex. Clini-
cally, HS is associated with longstanding complex partial seizures, most often
beginning near the turn of the first decade of life. Radiographically, the hip-
pocampus is small, and affected structures show T2 and FLAIR hyperintensity
and evidence of hypometabolism. Surgical treatment often involves amygdalo-
hippocampectomy. Histologically, the hippocampus shows variable neuronal
loss and gliosis, most pronounced in areas CAl (Sommer's sector) and CA4
(the endfolium of Ammon's horn) (e-Fig. 41.67). In more advanced cases, area
CA3 and the subiculum may be similarly affected. Neurons within the dentate
gyrus are often reduced in number and, in some cases, may appear dispersed or
split into two layers. Because this diagnosis requires accurate identification of
neuronal populations defined only by anatomic landmarks, the hippocampal
specimen must be carefully oriented when it is sectioned for histology; ide-
ally, it is resected in one piece that can be sectioned perpendicular to its long
axis to reveal the desired classic "seahorse" architecture. Not uncommonly, HS
resection specimens demonstrate a second pathology (a minute tumor or a mal-
formation of cortical development [MCD]) that could otherwise independently
account for seizures; such findings mirror the clinicopathologic experience that
seizure activity can both engender and arise from HS. After resection, the vast
majority of patients experience a significant reduction in seizure frequency or
are cured altogether.
B. Malformation of cortical development (MCDicortical dysplasia) is a general term
applied to a wide range of developmental abnormalities that affect the cortex.
These abnormalities come to attention most often as epileptogenic foci, and
are thought to account for up to 25% of cases of intractable epilepsy.
Grossly and radiographically, MCD may appear as (1) ectopic bands or
nodules of cortical tissue in the white matter (heterotopias); (2) a fum, focal
cortical expansion (cortical tuber); (3) an area of smooth, unfolded cortical
ribbon (agyriallissencephaly); (4) an area of cortical ribbon with few, broad
gyri (pachygyria); (5) an area of cortex with many small irregular gyri and
fused sulci (polymicrogyria); (6) an area with a blurred gray-white junction
(some focal cortical dysplasias, FCDs); or (7) ostensibly normal tissue (other
FCDs).
Histologically, MCD shows similar variability. Heterotopias contain gray
matter neuropil and abnormally arranged neuronal elements. Cortical tubers
are formed by aggregates of large "balloon cells" with abundant glassy
eosinophilic cytoplasm and an eccentrically placed neuron-like nucleus (e-Fig.
41.68). Tubers may occur sporadically in isolation or in greater numbers in
the setting of TS; the lesion itself is histologically identical in the two settings.
Agyria and pachygyria appear most commonly as a four-layered (rather than
six-layered) cortical ribbon. Polymicrogyria has been described as having two-
layered and/or four-layered areas within the cortex, but the most consistent
feature is a fusion of two adjacent molecular layers through what would oth-
erwise be a sulcus.
FCD, like MCD, represents a collection of entities; these have been con-
veniently organized within the classification of Palmini et al. (Neurology
2004;62[Suppl3]:S2). Particularly in subtle cases, immunohistochemistry for

NeuN (to accentuate neuronal cytoarchitecture), NF protein (to stain the
somata of enlarged, atypical neurons), GFAP and CD34 (to reveal poorly
defined, highly branched cells associated with MCDs) may facilitate diagnosis
(Acta Neuropathol. 1999;97:481).
C. Other seizure-associated disorders encountered less often are hemimegalen-
cephaly, Rasmussen encephalitis, hypothalamic hamartoma, and Sturge-
Weber angiomatosis.
VIII. VASCULAR DISORDERS
A. Vascular malformations are classified into four groups: arteriovenous malfor-
mation (AVM), cerebral cavernous malformation/cavernoma (CCM), capillary
telangiectasia (CT), and venous angioma (VA). CT and VA are asymptomatic
and virtually never encountered in surgical pathology, so will not be discussed
further.
1. Arteriovenous malformations (AVMs) are usually supratentorial and are
thought to be congenital. About 50% come to clinical attention in the
third, fourth, or fifth decades due to hemorrhage. Other AVMs induce
seizures ot; less often, slowly grow and cause a constellation of progres-
sively worsening headaches and focal neurologic deficits. Angiography
and MRI provide a diagnosis, reveal abnormal flow characteristics, and
identify the supplying and draining vessels of these lesions to facilitate inter-
vention strategy. Grossly, superficially located AVMs typically show dilated,
thick-walled draining veins on the cortical surface; cut sections show a dis-
organized mass of blood vessels of varying mural thickness and diameter,
with associated atrophic, relatively firm (gliotic) brain parenchyma. Histo-
logically, AVMs are characterized by an array of abnormal blood vessels that
show evidence of remodeling and degenerative changes (arteries with medial
hyperplasia, collagen deposition, and complex restructuring of the internal
elastic lamina; veins with thick collagenous walls). In some vessels, mixed
arterial and venous features may be identified. Amongst the abnormal blood
vessels, the intervening parenchyma shows atrophy, astrocytosis and, often,
hemosiderin deposits. Masson trichrome stain and an elastin stain Verhoeff-
Van Gieson (VVG) can be used to reveal features of the vascular pathology
that are subtle on routine sections. If embolization has been performed prior
to resection, intravascular foreign material may be identified; if sufficient
time has elapsed, a foreign body giant cell reaction may be visible.
2. CCM (also called cavernous angioma, cavernous hemangioma, and caver-
noma) is a vascular malformation characterized by large, thin-walled ectatic
vessels or sinusoids that are closely packed with minimal intervening brain
parenchyma (e-Fig. 41.69). CCMs are usually supratentorial, but may occur
anywhere in the CNS or leptomeninges. Autopsy studies have estimated
the prevalence of sporadic CCMs to be approximately 1 in 200 individ-
uals. As many as 50% of the patients with these lesions develop seizures
and/or recurrent headaches. Additionally, these lesions exhibit an annual
1% chance of hemorrhage. Because such hemorrhages are not at high pres-
sure, they are usually not fatal; however, they may cause focal neurologic
deficits and do increase the risk of seizures and subsequent hemorrhage
severalfold. Brainstem lesions are associated with worse prognosis. Radio-
graphically, T2-weighted images show a heterogeneous center with a sur-
rounding rim of hypodensity that corresponds to deposits of hemosiderin;
gradient-echo MRI shows greater sensitivity for this ferruginous penumbra
and therefore can detect smaller lesions. Grossly, CCMs appear spongy and
dark red with golden brown hues due to hemosiderin deposition and, occa-
sionally, calcifications. Microscopically, the vascular component shows only
a single layer of endothelium and lacks a muscular layer and an internal
elastic lamina. The peripheral rim of resected tissue contains hemosiderin-

laden macrophages and gliotic parenchyma with axonal spheroids. Areas
of fibrosis, xanthomatous degeneration, and calcification are common.
Although most cases are sporadic, some are familial with an autosomal
dominant inheritance, and affected individuals often have multiple CCMs.
B. Primary angiitis of the CNS (PACNS). Inflammation of the CNS vasculature may
occur as part of a systemic vasculitis (e.g., Giant cell arteritis, polyarteritis
nodosa) or may be CNS specific (PACNS). PACNS more commonly afflicts
males in late middle age. Symptoms are highly variable, but are generally multi-
focal, intermittent, progressive and, by definition, strictly neurologic. On MRI,
PACNS may show nothing more than multiple ischemic microinfarcts; the
angiographic finding of alternating luminal stenosis and dilatation (beads on a
string), while characteristic, is neither sensitive nor specific for the disorder. CSF
cytology may show mild lymphocytic pleocytosis, another nonspecific finding.
To establish the diagnosis, meningeal/brain biopsy is performed. Histologi-
cally, the key feature of PACNS is segmental granulomatous inflammation of
small- and medium-sized arteries featuring transmural lymphocytic infiltrates
and expansion of the intima by histiocytes and multinucleated giant cells. Fib-
rinoid necrosis is characteristic but not required for diagnosis. The associated
parenchyma often shows changes associated with ischemic injury. As is the case
for sarcoidosis, other potential causes of granulomatous vasculitis (including
acid-fast bacilli, fungus, and intravascular amyloid) must be excluded, and the
most appropriate pathologic diagnosis is often simply "granulomatous vas-
culitis." Although prognosis is poor without treatment, immunosuppression
(prednisone, with or without cyclophosphamide) has been effective in some
cases.
C. Congophilic (or cerebral) amyloid angiopathy (CAA) is characterized by deposits
of amyloid in leptomeningeal and superficial cortical vessels. By far, the most
common substrate of this disease is amyloid-beta 1 to 40; ""80% of the neu-
ropathologically confirmed AD cases show CAA at least focally. Not surpris-
ingly, therefore, CAA commonly affects the elderly. These deposits can disrupt
physiological autoregulation of blood flow and may contribute to cognitive
dysfunction. Most often CAA comes to clinical attention only when it becomes
severe enough to compromise vessel integrity. The superficial lobar hemorrhage
that can result from CAA can cause mass effect and may resemble intratu-
moral hemorrhage; consequently, surgical evacuation is often performed for
decompression, occasionally with some degree of parenchymal resection to
determine the cause. Parenthetically, this circumstance illustrates why CNS
hematoma evacuation specimens must be examined very thoroughly for even
the smallest fragments of brain tissue. Microscopically, involved vessels show
amyloid deposits that can be appreciated on routine and special stains (Congo
red and thioflavin S) (e-Fig. 41.70). Immunohistochemistry can be used to iden-
tify the amyloidogenic peptide involved; cases that do not show reactivity for
amyloid-beta may be tested for rarer forms that are caused by cystatin C (Ice-
landic type), integral membrane protein 2B (ITM2B, British type or Danish
type), gelsolin (Finnish type), or transthyretin (meningo-vascular amyloido-
sis). Although inflammation is usually absent, specimens must be evaluated
for amyloid-beta related angiitis (ABRA), a form of granulomatous vasculitis
(e-Fig. 41.71) that may respond to immunosuppressive therapy.
D. Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoen-
cephalopathy (CADASIL) (e-Fig. 41.72) is caused by missense mutations of the
Notch3 gene. The abnormal protein accumulates in granules (PAS-positive,
osmiophilic) within the walls of small arteries, the smooth muscle layer degen-
erates, and the vessel walls become thickened and fibrotic; this process grad-
ually diminishes capacity for and regulation of perfusion. CADASIL usually
presents in the fihh or sixth decade of life with classic migraine, multiple sub-
cortical ischemic strokes, psychiatric disturbance, and later, dementia. In this
context, T2 hyperintensities in anterior temporal lobes and external capsules on
MRI are strongly suggestive. Although the symptoms of CADASIL arise from
the brain, the disease also affects peripheral tissues including the skin. His-
tologic and ultrastructural examination of arterioles in the skin or brain pro-
vides the diagnosis. Notably, a rare, otherwise identical clinical/radiologic syn-
drome with an autosomal recessive inheritance pattern (appropriately named
CARASIL, linked to the HTRA1 gene) lacks granular osmiophilic inclusions.
E. Cerebral infarcts. Most often, biopsies of infarcts are performed when "glioma"
is in the differential diagnosis, particularly in young patients. Gross and micro-
scopic features of a cerebral infarct change over time. In the acute stage
(1 to 3 days), red necrotic neurons, vacuolated neuropil, and variable neu-
trophilic infiltrates are present. Subacute infarcts (days to weeks) demon-
strate capillary proliferation with prominent endothelial cells, numerous foamy
macrophages, and reactive astrocytes that are typically more common at the
periphery. Because such reparative lesions may exhibit mitotic figures, necro-
sis, and microvascular proliferation consistent with GBM, care must be taken
to recognize the reactive and histiocytic components, particularly during an
intraoperative microscopic evaluation. Large infarcts eventually evolve into
cystic lesions with a weblike network of delicate blood vessels, sparse gliotic
parenchymal remnants, and a few residual macrophages. Infarcts disrupt and
destroy axons, a feature often helpful in distinguishing infarcts from demyeli-
native processes such as MS in which the axons are relatively spared. Once an
infarct is diagnosed, identifying its cause is the next step; if the specimen at
hand provides no insight, clinical correlation is required.
IX. NEURODEGENERATIVE DISORDERS. Although some of these affect children and ado-
lescents, most come to clinical attention after the fifth or sixth decades. An increas-
ing number of neurodegenerative diseases can be diagnosed by identifying changes
in DNA, biologic fluids, peripheral biopsy material, or radiographic images. Nev-
ertheless, few treatments are currently available in clinic for neurodegenerative
disorders, and most of them remain idiopathic, progressive, and fatal. Conse-
quently, CNS biopsy performed explicitly to diagnose a neurodegenerative disease
is rare. These disorders usually appear in a surgical pathology specimen inciden-
tally (e.g., when some brain is removed to evacuate a life-threatening hematoma)
or when unusual clinical/radiologic data suggest that securing a definitive tissue
diagnosis warrants the risk of brain biopsy. Recognizing these disorders in surgical
specimens may become more critical as treatments become available.
A. Alzheimer disease (AD}, the most common neurodegenerative disorder, is char-
acterized histologically by neuron loss, gliosis, and the presence of extracellular
beta-amyloid peptide deposits (diffuse, cored, and neuritic plaques) and NFTs
(NFT, intraneuronal aggregates of hyperphosphorylated tau protein) in the
neocortex and hippocampus (e-Fig. 41.73). Plaque deposition begins 10 to
15 years before the onset of very mild dementia of the Alzheimer type (DAT,
the appropriate term for the clinical manifestations of AD). The degree of
cognitive impairment appears to correlate with the severity and extent of NFT
pathology, which begins in the mesial temporal lobes and subsequently spreads
outward to involve the parietal and frontal lobes; the occipital lobe is relatively
spared. Neuroimaging techniques (to assess atrophy, metabolism, and amyloid
burden within the brain) and CSF biomarker measurements (e.g., of amyloid-
beta peptide and tau protein) are likely to allow reasonable antemortem con-
firmation of AD pathology in the near future (Nature 2009;461:916-922), but
recognition of the histologic changes will remain important.
Currently, there are three leading sets of criteria for the neuropatho-
logic diagnosis of AD: Khachaturian, Consortium to Establish a Registry for
Alzheimer's Disease (CERAD), and National Institute on Aging (NIA/Reagan).

Technically, the Khachaturian system, which evaluates only the presence of
amyloid plaques within any portion of neocortex, is the only one that can be
applied faithfully to a limited brain biopsy; officially, CERAD requires sam-
pling of numerous cortical areas (possible only at autopsy) for a semiquanti-
tative assessment of neuritic plaques, and NIA/Reagan additionally requires
assessment of the anatomical distribution of NFT pathology within the brain.
Nevertheless, since NFT pathology correlates most reliably with the degree
of cognitive impairment and the frontal lobe is generally affected later in
the disease process, the presence of plaques and NFTs in a biopsy specimen
from the frontal lobe should suggest that AD pathology may be responsible
for/contributing to a patient's cognitive symptoms. The deposits of AD are best
visualized by modified Bielschowsky silver staining, thioflavin S or T staining
(using fluorescence microscopy), or by immunohistochemistry for amyloid-
beta peptide and for phosphorylated tau protein. As discussed above, CAA
often accompanies AD pathology.
B. Lewy body disease (LBD). This generic term is intended to unify several clinico-
pathologic diagnoses that feature this intraneuronal inclusion and the related
Lewy neurite (e-Fig. 41.8). Lewy body pathology, either alone or in conjunc-
tion with AD, is associated with about 10% to 15% of dementia cases. LB
pathology has been proposed (in the setting of clinical PD) to begin within a
few selectively vulnerable brainstem nuclei and to spread rostrally, reaching the
prefrontal neocortex in later stages (Neurobiol Aging 2003;24:197). In paral-
lel, many patients with PD develop dementia about 10 years after motor symp-
toms. Therefore, identification of these lesions in a frontal lobe biopsy specimen
at sufficient density provides reasonable evidence that a patient may be suffer-
ing from dementia related to LBD (Neurology 2005;65:1863). Because these
lesions are very subtle on H&E sections of the neocortex, and because they
are not argyrophilic on Bielschowsky-stained sections, immunohistochemistry
for (ideally, phosphorylated) alpha-synuclein is required for definitive iden-
tification; this stain should therefore be applied routinely, particularly when
symptomatology is suggestive of PD or dementia with LBs, or when other
relevant pathologies are not detected.
C. Creutzfeldt-Jakob Disease (CJD) (e-Fig. 41.74). The need to diagnose or rule out
C]D as a cause of rapidly progressive dementia (developing in ""2 years or less)
is one of the leading clinical scenarios prompting brain biopsy. C]D is a trans-
missible spongiform encephalopathy (TSE; other types include Gerstmann-
Strauss-Scheinker [GSS] syndrome, fatal insomnia, and kuru) that is classically
characterized by a clinical triad of myoclonus, periodic shortwave EEG activ-
ity, and rapidly progressive dementia. Diffusion weighted (DWI) and FLAIR
MRI commonly show restricted diffusion within the striatum and cerebral cor-
tex. Although the condition is transmissible, most cases are sporadic, affecting
one person per million individuals per year with a peak age of onset at age 60;
10% of the cases are familial and are attributable to mutations in the prion pro-
tein. Only rare examples are iatrogenic, resulting from inoculation with tissues
or instruments inadvertently contaminated by protease-resistant prion pro-
tein from another individual with CJD. Microscopically, CJD cases show neu-
ronal loss, gliosis, and spongiform change consisting of small, sharply defined,
punched out vacuoles. These histologic changes appear in an anatomic pattern
suggested by radiographic changes, but evaluation is typically limited to a right
frontal cortex biopsy specimen. Within the cortex, the spongiform change of
C]D should span the entire thickness of the cortical ribbon; other dementing
conditions may show vacuolation that appears similar and is limited to super-
ficial layers. It is worth noting, however, that the histologic changes in CJD
may be anatomically patchy, and therefore very subtle or invisible in a limited
biopsy specimen. It is also worth noting that biopsies of individuals with CJD
may also show preexisting histologic changes associated with other dementing
illnesses, therefore, identification of abundant amyloid plaques, NFTs, or LBs
does not rule out the possibility of CjD. For this reason, immunoblotting and
immunohistochemical analysis for the abnormal protease-resistant protein are
essential for definitive diagnosis.
D. New variant CJD (nvCJD) is a rare form of TSE linked to bovine spongiform
encephalopathy (BSE). Unlike CJD, nvCJD affects a younger demographic
(adults younger than 40 years), typically has a longer clinical course charac-
terized by behavioral changes more often than dementia, and shows an addi-
tional thalamic abnormality on DWI, T2, and FLAIR MRI. Microscopically,
the most characteristic feature is the presence of numerous amyloid plaques
(the so-called florid plaques that are PRP-positive, amyloid-beta negative) in
the cerebral and cerebellar cortices. The few hundred cases reported since 1996
have been clustered in the UK and France and are thought to stem from an
outbreak of BSE that affected the UK cattle industry in 1986. Perhaps reflecting
the incubation period and changes in cattle industry practices, the incidence of
nvCJD in the UK has dwindled over the last decade, so the diagnosis is now
quite rare.
Cytopathology of the Central

Nervous System
Souzan Sanati and Lourdes R. Ylagan
I. SPECIMEN TYPES. The most common specimen type obtained for cytologic evalua-
tion of pathologic conditions of the central nervous system (CNS) is cerebrospinal
fluid. Rarely, stereotactic fine needle aspiration (FNA) of cysts or masses of the CNS
is used to provide specimens for cytologic diagnosis.
A. Cerebrospinal fluid (CSF}. Several methods have been developed in the last decade
for concentration of samples obtained via lumbar puncture, including cytocen-
trifugation and membrane filtration. Slides prepared via the cytocentrifugation
technique using a cytofunnel (http://www.shandon.com) are usually Diff-Quik
stained since leukemic involvement of the CSF is the most common malignant
diagnosis; the limited amount of material obtained (0.5 to 1.0 ml) usually pre-
dudes allocating material for Papanicolaou and other stains. Although the precise
proportions vary by practice setting, most CSF specimens obtained for headaches
and mental status changes show reactive (e-Fig. 41.75) or nonspecific features,
and usually contain benign lymphocytes andmonocytes (e-Fig. 41.76). Other nor-
mal cells which can be found in CSF specimens include ependymal and choroidal
cells (e-Fig. 41.77).
1. Traumatic tap. CSF samples that consist predominantly of peripheral blood are
diagnosed as "negative for malignant cells" with the caveat that the sample
may represent a traumatic tap (which occurs in about one-quarter of the cases).
When a CSF sample shows leukemic involvement in the presence of peripheral
blood, the possibility of CSF sample contamination by peripheral blood blasts
should be raised and a repeat sample should be obtained.
2. Infectious processes. Peripheral blood contamination can result in the presence
of variable numbers of polymorphonuclear neutrophils in a CSF sample; how-
ever, abundant neutrophils in the absence of blood contamination should raise
the possibility of acute meningitis. Prompt diagnosis is crucial, since it can be
fatal if not treated promptly (e-Fig. 41.78). Aseptic meningitis gives a picture
of an increased number of mature appearing lymphocytes and monocytes

(e-Fig. 41.79). Suspicion of an infectious process should prompt additional
work up including microbiologic cultures and molecular methods to identify
the source of infection. With the advent of highly active antiretroviral ther-
apy (HAART), AIDS related CSF lesions have become rare in recent years
(J Neurovirol. 2005;11(Suppl3):72).
3. Lymphoma and leukemia. A primary diagnosis of lymphoma or leukemia should
not be made in the absence of flow cytometric analysis of the cells to demon-
strate a clonal process. A diagnosis of involvement by lymphoma or leukemia
may be rendered if malignant appearing lymphoid cells or cells consistent with
blasts are present (e-Figs. 41.80 to 41.85) in a patient with a previously proven
history of lymphoma or leukemia.
4. Metastasis. Cytologic examination of CSF can be used to document CNS
involvement in patients who have a known history of metastatic carcinoma
(e-Fig. 41.86) or melanoma (e-Fig. 41.87), as well as leukemia or lymphoma
(e-Fig. 41.88) as noted above. In cases of metastasis, the diagnosis should be
confirmed by immunocytochemical stains or by comparison of the cytologic
material with the patient's primary malignancy. The diagnostic approach to
the rare metastatic tumors of unknown origin that present in the CSF is the
same as for tumors of unknown origin presenting at other sites (Semin Oncol.
1993;20:206).
B. Stereotactic brain FNA is an uncommon procedure, and the choice of the prepara-
tory method is dependent upon the type of tissue aspirated. Involvement by a
known primary lesion should be confirmed by comparison of the cytologic spec-
imen with the prior diagnostic material. Fluid or purulent fine needle aspirates
should be cultured as well as submitted for cytologic evaluation.
C. Diagnostic categories
1. Negative for malignancy. This diagnosis is rendered for CSF specimens in which
only benign cellular elements, and primarily mature appearing lymphocytes
and/or monocytes are present.
2. Atypical cytology. This diagnosis is rendered when there are rare atypical cells
but for which there is inadequate material for ancillary studies. This diagnosis
should prompt the collection of additional material for either flow cytometric
or immunocytochemical evaluation.
3. Suspicious for malignancy. This diagnosis is rendered when there are rare cells
highly concerning for involvement by a malignant process, but when the find-
ings are nonetheless insufficient for definitive diagnosis.
4. Positive for malignancy. This diagnosis is rendered when there is both quan-
titative and qualitative evidence of a malignant neoplasm. Ancillary tests can
be used to provide a definitive diagnosis as to the type of neoplasm.
D. Special techniques. Special techniques, such as confirmatory FISH, can be per-
formed on CSF samples (e-Fig. 41.89).
Nerve Biopsies
Robert E. Schmidt
Successful use of nerve biopsy requires a discussion between the clinician and neu-
ropathologist since the clinical differential diagnosis may dictate an unusual sampling
scheme, for example, vasculitis in which multiple cross-sections may be required for
diagnosis.
I. NORMAL PERIPHERAL NERVE AND METHODS OF ANALYSIS. The sural nerve, a cuta-
neous sensory nerve to the lateral foot, is typically biopsied without stretching
or use of intraneural anesthetic. The nerve is divided into a portion for fixation
in formalin for paraffin embedding, and a portion for fixation in glutaraldehyde
for plastic sections and possible electron microscopy. The entire nerve is typically
used rather than dissected into individual fascicles. The portion {"'1 em) of the
nerve fixed in formalin should be cut into 3 mm segments and examined in cross-
section with paraffin-embedded H&E sections, thioflavin-S histochemistry, and
possible immunohistochemistry.
An H&E-stained cross section of the sural nerve {e-Fig. 42.1)* contains 6 to 12
fascicles, each surrounded by flattened perineurial cells. Outside the perineurium
is the epineurium, containing connective tissue and an anastomotic vascular net-
work. Often individual myelinated axons can be seen in H&E-stained material
{e-Fig. 42.2). The endoneurium {the space inside of the perineurial cell layer and
outside the axon/Schwann cell units) contains fluid, collagen, capillaries, venules,
fibroblasts, macrophages/monocytes, and scattered mast cells. Immunohistologic
localization is useful for the demonstration of neurofilaments {a rough measure of
axon number, e-Fig. 42.3 ), amyloid, immunoglobulins, subtypes of inflammatory
cells, and growth factors and their receptors.
Plastic-embedded sections of 1 J.Lm thickness {e-Fig. 42.4) provide a wealth
of information. Myelinated axons range in diameter from 2 to 18 J.Lm and are
coarsely separated into small {mean of 4 J.Lm) and large {"'12 J.Lm) myelinated
axon populations {e-Figs. 42.5 and 42.6) with myelin thickness related directly
to axonal diamete.t The patterns of nerve damage visible in plastic sections may
be characteristic {although rarely pathognomonic) of certain disease entities or
pathogenetic processes. Qualitative information is provided on the degree of
myelinated axon loss, distribution of axon loss, presence of active axonal degen-
eration {AD) or demyelination, identification of regenerative clusters of axons,
swollen axons, onion-bulb formation, the nature of cellular infiltrates, or amy-
loid deposition. Groups of unmyelinated axon populations are detectable in plas-
tic sections {e-Fig. 42.6).
Ultrastructure provides additional detail and is the only definitive method
to evaluate unmyelinated axons {e-Fig. 42.7), which are three- to fourfold more
numerous than myelinated axons in the sural nerve and typically are <2 J.Lm in
diameter.
If necessary, lengths of individual lightly fixed, osmicated myelinated axons can
be dissected or "teased, out of a fascicle with pins. Each myelin internode, main-
tained by a single Schwann cell, ranges from 0.2 to 1.8 mm in length, increasing
673
Figure 42.1 Selective loss of small myelinakd axons. Normal fascicle is on the left.
linearly with axon diameter. Ongoing activity or residua of past episodes of

demyelination or AD are readily identified.
Morphometry provides quantitative data concerning axon number and axon
size-frequency distribution. Large axons are lost in uremia, abetalipoproteinemia,
thallium, arsenic, acute intermittent porphyria, cisplatin, vincristine, and Friedre-
ich ataxia. Small axons are selectively damaged (Fig. 42.1) in amyloidosis, some
forms of diabetic neuropathy, acute pandysautonomia, Fabry disease, and hered-
itary sensory and autonomic neuropathies (HSANs).
II. THREE BASIC PATHOLOGIC MECHANISMS CHARACTERIZE NEUROPATHIES
A. Axonal degeneration is the most common pattern in biopsies, resulting in degen-
eration of the axon and its myelin sheath (e-Figs. 42.8 and 42.9). Often
the distal portions of the longest axons are preferentially involved (i.e., dis-
tal axonopathy or dying-back neuropathy). Myelin destruction and its early
catabolism occurs in the Schwann cell, producing the myelin "ovoid" of teased
fibers and is subsequently continued in hematogenously and endogenously
derived macrophages which engulf the debris. Early regenerative events begin
immediately with the proliferation of Schwann cells and their processes which
accumulate within the original basal lamina of the axon/Schwann cell unit
as "bands of Biingner" which are conduits for regenerating axonal sprouts.
Schwann cells increase their synthesis of growth factors as trophic and tropic
stimuli. Maturing axons form regenerative dusters (e-Fig. 42.10) of thinly
myelinated axons within the original Schwann cell,s basal lamina and, with
time, one axon emerges and begins to function and the others regress. Such a
regenerated axon characteristically has a myelin sheath relatively thin for its
axon,s caliber. Teased fiber preparations show a distinctive and uniform intem-
odallength regardless of axon diameter. The end-stage of a chronic neuropathy
may consist of rare preserved axons, scattered fibroblasts, and Schwann cells
and may provide little information concerning the process which preceded it.
AD shares mechanisms with Wallerian degeneration (WD) which is the
reaction of axons and myelin distal to a crush injury. In WD, many axons
show simultaneous involvement and are often at the same histologic stage; in
AD, it is typical to find degenerating, regenerating, and normal axons together,
some of which may have a distinctive pathologic signature (Fig. 42.2).
B. Seamental demyelination represents preferential damage to one or several
internodes of the myelin sheath, directly to myelin or to its Schwann cell, with
relative axonal sparing as seen in this teased fiber preparation (e-Fig. 42.11).
Schwann cell proliferation results in the replacement of each lost myelin intern-
ode by several shorter internodes resulting in variation in internodal length
along individual teased fibers.
Chapter 42 • Nerve Biopsies I 675
Axonal Wallerian
Degeneration Degeneration
Figure 42.2 AD vs. WD. AD is typically characteri2ed by simultaneous
AD, regenerative clustm~ (arrow), and axons with pathologic signatures
(crosshatched) compared with synchronous and uniform degeneration of
most axons in a fascicle in WD.
C. Secondary demyelination preferentially, nonrandomly involves selected axons,

which may be atrophic or damaged, while entirely sparing others. It may reflect
an abnormal axon-Schwann cell interaction resulting in secondary myelin loss.
Uremic neuropathy is the prototype.
Ill. TOXIN-INDUCED NEUROPATHIES. A huge number of toxins and drugs (Table 42.1),
including many whose neuropathic toxicity limits their clinical usefulness, pro-
duce neuropathy, likely with different mechanisms. One group of ADs targets
axonal transport, and others selectively affect neurofilaments or microtubules.
Acrylamide can result in distinctive neurofilament and tubulovesicular aggregates
involving distal portions of axons. Zinc pyridinethionine and bromophenylacety-
lurea preferentially target reversal of the polarity of axonal transport result-
ing in terminal swellings. Lead, diphtheria toxin, perhexiline, lysolecithin, and
hexachlorophene are prominent toxins preferentially directed at the Schwann
celVmyelin sheath, resulting in demyelination. Many therapeutic agents may
induce peripheral nerve disease. Heavy metals (e.g., arsenic, mercury, thallium,
gold) are well known for their toxic effects on peripheral nerves. Neuropathic
industrial and environmental agents have resulted in epidemics of toxic neuropa-
thy.
IV. ISCHEMIC NEUROPATHIES. The nerve vascular supply (i.e., vasa nervorum} is rich
and anastomotic, requiring a substantial decrease in nerve blood flow to interrupt
function. Vasculitis {i.e., vasculopathy with angionecrosis) is frequently patchy,
resulting in asymmetric nerve involvement (mononeuritis multiplex), emphasiz-
ing the need for thorough sampling of the nerve biopsy. Polyarteritis nodosa, the
vasculitic prototype, is characterized by epineurial arteries damaged by polymor--
phonuclear leukocytes, macrophages, monocytes, fibrin (e-Figs. 42.12 and 42.13 ),
and a range of axonopathy rarely culminating in infarction. There may be inter-
or intrafascicular variability in axon loss (Fig. 42.3, and e-Figs. 42.14 and 42.15).
Fibrotic recanalized vessels mark previous sites of vasculitic damage.
Patients with collagen vascular diseases may clinically present with mononeu-
ritis multiplex, histologically comparable to that of polyarteritis nodosa or a min-
imal epineurial perivascular mononuclear cell infiltrate (i.e., microvasculitis) lack-
ing angionecrosis. Epineurial perivascular collections of a few mononuclear cells
are common, and some authors consider them, in isolation, to be of little patho-
logic importance in the absence of loss of vascular continuity or vasculopathy.
Alternatively, they may represent an early or mild vascular injury, or represent
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Chapter 42 • Nerve Biopsies I 87 7
Figure 42.3 Ischemic pattern of axon loss. Intra- and inrerfascicular variability of axon loss
is typical but not pathognomonic of a vasculiti.c/ischemi.c pathogenesis.
resistance to degradation because of the formation of advanced glycation

end products. Some patients with painful dysesthesias and a normal elec-
trophysiologic exam show loss of intraepidermal ax:ons in skin biopsies, a
procedure becoming routine practice in some institutions. Multiple ischemic
insults may summate to produce a symmetrical and uniform axon loss
distally, but this mechanism has not been universally accepted. Diabetes-
induced oxidative stress may reflect deranged metabolism and/or mitochon-
driopathy.
2. Asymmetric neuropathies include radiculoplexus neuropathy (diabetic amy-
otrophy), truncal radiculopathy, upper limb mononeuropathy, and cranial
nerve (chiefly III) palsies characterized by an inflammatory microvasculitis
and/or perineuritis, possible immune complex and complement deposition,
and a mild to marked axonopathy.
3. Diabetic autonomic neuropathy produces a large range of symptoms con-
tributed by the sympathetic and parasympathetic nervous systems (also
enteric and visceral sensory). Autonomic axons may be involved in sym-
metrical sensorimotor polyneuropathy or as more restricted involvement
of autonomic fibers in the gut, bladder, penis, and other organs. Studies of
human diabetics and rodents show markedly swollen dystrophic axons and
synapses in prevertebral sympathetic ganglia.
B. Uremia results in demyelination of relatively atrophic ax:ons (i.e., secondary
demyelination).
C. Others. Deficiencies of pyridoxine, thiamine, vitamin E (found in abetalipopro-
teinemia, cystic fibrosis, and biliary atresia), niacin, cobalamin, and multiple
nutritional deficiencies (apparently resulting in an epidemic in Cuba) are asso-
ciated with several forms of neuropathy. Hypothyroidism, acute intermittent
porphyria, galactosemia, hepatic failure, acromegaly, chronic respiratory insuf-
ficiency, and critical illness may also be associated with neuropathy.
Figure 42.4 AIDP. Patchy loss of myelin in large and small axons is accompanied by an inflam-
matory infiltrare (solid dots). Normal fascicle is on the left.
VI. NEUROPATHIES WITH AN IMMUNE-MEDIATED MECHANISM

A. Guillain-Barri syndrome (GIS)
1. Acute inflammatory demyelinating polyneuropathy (AIDP) results in patchy
myelin loss with residual debris and the appearance of naked demyelinated
axons (e-Figs. 42.17 and 42.18). A perivascular epineurial and endoneuria!
T-cell rich infiltrate is coupled with macrophage infiltration (e-Fig. 42.19)
and results in distinctive macrophage-mediated stripping of layers of oth-
erwise normal appearing myelin (Fig. 42.4, e-Figs. 42.20 to 42.22). Circu-
lating antibodies against endoneuria! targets may secondarily gain access
through a damaged blood nerve barrier or be locally synthesized. Plasma-
pheresis and intravenous immunoglobulins may be useful treatment modal-
ities. Eventually, Schwann cells proliferate and remyelinate the denuded
internode (e-Fig. 42.23). Axonopathy and axon loss is seen, probably
because of a noxious endoneuria! environment containing cytokines and
oxidative mediators.
2. Acute motor axonal neuropathy (AMAN) and acute motor and sensory axonal
neuropathy (AMSAN). In these axonal forms of GBS, the axon rather than
myelin is the primary target and may produce AMAN or AMSAN dys-
function. Macrophages may be found adjacent to nodes of Ranvier or
entering the periaxonal space, possibly targeting axons with a variable
degree of AD. Antibody may target a constituent of the node of Ranvier
to which it binds, fixes complement forming a membrane attack complex,
recruits macrophages, and results eventually in AD. Axonal GBS may be
associated with a more aggressive course with poorer outcome, electro-
physiologic and pathologic evidence of prominent axonopathy, relatively
increased incidence of enteric Campylobacter jejuni infection, and a role
for antibody directed against GMt ganglioside. Different patterns of nerve
injury seen in GBS may reflect the strain of infecting organism or host-HLA
alleles.
3. Millar Fisher variant of GBS is distinguished from AIDP/AMAN by a distinc-
tive presentation of ataxia, areflexia, and ophthalmoplegia, and antibodies
against the ganglioside GQtb which is concentrated at the nodes of Ran viet;
particularly those of the oculomotor nerves.
B. Chronic inflammatory damyalinalinl polyradiculonauropathy {CIDP) is character-
ized by a symmetric progressive or relapsing/remitting, sensory and motor
polyradiculoneuropathy, particularly involving proximal nerves. Perineurial
inflammation and onion-bulbs reflect its chronicity, but do not occur in all
Chapter 42 • Nerve Biopsies I 679
cases. CIDP may be a chronic form of GBS in which there is failure of regulatory
T-cells to suppress the typically monophasic attack of GBS. Its autoimmune
pathogenesis involves CD4+ and CD8+ T-cells, and possibly also antimyelin
antibodies. HLA subtype frequencies (e.g., HLA-DR2) differ between GBS and
CIDP.
C. Antimyelin associated glycoprotein (MAG) neuropathy is a slowly progressive
demyelinating neuropathy with an associated lgM monoclonal gammopathy
(monoclonal gammopathy of unknown significance, MGUS) in which the anti-
body is directed against MAG, a molecule which supports myelin integrity.
Anti-MAG antibodies deposited on portions of the myelin sheaths of periph-
eral nerve axons result in altered myelin periodicity, demyelination, and AD.
Immunosuppression and plasmapheresis have resulted in transient improve-
ment. Rituximab, a mouse-human chimeric antibody against the B cell surface
marker CD20, is reported to have clinical benefit.
D. Antiglycolipid, antisulfatide, and antiganglioside neuropathies have been associ-
ated with a wide range of clinically identifiable acute and chronic neuropathy
syndromes.
E. Anti-Hu antibody neuropathy is associated with paraneoplastic sensory neuropa-
thy and likely reflects development of antibodies against shared antigens of
small cell lung carcinoma and dorsal root ganglion neurons. Infiltration of
CD8+ T cells is also present.
F. Autoimmune autonomic neuropathy resulting in autonomic failure following a
viral illness may be mediated by circulating antibodies against ganglionic nico-
tine acetylcholine receptor.
VII. GENETIC NEUROPATHIES
A. Hypertrophic (onion-bulb) neuropathies
The concentric proliferation of Schwarm cells in response to multiple
episodes of demyelination and subsequent remyelination results in onion-
bulbs, the defining pathologic hallmark of a group of neuropathies. Onion-
bulbs are visible in H&E-stained nerves (e-Fig. 42.24), a pattern accen-
tuated with immunolabeling for Collagen type IV (e-Fig. 42.25). Onion
bulbs are best illustrated in plastic embedded nerve (e-Fig. 42.26) and
with electron microscopy (e-Fig. 42.27). Nerves may be palpably enlarged
and conduction velocities markedly decreased. Genetic analysis has iden-
tified mutations in myelin constituents (Po, PMP22, myelin basic pro-
tein), including those localized to noncompact myelin near the paranode
(MAG, cormexin 32, neurofascin 155) and axonal proteins (Caspr, contactin,
kinesin). A more complete catalogue can be found at several websites (e.g.,
http://www.molgen. ua.ac.beiCMTMutationsl).
1. Hypertrophic Charcot-Marie-Tooth disease (CMT 1) is inherited as an auto-
somal dominant condition resulting from a duplication or point muta-
tion in the PMP-22 gene or a point mutation in Po myelin protein (both
needed for myelin compaction), a defect in the early growth response 2
gene (EGR2), or mutations of periaxin (a Schwarm cell protein regulating
its shape and axonal communication). Animal models have established a
role for macrophage and lymphocytic infiltration in disease pathogenesis.
X-linked CMT demonstrates a defect in the gene for cormexin-32.
2. Other types (Table 42.2). Dejerine-Sottas disease (DSS, includes HMSN-
ill+) begins in early life, some cases of which have a demonstrable genetic
defectinPMP-22. periaxin, GDAP1. EGR2. or myelin protein Po. Refsum's
disease, caused by phytanic acid oxidase deficiency, also results in an onion-
bulb neuropathy.
B. Hereditary neuropathy with pressure palsies (HNPP). Teased fiber preparations
of this dominantly inherited neuropathy demonstrate marked focal hyper-
myelination (called tomaculi or sausages) characterized by redundant myelin
110 I SECTION IXt NERVOUS SVtHE ...
O..IIN"M
CMTIA
CMTIB
CMTIC
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7. """"""lo nou"''"1f1Y Ad111nood HIV
8. ~ lftlllr-lfmphoqlosls Modorlll~ llMnc<ld
682 SECTION IX: NERVOUS SYSTEM
D. Lyme disease. Cranial and peripheral nerves show an epineurial or perineurial

perivascular lymphocytidplasmacytic infiltrate (perivasculitis) and AD. Organ-
isms are not typically seen in involved nerves.
X. TRAUMATIC NEUROPATHY exists in many forms. Chronic nerve compression and
entrapment is characterized by focal loss of myelin. More severe trauma with loss
of nerve continuity may produce a traumatic neuroma, that is, a combination of
degenerative and regenerative responses resulting in a disorganized aggregate of
collagen and axonal "minifascicles" (e-Figs. 42.31 and 42.32). The distinctive
appearance of minifascicles can be seen with immunohistochemistry for neurofi1-
aments (which stains axons, e-Fig. 42.33) or epithelial membrane antigen (EMA,
which stains the perineurium, e-Fig. 42.34), and in plastic sections (e-Fig. 42.35).
SUGGESTED READINGS
Kennedy WR. Opportunities afforded by the study of unmyelinated nerves in skin and other organs.
Muscle Nerve. 2004;29:756-767.
Midroni G, Bilbao JM. Biopsy Diagnosis of Peripheral Neuropathy. Boston, MA: Butterworth-
Heinemann; 1995.
Pestronk A. Neuromuscular Division Website, Washington University School of Medicine,
http://www.neuro.wustl.edu/neuromuscular/
Spencer PS, Schaumburg HH, Ludolph AC. Experimental and Clinical Neurotoxicology. New
York, NY: Oxford University Press; 2000.
Zochodne DW. Diabetes mellitus and the peripheral nervous system: Manifestations and mecha-
nisms. Muscle Nerve. 2007;36:144-166.
SECTION X
Hematopoietic System
Lymph Nodes
Anjum Hassan and Friederike Kreisel
I. NORMAL ANATOMY. Lymph nodes are the most widely distributed collections of
lymphoid tissue within the lymphoreticular system, which also includes the thy-
mus, tonsils, adenoids, spleen, and Peyer patches. Due to their easy accessibility,
lymph nodes are the most frequently examined lymphoid tissue for a lymphoretic-
ular disorder. Microscopically, the lymph node shows four compartments: The
most obvious are the primary and secondary follicles, which are usually found
near the capsule, surrounding the follicles and extending deeper into the node
is the paracortex. The third and fourth compartments represent the medullary
region and the sinuses, respectively (e-Fig. 43.1).*
II. GROSS EXAMINATION, TISSUE SAMPLING, AND HISTOLOGIC SLIDE PREPARATION.
Fresh lymphoid tissue should be examined by gross inspection, touch prepara-
tion, or frozen section examination to assess whether the:
A. Tissue represents adequate sampling, and
B. Tissue needs to be allocated for various ancillary studies essential to a correct
diagnosis. The fresh lymph node should be cut perpendicularly along the long
axis, and material for ancillary studies procured as follows:
1. Wet touch preparations fixed in 95% alcohol or formalin, for H&E or
Papanicolaou staining
2. Air-dried touch preparations for Giemsa or Wright-Giemsa staining, cyto-
chemistry (myeloperoxidase, nonspecific esterase, etc.), or cytogenetics [i.e.,
fluorescence in situ hybridization (FISH)]
3. Rapidly frozen tissue for immunohistochemistry, cytochemistry, or genetic
analysis
4. Fresh tissue (in RPMI 1640 medium or saline) for flow cytometry
5. Sterile fresh tissue for microbial cultures or cytogenetic analysis (karyotyp-
ing or FISH)
6. Thin-shaved tissue rapidly fixed in glutaraldehyde for electron microscopy
7. Paraffin-embedded tissue after fixation for routine H&E-staining, immuno-
histochemistry, and special stains [e.g., Giemsa, periodic acid-Schiff (PAS),
elastin, trichrome, Leder, etc.]
Procuring tissue for histology takes priority over other studies. The most
commonly used fixative for permanent sections is 10% neutral buffered for-
malin. B5 fixative is commonly used in addition to 10% neutral buffered
683
684 I SECTION X: HEMATOPOIETIC SYSTEM
formalin because of the sharp nuclear detail it produces. Howevet; this mer-
curic chloride-based fixative is very expensive and poses an environmental
hazard. Furthermore, molecular studies cannot be carried out on B5-fixed
paraffin-embedded tissue, because it will generally yield poor polymerase
chain reaction (PCR) amplification results.
Ill. DIAGNOSTIC FEATURES OF COMMON BENIGN DISEASES OF LYMPH NODES. In reactive
lymphadenopathy, there are five different architectural patterns to be recognized,
with many showing a mixed pattern of response.
A. Follicular hyperplasia (e-Fig. 43.2) is characterized by an increase in number
and size of B-cell germinal centers and is common in lymph node-draining sites
of chronic inflammation. This pattern is also present in syphilitic lymphadeni-
tis where, in addition to the marked lymphoid hyperplasia, thickening of the
capsule by chronic inflammation, fibrosis, and neovascularization with arteritis
and phlebitis, and a marked plasma cell infiltrate in the medullary region pre-
dominate. Rheumatoid lymphadenopathy and acute human immunodeficiency
virus (HIV) lymphadenitis are other examples of marked follicular hyperplasia.
B. Diffuse (paracortical) hyperplasia shows expansion of the T-cell paracortical
areas. This pattern is commonly seen in viral lymphadenitis [Epstein-Barr virus
(EBV), cytomegalovirus (CMV), herpes] and vaccinia lymphadenitis revealing
an expansion of the paracortex with increased immunoblasts, imparting a mot-
tled appearance. Follicular hyperplasia and sinus dilation are often concurrent
findings in this entity, resulting in a mixed pattern of lymphoid hyperplasia.
Phenytoin lymphadenopathy represents a relatively pure diffuse hyperplasia
showing an expanded paracortical T-zone with numerous large immunoblasts,
eosinophils, plasma cells, and neutrophils.
C. Sinus hyperplasia describes increased cellularity within the medullary sinuses
of lymph nodes. Sinus histiocytosis is seen in numerous nonspecific responses
to chronic inflammation, as well as in lymph nodes draining solid tumors.
Sinus histiocytosis with massive lymphadenopathy (Rosai-Dorfman) disease
is characterized by markedly dilated sinuses filled with CD68+, S100+ histio-
cytes showing emperipolesis. Lipophagic reactions causing sinus histiocytosis
with accumulation of phagocytosed fat include mineral oil ingestion, Whipple
disease, and lymphangiography procedures. Prominent vacuoles in the histi-
ocytes can generate a signet-ring cell histiocytosis, a pattern that should not
be confused with signet-ring-cell carcinoma. Finally, vascular transformation
of lymph node sinuses (e-Fig. 43.3) shows a sinus pattern, and it is important
to distinguish this entity from Kaposi sarcoma (e-Fig. 43.4). The latter can be
distinguished from the former by the proliferation of spindle-shaped Kaposi
sarcoma cells forming cleft-like vascular spaces that contain erythrocytes, many
of which are extravasated.
D. Granulomatous lymphadenopathy describes formation of epithelioid granulomas
in lymph nodes. Caseating granulomas are epithelioid granulomas that form
central necrosis and caseation and are typically seen in Mycobacterium tuber-
culosis lymphadenitis. Special stains for mycobacteria in paraffin-embedded
tissue (Ziehl-Neelson, Fite--Faraco) will detect the bacilli as bright red, slen-
der, beaded microorganisms (e-Figs. 43.5 and 43.6). Mycobacterium leprae
lymphadenitis and histoplasma lymphadenitis are other examples of caseat-
ing granulomatous inflammation. Necrotizing, noncaseating granulomas are
present in cat-scratch disease (e-Fig. 43.7) caused by Bartonella henselae. in
which suppurative granulomas with stellate microabscesses surrounded by pal-
isading histiocytes predominate. Kikuchi-Fujimoto lymphadenopathy is char-
acterized by necrotizing granulomas containing karyorrhectic debris, but lack-
ing neutrophils; this form of necrotizing lymphadenitis characteristically occurs
in young Asian women. Nonnecrotizing, noncaseating granulomas are charac-
teristic of sarcoidosis lymphadenopathy (e-Fig. 43.8), composed primarily of
epithelioid histiocytes with scattered multinucleated giant cells, lymphocytes,
Chapter 43 • Lymph Nodes I 6 85
and plasma cells. This type of epithelioid granuloma can also be seen in drain-
ing lymph nodes of Crohn's disease.
E. Acute lymphadenitis is an acute inflammation in lymph nodes draining an
infected focus. Acute lymphadenitis is almost exclusively bacterial in nature,
and morphologic features range from focal infiltration by neutrophils to necro-
sis and suppuration with abscess formation.
It should be mentioned that in most cases of benign reactive lymphadenopa-
thy, a combination of more than one architectural pattern is present in the same
lymph node.
IV. INCIDENCE AND EPIDEMIOLOGY OF NON-HODGKIN LYMPHOMAS
A. 8-celllymphomas. B-celllymphomas constitute the vast majority of lymphomas
in North America and Europe, accounting for nearly 90% of all lymphomas.
Diffuse large B-celllymphoma (DLBCL) (""31 %) (e-Fig. 43.9) and follicular
lymphoma ("'-'22%) (e-Fig. 43.10) are the most common types. Immunosup-
pression, specifically due to HIV infection and immunosuppressive therapy to
prevent graft versus host disease, is associated with a markedly increased inci-
dence of mature B-celllymphomas, particularly DLBCL and Burkitt lymphoma
(e-Fig. 43.11).
Some of the low-grade B-celllymphoproliferative disorders include follicu-
lar lymphoma grades 1 and 2, chronic lymphocytic leukemia/small lymphocytic
lymphoma (CLUSLL) (e-Fig. 43.12), nodal and extranodal marginal zone B-
celllymphoma (e-Fig. 43.13), and lymphoplasmacytic lymphoma, which are
generally indolent, but incurable and usually present in a disseminated stage
with bone marrow involvement. Mantle cell lymphoma {e-Figs. 43.14 and
43.15) and DLBCL represent "intermediate-grade" B-celllymphomas that
generally show a more aggressive clinical behavior, but are potentially cur-
able. The same applies to high-grade B-celllymphomas, which include Burkitt
lymphoma and precursor B-lymphoblastic leukemia/lymphoma.
B. T-celllymphomas. Mature T-cell and natural killer {NK)-cell malignancies are
rare, accounting for only 10% to 12% of all non-Hodgkin lymphoma (NHL),
and usually are more aggressive than B-celllymphomas. The most common
subtypes are peripheral T-celllymphoma, unspecified {"'4% of all adult NHLs
and "'30% of peripheral T-celllymphomas) {e-Fig. 43.16) and anaplastic large
cell lymphoma ("'3% of all adult NHLs) (e-Fig. 43.17). In general, T-cell
and NK-cell malignancies are much more common in Asia and are linked to
viral infection with EBV (NK-celllymphomas) {e-Fig. 43.18) and human T-cell
leukemia virus (HTLV-1) (adultT-cellleukemia/lymphoma) (e-Fig. 43.19). The
current 2008 World Health Organization classification of lymphoid malignan-
cies is summarized in Table 43.1.
V. PATHOPHYSIOLOGY OF NON-HODGKIN LYMPHOMAS
A. B-cell lymphomas. Mature B-cell malignancies mimic stages of normal B-cell
differentiation; therefore, classification is generally based on their morpho-
logic and immunophenotypic resemblance to the normal B-cell counterpart
(Fig. 43.1) (Best Pract Res Clin Haematol. 2005;18:11). Normal B-cell devel-
opment begins in the bone marrow where precursor B-lymphoblasts undergo
immunoglobulin VDJ gene rearrangement and develop into IgM+, IgD+ na1ve
B-cells with surface immunoglobulin light chain and CDS expression. These
resting B-cells circulate in the blood and occupy primary follicles and man-
tle zones of secondary follicles. Malignant counterparts of CDS+ naive B-
cells are believed to be CLL and mantle cell lymphoma. Upon antigen stim-
ulation, naive B-cells undergo blastic transformation, migrate into the cen-
ter of the primary follicle, and form the germinal center. These centroblasts
are large lymphoid cells with vesicular chromatin and several eccentrically
located nucleoli. They express BCL-6 and CD10 but switch off BCL-2 pro-
tein expression, therefore becoming susceptible to death through apoptosis.
Centroblasts undergo intense proliferation that is accompanied by somatic
II& I SECTION x.. HEYATQPQIEliC SY5TEU
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Normal and Abnormal Counterparts of B-Cell Progenyt

..-...- Early Differentiation in Bone Marrow _.,....,.
Stem Progenitor Precursor Pre
Cell B-cell B-cell B-cell
4 lgHGR +/- kA,ttt --~

Acute lymphoblastic lymphoma (ALL)
.__..,._. Early differentiation is antigen independent
. - - - - - - Mature 8-lymphocytes - - - - -..
Early B-cell Mature B-cell Activated B-cell Plasmacytoid

lymphocyte
. .- -NHL, PLL, HCL****' -.....MIII - or

lympho-
plasmacytic
Mature plasma cell lymphoma
• ~Multiple~
l ~myeloma:-j
t Only positive markers are noted; *Cytoplasmic heavy chains

+1- indicates partial expression **Markers abnormally expressed in neoplastic 8 cells
tt Surface immunoglobin or plasma cells
ttt Immunoglobulin heavy chain (VDJ) gene ***Cytoplasmic immunoglobin
rearrangement with or without light chains **** Non-Hodgkin's lymphoma, prolymphocytic lymphoma.
tttt lsotype switch, lgH and kA. rearrangements hairy cell leukemia, etc.
Figure 43.1 Brief overview of normal B-cell progeny and malignant counterparts.
NK.-cell malignancies (Fig. 43.2). For example, hypercalcemia is associated

with adult T-cellleukemia; hemophagocytic syndrome occurs more frequently
in cytotoxic T-cell or NK-cell malignancies; persistent severe neutropenia is
a relatively common clinical feature in T-cell granular lymphocyte leukemia
(e-Fig. 43.22); and systemic symptoms of edema, pleural effusion, ascites,
arthritis combined with anemia, and polyclonal hypergammaglobulinemia are
relatively specific for angioimmunoblastic T-celllymphoma (e-Fig. 43.23).
Normal T-Cell Progeny rr
Bone marrow precursor In the thymus
Migrate
to th ymus
loll Subcapsular ,. I,. Inner ,. 1111( Medulla ••
cortex cortex
Stem cell Precursor T cell
I ll( TCR** gene-rearrangement: 1st y8 *** or if yo unsuccessful, Ill I

then ~ and then a
t Mature T-helper
double ~ double 4/8
I .r 4/8
negative
,. positive
80% a~
--
,. (CD4+) or
T-suppresso~
* Cytoplasmic CD3 (CDS+) with
** T-cell Receptor aWCD3 complex
*** Gamma-Delta chains expressed on immature thymocytes and Natural Killers cells only
t Mature T-cells can circulate in peripheral blood and tissues
tt Only positive markers are noted. Markers partially expressed or about to lose expression are noted by a"+/-".
en
os Figure 43.2 Brief overview of normal T-cell progeny.
co
110 I SECTION x.. HEYATQPQIEliC SY5TEU
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3. CDl 0 (CALLA, common acute lymphoblastic leukemia antigen); expressed

on marrow pre-B-cells and mature follicular center B-cells)
4. CD22 (present on pre-B-cells)
5. Cfk (cytoplasmic fk heavy chain)
c. Primarily B-cell associated markers
1. CD19 (present on marrow pre-B-cells, mature B-cells, but not on plasma
cells; no paraffin-reactive antibody available)
2. CD20 (present on marrow pre-B-cells, mature B-cells, but not on normal
plasma cells)
3. CD79a (present on mature and pre-B-cells, as well as on plasma cells)
4. CD22 [transmembrane molecule, present on mature B-cells (cCD22) and
pre-B-cells (cCD22)]
D. Markers helpful in subclassifying mature B-cell lymphomas
1. CDS (expressed on all T-cells and small subset of B-cells, expressed on
neoplastic CLL and mantle cell lymphoma cells)
2. CD10
3. CD11c (expressed in high levels on monocytes, macrophages, and NK
cells, as well in moderate levels on granulocytes; expressed on subsets of
T and B-cells)
4. CD23 (present on activated mature B-cells)
5. CD38 (primarily expressed on mature B-cells and plasma cells)
6. CD43 (leukosialin, expressed on the surface of all leukocytes except resting
B-cells; CD43 expression in B-cell lymphomas is highly correlated with
CDS and is therefore an indicator of aberrant B-cell populations)
7. BCL-6 (expressed normally in germinal center lymphocytes in the normal
lymph node; it is distributed in a pattern reciprocal to that of BCL-2)
8. BCL-2 (present on T-cells and normal mantle B lymphocytes; aberrantly
expressed in majority of B-celllymphomas)
9. Cyclin Dl (cell cycle regulatory protein rearranged through the t(11;14) of
mantle cell lymphoma; also expressed weakly in hairy cell leukemia and
plasma cell dyscrasias; nuclear protein detectable in paraffin-embedded
sections)
1D. CD138 (Syndecan-1, expressed on most cases of myeloma, but also present
in carcinomas)
E. Markers of clonality: " and ).. immunoglobulin light chains
F. Primarily T cell- and NK cell-associated markers:
1. CDl (expressed on cortical thymocytes and Langerhans cell histiocytes)
2. CD2 [present on all T-cells (thymic and peripheral T-cells) and NK cells]
3. CD3 (lineage-specific marker forT-cells; cytoplasmic form also expressed
in NK cells)
4. CDS (expressed on all T-cells and a small subset of B-cells)
5. CD7 (expressed on all T-cells and a small subset of myeloid precursor cells)
6. CD8 (present on cytotoxic subset of peripheral T-cells and on a subset of
thymocytes and NK cells)
7. CD16 (present on NK cells and granulocytes)
8. CDS6 (present on NK cells and subset ofT lymphocytes; also present on
myeloma cells)
9. TIA-1 (cytotoxic granule-associated RNA binding protein), granzyme B,
and perforin (proteins released by cytotoxic T-cells, inducing apoptosis) are
expressed in cytotoxic T-cells and NK cells
G. Flow cytometry can easily detect clonality in B-cell malignancies (normal" to)..
light chain ratio ranges between 1:1 and 4:1). Clonality forT-cell malignancies
can only be detected by molecular or cytogenetic studies.
H. Southern blot analysis is considered the gold standard for identifying clonal
immunoglobulin heavy chain (IgH) gene or TCR gene rearrangements.
T lymphocytes normally rearrange four different TCR genes (TCRa. {3. y. and
8) to encode a unique antigen receptor expressed on their surface, and South-
ern blot analysis targets the TCR/3 gene. In contrast to normal tissue, where
only one band corresponding to the germline configuration is visible with this
technique, both B-cell and T-cell malignancies will yield an additional band
corresponding to the clonal population.
I. PCR. Because Southern blot analysis is labor intensive and expensive to per-
form, many laboratories have turned to PCR amplification methods to detect
clonaligH and TCR gene rearrangements.
The 200 variable segments of the lgH gene contain three highly variable and
mutation-prone regions called complementary determining regions (CDR1, 2,
and 3) that are interspersed between four conserved framework regions (FR1,
2, 3, and 4) that provide reliable targets for consensus primers. About 70%
of B-cell malignancies harbor donal rearrangements that are detectable with
framework 3 primers, and about 15% to 20% of additional donal rearrange-
ments can be detected when framework 2 primers are used. Therefore, most
laboratories use primers targeting frameworks 2 and 3 to detect at least 80%
of all B-celllymphomas.
The rearranged TCRy gene is most suitable for clonal detection by PCR
amplification in T-cell malignancies, because most T lymphocytes harbor rear-
rangements in 1 of the 11 y segments. Because many of these segments are
homologous to one another, they can be targeted by a single consensus primer
set. TCR8 and TCRcx cannot be targeted because TCR8 is deleted during TCRcx
gene rearrangement and TCRcx has such a diversity of possible rearrangements
that these cannot be covered by current probe technology.
J. Cytogenetic analysis is a morphologic study of chromosomes to assess changes
in their number and structure.
1. Conventional karyotyping requires viable cells, and analysis of chromosomes
is performed according to their size and banding pattern after chromosome
staining. Although this study has become critical in the diagnosis and clas-
sification of acute leukemias, it is not frequently used as part of a routine
diagnostic workup for lymph node biopsies.
2. FISH uses one or more labeled probes directed toward specific portions of
chromosomes. It is helpful in locating specific translocations, deletions, and
amplifications of chromosomal regions. The advantage of FISH (see Chap.
59) is that it can be performed on nondividing nuclei (interphase nuclei),
eliminating the need for cell culture. Furthermore, interphase FISH can be
performed on a wide range of specimen types, such as peripheral blood
smears, cytospin preparations, or paraffin-embedded tissue. Many of the
classic chromosomal translocations that characterize specific lymphomas
can be identified with interphase FISH techniques. Tables 43.3 and 43.4
summarize the characteristic morphologic, immunophenotypic, and genetic
features of B-cell and T/NK-cell malignancies, respectively.
VII. PLASMA CELL NEOPLASMS. Plasma cells represent terminally differentiated B-
cells. Their neoplasms are characterized by secretion of a single homogenous
immunoglobulin product known as the monoclonal or "M" component, which
can be detected by serum and urine protein electrophoresis. The World Health
Organization categorizes plasma cell neoplasms into monoclonal gammopathy of
undetermined significance (MGUS), plasma cell myeloma, plasmacytomas (PCs),
immunoglobulin deposition diseases, and osteosclerotic myeloma (POEMS syn-
drome).
A. Patients with MGUS are asymptomatic with no evidence of lytic bone lesions,
but reveal a clonal plasma cell population in the bone marrow of< 10% and an
associated small M component of <30 FJL. No lytic lesions or myeloma-related
organ damage is present and a B-celllymphoma with plasmacytic differentia-
tion should be excluded.
o-..
ClUil!.
Cl'-ll>lllf _ _,.,
Mosllt Oljmpll>mollt, tni!Y
IIIU\11li>IIIJ
PB: Srnal colo w!th<OelliO\Y
...........l>l.nlt
al20 (-11), CD19,f&M,
.......
Trllomy 12, dlllllonut
<limped <111>"'"'*' end
~- fl'llll>oo,
.............. homob1i= h»mplcuous nu<!ool
(tO, alll2 (-11). CD~;,
ali'3, CD79o. 0043
13ql4,d-Oil
llqll2-2il
Prolfmp"""¥11c
........... he!>etool>lenom...W.
tlmPhlldenolii>IIIY: 1><1111>11era1
•blciiD lfm"""'*"l&
Mllbd 'l>lln«i"'I\Y...m...l
---of
LN:Prolmte~•~
"paiUnmul'tllllula" •I'd
prot/mpl!oq1as
PB: IIIII! kim..-- o.t11 al20 (brV!I), CD19, ~.
~IIIII&~
~no.llfi
doNIf ,..,.l'fl'd
canp~aa ""~ ...
tlmpllod ....pallly, flll'lcly round nuclous, ~ al22, CD711o, FMC7, common~ JbNM'ml!liM: of
IIMNmll
liiii'C lfmphocoJto eoijnl _oll,..,_,..,d ali'31yplcallr ·-cos TP63 In -tiO'J.. I(H
C>100 " 10'/l.l promNI!t <lllltlel nucteollo Pt&0011tln113ol- cb\aly re.lrnlrcod
tllkYooiiiMJklmll Prodomba.mloid~ men. PB: Sm«J. to MICIIJm-ol!od col& all&, CD20, C022 (b!Wrtl, Nool*llle~
__
Ol'i&lmllllllllr. PII""'''II)f)llls wtth""'rrrfetenllslliii!Y al79tl. CDlie(~. «b_,.lty,llfi clcNiy
LJ!nplql~
t.mphornll
Walkno111!m
wtth~penla
""'·~..ttl-lad
,.,.........,
I!IW.tloooty eym!*mt
pro)dono
Spleen: Rod !>liP ln11-
Crna=ol')': "'JI>ody 11.....1
Mll:lln of """!111m~
~'""""'"'
..... "*- lfmpllo<:JIOO.
Dln:het b>dlao
end
-..
al2S lblWI1!, CD10:!1,
TRAI\DBM4lnllul»
all9, CDII), C022. CDl'!le.

alilll. ,.,_end
"ltX>Piltrnle IBM. no
""'"""~~~~'~
No 1podilc .,.llltic
«b""""lty.llfi doNIf
I<IOiit«JWWd
,...,.,...b<JIInomlo ~o!COS.CD:zil,
Of COlO
Splon~ maiJ,NI Splono,....lf, . , , _ . PB: SmiJ. to MICIIJm--- allS., CD20, 007llo, IJM Allelic ""'old!_,.
mnoB-coll -101111•-.rnune with pollor Yl!l (¥1- and (tO, no - - n o f 7<!21-a2ln40\lloof..-.
t.mphorno lll,..,bo<:ylo..,.... or""""""· IJm~
Spbon: Both-pllpond Old
al6, CD23, COlO, or tbomy31nrn-IJH
doMif ...mlrco<l
""''~ lfmp-nopotny C043
1111t> lnl'l!nltlon
....•
IMOrnii'IOtl
(oMII'o<lold)
••
• 1m=! • '1:5!11 c~~a,.d••tooucr- <If DillS a~ ~.con Mall.ll&tlld• <m•..o
D•a. ~1111 ..1Jn'Millotta.t 1111,..1111J ••••~'"~,. Caallfn
Ealno<llll mqn&l Hlllay alclwnl: lnfllm""""'Y i'olymofplloulln- a1 Cl>19, CD20, CDi'!IQ, .,., no 1(11;lll)(q21;<121) In -25%
"""" BGI Of •!b*nmune diad.,. ~llo> <olb, ""''"""lonal COO, CD23, to !iO'll.o f -
t.mphom.o(INI\1 ~P)'ftlrl"'""' mo~<ell..,al CI>IO 1(14:115)(q32;~.1)
bmphoft\ll Haohmolo111midlll. S!Oiretl limP~ -..s ~1\i.nl, lllti c:lcNIY
~-l. Olnctll'lO'It mmunol>hblo end limr>llcid r&~~mu'CIId
comnm.aball- '*With...,,~
-
dftltwtlltlon, .CIItlfl'UI\
'*""(limpliooplhol~l
~ ,...lltlal zone l<>:$1md OfiOnOrdmd Milllbll zone 0111d ~- CDI9, CD:!l>, CDl'911. _,.,no The U'OIM~
wtlll ~al MZI. _,..
....,...led
lkoiiiJmDhoma 1/mlhdo~ .,... ~llnl1odb!' ""''l'&lllllorl al COS, CDit),
~l .... lkolk, CDIO not .W""''; laH elonll!r
,...,~ lk>el&, til amd roononpd
limP,.,.,_ plosni1 col
d i l l . . - mi!Y 1» ~
Fol...,r~phomo Wkllsprood poolpllonillllld Fdwlllr )llltl.wn alclosoly Cl>19, CD:!l>, CD22, CD7'll, 1(14;lll)(q32,"<!21) ....
<Onbal bmpllad""pa111y, pocl;od ""'plint!o l'dll::loo BCL4 BC~ CDIO RWIIT8l-il8rlt of the
-
""""mlliTOII' -ln40'll'. """poood " ' - ..... BCL·2 IOnOiood~ to
"'- den..-:d ~ « ••
G-l:0-6oo-l!llllllpf
--lorlal111o
BCL-2 prolaln ill1d s..WOI
""........,...lpnl
Gredo2:6-15-pt
Gredo3: >15 -IMb'llpf ...,.,...
c:olb; I&H _I.,
Marllo ..alimpllomo ~..-1>11111 ......

commonGCtrllllOdlllet&lb111o
hloy-"""""'
«
dl'llllo
nodi""·
zone
m.ol'ido Plltlam.
CD19, CDi!O, C06, FloiC:.7,
C043, cydiii-Dl.lllck of 11'01-
1(ll;l4)(ql3;q32).
b~-"'1IIH
G11rllct trw-.,1& modklm-Czod lfmlilold <oil! ""''l'&lllllorl al coza. CDIO, and BCL·l-: IIH
lfmfilom"""" .,..,.., IOI!IIm(pl~r nidw-l.ft, BCL-6 elonll!r """'"'11111'1
rnoy 1» blaslllld In""_."""
emu. ilrplk:ll Rtlltt\1' onlllratna. oftln -olliqlcolllwlllnll:loor C019, C021>, C02:!, CD1'ill, I&H cl:lnl!ly rarr.,.,od,
t.mphorna (DLSCL) """pb>o1111io ITlOIO, oftEon olle equal or_,,. oormlll COlO, olld BCL~ -30ll'> l'llll atonormlllniooo
dbooml_d_
"""""""""'"-
Morf)hololllo ""olo!Mol:
Colntrobll.oek, l'nmlllObldo,
--:pnnhol
...,,., &-lb (bellor--.1
~11Mn~
o1 ~27 """"lowoMna
BCI.6
T -~ rk:ll, ,,.plulle 9~)
- llwl~l:) Moolly_,.n In lhN tloiniiD Lilli" COils ..til MOlCilbd C019, C02D, C030 6oMk!, I&H..,..IIfr•~;
lllrp-
t.mphoma
IIIIo d -.....-
msdll.litnlll me.-. wmaltnes
wtllllml>illldnreuDtriorYBnll
~~-
..
d-~n~oo.-n~~~os~s
f>OOIII>Io"""'""_'_"'
"""' tloymlo """"nil.
bloi>O!'"' mDilo diM omd and
la•na HlA...,.tnll
-lonboftonebloorrt
IPJI>onl!plold .. l)'lllypo.
pNin<-9p
f)fc11.oo!o-""*
-----
obleond 1>,-
and"""" aJ!!fal:t
Buotoll t.mploom• Hfllo\1' .......... tlmplooml C019, C02D, C02:!, COlD, I&H donoollf ••IYM.II'd;
obn ~na.tm•omel booplollc -IIOIWd C{)JfMJ BCL-61 no 1(11;1<4) Inmoool.-,
- o r • . . . llullllmloo, rllk ~~~ anc1 racukr nii::W -lonoiTdl, BCL-l!, 1(2:11) and t(ll;22) ..,.
for ...- ........... "¥"f'ro wth -...nl .srna!ll'ltl.ldloll 10-Qind.. of IOO!r.
~-.....-. "'lo<<omk:'" 8uri<ltt
tlfnplooma wttn ~em ol
Unalbb-body m•"""'-
lmPIII11!1I._I!Y-OiiY"
- llld ot""' fldll
llln.lo:llns, • •porut t::" 8 uri<ltt
•-mnco
tlfnploomo wtttl•bdomlnol
on ....
CU.- tim~ ....lolmiii;$U. omell irlrcl~ IJm- MAll, onu0011a 1111 ><laW IJm-lllooe; II. Ion,..,oqt:o~xofn QI, iiiiiNiualhot f8, ~I
- Lll; hilt, hlii!--MI; TRAP. tl1lr<lln-.~dl- HLA. humoon lou-•~ Tdl, amlwl~ ~--
....•
i r1'i~Q~:~)~~:f~·S~X~li!}l
o-..
CllaJtdelll1lc Fullnl'lf ~""""""" T~llllld fjlt~IIIIIIIIII'Odet
Cllllr.l Ifill rtt'IM ..,..,,. __,_,.,.. _,.,
r....nprot~mp!Jo<;)11o 11#-d->Oith PS: Mlldl..,..lzld colla wll\ C03, C02, C07, C04+1C08- ln'<(14)(q11;qW In 110\11,
le<Rmla ho~l&non-tt and .......... C$>plinm •.tllblo _, G)'ll',, CD4+1Cil!l+ In 1(14:14) (qll:<jW In 10'll.
IIIII'MIIIIod tim~.
mllbd tim~ ,.Uitj
""'*"'"lo
nucloolle. ellll
I>IOCI\IIJIOno«bloll&
:I:Slt>, co.h-tD$+ In IS'l(. ~ltR...,,6ond
ret.-1. deNlireamtnft)d
>100 X 10'11. · -llld TCR
lltromboc>yt.oponla
r... u~a,...,..,..,, Indo,.,. dnbJ COU/lit~ .tMIIl'6 pg, Llrp .,.,••" tlmpllcq1as coa, TCR...t,6, coa In 110'11., CloNJi ,..,.,pd ltR, no
tim~ wll\ ol>ulllont~ end
~·
-nmlooolTIA-1, unlqw eylopholl<:
lcMicomll onomll, mltl II> m - ,.,80f 0011/118 ODI'Ophk CD57, flOf(orln, IV""'Yl\1& 1\bnonnal~
tlm~ modenl1o !Pralilo:BM:~I B. Fao !CD95l ellll Faol.
'l'iem"'""'lf, rlwJmmld lnfl-
l!lrthrtt:I.!Ji~ du.lfldttrlmmune
compllilcas, rw-IPJI'In...
l)oll..,..,..
_._NKoall Morep-nlomqMiilm; pg, (Jmpllold-lo!p<tllen C02, cCD3 ,, C056~ 'T'JA..lf TCR no! cbldy ,..,......,,
lcMicomlo -.loumlo blood ploUo, nonnal LGL wll\ •1\Chi!J . .noymo 8, porto!fn, EBY. ESV ~ h <~IIIII
"'mill~! "''fllploms.
_
b.,opl\lt::'*"'IMm
_,[f11IJillnU[\)O, no""~'""""".,""'"""' oploomall'onn
~·· hoTJO.toeolr
lm'IIIIII!Y. IIIJiiotpn flllu,.. hypotdl- .......
C03«C057
.... tot 101\m solJblo Fe
-ion
ilpndHI
NluiT-all.......,ll £ndornlo In Jlpon,llto Cort!I>Nn pg, Mlldllm·lo IIQilHIZod colla Cl>2, CD3, 0051 CD4, C025, CloNJi lra.,..tld Kll\41,
bonln, porto of C8n1nll-.
tllild '> HTI.V-1. ('11_...,,
wll\ polf!ollobid - -
lodl C07 '*"'".,...."'"""' ltR
li!!Potoe~ll.
llol>'bi>IMon'IIIIII!Y, ti.CH,
.....,lotldT....a
lmmunccloll:ilnc!rwll\
oppo11111lrllo , _
i:XW~I NK-11"-<>ll M010 P,_l'lt b Mil.,..,., ~~~~r.t~. Cl>2. CD56. cCIXh. 11:R notc:ioMIY lllllmii'Cild.
ttmphoiNI, nlllll Centro~,
_ _and m_S..IAI!Amlllb,
__ tnfjocoll!!t: ~nd VIN!JimO 8, TV\-I, £8V P<-ln elonll
t)l>o
·~~ pMo/11\ 111Jifoo&C03 I'd "11-lfann
- ... -
Gl not
cdrancdalllloo
lnc:luda ..... taft t:tasw, tasl:tt,
pc!lom, hypon:llnomllll::
....~~~~~, lnlllmbalcl
lnflomrnalory....
"''"-
~T-<>&1 Abeocla1od Willi <:ollie d lloolt» U~m-lma•b Cl>). CI>8-H-. Cl>l<G. ClcNiin~~~1111n.DldTCR. HLA
ttmphoft\a end~~l'llk )o)Jnum or him. btoed TIA-1. PI\IYIIIO, perfoltl IXIA1'050l.IJQB1'1)21)1
~~ •pedrum,lnbtmlad pno1ype II""!"".......
lnillmrnalo'Y celo
~pllni:T<III
ttmphoiTII
PMk lndd""" ln•do-
olldyou,.od...,morlold
IIIPI1DII!>Iotuo.,..tf"""
tlm~.rn«e
oomnm In.,..- ..ttl
lmmiiii*ITJIIIOIIOion
""'-...,..,..
S*ILIIIOIIIII!nlllrwllonalllw,
oploon, •lid lloni> ITIIrrow by
mol(fii!Himd ....
CD3, TCR:-y/1, llA·l,
l'lllllf1Nv "" 004, CD&
TCR-«<ft, •lid podwln
ClcNJyrumo~
bodw'omotOiillD
trbomy8
TCR,
7q,
subaill..... Mulllplo ,.-.,.nod..., OIIIUie, ...... ' " " - ' ·

C03, COS, -oym• 8, ClcNJy,...,.~ TCR
ponrCIIIk-ll<a M"f boo-ted oolth ..- wlt"""' ,pou~,. f)tlfool" nA-t
-~ "'"'""""
T<CIIItlmpi>Oin& saptel; 6pfd~tmlt urlrwol\'ld;
1'1111 pon<:)ll;'l""""' - · ....S """""'· """""-. rtnmhf
IIIPI1Dill>lotu II....\I' orl<rtoais
t.ttcoollfU~ l.or'C 11111111111 hlltoly, mulllplo ~ ~ldennOO'OO~ ln11i!MI oftmor Cl>2. CD3. CDS. 004, ClcNIY lllllmii'Cild TCR
IGIJioM.I'OIIdl-+ PMUO ... <o~l~dll..,.reMJorm• nue1a1. TCR-«<ft. no <lM181'1Jio~of
t<mr,11'11quenllyon lnllk Pll~t'IJJ mb'adbsca•e CD7
_ , oync~ ..... Aai&llt:w'AI!Wrtof~ PB, l.llnmurn ai!OOO&Izo'Y I n c - CD4.C08 rllll> ClcNJyru"""'JIJI TCR
f-lclol;~. ce!ll per mml. •carebrtfofm,. loci<l,.
lllf1!1101b1 of CD7
tlm~.ond -lei
<R<IIr<thf$6my....
..•... [0>1-
•
••
Pli.t:!l:(£tj( ~FIIIIIIniGfDtft'aJ&tT.Qilland .. K-alliMIIf.&nand•~
--a!
'
DlleiA Clllll:ll , _ ' - II.,_IDD ...nplllaaw- a-11:1
Mllclmmunobllo11o lmmu~..c.,ncloiiY" Mtt1Un>of 1>011m01~ Cl>2. Cf>). C04l0tll ClcNIY ,..mii'Cild '!CR.
T,.,.ll tlfnplloma tlfnplloma, 111m lUll,...,..,., tim~ - • e\ur tltlomy S,lrlllt>my S,
plolftle1\lalon,aaellot, ~plolm,.....,.....,
COlO,
folk:tMr dendrt!Sc oolll .,. addlloNIX ell,...,...,.•
qlojlonfll, ~I -pltlo, ond hW!IWr\111 byCI>21,
~mmq,lol>llllnomll, + lmrr&lnobllol>; -~ HlV; CI>ZI, coa!i,
''"""-- proml,.,...,ol FDC """"-k lmmuno-.,CI>20+
o.-.lcla i)tlcia& Oll1d EBV+
Pel\?hint r....11 PradomNI!Cii nodlll-buucl BIOOid ~ -m. T,zono Ct>2. Ct>3. ~ C04, mw Re.!mll\,lld TCR
tlfnpmiNI,
-11\od
AN!>Inllc lqll <Oil
tlfnpmm.s
--"'·--
tlfnpllomu lhot oonrd bo
·--of
diM.ua
cllltlhoo<l
tlfnpllomo- 114ip (Ill
or M will froquont8ldnlnodal
...- . P<....wlio• tl
fdllcM. LMI'MI wi!MtlOtll
many aplholbld ellalrua
Kllmlrl: ..,~ = lrup .... with
abllndlrt ~1o1m end
hoo...._.holped nldil
--coso
Cl>30 (momblllno and Gol&f),

ALK:l (60ll-«ill>),
EMA+. CD43. C02. CD4
Ra.t"""'fl'l TCR
1(2;5) ~ NPM aNI ALK
II'I'IO!f<IIMI'It
tf(. omll'll kllllr;.HT~hurNn T-<011 lill.bmlt>iN'< LDH, LGl,lll"ffO-1; fB, pell""""l-; Bioi, lloMmo"""' IJ:I, q .,.nu~rljn\"'= HE\\ h~
ll'ollolllollll.._.., :::·< _ r _ l f t l e _ TCR, T- """''b, nA, T-<OIIrlll!t:Oid 1-lullro~i':... al, F•...l>d; £8V, (jloall...a..r ; Al.K:l, ll\lpluli>
lllpc•ll-1; IMA,'I'"""~I memll-. . . .; teL• I, T...al6illo!MII/Ijmpltomo I; HLA, IIIJINf'ollulcoojl&l._,.
~ 43 • I.J'Mp!l Hedee I •••
''I.I II ( fHI Q11111a fof 1M Dlll!*fuf " ' - tdlo!Jeloma
f)o.... • ,..._ ooll ,.,.._
PC WI tb:lvt *'PI'I«
donll bono"""""' plurne<:)tol\11 ("""'ly >lOll plume.,.lll)
M~><*lh In ooturn « u'*">: uouetJ Ontt nct•~•ennn 1&0 >30 etdl «ItA >2Srfdl.
• or~ IIB!rtoholn - · :.I.I)Wd"' ~ t.r!M -.p~~o,_
Rtlobd - · orlliale mpolmum(llnemla, ~mil,
r.curnllnl: fnfec;:tk)ns, ek:.)
t111t-..,
-lnollf!ldlnoy,
Allllf!I'Fff<(l.. ll!rlrcl . . . . . . .,...,...

M~><*ln In •urn at myalorne - (>30 IIIJ
Olld/llt
CiollolboM IMIOW ratno.qttlll& (10!1:. t1 motO)
No rolnod 0111'1' ortlalua mpdnwt
llllta• Orpur11aau
~ ,....._.. S.. M c.j~alllt lytle IIIII au llfllllr•n
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extracellular matrix as ,8-pleated sheets containing amyloid P component. The

deposits are visible as homogenous, pink material in tissue sections that are
green birefringent by Congo red staining. The clinical consequences are related
to deposition of amyloid in organs, resulting in organomegaly and organ
malfunction. Rarely, plasma cell or lymphoplasmacytic neoplasms secrete
abnormal light or, less often, heavy chains or sometimes both, which may
cause similar but nonamyloid deposits in organs, most commonly in kid-
neys (Hematol Oncol Clin North Am. 1992;6:323; N Eng] Med. 1993;329:
1389).
E. POEMS syndrome represents an osteosclerotic myeloma with associated clin-
ical features including polyneuropathy, organomegaly, endocrinology, mono-
clonal gammopathy, and skin changes. Lymph-node biopsies in these patients
may demonstrate Castleman-like features, specifically its plasma cell variant
(Blood. 2003;101:2496-2506). The bone marrow shows plasmacytosis only
in the vicinity of sclerotic lesions with <5% plasma cells elsewhere (Blood Rev.
1996;1 0:75-80).
F. lmmunophenotype. Neoplastic plasma cells typically express monotypic cyto-
plasmic immunoglobulin and lack surface immunoglobulin expression. Most
cases lack CD19 and CD20 expression, whereas CD79a, CD38, and CD138
(syndecan-1) are found in the majority of cases. CD56 is aberrantly expressed
in up to 70% of myelomas. Expression of cyclin D1 is also a feature of plasma
cells, specifically in multiple myeloma (see Section VII.G), and diagnosis of
mantle cell lymphoma must be avoided on the basis of an isolated finding of
cyclin D1 positivity. Other antigens that are aberrantly expressed and could
be helpful in detection of minimal residual disease or targeted therapy include
CD20, CD117, CD52, or CD10 (Am] ClinPathol. 2004;121:482-488;Blood.
2008;12:4017-4023). CD56 negative myelomas show more extensive marrow
infiltration, lower osteolytic potential, frequent plasmablastic morphology, and
a tendency toward leukemic transformation (Br] Haematol. 2002;117:882-
885).
G. Myeloma genetics. Plasma cell neoplasms demonstrate clonally rearranged
immunoglobulin heavy and light chains by molecular studies; cytogenetic
methods to detect chromosomal abnormalities include cytokine-stimulated
bone marrow biopsy for conventional karyotyping and FISH. Combined, these
modalities increase the frequency of detectable genetic aberrations in myeloma
to over 90% (Cancer Res. 2004;64:1546-1558). Complex karyotypes with
multiple chromosomal gains and losses are the most frequent abnormalities,
the most common being gains in chromosomes 3, 5, 7, 9, 11, 15, and 19 and
losses in chromosomes 8, 13, 14, and X (Blood. 1995;85:2490). Monosomy
or partial deletion of 13 (13q14) is the most common structural chromosomal
abnormality. In about 40% of cases, translocations involving one of five recur-
rent oncogenes are seen. The most common translocation is t(11;14)(q13;q32),
involving the BCLl locus on chromosome 11q13 (CYCLIN Dl) and the
immunoglobulin heavy (IgH) chain locus on 14q23, which leads to over expres-
sion of cyclin-D1 (Brit] Hematol. 1998;101: 189; Blood. 2002;99:2185;] Clin
Oncol. 2005;23:6333). Other translocation partners for the IgH locus are
4p16.3 (FGFR-3 and MMSET), 6p21 (CCND3), 16q23 (C-MAF), and 20q11
(MAFB) (Table 43.7).
VIII. CASTLEMAN DISEASE. Castleman disease is an important consideration in the dif-
ferential diagnosis of lymphomas due to its clinical presentation. It can occur at
any age but has a predilection for young adults, often presenting as a mass, most
often in the mediastinum, although cervical, abdominal, and axillary nodes may
all be the sites of origin (Cancer. 1972;29:670; Semin Diag Pathol. 1988;5:346;
Mod Pathol. 1992;5:525). In some cases, generalized lymphadenopathy with
B-symptoms, anemia, thrombocytopenia, and pemphigus mimic the clinical
~43•1.J'Mp!l Hedee I 101
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type Castleman disease can be cured with surgical excision, systemic therapy may
be required in multicentric forms.
IX. HODGKIN LYMPHOMA. Hodgkin lymphoma typically presents as a primarily nodal
disease characterized by a predominance of reactive cells with a paucity of neo-
plastic Reed-Sternberg (RS) cells or variants. Hodgkin lymphoma comprises two
distinct entities: classical Hodgkin lymphoma (CHL) and nodular lymphocyte-
predominant Hodgkin lymphoma (NLPHL). These two entities differ in their
clinical appearance, morphology, immunoglobulin transcription of the neoplastic
cells, and immunophenotype.
A. CHL accounts for "'95% of Hodgkin lymphoma and is subdivided into four
categories: nodular sclerosis ("'70%), mixed cellularity ("'20% to 25%), lym-
phocyte rich ("'5%), and lymphocyte depleted (<5%). The neoplastic cell is
the classical RS cell or its variants (mononuclear forms, mummified forms, or
lacunar cells).
1. Nodular sclerosis CHL commonly presents with a mediastinal mass. His-
tology reveals scattered RS cells embedded in a mixed inflammatory back-
ground of mostly T lymphocytes, plasma cells, eosinophils, neutrophils,
and histiocytes, compartmentalized into nodules by thick strands of colla-
gen fibrosis (e-Figs. 43.33 to 43.35).
2. Mixed cellularity CHL presents with scattered RS cells in a diffuse or
vaguely nodular mixed inflammatory background without nodular scle-
rosing fibrosis.
3. In lymphocyte-rich CHL, the nonneoplastic cellular background is com-
posed of predominantly T lymphocytes. Other inflammatory cells are rare.
4. Lymphocyte-depleted CHL is rare and occurs most commonly in association
with HIV infection. Histologically, sheets of RS cells predominate without
a significant lymphocytic or mixed inflammatory infiltrate.
The main clinical findings of CHL at diagnosis are painless peripheral
lymphadenopathy, most commonly involving the cervical region, and con-
stitutional symptoms such as fever, night sweats, weight loss, and infec-
tions due to immunosuppression. Many of the pathologic and clinical fea-
tures reflect an abnormal immune response due to the wide variety of
cytokines and chemokines (Table 43.8), produced by the RS cells (Blood.
2002;99:4283). The etiology of CHL is largely unknown. Studies based
on the month of diagnosis have revealed peaks in February and March,
and the lymphoma appears to be more prevalent in adolescents and young
adults with higher socioeconomic status (Hematol On col. 2004;22: 11). The
association between EBV and CHL has been demonstrated in numerous
seroepidemiologic studies in which antibody titers to EBV and viral capsid
antigens have been consistently found to be higher in the lymphoma cases
compared with controls. There also have been reports of clustering of CHL
within the same family, especially among siblings of same sex and close age.
Certain human leukocyte antigen (HLA) types, such as Al, B5, and B18,
appear also to be clearly associated with this lymphoma (Hematol Oncol.
2004;22: 11).
B. Nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) is characterized by
scattered large neoplastic L&H cells (lymphocytic and/or histiocytic cells, pop-
corn cells) residing in nodular meshworks of follicular dendritic processes, filled
with nonneoplastic lymphocytes mostly of B-celllineage (e-Figs. 43.36 and
43.37). Clinically, patients present with localized peripheral lymphadenopa-
thy. The disease develops slowly, with fairly frequent relapses, but remains
responsive to chemotherapy and usually is not fatal. About 3% to 5% of cases
evolve into DLBCL.
C. Pathophysiology. The cellular origin of neoplastic cells in both CHL and
NLPHL was finally determined when clonally rearranged immunoglobulin
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infraclavicular; mediastinal; hilar; para-aortic; mesenteric; right pelvic; left

pelvic; right inguinofemoral; and left inguinofemoral.
2. Definition of extranodal involvement (such as by direct extension into lung,
thyroid, liver, etc.) is designated by an "E" alongside the anatomic stage.
Involvement of bone marrow, cerebrospinal fluid, liver, or pleura equates
to stage IV by convention.
B. Staging of multiple myeloma. The AJCC recommends the Durie-Salmon staging
system for multiple myeloma. This system takes into account various com-
plete blood count variables, blood chemistries, M-protein levels, and imaging
studies.
A newer, simpler, and more cost-effective alternative is the International
Staging System (ISS), previously the International Prognostic Index. This sys-
tem uses two simple blood parameters, specifically fh microglobulin (.82-M)
and albumin. The ISS has been proven to be useful for staging myeloma (] Clin
Oncol. 2005;23:3412) and is currently recommended for widespread use. A
comparison of these myeloma staging systems is depicted in Table 43.10.
Lymph Node Cytopathology

I. INTRODUCTION. Diagnosis of NHL by FNA requires a multiparametric approach

that couples cytomorphology with ancillary studies, including flow cytometry,
immunohistochemistry, FISH, cytogenetics, and molecular studies. Using a com-
bined approach, diagnosis of NHL based on FNA and core needle biopsy speci-
mens has >96% sensitivity and specificity when compared with excisional biopsy
(Am J of Clin Pathol. 2011;135:4). Proper triage of a lymph node FNA at
the time of immediate evaluation depends on the distinction between reactive
lymphadenopathy, a lymphomatous process, or a metastasis from a known or
unknown primary to direct subsequent distribution of tissue for appropriate stud-
ies (cell block, flow cytometry, FISH, etc.).
The lymphoid cells from FNA specimens characteristically are dyshesive, and
the background frequently shows lymphoglandular bodies (cytoplasmic fragments
of disrupted lymphocytes). In general, NHL exhibits a monotonous population
of atypical lymphoid cells in contrast to the polymorphous appearance of normal
and reactive lymph nodes. Cytologically, NHL is classified as small cell lymphoma,
intermediate cell lymphoma, and large cell lymphoma on the basis of the cell size
in comparison with histiocytes. Small cell lymphoma includes small lymphocytic
lymphoma, lymphoplasmacytic lymphoma, follicular lymphoma (grade 1), mantle
cell lymphoma, and marginal zone lymphoma. Intermediate cell lymphoma is fur-
ther divided into lymphoblastic lymphoma and Burkitt lymphoma. Large cell lym-
phoma usually refers to DLBCL, but also includes T-cell and NK-celllymphoma.
II. REACTIVE LYMPHADENOPATHY. The aspirate is characterized by a polymorphous
population of lymphoid cells with variable size and shape that predominantly con-
tains small lymphoid cells with smooth nuclear contours and coarse chromatin,
intermixed with scattered intermediate cells and large immunoblast-like cells. Scat-
tered plasma cells, histiocytes, and tingible body macrophages are present. Lym-
phohistiocytic aggregates are also seen, consisting of collections of lymphoid cells
and histiocytes held together by dendritic reticular cells (e-Fig. 43.38) (Acta Cytol.
1987;31: 8).
Clusters of neutrophils are indicative of suppurative lymphadenopathy. Collec-
tions of epithelioid histiocytes are consistent with granulomatous lymphadenitis
(e-Fig. 43.39) (Sarcoidosis. 1987;4:38) and may be seen in infectious and nonin-
fectious processes; special stains and culture are required for identification of fungi
and mycobacteria. Fungal hyphae and yeast forms can occasionally be identified
as negative images on Diff-Quik--stained smears (e-Fig. 43.40).
Ill. SMALL-CELL LYMPHOMA
A. Small lymphocytic lymphoma. The aspirate contains a monotonous population of
small lymphoid cells with round nuclear contours, characteristic checkerboard-
like clumpy chromatin, and indistinct nucleoli (e-Fig. 43.41). Scattered large
prolymphocyteslparaimmunoblasts with vesicular chromatin, prominent cen-
tral nucleoli, and pale cytoplasm are present.
B. Lymphoplasmacytic lymphoma. The smear contains a mixed population of small
lymphoid cells, plasmacytoid cells, and plasma cells.
C. Follicular lymphoma. The aspirate is composed predominantly of small centro-
cytes (that have irregular and grooved nuclear membranes) that are intermixed
with scattered centroblasts (that have a large noncleaved nuclei with multi-
ple small peripheral nucleoli) and immunoblasts (that have single prominent
central nucleolus) (e-Fig. 43.42). Lymphoid aggregates without abundant histi-
ocytes are not infrequently found (Cancer. 1999;87:216; Cancer. 2006;108:1).
Although the cytologic criteria for grading follicular lymphoma are not well
established, the percentage of centroblasts on smears has been proposed as a
basis for grading by several groups (Am] Clin Pathol. 1997;108:143; Clin
Pathol. 2002;117:880).
D. Mantle cell lymphoma. The aspirate is composed of a monotonous population
of small to intermediate-sized lymphoid cells with variable nuclear membrane
contour irregularity, dispersed chromatin, and inconspicuous nucleoli (e-Fig.
43.43) (Cancer. 1999;87:216).
E. Nodal marginal zone B-cell lymphoma. The aspirate contains a rather poly-
morphous lymphoid population composed predominately of small lymphoid
cells with round nuclei and clumpy chromatin, with scattered plasmacytoid
cells and immunoblast-like large cells (Diagn Cytopathol. 1999;20:190). Some
intermediate-sized lymphoid cells that have a monocytoid appearance and
exhibit a moderate amount of pale cytoplasm are also present (e-Fig. 43.44).
Lymphoepitheliallesions are not identified on smears. Due to its polymorphous
appearance, diagnosis based on the cytopathologic findings alone is challenging
without ancillary studies.
IV. INTERMEDIATE CELL LYMPHOMA
A. Lymphoblastic lymphoma. The aspirate is composed of a monotonous pop-
ulation of intermediate-sized lymphoid cells with fine granular chromatin,
irregular nuclear membrane contours, inconspicuous nucleoli, and scant
basophilic cytoplasm (e-Fig. 43.45). Mitoses are frequently identified (Acta
Cytol. 1992;36:887).
B. Burkitt lymphoma. The aspirate contains a monotonous population of
intermediate-sized lymphoid cells with round nuclei, finely dispersed chromatin,
and multiple distinct nucleoli. The cytoplasm is scant, deep blue, and contains
small lipid-filled vacuoles (e-Fig. 43.46). Tingible body macrophages and mito-
sis are prominent (Cytopathol. 1995;12:201) and are indicative of brisk cell
turnover.
V. LARGE CELL LYMPHOMA
A. DLBCL. The aspirate shows a monotonous population of large atypical lymphoid
cells. The centroblastic variant contains centroblasts with irregular nuclear
membrane contours, coarse chromatin, and multiple small nucleoli (e-Fig.
43.4 7). The immunoblastic variant shows immunoblasts with irregular nuclear
membrane contours, open chromatin, and single prominent nucleoli. The T-cell
rich B-celllymphoma variant is composed predominantly of small mature lym-
phocytes and scattered large immature lymphoid cells with polymorphic nuclei
and prominent nucleoli. The cytological diagnosis of this variant is challenging
(Diagn Cytopathol. 1998;18:1).
B. Peripheral T-cell lymphoma, unspecified type. The aspirate is polymorphic and

contains scattered large atypical lymphocytes with convoluted nuclear mem-
branes. The background shows a reactive pattern and is composed of small
mature lymphocytes, plasma cells, neutrophils, eosinophils, and macrophages.
Ancillary studies are necessary for diagnosis (Diagn Cytopathol. 2000;23:375).
C. Anaplastic large cell lymphoma. The aspirate shows both singly dispersed
and poorly cohesive groups of malignant cells with large and pleomorphic
nuclei, prominent nucleoli, and a moderate amount cytoplasm. Horseshoe-
and "donut" -shaped nuclei, as well as binucleated and multinucleated RS-like
tumor cells, are also present. Immunostains are necessary for diagnosis (Acta
Cytol. 1996;40:779). Mistaking this entity for carcinoma may be a diagnostic
pitfall due to the cellular cohesion and pleomorphism.
D. Myeloid sarcoma is an extramedullary solid collection of myeloid cells. The
disease may be present in a lymph node and may be a harbinger of leukemia or
evidence of relapsed disease. Aspirates show tumor cells in any stage of myeloid
differentiation ranging from blasts to more mature myeloid forms; however, in
some cases, myeloid differentiation may be entirely absent. The differential
diagnosis often includes large cell lymphoma; correct characterization rests on
histochemical and flow-cytometric analyses (Ann Diagn Pathol. 2000;4:1 ).
VI. HODGKIN LYMPHOMA. The aspirate of classic Hodgkin lymphoma contains rare
to scattered RS cells. The classic binucleated RS cells have markedly enlarged
nuclei, macronucleoli, and a moderate amount of basophilic cytoplasm (e-Fig.
43.48). The mononuclear variant has cells with markedly enlarged, irregular, and
multilobated nuclei with macronucleoli (e-Fig. 43.49). The characteristic reactive
lymphoid background includes mainly mature lymphocytes, scattered eosinophils,
neutrophils, plasma cells, and occasional epithelioid histiocytes (e-Fig. 43.49)
UClin Pathol. 1986;86:286). Diagnosis is challenging due to the scarcity of the RS
cells, and the cytopathologic diagnosis of NLPHL from FNA specimens is very dif-
ficult if not impossible. Nonetheless, when FNA biopsy is used in conjunction with
immunohistochemistry as a screening test for Hodgkin lymphoma, the approach
has a positive predictive value that has been reported to be >90% in some studies
(Cytopathology. 1994;5:226; Acta Cytol. 2001;45:300).
VII. METASTATIC MALIGNANCY. The most common indication for lymph node FNA
biopsy is metastatic malignancy. The reported sensitivity of diagnosis based on
FNA specimens in this clinical setting is >90%, with a specificity of >98%
(Cytopathology. 1996;15:382; Diagn Cytopathol. 2003;28:175). The aspirate
shows a nonlymphoid population of cells with malignant cytological features. An
absence of a background of lymphocytes and nodal elements is important to report
if it is uncertain whether the epithelial malignancy represents a nodal metastasis;
examples include breast cancers present in the axillary tail which may be mistaken
clinically for axillary nodal metastases, and hilar lung masses which may represent
central tumors versus metastatic involvement of hilar nodes. In the case of nodal
effacement by tumor, this distinction may not be possible.
In the case of a known primary, comparison with the primary malignancy usu-
ally will confirm the diagnosis. For patients with a history of multiple malignancies,
or cytomorphology that is not congruous with the reported history, immunostains
must be employed to confirm the site of origin. If clinical uncertainty is present
at the time of immediate evaluation, directing additional passes for cell block
preparation will increase the diagnostic yield of material available for immuno-
histochemical profiling.
SUGGESTED READINGS
Immunobiology, (6th ed.) New York and London: Garland Publishing; 2005:149-153.
Swerdlow SH, Campo E, Harris NL, et al., WHO Classification of Tumors; Pathology and Genetics;
Tumors of Hematopoietic and Lymphoid Tissues, 4th ed. Lyon, France: IARC Press; 2008.
Bone Marrow Pathology
Jeffery M. Klco and John L. Frater
I. NORMAL GROSS AND MICROSCOPIC ANATOMY. The bone marrow is generally consid-
ered the fourth largest organ in the human body and is composed of cells derived
from a variety of lineages including stromal cells, adipocytes, lymphocytes, and
hematopoietic precursors. The most frequently sampled areas are the posterior
superior iliac crest and, much less frequently, the sternum and long bones. The
bone marrow has an orderly microscopic anatomy. The most superficial part con-
sists of a layer of dense cortical bone with an adjacent cover of dense fibrous
periosteum. Deep into the cortex are the bony trabeculae, which consist of thin
trabecular bone and the marrow cavity itself. The marrow cavity contains islands
of maturing hematopoietic cells with intervening areas of fat, the latter of which
increase with age.
The cellularity of the bone marrow is defined as the percentage of the marrow
cavity composed of hematopoietic cells. In biopsies from the posterior iliac crest,
marrow cellularity decreases with age, and is expressed by the formula: Marrow
cellularity= (100- patient age)% ± 20%. Thus, a 50-year-old individual would
be expected to have a marrow cellularity of approximately (100- 50)% ± 20%,
or 30% to 70%.
Under normal circumstances, maturing myeloid and erythroid elements occupy
different regions of the marrow cavity. Myeloid precursors lie adjacent to the tra-
becular bone, and erythroid elements form "islands" of cells between trabeculae.
The ratio of myeloid to erythroid elements is roughly 2:1. Megakaryocytes are
irregularly distributed throughout the bone marrow. Under normal circumstances
they are not present in clusters.
It is important to be cognizant of the multidisciplinary nature of hematopathol-
ogy. Diagnoses in bone marrow pathology are not generally the product of mor-
phologic analysis of the bone marrow core biopsy or peripheral blood and bone
marrow aspirate smears alone. Clinical history is extremely important in sepa-
rating morphologically similar diseases and should be provided by the patient's
physicians. Also, the pertinent features of the physical examination, such as lym-
phadenopathy, splenomegaly, and hepatomegaly, are of importance. Other clinical
laboratory information, such as complete blood counts and serum and/or urine
protein electrophoresis, are often of interest. Radiographic data are of importance,
particularly in the evaluation of a monoclonal protein.
II. GROSS EXAMINATION AND TISSUE SAMPLING. The same pathologist should review
both the bone marrow aspirate smears and the core biopsies whenever possible to
avoid ambiguities or outright contradictions. The bone marrow aspirate is generally
performed before the biopsy. There are two kinds of aspirate smears: smears pre-
pared directly from the specimen without pretreatment and smears prepared from
concentrated aspirate fluid. Concentrated bone marrow aspirate smears and touch
preparations prepared from the bone marrow core biopsy are particularly useful
in the evaluation of specimens diluted with peripheral blood. Three to five smears
are stained using the Wright-Giemsa or similar technique, one is stained for iron
using the Prussian blue technique, and additional unstained smears are reserved
for ancillary techniques such as fluorescence in situ hybridization (FISH), if nec-
essary. Although examination of bone marrow cells with enzyme cytochemistry
is likely to be rendered obsolete in the coming years, its use is still highly recom-
mended by the World Health Organization (WHO) committee for the diagnosis of
709
acute myeloid leukemia, and high-quality aspirate smears should be reserved for
this purpose when an acute myelogenous leukemia is suspected. Additional bone
marrow aspirates are obtained for flow cytometric, cytogenetic, and/or molecular
genetic studies. Aspirate smears are generally reviewed using high power (600x to
1000x) and are important for evaluating individual cell detail. However, because
the process of aspiration disrupts cell cohesion, the relationship of the various cell
types and the marrow cellularity cannot be reliably assessed. An adequate bone
marrow core biopsy adds this important information.
Ill. DIAGNOSTIC FEATURES OF COMMON BENIGN DISEASES. The number and scope of
nonneoplastic bone marrow disorders are vast. Emphasis is given to commonly
encountered bone marrow diseases and conditions that may simulate neoplasia.
A. Megaloblastic anemia. It is often not necessary to perform a bone marrow biopsy
in patients who present with anemia, because the most common forms of anemia
(iron deficiency, megaloblastic, and anemia of chronic disease) may be diagnosed
by laboratory analysis of the peripheral blood, clinical history, and response to
iron, vitamin B12 , and/or folate replacement. Bone marrow biopsy is performed
for patients with anemia that is unexplained or therapeutically resistant.
It is important to note that megaloblastic anemia may simulate a neoplastic
condition, particularly a myelodysplastic syndrome. Patients with megaloblas-
tic anemia may present with marked cytopenias and pronounced dyspoiesis
(e-Figs. 44.1 and 44.2).* Clues to discriminate this benign condition from a
myelodysplastic syndrome include normal blast percentage, absence of kary-
otypic abnormalities, and improvement of cytopenias following administration
of vitamin B12/folate in megaloblastic anemia. Also, the degree of dyspoiesis in
megaloblastic anemia often exceeds that encountered in most cases of neoplastic
myelodysplasia.
B. Benign causes of lymphocytosis. Numerous benign conditions may cause a tran-
sient increase in benign lymphocytes in the peripheral blood and can simu-
late chronic lymphocytic leukemia/small lymphocytic lymphoma (CLUSLL),
T-celllarge granular lymphocytic leukemia, or other lymphoid leukemias. Stress
lymphocytosis is a transient increase in morphologically normal peripheral
blood lymphocytes encountered in individuals subjected to physiologic stresses,
including individuals presenting to hospital emergency departments. An abso-
lute increase in lymphocytes accompanied by "reactive" forms with increased
basophilic cytoplasm and inconspicuous nucleoli may be seen in patients infected
with Epstein-Barr virus (EBV, e.g., infectious mononucleosis) or, less commonly,
cytomegalovirus or other viral pathogens. Interestingly, in cases of EBV infec-
tion the virus particles are present in morphologically normal lymphocytes, and
the reactive lymphocytes represent cytotoxic T cells directed at the infected cells.
Correlation with serum viral antibody titers is useful for arriving at the correct
diagnosis and for avoiding unnecessary procedures such as bone marrow and
lymph node biopsies. Many other infections are associated with lymphocytosis,
including pertussis, in which the lymphocytes have characteristic clefted nuclei
and thus may simulate peripheral blood involvement by a non-Hodgkin lym-
phoma. Persistent polyclonal B-cell hyperplasia, as its name implies, is often of
longer duration than other benign forms of lymphocytes. It frequently affects
women who smoke, and is associated with the human leukocyte antigen (HLA)-
DR7 phenotype.
C. The granulocytic 111eft shift." Other cases of leukocytosis represent a granulocytic
shift to immaturity (colloquially known by the archaic term "left shift" that
refers to the traditional placement of immature myeloid cell percentages to the
left of neutrophils in classical peripheral blood smear reports). Because many
Chapter 44 • Bone Marrow Pathology I 7 11
of these cases also demonstrate eosinophilia and/or basophilia, it is important

to distinguish them from neoplastic conditions, in particular chronic myeloge-
nous leukemia or other myeloproliferative neoplasms. Most commonly, espe-
cially in hospitalized populations, a granulocytic shift to immaturity repre-
sents an acute response to bacterial or other infections. Transient increases in
mature demarginated neutrophils may also follow surgery or other physical
trauma. Unusual causes of increased peripheral blood neutrophils with or with-
out immature granulocytes include chronic idiopathic neutrophilia, hereditary
neutrophilia, and leukocyte adhesion factor deficiency.
D. Benign causes of erythrocytosis. Under normal conditions, the circulating red
blood cell mass is maintained at a constant level by the actions of the cytokine
erythropoietin, which is produced by renal peritubular cells. An absolute
increase in circulating red blood cells (polycythemia) may be primary (most
commonly due to the myeloproliferative neoplasm polycythemia vera) or sec-
ondary (due to increased production of erythropoietin). Thus, in establishing a
diagnosis of polycythemia vera, a number of conditions must be excluded that
may cause secondary erythrocytosis, including smoking, living in a high-altitude
environment, and high oxygen-affinity hemoglobins.
E. Benign causes of thrombocytosis. There are numerous causes of benign thrombo-
cytosis (defined as a peripheral blood platelet count in excess of 450 x 109/L).
Common reactive causes of peripheral thrombocytosis include childbirth, major
hemorrhage, iron deficiency anemia, chronic inflammatory conditions, infec-
tion, and acute stress events.
F. Aplastic anemia. Bone marrow aplasia is usually associated with hi- or pan-
cytopenia rather than isolated anemia, and may be identified in a variety of
clinical settings. It may occur secondary to a variety of drugs, most commonly
chemotherapeutic agents, benzene, alcohol, and arsenic. It may also occur sec-
ondary to exposure to radiation, viral infection (e.g., viral hepatitis, EBV), or
tuberculosis. An important cause of infection-mediated isolated anemia is infec-
tion with parvovirus B19. Other causes of bone marrow hypocellularity include
paroxysmal nocturnal hemoglobinuria, Fanconi anemia, dyskeratosis congeni-
tal, and other very rare inherited bone marrow failure syndromes (lnt] Hematol.
2002;76, Suppl1:207).
The common morphologic finding in bone marrow biopsies from patients
with aplastic anemia is marked panhypoplasia; a notable exception is parvovirus
infection in which there is selective suppression of erythroid precursors. These
specimens should be carefully scrutinized for evidence of significant dyspoiesis
or increased blasts because a minority of myelodysplastic syndromes and acute
myeloid leukemias present with markedly hypocellular bone marrow biopsies.
These specimens should also be examined for evidence of infection as evidenced
by granuloma formation in the case of tuberculosis, or intranuclear inclusions
in the case of viral infection. The inclusions of parvovirus are ill defined and are
localized to the nuclei of proerythroblasts and can be illuminated by immuno-
histochemistry against viral capsid proteins.
G. Serous degeneration (serous atrophy) is a pattern of bone marrow injury most
often associated with acquired immunodeficiency syndrome (AIDS) and states
of chronic nutritional deficiency such as starvation, chronic alcoholism, and
anorexia nervosa (Arch Pathol Lab Med. 1992;116:504). The primary morpho-
logic finding in the bone marrow is stromal edema with associated microvesicu-
lar change. The bone marrow is hypocellular in these regions due to loss of nor-
mal hematopoietic elements. The findings may be focal, alternating with regions
of relatively preserved normal hematopoiesis. It is important to recognize that
the subcortical bone marrow is normally hypocellular and occasionally demon-
strates edema, possibly related to trauma associated with the biopsy procedure,
and thus may mimic serous degeneration.
H. Granulomas may be encountered in bone marrow core biopsies and are occa-
sionally noted in aspirate smears. Granulomas may be quite subtle and are
usually composed of admixed histiocytes, lymphocytes, and plasma cells. Some
granulomas contain foci of necrosis and infiltrating neutrophils. The causes of
granuloma formation in the bone marrow are similar to those in other sites;
the most common etiologies are infectious, autoimmune, or idiopathic. In addi-
tion, granulomas are occasionally encountered in the bone marrow of patients
with Hodgkin lymphoma, although their presence is not indicative of marrow
involvement by disease. Because it is impossible to predict with certainty the
etiology of bone marrow granulomas, their presence usually warrants the use of
stains to aid in the identification of acid-fast bacilli or fungi. However, it should
be noted that special stains are far less sensitive than microbiologic culture in the
detection of most infectious agents in the bone marrow, so microbiologic anal-
ysis of fresh bone marrow tissue is recommended when an infectious etiology
is considered. An important item in the morphologic differential diagnosis of
granulomas is the lipogranuloma, which is generally considered to be nonpatho-
logic and consists of a collection of histiocytes and lymphocytes surrounding an
area of fat demonstrating microvesicular change.
I. Benign lymphoid aggregates are present in the bone marrow of healthy individ-
uals and at increased incidence in elderly individuals. Because they are com-
mon, it is important to recognize the attributes of benign aggregates and dis-
tinguish them from their malignant counterparts. Benign aggregates are often
well circumscribed and are small to medium sized. They are composed of an
admixture of cell types including small and mature-appearing lymphocytes (typ-
ically CD3+ T cells), histiocytes, and granulocytes. They frequently contain a
central small-caliber vessel. Although they occasionally abut bony trabeculae,
they do not demonstrate the paratrabecular pattern of growth seen in follicular
lymphoma and other non-Hodgkin lymphomas involving the marrow. In some
cases, immunohistochemistry or in situ hybridization forK and).. immunoglob-
ulin light chains can be used to further evaluate aggregates.
J. AIDS. Since the first cases of AIDS were reported in 1981, a number of associated
diseases have been identified in the bone marrow. Commonly encountered mor-
phologic changes in the bone marrow of human immunodeficiency virus (HIV)-
infected individuals include granulomas, lymphoid aggregates, plasma cell
aggregates, and dysplasia in one or more hematopoietic lineages (Haemophilia.
2001;7:47). Since the development of combined drug therapy, the incidence of
secondary infections has decreased. However, infectious agents are still encoun-
tered in the bone marrow of individuals infected with HIV and, because impaired
inflammatory responses are a hallmark of HIV infection, it is recommended that
all bone marrow biopsies from patients with HIV be examined with special stains
for fungi and mycobacteria.
Lymphoid aggregates occur with increased frequency in the bone marrow of
HIV-infected individuals, are sometimes large with ill-defined borders, and grow
along bony trabeculae; these are all features suggestive of malignancy. They may
be extremely difficult to evaluate, especially in view of the increased incidence
of non-Hodgkin lymphomas in this population. Ancillary studies may be useful
in the evaluation of monoclonal B-cell or phenotypically abnormal/monoclonal
T-cell populations.
Dyspoiesis is a common finding in the bone marrow of patients with
HIV infection and can be seen in other viral infections such as hepatitis C.
The significance of this finding may be difficult if not impossible to inter-
pret because of the increased incidence of neoplastic myelodysplasia and
acute leukemias in the HIV+ population. Blasts are not typically increased
in HIV-associated dysplasia, and clonal cytogenetic abnormalities are not
present.
K. Hemophagocytic syndrome is a potentially deadly condition in which cytokine-

stimulated benign histiocytes phagocytose other hematopoietic cells in an uncon-
trolled fashion. The most common causes of hemophagocytic syndrome are
related to activation of benign macrophages by cytokines produced by malig-
nant cells, and unregulated phagocytosis by macrophages following infection.
Malignancy-related hemophagocytic syndrome is most commonly associated
with peripheral T-celllymphoma, although other hematopoietic and lymphoid
malignancies are also rarely associated with this complication, including acute
myeloid leukemias with the translocation t( 16;21 )(p 11 ;q22 ). The most common
infectious cause of hemophagocytic syndrome is EBV. Regardless of the under-
lying cause, hemophagocytic syndrome presents with splenomegaly, fever, and
wasting. Pancytopenia, elevated liver function tests, and coagulopathy are vari-
ably present. Evaluation of the bone marrow aspirate and core biopsy reveals
variable numbers of macrophages containing phagocytosed hematopoietic cells
(Am] Surg Pathol. 2001;25:865).
L. Chediak-Higashi syndrome is an autosomal recessive inherited disorder that is
caused by a mutation in the CHS1-LYST gene located at chromosome 1q42. The
clinical features of this syndrome are related to abnormal lysosomal trafficking
and include recurrent infection, oculocutaneous albinism, neurologic disorders,
and a bleeding diathesis. Granulocytes, monocytes, and lymphocytes contain
abnormal large granules derived from secondary or cytotoxic granules (e-Fig.
44.3) (Platelets. 1998;9:21 ).
M. Mucopolysaccharidoses. Alder-Reilly anomaly is identified in the granulocytes
of patients with a group of uncommon diseases characterized by X-linked or
autosomal recessive transmitted defects in the enzymes involved in mucopolysac-
charide metabolism (Table 44.1A) (Clin Lab Haematol. 1996;18:39). Granulo-
cytes in the peripheral blood and bone marrow have coarse azurophilic gran-
ules superficially resembling normal primary granules. Classification of cases of
mucopolysaccharidosis requires chemical and/or molecular analysis.
N. Lipid storage disorders. There are numerous genetically mediated conditions
related to defects in enzymes comprising the pathway of lipid metabolism (Table
44.1B). The most commonly encountered are Gaucher disease and Niemann-
Pick disease. Gaucher disease demonstrates an autosomal recessive pattern of
inheritance and is caused by a defect in the enzyme ,8-glucocerebrosidase. Clin-
ically, patients present with bone pain and splenomegaly related to the prolifer-
ation of morphologically abnormal histiocytes at these sites. The cytoplasm of
the macrophages has a "wrinkled tissue paper" appearance which represents
the accumulation of glucocerebroside in these cells (e-Fig. 44.4). Occa-
sional cases are associated with B-lineage malignancies (including plasma cell
dyscrasias) and light chain amyloidosis UIntern Med. 1999;246:587).
Patients with Niemann-Pick disease typically present with organomegaly,
neuropathy, and abnormal laboratory findings similar to those identified in
Gaucher disease. The pathophysiology of Niemann-Pick disease is related to
autosomal recessively inherited defects in the enzyme sphingomyelinase, with
the presence of sphingomyelin in affected cells. Macrophages in this disorder
have been described as "sea-blue" due to the cytoplasmic accumulation of PAS-
positive material representing sphingomyelin (Ann Hematol. 2001;80:620).
The remaining lipid storage diseases, which are somewhat less common, have
clinical presentations similar to those of Gaucher and Niemann-Pick diseases.
Some are associated with additional findings. For example, Hermansky-Pudlak
syndrome is associated with platelet storage pool deficiencies and oculocuta-
neous albinism (Platelets. 1998;9:21).
IV. DIAGNOSTIC FEATURES OF MALIGNANCIES
A. Myelodysplastic syndromes are clonal hematopoietic disorders characterized in
most cases by peripheral cytopenias (hemoglobin <10 gldL, absolute neutrophil
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Detection of a cytogenetic abnormality is helpful, although the majority of

cases of "low-grade" myelodysplasia (refractory cytopenia with unilineage dys-
plasia [i.e., refractory anemia], refractory anemia with ringed sideroblasts, and
refractory cytopenia with multilineage dysplasia) have normal karyotypes. Com-
mon cytogenetic abnormalities include partial or complete deletions of chromo-
somes 5 and 7, del(20q), and/or trisomy of chromosome 8. Abnormalities of
chromosome 7 or complex karyotypes (>3 cytogenetic abnormalities) carry a
poor prognosis. 5q-syndrome is a special type of myelodysplastic syndrome in
which dyspoiesis is most prominent in the megakaryocytic series, accompanied
by thrombocytosis; this form of myelodysplasia is associated with long survival.
"High-grade" myelodysplasia (refractory anemia with excess blasts types
1 and 2) presents with a greater percentage of peripheral blood/bone marrow
blasts compared to low-grade cases. In general, refractory anemia with excess
blasts is more clinically aggressive than that with or without ringed siderob-
lasts, and has a variable propensity for progression to acute leukemia or bone
marrow failure, both of which are essentially untreatable by any means short
of a bone marrow transplant. Outcome in the myelodysplastic syndromes may
be predicted using the International Prognostic Scoring System (IPSS), which
takes into account blast percentage, karyotype, and the presence of cytopenias
(Leukemia. 2005; 19:2223).
B. Myeloproliferative neoplasms. The myeloproliferative neoplasms (formerly
chronic myeloproliferative disorders) are characterized by an expansion of one
or more of the hematopoietic lineages as evidenced by increased bone marrow
cellularity and increased circulating white blood cells (usually granulocytes),
erythrocytes, and/or platelets. Initially, blasts are present in normal to slightly
increased numbers in the marrow. Although these are malignant disorders, the
affected cellline(s) are usually morphologically normal, and pronounced dys-
poiesis is not a characteristic feature of the chronic myeloproliferative disor-
ders. In contrast to myelodysplasia, organomegaly (splenomegaly and/or hep-
atomegaly) is a common feature of this disease, and becomes more pronounced
with disease progression as the bone marrow becomes dysfunctional and/or
fibrotic.
Particularly in their early stages, it may be difficult to distinguish these dis-
eases from reactive conditions affecting the marrow. Also, because the various
myeloproliferative neoplasms have overlapping morphologic features, distin-
guishing between them may be very difficult. However, it is important to rec-
ognize and distinguish between the various members of this family of disorders
because the different malignancies have variable propensities for high-grade
(i.e., acute leukemic) transformation and bone marrow failure. Accordingly,
the identification of disease-specific molecular markers in this family of dis-
eases is the subject of intense scrutiny, and important molecular lesions have
been identified. The first, BCR-ABL, has been known for many years, and is
largely limited to chronic myelogenous leukemia and occasional cases of acute
myeloid leukemia, precursor B-lymphoblastic leukemia/lymphoma, and biphe-
notypic acute leukemia, at least some cases of which likely represent blast phases
of clinically silent chronic myelogenous leukemia.
BCR-ABL is most frequently encountered as the classic Philadelphia chro-
mosome, t(9;22)(q34;q11), which juxtaposes the break cluster region gene
(BCR) at chromosome 22q11 with the Abelson tyrosine kinase gene (ABL)
at chromosome 9q34. The resultant BCR-ABL fusion protein is a constitutively
activated tyrosine kinase directly responsible for the manifestations of disease
in chronic myelogenous leukemia. In chronic myelogenous leukemia, BCR-ABL
is typically of the 210 kd (p210) form, compared with BCR-ABL in precursor
B-lymphoblastic leukemia/lymphoma which is generally 190 kd (p190). The
p230 (230 kd) form of BCR-ABL is identified in cases of chronic myelogenous
leukemia with a predominance of mature neutrophils rather than immature

granulocytes. Previously, cases of chronic myeloproliferative disorders with the
p230 form of BCR-ABL and neutrophilia were classified as chronic neutrophilic
leukemia, but the WHO classification now recommends that such cases be clas-
sified as chronic myelogenous leukemia.
More recently, point mutations have been identified in Philadelphia
chromosome-negative myeloproliferative neoplasms, the most prominent of
which is the ]AK2 V617F point mutation involving the Janus tyrosine kinase 2
(Am Soc Hematol Educ Program. 2006;240). Other mutations have recently
been identified in several genes, including myeloproliferative leukemia virus
(MPL), TET oncogene family member 2 (TET2), additional sex combs-like
1 (ASXLl), casitas B-lineage lymphoma proto-oncogene (CBL), and the lym-
phocyte adaptor protein LNK, although none of these mutations appear to be
specific for myeloproliferative neoplasms.
1. Chronic myelogenous leukemia is a well-characterized clinicopathologic entity
because essentially all cases have the characteristic Philadelphia chromo-
some t(9;22)(q34;q11) and corresponding rearrangement of BCR-ABL (Br
] Haematol. 1991;79, Suppl 1:34). This is a disorder derived from clonal
expansion of an abnormal pluripotential hematopoietic stem cell. Affected
individuals typically present with nonspecific constitutional symptoms and
on physical examination are frequently found to have an enlarged spleen.
Patients most often present in the chronic phase characterized by a periph-
eral leukocytosis composed of granulocytes (in various stages of maturation)
with peripheral and bone marrow basophilia and eosinophilia (e-Fig. 44.9).
The blast percentage in chronic phase is usually <2% of white blood cells
in the peripheral blood and <5% in the bone marrow. Untreated, cases of
chronic myelogenous leukemia invariably acquire additional genetic lesions
(i.e., additional Ph chromosome, isochromosome 17q, or trisomy 8) result-
ing in a maturation arrest in the malignant population and terminate in an
acute leukemic (blast) phase. The blast phenotype is myeloid in ""80% of
cases and lymphoid in most of the remaining cases, and has a biphenotypic
or ambiguous phenotype in rare individuals. Chronic myelogenous leukemia
in blast phase is essentially untreatable by any means short of a bone mar-
row transplant. Increasingly, patients presenting with chronic myelogenous
leukemia in chronic phase are treated with small molecular inhibitors such as
imatinib mesylate, a tyrosine kinase inhibitor with a high degree of specificity
for the BCR-ABL-encoded tyrosine kinase. However, an increasing problem
is the development of resistance to imatinib mesylate, for which second- and
third-generation tyrosine kinase inhibitors have been developed.
2. Polycythemia vera is a clonal stem cell disorder in which the majority of dis-
ease manifestations are related to expansion of the cells of the erythroid lin-
eage as evidenced by increased hematocrit, blood volume, and blood viscos-
ity (Blood. 2002;100:4272). The clinical features of this disease are related
to this phenomenon. The skin has a plethoric appearance and there is an
increased propensity for vascular thromboses, hemorrhage, and central ner-
vous system phenomena including stroke, tinnitus, headache, and vertigo.
The peripheral blood manifestations of disease are related to expansion of
red blood cell mass. There is an increased red blood cell count (approximately
7,000,000 to 10,000,000/mm3 ) and hemoglobin (usually> 18.5 gldL in men
and> 16.5 gldL in women) without a corresponding increased reticulocyte
count. Because polycythemia vera is related to expansion of a pluripoten-
tial hematopoietic stem cell, other cell lines are variably affected. Thus, some
patients have an associated neutrophilic leukocytosis and/or thrombocytosis.
Other clinical laboratory features of disease include an increased leukocyte
alkaline phosphatase (LAP) score and increased serum vitamin B12 levels, the
latter due to an increase in transcobalamin 1. Bone marrow analysis is per-

formed to exclude other forms of myeloproliferative disorder and to assess
baseline fibrosis. Usually, initial bone marrow analysis reveals increased bone
marrow cellularity with multilineage expansion of hematopoiesis and mini-
mal fibrosis.
In establishing a diagnosis of polycythemia vera, nonneoplastic erythro-
cytosis and erythrocytosis due to cytokine production by nonhematopoietic
malignancies must be excluded. Common causes of secondary erythrocytosis
include pathologic conditions (such as chronic pulmonary disease and smok-
ing) and physiologic conditions (such as living in high-altitude locations). A
JAK2 mutation (most commonly V617F but mutations in exon 12 have also
been identified) is found in the vast majority of cases and thus is now a major
criterion for diagnosis.
In most cases, polycythemia vera is a clinically indolent condition. The
treatment of choice for most patients is periodic phlebotomy to minimize the
risks associated with increased blood viscosity. Less than 10% of patients
develop bone marrow failure or acute leukemia.
3. Essential thrombocythemia is a clonal neoplasm derived from a pluripotential
hematopoietic stem cell in which the majority of clinical and pathologic fea-
tures are related to morphologically and physiologically abnormal megakary-
ocytes and platelets (Haematologica. 1999;84:17). There is usually a marked
peripheral thrombocytosis, generally in excess of 1,000 x 109/L, although a
sustained count of >450 x 109/Lis sufficient for diagnosis. The platelets are
frequently morphologically abnormal, including forms with decreased gran-
ularity. The most prominent characteristic of the bone marrow is marked
megakaryocytic hyperplasia. The megakaryocytes are frequently morpholog-
ically abnormal, with large overall size and hyperlobate (staghorn) nuclei. A
definite diagnosis can be established after other myeloproliferative neoplasms
are eliminated from consideration with ancillary studies. JAK2 V617F muta-
tions are found in "'50% of cases (both of essential thrombocythemia and
primary myelofibrosis). The major pathophysiologic consequences of the
thrombocytosis and proliferation of megakaryocytes are episodic bleeding
and thrombosis, which are major causes of morbidity and mortality. Less
than 1% of cases progress to acute leukemia.
4. Primary myelofibrosis (chronic idiopathic myelofibrosis, agnogenic myeloid
metaplasia, myelofibrosis with myeloid metaplasia) is another myeloprolif-
erative neoplasm attributed to transformation of a pluripotential hematopoi-
etic stem cell. Classically, there are two phases in the natural history of
disease (Hematol Oncol Clin North Am. 2003;17:1211). The first phase,
referred to as the cellular phase, is characterized by a marked expansion
of all hematopoietic lineages; it is followed by the spent phase, character-
ized by progressive bone marrow failure and fibrosis. Because the cellular
phase is often clinically silent, the majority of clinical findings are related
to increasing bone marrow fibrosis, and include fatigue (due to anemia),
bleeding (due to thrombocytopenia), and infection (due to granulocytope-
nia). Hepatosplenomegaly due to extramedullary hematopoiesis is a common
physical finding.
The cellular phase is often characterized by peripheral granulocytic
leukocytosis and/or thrombocytosis. The former is sometimes accompa-
nied by eosinophilia or basophilia clinically mimicking chronic myelogenous
leukemia. In the cellular phase, the bone marrow is hypercellular due to a
panhyperplasia of all hematopoietic lineages. Megakaryocytes cluster in the
bone marrow and are characteristically highly atypical including large forms
with bulbous/hyperchromatic nuclei. Fibrosis is minimal at this stage of dis-
ease. With time, the bone marrow becomes increasingly fibrotic (which is
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(i.e., with blast percentages of at least 20%) and are broadly categorized as
myeloid or lymphoid according to their immunologic and enzyme cytochemical
properties, the characteristic features of which are summarized in Tables 44.5
and 44.6. There are two important exceptions to the requirement of at least 20%
blasts in acute myeloid leukemia. In the WHO classification, the presence of
t(8;21)(q22;q22) [AML1-ETO], inv(16)(p13;q22)/t(16;16)(p13;q22) [CBFfJ-
MYH11], and t(15;17)(q22;q12) [PML-RARa] is classified as acute myeloid
leukemia regardless of the percentage of blasts (e-Figs. 44.13 to 44.17). The
second exception involves acute erythroid leukemia, for which blasts comprise
at least 20% of the nonerythroid cell population, rather than at least 20% of
all cells; because another requirement for acute erythroid leukemia is that at
least 50% of all cells are erythroid precursors, cases with very high erythroid
percentages may qualify as acute leukemia even in the presence of a very modest
blast percentage. Immunophenotypic analysis of acute leukemia is discussed in
Chapter43.
Acute leukemias, particularly in children and the elderly, may present with
isolated anemia and a lack of circulating blasts. Diagnosis requires detection
of at least 20% blasts in the bone marrow and/or the presence of an acute
myeloid leukemia-specific cytogenetic abnormality. Although replacement of
the bone marrow by fibrosis, carcinoma, sarcoma, or other nonhematopoietic
cell may result in anemia, it is generally accompanied by leukopenia and/or
thrombocytopenia reflecting indiscriminate displacement of normal marrow
constituents.
1. B-lymphoblastic leukemia is arbitrarily separated from its tissue analogue,
lymphoblastic lymphoma, by the presence of a tissue mass and :::;20% bone
marrow blasts in the latter (e-Fig. 44.18) and is no longer subdivided on
the basis of immunophenotype since genetic features are more predictive of
outcome. B-lymphoblastic leukemia is most common in children <6 years
old; patients with purely lymphomatous disease are somewhat younger on
average. Most cases express HLA-DR, terminal deoxynucleotidyl transferase
(TdT), CD10, CD19, CD24, and cytoplasmic CD79a. CD10-negative cases
often demonstrate rearrangements of the mixed-lineage leukemia (MLL)
gene and coexpress one or more myeloid-associated antigens. In the pediatric
population, cytogenetic and molecular genetic findings are highly predictive
of disease outcome. Predictors of good outcome include the following: age 4
to 10 years at diagnosis, hyperdiploidy (51 to 65 chromosomes) in the blast
population, and presence of translocation t(12;21)(p13;q22) [TEL-AML1].
Predictors of poor outcome include the following: age <4 years or > 10
years, hypodiploidy, t(9;22)(q34;q11.2) [BCR-ABL], t(4;11 )(q21;q23) [AF4-
MLL], and t(1;19)(q23;q13.3) [PBX-E2A]. The prognostic significance of
t(1;19) is controversial. Recently, mutations/deletions in the lymphoid tran-
scription factor gene IKZF1 (IKAROS) have been shown to be associated
with a high rate of leukemic relapse and a poor outcome. Mutations in PAXS
are also common.
A special category of lymphoblastic leukemia is the leukemic analogue
of Burkitt lymphoma. Patients with this disease commonly present with
an abdominal mass (Western Europe and North America) or a jaw lesion
(Africa). The bone marrow is commonly involved, and the blasts have deeply
basophilic cytoplasm with many lipid vacuoles, express CD10 and mono-
clonal surface immunoglobulin light chain, and are generally CD34 and TdT
negative (e-Fig. 44.19). Diagnosis requires demonstration of the transloca-
tion t(8;14)(q24;q32) [MYC-IGH] or, less commonly, the variant transloca-
tions t(2;8)(q11;q24) or t(8;22)(q24;q11) involving MYC and the K and A.
light chain genes, respectively.
.....
,. DilliN
A._.....,. _ ..... CIIIICII
,.lll .... lllc ... ___
-MIDJf .....,....,.
k>bln'f'llold 1111/loltrole Mil Often - M i l Blu1* Mllloi'C lllander J\uar CD~. CD$+, MPO+. F""'r«blo
t!S;21l!<!22;<1221: G«<011-..IIIY<hlo.10 roda. «bl'lOITMI.croniiCicll C()~,CD$4+,C056+
(RIJNXUIUNXlTCJ
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-lfllslrw(l6)(pl3q22) .. lobotlon CDII~.CDllo+.C~
ttl&;lli)(pl3:q:i2) C036+.~
rtlBF,IIrl'tHllJ
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CAloR 10!1111(15;17!(<122;ql2);
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(l!y_..,u~rVINJil;
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hl!tl
F~ca:~nln dllldren
m\Oifpl!o A"" rod& ptedomlnd&
lllo,.,.,UC - fi'OIIOmN11
(lr1.!H), HLA-DR-, COS4-
V•lltbl& cota .,d c~.

CD4+. all4+. CDII~.
.._ ...
"'"""
1(9;11)
..,._,....
CDlie+, C064+, elm+•
k>bln'f'llold 1111/loltniD Mil P~ll B&aoPIIIIlllOith mulllllneo.li& CD~. CD$+, ~

1(6,9)~:q341; DEK-tiUP214 rrryojo~. HLA-OR+.
k . . .,.lold 1111/l:lmll Mil N"'"""l'olorobld ~ <01111

""'*""'
,.,. ._ . . . . .,. lndudlrw
CO:W...,C0117+, Tc!T±
CD~, C~, HLA-DR+, Poor
ln.Q)(q21,q26.2) or 1(3;3)
(~; q26.2); Rffll-E'ill
- c~r~........,..pololls CD34+, C0117+, CD411
CD61±
k*~nY<~Iold 1111/loltniD Mil lnfamul'd l'OIII&<Iilkhll: Mleiw)'oi)IIIJCic CD411C061+. CD~. CDU+-. ~~~-
tti:22KPI3;QU~ hof)O)IJDMo"-lflo C()S4-, HlA-DR-
IISiollSMU(l <011\MI"
.__..,..... -·--~~-..lool·
I. fdlowii'C a ln'f'l~luCio oYIIdrom& Cl.ltl$l or ~ld>'rn~~~~dklordor
2. I.IDS-nolobd e,t:el- •lln-ltj
!. l.lull ....... d)OI)i&IM ~I&IM In . t - SO'l!. ofh-ln!oo> eollll,_,
e. ..................... .,.. ,......,.., ,tiiiC.., lilllltlwl
• AI~ ...rt...llled; typlco!ly hu• >10,... lotlnq!wttl\ toMOS ....
• ~~II _,hlbloHalaD:I (tome ma:f ba tlfnphold); 1~ lllbio<;ywttl\ no t-MDS phooo. """'""""""' ba~,_,,....-,.ln<:lud~
l!Q~•tmm.llm
l._.....,...
• OIIwMM
._laNII-CII:Ji•IDII
Aclto rnyolotllolloltr:le UollltJ - I n d:.lil>ood, -dlt ol ~ MPO+. dlt o! Ofton CD13+. co~. CD117+. u,.,.,.,bl&
mblmatJcltiW•:d:llad cytoc,oilll:& bhbto NBE+ C())4+, Cll31k-, HLA-DR+
Ai:lb rnyolotllo- - I t UollltJ p-In ad :.~~hood, ..mpr~sa ~ o1
s~~.m Ol!on C013+, co~. C014+, Ullk..,nlblo
IN1:ntlcn qtopolniM, - ! I f..til no!101)11mll "'*; :!:!!!'. ol C0117+, MPO+
mlfk:lcl.,.lna : cf W'8C blull MPG+, ,;;?."of blalll
NBI".t
N.bi!I)'O!iotlla-""" Vorllblo""r'"'P•rol ~ ol bl:lsls MPO+. ~ ol !Ju:llfCD13+, CD~. COlli+; Vllllablo
mav0!11on ~I!IO~io&Y bhbto NBE+: >l'"' ml:tll1rc Y>i:l::bD CD117+, CD34+,
~t~enu~cel& HlMlR+
Ai:lb rnyolom~ IMJI<.eml! AMml!, r-. ~,wac >20!i.blodtO..,....rc U.U.IfCDI3+, CDS3+; 01lan Vuleblo
....I t t - prornoncqte.s); ==~ CD4+, C014+, COil b+,
monocytes. aI'd PMCUJ'SOI'I; CD!l'*' C064+, C0!6+,
llJid ~ noulropl:llo and tfiOZlmO+
P"""'"""" !:3l!'. ol biMI>
MPO+. !:3!1'. o l - "'""!If
NBE-+"
Aclto m~ il<lk.eml! Moot...,mon lndikhn.., ~ ~C>IIIh. olwl:lell V•:tellD CD13+. Cll33+. U~l&
p---"""*'IMy
d~ l:l:!edlrc d~son~
...
~~ arw; mono~ ....::20%
-pi:!~ and PlliCU/liOI'Ii
CD117+; Ol!on C014+,
CD4+, COlli>+, COlle+,
<3~ oT blab MPO+, !!3~ of CD64+, C06&t, CD36+,
bl:lm NBE+ ~
N.bm~l!umlo -common ln...,b. ollan ~~ ..lb. olwl:tl: V•rlablo COI3+. Cll33+. Unf-ru>lo
p-wtll\-tnll(lllfliiY 111o mo)o:l!yer& 1>10~ C0117+: Ofton CD14+,
dr-,l:l:!edlrcd~am~ ... <20% noWophlund CD4+, COlli>+, COlle..,
.... prea.ll'II:I/Bj ~of blot&

MPG+, ?:>1" of blalll NilE+
CD64+, COQI+, CD36+,
~
,_
........ IW'!I:J;~!:II --af-lliiJIIDid........,llo~l
Ao.... cwylllrold - ... Adulb:•nomla
..
~f.O'I. <Jf erth nl.daretDd ~-.,..~A+ Unfm>robb
Cs/yll\roid/rn'f<llotl) rx>o*lcllll eM!r'okl and ~nd hetiiORlO!i~A+:
~~ln'f<l~-. ln'f<l~ m COI3+.
"""'1)111'* poi>&Mllono ~"
ofblula ny be MPO+
0033+, COI17+, ~nd MPO+
P&nll)1lruld lou- >80!(; of colt nlmmllu"' Blula.,. . , m _flljtojlliofln U,_.blo
flciJ •• ""....
e1)11lrold celll;.., ollnlllcont A+ •rd ,..,.,.,.,bin
At;
m)Oioblostcom"""""" <31!'.
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Aoib m~ lellk.emll ~penI!.&

NBE+
fl!ol>lulle~
- - -qtoplamlc
r••noloporl&. A!>nOtn'lll
ploColob encl """'"'(YQCJ~o
Uo~CD41+, CD61+;
....,lialalyCDI*, Cllil3+;
coa4-, CD45-, HlkOR-
-
11118mem&ln 1>0/\0henll blood:
uo~ ~of bkm MPO+
erd >!I% ofbl&ab NBE+
Ao• bMolitllc lou- Ve/yi'OI['I) -·l&~ldf/11) Iiiii>+; UI~CDI3+, CW3+. CC$4+, DIII:Ut'>
IBUitj dll. of blu1s MPO+, HlA-DR+,C~ predk:t
<ll"ofblull NBE+ diaiD law
n\I'Tiblf rR
rvpo!1od
00.1<10.
ProbeillY
Aoib P"""~"'"* with

nr,'lla!lbtolll
Vfly ,.,... ildullo, ~~~
will nolm-.m.~ lfllo""""''P''.r
Poi'ohype~J>iua, ~pilotle
........I)OX)1ols, lncrusall
.-.unfll>mlo
C013+, C039+, C0117+,
MPO+; tom&CII.Jilllrt t:q~tWt
Cllyillrold"' m~
•rllpona
- poor
MPO, myola_.., WBC,w!l;to-

-nt~Wcr1d Heelt\ O(JIII'Ii:2:irti:l\ •to.. h
'*'""" NB£, ~,., tart,rlta- HIJI. nu_, ieu-diJII\
ClilliUIB Of aoAe tn'JEID'nOnc:ae,tie llklk:I!Wnia ill\ th&~ Of NBE ~I th& ~I! tl'liEISt ~ tdleti& b' {'(r(JJNX.1*-•
Fta'l'l: s.atib.r$, Cl!ltl'f,O E. Hetri&. Nl.4lt:M., ads. WHO Oflss·~ ttl T~d~lfftd l.ymp/'1(1idT--.I.Jarte IARC P1'81!1e.; 2DQS.. Lll!a:Jwit\ j!atl'lillliol\.
NPI.Il (Nueleojli\CIImln)• 25-3!'.'11.
Clllclll
1'1~do ,.,.,.......,., hi!tl wac
C,...lllll:e
Froquorily ...,.,
a•c••
F...,.bl&
ond ln)Oib'nonoqllc: mol)ll\ol:lf.y
cunmon; blolb ~ C034-
C€BPA (CCMTienhl""" bfld~ 1>-IOlr. H\lh I'll blost""'nt: ~l<:oly AML Frequorily nonnal f ......blo
~...phi). ~It !Mtfdon
I'I7).1T!lli'M$-Ib~ 3)-31)'11. Hl.ll\ WBC: WJied rn~ VAJ1ed Po«
klrwe-Momolllndom duplootlan
/OHl/10112(-A~ I I~HI: -1110; IDH2: ...a'll.

.......,
(lndlll~-~
ljplcatjAML- Typlco!lfnonnol(~!lyca:.n TBD

""".u...
-
ti\12)C mllturel:kln; uncommon In 11'0111!tSomwtil
"''"'""'lie·- -'lo• 1lT.l!)
C/NIIt1"311 {I)NA "'~"' 3ol ~,..
tommon In myolom~ Nonnal ~- (BdiOII''"''
...,~ .. fOOd ""' """mont....-
elmnnelltl AM\.01
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7£7:1 (TET .,._tomtj mombef 2) -7'11. No d~de!lnk:oll p o - ~!lfnonnol(~cee~n TBD
IOith .u...
11'0111!tSomwtil
-'loniVHmW!tlm)
•ou....m.-... litel-ln AMLincUie c-«<T, H-IlA$. K-IIAS. WTJ, fl(fl{ll.f n1 pe11111- dupl:llion of NU, "'"" m -
1':"1""*tll)l!l:o""' 1111!diD l>e cldmnh:d •IIIII" cohO!tof AML pe1len1o. '"'root""""'10rANL end-
Atulel1l!<i:ll:lleu""""'""'- -lerd C<lii'Ae"'p"""-...- In WI«) (20011.
'IDHJ,/D/12., D - end m:~...-.. h..,. eloo been litel-lndllet m)'dold mel[tnoncb.
~1n>m.ICC.20ll;a47S-486:-~Il:ll&~ .
....
2. T-lymphoblastic leukemia/lymphoma is less common and involves an older

demographic than its B-cell counterpart. Patients typically present with abun-
dant blasts in the peripheral blood and a mediastinal mass. The blasts
are generally TdT positive and demonstrate variable expression of other
T-lineage antigens, most commonly CD3 and CD7. Genetic aberrations iden-
tified in this entity include translocations between TCR{j and TCR~ genes
and a variety of partners; microdeletions of TAL1; activating mutations in
NOTCH; and del(9q), which results in deletion of the tumor suppressor gene
CDKN2A. Although T-lymphoblastic malignancies have historically had a
poor prognosis, recent innovations in treatment have improved outcome.
E. CLLISLL and related disorders. In patients (particularly elderly individuals) with
unexplained lymphocytosis, an absolute increase in lymphocytes may repre-
sent peripheral blood involvement by low-grade non-Hodgkin lymphoma or
leukemia such as CLUSLL (Table 44.7). For most non-Hodgkin lymphomas,
diagnosis is made following biopsy of an involved lymph node or extramedullary
focus of disease, and bone marrow biopsy is performed for staging rather than
precise classification. An exception to this is CLUSLL, in which primary diagno-
sis is often made following flow cytometric analysis of the peripheral blood and
demonstration of the characteristic immunophenotype (CDS+, CDlO-, CD19
bright+, CD20 heterogeneously+, CD23+, FMC7-, and CD79b-).
Various non-Hodgkin lymphomas have different patterns of marrow
involvement. Because B-celllymphomas are more common than T-cell tumors,
the former are better characterized. For example, follicular lymphoma and
mantle cell lymphoma have a paratrabecular pattern of marrow involvement,
whereas CLUSLL is never paratrabecular (e-Fig. 44.20 ). In the case of CLUSLL,
the pattern of involvement of the marrow may be predictive of prognosis: cases
with predominantly focal lesions are more indolent, whereas examples with
a diffuse pattern of involvement are more aggressive. As in lymph nodes, the
bone marrow infiltrate of CLUSLL may contain proliferation (or growth) cen-
ters that represent aggregates of prolymphocytes (paraimmunoblasts) and pre-
sumably represent the proliferative component of the tumor mass (e-Figs. 44.21
and 44.22). Marginal zone lymphomas may demonstrate follicular colonization
when they occur in bone marrows with lymphoid aggregates. Overall, the like-
lihood of marrow involvement by non-Hodgkin lymphoma is highly variable
depending on type, ranging from very common (e.g., CLUSLL) to rare (e.g.,
extranodal marginal zone lymphoma). Immunohistochemistry is typically not
required in non-Hodgkin lymphoma staging biopsies, although it is occasionally
helpful in delineating benign from malignant lymphoid aggregates.
F. Hairy cell leukemia is a rare form of chronic leukemia. Most commonly, the
affected individual is a middle-aged man presenting with pancytopenia, includ-
ing lymphopenia and monocytopenia, and an enlarged spleen. Occasionally,
patients present with a leukemic blood picture mimicking CLL. The bone mar-
row is virtually always involved. Although identified in bone marrow aspirate
preparations, hairy cells are more easily identified in the peripheral blood by
the presence of cytoplasmic projections (hence the name "hairy cell leukemia,"
e-Fig. 44.23 ). The malignant cells frequently have an interstitial pattern of
involvement of the bone marrow and for this reason are occasionally over-
looked. Sometimes the pattern of marrow involvement recapitulates the pattern
of splenic involvement by this malignancy with collections of extravasated red
blood cells surrounded by ill-defined collections of hairy cells (e-Figs. 44.24 and
44.25). The malignant cells are positive with tartrate-resistant acid phosphatase
(TRAP) enzyme cytochemistry. Flow cytometry identifies a characteristic pat-
tern of reactivity: the hairy cells are CDS, CDlO, and CD23 negative, but pos-
itive for the pan B-cell antigens CD19 and CD20. In addition, they typically
demonstrate bright coexpression of CDllc and CD25, and are also positive for
......,mill- lla,.M.,..
llllllnIll pill! 1:1
DlleiA ""'lllii7PI lllll]liiiiiDD ~1:1 Cll&llftca.lcll I~DII-IM)
CIVIl'* limPho<:Yiic lilllcetnle/ 00&+-,CI>IG-, COl~!~. U!Jwlt anoil 11\AUH Pleo!lt\l: 13QI4 d e - Cfowonlblf) ~ tA Ill htlaW
srnlll tympllae)1i:
tymph>INI
CD20+ (dm), c~.
FM07-, C0'/91>-, siJIWI!
<lllln+ (dim)
tymphoq!M; dlftWa,
IIU~ or""""~' .-,n&
of morrow 1-monl c-
a&llo»mo),+12
(mOfplioicP:alf~l,
unfi'IIIWtl'-ot.*::ornl),
,.,.m.-
<llolln Mbl&
(1•"'"'bit)
Cllilll± (u-.,blo H+1, ponlnobculllf) 17p (ll'S3) clo-
Z.p70 :1: (unf...,rlll>lo H+l (111fm><oblol
Mal'llo ... b'rnPIIoiNI CO&+-, CI>IO-, COl~. C02!>+ U.Witanoll lim~""" ~ 11;14l t8CU -Iaiii
(b!W\1l, Cl>lt)-, FMC7+, cleftodll)ldad ~ dtllloa.
Cl77il>+-, siJIWI! dlah+ 1\ttdi.Mr. lrihw~ or
(WaleIf bo11J11!, qcln Dl+ ~beo::ollu pol!Mno a!
MlmlW-
Fol...,r tymph>mo CD!;-, CI>IO+, COIH, C020+, VIll1oblo "'"""''l'holoilr; dl'fllll, ~14;18) fJlCL2o/GHI
COZI-, '" 111111 <"'*'+ nodiAar, 1~ or
ponrtnobo<:<llllf palbifm a!
M«JJ'CW~
MarltlllZIII» lk>ell COh Cl>lo-, COI9+. C020+, Smlllll)m~. -with ~ll;l8l(Q21:<121l
tymph>INI COl!!-, '"llgttehMI+ omplo~ !API2..uou:rn lclon1Hitd 1n
·-<tf-...,11
""'"*"'"""" tympmm•
LJmplql~ C06-,CI>lll-, COIH.C020+, Smaii1Jm~,....,- t!9:141(ql3;1j32)
tymph>moiW<Itlomlnlm COZ3-,I- illlht<"'*'+ plosm~ lill1lnl ~~ tloi'AIIod ••
~I>Uihomll ~tA-bla II not
llmbd 101llk""" a1
""'"'"'""'
..... tiJ, _,...,.......,....lin; II. hmu-lln.
F"'"': -lowS. C.. E. lllnto Nl.dtL, ods. WHO_, of liRr>ol.l'odHo""'fojootlloM!d ljql>p/lold-., ~ IAJ\C -lllOII. Uoohlh pomhom
CD103 and FMC7 (Semin Oncol. 1998;25:6). Identification of this pattern of

reactivity in the presence of appropriate cytomorphology essentially excludes
other types of B-cell neoplasia, such as splenic marginal zone lymphoma and
prolymphocytic lymphoma. Although hairy cell leukemia is largely resistant to
conventional chemotherapies, it is sensitive to purine analogs (i.e., cladribine)
and is associated with a prolonged median survival.
SUGGESTED READINGS
Foucar K. Bone Marrow Pathology. Chicago, IL: ASCP Press; 2001.
Jaffee ES, Harris NL, Stein H, et al. World Health Organization Classification of Tumors. Pathology
and Genetics of Tumors of Haematopoietic and Lymphoid Tissues. Lyon, France: IARC Press;
2008.
Knowles DM, ed. Neoplastic Hematopathology, 2nd ed. Philadelphia, PA: Lippincott Williams &
Wilkins; 2001.
Spleen
Mohammad 0. Hussaini and Anjum Hassan
I. NORMAL GROSS AND MICROSCOPIC ANATOMY. The spleen is the largest lymphatic
organ. In a normal adult, it weighs 50 to 250 g. Anatomically, the spleen is divided
into white and red pulp, separated by an ill-defined interface known as the marginal
zone (e-Fig. 45.1A).* For a schematic of splenic architecture, see Figure 45.1.
A. White pulp. The white pulp consists of periarteriolar lymphoid sheets (PALS),
which contains lymphoid follicles. T lymphocytes are predominately located in
periarteriolar lymphoid nodules and B lymphocytes are predominately located
in the lymphoid follicles. The latter may contain germinal centers that become
visible to the naked eye when enlarged, forming splenic nodules (malpighian
corpuscles). In routine hematoxylin and eosin (H&E)-stained sections, the white
pulp appears basophilic due to the dense heterochromatin in lymphocyte nuclei
(e-Fig. 45.1B).
B. Red pulp. The red pulp has a red appearance in fresh specimens and in histologic
sections because it contains a large number of red blood cells (e-Fig. 45.1B).
It consists of splenic sinuses separated by splenic cords (the cords of Billroth),
which are composed of a loose network of reticular cells and fibers with a large
number of erythrocytes, macrophages, lymphocytes, plasma cells, and granu-
locytes. Special endothelial cells that express both endothelial and histiocytic
markers (known as Littoral cells) line the sinuses. The sinusoidal lining epithe-
lium is discontinuous, allowing for transport of blood cells between the splenic
cords and sinuses.
A. Biopsy and fine needle aspiration cytology. These procedures are rarely attempted
because of the risk of hemorrhage and the likelihood of undersampling. How-
ever, some studies suggest increased chances of a definitive diagnosis when fine
needle biopsy is combined with flow cytometry, with an overall accuracy of 91%
and a major complication rate of< 1% (Am] Hematol. 2001;67:93 ).
B. Splenectomy. Trauma, staging procedures, and surgical convenience account
for >50% of all splenectomies. Therapeutic splenectomy for known diagnoses
(idiopathic thrombocytopenic purpura [ITP], chronic myeloproliferative dis-
orders, lymphomas, etc.) accounts for most of the remaining cases (Cancer.
2001;91:2001). Unexpected pathology is rarely found in splenectomy speci-
mens, but significant splenomegaly (weight >300 g) or localizing lesions warrant
careful prosection and ancillary studies.
C. Processing. The spleen is weighed, and its outer dimensions are recorded. Hilar
fatty tissue is removed and processed for lymph nodes. The capsule should be
described, noting texture and intactness. The spleen should be thinly sliced (every
2 to 3 mm); lesional distribution should be noted, followed by a description of
the uninvolved spleen. Sections of any lesions (preferably following overnight
formalin fixation of thin slices) and two representative sections of uninvolved
spleen should be submitted for microscopic examination.
If clinically indicated, fresh lesional tissue in 1 mm pieces should be placed
in RPMI medium and directed immediately to the flow cytometry lab with
instructions regarding the appropriate protocol. Cytogenetic studies may be
729
sc - .
FT- F
TA-l
TV-.
LN -I
PALS
WP-
RP - I
S- Si
CB - '
Figure 45.1 Diagram of the spleen showing important anatomic landmarks and&- and T-cell
distribution.
useful, especially for diagnosis of hematologic malignancies; using sterile tech-

nique, lesional material should be procured immediately after removal of the
spleen in the operating room and directed to the cytogenetics lab. Samples can
also be frozen, or fixed for electron microscopy. Freezing preserves many of
the antigens for immunohistochemical evaluation in hematopoietic malignan-
cies and enhances nucleic acid recovery for DNA- and RNA-based molecular
diagnostic techniques (although most diagnostic molecular tests can be reliably
performed on formalin fixed, paraffin embedded material).
Ill. GENERAL CONSIDERATIONS
A. Splenomegaly. Often the spleen becomes enlarged due to infectious causes or
congestive states. Red pulp congestion is frequently observed and is the most
common finding in such cases (Table 45.1).
B. Hypersplenism. Hypersplenism refers to destruction of one or more blood cell
lines by the spleen (Eur] Gastroenterol Hepatol. 2001;13:317). It is the most
important indication for elective splenectomy. Diagnostic criteria for hyper-
splenism include cytopenia of one or more blood cell lines, bone marrow
I. I -
A. lnfodblo611docordlll
8. lnftd:but rnonon~
C. Tubotcal-
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Ill. H.,..,..,... molllnon<Y
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A. CoriJIInbl dllmln<llll)lllroqlos (llndla!yl!>h~ ell-.~;
ho""'P>bl"'pcllb, 0.1.. - c o l d - llllbllllo hllf110I)oblnl)
B. Ac~~uhd lkltderoot ·~~ !olttommLI\0 hlllnCIMI:•n........, molllllo. bobeobhl
c.
Auldmmune11u~~lo. UidJ<lf ,_.,""""'
11. Alln..,.l '11\een OIJ.IIIntaqu-.n of """"II blood eoll!;
A. Dllmlnot lilt monoq1a/lnlla0jlhqo aystem (ell rank:~. """"""d-..,
ponulllc: lnfeciSonl, LCH, ell:.)
B. Moll,lnorrt lnllllr- dllcrdtn (loula>mlln, t~mpllomu, plalml cdl dp:toolat,
r-..: Cllcincmol
C. EX!runedLllaiYhornotoi>Oieolo <-•
helno!l1!o...,., ohrank: l:lloiollllb lll)lliollbrool!l
1>. Clu1>nle ln-..,1\r-mp"" tubore<Jiadl, ,_Ia
E. -lillloo•oalabnonm- (-.lubunM, polboll, l!llonlc: eysb, hamii!Dnw)
Ill. Ma..cllsneot.acondii:Sonl
A. HJpoojl!fl"ldhm
B. ~rne.P>IIuln"'*
c. Pl-t;e rnu.,..llllldooonOIICih•I>OCiti
'JXf I SECTION x.. HEYATQPQIEliC SY5TEU
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II. Acquired
A. ~lot-IOCiorrry
8. AoQulredmDh:t•hdA?rl'lfaldlon
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6. -lm11'01no d - .
6. lmodlodoll
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II. Cllronlo elcollollam
9. Hypcpi:LitoNm
C. Funcllonlhooplenlo will n..,.l..illoclor tnllrpd SjliMn
I. lnft!IR1!onlly loulo!mla.l!mplloma. mulllplo II1)'<Qnll, mli--jll>ll>
2. Elrlf!ol>bnomoplc)-... d -
s.~
4.So~
S. &rii!Jund md.i!l1111t -~r1>1mo11
6. Mlllblorpllon synd,..,.
D. Ooproooadmmunefll1cl!on
I. AIDS
2. S111U41>001
b. '**""" _,_,.1"1
•• llt&dlo1ion
c.lmmunoou-lwlqenll,ln-na-rold&
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b. liYI>®IIIIbJtorn
•. Die- rrwlltuo
-4. Cllronlo elcollollam
Chapter 45 • Spleen I 7 33
Usually benign incidental findings, their clinical importance lies in their potential
to mimic neoplastic and nonneoplastic splenic lesions.
IV. REACTIVE SPLENIC DISORDERS. These can be divided into diffuse and localized pro-
cesses. Diffuse disease entities include reactive lymphoid hyperplasia (e-Fig. 45.7A),
follicular hyperplasia, and disorders such as Castleman disease (see Chap. 43 ).
Localized disease includes granulomatous disorders and infectious processes simi-
lar to those in other locations (see Chap. 43 ).
A. Diffuse reactive processes. Reactive lymphoid hyperplasia in the spleen may
occur with or without germinal center formation. Because of the reactive nature
of the latter, and its histologic lack of maturing germinal centers, this entity is
variably referred to as "activated A," "early activated immune reaction," "reac-
tive nonfollicular hyperplasia," and even "immunoblastic hyperplasia" (Am ]
Surg Pathol. 1981;5:551). Nongerminal center hyperplasia is often associated
with viral infections, especially herpes simplex virus and Epstein-Barr virus
(e-Fig. 45.7B), which explains the common occurrence of splenomegaly in
patients with infectious mononucleosis.
Reactive lymphoid hyperplasia with germinal center formation (e-Fig.
45.7C) is commonly referred to as "follicular" hyperplasia. It is the most com-
mon pattern of lymphoid hyperplasia in the spleen and is seen in both acute
and chronic immune reactions. Follicular hyperplasia is frequently observed in
bacterial infection, often as an incidental finding. In fact, splenomegaly is char-
acteristic in subacute bacterial endocarditis, alerting the clinician to this process
in the appropriate clinical context.
B. Focal reactive processes. Localized reactive splenic processes can also present
with splenomegaly.
1. Granulomas. The most common form of a focal benign process is granu-
lomatous inflammation, ranging from lipogranulomatous inflammation (of
unknown etiology) to caseating or noncaseating granulomatous inflamma-
tion. Caseating granulomas are primarily due to infectious disease, including
tuberculosis and fungal infection; however, they are also seen in X-linked
chronic granulomatous disease. Noncaseating granulomatous disease is most
frequently associated with sarcoidosis (e-Fig. 45.8). For most granulomas
in the spleen, no known etiology can be found (Arch Pathol Lab Med.
1974;98:261 ).
2. Infarcts. The spleen is a frequent site of systemic emboli, which commonly
arise from cardiac valve lesions or mural thrombi. Infarcts are usually wedge-
shaped with a hemorrhagic to pale-tan to fibrotic appearance depending
on the age of lesion (see e-Fig. 45.9). Non-wedge-shaped infarcts arise in a
variety of intrinsic hematopoietic and nonhematopoietic processes. Essential
thrombocythemia and chronic idiopathic myelofibrosis are the hematopoi-
etic disorders most frequently associated with infarcts; less common causes
include paroxysmal nocturnal hemoglobinuria, sickle cell disease, and aplas-
tic anemia. Among nonhematopoietic etiologies, vasculitides (e.g., polyarteri-
tis nodosa, infections, and TTP/ITP associated) and splenic artery aneurysms
are common culprits.
V. NEOPLASTIC DISORDERS OF THE SPLEEN
A. Lymphoid neoplasms. Lymphoid neoplasms can involve the spleen as primary
disease or as a part of a generalized lymphomatous process. Splenomegaly is a
nonspecific but classic component of many hematolymphoid disorders. A brief
approach to the evaluation of the normal and neoplastic components of the
spleen is presented in Table 45.4.
1. Primary splenic lymphomas. These account for <1% of all lymphomas and
can of B- or T-cell origin (see Chap. 43). Not unexpectedly, B-celllymphomas
are more common, and diffuse large B-celllymphoma (which usually presents
as single or multiple circumscribed nodules of varying sizes) accounts for the
ClonoraJ Guld&IN&lO Ewluota
lhostran.l Ccmpollmont
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ean-lltoo•- of
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lnlhofollcliD.
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"'""'"II<Y be oubtYI>od? clloracUnze t>llenottl>6of""llouo llmrilorn .. (Ooo•loo
ClloP. ol3, T®loo o13.3 01\d o13.4l
s.mr. er.mp~oooro In -~ dll«dool ..til lallll"""mpon.,.. of ,.,uld\&
wllll Pnodomnontly Rod col& h - mon><ocl..- red pulp 1.-mert.
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bl Hapodooplorrlc r....u
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~. C04-. CO!!-, ollan Cllli6+.
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COI58
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ftJ' P"""""" tJrnplrolclloulc.emtat: Tdt, COi'!lt, CDS, COlO
vast majority of cases (e-Fig. 45.10). The diagnostic approach to primary

splenic lymphomas is identical to lymphomas presenting elsewhere
(see Chap. 43 ).
2. Secondary splenic lymphomas
a. Hodgkin lymphoma {HL). The spleen is the most common extranodal
site of HL, both classic (e-Fig. 45.11A) and lymphocyte-predominant
Hodgkin disease (LPHD), although the latter is extremely rare (Cancer.
1971;27:1277; Cancer. 1987;59:99). Diagnostic Reed-Sternberg cells or
variants (e-Fig. 45 .11B and C) are a requirementfor the diagnosis of classic
HL, especially in cases without previously documented history. Immuno-
histochemical evaluation is often very helpful in the differential diagnosis
of classic HL versus LPHD.
b. Non-Hodgkin lymphomas. More than 50% of cases of low-grade lym-
phomas show splenic involvement either in the form of splenomegaly or
splenic hilar lymph node involvement. Splenic involvement can be cate-
gorized as focal or diffuse, the latter mode being more common. Usually
there is initial expansion of white pulp in a nodular pattern in cases of low
grade B-celllymphomas (e.g., small lymphocytic, mantle cell, follicular,
or marginal zone) eventually evolving into a diffuse pattern. Both pat-
terns are often observed in appropriately sampled splenectomy specimens.
Intermediate and high-grade lymphomas (large cell, Burkitt, lymphoblas-
tic) tend to form single or multiple tumor masses. General histopathologic
considerations for the diagnosis of these lymphomas are similar to those
occurring in lymph nodes (see Chap. 43 ).
3. Lymphomas presenting with prominent splenomegaly
a. Splenic marginal zone B-celllymphoma {SMZL). SMZL is an indolent dis-
ease, and splenectomy results in long-term remission (Semin Diagn Pathol.
2003;20:83). SMZLis sometimes accompanied by autoimmune thrombo-
cytopenia or anemia, and circulating villous lymphocytes (with polar pro-
jections) are sometimes seen in peripheral blood. Cytogenetically, allelic
loss of chromosome 7q31-32 has been described in up to 40% of cases,
and complex karyotypic abnormalities appear to carry a worse prognosis
(Discov Med. 2010;10:79; Blood. 2010;116:9).
Histologically, expanded periarteriolar lymphoid sheets are seen, com-
posed of small lymphocytes and monocytoid lymphocytes (e-Fig. 45.12).
Immunophenotypic studies by flow cytometry should demonstrate a
clonal B-cell process (surface light chain restriction), usually lacking coex-
pression of CDS, CD10, and CD23. Alternatively, immunohistochemistry
can be performed (to document a B-cell phenotype and coexpression of
CD43) coupled with in situ hybridization studies (for kappa and lambda
light chains); the latter is usually helpful in highlighting the clonal plasma
cell population, which forms part of the spectrum of B-cell differentiation
in marginal zone B-celllymphomas.
b. Hepatosplenic T-celllymphoma {HSTL). This rare lymphoma has a clinically
aggressive course and usually afflicts young males. It is characterized by a
triad of peripheral cytopenias (anemia and thrombocytopenia), sinusoidal
tropism, and hepatosplenomegaly. The disease is somewhat more common
in immunosuppressed settings, such as post solid organ transplantation,
and splenomegaly may exceed 3000 g. The neoplastic process is based
on the red pulp, with conspicuous infiltration of sinuses (e-Fig. 45.13).
The differential diagnosis includes other red pulp-based diseases such as
hairy-cellleukemia. Immunophenotypic and genetic studies demonstrate a
clonal population ofT cells, often double negative for CD4 and CDS, with
the majority showing T-cell receptor y8 gene rearrangements. Isochromo-
some 7q10 and trisomy 8 are the strongest genetic associations (Nat Rev
Gastroenterol Hepatol. 2009;6:433 ).
c. Mantle cell lymphoma (MCL}. In mantle cell lymphoma, prominent

splenomegaly usually represents the leukemic phase or stage III or N
disease. Consequently, the morphologic pattern of involvement can be dif-
fuse, nodular, or both (e-Fig. 45.14 ). Splenic involvement may occur in the
absence of significant lymphadenopathy (Virchows Arch. 2000;437:591).
The usual constellation of morphologic, immunophenotypic, and cytoge-
netic findings is required for diagnosis (see Chap. 43).
d. Hairy cell leukemia (HCL}. HCL classically presents with peripheral cytope-
nias, particularly monocytopenia (Leuk Lymphoma. 1994;13:307), and
splenomegaly in a young male with recurrent opportunistic infections (Am
J Clin Pathol. 1977;67:415). In the spleen, HCL involves and expands the
red pulp; the white pulp is usually inconspicuous (e-Fig. 45.15). The clas-
sic immunophenotype by flow cytometry (CD103+, CD11c+, CD25+)
can be easily demonstrated utilizing peripheral blood in the presence of
circulating "hairy cells" and is required for diagnosis. This immunophe-
notype is also helpful in distinguishing hairy cell leukemia from HSTL,
T-large granular lymphocyte lymphoma (T-LGL), other T-cell neoplasms,
and mast cell disease, all of which can morphologically mimic hairy cell
leukemia in spleen. By immunohistochemistry, DBA 44, Annexin A1, and
cyclin D1 stains are helpful (Lancet. 2004;363:1869; Hematol Oncol Clin
North Am. 2006;20:1051). No common recurrent cytogenetic abnor-
mality is specific, however, numerical abnormalities of chromosomes 5
and 7 have been reported (Hematol Oncol Clin North Am. 2006;20:
1011).
4. Other B-and T-celllymphomas. T-prolymphocytic leukemia and T-LGL com-
monly involve the spleen (see Chap. 43) as they are usually leukemic at pre-
sentation. Likewise B- cell lymphomas, presenting at stage III or N (see Chap.
43) can also involve spleen. An example of stage IllES follicular lymphoma
involving spleen is shown in e-Figure 45.16.
B. Myeloid neoplasms
1. Chronic myelogenous leukemia (CML}. CML is classically associated with
splenomegaly. The spleen is also the most common extranodal site of involve-
ment in the blast phase of CML (see e-Fig. 45.17). Morphologic findings
leading to the diagnosis of CML are best evaluated in touch preparations,
although histologic sections are also easy to interpret. Touch preparations
are required for optimal enumeration of blasts. Immunohistochemical stains
(CD34, c-kit) and cytochemical stains (Leder, myeloperoxidase) can also be
useful in highlighting the blast population. Disease progression (accelerated
phase) and transformation in CML is usually obvious both clinically and
morphologically.
2. Acute leukemias. Acute leukemias present as diffuse involvement of the red
pulp (e-Fig. 45.18). The spleen is rarely a primary site of myeloid disease, and
involvement reflects systemic disease. Histopathologic and immunopheno-
typic considerations are similar to acute leukemias presenting with peripheral
blood and bone marrow involvement (see Chap. 44). Evaluation of spleen
specimens for commonly occurring cytogenetic abnormalities, by conven-
tional cytogenetics or fluorescence in situ hybridization (FISH), is a standard
part of the work up unless there is already a prior history of leukemia. Care
must be taken to appropriately evaluate for therapy-related morphologic and
phenotypic changes and clonal evolution that may alter treatment course or
effectiveness for targeted therapies.
3. Mast cell disease (MSD} andlor systemic mastocytosis (SM}. These myelo-
proliferative neoplasms frequently involve the spleen. The morphologic
patterns of involvement vary from isolated white pulp accentuation with
fibrosis, to red pulp involvement with diffuse infiltration, fibrosis, and/or
nodular perivascular infiltrates. The presence of eosinophils, plasma cells,

and fibrosis are all clues that point to the presence of mast cells. Flow
cytometry is often not helpful for mast cell disease primarily because of
technical difficulties in gating the desired population, although expression
of CD2 and CD25 by flow cytometry is a feature specific to neoplastic mast
cells (both benign and neoplastic mast cells are CD45+, CD33+, CD68+,
and CD117+ ). In tissue sections, Leder stain (naphthol ASD chloroacetate
esterase), CD117, and tryptase are helpful in highlighting mast cells.
C. Nonhematopoietic neoplasms and pseudoneoplasms. A wide variety of mesenchy-
mal cell types form the complex reticular support network of splenic pulp and
consequently a wide variety of mesenchymal tumors can occur as primary splenic
neoplasms. These can generally be divided into stromal lesions, vascular lesions,
and tumor-like lesions.
1. Stromal lesions
a. Dendritic cell tumors. Two different kinds of dendritic cells exist in the
normal lymphoid support network: interdigitating dendritic cells (IDCs)
which are normally S-100 protein and MHC ll positive, and follicular
dendritic cells (FDCs) which express CD35 and CD21. Splenic involve-
ment can be seen in neoplastic disorders of either IDC and FDC (Can-
cer. 1997;79:294; Am I Surg Pathol. 2002;26:530). Grossly, involvement
is usually nodular, although disseminated systemic disease may present
with diffuse splenic involvement. Dendritic cell tumors tend to behave in
an aggressive manner despite their bland histologic appearance. In the
absence of a preceding history, dendritic cell tumors are diagnoses of
exclusion, mandating a thorough immunophenotypic workup to exclude
myeloid malignancies, lymphoid B- and T-cell malignancies, and non-
hematopoietic malignancies. It is important to note that FDC neoplasia
may be associated with the hyaline vascular type of Castleman disease
(Adv Anat Pathol. 2009;16:236).
b. Histiocytic lesions. These lesions range from Langerhans cell histiocytosis
(LCH) to histiocytic sarcomas including Langerhans-cell sarcoma. Splenic
involvement is rare, usually occurring in the setting of disseminated disease
and grossly presenting as single or multiple solid nodules.
Expression of S-100 and CD1a by the neoplastic cells is consistent
with a diagnosis of LCH. The sarcomatous forms can be less differentiated
and may variably show expression of HLA-DR, CD45, CD68, PLAP, and
vimentin (Am I Surg Pathol. 2004;8: 1133 ).
2. Vascular lesions. Vascular tumors are common in the spleen given its rich
vascular framework. Both benign and malignant vascular lesions may present
in the spleen. (Am I Surg Pathol. 1997;21:827).
a. Benign lesions
i. Littoral cell angioma is unique in its presentation in the spleen and
grossly is characterized by multiple spongy, cystic nodules. The cys-
tic spaces are lined by cuboidal epithelium with intracytoplasmic
eosinophilic globules. The lumina often contain abundant desqua-
mated cells. Vascular markers (CD31, Factor VITI related antigen)
are characteristically expressed; expression of CD68 and CD21 is
more variable. CD34, commonly expressed in normal sinusoids, is
uniformly negative.
ii. Peliosis, characterized by ectatic sinusoids and blood-filled cysts, can
involve the spleen. The location of the cysts (adjacent to PALS and fol-
licles) is helpful in establishing the diagnosis. The clinical importance
of this lesion lies in its propensity to undergo spontaneous rupture.
iii. Hemangiomas are a frequent incidental finding at splenectomy (e-Fig.
45.19).
iv. Sclerosing angiomatoid nodular transformation {SANT} is a rare nonneo-

plastic lesion characterized by angiomatoid nodules surrounded by
sclerotic stroma and a lymphoplasmacytic infiltrate (e-Fig. 45.20). It
should be differentiated from vascular neoplasms of the spleen and
lymphomas showing lymphoplasmacytic differentiation (Am ] Surg
Pathol. 2004;28:1268).
b. Malignant lesions
i. Littoral cell hemangioendothelioma and angiosarcomas rarely present in
spleen (Am] Surg Pathol. 2006;30:1036). They are usually solid, often
prompting a differential diagnosis that includes other spindle cell sar-
comas. CD31 and Factor VIII related antigen immunostains establish
the vascular nature of these otherwise undifferentiated malignancies;
some cases may also show CD34 expression. A translocation involv-
ing chromosomes 1 and 3 has been reported in a few cytogenetically
analyzed hemangioendotheliomas (Am] Surg Pathol. 2001;25:684).
Complex cytogenetic abnormalities, none consistent from case to
case, have been reported in angiosarcomas (Cancer Genet Cytogenet.
1993;63:171; Cancer Genet Cytogenet. 1998;100:52; Cancer Genet
Cytogenet. 2001;129:64).
ii. Kaposi sarcoma, in the setting of HIV/AIDS, must be considered in the
differential diagnosis of any splenic vascular lesion. Kaposi sarcoma
usually shows positive immunohistochemistry for HHV-8 and a variety
of vascular markers.
3. Pseudoneoplastic lesions. Examples include splenic hamartoma (well-
circumscribed lesions with an angiomatoid lobular-nodular configuration,
resembling red pulp; usually CDS+ and CD68+ ), splenic cysts (with or with-
out an epithelial cell lining; when an epithelial lining is present it is usually
cytokeratin positive; e-Fig. 45.21), angiomyolipoma (focally HMB-45 posi-
tive), and lymphangioma.
Inflammatory pseudotumor is a reactive nodular process that shows a
predominance of benign inflammatory cells and stromal cells with sclero-
sis (e-Fig. 45.22). Inflammatory pseudotumor must be distinguished from
inflammatory pseudotumor-like FDC tumor (which is usually EBV-associated
and shows immunohistochemical expression of CD21 and CD35) and lym-
phomas with lymphoplasmacytic differentiation (MZL and LPL, see Chap.
43).
4. Metastatic tumors. A variety of carcinomas and sarcomas can metastasize to
the spleen, although the lack of afferent lymphatics renders the spleen gener-
ally less amenable to metastatic disease. Metastases therefore commonly arise
in the setting of disseminated disease. The most common epithelial metastatic
tumors are carcinomas of breast or lung origin. Sarcomas involving the spleen
tend to be of dendritic/histiocytic or vascular lineage.
5. Other neoplasms. Benign fibromas, osteomas, and chondromas can also occur
in the spleen.
SUGGESTED READINGS
Bowdler AJ. The Complete Spleen. 2nd ed. Towota, NJ: Humana Press; 2001.
Neiman RS, Orazi A, eds. Disorders of Spleen. 2nd ed. Philadelphia: W.B. Saunders; 1999.
Rosati S, Frizzera G. Pseudoneoplastic lesions ofhematolymphoid system. In: Wick MR, Humphrey
PA, Ritter JH, eds. Pathology of Pseudoneoplastic Lesions. New York: Lippincott-Raven;
1997:449.
Swerdlow SH, Campo E, Harris NL, et al. WHO Classification of Tumors and Haematopoietic
and Lymphoid Tissues. Lyon: IARC; 2008.
SECTION XI
Soft Tissue and Bone
Soft Tissue
John D. Pfeifer and Louis P. Dehner
If the term "soft tissue" were restricted to only mesodermally derived structures, then
nerves and neural tumors would not be considered in the discussion. Similarly, there
are other examples of "soft tissue tumors (SITs)" which are present in organs but
may not be derived from mesoderm. By convention, however, neural tumors, gastroin-
testinal stromal tumm; melanoma of soft parts, perivascular epithelioid cell tumor,
and so on are regarded as "soft tissue neoplasms" despite the absence of any evidence
of a well-characterized progenitor cell or mesenchymal stem cell (MSC, characterized
immunophenotypically by reactivity for CD73, CD90, and CD105) with the multipo-
tentiality to differentiate into a variety of tissues such as muscle, blood vessels, and fat
(Stem Cells. 2011;29:397;] Pediatr Hematol Oncol. 2008;30:301 ).
I. TISSUE PROCESSING
A. Biopsy specimens. lncisional or core biopsy of a suspected soft tissue neoplasm
is performed to determine the appropriate management based on the patho-
logic type. The biopsy tissue should be placed immediately into 10% forma-
lin or other appropriate fixative. The number of biopsy fragments should be
recorded, as well as their aggregate dimension, and all the submitted tissue
should be processed. Three H&E levels should initially be prepared for micro-
scopic examination. For very small specimens, in order to avoid wasting tissue
when refacing the block, it is strongly recommended that additional unstained
slides be cut from the block during initial sectioning in the event additional
studies such as immunohistochemistry are needed.
B. Resection specimens. Excisional specimens are often complex and varied, and
the macroscopic examination should be guided by tumor location, extent,
and type. The margin of all intact specimens should be inked, and the gross
distance from the tumor to the closest margin documented. The maximum
dimension of the tumor should be recorded, as well as the color and consistency
of the cut surface, and presence of hemorrhage and necrosis. In general, it is
recommended that one section per centimeter of tumor should be submitted for
microscopic examination (scout sections can be used to evaluate whether such
thorough sampling is required for definitive diagnosis). The closest surgical
margin should be evaluated by either shave or radial sections depending on
the nature of the specimen.
For those tumors in which a biopsy did not permit definitive diagnosis,
tissue should be collected and processed for electron microscopy. Considera-
tion should always be given to the need to send a sample of viable tumor for
739
'JCO I SECTION .... SOFT TISSUE AND BONE
I' I
1,1,! I (!-I\ ~~~~uc MHwlltlet a-c~e~~.uc of"'lllooa l1olt
Tl.a r~!~Pol ._lilt •IIIIIIIIIG a lad flriGIMol
_ , rhlbdoiT(jOMn:oma 1(2;13)(q36;q14) PAXJ.FOXOll'wlon
10;13l[l>36;ql4) I'AX7-ffl'(!)ll'wlon
-rdDI!ItWOOitlll d..O7ltOC; 17)(~11.2;<!2.Sl MPI.-17'D Mlon
Alljjoln.mld tbii>UI: t02;22Kqllt\ql2) EWSRZ..ATI'l1\.u!lan
hldSoq!Dma 1(2;22)~;q12) EWI/Sl.ciiEBl
102;16Kql'%qlll FUI'.41l'll'llllon
Al)lbll_.,niiiiDus ~l"ff\um""'Y nns•ndlor Am~ of MOM2.
tumw/ul :lff&ui!UI1Dd m-<ttrar.OIIOIIIOS CDI(4,H/116112p8t
.-....,...,. W.TNroLPSl lOith ampl~- of
12ql4-ql5
C!Mr oelun:orn• t02;22Kqllt\ql2l EWSRZ..ATI'l1\.u!lan
(mollnorna Dl tDII palll) 1(2;22)~;q12) EWSJ!l.cREBll'llllon
aon...,llllnfonlllo 1(12;15)(pl3;qlfi) EJW.Nil!l<3,...,
n"""""""""
Dodl'l&lllfldotod ll_r...,. ~-um•OIY ~nundlor
mill«< <II_,.,
lOith •mpl~- of
12q14-q15
-
O.niiiiDft-rama 107;22Kq:!21q13l•l'od COL!Al-P£161'B l'wlon
~bcnnlfPont ...n d_n,..n...,.,..
ftbloblulomo
Doomol:l ftlllrAnirlcoll Tr1oolny 8 .. 20: .... "'Sci Soma11c C'INNIJlor APC
mwtlona (only In deop
Ill moll)
Doomopl.ult:..,.l """" 101;22Ki>l<%q121 EWSRl-WTl fulbn
ot~lllllnv
Eloo$>ftblolna IQ allno!Tnlillleo UnknOWII
E'lnbi)»rroo L.ooo of ldow<a</Z!fPitt-t.et UnknOWII
rhlbdo-- llpl5; IIIN of 2, 7, 8,
II, 12, 20, 21, 19<121,
20;-ort~
7, ~ 9q22,14q21..:12,
17
~!lllalold hemlll!»- 10;3)(~25)
••do111..,.,.
8~-··-·"'
Epll:t\.ld MR:OI'nlllJ Ah/ilii<N; "'22qll.2
prad'liiii!Jpo h6NFSRN!l
__,,_
ilot1r U/QOOI illp/tnlllwl 101;22Kq24;q121 EWSJ!l-l'llll'wlon
nouro&~;tldaimeltumor 1(21;22)(q221q12) EWSRI-El!G l'wlon
~~~-··-·ot
Elc!nl11110lltiiii!Nilt AIB!Ilbooof22Qil.2
rhlbdold 1llnor hSNFS/IN/1
t(!I;22)(Q22;ql2) EWSR 1-NRIASfu&loll
chondi'Oiiii'CI:Ifnl 1(!1;17)(Q22;q11.2) TAF2N-NR4A3,...,
1(!1; IS)(Q22;q21) TCFl2-NR4113-
Gllntot~lllllnvof T~tnolol:1/l!omiiMIMIW CSF'lflllbl~ -k\,t
ollorlllld~ &1oM ll>l~.lnwdiiW CSFI-C014113
""lllllnva t(l;2)(pl~l3Sl
~m.-...y n.no-.r..IIM>Mre
mya11btol>lullet:Lmor 21>2$
a..~.46· Sottr-.e. I .,.,
I'1.1 II( !-f\1 ~:~c.r:~nuet a-c~e~~auc ot'llllo<a Soft
-n.orlJIII C)loalo,.~ ••~... •...u.-..
SW<:tu11111allol1!tiomof I,
7, 10. 13. 14
Reu..rwo-ofll<ll2
RMinll\lll-
....
R001t1181nlfllt <>I I'I.Ml
HMSA2andHIIIGA1
12q14-ql5 and ilslona; unlcncMTI
6p2142;d-!l
13ql2-14
i.oW-1111dotb~ 117;16l<Q33:Plll
u""""" l(ll;l£)(~11;pll)
Malpnt .-llf\e<OI""""'
t.t'IMth tumor
"'"""'ldlrwncl<d IU2:16J(ql3:pll) RJS.CDJT3futb!
llpolll""""" 1{12:22] (q 13;ql2) Ew.SRl.OOlT3,....,
Nooo!>hiMlliiOI (loN II IP, 7q, lOa, 12<1, ~m...._,l'l
lllllo4\b~ 1611. !tiel. 17q, 19~. CTNI{9J
21lq, 22'1
Softlloul-11111111 Tnonllio<>ltlorotl.-...lflna Ew.SRlfW>no\O!Ih •
tumor 22ql2 WJIIItyof cthlr•n•
SSKlii-SSlfJ, .s:sx:!B-
~~--
10(;18)(pll:qll)
S$1o2 $$XIB-SSJI4
ilolono
'JC2 I SECTION .... SOFT TISSUE AND BONE
UJIISI!t-
lollp
u-
Uoomow
Upo~•llotooll of,....
Upol>lu1rma.41po-
An&loll-
M)'aii-
Chonct'oldlpoma
E'lc!nl-r1flal•*m:ldi-
Elc!nHdiWIITI'.IOitil-
Spindle oollp"""'OI!JIIIc 1-..m•
Hlllmorn.t
-lllttloaiP.rqpoooho)
Al)lbii_,.INIIDustumOf--lpooa"""'o
......It
o.dllo!WoliDdll--
u--
~ llpauu:ou"""'"nd a!llpo~~,..,.,,
Ploom,.plllc llpoouama
M!Jal~l-mo
~~-....,.,......-1:1
IIIIP
...
110t OChenol8o C>dod
-~~~fuellila
Prolf--
Prolf-m)Ottil
~oalllcoM
Flbtt>-.a
lld>omlc-
""""'""'mor II dil!lt&
EIIoloftlmnl
Flb!out lllmoltomo II Infancy
M)'ofli>fon'6'rn:lollb.... -
F1-coll
JIJYM!Io ...,...... ftbllli'IIO-
~ bodytbll>lrt-
Fl-. aflon6on sh-
Ocomoplllll: f l b - •
Mlm . .!Y'lype m)IOII-•
COic&'JinUb'"IOIJIOCSc ~blr.lno
Anatou~brobiMtM'B
COIIJ~r IJl!jallbloma
Nudl.tlqpo flb"""1
Gardner flbroma
CllcfJina flbn>llstum"'
Glln!Q!IIInaloflbloml
I IIIIi lOII IIIt Ooall.r IJIJIIIII'I)
f>upelfte~lftbla- (polmulp~rml
Ocomolii~P"ft
Upoft-
a..~.46· Sottr-.e. I .,.a
'' f.t:] I (!-Ill WHO C&Mik&tJon of Soft 11at n.nca (fcrM*Mf)
-Crnl.r·-·"tl
Soaalyftlln>uotumor and llcml'*~
lntllmnWoly ll'l)"ftbn>blullc tumo<
Low-indo ml'lf\brobloo1ic ""'""""
tot..>l.-mt:IO!yftbrol>l&ade arcol!\1
•-nb......""'
lllllloool
Aclu~ n-,...,.
~-,..,.,.
Low-indoftb~ 01/~lrM\.c~lo col tumOf
.......
.... _loti,.._
~·ptilelklidft.....,_
Glontc:lllll tumor of ton lion oh-

~D&a!artcol111nor
-Crnl.r·-·"tl
DMc> b&n\111 ftbrouo. hlllloqtomo
Ploodfonn n-•llcqllt lllmar
Glontc:lllll tumor of tollllauao
lllllloool
Pleom01plile MFHAindlll,_ ~hie ..,..,.
Glint c:lllll MFHAindlllerwrtilllll plao"""'>hlc •ramo ..til pnl Clllls
·-·--...
lnftamii'IIIDiy MFHiund- plaornorplllc..,...... ..til pnrnlft01111nlllm1Ntt>n
AnaloWonTIOIM
DMc> loiom)ooiNI
Gtnlltl i&b'nJr10ml
Ltioii1)'0LI,..,.
..... ~
Olcmuo !UmOf
...,.lftO
Malinont alotrrlla 1llnor
·-·-"11-
....
""-"""""'"
Rhobdo"""'""' Ct4/l""". fiiiBIIYI>O.IIIIitel typo)
lllllloool
EmbljiOnal ltlahd..,_..INI tmblll& '!>Indio oel, lmyokl, •napl&ade)
A - rhlbdrlri)'OOircotnl
....
PlaomorpNc rhllxbli)'OII/~'1111
-.~~r ....
HomarWOmuda~b<Wn-p...rttllau•
C.plltry
eo.omo.a
Ario-n..,.
Ve.....
lntom~~J~C~Jittr
~·
Epltlotidhlma'*ma
An&m-ILA
Ljmpl\l ......
,.... I SECTION .... SOFT TISSUE AND BONE
,, (.1:! I (!-JJI WHO cua:atJon of Soft n.t n.nca (ffrM*Ml

lillf••lllo ~" . ."""..,
IC:Ipoollonn liome.....,nilalluilome
--IIIIo
Rtd'onn INtiJ - - -
hOIIIIII\I.bllidollieilorno
Popl~ry liltrll.wTn~ ~lorn·
Com..,... h.,.nfllaonddl,...••
IC:Ifl(lll ..........
- ......
llllp.tlt
~!lllolold heml/liSOGIII:Iallellol!lll
Alijjou""""" o l d - ·
SilftU...O<hon<*'ome
Mooml:!iymll oliond"""'"""""
......... _ ..... 11111 . . .
-.ra
-~~~-
Ji.iidu_,., m-
o...,. (lui""'IIO) ...... _ .
Pleonlotlllilc ll';all'illln..~111n«
~h-uo11tJrnomo
lillfi ... lllo"'""' - - -

Aii.....,._,ld nb...... nliiiSoq!Dma
~i'C flb...,)llllld IUiliQ'
Mlial llimoliii'I)OOpllidlornii/pliii<lUcbna
llllp.tlt
$fllowlai1U......
~!lllolold ........,.
_,soft pcrt .........
Clolr cd ,.......,. Ill softU...O
~ ~ dloll<toootool!lll
PNET~I Elotra:lllmor
O.mopl.&.d! .ltnd roond oe!ltumor
EllnMINI ill.t- bimor
,.,...,.,.. ,_nd!ymoma
........,.
~.,...., poilvuclllar apllid~ odl • - (PEtonwil
Clolr cdlli)'Om'*"""')tic tiiTIOf
~,.,
(hlllo!Qii tpootllc). n1lollc ~. 1nd -of
niUC -111. The FNCL<:C.,..S. ~~-- llythrM ponune!Ms: dll'lera1tlltlon
-.0111. ~ peramdllf ls...,.od:
dllar-n (1-3). mllollc adMif (1-3). end ,_..Ia (N).
~-1)1do.
Tho...,,..
uo !lllmmod to
Gro&XGruloOMinotb&e_...
Gro&l2tr$
Grldo24or5
Grldoa&-a
.......,..., TLmOr <II!IIUI- b """""'ljiOdllc end bl'notll~ ""'""'at -.•,
Sootlll S.l'tOI"'''M doistJly r81&1embllrw nomw, metuhl ~\altlllilu•
Scora 2 s........ of_.. -lo0>1!!1>6
Sccf'a 3 s,onc:MII ~ tmtr)a1.11.u.rmmu, undlft'•ilt'tllled urc:om~~, Jnd u~ of
unknownldoullti\11we
.lllllllft-ln 11M> moot mbCI!:otJ . . - . ,.. of tho"""""'., !811-M> hlat>-
fWda {HPft) m •••ud "*'£ • 4fit ob}ed:W.
Scora 11)..0 m-10 HPF&
Scora 210.1; m~O HPFs
Scora a20"' mora miDMif!O HPFs
,..,_,a rrAt EW~Iuat&d on ITOiltEU.Illll'liltkJn tnd w.ldltiDdwth hM'"'* w:t1on.s.
Score 0 ~tum« noe<*
Sootlll ~~tum«~
Sootll2 >~ t.rmor rtl!la'osk
746 I SECTION XI: SOFT TISSUE AND BONE
adipocytes separated into complete and incomplete lobules. Numerous his-

tologic subtypes have been described but none has prognostic significance,
including the following (Adv Anat Pathol. 2006;13:279).
a. Angiolipoma typically occurs in the subcutaneous tissue and consists of
mature adipocytes with a variably prominent capillary network with
scattered microthrombi. These vessels are typically found at the periph-
ery of the lobules (e-Fig. 46.1)."' These tumors may be multiple in the
subcutis, and also occur in and around the spinal cord (Childs Nerv Syst.
2002;18:725).
b. Myolipoma (intramuscular lipoma) is found in the deep soft tissues of the
abdominal cavity, inguinal region, and retroperitoneum. Mature adipose
tissue is intermixed with mature smooth muscle or skeletal muscle.
c. Chondroid lipoma occurs in the limb girdle and proximal extremities.
Cords and nests of lipoblasts as well as mature adipocytes are present in
a myxoid to hyalinized chondroid matrix in the absence of true hyaline
cartilage. Despite the presence of immature fat cells, surgical excision is
curative. This tumor is distinct from chondrolipoma, which is character-
ized by hyaline cartilage within a lipoma.
d. Spindle cell lipoma/pleomorphic lipoma occurs predominantly on the pos-
terior neck and shoulder area in middle aged and elderly men (only 10%
of cases occur in women). The tumor presents as a mobile dermal or
subcutaneous nodule that often has been present for many years. The
microscopic features are variable: at one end of the spectrum are tumors
composed of bland spindle cells with associated dense collagen bundles
between mature adipocytes; at the other end are tumors with small hyper-
chromatic cells admixed with multinucleated giant cells between mature
adipocytes. The spindle cells are immunopositive for CD34, and in some
cases also for S-100 protein. This neoplasm is one of several cutaneous
and STTs, which express CD34 (] Cutan Pathol. 2009;36:89). In the
presence of atypicallipoblasts, spindle cell LPS must be considered since
the latter tumor also rises in the subcutis (Mod Pathol. 2010;23:729).
2. Lipomatosis occurs in several different clinicopathologic settings, all of
which are characterized by a diffuse overgrowth of mature adipose tissue.
Regardless of the clinical subtype, the neoplastic cells are indistinguishable
from those found in lipomas, which emphasizes the role of clinical history
in arriving at the correct diagnosis.
a. Diffuse lipomatosis preferentially occurs in children <2 years old, and
involves a substantial part of an extremity as well as the trunk, head
and neck, pelvis, abdomen, or intestinal tract. In this setting, the phos-
phatase and tensin homolog (PTEN) hamartoma tumor syndrome as
expressed in the Proteus syndrome, encephalocraniocutaneous lipomato-
sis, Bannayan-Ruvalcaba-Riley syndrome, and Cowden disease should
be considered (Genet Med. 2009;11:687).
b. Symmetric lipomatosis occurs predominantly in middle aged men of
Mediterranean ancestry, and is characterized by symmetric deposition
of fat in the upper body.
c. Pelvic lipomatosis, which affects black males over a wide age range, usu-
ally manifests as an overgrowth of fat in perirectal and perivesical areas.
d. Steroid lipomatosis occurs in the setting of adrenocortical hormonal ther-
apy or with endogenous endocrine abnormalities, and characteristically
involves accumulation of fat in the face, sternal region, or middle of the
upper back (the so-called buffalo hump).
Chapter 46 • Soft Tissue I 74 7
e. HIV-Iipodystrophy in patients with AIDS occurs in those undergoing treat-

ment with protease inhibitors or other forms of antiviral therapy, and is
characterized by the accumulation of visceral fat with fat wasting in the
face and limbs.
f. Nevus lipomatosus is a developmental anomaly presenting as a yellowish
polypoid lesion of skin, typically in the lower abdominal-sacral-pelvic
region (] Dermatol. 2000;27:16). It is characterized by mature adipose
tissue in the papillary and reticular dermis. Soft fibroma and acrochordon
have some overlapping features, but adipose tissue is not found in the
papillary dermis in these latter two lesions.
3. Lipomatosis of nerve (neural fibrolipoma, fibrolipomatous hamartoma) is
noted at birth or in early childhood, but is seen through the fourth decade.
The median nerve and ulnar nerves are the usual sites of involvement, and a
subset of cases is associated with macrodactyly. Perineurial and epineurial
infiltration by a mixture of mature adipocytes and fibrous tissue typically
separates individual nerve bundles. Another congenital fatty lesion related
to peripheral nerves is lumbosacral lipoma in infants, which is character-
ized by a tethered filum terminale or conus medullaris (Childs Nerv Syst.
1997;13:298).
4. Lipoblastoma occurs in children (90% of cases occur in children under
the age of 10 years) with a predilection for the lower extremity, but can
also involve the trunk, mediastinum, abdomen-retroperitoneum, and head
and neck (Am] Surg Pathol. 2009;33:1705). It is either a localized, well-
circumscribed tumor (lipoblastoma) or has a diffuse infiltrating pattern
(lipoblastomatosis). Like other fatty tumors, it has a lobulated architecture
and is composed of a mixture of cell types including mature and immature
adipocytes, a variable number of lipoblasts, multinucleated cells, signet-
ring lipocytes, and stellate mesenchymal cells (e-Fig. 46.2). Grayish myxoid
areas noted grossly have a resemblance to myxoid LPS, which can be prob-
lematic. Despite the presence of immature fat cells, this tumor is benign and
does not metastasize, although approximately 20% to 25% of cases recur
(most are examples of lipoblastomatosis). These tumors have the potential
for maturation, and some cases have predominantly lipomatous features
with residual immature myxoid areas at the periphery of the lobules. These
tumors express for S-100 and CD34.
5. Hibernoma is a neoplasm composed of brown fat (adipocytes with multi-
vacuolated granular cytoplasm) admixed with conventional adipose tissue.
It occurs in young adults and is found in the neck, axilla, thigh, retroperi-
toneum, head and neck, trunk, and upper extremities (Am] Dermatopathol.
2009;31:685). The cut surface has a yellowish to brownish appearance, is
usually oily and spongy; may be lobulated but is well demarcated; and can
measure over 20 em. Microscopically, lobules of brown fat are separated
from conventional adipose tissue (e-Fig. 46.3).
B. Intermediate (locally aggressive)
1. Atypical lipomatous tumor/well-differentiated LPS (ALTIWDLPS) (50% to 55%
of cases) occurs in adults in the fifth through eighth decade of life (Cyto-
genet Genome Res. 2007;118:138). The deep soft tissues of the lower
extremity and retroperitoneum are usual primary sites; the paratesticu-
lar region and mediastinum are less common sites. Tumors arising in the
retroperitoneum may attain sizes in excess of 20 em and weigh 500 to
1000 g. The tumor has a lobulated, yellow to white, soft to firm cut sur-
face that varies on the basis of lipomatous, fibrous, and myxoid com-
ponents, and discrete margins are often difficult to discern from gross
examination. Microscopically, the tumor is composed of cells with lipoma-
like features except for the presence of scattered hyperchromatic, often
multinucleated and vacuolated cells with features of atypical lipoblasts.

Four histologic subtypes are designated, namely adipocytic (lipoma-like),
sclerosing, inflammatory, and spindle cell types, but more than one pat-
tern may be present in the same neoplasm. Sclerosing foci are helpful in
diagnosis because the areas of collagen contain atypical stromal cells (e-
Fig. 46.4). Fluorescence in situ hybridization (FISH) for MDM2 amplifica-
tion characteristic of ALTIWDLPS is a sensitive and specific tool for dis-
tinguishing ALTIWDLPS from benign lipomatous neoplasms (Mod Pathol.
2008;21:943; Adv Anat Pathol. 2009;16:383).
Prognosis is largely determined by anatomic site and size. Smaller, more
superficial tumors can be locally resected with negative margins, but those
in the retroperitoneum are likely to recur because of positive surgical
margins. Recurrent tumors may show evidence of so-called dedifferenti-
ation (see below) which clearly demonstrates the overt malignant poten-
tial of ALTIWDLPS. Multifocal ALTIWDLPS is an uncommon but well-
documented presentation.
C. Malignant
1. Dedifferentiated LPS shows a transition from ALTIWDLPS to a pleomorphic
and/or high-grade spindle cell sarcoma (e-Fig. 46.5), either in the primary
tumor (85% to 90% of cases) or in a recurrence (10% to 15% of cases) (Vir-
chows Arch. 2010;456:167). The transition in pattern from ALTIWDLPS
to high-grade sarcoma is usually abrupt. By convention, the focus of ded-
ifferentiation should be at least several millimeters in greatest dimension.
Because the area of dedifferentiation may be limited, thorough sampling and
careful microscopic examination of all ALTIWDLPS is required in order to
exclude the presence of dedifferentiation.
2. Myxaid LPS/raund cell LPS peaks in incidence at the age of 30 to 40 years,
occurs predominantly in the deep soft tissues of the extremities (more than
two-thirds of cases arise in the musculature of the thigh), and is the most
common type of LPS in the first two decades of life (Ann Diagn Pathol.
2000;4:252; Am] Surg Pathol. 2009;33:645). If this tumor is discovered
in the retroperitoneum, it likely represents metastatic disease rather than a
primary tumor (Mod Pathol. 2009;22:223 ).
The cut surface of myxoid LPS is tan, glistening, and gelatinous; the
round cell morphology is associated with a fleshy, white cut surface. Myx-
oid tumors are composed of uniform, round to oval, primitive nonlipogenic
mesenchymal cells and smalllipoblasts embedded in a myxoid stroma with a
delicate arborizing capillary network (e-Fig. 46.6). In contrast, areas com-
posed of sheets of high-grade primitive round cells are not accompanied
by a myxoid stroma. It is not uncommon to identify a subset of round
cells in myxoid LPS, usually in a perivascular distribution, but by conven-
tion round cell LPS has a composition of 80% or more of round cells.
Immunohistochemically, the round cells are often S-1 00 protein positive. A
poorer outcome is associated with round cell LPS, unlike the more favor-
able prognosis of pure myxoid LPS. Although the same two translocations
are characteristic of both tumor types (Table 46.1), they do not correlate
with prognosis (Cytogenet Genome Res. 2007;118:138).
3. Pleomorphic LPS, as the name implies, is by definition a high-grade sarcoma
with a variable number of convincing pleomorphic lipoblasts. This tumor
has a preference for the extremities, usually measures in excess of 10 em,
and primarily occurs in individuals over 40 years old. The tumor is either
a well-circumscribed or an infiltrative mass with a variable appearance on
cut surface ranging from solid to cystic, to necrotic, to hemorrhagic, to
myxoid. Pleomorphic lipoblasts with enlarged hyperchromatic nuclei that
are scalloped by cytoplasmic lipid vacuoles are not always numerous in the
background of highly atypical, even anaplastic round cells, spindle cells,

and multinucleated tumor giant cells (e-Fig. 46.7). These tumors may have a
prominent inflammatory infiltrate. Atypical, even bizarre mitotic figures are
often present. Well-differentiated LPS can dedifferentiate to pleomorphic
LPS (Am] Surg Pathol. 201 0;34: 1122; Am] Surg Pathol. 201 0;34:83 7). In
the absence of identifiable lipoblasts, these neoplasms are otherwise diag-
nosed as pleomorphic undifferentiated sarcomas but there is marked mor-
phologic overlap between these tumors (Am] Surg Pathol. 2009;33:1594).
IV. FIBROBLASTICIMYOFIBROBLASTIC TUMORS
A. Benign
1. Nodular fasciitis occurs in all age groups but has a predilection for young
adults. It usually involves the subcutaneous tissue of the head and neck
(especially in children), trunk, or upper extremities (e-Fig. 46.8). Dermal
involvement is uncommon, but deeper fascial or intramuscular tumors are
other presentations. Similar lesions may involve small to medium sized
veins or the soft tissue of the outer table of the scalp (infantile cranial
fasciitis), as intravascular and cranial fasciitis (e-Fig. 46.9), respectively.
Uncommonly, the tumor develops at intraneural and intra-articular sites.
These circumscribed, minimally infiltrative spindle cell proliferations
have a fibrous to myxoid cut surface, and most are <2 em in greatest
dimension though some lesions can exceed 5 to 6 em. Cystic degeneration
is an uncommon gross feature, but one of the microscopic hallmarks is
the presence of microcysts among the more cellular foci (Arch Pathol Lab
Med. 2008;132:579). Collections of inflammatory cells, extravasated red
cells, or osteoclast-like giant cells may be associated with the microcysts.
Compactly cellular foci with storiform profiles or interlacing fascicles may
reside adjacent to individual cells with a myxoid background with a tissue
culture-like appearance (e-Fig. 46.8 and e-Fig. 46.9). More collagenized
foci resemble a keloid. Mitotic figures, but not atypical ones, are expected
in variable numbers. The spindle cells are strongly reactive for smooth
muscle actin (SMA); unlike desmoid tumor, the cells are nonreactive for
,8-catenin (Histopathology. 2007;51:509).
2. Proliferative fasciitis and proliferative myositis primarily occur in middle
aged and elderly patients. The subcutis in the upper extremity is the most
common site of proliferative fasciitis, but some cases involve the trunk
or lower extremity. Proliferative myositis is intramuscular, and primarily
involves the trunk, shoulder girdle, and upper arm. Both lesions grow
rapidly, measure between 3 and 5 em, and are composed of plump fibrob-
lastic and myofibroblastic spindled cells as in nodular fasciitis. However,
the hallmark of proliferative fasciitis and proliferative myositis is the pres-
ence of large ganglion-like cells with an uneven distribution within the
lesion. The ganglion-like cells may be mitotically active but atypical mitotic
figures are not present. In addition to SMA, CD68 may be expressed in
the ganglion-like cells.
3. Ischemic fasciitis occurs over bony prominences, usually due to impaired
circulation and prolonged pressure in immobilized, often elderly individ-
uals. A zonal architecture consists of central areas of coagulative necrosis
and myxoid change, with fibroblastic and vascular proliferation at the
periphery (Am f Surg Pathol. 2008;32:1546). There is some resemblance
to deep granuloma annulare.
4. Myositis ossificans and fibro-osseous pseudotumor of digits are related
lesions that occur in a broad age range of patients, though young adults
are most frequently affected. Myositis ossificans has a propensity for the
extremities, trunk, and head and neck, while fibro-osseous pseudotumor
primarily occurs, as its name indicates, in the subcutis of the proximal
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visceral organs, and the central nervous system. Individual tumors range
from < 1 to over 7 em in greatest dimension and have a cut surface that
ranges from firm and fibrous, to cystic and hemorrhagic. Immature plump
to spindled cells are arranged in whorls and fascicles within a fibromyxoid
stroma; smaller nodules at the periphery of the mass may be associated
with a vessel(s) to suggest an angiocentric origin (e-Fig. 46.13). Hyper-
cellular spindle cell foci are present in some cases with a resemblance to
congenital infantile fibrosarcoma, but the tumor cells lack the t(12;15)
translocation characteristic of the latter tumor. Other findings include a
hemangiopericytoma-like pattern centrally, often with ischemic or hem-
orrhagic regions, dystrophic calcifications, and hyalinization. The spindle
cells express vimentin and SMA, whereas the hemangiopericytoma-like
foci are variably CD34 positive. The differential diagnosis of myofibroma
includes nodular fasciitis; the distinction can be problematic since the two
lesions have some overlapping morphologic and immunophenotypic fea-
tures (j Clin Pathol. 2009;62:236).
8. Angiomyofibroblastoma, a rare tumot; occurs in women of reproductive age,
where it arises in the rises in the pelviperineal region (vulva and vagina)
as a painless, well-circumscribed, slowly enlarging mass (IntI Gynecol
Pathol. 2005;24:26). In men, the neoplasm usually involves the parates-
ticular soft tissues or scrotum. Round to plump spindled myofibroblasts
tend to cluster around blood vessels (e-Fig. 46.14); a subset of cases has
a mature fatty component (IntI Gynecol Pathol. 2005;24:196). Binucle-
ate and multinucleate cells are common in the absence of mitotic activity.
These tumors are immunoreactive for desmin and estrogen and proges-
terone receptors. Some cases have an overlap with cellular angiofibroma
and deep aggressive angiomyxoma (see below).
9. Cellular angiofibroma involves the superficial soft tissues of the vulva or
inguinoscrotal region as well as the perineum, retroperitoneum, and sub-
cutis of the chest U Cutan Pathol. 2003;30:405). Grossly, the tumor is
a well-circumscribed mass that usually measures <3 em in diameter in
women and <10 em in men. The cut surface of the tumor has a yellow to
tan-brown, soft to rubbery appearance. The tumor is composed of plump
spindled cells with minimal eosinophilic cytoplasm, little cytologic atypia,
and few mitotic figures; a background of delicate collagen fibers is present
in tumors in females. The vascular component of the tumor is composed of
small- to medium-sized vessels, with or without prominent hyaline walls,
and is usually present throughout the entire lesion. Regressive/degenerative
changes, including extravasated erythrocytes, hemosiderin deposition, cys-
tic change, and intravascular thrombi are also present.
10. Giant cell angiofibroma, a slowly growing occasional painful tumor, has a
predilection for the eyelids and orbital region of adults, although it has
been identified in a number of other anatomic sites. The tumor is usually
about 3 em in greatest dimension, well-circumscribed, variably encapsu-
lated, with cystic and/or hemorrhagic areas. Microscopically, cellular areas
of round to spindled cytologically bland cells and multinucleated stromal
cells (which often line pseudovascular spaces as is common in giant cell
fibroblastoma) are set in a background of myxoid to collagenous stroma
with small- to medium-sized blood vessels. Both the mononuclear and
multinucleated cells are immunoreactive for CD34 and CD99, and occa-
sionally also for BCL2.
11. Nuchal-type fibroma and Gardner-associated fibroma are virtually identical
in terms of the microscopic features of dense, paucicellular collagenous
bundles, which overgrow and occupy the dermis, subcutis, and deep soft
tissues (Cancer. 1999;85:156). These tumors do not have the infiltrative
features of desmoid fibromatosis, however, a recurrence may be indis-

tinguishable from a desmoid. As in desmoid tumors, there is nuclear
immunopositivity for ,8-catenin. The possibility of Gardner syndrome
should be raised in the presence of this tumor (Am ] Surg Pathol.
2007;31:410). In the latter, multifocallesions are seen.
B. Intermediate (locally aggressive>
1. Superficial fibromatoses. Palmar-plantar fibromatosis develops in males over
30 years of age (with a male to female ratio of 4:1) as an asymptomatic, iso-
lated firm nodule, which evolves into cord-like bands between the nodules
involving adjacent fingers. Plantar fibromatosis is seen more often in chil-
dren and adolescents as painful subcutaneous nodules (Am] Surg Pathol.
2005;29:1 095 ).
The microscopic features evolve over time. Greater cellularity is present
early on, consisting of bland plump to spindle cells that have a low mitotic
rate and are set in a background of collagen and elongated vessels (e-Fig.
46.15). Older lesions are much less cellular and have a stroma that consists
of dense, often hyalinized collagen. The extent of surgical excision is the
primary determinant of the rate of recurrence. These tumors may demon-
strate focal rather than diffuse nuclear positivity for ,8-catenin (Mod Pathol.
2001;14:895).
2. Desmoid-lype fibromatosis (desmoid tumor, musculoaponeurotic fibromato-
sis) usually involves the head and neck region in children, and the proximal
extremities and abdominal wall in adolescents and older females (Hema-
tol Oncol Clin North Am. 2005;19:565). Both soft tissues and mesenteric
desmoids may be associated with Gardner syndrome. A circumscribed mass
measuring 5 to 10 em with a firm white trabeculated surface is the typical
gross appearance; the macroscopic circumscription may be deceptive since
subtle and extensive infiltration into the interstitium between muscle bun-
dles and along fascial planes is often present microscopically. Although
desmoid tumor is classically a proliferation of fibroblasts, a number of pat-
terns are seen, ranging from spindle cells forming bundles and fascicles in
a dense collagenous background, to plump fibroblasts in a pale less fibrotic
stroma (e-Fig. 46.16). Small blood vessels may be conspicuous, and when
red blood cell extravasation is present, the lesion can resemble nodular
fasciitis. Some mitotic figures may be present, and scattered small lymphoid
nodules may be noted at the interface with surrounding normal tissues. If
skeletal muscle is involved, it is usually infiltrated with remnants of mus-
cle embedded in the fibrous proliferation (e-Fig. 46.17). SMA expression is
common, and 46% to 50% of cases have nuclear reactivity for ,8-catenin
(Am] Surg Pathol. 2007;31:1299).
3. Lipofibromatosis (infantile subcutaneous fibromatosis}, another fibrous tumor
of childhood, is a slowly growing, painless, ill-defined neoplasm occurring
in a variety of sites including the distal extremities (Am ] Surg Pathol.
2000;24:1491 ). Grossly, the tumor is an ill-defined white-tan to yellow mass
that usually measures <5 em in greatest dimension. Spindled fibroblastic
cells form bands that surround and may separate lobules of fat; the growth
pattern resembles fibrous hamartoma but without the nodules of immature
mesenchyme. The tumor may express CD34, BCL2, S-100, actin, epithelial
membrane antigen and CD99, an unusual phenotypic profile for a fibrous
tumor.
4. Infantile digital fibroma-fibromatosis (inclusion body fibromatosis, recurring
digital fibrous tumor of Reye) presents in the fingers and/or toes (10%
to 30% of cases are multifocal), excluding the thumb and great toe, as
a firm nodule or nodules. Rare examples of extradigitallesions have been
reported (Am] Surg Pathol. 2009;33:1 ). The fibrous proliferation resembles
a desmoid tumm; with confluent infiltration and replacement of the dermis

and deeper soft tissues including the skeletal muscle. Isolated adnexal struc-
tures are surrounded by the moderately cellular spindle cell proliferation.
Paranuclear bodies consisting of actin microfilaments are one of the unique
features of this tumm:. These tumors are immunopositive for SMA, calponin,
CD99, and CD117.
5. Juvenile nasopharyngeal fibroma occurs almost exclusively in adolescent
males, often presenting with epistaxis. Extensive local growth occurs in
the confined spaces of the nasopharynx and into the paranasal sinuses and
pterygopalatine fossa (lnt J Pediatr Otorhinolaryngol. 2011;75:1088).
These tumors are firm and have a white-tan cut surface. A diffuse bland
fibrous proliferation with a prominent component of small blood vessels
is the typical microscopic appearance. This tumor is seen in the setting
of familial adenomatous polyposis. The stromal cells express nuclear
,8-catenin.
C. Intermediate (rarely metastasizing}
1. Solitary fibrous tumor (SFT} and hemangiopericytoma (HPC} are grouped
together as related (if not identical) neoplasms in the WHO classification
(Histopathology 2006;48:36).
a. SFT is classically found on the pleura (Semin Diagn Pathol. 2006;23:44 ),
but has been reported in many different locations (including the dura,
intestinal tract, mesentery, liver, skin, thyroid, lung, and orbit) in a broad
age group. The tumor is a well-circumscribed, nonencapsulated, firm,
white mass measuring 8 em or less, and may show hemorrhage and
focal myxoid change. Bland plump to spindle-shaped cells with a pat-
ternless architecture surround branching blood vessels of the type associ-
ated with HPC (e-Fig. 46.18). The cellularity often varies within individ-
ual tumors, and the hypocellular background stroma can have a myxoid
fibrous appearance, and can resemble a nerve sheath tumor or low-grade
fibromyxoid sarcoma (LGFMS). The tumor cells are immunoreactive for
CD34 and CD99, but a subset of tumors also shows reactivity for SMA,
BCL2, EMA, and even focal positivity for desmin, cytokeratin, and/or
S-100. D2-40 may be positive in a small subset of SFTs, but immunoreac-
tivity is more common in mesotheliomas (Appllmmunohistochem Mol
Morpho/. 2010;18:411).
Malignant SFTs are usually 10 em or greater in size and have increased
mitotic activity (~4 mitoses per 10 HPFs), focal necrosis, increased cellu-
larity, marked cytologic atypia in a patchy distribution, infiltrative mar-
gins, and show p53 expression (Arch Pathol Lab Med. 2010;134:1645).
However, the clinical behavior of an individual tumor is not always cor-
related with the histologic features. Some tumors have a solidly cellular
fibrosarcoma-like pattern yet behave in a relatively innocuous fashion.
b. By the current WHO classification scheme, the diagnosis of HPC is lim-
ited to STTs that morphologically resemble the cellular areas of SFT
(e-Fig. 46.19), and that are composed of cells that have the immuno-
profile of SFT. By this definition most cases of HPC occur in the deep
soft tissues, primarily in the retroperitoneum of the pelvis, but also in
the limb girdle and proximal upper or lower extremity. Most tumors are
<15 em in greatest dimension. Microscopically, the tumor cells have a
uniform ovoid to spindled appearance in a background of small clefted
to branching vascular spaces which are highlighted by CD34 immunore-
activity. Variable immunostaining for SMA is present. Mature adipose
tissue is found in the lipomatous variant of HPC. Other settings for HPC
include dural-intracranial, sinonasal, or infantile presentations; the lat-
ter is regarded as a pattern of infantile myofibroma-myofibromatosis
(] Clin Neurosci. 2010;17:469;/ Pediatr Hematol Oncol. 2011;33:356;

Am J Surg Pathol. 2003;27:737). As a final note, an HPC-like pattern of
growth is seen in synovial sarcoma, malignant peripheral nerve sheath
tumor (MPNSTs), mesenchymal chondrosarcoma, infantile fibrosar-
coma, thymoma (spindle cell), and endometrial stromal sarcoma, empha-
sizing the need for careful microscopic examination, immunohistochem-
istry, and molecular studies for correct diagnosis.
2. Inflammatory myofibroblastic tumor (IMT) has a predilection for the mesen-
tery, small intestine (ileocecal region), lung, and bladder, throughout child-
hood and into young adulthood. Approximately 5% to 10% of IMTs
have systemic, constitutional manifestations (fever, anorexia, weight loss,
hypochromic microcytic anemia, and/or hypergammaglobulinemia, throm-
bocytosis), likely due to IL-6 production by the tumor.
The tumor forms a white to tan, whorled, fleshy to myxoid, circum-
scribed mass measuring 1 to 25 em in diameter that may also show areas
of necrosis, hemorrhage, and calcification. Three basic histologic patterns
can be found in any one tumor, although one or two may dominate (e-Fig.
46.20): dense fascicles of spindle cells with a mixed population of plasma
cells, lymphocytes, and eosinophils in the background; loosely cellular foci
with a myxoid and edematous background resembling nodular fasciitis; and
hypocellular, collagenized foci with minimal inflammation and dystrophic
calcifications. Mitotic figures are found among the spindle cells, but they
are not atypical. Some tumors have a predominant round cell or epithe-
lioid pattern, which should raise the possibility of an anaplastic lymphoma
kinase (ALK)-positive inflammatory myofibroblastic sarcoma with only a
minor spindle cell component (Am J Surg Pathol. 2011;35:135). The dif-
ferential diagnosis of IMT includes two unrelated lesions, calcifying fibrous
pseudotumor with psammomatous calcifications and inflammatory fibroid
tumor, the latter usually is present in the stomach or in the small intestine
(Int J Surg Pathol. 2002;10:189; Adv Anat Pathol. 2007;14:178).
Immunohistochemically, a subset of tumors (approximately 50% to
60%, corresponding to those cases that harbor a rearranged ALK gene)
shows cytoplasmic staining for the ALK gene product (Am J Surg
Pathol. 2001;25:1364). However, virtually all cases are immunoreactive
for vimentin; most show reactivity for SMA and variable reactivity for
muscle-specific actin (MSA) and desmin; and 25% to 30% are reactive
for cytokeratin. ALK-positive tumors may have a more favorable clinical
outcome than the ALK-negative tumors (Am J Surg Pathol. 2007;31:509).
Some IMTs may have a population of lgG4 plasma cells, but this finding
does not imply a relationship to the lgG4-related sclerosing disorders (Mod
Pathol. 2011;24:606).
3. Congenital infantile fibrosarcoma (CIF) occurs in children aged 2 years or
younger (and can be present at birth) and usually presents in the superficial
or deep soft tissues of the distal extremities, although cases also present
in the trunk, head and neck, intestinal tract, deep pelvic tissues and heart
(J Clin Oncol. 2010;28:318). This tumor can be mistaken clinically for a
vascular tumor.
The cut surface of the tumor (which can measure up to 10 to 15 em
in diameter) ranges from white to tan, fleshy to firm, and may show areas
of hemorrhage, necrosis, and/or myxoid and cystic degeneration; careful
gross examination usually shows that the tumor has an infiltrative irregular
margin. Several histologic patterns are seen. Interlacing, broad fascicles of
spindle cells resembling the herringbone pattern of adult-type fibrosarcoma
or monophasic synovial sarcoma are present in some tumors (e-Fig. 46.21).
Alternatively, the tumor may be composed of more primitive appearing,
a..~.46· Sottr-.e. I 71&
I' I.I II ( !-ltl Dlll'er•tlllllmmuMIIIIIIKIIImlclll Profile ol AcLII..~ F l - -

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background have been reported in LGFMS (Ann Diagn Pathol. 2011; Am

I Surg Pathol. 2007;31:1387).
3. Low grade fibromyxoid sarcoma (LGFMS}, another variant of fibrosarcoma,
presents as a painless deep soft tissue mass (intra or extramuscular) that has,
in some cases, been present for many years. Other sites of origin include
the chest wall, thoracic cavity, head and neck, and retroperitoneum. Most
tumors present in the fourth to fifth decades, but 5% of cases are diagnosed
as early as the second decade; in younger patients, LGFMS is smaller and
more superficial. Microscopically, a prominent capsule or pseudocapsule
surrounds an 8 to 10 em mass, which is composed of bland spindle cells
with alternating and blending, fibrogenic and myxoid areas, with variable
cellular density from one area to another (e-Fig. 46.23). Mitotic figures
are sparse in number. A subset of tumors (""40%) contains poorly formed
giant collagen rosettes consisting of a central hyalinized core surrounded by
a rim of fibroblasts with epithelioid features. Another small subgroup has
foci of increased cellularity and cytologic atypia of the type usually found
in intermediate grade fibrosarcomas, but the prognostic significance of this
finding has yet to be established. Both the t(7;16) and t(11 ;16) translocations
are present in LGFMS with or without giant collagen rosettes (Virchows
Arch. 2010;456:153). Fusion-gene positive tumors are EMA positive; are
immunoreactive for CD99 and BCL2; but do not express SMA, S-100, and
desmin (Arch Pathol Lab Med. 2006;130:1358).
4. Myxofibrosarcoma {MYFS) occurs over a wide age range, but has a predilec-
tion for individuals over 60 years of age; this tumor is also seen in young
adults. Most cases present in the dermis and subcutis; only about one-third
occur in the deep soft tissues of the lower (50% to 60% of cases) and upper
extremities (25% to 30%). Rare cases occur in the breast, heart, retroperi-
toneum, paratesticular tissues, and orbit. Most tumors have a myxoid and
infiltrative appearance, but more solid foci are present on gross examina-
tion. A multinodular growth pattern, variable cellularity within a myxoid
stroma, and incomplete fibrous septa that course through the tumor are
consistent microscopic features (Hum Pathol. 2004;35:612). Low-grade
tumors, generally smaller and superficial, are hypocellular with a promi-
nent myxoid matrix that contains only scattered plump or stellate tumor
cells, with a relatively low number of mitotic figures. High-grade MYFS,
generally larger and deeper, show marked cellular pleomorphism, multinu-
cleated giant cells, a high mitotic rate (with easily identified atypical mitotic
forms), and areas of solid growth (e-Fig. 46.24). The tumor cells express
vimentin, but otherwise do not have a characteristic immunophenotype.
Progression to a higher-grade sarcoma is seen in recurrent MYFS. Complex
karyotypes have been identified and are similar to those seen in leiomyosar-
coma (LMS) and undifferentiated pleomorphic sarcoma (Virchows Arch.
2010;456:201).
5. Acral myxoinflammatory fibroblastic sarcoma (inflammatory myxohyaline
tumor) is a low-grade neoplasm in most cases arising in an articular-
juxtaarticular location of the hands, feet, or digits of adult although exam-
ples in nonacral sites are seen (Ann Diagn Pathol. 2002;6:272; I Cutan
Pathol. 2008;35:192). Clinically, the tumor is a dermal-subcutaneous nod-
ule, measuring 2 to 3 em, with a multinodular architecture. Microscopically,
the tumor has a fibrous and myxoid stroma containing a mixed population
of spindled to epithelioid cells, ganglion-like cells, Reed-Sternberg-like cells,
bizarre multinucleated giant cells, and multivacuolated lipoblast-like cells.
The tumor cells are immunopositive for CD34 and CD68.
V. FIBROHISTIOCYTIC TUMORS. As the name suggests, these neoplasms are composed
of cells that have fibrohistiocytic morphology. However, electron microscopy and
immunohistochemistry have firmly established that the cells comprising these

tumors (other than the foamy macrophages) are, in fact, not histiocytes but rather
primitive mesenchymal cells, fibroblasts, and myofibroblasts. The circulating or
bone marrow fibrocyte is another potential candidate progenitor cell for this group
of tumors (Lab Invest. 2007;87:858).
A. Benign
1. Synovial tendon-based tumors. It is uncertain whether synovial tumors are
best classified as fibrohistiocytic tumors (as in the WHO classification) or
as tumors differentiating toward synovial cells. In any event, this fam-
ily of articular and extra-articular neoplasms includes giant cell tumor of
tendon sheath (GCTTS) and diffuse-type giant cell tumor. Both tumors share
translocations of chromosome 1p13 (Table 46.1), which often also involve
2q35 resulting in formation of a COL6A3-CSF1 fusion gene (Proc Nat/
Acad Sci USA. 2006;103:690).
a. GCTTS (also known as giant cell tumor of tendon sheath, localized type;
nodular tenosynovitis) is a localized tumor that arises from the synovium
of joints, bursae, and tendon sheath or adjacent tissues. The hand, and
less often the wrist, ankle, foot, knee, elbow, and hip, are various sites of
the tumo.t;, which clinically presents as a painless mass, mainly in adults.
GCTTS forms a firm lobulated yellowish-brown to tan circumscribed
mass that measures 0.5 to 4 em; erosion into adjacent bone is seen in
larger lesions. Mononuclear cells with rounded to plumb-spindled fea-
tures, foamy macrophages, siderophages, and multinucleate giant cells
with a variably prominent hyalinized stroma (e-Fig. 46.25) are the micro-
scopic elements of this multinodular neoplasm.
b. GCTTS, diffuse type (pigmented villonodular tenosynovitis} presents as an
intra-articular proliferation predominantly in the knee and hip joint,
whereas the extra-articular tumor predominantly involves the periartic-
ular soft tissues in the region of the knee and thigh (some extra-articular
tumors have been localized to muscle and subcutis). This tumor has
been reported in Noonan and NF1 syndromes. The tumor has a resem-
blance to GCTTS, although giant cells are less numerous or absent
altogether. Pseudosynovial and blood-filled pseudoalveolar spaces are
a common finding in the otherwise monotonous mononuclear prolif-
eration (e-Fig. 46.26). Mitotic activity is usually present (in rare cases
>5 mitoses per 10 HPFs), but atypical mitotic figures are absent. The
immunoprofile of CD68, MSA, and desmin expression is the same as for
GCTTS.
Classification of this neoplasm as an intermediate (locally aggressive)
tumor is based on the fact that over 45% of intra-articular and up to
50% of extra-articular tumors recur. Clearly malignant tumors are asso-
ciated with the metastasis; this subgroup has a high mitotic rate (over
20 mitoses per 10 HPFs), necrosis, and cellular atypia (Am] Surg Pathol.
1997;21: 153 ).
2. Deep benign fibrous histiocytoma is very likely the subcutaneous and deep
soft tissue counterpart to cutaneous fibrous histiocytoma-dermatofibroma.
There is a predilection for the extremities and head and neck region,
although this tumor is also seen in the retroperitoneum and mediastinum
(Am] Surg Pathol. 2008;32:354 ). The tumor is a circumscribed mass, com-
monly <4 em in greatest dimension, that displays a similar range of histo-
logic features as in the dermal-based lesions including a storiform architec-
ture of spindle cells intermixed with mononuclear and multinucleated giant
cells, hemosiderotic macrophages, xanthomatized histiocytes, and aneurys-
mal erythrocyte-filled spaces. Necrosis and frequent mitotic figures should
be noted with concern.
3. Fibrohistiocytic lesions that typically occur in the skin (see Chap. 38) are
dermal dendrocytic proliferations like the reticulohistiocytoma, Langer-
hans cell histiocytosis, and juvenile xanthogranuloma (Am] Surg Pathol.
2003;27:579; Histopathology. 201 0;56:148).
B. Intermediate (rarely metastasizing)
1. Plexiform fibrohistiocytic tumor (PFH) primarily occurs in the first three
decades of life with a preference for the extremities and head and neck
(Ann Diagn Pathol. 2007;11:313). Grossly, it is a firm, multinodular and
poorly circumscribed mass that usually measures <3 em in greatest dimen-
sion in the dermis and subcutis. Microscopically, the architectural pattern is
that of nodules or elongated groups of cells with a plexiform arrangement.
Three cell types are present in the nodules: central multinucleated giant
cells, mononuclear histiocyte-like cells, and spindled fibroblast-like cells
(e-Fig. 46.27). Since one of the cell types may dominate, an appreciation
of the overall architecture is important for diagnosis since the nodules can
have few, if any, multinucleated cells and the spindle cell foci can resemble
fibromatosis. The tumor cells express SMA and vimentin; CD68 reactivity
is also present but is confined to the giant cells and mononuclear histiocyte-
like cells. Approximately 30% to 35% of cases locally recur and <5%
metastasize to regional lymph nodes and beyond (Arch Pathol Lab Med.
2007;131:1135; Am] Dermatopathol. 2004;26:141). A histogenetic rela-
tionship between PFH and cellular neurothekeoma has been proposed since
there are overlapping microscopic and immunophenotypic features (Am]
Surg Pathol. 2009;33:905; Am] Surg Pathol. 2007;31:329).
2. Giant cell tumor of soft tissue occurs in the superficial soft tissue of the
lower and upper extremities, and occasionally at other sites. It is a cir-
cumscribed, nodular mass that has a soft, fleshy, gray to red-brown cut
surface. Individual nodules of the mass measure up to 1.5 em and are
separated by fibrous septa that contain hemosiderin-laden macrophages.
Microscopically, admixed mononuclear round to oval cells and multinucle-
ated osteoclast-like giant cells are set in a richly vascular stroma. Mitotic
activity can be brisk (up to 30 mitotic figures per 10 HPFs are often present),
but cellular pleomorphism and atypia are absent. Metaplastic bone is noted
in up to 50% of cases, and definitive foci of vascular invasion are identi-
fied in 30% of cases. The multinucleated giant cells are strongly CD68
immunopositive, whereas the mononuclear cells express SMA and vimentin
but show only focal CD68 immunoreactivity.
c. Malignant
1. Pleomorphic malignant fibrous histiocytoma (MFH)Jundifferentiated high-grade
pleomorphic sarcoma is the diagnostic term reserved for those obvious high-
grade sarcomas typically presenting in the deep soft tissues, which lack
a lineage-specific immunophenotype. Considerable reassessment and revi-
sionism have been directed toward the question of whether MFH represents
a specific tumor type or a final common pathway of high-grade sarcomas
(Am] Surg Pathol. 1992;16:213; Am] Surg Pathol. 1996;20:131; Am]
Surg Pathol. 2001;25:1030). These neoplasms are characterized by com-
plex cytogenetic rearrangements that can involve over 30% of the genome
(Virchows Arch. 2010;456:201).
The tumor preferentially arises in the extremity and trunk, including the
retroperitoneum. Individuals over 40 years of age present with a rapidly
enlarging mass, and about 5% have metastatic disease at diagnosis, usually
in the lung. The tumor measures 15 to 20 em, if not greater, and has a
white to tan-white, fleshy to fibrous cut surface, with areas of necrosis and
hemorrhage. The microscopic pattern is often complex (e-Fig. 46.28), with
field to field variation in terms of spindle cells, rounded to ovoid cells, and
multinucleated cells. Myxoid foci can resemble areas of pleomorphic LPS

but have an absence of convincing lipoblasts. One consistent microscopic
feature of the tumor is high nuclear grade with numerous atypical mitotic
figures; extensive necrosis with cystic degeneration and hemorrhage are
other common features. Some of these tumors are immunoreactive for SMA
and/or desmin, and so the differential diagnosis includes pleomorphic LMS
or myofibrosarcoma (Histopathology. 2001;38:499). The CD68 positivity
present in most cases is regarded by many as nonspecific. In the retroperi-
toneum, sarcomatoid carcinoma of the kidney, pancreas, and gallbladder
should be excluded.
2. Giant cell MFH/undifferentiated pleomorphic sarcoma with giant cells is a rare
neoplasm, which is composed of undifferentiated pleomorphic cells with
associated stromal osteoclastic giant cells (in contrast to pleomorphic mult-
inucleated tumor cells). Since it is now recognized that a prominent stromal
osteoclastic giant cell reaction is a feature of many poorly differentiated
carcinomas as well as sarcomas, putative cases of giant cell MFH should
be carefully studied to exclude the presence of an even focal definable line
of differentiation indicative of a specific carcinoma or sarcoma type. LMS
and carcinoma of the breast, pancreas, biliary tract, and urinary tract are
reported with similar osteoclast-like giant cells.
3. Inflammatory MFH/undifferentiated pleomorphic sarcoma with prominent
inflammation is a diagnosis now reserved for an undifferentiated high-grade
pleomorphic sarcoma with a prominent neutrophilic infiltrate, in addi-
tion to histiocytes, eosinophils, and xanthoma cells. Most cases arise in
the retroperitoneum. Some of these tumors may be dedifferentiated LPS
(] Pathol. 2004;203:822).
VI. SMOOTH MUSCLE TUMORS. Smooth muscle tumors are neoplasms in most cases,
but a small subset of cases are hamartomas that usually present in children. Neo-
plasms in this category are ubiquitous in distribution from the skin and soft tissues,
to visceral sites including the gastrointestinal and female reproductive tracts. In
most cases, the pathologic distinction between the benign and malignant smooth
muscle neoplasm is reasonably straightforward, with the exception of the leiomy-
oma of deep soft tissues and some tumors arising in the uterus (Histopathology.
2006;48:97). Many of the biologic features of this group of tumors are perplex-
ing. For example, superficial, often small LMSs of skin may have the same micro-
scopic features as their large deep counterparts even though the former often have
a favorable prognosis(] Am Acad Dermatol. 2002;46:477). Similarly, an entirely
benign appearing leiomyoma of the uterus can have an intravascular growth pat-
tern with a capacity to "metastasize" to the heart and lungs ( Obstet Gynecol Surv.
2010;65:189).
A. Benign
1. Angioleiomyoma (vascular leiomyoma) is a deep dermal or subcutaneous
neoplasm often associated with pain that in women typically occurs in the
lower extremities, but in men more often occurs in the head and upper
extremities (Skeletal Radiol. 2008;37:339). These tumors, usually mea-
suring 2 em or less, consist of three subtypes: a solid subtype of mature
smooth muscle cells with virtually no mitotic activity and small slit-like
vascular channels; a venous subtype composed of less compact smooth
muscle bundles, with venous-type vascular channels that blend with the
intravascular smooth muscle; and a cavernous subtype with dilated vascu-
lar channels with indistinct smooth muscle walls. The differential diagnosis
includes angiomyomatous hamartoma, which is an inguinal or popliteal
lymph node-based lesion (Ann Diagn Pathol. 2008;12:372).
B. Leiomyoma of deep soft tissue is a rare neoplasm that, as its name suggests, usu-
ally develops in deep subcutis, skeletal muscle, the pelvic retroperitoneum,
and the abdominal cavity (typically in the omentum and mesentery). This
tumor can measure in excess of 30 em. It is composed of bland appearing
highly differentiated smooth muscle cells that have minimal atypia and a very
low mitotic rate (for tumors in the extremities and intra-abdominal tumors in
males, the mitotic rate is < 1 mitosis per 50 HPFs; for peritoneal/retroperitoneal
tumors in females, the mitotic rate is ::::;5 per 50 HPFs). While degenerative
changes such as fibrosis, myxoid change, hyalinization, and calcification may
be present, necrosis is absent. The diagnosis of a deep leiomyoma should be
approached with caution and should rely on the absence of virtually any mitotic
activity and atypia (Adv Anat Pathol. 2002;9:351; Ann Diagn Pathol. 2003;
7:60).
Pilar leiomyoma and uterine smooth muscle tumors are discussed elsewhere
(see Chaps. 39 and 33, respectively). There is a substantial literature on the
cytogenetics of uterine leiomyomas, but considerably less on the extrauterine
counterparts (Cancer Genet Cytogenet. 2005;158:1).
C. Malignant
1. Leiomyosarcoma (LMS} is typically a malignancy presenting in mid-life and
beyond. There are four clinicopathologic settings: a retroperitoneal mass;
a tumor arising from a large blood vessel with a preference for veins of the
lower extremity and inferior vena cava; a subcutaneous or intramuscular
tumor of the extremities; and a dermal-based neoplasm. Uterine and intesti-
nal LMSs are not considered in this chapter, although the uterus is the most
common primary site overall for LMS (Adv Anat Pathol. 2011;18:60).
Except for cutaneous LMS, these tumors usually measure in excess of
6 em and have a trabeculated or smooth, glistening, white-gray cut surface.
High-grade tumors are accompanied by necrosis and hemorrhage. Archi-
tecturally, the tumors are composed of intersecting bundles of eosinophilic
spindle cells with elongated nuclei with blunted ends (e-Fig. 46.29). Nuclear
pleomorphism and an increased mitotic rate with atypical mitotic forms
are characteristic. Poorly differentiated LMS is a pleomorphic high-grade
sarcoma whose smooth muscle differentiation is highlighted by immunore-
activity for SMA, desmin, and h-caldesmon. Focal expression of keratin,
EMA, CD34, and S100 may be present. These tumors typically have
complex karyotypes with gains and losses across multiple chromosomes
A distinct clinicopathologic group of smooth muscle neoplasms are rec-
ognized in the immunocompromised setting; these tumors are associated
with genomic integration of Epstein-Barr virus (EBV) (] Clin Pathol. 2007;
60:1358; Am] Surg Pathol. 2006;30:75;] Cutan Pathol. 2011;38:731) and
have been described in both visceral, soft tissue and cutaneous sites. These
tumors have low-grade spindle cell features to the extent that the traditional
criteria for the diagnosis of LMS are not met in all cases, and are designated
as EBV-associated smooth muscle tumors in some cases.
VII. PERICYTIC (PERIVASCULAR} TUMORS. Tumors in this category show evidence of
myoid/contractile perivascular cell differentiation. Morphologically, they have a
tendency to grow in a circumferential perivascular pattern.
A. Benign
1. Glomus tumor usually occurs in the subungual region of the hand, wrist, and
foot, but not to the exclusion of other sites including the bone, lung, and
intestinal tract (Arch Pathol Lab Med. 2008;132:1448). Those tumors are
< 1 em in greatest dimension in most cases, and are painful with minimal
tactile stimulation or exposure to cold. Microscopically, these tumors con-
sists of a mixture of glomus cells (characterized as uniform, small, rounded
cells with eosinophilic to amphophilic cytoplasm and a central nucleus),
smooth muscle cells, and central vascular space (e-Fig. 46.30). On the basis
of the relative proportion of these three elements, there are three subtypes
of glomus tumor: solid, composed of nests of glomus cells surrounding
capillary-sized vessels; glomangioma, composed of small dusters of glomus
cells surrounding dilated vessels; and glomangiomyoma, in which there is a
transition from typical round glomus cells to elongated cells that resemble
mature smooth muscle. Glomangiomas may be multiple or familial, and are
likely vascular malformations (Arch Dermatol. 2004;140:971).
2. Myopericytoma, previously interpreted as a hemangiopericytoma, usually
arises in the subcutis of the distal extremities U Clin Pathol. 2006;59:67).
Less than 2 em in greatest dimension, this tumor is unencapsulated
but well circumscribed. Microscopically, the tumor is composed of a
densely cellular population of oval to spindle-shaped cells with eosinophilic
to amphophilic cytoplasm arranged in multilayered concentric profiles
around compressed blood vessels. Mitotic figures are inconspicuous. There
is diffuse immunopositivity for SMA and focal immunopositivity for
CD34.
B. Malignant. Malignant glomus tumor (glomangiosarcoma) is rare and seemingly
arises from a benign-appearing glomus tumor. The characteristic findings are a
visceral or subfascial origin, a size >2 em, marked nuclear atypia, and atypical
mitotic figures. However, a subset of tumors does not have all of these findings;
these cases are designated as tumors of uncertain malignant potential (Am]
Surg Pathol. 2001;25:1). The round cell type is composed of poorly differen-
tiated round cells, so SMA and pericellular type IV collagen are required for
diagnosis. A spindle cell variant has features of LMS or fibrosarcoma.
VIII. SKELETAL MUSCLE
A. Nonneoplastic disorders
1. Preparation of skeletal muscle biopsies. Histopathologic examination of
skeletal muscle biopsies, whether obtained through an open biopsy or
through a needle biopsy, continues to have a critical role in the evaluation of
patients with suspected myopathy (Curr Neuro Neurosci Rep. 2004;4:81).
One portion of the biopsy should be formalin fixed and paraffin embed-
ded according to standard laboratory protocols; another should be frozen
for enzyme histochemistry studies; and the remainder should be fixed in
glutaraldehyde for electron microscopic studies. While the histopathologic
evaluation of skeletal muscle biopsies is a key component in the evaluation
of neuromuscular disorders, and can be used to diagnose a variety of inher-
ited, inflammatory, and toxic myopathies, it should only be performed in
the context of a thorough history and clinical examination that has included
appropriate laboratory studies (including measurement of serum creatine
kinase) and electromyography. A number of inflammatory, toxic, and axial
myopathies have characteristic histopathologic findings, and immunohis-
tochemical studies can be utilized to characterize inflammatory infiltrates
when present (Autoimmunity. 2006;39:161).
2. Muscular dystrophies and other congenital myopathies are diagnosed pri-
marily on the basis of clinical and electromyographic features. Increasingly,
the histopathologic evaluation of muscle biopsies in these settings has been
supplanted by genetic testing as an increasing number of diseases has been
characterized at a genetic level (Pediatr Dev Pathol. 2006;9:427; Brain
Pathol. 2001;11:206).
3. Mitochondrial myopathies, which traditionally have been diagnosed on
the basis of the demonstration of so-called ragged-red fibers by Gomori
trichrome stain, are increasingly diagnosed on the basis of genetic testing
since the genetic abnormalities in mitochondria that underlie these disease
have recently been characterized (Submicrose Cytol Pathol. 2006;38:201;
Biosci Rep. 2007;27:23).
B. Benign
1. Rhabdomyoma by definition shows skeletal muscle differentiation, and
includes cardiac rhabdomyoma and extra-cardiac rhabdomyoma (which
has two subtypes: fetal and adult). Both cardiac and fetal rhabdomyomas
may be hamartomas rather than true neoplasms; tuberous sclerosis and
nevoid basal cell carcinoma syndrome accompany these tumors, respec-
tively (Ear Nose Throat]. 2004;83:716; Pediatrics. 2006;118:e1146; Con-
genital Heart Dis. 2011;6:183).
a. Fetal rhabdomyoma occurs almost exclusively in children under the age
of 10 years, with a predilection for the postauricular region and chest
wall (Am] Surg Pathol. 2008;32:485). The tumor forms a nodule in the
subcutis or deeper soft tissues that has a circumscribed noninfiltrative
pattern and is composed of bundles of fetal myotubes with interspersed
small immature-appearing mesenchymal cells. The classic or myxoid
pattern must be differentiated from ERMS, the latter of which has a
less well-organized pattern, hyperchromatic nuclei, and mitotic figures.
The so-called intermediate or cellular pattern (juvenile rhabdomyoma) is
characterized by skeletal muscle differentiation beyond the fetal myotube
stage.
b. Adult rhabdomyoma is solitary in 75% to 80% of cases, but in about
25% of cases is multinodular, even multicentric. There is an overwhelm-
ing predilection for the head and neck region (especially the larynx and
pharynx) (Am] Otolaryngol. 2011;32:240). Microscopically, lobules of
large uniform polygonal cells with abundant granular, vacuolated, or
eosinophilic cytoplasm; well-defined cell borders; and round nuclei with
a prominent nucleolus are present. Cytoplasmic cross striations and rod-
like inclusions are also present, in addition to abundant glycogen. Com-
plete excision is recommended, since recurrence occurs in >40% of cases
that have been incompletely excised.
c. Genital rhabdomyoma presents almost exclusively in the vagina in women
between the ages of 35 and 50 years as a solitary 1 to 3 em polyp that may
have been present for several years. Microscopically, bland, interlacing,
haphazardly arranged rounded to strap-like cells that have abundant
eosinophilic cytoplasm with cross striations, cytoplasmic glycogen, and
a centrally located round nucleus with a prominent nucleolus are present.
The tumor cells are embedded in a fibrous stroma with dilated vessels.
Local excision is curative.
c. Malignant
1. Embryonal rhabdomyosarcoma (ERMS} is the most common soft tissue sar-
coma of childhood (50% to 60% of cases), typically presents before
10 years of age, and accounts for 75% to 85% of all rhabdomyosarcomas
in children. The other types of rhabdomyosarcomas, alveolar and pleomor-
phic types, comprise 20% to 30% and 5% or less, respectively. ERMS has
a predilection for the head and neck (25% to 35% of cases) and pelvis-
genitourinary tract (30% to 40% of cases). Unlike alveolar rhabdomyosar-
coma (ARMS), a minority of ERMS presents in the peripheral soft tissues.
The three basic microscopic patterns of ERMS are botryoid, spindle, and not
otherwise specified (NOS), which proportionately account for 5%, 10%,
and 85% of cases, respectively. The botryoid and spindle subtypes have a
"superior" outcome, whereas the NOS subtype has an "intermediate" to
"poor" prognosis (Pediatr Dev Pathol. 1998;1:550).
The preoperative-pretreatment staging of childhood RMS is a multitier
system, which includes the status of the tumor before treatment utilizing a
TMN-based system (Table 46.6). The prognostically favorable sites of RMS
in children include the orbit, nonparameningeal head and neck, common
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neuroectodermal tumor (EWSIPNET) and malignant rhabdoid tumor.

Where there is stroma, the tumor cells are larger, more compact, and more
likely to have bright eosinophilic cytoplasm. In many tumors, the fact that
no two high magnification microscopic fields have identical features reflects
the polymorphous character of many ERMS. Diffuse and/or lobular growth
patterns are seen either as a dominant or as a mixed feature. Those tumors
composed predominately of spindle cells resemble a LMS or fibrosarcoma,
but with immature rhabdomyoblasts present among the spindle cells.
A difficult assessment is the determination of "viable" tumor in a
posttreatment biopsy. By convention, if the biopsy contains tumor cells
with features seen in the pretreatment biopsy, residual viable tumor is the
appropriate interpretation. Differentiated rhabdomyoblasts with negative
Ki-67 (MIB-1) staining are considered nonproliferative and nonviable.
The immunophenotype correlates with the degree of skeletal muscle dif-
ferentiation. Primitive ERMS may show only vimentin expression, although
myoD1 and/or myogenin nuclear positivity is often present, but in a minor-
ity of the tumor cells. Most ERMSs are positive for desmin in 15% to 50%
of tumor cells, whereas myogenin tends to display a bright nuclear signal in
15% to 30% of cells in contrast to ARMS in which myogenin shows near
100% nuclear positivity (Am I Surg Pathol. 2008;32: 1513 ). Differentiating
rhabdomyoblasts show desmin and/or MSA expression, while differentiated
cells also are immunoreactive for myosin.
2. Alveolar rhabdomyosarcoma (ARMS), like EWS/PNET and hematolymphoid
malignancies, is one of the quintessential malignant round cell neoplasms
of childhood. Most cases are diagnosed in adolescents and young adults,
although ARMS is seen infrequently in infants (Cancer. 2011;117:3493).
Primary sites include the extremities, paraspinal and perineal regions, and
the paranasal sinuses. Microscopically, the tumor has two general patterns:
solid sheets of uniform high-grade round cells without extracellular mucin,
and the classic alveolar pattern with tumor cells attached to incomplete
fibrovascular septa dividing the loosely arranged cells into nests (e-Fig.
46.32). Both patterns can be seen in metastatic ARMS (the initial encounter
with the tumor may be in a lymph node or bone metastasis since ARMS may
present with disseminated metastatic disease). Most ARMSs are strongly
and diffusely immunopositive for desmin and myogenin (Pathol Oncol Res.
2008;14:233 ). ALK protein overexpression is found in 40% to 50% of cases
(Pediatr Dev Pathol. 2009;12:275).
The demonstration of the FOX01 (FKHR) breakapart by FISH is an
important adjunct in the diagnosis of ARMS, with a positive result in
approximately 75% to 80% of cases (Am I Pathol. 2009;174:550). The
PAX3-FO XO1 fusion gene (resulting from t(2; 13)) is more common (present
in 70% to 80% of fusion positive cases) than the PAX7-FOX01 fusion
gene (resulting from t(1;13)). Fusion-negative ARMS has a gene profile and
prognosis more like ERMS, but the significance of this observation remains
unsettled U Clin Oncol. 2010;28:2151) since specific fusion type and pat-
tern of ARMS do not appear to correlate with outcome. Rarely RMS has
a mixed pattern of ARMS and ERMS; these tumors represent a diagnostic
dilemma, which may be resolved by FISH to determine the fusion status of
the tumor.
3. Pleomorphic rhabdomyosarcoma is a high-grade sarcoma that presents in the
deep soft tissues (Am I Surg Pathol. 2009;33:1850). These rare neoplasms
occur overwhelmingly in adults. A poorly differentiated pleomorphic sar-
coma resembling pleomorphic undifferentiated sarcoma and/or an equally
high-grade spindle cell sarcoma are the histologic features. The eosinophilic
cytoplasm may or may not have cross striations. Both desmin and
myogenin are positive; pancytokeratin and CD34 are reactive in a minority

of cases.
4. Sclerosing rhabdomyosarcoma, a rare putative variant of rhabdomyosar-
coma, may have a pattern of gene amplification of chromosome 12q more
like LPS (Hum Pathol. 2009;40:1347).
5. Ectomesenchymoma is a rare neoplasm that occurs in children for the most
part. It may have ERMS- or ARMS-like features, with populations of cells
with a range of neural differentiation, from ganglion cells to more primitive
neuroectodermal cells that express a variety of neural and myogenic mark-
ers. This tumor has a predilection for the head and neck including the CNS
(Pediatr Dev Pathol. 2000;3:290; I Neurosurg. 2011;7:94). These tumors
seem to bridge the molecular interface between RMS and MPNST (Diagn
Mol Pathol. 2007;16:243; Acta Neuropathol. 2007;113:695).
IX. VASCULAR TUMORS. Vascular or vasoformative lesions constitute a pathogeneti-
cally heterogenous group of lesions composed of endothelial-lined vascular spaces
with a circumscribed or more diffuse pattern of growth. These lesions are solitary
or less often multifocal. They may be initially noted at or shortly after birth or
become clinically evident in later childhood or adulthood. The skin and under-
lying soft tissues are the most common sites of presentation, but there are few
anatomic sites or organs that are excluded from involvement (Am I Surg Pathol.
2010;34:942). In terms of pathogenesis, vascular lesions are reactive (etiology
either known or unknown), malformative, or neoplastic.
A. Inflammatory lesions. Inflammatory disorders of the vessels, including the
vasculitides, are covered in the chapter on the cardiovascular system (see
Chap. 9).
B. Reactive vascular proliferations. This heading includes lesions that are gener-
ally thought to be reactive and nonneoplastic, although many mimic vascular
tumors clinicopathologically.
1. Papillary endothelial hyperplasia {PEH, Masson vegetant hemangioma) is usu-
ally subdivided into a primary type (occurring in a vein in the head and
neck or fingers) and a secondary type (arising in a preexisting heman-
gioma, hemorridal vein, or thrombohematoma) (Auris Nasus Larynx.
2009;36:363 ). Microscopically, numerous, small, delicate papillae with
hyaline cores project into a vascular lumen (e-Fig. 46.33). Often there is
a transition between an organizing fibrin clot and the PEH. A single layer
of plump endothelial cells without appreciable cytologic atypia is charac-
teristic, and helps differentiate it from angiosarcoma, which does not arise
from a blood vessel or hematoma. With continued organization of the clot,
the papillae fuse and form an anastomosing network of vessels with even-
tual recanalization.
2. Glomeruloid hemangioma is an uncommon, multifocal, primarily intravas-
cular capillary proliferation that is associated with Castleman disease and
the POEMS (polyneuropathy, organomegaly, endocrinopathy, M-protein,
and skin changes) syndrome. Glomeruloid capillary proliferations within
blood vessels of the dermis, deep soft tissue, or viscera are the micro-
scopic features. Glomeruloid capillary tufts have an outer layer of peri-
cytes but are lined by bland endothelial cells with or without cytoplasmic
vacuolation.
3. Lobular capillary hemangioma (LCH) is one of the more common vascular
lesions which is seen primarily in the skin where satellite lesions may occur,
but the anatomic distribution is widespread in the head and neck (nasal and
oral cavities), liver, and gastrointestinal tract. Microscopically, the lesion is
pedunculated with a collarette of epidermis and is composed of lobular
arrays of uniform capillary-sized vessels with a central thick-walled feeder
vessel (e-Fig. 46.34). Ulceration of the epidermis with inflammation and
hemorrhage can obscure the underlying pathology, with superim-

posed granulation tissue U Eur Acad Dematol Venereal. 2001;15:106).
Extramedullary hematopoiesis may be seen. Other presentations include as
a more dermal-based tumor producing a nodule rather than a polyp, and as
an intravascular process. Lesions with more slit-like vascular spaces with a
retained lobular architecture can resemble Kaposi sarcoma.
4. Bacillary angiomatosis has a close clinical and pathologic resemblance to
LCH, but also features an interstitial neutrophilic infiltrate and amorphous
deposits of Warthin-Starry positive aggregates of Bartonella henselae or B.
quintana. Virtually all cases of bacillary angiomatosis occur in immunocom-
promised individuals (Am] Dermatopathol. 2011;33:513; Clin Dermatol.
2009;27:271).
5. Vascular transformation of lymph nodes (nodal angiomatosis) occurs as a
secondary change in lymph nodes due to lymphatic and/or venous obstruc-
tion. Ectatic capillary-sized vessels within the subcapsular space and nodal
sinuses are the microscopic features. The differential diagnosis is Kaposi
sarcoma.
C. Benign
1. Hemangioma is the most common cutaneous and soft tissue tumor of infancy
and childhood, and accounts for "'7% of benign STTs overall. Although
most hemangiomas are superficially located, the liver and parotid gland
may be involved in infants UPediatr Surg. 2007;42:62; Ann Diagn Pathol.
2002;6:339). There has been substantial revision in the classification of
vasoformative lesions into vascular neoplasms or malformations based in
part on glucose transporter 1 (GLUf-1) immunoreactivity in the major-
ity of vascular neoplasms and its absence in malformations (Hum Pathol.
2000;31:11; Clin Plast Surg. 2005;32:99).
a. Capillary hemangioma is the most common type of hemangioma and has
a predilection for the head and neck. It is usually diagnosed in infants
and young children, hence the designation infantile hemangioma. These
lesions have rapid proliferative and prolonged involutional stages; the
latter can continue into early to late childhood. The tumor is clinically
a purple to reddish macule or nodule centered in the skin or subcuta-
neous tissue. Microscopically, cellular lobules during the proliferative
phase may have few identifiable vascular lumina and readily identifiable
mitotic figures. With involution, the vascular spaces are well defined,
mitoses are absent, and a collagenous stroma becomes more prominent
in the interstitium. The current classification of infantile hemangioma
as a benign vasoformative lesion reflects the observation that GLUf-1
immunoreactivity is present in the majority of cases (Clin Plast Surg.
2011;38:31 ).
b. Congenital hemangioma is present at birth and has two subtypes, invo-
lution congenital hemangioma and noninvoluting congenital heman-
gioma (]Am Acad Dermatol. 2004;50:875; Cardiovasc Pathol. 2006;
303:2006). The two subtypes have distinctive microscopic features and
are GLUf-1 immunonegative unlike infantile hemangioma (Arch Facial
Plast Surg. 2005;7:307).
c. Tufted angioma typically presents in infancy and is characterized by
scattered cellular vascular nodules associated with variably prominent
dilated lymphatic spaces in the dermis and subcutis (Arch Dermatol.
2010;146:758). The endothelial cells may be spindled as in the related
kaposiform hemangioendothelioma (KHE), or more epithelioid in
appearance (e-Fig. 46.35). The tumors have in common expression for
D2-40 and may be complicated by Kasabach-Merritt syndrome. Neither
tufted angioma or KHE is GLUT-1 positive.
d. Verrucous hemangioma is composed of a mixture of cavernous and capil-

lary vessels immediately beneath a hyperkeratotic, acanthotic epidermis.
e. Cherry angioma is suspected to represent an involuted LCH (] Cutan
Pathol. 2011;38:740).
f. Cavernous hemangioma (likely a vascular malformation) occurs in the
same age range and anatomic distribution as infantile hemangioma, but
is less common; shows virtually no tendency to regress; and may be
locally destructive due to extrinsic pressure on adjacent structures. A pat-
tern of grouped, dilated, thin-walled blood vessels with an inconspicuous
endothelial lining is the microscopic appearance. The blue-rubber-bleb
nevus syndrome (characterized by cavernous hemangiomas of the skin
and gastrointestinal tract) and Maffucci syndrome (cavernous heman-
giomas and enchondromas) are two associated syndromes.
g. Arteriovenous hemangioma, also known as arteriovenous malformation,
is associated with arteriovenous shunts. It is found more commonly in
the skin than in the deep soft tissue where it is composed of a mixture
of arterial vessels and thick-walled veins with variable circumscription.
Focal vascular thrombosis, PEH, and dystrophic calcifications may be
present.
h. Venous hemangioma (venous malformation) typically presents during
adulthood in the deep soft tissue. Ectatic vessels often show thrombosis,
PEH, and dystrophic calcification.
i. Spindle cell hemangioma occurs in young adults and is found in the skin
and subcutis of the distal extremities, especially the hand. Thin-walled
cavernous vessels lined by bland flattened endothelium are admixed with
solid areas composed of plump spindle cells resembling Kaposi sarcoma.
The tumor involves a large preexisting vessel in many cases. Recurrences
are common (>50% of cases), often with a discontinuous growth pat-
tern. This lesion should be distinguished from KHE, which has spindled
endothelial cells.
j. Synovial hemangioma, as the name implies, arises in synovial-lined spaces,
most commonly in the knee in the second decade of life. Microscopi-
cally, variably sized thin-walled vascular spaces occupy the stroma of
hyperplastic synovium with associated marked hemosiderin deposition.
The differential diagnosis includes pigmented villonodular synovitis and
trauma-associated hemarthrosis as in hemophilia.
k. Intramuscular angioma (intramuscular hemangioma, skeletal muscle
hemangioma) most frequently arises in the lower extremity, particu-
larly the muscles of the thigh. It is a poorly circumscribed and diffusely
infiltrating mass in the muscle composed of variably sized vessels rang-
ing from large thick-walled veins to cavernous vascular spaces, small
arteries, and capillaries, and thus has features more in keeping with an
arterial-venous malformation than a true neoplasm. Despite the presence
of mitotic activity and intraluminal capillary tufting, freely anastomos-
ing vascular channels are not present. Mature adipose tissue may be a
component of the intramuscular mass.
2. Epithelioid hemangioma occurs mainly in females between 20 and 40 years
of age as a small, dull erythematous plaque of the head and neck. Micro-
scopically it is a well-circumscribed nodule in the dermis or subcutis, or
less frequently in the deep soft tissue, that has a vague lobular pattern of
clustered small capillary-sized vessels around a feeder vessel. The endothe-
lial cells are plump and have abundant cytoplasm with impingement upon
the lumen of the vascular channel (so-called tombstone appearance). The
neoplasm occurs rarely in the skeletal system and penis (Am I Surg Pathol.
2004;28:523; Am I Surg Pathol. 2009;33:270). When lymphocytes and
eosinophils are prominent, the lesion is an example of angiomatoid hyper-

plasia with eosinophilia(] Cutan Pathol. 2010;37:1045).
3. Angiomatosis is, by definition, a poorly circumscribed, diffuse network
of vascular structures within a soft tissue site. In virtually all cases it
presents during childhood or adolescence as diffuse soft tissue swelling.
The demonstration that the vessels are GLUT-1 immunonegative is con-
sistent with the interpretation that angiomatosis is a malformation. There
are two histologic patterns: mixed-vessel-type lesions resembling intramus-
cular angiomas, and capillary-predominant type lesions with a lobular
pattern. The various syndromic associations include Klippel-Trenaunay-
Weber, Parkes-Weber, Bannayan-Riley-Ruvalcaba, Maffucci, and Proteus
syndromes (Semin Musculoskelet Radio/. 2009;13:255).
4. Lymphangioma is a cavernous and/or cystic vascular lesion composed of
dilated lymphatic channels that occurs most commonly in the head and neck
of young children (e.g., a hygroma). Thin-walled dilatedlymphaticvessels of
varying size are lined by flattened endothelium that is strongly immunopos-
itive with the D2-40 antibody (Histopathology. 2005;46:396). Scattered
lymphoid aggregates may lie beneath the endothelium (e-Fig. 46.36). Lym-
phangioma is regarded as a lymphatic malformation.
5. Atypical vascular lesion of the breast presents as papules, nodules, or ery-
thema several years (average 5 years, range 2 to 20 years) after external
radiation for carcinoma of the breast. This lesion is composed of thin-walled
vascular spaces resembling lymphatics and is confined to the superficial der-
mis. An infiltrating, more diffuse pattern into the deeper dermis and atypia
of endothelial cells should be viewed with concern (]Am Acad Dermatol.
2007;57: 126).
6. Several other lesions have been added to the seemingly ever expanding uni-
verse of vascular tumors. Epithelioid angiomatous nodule presents as one,
or less often multiple, nodules with a wide distribution (Pathol Res Pract.
2009;205:753). The cellular nodules occupy the dermis and are composed
of epithelioid endothelial cells. Some mitoses are present, and overall the
lesion has features of an epithelioid hemangioma. Pseudomyogenic heman-
gioendothelioma arises in the extremities, for the most part as a dermal-
subcutaneous or deep soft tissue mass. Plump spindle cells with prominent
eosinophilic cytoplasm are arranged in loose or storiform fascicles (Am ]
Surg Pathol. 2011;35:190). Epithelioid cells are seen as a minor compo-
nent, as are neutrophils. The tumor is immunopositive for AE1/AE3, FU1,
and CD31 expression. Polymorphous hemangioendothelioma is a lymph
node-based vascular neoplasm (Am] Surg Pathol. 1997;21:1083). Com-
posite hemangioendothelioma is, as its name implies, a vascular neoplasm
with epithelioid, spindle cell, and angiosarcoma-like features (Pathology.
2011;43:176).
D. Intermediate (locally aggressive}
1. Kaposiform hemangioendothelioma (KHE}, a rare locally aggressive neoplasm,
presents most commonly in the first year of life in the extremities or
trunk (75% of cases) or retroperitoneum (20% of cases) (Bur] Int Med.
2009;20:106). One or multiple masses, or a more diffusely infiltrative pro-
cess, are the various clinical manifestations. As the designation KHE implies,
the individual lobules consist of a mixture of capillary-sized vessels that
blend with more slit-like spindled vessels, with an absence of mitoses and
an absence cytologic atypia (Am] Surg Pathol. 2004;28:559). KHE is
immunoreactive for D2-40 (Am] Surg Pathol. 2010;34:1563). Approx-
imately 20% of cases are complicated by the Kasabach-Merritt syndrome.
A possibly related entity with a similar coagulopathy is multiple lymphan-
gioendotheliomatosis (Arch Dermatol. 2004;140:599).
Chapter 46 • Soft Tissue I 769
E. Intermediate (rarely metastasizing)

1. Retiform hemangioendothelioma, an uncommon neoplasm, arises in the skin
of the distal extremity or trunk in young to middle-aged adults, and has a
recurrence rate of 40% to 50% without wide excision. Grossly, the tumor
is a reddish-purple slowly growing plaque centered in the reticular dermis,
usually less than 2 to 3 em in maximal dimension. Microscopically, the
tumor is characterized by elongated, arborizing, narrow vessels that resem-
ble the rete testis. The endothelial cells are monomorphic with a low mitotic
rate, hyperchromatic nuclei, and hobnail morphology. The stroma between
the vascular channels is prominent and often shows a pronounced lympho-
cytic infiltrate, but unlike angiosarcoma, the vascular spaces do not dissect
through the dermal collagen. The tumor is immunopositive for CD31, but
not for D2-40 or VEGFR-3 (Am J Dermatopathol. 2008;30:31).
2. Papillary intralymphatic angioendathelioma (Dabska tumor) occurs in infants
and children in the skin (reticular dermis) as a slowly growing nodule
or plaque without a site predilection (Dermatology. 2000;201:1). Thin-
walled vascular spaces contain intraluminal papillary tufts of endothelial
cells with a hobnail morphology as in retiform hemangioendothelioma.
Papillary intralymphatic angioendothelioma (as well as retiform heman-
gioendothelioma) have been reported in bone (Orthopedics. 2004;27:327;
Pathollnt. 2011;61:382).
3. Kaposi sarcoma is a low-grade donal endothelial proliferation caused by
human herpesvirus 8 (I-lliV8) infections. The clinicopathologic features
of Kaposi sarcoma are discussed in more detail in the chapter on non-
melanocytic tumors of the skin (Chap. 39).
F. Malignant
1. Epithelioid hemangioendothelioma (intravascular bronchioloalveolar tumor)
is a low-grade malignant endothelial neoplasm that occurs in patients of
all ages, although it is rare in early childhood. The superficial and deep
soft tissues, viscera (liver), and bone are various primary sites. About
10% of patients have multiorgan disease at the time of presentation. A
t(1;13) translocation that produces a WWTR1-CAMTA1 gene fusion has
recently been shown to be a characteristic feature of the tumor, regardless
of anatomic site (Genes Chromosomes Cancer. 2011;50:44).
Grossly, the tumor presents as a poorly circumscribed multilobular infil-
trative mass, measuring up to 10 em in greatest dimension. Microscopi-
cally, the tumor consists of cords, short strands, solid nests, or individual
cells that have rounded to slightly spindled features (e-Fig. 46.37). The cells
are low grade with a low mitotic rate. Endothelial differentiation is evident
by the formation of intracytoplasmic lumina (producing signet ring-like fea-
tures) but distinct endothelial-lined vascular channels are not prominent.
The neoplastic cells are embedded within a chondroid-like to hyalinized
stroma. Atypical morphologic features, including an increased mitotic rate,
increased nuclear pleomorphism, more spindled cytology, and necrosis cor-
relate with more aggressive behavior. The tumor cells express CD31, CD34,
and Ulex europaeus antigen, and variably express factor Xlll-related anti-
gen. Of note, 25% to 30% of tumors show focal cytokeratin expression
which can lead to an incorrect diagnosis of metastatic signet ring cell car-
cinoma. The distinction from an epithelioid hemangioma is not obvious in
all cases.
2. Angiosarcoma, a rare malignancy composed of endothelial cells, recapitu-
lates the functional and morphologic features of normal endothelium to a
variable degree (Lancet Oncol. 2010;11:983). The tumor is divided into
several groups: cutaneous angiosarcoma (with or without an association
with lymphedema-Stewart-Treves syndrome); angiosarcoma of the breast;
radiation-induced angiosarcoma; and angiosarcoma of the deep soft tissue.

The malignancy is a poorly circumscribed hemorrhagic mass that mea-
sures from 1 to 2 em to > 10 em, with histologic features that vary from
hemangioma-like (but with scattered, enlarged, atypical endothelial cells
with occasional mitotic figures and an infiltrating growth pattern) to a high-
grade spindle cell sarcoma with a hemorrhagic background (e-Fig. 46.38).
Intraluminal papillae lined by hyperchromatic cells are sometimes present.
Vascular channels with a dissecting pattern of infiltration through the der-
mis or other surrounding tissues should be viewed with concern (see atypical
vascular lesion of breast discussed above).
Epithelioid angiosarcoma, a well-recognized variant of angiosarcoma
in the deep soft tissues, is composed of malignant epithelioid cells with
abundant eosinophilic or amphophilic cytoplasm, large vesicular nuclei, and
prominent nuclei (e-Fig. 46.39). In sites such as a pleura and peritoneum, the
initial impression may be malignant mesothelioma or metastatic carcinoma
(Arch Pathol Lab Med. 2011;135:268).
X. CHDNDRD-DSSEDUS TUMORS
A. Benign
1. Chondroma occurs over a broad age range usually in the fingers and toes,
with a juxtaarticular and tendinous predilection. Chondromas are typically
composed of lobules of mature hyaline cartilage, but there are histologic
variants: chondroblastic chondroma (when the lesion is cellular), fibrochon-
droma (when there is prominent fibrosis), and osteochondroma (when there
is prominent ossification). Chondromyxoid fibroma and chondroblastoma
typically arise in bone but have been reported in juxtacortical or soft tissue
sites (Am] Surg Pathol. 2007;31:1662; Diagn Cytopathol. 2003;28:76).
If the chondroid tumor is from the base of the skull, chondroid chordoma
should be considered in the differential diagnosis.
B. Malignant
1. Mesenchymal chondrosarcoma is a neoplasm of the soft tissues or other
extraosseous sites (25% to 30% of cases) or bone (70% to 75% ), with
some preference for the head and neck (meninges, orbit), spine, and
lower extremities. Most cases are diagnosed between 15 and 40 years
of age, but the tumor may be seen in the neonate (Pediatr Dev Pathol.
2008;11:309). Microscopically, nodules of neoplastic hyaline cartilage are
separated by undifferentiated malignant small round cells, with or without
a hemangiopericytoma-like appearance. The tumor cells are immunopos-
itive for vimentin, desmin, EMA, and SOX9, and focally also express
.B-catenin (Hum Pathol. 2010;41:653; Ann Diagn Pathol. 2010;14:8). The
tumor cells are nonreactive for FLil (Appl Immunohistochem Mol Mor-
pho/. 2011;19:233).
2. Extraskeletal osteosarcoma usually arises in the deep soft tissue, most com-
monly in the thigh, although the buttock, shoulder girdle, trunk, and
retroperitoneum are other sites in individuals between 40 and 60 years of
age(] Thorac Oncol. 2009;4:927). About 10% of patients have a history
of prior radiation or trauma to the site. Malignant bone is usually most
prominent in the center of the tumor, while more peripheral regions of the
neoplasm tend to be more densely cellular (this zonation is the reverse of
the pattern present in myositis ossificans and can be a useful feature for
differential diagnosis).
XI. PERIPHERAL NERVE TUMORS. The tumors of presumed peripheral nerve or nerve
sheath derivation comprise some of the more common soft tissue neoplasms in
routine practice.
A. Traumatic neuroma is a nonneoplastic proliferation in response to nerve injury,
often after a surgical procedure. It is a firm, tender nodule measuring <5 em
Chapter 46 • Soft Tissue I 771
that consists of small crowded nerve fascicles arranged in a haphazard architec-

tural pattern, that are composed of Schwann cells and fibroblasts in a fibromyx-
oid stroma (e-Fig. 46.40).
B. Mucosal neuromas on the lips, mouth, eyelids, and intestines are manifesta-
tions of multiple endocrine neoplasia llB. The irregular nerve bundles in the
submucosa have a prominent perineurium. The stroma often has a myxoid
quality.
C. Neurofibromas are divided into three types on the basis of a growth pattern:
localized, diffuse, and plexiform.
1. Localized neurofibroma is sporadic, usually superficial and solitary, and unas-
sociated with a genetic syndrome. The dermis and subcutis are the sites
of predilection. Microscopically, the circumscribed nodule is composed of
spindle cells with wavy nuclei and strands of collagen in a neurofibrillary
background with scattered mast cells. The differential diagnosis is a dermal
melanocytic nevus with extensive neurotization.
2. Plexiform neurofibromas are manifestations of neurofibromatosis 1 (NF1),
as are diffuse neurofibromas (Lancet Neurol. 2007;6:340). These tumors
develop in early childhood as superficial soft tissue masses, and vary in
size and extent of local involvement. An entire length of nerve can be trans-
formed into the so-called bag-of-worms appearance in which multiple nerve
bundles characteristically show expansion of the endoneurium by a myxoid
stroma, with later extension beyond the perineurium into the adjacent soft
tissue (e-Fig. 46.41). The interstitium is composed of thickened irregular
fibrous bundles with interspersed short to fusiform spindle cells, and a dif-
fuse pattern may accompany the plexiform component in the surrounding
soft tissues. Foci of increased cellularity with associated enlarged, hyper-
chromatic nuclei should be viewed with concern for malignant transforma-
tion (e-Fig. 46.42). Nuclear reactivity for p53 is an additional finding of
concern.
3. Diffuse neurofibroma has a predilection for the head and neck as a plaque-
like elevation of the skin. The dermis and subcutis are effaced by a uni-
formly cellular proliferation within a fibrillary collagenous matrix contain-
ing Schwann cells with short uniform nuclei (e-Fig. 46.43) and occasional
Meissner body-like formations. Some tumors are associated with other mes-
enchymal elements including ectatic vessels and mature entrapped adipose
tissue. The overgrowth of subcutaneous fat is similar to that seen in der-
matofibrosarcoma protuberans and infantile subcutaneous fibromatosis.
D. Schwannoma (neurilemoma) most often occurs in patients between 20 and
40 years old, but is also seen in children. Most schwannomas present as a
solitary mass in the head and neck, on the flexor surfaces of the upper and
lower extremities, or in association with spinal and paraspinal sensory nerves.
Multiple or bilateral schwannomas are manifestations of NF2 and schwanno-
matosis (Annu Rev Pathol. 2007;2:191; Rev Neurol Dis. 2009;6:E81). Cellular
and plexiform schwannomas may occur in children and in NF2; pigmented or
melanotic schwannomas are one of the features of the Carney complex (Ann
Endocrinol. (Paris) 2010;71:486).
The tumors vary in size, but virtually all are invested by a true capsule
of the epineurium which can be difficult to demonstrate in some sites in the
head and neck. A white to yellow-white mucoid cut surface with or without
cystic degeneration and hemorrhage are the gross features. Most tumors are
<5 em in greatest dimension, although those that arise in the paraspinal
retroperitoneum can be larger. The characteristic low-power microscopic pat-
tern (e-Fig. 46.44) consists of alternating Antoni A areas (consisting of orga-
nized spindle cells with (e-Fig. 46.45) or without Verocay bodies) and Antoni
B areas (consisting of less cellular regions in which the oval to spindled cells are
arranged more haphazardly in a loose fibrous matrix). Thickened hyalinized

vessels are prominent in many cases. There is considerable histologic variability
in schwannomas; despite atypical findings, especially in so-called ancient-type
schwannomas, these tumors rarely undergo malignant transformation (char-
acterized by invasion through the capsule, epithelioid or small cell transfor-
mation, high-grade nuclear abnormalities, and atypical mitotic figures). It is
noteworthy that epithelioid morphology alone is an insufficient criterion for
malignancy because there is an epithelioid variant of schwannoma. The tumor
cells are diffusely immunopositive for S-1 00 protein and collagen type IY, with
delicate pericellular reactivity reflecting basement membrane-like deposition.
E. Granular cell tumor occurs in virtually all age ranges, although more frequently
in adults than children. There is a predilection for the skin and sites in the head
and neck, but this tumor occurs throughout the body. One unique type, the
congenital epulis of the anterior gingival region in neonates, does not express
S-100 protein (nonneural granular cell tumor). Several histologic patterns are
seen, including closely apposed nests and ribbons, infiltrating cords, and con-
fluent sheets of uniform polygonal to slightly spindled cells with a central
nucleus that is surrounded by abundant uniform granular cytoplasm (e-Fig.
46.46). Pseudoepitheliomatous hyperplasia may be present in the overlying
epidermis or squamous mucosa. Malignancy is rare, but is well documented
(Am I Surg Pathol. 1998;22:779). Strong immunoreactivity for S-100, neuron
specific enolase, and CD68 is the usual phenotype of the neural granular cell
tumor.
F. Neurothekeoma (nerve sheath myxoma), a rare tumor in children and young
adults, presents in the head, neck, and shoulders. Microscopically, nodules
of bland, plump spindle cells are present in a myxoid matrix separated by
fibrous connective tissue (e-Fig. 46.47). The cells show little pleomorphism
and have a low mitotic rate. Increased cellularity, cytologic atypia, increased
mitotic figures, extension into adjacent or deep soft tissues, and even vascular
invasion are regarded as atypical variants (Am I Surg Pathol. 1998;22:1067).
G. Perineuriama is an uncommon neoplasm composed of perineural cells whose
expression of epithelial membrane antigen is similar to meningothelial cells.
The two types of perineurioma are those that arise within or external to a
nerve (Arch Pathol Lab Med. 2007;131:625). The tumor has several histo-
logic patterns including a lacy and reticulated network of delicate to somewhat
plump fusiform spindle cells, and a more cellular pattern of storiform perivas-
cular spindle cells. The background may be sclerotic or more typically myxoid.
There are now several examples of peripheral nerve sheath neoplasms which
have hybrid features of schwannoma and perineurioma (Am I Surg Pathol.
2009;33: 1554); these tumors arise in the peripheral soft tissues and even within
lymph nodes (Pediatr Dev Pathol. 2011 ).
H. Malignant peripheral nerve sheath tumor (MPNST) that arise from peripheral
nerves are currently classified as MPNSTs. A sarcoma can be classified as an
MPNST if it has arisen from a peripheral nerve, if it has arisen from a pre-
existing benign nerve sheath tumor (in most cases, a neurofibroma), or if it
demonstrates histologic and immunophenotype features that reflect Schwan-
nian differentiation. Overall, 5% to 10% of STSs are MPNSTs, especially in
late adolescence and early adulthood.
These tumors are typically large (in excess of 5 em) with a fleshy mucoid cut
surface and large areas of hemorrhage and necrosis. Microscopically, they are
composed of fascicles or broad sweeping profiles of malignant spindle cells that
have wavy or comma-shaped nuclei. The cellular density often varies producing
a "light-cell, dark-cell" appearance (e-Fig. 46.48). Nuclear palisading is present
in some tumors, as are hyalinized cords and nodules that may resemble large
rosettes. The immunophenotype tends toward Schwannian differentiation with
S-100 positivity (only in 50% of cases), but the tumors are more commonly
positive for CD57 (Leu 7), collagen type N, and myelin basic protein (Arch
Pathol Lab Med. 2009;133:1370). Chromosomal losses are more common
than amplifications (Virchows Arch. 2010;456:201).
Variant patterns of MPNST show rhabdomyoblastic differentiation (malig-
nant triton tumor), glandular formation, and heterotopic bone and/or cartilage
formation. These variants are encountered with some frequency in the setting
of NF1 (the setting in which 40% to 50% of all MPNSTs are seen) (Neurosurg
Clin N Am. 2008;19:533).
XII. TUMORS OF UNCERTAIN DIFFERENTIATION. For many of the tumors in this category,
either the cell type that produces the tumor or the normal cell type that the tumor
is recapitulating, or both, is unknown.
A. Benign
1. Intramuscular myxoma occurs primarily in adults between 40 and 60 years
as a solitary, painless, slowly growing mass within the muscle of the thigh,
buttocks, or limb girdle. Most tumors measure 5 em in maximal dimen-
sion, and are well circumscribed with a pale gelatinous or myxoid cut sur-
face. The tumor cells are small, bland, stellate to spindle in shape, have
a very low mitotic rate, and are present in an abundant myxoid matrix
that contains an inconspicuous vascular network (e-Fig. 46.49). Focal areas
of increased cellularity and vascularity with collagenous background are
present in some cases whose presence may cause concern (Am I Surg Pathol.
1998;22:1222). Simple excision is followed by a recurrence rate of <5%
of cases (Histopathology. 2001;39:287). The differential diagnosis of myx-
oma includes many other myxoid tumors of the soft tissues (Histopathology.
1999;35:291; Ann Diagn Pathol. 2000;4:99). One or more myxomas of the
soft tissues with fibrous dysplasia defines Mazabraud syndrome. Multiple
cutaneous and mucosal myxomas are manifestations of Carney complex
(Lancet Oneal. 2005;6:501; Neuroendocrinology. 2006;83:189).
2. Juxtaarticular myxoma, as the name suggests, usually arises in the vicinity of
a large joint (primarily the knee). The lesion has a predilection for males
between 20 and 40 years of age, and is usually <5 em maximal dimension.
This tumor is indistinguishable from an intramuscular myxoma, but is prob-
ably a different entity than the latter (Virchows Arch. 2002;440:12). Some
cases may have hemorrhage and hemosiderin deposition, chronic inflam-
mation, and fibrosis.
3. Digital myxoma (digital mucus cyst), usually occurs on a finger, and in adult
women as a painful nodule < 1 em in diameter. Histologically, it resem-
bles intramuscular myxoma. Cutaneous myxoma (also known as superficial
angiomyxoma) is a cutaneous or subcutaneous lesion which occurs primar-
ily in adults between 30 and 50 years of age; it must be differentiated from
nerve sheath myxoma of peripheral nerve sheath origin and neurothekeoma
(Mod Pathol. 2011;24:343).
4. Deep aggressive angiamyxoma is a slowly growing locally infiltrative tumor
that usually occurs in the pelviperineal soft tissue of women, but is also
seen as a scrotal mass (Int I Gynecol Pathol. 2005;24:26; IntI Urol.
2003;10:672; Histopathology. 2009;54:156). It forms a soft poorly cir-
cumscribed mass measuring 10 to 20 em in greatest dimension that has a
gelatinous cut surface. Small stellate to spindled cells with ill-defined cyto-
plasm and bland nuclei are set in a myxoid stroma rich in collagen fibers
(e-Fig. 46.50). Numerous, variably sized, thick or thin walled vascular chan-
nels are present, from which smooth muscle seems to spin off and merge
with the surrounding stroma. The tumor cells express vimentin and desmin,
and also usually show nuclear expression of estrogen and progesterone
receptors.
The tumor has a propensity to recur, which is not unexpected given

its infiltrative and poorly defined margin. Local control is usually possible
without radical surgery, and the tumor does not metastasize.
5. Pleomorphic hyalinizing angiectatic tumor of soft parts (hemosiderotic fibro-
histiocytic lipomatous lesion), a slowly growing tumor, usually arises in
the subcutis of the lower extremity in adults. There is a predilection
for the foot and ankle. The neoplasm is characterized by a background
of thin-walled ectatic blood vessels within a stroma that contains large,
pleomorphic, plump spindled to round tumor cells with a low mitotic
rate. Although the tumor may recur after excision, aggressively in some
cases, its potential for metastatic spread appears limited (Am] Surg Pathol.
2004;28:1417).
B. Intermediate (rarely metastasizing)
1. Angiomatoid fibrous histiocytoma {AFH), a slowly growing tumor usually aris-
ing in the lower dermis or subcutis of the limbs, trunk, or head and neck,
occurs in children and young adults (Arch Pathol Lab Med. 2008;132:273).
Most cases measure 4 em or less. The cut surface is multinodular and
hemorrhagic; blood-filled cystic spaces are a common feature, but non-
hemorrhagic tumors are also seen. The neoplastic population consists of
plump to spindled cells, with interspersed pseudoangiomatoid or aneurys-
mal spaces (e-Fig. 46.51). A minority of cases feature nodules without hem-
orrhage that are composed of cells with enlarged hyperchromatic nuclei. A
minority of cases also feature prominent mitotic figures. The tumor cells
express vimentin, desmin, CD99 (with membranous staining), and CD68.
This tumor is one of several STTs that harbor an EWSR1 translocation
2. Ossifying fibromyxoid tumor is seen in adults as a long-standing painless
extremity mass measuring 3 to 5 em in greatest dimension. The extremities
(70% of cases), head and neck, and retroperitoneum are various primary
sites of the tumor (Am] Surg Pathol. 2008;32:996). Microscopically, lob-
ules of uniform, round to spindled cells are arranged in cords and nests
in a fibromyxoid to collagenous matrix. An incomplete fibrous pseudocap-
sule or shell of lamellar bone is present around the periphery. Although
microscopic features do not reliably predict behavior, increased cellularity
(resembling EWS/PNET) and an increased mitotic rate (up to 10 mitotic
figures per 10 HPFs) are associated with a higher rate of recurrence. These
tumors have a variable immunophenotype, but can express cytokeratin,
S-100 protein, glial fibrillary acidic protein, and CD99.
3. Myoepithelial neoplasms have a number of different designations including
soft tissue myoepithelioma, parachordoma, and cutaneous mixed tumor.
These tumors comprise a heterogenous clinicopathologic group of neo-
plasms, all of which display varying proportions of epithelial and/or myoep-
ithelial elements (Ann Diagn Pathol. 2007;11:190; Am ] Surg Pathol.
2003;2 7:1183 ). All present as a painless swelling in the subcutis or
deep soft tissues of the extremities that has often been present for sev-
eral years. Most tumors present primarily in adults, but up to 20% of
cases are seen in children under 10 years old (Am] Surg Pathol. 1997;
21:13).
There is a resemblance to pleomorphic adenoma of the salivary glands
in that the epithelial and myoepithelial elements characteristically form a
wide range of architectural patterns, divergent differentiation, and a low
mitotic rate. Malignant degeneration into either a carcinoma or sarcoma is
occasionally observed, although it is clear that morphologic features cannot
be used to reliably predict prognosis since a subset of histologically benign
cases also recur and metastasize. Despite the similarities between soft tis-
sue myoepithelial tumors and pleomorphic adenoma of the salivary glands,
the presence of EWSR1 rearrangements in myoepithelial neoplasms (rather

than the PLAG1 and HMGA2 rearrangements characteristic of pleomor-
phic adenoma) emphasizes that there is no pathogenic relationship between
the tumor types (Genes Chromosomes Cancer. 2010;49:1114).
4. Giant cell fibroblastoma is the unique morphologic manifestation of der-
matofibrosarcoma protuberans that presents in early childhood as a soft
tissue mass in the subcutis of the trunk, thigh, and perineum. It is an
ill-defined tumor composed of delicate to spindle cells within a fibrous
to fibromyxoid to hyalinized stroma, with scattered multinucleated giant
cells associated with nonvascularized spaces. Similar cells are found in
giant angiofibroma, pleomorphic lipoma, neurofibroma, and collagenoma
(Diagn Pathol. 2007;2:47; Am] Dermatopathol. 2004;26:141). A subset
of giant cell fibroblastomas are accompanied by classic dermofibrosarcoma
protuberans, which is not surprising since these tumors share the same
t(17;22)(q22;q13) translocation.
5. Perivascular epithelioid cell tumors (PEComas} are a family of presumably
related neoplasms with morphologic and immunophenotypic similarities
(Histopathology. 2006;48:75; Semin Diagn Pathol. 2009;26:123; Virchows
Arch. 2008;452:119). Angiomyolipoma, lymphangiomyomatosis, dear cell
"sugar" tumor of the lung, and so-called myomelanocytic tumor of the lig-
amentum teres are the various specific morphologic subtypes of PEComa;
generic-type PEComas are commonly found in the female reproductive
tract, kidney, heart, bladder, and bone. Microscopically, PEComas are com-
posed of variable proportions of epithelioid and spindle cells with clear to
eosinophilic granular cytoplasm and round nuclei. The tumor cells can have
a perivascular orientation around a thin-walled blood vessel, or a vague
nesting pattern with a resemblance to a paraganglioma. Necrosis and vas-
cular invasion are seemingly reliable features of malignancy.
The tumor cells are immunopositive for the melanocytic markers HMB-
45 and melan-A, for SMA, and less often for desmin. The tumor cells are
nonreactive for S-100, neuroendocrine markers, and cytokeratin. A subset
of PEComas harbor TFE3 fusion transcripts, which suggests a molecular
relationship to melanotic Xpll translocation renal carcinoma and alveolar
soft part sarcoma (Am] Surg Pathol. 2010;34:1395; Am] Surg Pathol.
2009;33:609; Arch Pathol Lab Med. 2010;134:124).
C. Malignant
1. Ewing sarcoma/primitive neuroectodermal tumor (EWSJPNED presents in the
soft tissue, bone, viscera, skin, and other anatomic sites in children, ado-
lescents, and young adults. This tumor accounts for about 20% of STS in
the first two decades of life, but is also recognized throughout adulthood.
Whether in bone or soft tissue, the tumor is usually in excess of 6 em; larger
tumors occur in anatomically silent locations like the paraspinal region or
pelvic retroperitoneum.
A biopsy followed by posttreatment resection is the management
sequence in most cases (since few of these tumors are candidates for pri-
mary resection); these biopsies offer many diagnostic challenges because of
the frequently associated necrosis and compression artifact. Typically, the
tumor is composed of uniform round cells with clear to finely vacuolated
cytoplasm, and a central nucleus with fine to slightly coarse chromatin.
Mitotic figures are present but are not numerous in most cases. Architec-
turally, broad sheets, lobules, nests, and strands are some of the growth
patterns (e-Fig. 46.52); other features include Homer-Wright rosettes, con-
densed small cellular nodules within otherwise monotonous sheets of round
cells, some spindling (possibly an artifact), and a pale mucoid background.
Clear cytoplasm is usually an indication of abundant diastase digestible
glycogen. Vimentin and CD99 immunostains are diffusely positive with
'J'J& I SECTION .... SOFT TISSUE AND BONE
I'1,1,! (!·Ill :..'1"-r::,U,:

I lmmuMIIIItocllemlclll ~ olllll(llwt lltound
VIII Cl CII4S - Cllll Wf~ all Tllf Cl* lii'MI8ll

RMS + +
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epithelial-like glands (e-Fig. 46.54) or nests]. A third type, poorly differenti-

ated SS, has features of a primitive round cell sarcoma. The immunopheno-
type, regardless of subtype, includes the expression of vimentin, cytokeratin
(CK7), and/or epithelial membrane antigen and CD99. The t(X;18) is char-
acteristic of the tumor, which can be helpful for distinguishing the tumor
from fibrosarcoma, MPNST, LGFMS, and SFTihemangiopericytoma.
4. Clear cell sarcoma of tendon and aponeuroses (melanoma of soft parts)
typically presents in the soft tissues of the foot and ankle of children and
young adults. However, the neoplasm is also recognized in the intestinal
tract (often with osteoclast-like giant cells), kidney, and other sites includ-
ing the head and neck (Arch Pathol Lab Med. 2007;131:152;] Clin Pathol.
2010;63:416; Ann Diagn Pathol. 2009;13:30). Nests to broad fascicles of
plump spindled to polygonal cells with abundant eosinophilic to dear cyto-
plasm, with vesicular nuclei with a prominent eosinophilic nucleolus, are the
histologic features (e-Fig. 46.55). Multinucleated cells may be found. Like
cutaneous melanoma, the immunophenotype includes vimentin, HMB-45
and melan-A (MART-1) immunopositivity, but these two tumors are distinc-
tively different at the molecular genetic level. The balanced translocations
involving the EWSR1 gene in dear cell sarcoma (Table 46.1) are not present
in cutaneous melanoma (Virchows Arch. 2010;456:219).
5. Alveolar soft part sarcoma, most commonly seen in patients between the ages
of 15 to 35 years, has a predilection for the orbit and base of the tongue
in children, and the deep soft tissue of the extremities (especially the thigh)
in adults (Arch Pathol Lab Med. 2007;131:488). Other than a mass with
varying dimension, there is nothing specific about the gross features of the
tumor. Microscopically, uniform large epithelioid or polygonal cells with
abundant eosinophilic granular to dear cytoplasm are typically arranged
in nests separated by delicate fibrous septa and sinusoidal vessels (e-Fig.
46.56). However, the tumor may have a more diffuse, nonalveolar pat-
tern, especially in children; anaplasia, mitotic activity, necrosis, and vascular
invasion are other common features (] Clin Pathol. 2006;59:1127). Rod-
shaped or rhomboid crystalline inclusions, when present, are highlighted
by a PAS stain after diastase digestion. The nonreciprocal der(17)t(X;17)
translocation is the characteristic cytogenetic feature of this tumor (Vir-
chows Arch. 2010;456:153).
6. Extraskeletal myxoid chondrosarcoma, typically arising in the deep soft tis-
sues of the proximal extremities, usually presents between 35 and 60 years
of age and has a male predilection. Uncommon other primary sites include
the nasopharynx, vulva, heart, and retroperitoneum. Grossly, the tumor is
multilobulated, and its myxoid gross appearance is reflected in its micro-
scopic composition of small, uniform, round cells with minimal atypia,
finely granular eosinophilic cytoplasm, and bland round to oval nuclei that
are arranged in dusters, cords, and delicate networks within a prominent
myxoid matrix (e-Fig. 46.57). Cartilaginous differentiation is not present
despite the appellation.
The tumor is associated with a limited set of translocations (Table 46.1)
that produce rearrangements of the CHN gene (Pathol Int. 2005;55:453).
The same rearrangements are not present in chondrosarcoma of skeletal
origin or extraskeletal mesenchymal chondrosarcoma.
7. Extrarenal malignant rhabdoid tumor is an extremely aggressive neoplasm of
the liver, axial soft tissues, and central nervous system (in the latter loca-
tion it is named atypical teratoid-rhabdoid tumor) in children, typically
under the age of2 years (Cancer. 2007;110:2061; Pathol Int. 2006;56:287).
This tumor can present in newborns with disseminated disease including
placental involvement, and is in all respects identical to renal malignant
rhabdoid tumor. Grossly, it is a poorly circumscribed, highly infiltrative neo-

plasm has a soft, grayish tan, and often necrotic cut surface. Various micro-
scopic patterns may be associated with this high-grade malignant round cell
neoplasm. Few or many obvious rhabdoid cells that have an eccentric vesic-
ular nucleus, prominent nucleolus, and the characteristic eosinophilic fila-
mentous inclusion (which contains vimentin or cytokeratin) may be present.
Oftentimes, immunostaining for vimentin and/or cytokeratin highlights the
fact that many more cells have inclusions. The tumor cells also express
CD99 with a patchy or focal pattern.
Somatic biallelic inactivation of the hSNF5flNI1 gene is the underly-
ing genetic feature which can be demonstrated by an absence of nuclear
immunoreactivity for BAF4 7. Molecular demonstration of alterations
involving this locus can be used to differentiate extrarenal malignant rhab-
doid tumor from poorly differentiated carcinomas and sarcomas in adults
that have a rhabdoid phenotype as an epiphenomenon. The distinction
between extrarenal malignant rhabdoid tumor and epithelioid sarcoma has
become blurred on the basis of recent immunophenotype and molecular
genetic findings.
8. Epithelioid sarcoma typically presents in individuals in the second through
the fourth decade. The distal or conventional type of epithelioid sarcoma
is found in the fingers, hand, or wrist, or the equivalent sites in the distal
lower extremity. The more aggressive proximal type involves the pelvis, per-
ineum, and genital tract with overlapping clinical and pathologic features
of extrarenal malignant rhabdoid tumor (Cancer Res. 2005;65:4012).
In the distal type, arcades and nodules of uniform epithelioid cells with
an eosinophilic cytoplasm may show a transition into areas of spindled
cells, both set in a background of abundant collagen (e-Fig. 46.58). The
nodules can undergo central necrosis, creating an appearance that resembles
a necrobiotic granuloma (with the potential misinterpretation as granuloma
annulare since both lesions present in the fingers or toes) (Adv Anat Pathol.
2006;13:114). Recurrent epithelioid sarcoma may have features of a high-
grade pleomorphic sarcoma with only a remote resemblance to the primary
tumor.
In the proximal type, the tumor is composed of sheets of large cells
with prominent rhabdoid morphology in the absence of a granuloma-like
pattern (e-Fig. 46.58).
Both the distal and proximal types of tumor express EMA, vimentin,
cytokeratins, and CD34 (in 40% to 50% of cases). There is general agree-
ment that both types consistently display loss of BAF47 reactivity (80%
to 90% of cases) that reflects inactivation of the hSNF5/INI1 gene (Am]
Clin Pathol. 2009;131:222; Am] Surg Pathol. 2009;33:542; Hum Pathol.
2009;40:349).
9. Undifferentiated-poorly differentiated sarcoma is a pathologic diagnosis of
last resort. The morphologic scenario usually differs on the basis of age: in
children, the problematic tumor is usually a primitive or high-grade round
cell neoplasm, while in adults it is an equally high-grade pleomorphic neo-
plasm with anaplasia, bizarre mitoses, and necrosis.
In children, the diagnosis of "undifferentiated round cell sarcoma" or
"sarcoma of undermined histogenesis" becomes the interpretation once the
various differentiated malignant round cell neoplasms have been excluded
by immunohistochemical and molecular testing (Table 46. 7), and both
are ever shrinking categories (] Pediatr Hematol Oncol. 2001;23:215;
Hum Pathol. 2009;40:1600). In these undifferentiated tumors, loss of
hSNF5/IN11 expression in infants and young children seems to be asso-
ciated with a better prognosis (Mod Pathol. 2009;22:142).
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SUGGESTED READINGS
Dehner LP. Soft tissue. In: Stocker T, Dehner LP, Husain AN, eds. Stocker and Dehner Pediatric
Pathology, 3rd ed. Philadelphia: Lippincott, Williams & Wilkins; 2011.
Fletcher CDM, Unni KK, Mertens F. Pathology and Genetics of Tumors of Soft Tissue and Bone.
World Health Organization Classification of Tumors. Lyon: IARC Press; 2002.
Kempson RL, Fletcher CDM, Evans HL, et al. Tumors ofthe Soft Tissues. Atlas of Tumor Pathology,
3rd Series, Fascicle 30. Washington, DC: Armed Forces Institute of Pathology; 2001.
Pfeifer JD. Molecular Genetic Testing in Surgical Pathology. Philadelphia: Lippincott, Williams &
Wilkins; 2006.
Weiss SW, Goldblum JR. Soft Tissue Tumors, 5th ed. St. Louis: Mosby; 2008.
Retroperitoneum
Catalina Amador-Ortiz, Louis P. Dehner,
and John D. Pfeifer
I. NORMAL ANATOMY. The retroperitoneal space (in some respects a virtual space) is
located between the posterior parietal peritoneum and the fascia that covers the
muscles of the lumbar region. It extends upward to the diaphragm, downward to
the base of the sacrum and iliac crests, and laterally to the external borders of
the lumbar muscles and the ascending and descending colon. The retroperitoneum
contains loose connective tissue surrounding lymph nodes, the aorta and inferior
vena cava with their vascular branches, the adrenal glands, the kidneys and ureters,
the pancreas, and portions of the duodenum. This chapter will focus on the entities
that arise from the tissues of the retroperitoneal space; disorders arising from the
organs that are completely or partially retroperitoneal are covered in the respective
chapters.
II. NONNEOPLASTIC DISEASES
A. Inflammatory
1. Idiopathic fibroinflammatory lesions constitute a morphologically similar cat-
egory of tumefactive processes, and a variety of diagnostic terms such as
sclerosing mesenteritis and/or panniculitis have been applied. The usual pre-
sentation is a mass in the retroperitoneum and/or root of the mesentery; scle-
rosing changes may extend into the inferior retroperitoneum with similar fea-
tures, if not identical, to idiopathic retroperitoneal fibrosis. A biopsy reveals
a variably intense mixed lymphocytic and plasmacellular infiltrate in a dense,
relatively hypocellular collagenous background. Residual adipose tissue may
have the features of fat necrosis with dystrophic calcification and panniculi-
tis. In the presence of lymphadenopathy, the possibility of lymphoma should
be excluded. Metastatic disease including poorly differentiated adenocarci-
noma of the pancreas, carcinoid, lobular carcinoma of the breast, and acinar
carcinoma of the prostate may be a accompanied by disproportionate fibroin-
flammatory reaction to the actual volume of tumor, and thus also need to be
excluded. If the biopsy has a prominently spindle cell component, inflamma-
tory pseudotumo.r, inflammatory myofibroblastic tumor (IMT), sarcomatoid
carcinoma, pleomorphic spindle cell sarcoma, and IgG4-related sclerosing
disorder are also in the differential diagnosis (Nat Rev Gastroenterol Hepa-
tol. 2010;7:401;] Clin Pathol. 2008;61:1093). Since the histologic findings
of idiopathic fibroinflammatory lesions are nonspecific, even after the other
possibilities have been excluded, the diagnostic line in the report may simply
read "chronic inflammation and fibrosis."
2. Malakoplakia occasionally affects the retroperitoneal soft tissues. The pro-
cess is a response to infection and grossly appears as a yellow plaque-like
lesion. Microscopically, it is composed of abundant inflammatory cells and
sheets of granular and vacuolated histiocytes that contain lamellated, PAS-
positive diastase-resistant inclusions known as Michaelis-Gutmann bodies.
These distinctive inclusions (e-Fig. 47.1)* are thought to represent the rem-
nants of bacteria within phagosomes that have been mineralized by calcium
and iron.
781
3. Retroperitoneal abscesses are generally secondary to infectious processes of

adjacent organs, most commonly of the kidneys. Less frequently, abscesses
originate from distant septic foci that propagate via a hematogenous route.
4. Other nonneoplastic conditions including hemorrhage, bile collection, and
extravasation of urine can be occasionally encountered in the retroperitoneal
space. Endometriosis may also involve the retroperitoneum.
B. Idiopathic retroperitoneal fibrosis (sclerosing retroperitoneal fibrosis, Ormond
disease) is an uncommon inflammatory process characterized by sclerosing fibro-
sis of the retroperitoneum that can ultimately cause constriction and obliteration
of the ureters.
Grossly, there is poorly circumscribed fibrosis, usually at the level of the
lower abdominal aorta and its bifurcation. Microscopically, the process is char-
acterized by dense fibrosis with collagen entrapment, with associated alternating
areas of prominent inflammation primarily consisting of plasma cells, histio-
cytes, eosinophils, and lymphocytes (e-Fig. 47.2). The plasma cells may be lgG4
positive in which case the disorder may be a member of the IgG4-related scle-
rosing disorders, especially in affected males (Am] Surg Pathol. 2009;33: 1833 );
periaortitis in association with retroperitoneal fibrosis may be part of the same
sclerosing disease spectrum (Am] Surg Pathol. 2008;32:197), as are scleros-
ing inflammatory diseases in the thyroid, mediastinum, and bile ducts. In the
remaining cases, the etiology is undetermined or associated with methysergide
and other drugs (and often regresses after cessation of the drug).
C. Cystic lesions. While most cystic lesions of the retroperitoneum represent a sec-
ondary or degenerative change within a benign or malignant neoplasm, several
benign primary cystic lesions also occur.
1. Cystic lymphangioma or cystic lymphatic malformation of the retroperi-
toneum accounts for 5% of all retroperitoneal cystic lesions and can occur
at any age. Involvement of the mesentery is an associated feature since this
malformation is not well circumscribed USurg Oncol. 1996;61:234). Patho-
logically, multi- or unilocular cysts contain clear or milky fluid, and are lined
with a single layer of flattened endothelium, which is 02-40 immunopositive
UPediatr Surg. 1999:34;1164).
2. Multicystic peritoneal inclusion cyst occurs predominantly in women of repro-
ductive age and, even though rarely seen in the retroperitoneum, is seen in the
differential diagnosis of cystic lesions (especially of multilocular cystic lym-
phangioma). Prior abdominal surgery is common. Histopathologically, the
lesion consists of mesothelial-lined cystic spaces usually containing watery
secretions, separated by a delicate fibromuscular stroma. The lining epithe-
lium is usually immunoreactive for calretinin. As discussed in some detail
in Chapter 11 (Section IY.B), some confusion exists regarding the proper
classification of multicystic peritoneal inclusion cyst, as demonstrated by the
fact that the lesion is also known as multicystic mesothelioma (Ann Diagn
Pathol. 2000;4:308).
3. Bronchogenic cysts can rarely occur in a subdiaphragmatic location, includ-
ing the retroperitoneum, as part of the morphologic spectrum of bronchopul-
monary foregut malformations. The related anomaly, extralobar sequestra-
tion with features of congenital cystic adenomatoid malformation type 2
presents a solid mass in the same site, usually in infants and young children;
its suprarenal location can lead to confusion with neuroblastoma U Pediatr
Surg. 2007;42:1627). The cysts are lined by respiratory-type, pseudostrati-
fied, ciliated columnar epithelium that can focally contain seromucous glands
and nodules of hyaline cartilage.
4. Mullerian cysts of the retroperitoneum are usually in excess of 10 em in great-
est dimension and are lined by cuboidal to columnar epithelium that often
contains ciliated cells; there is no atypia, although stratification and epithelial
Chapter 47 • Retroperitoneum I 7 83
tufting can be present as in tubal epithelium. Beneath the lining of a miillerian

cyst, loose fibrous tissue, dilated vessels, and incomplete smooth muscle bun-
dles can be seen (Hum Pathol. 2003;34:194). Mucinous cystadenoma with
ovarian type stroma (which is inhibin immunopositive) occurs rarely as an
extrapancreatic cyst in the retroperitoneum.
5. Enteric duplication cyst (EDC} is an extremely uncommon congenital lesion and
may be detected prenatally (Pediatr Radiol. 2000;30:671). Although usually
intra-abdominal and associated with the small intestine, it has sporadically
been described in the retroperitoneum. A spherical, tubular, or dumbbell-
shaped cyst with a smooth lining and viscous contents is the gross appearance.
Microscopically, the cyst wall has a variably differentiated enteric mucosa
consisting of a simplified cuboidal or cylindrical epithelium, to a more nor-
mal appearing mucosa with a muscularis mucosa and a muscularis propria.
Focal areas of squamous metaplasia and gastric-type mucosa have also been
described in retroperitoneal EDCs (fOP. 2006;7:492). Heterotopic pancreas
is another infrequent finding.
Ill. NEOPLASMS. Primary retroperitoneal tumors arise from the extravisceral tissues
that comprise the retroperitoneum; the neoplasms for the most part are malignant
in adults and originate from lymphoid and various soft tissue elements including
fat and neural structures. The histogenesis of some of the high-grade sarcomas is
not apparent, and the immunophenotype often does provide information that is
specific in terms of lineage.
In children and young adults, germ cell neoplasms can present in the retroperi-
toneum, but the most common neoplasms of the retroperitoneum in children
include the neuroblastic tumors (neuroblastoma and its variants) and high-grade
lymphomas, in particular Burkitt lymphoma, large cell lymphoma, and Hodgkin
lymphoma. The most commonly encountered primary retroperitoneal tumors in
adults are sarcomas, followed by lymphomas and germ cell tumors (Sarcoma.
2001;5:5). Overall, 70% to 80% of retroperitoneal tumors are malignant (and
thus the retroperitoneum is the only body site where the frequency of malignant
neoplasms exceeds that of benign tumors).
The anatomy of the retroperitoneal space makes it possible for primary (as well
as metastatic) tumors to attain substantial dimensions before clinical manifestations
become evident. Malignant retroperitoneal tumors have a generally poor prognosis
since they are often found at an advanced stage of disease, and even resectable
tumors often recur.
A. Soft tissue tumors. Primary sarcomas arising in the retroperitoneum constitute
10% to 20% of all soft tissue sarcomas in adults: liposarcoma (40% of cases),
leiomyosarcoma (30% ), and pleomorphic sarcoma-malignant fibrous histiocy-
toma (25%) (Expert Rev Anticancer Ther. 2009;9:1145). As a group, retroperi-
toneal sarcomas are often in excess of 10 em and weigh in excess of 400 to 500 g.
Complete resection is associated with the most favorable outcome, but size and
involvement of other structures often limits the extent and completeness of the
resection; local recurrence is guaranteed in the latter cases. With complete resec-
tion and no local recurrences, the 5-year survival is 40% to 50%. As for soft
tissue sarcomas arising in other anatomic sites, the American Joint Commit-
ted on Cancer (AJCC) staging system is utilized for staging of retroperitoneal
sarcomas (see Chap. 46).
1. Adipocytic tumors
a. Benign
i. Myolipoma, one of the more common benign retroperitoneal lipoma-
tous tumors, is more common than pure retroperitoneal lipomas. This
encapsulated lesion has a lobulated fatty appearance with bands and
nodules of white to gray-white tissue (World] Surg Oncol. 2005;3:72);
the large size, usually 10 em or greater, often raises the question of
liposarcoma. Microscopically, the tumor contains an admixture of

mature adipocytes and bundles of smooth muscle (which usually pre-
dominate), but there is an absence of mitotic activity and atypia. The
tumor is diffusely and strongly immunopositive for smooth muscle
actin (SMA) and desmin.
ii. Other mixed-type benign fatly tumors in the retroperitoneum include
myelolipoma and angiomyolipoma (AML) arising in the adrenal and
kidney, respectively; as these tumors enlarge, both may convey the
impression of a tumor based in the retroperitoneum. Examples of
purely retroperitoneal AMLs have been described (Abdom Imaging.
2004;29: 721 ). Microscopically, the presence of myeloid elements char-
acterizes myelolipoma; atypical appearing lipoblasts may lead to a
misinterpretation of liposarcoma in a largely fatty AML, but thick-
walled vessels should be the clue to the latter diagnosis. HMB-45
and Melan-A immunopositivity in the nonadipocytic areas of AML
also can be used to establish the diagnosis. Nuclear staining for TFE3
is another feature of AML. Perivascular epithelioid cell neoplasm, a
related entity to AML, has also been reported as a primary tumor of
the pelvis, mesentery, or retroperitoneum (Am] Surg Pathol. 2005;29:
1558).
iii. Lipoblastoma, a tumor of early childhood, presents in the retroperi-
toneum in 5% to 10% of the cases. From the retroperitoneum, the
tumor may extend into the mesentery and omentum. Lipoblastoma is
commonly in excess of 6 em in diameter, and grossly has a glistening
gray-white mucoid surface with faint lobulation. Microscopically, the
lobules contain a mixture of mature adipocytes and immature fetal
adipocytic cells with progressive differentiation toward mature adi-
pose cells. The lobules are usually separated by connective tissue septa
that contain small vessels. Some tumors have a striking resemblance to
myxoid liposarcoma, but lipoblastoma contains desmin-positive cells
and a PLAG1 mutation.
b. Malignant. As a group, liposarcoma is the most common primary nonvis-
ceral malignant neoplasm of the retroperitoneum and accounts for 30%
to 50% of retroperitoneal sarcomas.
i. Atypical lipomatous tumor/Well-differentiated liposarcoma (ALT/
WDLPS) is the most common subtype of liposarcoma in the retroperi-
toneum. It is the largest neoplasm of this site and can measure in
excess of 15 em and weigh 1 kg or more. Grossly, a thin external
membrane is usually the only structure serving as a "capsule" for
an otherwise well-circumscribed yellowish-white lipomatous mass.
The diagnostic challenge of ALTIWDLPS is the identification of
atypical lipoblasts with enlarged, hyperchromatic, irregular nuclei
with indented or scalloped nuclear membranes, especially in those
cases with a predominant mature lipomatous appearance and few
classic lipoblasts (e-Fig. 47.3). The paucity of lipoblasts may be
accompanied by fibrosclerotic foci containing atypical stromal cells,
a feature of equal diagnostic importance to the atypicallipoblasts.
ii. Dedifferentiated liposarcoma is identified by the presence of sharply
demarcated areas of nonlipogenic sarcoma adjacent to areas of well-
differentiated liposarcoma (e-Fig. 47.4). In the retroperitoneum, the
nonlipogenic sarcomatous component has a fibrosarcomatous or
undifferentiated morphology. Both ALTIWDLPS and dedifferentiated
liposarcoma harbor amplification of the MDM2 and CDK4 genes,
and immunohistochemically, both are positive for MDM2 and CDK4
expression.
iii. Myxoidlround cell liposarcoma is uncommon in the retroperitoneum.

On the basis of immunohistochemical and molecular analysis, it has
been shown that most presumed primary retroperitoneal myxoid/
round cell liposarcomas actually represent well-differentiated/
dedifferentiated liposarcomas (Mod Pathol. 2009;22:223).
iv. Pleomorphic liposarcoma in the retroperitoneum presents the differen-
tial diagnosis of pleomorphic undifferentiated sarcoma. Correct clas-
sification is often a matter of identifying convincing lipoblasts.
2. Smooth muscle tumors
a. Deep-seated leiomyoma (leiomyoma of deep soft tissue) is an extremely
uncommon benign tumor that primarily occurs in perimenopausal
women; only isolated cases have been reported in males. Microscopically,
the tumor resembles a uterine leiomyoma, exhibiting intersecting fascicles
of bland appearing smooth muscle cells. Mitotic activity is low (<5 mitoses
per 50 high-power fields [HPFs]), and the tumor is commonly positive for
estrogen and progesterone receptor expression. Deep-seated leiomyoma
will occasionally recur after incomplete excision, but metastasis does not
occur (Adv Anat Pathol. 2002;9:351 ). Explanations for a deep smooth
muscle tumor in the abdomen or retroperitoneum of a female include a
so-called parasitized leiomyoma from the uterus, which has essentially sep-
arated from the outer portion of the uterus and gained its principal blood
supply from an adjacent structure. Intravenous leiomyomatosis from
the uterus also presents as benign appearing smooth muscle tumors in the
pelvic soft tissues, often with extension into adjacent lymph nodes. The
essential pathologic feature is the identification of circumscribed nodules
of benign-appearing smooth muscle within vascular or lymphatic spaces.
b. Leiomyosarcoma (LMS) accounts for 20% to 30% of all retroperitoneal sar-
comas, is the most aggressive of retroperitoneum sarcomas, and in some
series is the most common sarcoma of this site. Like the other retroperi-
toneal sarcomas, dimensions and weights often exceed 15 em and 500 g,
respectively. There is commonly a pseudoencapsule and a nodular surface
with areas of hemorrhage, necrosis, and cystic degeneration. Histolog-
ically, the tumor is composed of highly atypical-appearing spindle cells
with elongated, blunt-ended nuclei and eosinophilic cytoplasm (e-Fig.
47.5). The presence of any cytological atypia, significant mitotic activ-
ity, or coagulative necrosis in a retroperitoneal smooth muscle neoplasm
warrants a diagnosis of LMS. Aggressive metastatic disease and a poor
prognosis are characteristic of retroperitoneal LMSs with even very low
mitotic activity (1 to 4 mitoses per 10 HPFs). It is worth mentioning that
LMSs in the retroperitoneum may present in the inferior vena cava, or as a
metastasis from a primary LMS of the uterus or paratesticular soft tissues.
3. Skeletal muscle tumors
a. Benign skeletal muscle tumors of the retroperitoneum are extremely rare.
Only isolated cases of rhabdomyoma have been reported, with similar
pathologic characteristics as when the tumor occurs elsewhere.
b. Rhabdomyosarcoma (RMS} of the embryonal or alveolar subtypes in the
retroperitoneum-pelvis is a neoplasm of children between 2 and 10 years
of age (e-Fig. 47.6). Approximately 10% to 15% of childhood RMSs arise
in the soft tissues of the pelvis and/or retroperitoneum, and are usually
10 em or greater in maximal dimension (Pediatr Blood Cancer. 2004;42:
618); the retroperitoneum is an unfavorable site for a childhood RMS
(/ Pediatr Hematol Oncol. 2001;23:215). Pleomorphic RMS in the
retroperitoneum may represent a component of a dedifferentiated liposar-
coma, a nongerminal pattern of malignancy in a germ cell tumor, or spread
of a malignant mixed miillerian tumor (Am J Surg Pathol. 2007;31: 1557).
4. Fibroblastic tumors
a. Benign fibroblastic tumors other than desmoid fibromatosis are rare in the
retroperitoneum. It may be difficult to differentiate between a presumed
fibrous tumor and a nonneoplastic fibroinflammatory process (e.g., an
IgG4 fibrosclerosing process).
b. Desmoid fibromatosis, IMT, extrapleural solitary fibrous tumor/hemangio-
pericytoma, and gastrointestinal stromal tumors {GISTs) are several examples
of intermediate behavior tumors that may involve the retroperitoneum.
Desmoid fibromatosis usually arises in the mesentery-omentum with
extension into the retroperitoneum, but occasionally is limited to the
retroperitoneum. Those desmoid tumors arising in the proximal lower
extremity can, in the course of several local recurrences, extend through
sciatic notch into the retroperitoneum as well; the presence of familial ade-
nomatous polyposis in these patients should be addressed. Desmoid fibro-
matosis is not encapsulated but is well circumscribed, except in these cases,
which represent a local recurrence. Its cut surface shows a firm, trabecu-
lated pattern resembling a smooth muscle tumot: Microscopically, dense
interlacing bundles of uniform spindle-shaped fibroblasts are present in a
collagenous, keloid-like, or myxoid stroma with no evidence of nuclear
atypia. Scattered mitotic figures may be identified and should not be a
source of concern unless they are atypical (e-Fig. 47.7). The tumor is
immunopositive for vimentin, SMA, and nuclear ,8-catenin staining. If the
tumor is positive for CD34 and CD117 (c-kit), GIST is the favored diag-
nosis as either a local recurrence or a rare primary extraintestinal GIST
(] Gastrointest Surg. 2009;13:1094).
Primary retroperitoneal extrapleural solitary fibrous tumor/hemangio-
pericytoma is uncommon, but documented (]Am Coli Surg. 2004;198:
322; Hum Pathol. 2000;31:1108). These CD34-positive (but CD117-
negative) neoplasms may be quite cellular and mitotically active, in which
case the most appropriate diagnosis is malignant solitary fibrous tumor.
Although grouped together as related (if not identical) tumors in the cur-
rent World Health Organization (WHO) scheme (see Chap. 46), the diag-
nosis of hemangiopericytoma is reserved for tumors that resemble the
cellular areas of solitary fibrous tumor with the characteristic network of
vascular profiles.
IMT, like desmoid fibromatosis, may present in the mesentery-
omentum with extension into the retroperitoneum. IMT typically presents
in children and young adults. Grossly, it is a circumscribed firm, gray-white
mass that measures 5to 10 em. Microscopically, it has a varied micro-
scopic pattern of spindle cells with accompanying plasma cells, myxoid
foci resembling nodular fasciitis, and dense hyalinized collagen. In addi-
tion to immunopositivity for vimentin and SMA, 50% or more of cases
are anaplastic lymphoma kinase (ALK-1) positive (Am] Surg Pathol.
2007;31:509). The presence of IgG4 plasma cells in IMT is seen in some
cases (Mod Pathol. 2011;24:606).
c. Malignant fibroblastic tumors are very rare in the retroperitoneum. Adult
fibrosarcoma is currently a diagnosis of exclusion and, according to recent
series, accounts for <5% of all retroperitoneal sarcomas; sarcomatous
transformation of a solitary fibrous tumor should be considered in the
differential diagnosis (Am] Surg Pathol. 2009;33:1314). Microscopically,
fibrosarcoma is composed of spindle-shaped cells with large elongated
nuclei and scant cytoplasm, arranged in a classic herringbone pattern.
Infantile fibrosarcoma is known to occur in the retroperitoneum, but
most commonly presents in the extremities; the identical neoplasm
arising in the kidney in infancy, cellular mesoblastic nephroma, has the
same morphology and also harbors the t(12;15) translocation (] Clin

Oncol. 2010;28:318-323). Other variants of fibrosarcoma, such as low-
grade fibromyxoid sarcoma and myxoid-type fibrosarcoma, are rarely
encountered in the retroperitoneum. In fact, most retroperitoneal tumors
that present histologic features of myxofibrosarcoma may actually repre-
sent dedifferentiated liposarcomas (Virchows Arch. 2001;439:141).
5. Fibrohistiocytic tumors
a. Pleomorphic undifferentiated sarcoma-malignant fibrous histiocytoma {MFH)
formerly accounted for up to 30% of sarcomas of the retroperitoneum.
Recent studies would seem to imply that many of these high-grade sar-
comas actually represent dedifferentiated liposarcomas. Thus, a careful
search for focal ALTIWDLPS, as well as immunostains for MDM2 and
CDK4 expression and/or cytogenetic analysis for chromosome 12 abnor-
malities (as discussed in Section Ill.A.1.b) should be performed to confirm
the diagnosis.
b. Inflammatory MFH is most commonly found in the retroperitoneum. Micro-
scopically, it is characterized by atypical spindle cells admixed with xan-
thomatous cells within an extensive acute and chronic inflammatory infil-
trate composed of neutrophils, eosinophils, lymphocytes, and plasma cells
(e-Fig. 47.8). The differential diagnosis includes LMS and liposarcoma,
both of which may have similar inflammatory reactions.
6. Neural tumors
a. Benign. Schwannoma accounts for approximately 1% to 5% of all
retroperitoneal tumors, and grossly appears as a large(> 10 em) solitary,
well-encapsulated, firm, round mass. Histologically, it presents with alter-
nating cellular areas with palisading nuclei (Antoni A areas), and myxoid
hypocellular areas (Antoni B areas). A characteristic feature of schwan-
nomas of the retroperitoneum is the presence of cystic change (Urology.
1986;28:529), as well as thickened hyalinized vessels. Ganglioneuroma
is a tumor with dispersed ganglion cells in a background neuromatous
stroma. Ganglioneuroma can be mistaken for a schwannoma in those
areas of the tumor with a paucity or absence of ganglion cells.
b. Malignant. Malignant peripheral nerve sheath tumor (MPNST) represents
5% to 10% of all retroperitoneal sarcomas. Microscopically, the tumor
features asymmetric spindle-shaped cells arranged in dense fascicles with
hyperchromatic nuclei and frequent mitoses (e-Fig. 47.9). In the setting of
neurofibromatosis type I, the tumor often has a plexiform growth pattern.
c. Tumors of sympathetic nervous tissue
i. Neuroblastoma may originate from the sympathetic nervous system
chain of ganglia, commonly in the retroperitoneum and without
involvement of the adrenal gland. In fact, the extraadrenal retroperi-
toneum is the site of origin of 30% to 35% of neuroblastomas in the
pediatric population. Adult neuroblastoma, though rare, is nonethe-
less in the differential diagnosis of malignant round cell tumors of
the retroperitoneum of adults. Nonetheless, there is a greater likeli-
hood that a paraspinal malignant round cell neoplasm in a young adult
is Ewing sarcoma/peripheral neuroectodermal tumor (EWS/PNET) or
lymphoma than a neuroblastoma.
ii. Extraadrenal retroperitoneal paraganglioma is an uncommon tumor, and
arises from the chromaffin cells located in the paraaortic sympathetic
chain and at the aortic bifurcation. Similar to adrenal pheochromo-
cytomas and extraadrenal paragangliomas elsewhere, the tumor may
be functional with associated characteristic signs and symptoms, but
most cases are nonfunctional. Grossly, the tumor is rubbery with firm
brown to tan cut surfaces, and in large specimens can exhibit a cystic
component secondary to central necrosis or hemorrhage. The tumor

is composed of well-defined nests of cells surrounded by sustentacu-
lar cells; the nests are usually composed of round cells with abundant
granular eosinophilic or basophilic cytoplasm (e-Fig. 47.10). Nuclear
atypia and vascular invasion may also be present. In the retroperi-
toneum, these tumors tend to have an aggressive course with local
invasion and a high incidence of local recurrence, and a higher rate
of metastasis than adrenal pheochromocytoma (20% to 42% for the
former versus 2% to 10% for the latter) (Urol Ann. 2010;2:12).
7. Vascular tumors. Most vascular tumors, ranging from hemangiomas to
angiosarcomas, have been reported as isolated cases in the retroperitoneum.
The vascular tumor that is particularly common in the retroperitoneum
is Kaposiform hemangioendothelioma, a tumor of childhood that is fre-
quently complicated by the Kasabach-Merritt phenomenon. Although ini-
tially described as a distinctive vascular neoplasm of the retroperitoneum,
it is now recognized to occur in skin and soft tissues of the extremities
(Amf Surg Pathol. 1991;15:982). In the retroperitoneum, the tumor tends to
be large, poorly circumscribed, and may involve adjacent structures including
the colon. Microscopically, the tumor is composed of fascicles of spindle cells
and narrow vascular spaces that have a lobular arrangement; microthrombi,
slit-like endothelial-lined vascular channels, and hyaline globules are also
present, and thus the tumor resembles Kaposi sarcoma. However, Kaposi-
form hemangioendothelioma lacks nuclear positivity for human herpes virus
8 (HHV-8).
8. Miscellaneous soft tissue tumors
a. Synovial sarcoma represents"' 1% of all retroperitoneal sarcomas. The true
incidence may be higher since cases formerly diagnosed as fibrosarcoma
may have represented examples of monophasic synovial sarcoma. A firm,
gray-pink, circumscribed mass measuring 8 to 10 em in greatest dimen-
sion is the typical gross appearance. In contrast to synovial sarcomas else-
where, retroperitoneal synovial sarcomas tend not to metastasize remotely,
but are difficult to control locally and thus have an overall poor prog-
nosis (Histopathology. 2004;45:245; J BUON. 2008;13:211). A poten-
tial diagnostic pitfall may arise since retroperitoneal schwannomas are
often AE1/AE3 cytokeratin immunopositive, but schwannomas are also
glial fibrillary acidic protein immunopositive unlike synovial sarcomas
(Mod Pathol. 2006;19:115).
b. Virtually all malignant round cell tumors have been described as primary
tumors of the retroperitoneum. EWSIPNET presents in this location in
rv5% of cases, usually as a paraspinal mass. Desmoplastic small round
cell tumor usually arises in the peritoneal cavity, but in some series up to
15% of cases originate in the retroperitoneum.
B. Germ cell tumors
1. Primary retroperitoneal teratomas typically occur in infancy and childhood
but are rare in adults. About 75% of cases occur in children younger than
5 years, and the incidence in females is twice that of males, as is also the
case for sacrococcygeal teratomas in children. Grossly, these tumors have a
mixed cystic solid appearance.
a. As elsewhere, mature teratomas contain mature tissue, often from all three
germ layers, with occasional calcification or ossification. Mature cystic
teratomas should be thoroughly sampled (one section per centimeter) in
order to exclude the presence of yolk sac tumor, especially in a young child.
Immature somatic elements such as primitive neural tubules or sheets
of neuroblasts with a fibrillary background should not be viewed with
concern in an infant or young child.
Chapter 47 • Retroperitoneum I 789
b. Immature teratomas contain immature or primitive tissue (derived from any

or all three germ cell layers) that is usually mixed with areas of mature
tissue. The most common immature neural tissues form rosettes, sheets of
neuroblasts, or tubules of primitive neural cells. The pathologic grading
of retroperitoneal teratomas, specifically in children, on the basis of the
extent of immature somatic tissues has no prognostic significance. On the
other hand, the presence of yolk sac tumor or endodermal sinus tumor
establishes the malignant nature of the neoplasm.
2. Secondary germ cell neoplasms. A teratoma in an adolescent or young male
may be a posttreatment "growing teratoma," which represents residual
metastatic disease from a malignant germ cell tumor of the testis. Similarly,
the presence of a retroperitoneal tumor in the absence of a known primary
testicular germ cell tumor should lead to careful evaluation of the testes since
they may harbor a scar with or without intratubular germ cell neoplasia as
evidence of spontaneous regression of a primary testicular neoplasm. In fact,
a retroperitoneal germ cell tumor, in a male, whether seminoma or of another
pattern, is metastatic disease from the testis until proven otherwise.
C. Lymphomas and other lymphoproliferative disorders
1. Both non-Hodgkin and Hodgkin lymphoma present in retroperitoneum with
lymphadenopathy, as a localized mass, or as retroperitoneal fibrosis. Bulky
retroperitoneal lymphadenopathy with or without intestinal or other organ
involvement in the first two decades of life usually represents Burkitt lym-
phoma. In adults, the same presentation usually represents small lymphocytic
lymphoma/chronic lymphocytic leukemia or one of the other B-cell lym-
phomas including follicular lymphoma. Diagnosis can be challenging due to
limitations on the adequacy of the specimen, a prominent fibrous reaction,
necrosis, or a nonrepresentative biopsy.
2. Castleman disease also occasionally presents in the retroperitoneum. The
hyaline type, which is most common in the retroperitoneum as elsewhere,
is usually localized to a single lymph node and tends to be asymptomatic.
The plasma cell type is multifocal and usually presents with a more aggressive
course and systemic manifestations. The hyaline type is characterized grossly
by homogenous orange-yellowish cut surfaces, and microscopically by giant
lymphoid follicles centered on a markedly hyalinized vessel surrounded by
lymphocytes arranged in an onion-skin pattern (e-Fig. 47.11). The plasma
cell type is grossly similar, but on microscopic examination contains more
plasma cells with less vascular hyalinization.
D. Tumors of miUierian type are occasionally described in the retroperitoneum. As
in the ovary, they are usually of serous, mucinous, or endometrioid type, and
can be benign borderline or malignant. In the retroperitoneum, borderline and
malignant tumors appear to be more common than their benign counterparts.
Tumors are usually large and unilateral, and tend to present with no concomitant
ovarian lesions.
E. Metastatic tumors. Various primary malignant neoplasms arising in retroperi-
toneal or posterior abdominal wall organs such as the pancreas, kidney, liver,
and adrenal gland often present with or are accompanied by direct extravisceral
invasion into the retroperitoneum. Retroperitoneal lymph node metastases are
also seen in association with many primary tumors of other sites.
Bone Neoplasms and
Other Nonmetabolic
Disorders
Omar Hameed and Michael J. Klein
I. NORMAL MICROSCOPIC ANATOMY. The bones are composed of compact bone, which
is derived from intramembranous ossification, and coarse cancellous bone, which
is the osseous remnant of endochondral ossification. Compact bone makes up the
cortices of long bones and constitutes their diaphyses and the surface portion of
their metaphyses, as well as the compacta of the flat and irregular bones. Cancellous
bone is present in the medullary cavity and is abundant at the ends of the long
bones. In bone, form follows function (Wolff's law). In the shafts of bones, most of
the forces act upon the surface. Here, the compact bone, which is 90% solid and
only 10% space, bears the compression, tension, shear, and torsional forces. The
medulla, shielded from forces, contains practically no bone at all. The ends of the
bones are supported by the vertical plates and horizontal struts of the cancellous
bone, yet cancellous bone is only 25% bone and 75% marrow by volume; here the
cortex is very thin.
Bone matrix is classified as woven or lamellar depending on the predominant
fiber arrangement of its collagen. In woven bone, the collagen fiber pattern is ran-
dom. This type of bone is found in the fetal skeleton and in processes in which
there is very rapid bone production. In lamellar bone, the bone collagen fibers are
arranged in stacks of tightly packed fibers that are parallel in the same stack. In the
next layet; the collagen fibers are also parallel to one another, but their direction is
different from the collagen in the previous stack so that the bone appears to be lay-
ered. Both compact bone and cancellous bone consist of lamellar bone after the age
of 3 years. After this age, woven bone is almost always pathologic, although the
etiology is often not discernible without imaging studies (Bullough PG. Orthopedic
Pathology. 5th ed. St. Louis: CV Mosby; 2010). In compact bone, the lamellae are
arranged concentrically around central vascular canals termed Haversian canals;
each vascular canal and its associated lamellae are referred to an osteon or Haver-
sian system. In cancellous bone, the lamellae are arranged in linear, parallel plates
(e-Fig. 48.1).* Adjacent osteons are separated from each other and from intersti-
tial lamellae (see the section on circulatory diseases) and circumferential lamellae
(which encircle the inner or outer cortex and are remnants of periosteal intramem-
branous ossification) by basophilic staining cement lines. Cement lines are sliding
planes that are richer in calcium than surrounding bone matrix but the exact com-
position of which is unknown; they are produced by osteoblasts when bone is
synthesized following osteoclast resorption (reversal cement lines) or after a period
of inactivity (arrest cement lines). In the former type, the lamellae are discontinuous
on either side of the cement line and in the latter the lamellae are continuous on
either side (e-Fig. 48.2).
A. Gross handling and selection of sections. The approach to specimen handling
is largely one of common sense. Small biopsy specimens should be submit-
ted for sectioning in their entirety. If there is any doubt about whether they
*All e-figures are available online via the Solution site image bank.
790
Chapter 48 • Bone Neoplasms and Other Nonmetabolic Disorders I 7 91
contain bone, they should be fixed, briefly decalcified, and rinsed. Most bone
biopsies performed with needles are sufficiently thin for adequate fixation and
decalcification, whereas curettings may sometimes need to be sliced into thin-
ner fragments. The amount of curettings to submit for sectioning depends
on their volume and the uniformity of the curettings. When it is feasible, all
curettings should be submitted. If the lesion curetted is a hyaline cartilage
tumor, as much of the histology as possible should be reviewed to identify
atypical chondrocytes as well as any subtle interface with normal surrounding
bone.
Other large specimens such as total joint replacements and bone resections
also need to be sliced into thinner fragments. Although this may be accomplished
with large band saws or other power-type saws, motorized saws are dangerous
and somewhat time-consuming to maintain properly, especially in a laboratory
that receives a limited number of bone specimens. Vibrating or oscillating saws,
which are usually available in autopsy suites, should be avoided if possible,
because they do not section uniformly and their oscillating movement creates
tension and compression artifacts that often make bone sections impossible to
interpret properly. A very easy approach is to hold bone specimens steadily in
a tabletop vise or clamp and to cut them with a hacksaw in which two fine-
tooth blades are separated by 2- to 3-mm-thick washers. Such an apparatus is
easily and cheaply made, although there are commercial instruments available
for the same purpose. It must be emphasized that double-bladed instruments
should be scrupulously cleaned between every specimen to avoid tissue cross-
contamination between different cases.
The handling and disposition of larger resection specimens depends on the
reason for the procedure. For malignant tumors in which patients have not
received neoadjuvant chemotherapy (after biopsy but prior to resection), grad-
ing, staging, and adequacy of resection are the major clinical issues. Amputations
from these patients should include sections from the soft tissue and vascular mar-
gins as well as those from the tumor itself. Tumor sections should be taken in
such a way as to document the pertinent tumor histology, whether the tumor
involves the medulla and/or cortex, and how far the tumor extends into soft
tissues. The specimen should be cut in such a way as to disclose the greatest
extent of tumor; review of the imaging studies can guide the selection process.
Careful attention should be paid to taking sections from any areas that are
grossly disparate from the appearance of the majority of the tumor. Radical
resections for malignant tumors that are not amputations need the same sec-
tioning methods, but any area of the resection constituting a margin must be
sectioned and appropriately designated. This includes the bone resection mar-
gin, overlying soft tissue dissection margins, and the margins of any skin and
soft tissue encompassing a prior biopsy site.
Specimens resected from patients who have received neoadjuvant chemother-
apy (currently used in osteosarcoma and in Ewing sarcoma/primitive neuroec-
todermal tumor [EWS/PNET]) need more extensive sampling to estimate the
extent of treatment-associated necrosis. This means that one or more thin slabs
should be cut through the entire extent of the bone and tumor, and that the
entire slab or slabs should be fixed, decalcified, mapped, and examined not
only for tumor stage, but also for the extent of necrosis. The slabs should be
photographed so as to produce a section map; if a specimen x-ray machine
is available, specimen radiographs can be used both as section maps and as
controls for adequate specimen decalcification (e-Fig. 48.3). Additional sections
may be taken if there are areas not in the slab selected that appear as though
they might be viable; the pathologist's task in this enterprise is to find viable
tumor if any is present. It is worthwhile to remember that to extrapolate the
degree of necrosis in a single slab into necrosis of the tumor as a whole makes
the assumption that what is present in that particular slab is representative of

the entire lesion.
B. Decalcification. The main difference between processing of bone specimens and
of softer tissues is the requirement for an extra step of decalcification. Removal
of calcium insures that bone collagen is no harder than the paraffin in which
it is embedded, and that microtomy of bone tissues will approximate that for
other types of specimens. Decalcification may be performed in a number of
ways. In acid decalcification, hydrogen ions are in effect substituted for cal-
cium ions. Electrolysis in effect accomplishes the same end, but is performed
in an electrolyte solution with a weak electrical current. Ionic exchange is the
slowest method but is the most gentle on tissue and results in the fewest arti-
facts. In practice, most histology laboratories rely on weak acid decalcification
because it is the quickest and there is pressure from eager clinicians for rapid
turnaround times in diagnosis. With use of acid decalcification methods, a few
caveats must be kept in mind. First, the tissue must always be fixed adequately
prior to decalcification to prevent artifacts that interfere with adequate stain-
ing or that can degrade the tissue after sections are prepared. This means that
the tissue must be adequately thin (no more than 3- to 4-mm thick) prior to
fixation, and that the tissue has remained in formalin or some other suitable
fixative for an interval adequate to coagulate the proteins for routine staining.
In addition, if immunohistochemistry needs to be performed, adequate fixation
helps to insure that the decalcification process will less alter tissue antigens.
Second, when decalcification is performed with acid solutions, specimens must
be rinsed in running water to ensure that the residual pH of the tissue is suffi-
ciently neutral for hematoxylin staining. Failure to neutralize the acid not only
results in understaining with hematoxylin, but also will cause stained sections
to lose their hematoxylin staining in an accelerated manner. If time is insuffi-
cient for adequate specimen rinsing, the specimen should be neutralized in a
dilute basic solution such as sodium bicarbonate. Third, if sections are left in
dilute acid for a much longer period than necessary for calcium removal, tissue
hydrolysis will remove the nucleic acids that cause nuclear hematoxylin staining
and nuclei will appear acidophilic. This so-called overdecalcification artifact is
generally not reversible. Overdecalcification may not interfere with many diag-
nostic interpretations, but it is important not to mistake this artifact for tissue
necrosis, particularly in postchemotherapy specimen interpretation.
Adequate decalcification will vary by the tissue being decalcified. For exam-
ple, woven bone, even though it tends to have higher calcium concentrations
than lamellar bone, will often section adequately with incomplete decalcifica-
tion because the former has less organization and less cutting resistance. The
decision regarding whether the tissue is ready for embedding is often subjective
and revolves around whether the tissue is pliable, trims easily, or can be pen-
etrated with a needle. Complete decalcification is best judged either by testing
the supernatant fluid with a colorimetric indicator or by comparing specimen
radiographs prior to and after decalcification. These tests are seldom practical
in a very busy general surgical pathology practice.
C. Approach to the interpretation of bane specimens. Patient complaints related to
the musculoskeletal system constitute nearly one-third of physician office visits
in the United States, so orthopedic problems are extremely common. While
surgical pathologists are often asked to rule out bone tumors as the etiology of
a clinical problem, it is useful to keep in mind that fractures alone are about
3000 to 4000 times more common than all primary bone tumors combined, and
that metastatic tumors to bone are at least 20 times more common than primary
bone tumors. The accurate diagnosis of bone diseases requires the correlation
of patient demographics along with the clinical history and imaging studies to
put the problem in its correct context prior to any histologic examination of the
Chapter 48 • Bone Neoplasms and Other Nonmetabol ic Disorders I 7 93
tissue. Symptoms and signs are fairly similar in orthopedic diseases; these consist
of pain, loss of function, deformities, and (in the case of tumors) sometimes a
mass or a sense of fullness. Pain is the most common symptom, and although it
may vary considerably, pain severe enough to wake a patient from sleep is the
type suspicious for neoplastic diseases.
D. Importance of radiologic findings. The surgical pathology of orthopedic diseases
most often consists of defining the nature of bone lesions that are space occupy-
ing on imaging studies, advising the clinician if an infection may be present, and
histologically documenting miscellaneous bone diseases that are not diagnosable
by imaging studies alone. Because surgical pathologists usually render biopsy
diagnoses with the assumption that a biopsy is representative of the pathologic
process, it is natural to assume the same parameters in bone biopsies. This
is a potentially dangerous assumption, because most orthopedic diseases are
invisible without imaging studies. This means that to assure that a biopsy is rep-
resentative of a process, the smaller the biopsy specimen, the greater the need to
review the imaging studies defining that process. Because bones are deep seated,
imaging studies are required to grasp the extent and behavior of bone lesions.
For a surgical pathologist, correlating imaging studies with histologic findings
depends on knowledge of normal bone and joint anatomy, normal anatomy as
represented in radiographic images or other imaging studies, and the rudimen-
tary alterations in imaging studies produced by pathologic processes (not only
what a process does to normal bone, but also how normal bone alters the pro-
cess) (Adv Anat Pathol. 2005;12:155). The majority of the radiographic image
produced by long bones is due to beam attenuation by cortical bone in the shafts
and by cancellous bone in the ends (e-Fig. 48.4). The attenuation produced by
flat bones is primarily due to cortical bone, and that of irregular bones depends
on the proportion of bone elements in any given part of the bone.
Space-occupying lesions within bone usually cause bone destruction, bone
production, or some combination of the two. Destructive lesions are not seen
in a single radiographic view until at least 40% of the bone in the path of
the x-ray beam is destroyed. This means that almost an entire thickness of
cortex must be destroyed to see the lesion if an intact and a destroyed cortex
are superimposed in one view, or that at least 40% of cancellous bone must be
destroyed in a bone end. It is partly for this reason that orthogonal views of bones
are taken (e.g., posteroanterior and lateral views) so that destructive lesions may
be isolated in routine radiographs. Radiodense lesions superimpose on the extant
bone, causing more attenuation and easier visibility on a routine radiograph. In
contrast, a lesion that is less dense than bone may fill the entire medullary cavity
of a bone, but if it does not destroy the cortex it will be invisible regardless of the
view because the dominant attenuator of the x-ray beam is the cortex, not the
medullary fat or marrow. It is for these reasons that other imaging studies such
as computed tomography (CT) scans and magnetic resonance imaging (MRI)
are performed. These studies yield information that is complementary to that
derived from routine radiographs. While they may be more sensitive in yielding
information, a particular type of imaging modality should be used in concert
with routine radiographs to answer a particular clinical question not answered
by the radiographs.
Ill. DIAGNOSTIC FEATURES OF BONE LESIONS. There are very few general categories of
bone disease (Table 48.1 ), although there are many individual diseases (McCarthy
EF, Frassica FJ. Pathology of Bone and joint Disorders with Clinical and Radio-
graphic Correlation. Philadelphia: WB Saunders; 1998). Most patients can be
separated into general diagnostic categories on the basis of their imaging stud-
ies. For example, traumatic diseases, which are among the commonest problems,
will demonstrate fractures (with or without bone displacement) or dislocations
on routine radiographs. Metabolic bone diseases (discussed in Chap. 49), which
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formation by tumor from that of trauma, although it takes some experience

to recognize them. Bone or osteoid production by tumor matrix is often lace-
like and becomes sheetlike as more bone is produced. Bone produced as a repair
phenomenon may be focally lacelike, but more often it rapidly acquires a micro-
trabecular architecture and then becomes trabecular as it matures. In reactive
bone there is almost always a zonation of maturity that is dependent on both the
area in the lesion sampled and the time from trauma. While bone and cartilage
are common findings in both osteosarcoma and fracture callus, cartilage tends
to disappear as callus matures but it persists in osteosarcoma (e-Fig. 48.6). In
addition, the progression from bone to cartilage and back to bone is orderly in
reactive processes but is totally random in bone tumors.
C. Circulatory disturbances
1. Bone necrosis. Osteonecrosis occurs in areas where the bone circulation has
an end arterial distribution. The most common sites are near the convex sur-
faces of joints where epiphyseal arterial branches supply the cancellous bone
in the distribution of a cone. When this area of bone is deprived of circula-
tion, avascular or aseptic necrosis of the bone results. The cancellous bone
up to the calcified zone of the articular cartilage, deriving its blood supply
from nutrient arteries to the epiphysis, undergoes infarction. The overlying
articular cartilage, which derives oxygen and nutrients from the synovial
fluid, remains viable. These changes are not immediately visible on routine
radiographs because there are no changes in density of the necrotic bone.
However, radionuclide bone scans do demonstrate early hypervascularity in
the zone surrounding the necrotic area, and MRI demonstrates edema and
loss of marrow fat because of early adipocyte necrosis. The wedge-shaped
area of radiodensity characteristic of late osteonecrosis develops for a vari-
ety of reasons, but deposition of calcium salts due to saponification of free
fatty acid esters may be of greatest importance (although it is often difficult
to recognize calcium salts in decalcified sections because they are dissolved
by the decalcification process). Clinical symptoms become severe when the
necrosis has extended to the articular cartilage with loss of congruency of
the usual convex-concave joint surface and destruction of the subarticular
plate. It is not uncommon for the articular cartilage and superficial subartic-
ular plate to detach from the underlying cancellous bone (because dead bone
matrix and living bone have the same inherent strength and stiffness, this
probably happens because the subarticular bone no longer has the capacity
for remodeling in the face of repetitive forces, and accumulated shear stress
causes it to detach). When detachment occurs, the radiodense subarticular
bone attached to the articular cartilage forms a crescentic shadow that may
be seen radiographically (e-Fig. 48.7).
2. Bone infarctions. Infarcts are also presumably the result of a disruption in
end arterial circulation. In the diaphysis, a bone infarct is largely confined to
the medulla. This portion of the bone derives its blood supply from nutrient
arteries that penetrate the cortex to supply the sinusoids of the medullary
cavity and the inner cortex. The saponified marrowfat resulting from fat
necrosis may appear to contain hazy or smoky radiodensities, and biopsies
will reveal fat necrosis and a few scant trabeculae with empty osteocyte
lacunae (e-Fig. 48.8).
The outer cortex is supplied mainly by perforating arterioles derived from
arteriae comitantes of the periosteum. The cortical portions of this circulation
travel longitudinally via Haversian canals and interconnect within the cor-
tex via the Volkmann canals. Because the circulation in the cortex is thereby
microscopically collateralized, the cortex of long bones is somewhat more
protected against infarction than is the medullary cavity. It is important to
remember that within the cortex there are interstitial lamellae derived from
the remnants of old Haversian systems, and inner or outer circumferential

lamellae that have not fully resorbed but have no active blood supply, and
because of this, physiologically there are lamellae that are devoid of osteo-
cytes and are physiologically dead (e-Fig. 48.9); since all cortical bone is
compact bone, this means that small foci of empty osteocyte lacunae within
the cortex do not necessarily imply that there is avascular necrosis even
though the bone is histologically dead. Ordinarily, it is necessary for both
nutrient and periosteal blood supplies to be disrupted to cause a true cortical
infarction. This happens most often in conjunction with trauma and with
infections.
D. Paget disease. This disease has some histologic features in common with high-
turnover metabolic bone diseases, but it is not a metabolic disease because it does
not diffusely affect the entire skeleton and has no known associated metabolic
defect (metabolic bone diseases are covered in Chap. 49). Paget disease is char-
acterized by an imbalance or uncoupling of osteoclastic and osteoblastic activ-
ities, with osteoclastic bone resorption predominating early in the disease and
osteoblastic activity persisting late in the disease. These histologic manifesta-
tions are correlated radiographically with characteristic radiolucency early in
the disease, radiodensity in the late stages, and a mixed pattern for most of the
interval between (Skeletal Radiol. 1995;24:173). Because the bone microarchi-
tecture is altered, there is loss of the normal bone contour radiographically, and
there is gradual loss of the normal cortical appearance and an increasingly coarse
appearance to the bone trabeculations. A biopsy from an early radiolucent lesion
demonstrates large bizarre osteoclasts producing large and irregular resorption
pits (Howship's lacunae) on trabecular surfaces. These are often accompanied
by paratrabecular fibrosis and dilated marrow sinusoids. As the resorption pits
become filled in by osteoblast activity, irregularly shaped cement lines (some-
times likened to grout lines in a mosaic) mark the demarcation between the old
and new bone. The bone on either side of these cement lines demonstrates either
lamellar bone, in which the layers are discontinuous on either side, or lamellar
bone on one side and woven bone on the other side. As the disease progresses
and osteoclast activity slows, the bone becomes thicker and more interconnected
than normal, but its arrangement and increased irregular cement lines make it
more prone to deformities and fractures (e-Fig. 48.10).
E. Infectious disorders. Infections of bone arise either by direct introduction of
organisms into the bone due to open trauma or overlying infections of soft tis-
sue, or by secondary hematogenous spread. Most hematogenous osteomyelitis
occurs in the first two decades of life. Its usual site in the bone is in the metaph-
ysis adjoining the growth plate of a long bone because the microcirculation is
stagnant in this area. Osteomyelitis due to open trauma can occur at any age;
osteomyelitis associated with overlying infections is most often associated with
peripheral vascular disease and so is seen later in life. Most infections of the
bone are bacterial, but infections with fungi and lower virulence organisms may
occur in immunocompromised hosts.
The vast majority of hematogenous osteomyelitis is due to coagulase-positive
Staphylococcus aureus, but many other organisms may infect bones. Histo-
logically, microorganisms are seldom seen in bone biopsies of patients with
osteomyelitis because the sheer number of organisms required for the sensitiv-
ity of high-power or oil-immersion microscopy to detect bacteria is very high.
Because of this, bacterial cultures should always be taken when infections of
bone are suspected clinically-preferably prior to the institution of antibiotic
therapy. A single bacterial culture is on the order of 10 million times more sen-
sitive than histology-even when special stains for organisms are added to the
regimen. Infections in bone are often accompanied by necrosis of at least some
of the affected bone; the primary reason for this is that edema accompanies
inflammation, and edema in the closed confines of the cortex compromises the
medullary nutrient arteries and sinusoids due to resulting increased pressure.
The innermost cortical circulation may be similarly compromised by increased
intramedullary pressure. If the pressurized exudate finds its way into empty
Haversian and Volkmann canals, it may push its way through these intracor-
tical spaces and eventually dissect the periosteum, and its perforating arteries,
from the cortex. If the cortex is deprived of its dual circulation, then it in turn
becomes necrotic; this necrotic bone is called sequestrum. The combination
of necrotic bone sequestrum, marrow fibrosis and/or fat necrosis, and mixed
inflammatory infiltrates (usually including neutrophils and plasma cells) pro-
vides good histologic corroboration of osteomyelitis, but the demonstration of
organisms is the gold standard for the diagnosis of infections (e-Fig. 48.11).
F. Iatrogenic disorders. Treatment-related disorders are seldom a major problem in
the pathologic diagnosis of orthopedic disease, provided that an accurate clinical
history is communicated to the surgical pathologist. For example, the diagnosis
of osteosarcoma would be very unusual in a patient of the sixth decade with-
out prior radiation of the site, or without some other underlying premalignant
bone lesion. Administration of various therapeutic regimens may lead to sec-
ondary alterations in bones; perhaps the most notable of these is the amyloidosis
of bones, tendon sheaths, and ligaments that develops from ,82-microglobulin
accumulation in long-term hemodialysis patients. Substances that have been
given parenterally but that are not metabolized may also be deposited in bones
or joints; without prior knowledge of therapeutic treatment, it may be difficult
to make an accurate diagnosis (e-Fig. 48.12).
G. Neoplastic and tumor-like lesions. Primary tumors of bone are quite rare,
accounting for only 0.2% of all malignancies, or an incidence of 1 per 100,000
individuals per year (Fletcher, CDM, Unni K, Mertens K, eds. Tumors of Soft
Tissues and Bone. Lyon, France: IARC Press; 2002). There is a bimodal age
distribution, with one peak in adolescence and a smaller one in patients older
than 60 years. Among other characteristics, each bone tumor has its own age
predilection, which is very useful from a differential diagnostic standpoint. Pri-
mary benign bone tumors are probably less common than primary malignant
tumors if the very common nonossifying fibroma, osteochondroma, and enchon-
dromas of the hands are excluded. In addition to benign and malignant bone
neoplasms, there are a number of nonneoplastic lesions that can present in a
manner similar to neoplastic conditions (Table 48.3 ); all of these lesions are
discussed below, and their main features are presented in Tables 48.4 and 48.5.
Pathologic stage is among the findings that are recommended to be reported for
bone tumors (Hum Pathol. 2004;35:1173) and the American Joint Commit-
tee on Cancer (AJCC) Tumor, Node, Metastasis (TNM) staging scheme (Table
48.6) and/or the simpler Musculoskeletal Tumor Society scheme (Table 48. 7)
can be used for this purpose.
1. Cartilage-forming tumors
a. Osteochondroma is a cartilage-capped bony protrusion (e-Fig. 48.13) that
arises from the surface of any bone that models or grows by endochon-
dral ossification. On imaging, osteochondromas demonstrate a marrow
cavity and a cortex continuous with those of the host bone. Although
classified as bone neoplasms, osteochondromas may also result from
displacements of the cartilaginous grown plate. This is consistent with
their metaphyseal location and the fact that they cease to grow after
skeletal maturation. Most osteochondromas are sporadic and solitary;
however, multiple lesions are present in osteochondromatosis, which is
an autosomal dominant hereditary condition. The presence of EXT-1
mutations in the germline of these patients has been used as evidence
of the classification of osteochondroma as a neoplasm U Clin Invest.
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cortical thickening. When peripheral, the cartilage matrix is usually

>3 em in thickness (e-Fig. 48.20).
Histologically, chondrosarcoma is composed of mature-appearing
hyaline cartilage except that the chondrocytes have varying degrees of
increased cellularity, nuclear atypia, and even mitotic activity. Chon-
drosarcoma can show variable degrees of differentiation, ranging from
minimally hypercellular tumors resembling enchondromas with scat-
tered enlarged hyperchromatic tumor cell nuclei that are sometimes binu-
cleate (grade 1), to unequivocally malignant tumors with markedly atypi-
cal cells and easily identifiable mitotic figures (grade ill). Grade II tumors
have features intermediate between the two (e-Fig. 48.21). Regardless
of their grade, chondrosarcomas (when sampled adequately) invariably
show permeation of existing marrow spaces between bony trabeculae
(e-Fig. 48.22); this is a very helpful feature for distinguishing low-grade
tumors from enchondromas, especially in the small bones of the hands
and feet where enchondromas may show a degree of hypercellularity
and/or binucleation quite reminiscent of that seen in low-grade chon-
drosarcomas of larger, more proximal bones. The presence of tumor cell
necrosis can also point to a diagnosis of chondrosarcoma. Nevertheless,
there are cartilaginous tumors that remain difficult to accurately cat-
egorize, especially when the radiologic features of malignancy are not
clearly evident. Occasionally, complementary imaging studies such as
CT scans may help to identify true bone destruction in a central carti-
lage tumor, or MRI will demonstrate the extent of a cartilage cap in a
peripheral cartilage tumor, which in turn can help to identify the true
biologic nature of the lesion when a small biopsy cannot (e-Fig. 48.23).
The prognosis of chondrosarcoma is mostly dependent on grade and
completeness of resection (as no other modality of treatment is effec-
tive), with 5-year survival rates ranging from 90% for low-grade tumors
to 53% for higher-grade tumors.
f. Dedifferentiated chondrosarcoma. In addition to areas of classic chon-
drosarcoma (usually low grade), this tumor is characterized by the pres-
ence of a distinct, second, clearly defined, high-grade, noncartilaginous
sarcomatous component (e-Fig. 48.24). The later is most frequently
represented by a malignant fibrous histiocytoma-like component, but
osteosarcoma, fibrosarcoma, and rhabdomyosarcoma have also been
reported. The tumor has a very poor prognosis.
g. Mesenchymal chondrosarcoma is a rare tumor also characterized by a
dimorphic pattern and is composed of a highly undifferentiated small
round cell component that is often arranged in a hemangiopericytoma-
tous pattern, intermixed with a variable number of islands of hyaline
cartilage (e-Fig. 48.25). Although it may occur at any age, the peak age
incidence of patients with this tumor (second and third decades) is earlier
than that seen in patients with other chondrosarcomas. Mesenchymal
chondrosarcoma has a high incidence of local recurrence and distant
metastasis, although the latter may not occur for 5 to 10 years.
h. Clear cell chondrosarcomais another rare type of chondrosarcoma; this
tumor shows a predilection for the ends of long bones after the growth
plates have closed (e-Fig. 48.26). It is characterized histologically by
the presence of abundant large round clear cells with well-defined cell
borders, intermixed with areas of conventional low-grade chondrosar-
coma (e-Fig. 48.27). The clear cells contain large amounts of intra-
cellular glycogen and stain strongly for S-100 protein. The prognosis
is similar to that of low-grade chondrosarcoma. Metastases occur in
about 20% of patients, and may behave indolently or aggressively. Clear
cell chondrosarcoma has a high predilection for metastasis to other

bones.
2. Bane-farming tumors
a. Osteoma is a well-circumscribed, radiodense, benign lesion that most fre-
quently arises in the jaws and paranasal sinuses (e-Fig. 48.28), but can
also be seen in long bones. Some cases are sporadic, whereas others
arise in association with familial polyposis coli (see Chap. 14). Histo-
logically, osteomas are composed of mineralized compact bone matrix
with a variable admixture of mature and immature bone but no cellular
stroma (e-Fig. 48.28).
b. Osteoid osteoma. This benign, self-limited tumor usually presents with
pain that often wakes patients from sleep, but is relieved by aspirin.
Although it usually involves the cortices of long bones, osteoid osteoma
has been reported in almost every skeletal site. Radiographically, there
is a central area of radiolucency surrounded by dense reactive sclerosis
(e-Fig. 48.29). The quantity of sclerosis varies by location in the bone. If
the lesion is cortical, the reactive sclerosis may obscure the lesion such
that it can only be seen by thin-cut CT scans. In the medullary cavity,
there may be little or no sclerosis.
Histologically, the radiolucent area, termed the "nidus," is composed
of vascularized fibroconnective tissue in which immature new bone is
being formed. This new bone is usually arranged in microtrabecular
arrays lined by plump appositional osteoblasts that lack nuclear pleo-
morphism (e-Fig. 48.30). Simple excision or curettage of the nidus of an
osteoid osteoma is curative.
c. Osteoblastoma is virtually identical histologically to osteoid osteoma
but, unlike the latter, is not limited in growth potential. When diag-
nosed, osteoblastomas are usually > 2 em in diameter. Radiographically,
they may resemble large osteoid osteomas, they may be expansile like
aneurysmal bone cysts (ABCs), or they may even appear as aggressive as
malignant tumors. There is also a predilection to involve the axial skele-
ton, especially the vertebral pedicles and arches. Occasionally, osteoblas-
tomas may be very cellular, and their osteoblasts may be several times the
size of usual osteoblasts. Tumors having predominant areas of this histo-
logic feature have been termed "aggressive osteoblastoma" or "epithe-
lioid osteoblastoma" (e-Fig. 48.31). The prognosis of osteoblastomas is
excellent if amenable to excision.
d. Osteosarcoma is the most common nonhematopoietic primary malignant
neoplasm of bone. The peak incidence of this tumor is late childhood
and adolescence; howeve.t; there is a second peak in patients older than
40 years, most cases of which develop secondarily in preexisting bone
lesions (e.g., Paget disease) or following irradiation. The metaphyses of
long bones (femur, tibia, and humerus) are the most common sites of
involvement; isolated diaphyseal involvement is rare, and involvement
of the epiphyses of long bones or small bones of the hands and feet is
exceptionally rare. Other sites of involvement include the jaws, skull, and
axial skeleton. Most cases of osteosarcoma present with pain (often dull
and unremitting) with or without a palpable mass. Radiologically, there is
almost always evidence of a destructive bony lesion, often with evidence
of new bone formation. There may also be an interrupted periosteal
reaction and soft tissue involvement (e-Fig. 48.32).
The histologic hallmark of osteosarcoma is the presence of osteoid or
bone formation directly by tumor cells. Osteoid appears as dense, pink,
amorphous material (resembling collagen or amyloid) that has a lacelike
or sheetlike appearance (e-Fig. 48.33). Intermixed within, and often in
direct contact with this osteoid matrix, are the neoplastic tumor cells
which can be quite variable in appearance and include polyhedral cells,
spindle cells with variable nuclear atypia, small blue cells (resembling
EWS, see below), and large markedly atypical cells. The predominant
matrix produced by the tumor can be bone or osteoid, cartilaginous,
or fibrous. Historically, osteosarcomas have been subclassified on the
basis of the predominant matrix production (osteoblastic, chondroblas-
tic, or fibroblastic), but this classification has no prognostic importance.
Instead, as described below, osteosarcomas are best classified on the basis
of radiologic and/or pathologic features that have been shown to have
distinct prognostic implications (Am] Clin Pathol. 2006;125:555).
i. Central osteosarcomas arise within the medullary cavity and include
the following:
(a} Conventional intramedullary osteosarcoma is the prototypical
osteosarcoma, for which most of the above information refers.
Neoplastic tumor cells in conventional osteosarcoma are often
polyhedral or spindle-shaped with unequivocally malignant fea-
tures. Given that preoperative chemotherapy for these tumors is
the current standard of care (and has significantly improved the
5-year survival rate of this tumor from 20% to over 80% ), it is
important to carefully map the tumor in the resection specimen (as
discussed earlier) to determine the extent of tumor necrosis com-
pared to the volume of viable plus nonviable tumor since a favor-
able long-term outcome is associated with > 90% tumor necrosis.
Additional chemotherapy is often offered to patients with less
necrosis as a second-line attempt to further improve survival.
(b) Law-grade central osteosarcoma is a rare type of osteosarcoma (1%
to 2%) composed of a variably cellular spindle celVfibroblastic
proliferation that lacks the degree of cytologic atypia seen in con-
ventional osteosarcoma. In addition, bone production is usually
evident as irregular, somewhat thick, anastomosing, or branching
bony trabeculae. These trabeculae simulate the woven bone of
fibrous dysplasia or the longitudinal seams of bone in parosteal
osteosarcoma (see below), and are separated by a spindle-cell
stroma. Review of the radiologic findings often reveals subtle
signs of malignancy that are useful in making the diagnosis
(e-Fig. 48.34). This tumor has a much more indolent course com-
pared with conventional osteosarcoma; however, there is still a
high recurrence rate if the tumor is inadequately excised, often
with associated grade progression.
(c) Telangiectatic osteosarcoma is characterized by large, blood-filled
spaces separated by highly cellular fibrous septae that contain
markedly pleomorphic cells with a variable amount of osteoid
production (e-Fig. 48.35). Radiologically, this tumor is radiolu-
cent and expansile, and resembles ABC. Compared to other
osteosarcomas, it is more likely to present with a pathologic frac-
ture. Although not necessarily associated with improved survival,
this aggressive osteosarcoma is very sensitive to chemotherapy.
(d) Small cell osteosarcoma is composed of small round blue cells
and histologically resembles EWS except that there is histologic
evidence for osteoid formation (although it is often scant). It is
usually entirely radiolucent. It has a capricious clinical behavi01;
often but not always resistant to usual osteosarcoma chemother-
apy. Although the usual reciprocal chromosomal translocation
described in EWS (see below) has not been generally observed,
this lesion has been shown to have membrane positivity for CD99,
which has led some authors to theorize that it is a variant of EWS
with divergent differentiation.
ii. Surface osteosarcomas. About 1 in 20 osteosarcomas occurs in asso-
ciation with the bone surface rather than in the medullary cavity. The
vast majority of these are low-grade tumors showing radiodensity and
osseous differentiation. They include:
(a) Parosteal osteosarcoma accounts for "'4% of osteosarcomas
and the majority of surface osteosarcomas. It characteristically
involves the posterior distal femur, is associated with the outer
fibrous layer of the periosteum, and tends to wrap around the
bone. Histologically, it consists of well-formed bony trabeculae,
often arranged in parallel streamers separated by a hypocellular
spindle stroma as seen in low-grade central osteosarcoma (e-Fig.
48.36). Cartilaginous differentiation is also common, often seen
as a cartilage cap and sometimes causing confusion with osteo-
chondroma. Radiographically, however, there is no continuity of
the interior of parosteal osteosarcoma and the medullary cav-
ity, the adjacent bony cortex is not continuous with the outside
of parosteal osteosarcoma (e-Fig. 48.37), and the intertrabecu-
lar spaces do not contain fatty or hematopoietic marrow. The
prognosis of parosteal osteosarcoma is similar to that of low-
grade intramedullary osteosarcoma. If inaccurately diagnosed as
benign, or if inadequately excised, these lesions will recur. Recur-
rences may be low grade, but they may also be high grade; low-
grade parosteal sarcoma undergoing high-grade transformation is
termed dedifferentiated parosteal osteosarcoma and has a prog-
nosis similar to conventional osteosarcoma.
(b) Periosteal osteosarcoma arises between the cortex and overlying
periosteum most commonly in the tibial or femoral diaphysis, and
is characterized by abundant cartilaginous matrix and a some-
what greater degree of cytologic atypia than is seen in parosteal
osteosarcoma.
(c) High-grade surface osteosarcoma is histologically identical to con-
ventional intramedullary osteosarcoma, except that it arises on
the bone surface. The prognosis is similar to that of conventional
intramedullary osteosarcoma.
3. Fibrous tumors and tumor-like conditions
a. Fibrous dysplasia. This space-occupying lesion has been classified vari-
ously as developmental, tumorous, or tumor-like. It usually presents as
a solitary lesion, although it may affect multiple bones in a single limb
bud distribution, or multiple bones without limb bud distribution. The
polyostotic form is one of the manifestations of the McCune-Albright
syndrome, which includes pigmented skin lesions and endocrinopathies.
The presence of activating G-protein mutations in both monostatic
lesions and McCune-Albright syndrome suggests that fibrous dysplasia
represents a true neoplasm UPediatr. 1993;123:509). Fibrous dysplasia
may be asymptomatic, but deformities, secondary fractures, and even
pain may be the presenting manifestation. Radiographically, the lesion
is almost always intramedullary, and it tends to affect those portions
of bone formed by endochondral ossification. While secondary corti-
cal atrophy may take place because of intramedullary expansion of the
lesion, fibrous dysplasia usually does not involve the cortex. Fibrous dys-
plasia is often expansile and results in modeling deformities of the host
bone. It is well circumscribed and radiolucent, but less radiolucent than
the underlying bone that it has replaced; radiologists often refer to its
appearance as having a ground glass quality (e-Fig. 48.38).
Histologically, fibrous dysplasia consists of various combinations of
any tissue present in bone, so fibrous tissue, bone, cartilage, and vascu-
lar tissues are produced in various combinations. The usual microscopic
pattern, however, consists of loosely arranged, vascularized fibrous tis-
sue in which disconnected curved microtrabeculae of bone are disposed.
These trabeculae are not only woven in their collagen fiber pattern, but
when a section is examined under polarized light, the fabric of their col-
lagenous background forms a continuum with the fabric of the fibrous
tissue (e-Fig. 48.39). Cartilageformation is not unusual, and occasionally
cartilage is formed in such excess that lesions may be mistaken radio-
graphically and histologically for cartilaginous neoplasms. Treatment is
usually focused on relief of deformities or other morbid symptomatology.
The prognosis is usually excellent.
b. Osteofibrous dysplasia is a fibro-osseous lesion and is invariably seen in
the tibia, fibula, or both. Its peak incidence is in the first two decades
of life. Radiographically, the lesion is radiolucent and usually based on
the cortex; it may extend to the medullary cavity. The lesion may be
unilocular or multilocular; while it tends to be circumscribed, it may
also diffusely involve the diaphysis and cause secondary bowing defor-
mities (e-Fig. 48.40). Histologically, osteofibrous dysplasia resembles
fibrous dysplasia except that the microtrabeculae of bone tend to be
rimmed by appositional osteoblasts even at their very earliest synthesis
(e-Fig. 48.41). The fibrous stroma tends to be more cellular than in
fibrous dysplasia, and is less contiguous with the trabeculae under polar-
ized light. The lesional bone also tends to mature at the periphery with the
surrounding normal bone. The lesional bone tends to undergo sponta-
neous involution with time, although sometimes it behaves more aggres-
sively.
c. Nonossifying fibroma {fibrous conical defect) is the commonest space-
occupying lesion of bone, estimated to affect one in four individuals.
Even though it is thought to be developmental, in rare cases, the lesion
behaves as a tumor of limited biologic potential. Unless it is associated
with a fracture, patients are generally without symptoms; the lesions are
typically discovered incidentally during the course of evaluation for some
other condition and the routine radiographs are virtually diagnostic. The
lesion is a well-circumscribed radiolucent defect in the metaphyseal cor-
tex with scalloped sclerotic borders, and it is almost always longer in the
cephalocaudal than axial direction (e-Fig. 48.42).
Histologically, the lesion consists of spindle cells arranged in a
distinctly storiform pattern. A fair number of multinucleated giant
cells, histiocytic cells with foamy cytoplasm, and histiocytes contain-
ing hemosiderin pigment (e-Fig. 48.43) may also be present. While bone
formation is not observed (hence the name), lesions that have fractures
or microscopic infarctions are admixed with reactive bone. Because this
lesion may be focally cellular, it is important to review the radiographs
to avoid misdiagnosis. Most nonossifying fibromas are self-healing and
do not require clinical intervention.
d. Desmoplastic fibroma. This rare tumor occurs in adolescents and young
adults, with the mandible being the most commonly affected site. Radi-
ologically, it often expands the involved bone, is entirely radiolucent,
and is usually well circumscribed. Histologically, it is composed of bland
fibroblastic or myofibroblastic cells in a background of collagen iden-
tical to that found in desmoid tumors and/or soft tissue fibromatosis
Chapter 48 • Bone Neoplasms and Other Nonmetabolic Disorders I 809
(e-Fig. 48.44). Although it is benign, it behaves similarly to fibromatosis

of soft tissue in that there is a high recurrence rate when not completely
excised.
e. Fibrosarcoma constitutes ""5% of all primary malignant bone tumors
with a relatively uniform age distribution between the second and sixth
decades. It usually involves the metaphyses of long bones resulting in
pain, swelling, and a destructive radiologic lesion without radiographi-
cally detectable matrix. Histologically, fibrosarcomas are usually quite
cellular with malignant spindle cells arranged in a fascicular or her-
ringbone pattern (e-Fig. 48.45 ). The differential diagnosis includes
other malignant spindle-cell tumors such as fibroblastic osteosarcoma,
leiomyosarcoma, malignant fibrous histiocytoma, and desmoplastic
fibroma.
4. Histiocytic and fibrohistiocytic tumors
a. Benign fibrous histiocytoma is a rare bone lesion that is histologically
similar to its soft tissue counterpart. There is a wide age distribution,
with more than half of the cases developing in patients older than
20 years. The tumor most commonly involves either the epiphysis or
the diaphysis of long bones, or the pelvis. Radiographically, the lesion is
well defined and radiolucent, and may expand the bone. Similar to benign
fibrous histiocytoma elsewhere, the lesion is composed of spindle-shaped
fibroblasts at least focally arranged in a whorled or storiform pattern,
intermixed with histiocytes and giant cells (e-Fig. 48.46). Mitoses may
be evident. The main differential diagnosis is a secondary fibrohistiocytic
reaction within another primary bone lesion, such as giant cell tumor or
nonossifying fibroma.
b. Malignant fibrous histiocytoma is also similar to its soft tissue counterpart,
with most cases developing in patients older than 40 years. It can also
complicate Paget disease, or occur after irradiation or infarction. Most
cases involve the long bones of the extremities or the pelvis. Histologi-
cally, there is a mixed population of spindle cells, histiocytic cells, and
giant cells. Pleomorphic tumor cells, abnormal mitotic figures, and the
characteristic storiform pattern of growth are also evident (e-Fig. 48.4 7).
Almost all of the histologic subtypes described in soft tissue have also
been described in bone. The management and prognosis of this tumor
most closely resemble osteosarcoma; the focal identification of osteoid
or bone matrix is sometimes the only histologic difference between these
two tumors, although most osteosarcomas arising in this age group are
secondary to a prior disease or treatment.
c. Langerhans cell histiocytosis comprises a group of neoplastic Langer-
hans cell proliferations that can be unifocal (solitary eosinophilic gran-
uloma), multifocal (Hand-Schiiller-Christian disease), or disseminated
(Letterer-Siwe disease). All can produce bone lesions that tend to present
early in multifocal and disseminated forms. Langerhans cell histiocyto-
sis most frequently involves the craniofacial bones, but other bones such
as the femur, pelvis, and ribs may also be involved. Radiographically,
the lesions appear radiolucent and rapidly destructive, sometimes with
associated exuberant periosteal new bone formation when they occur in
long bones (e-Fig. 48.48).
Histologically, there is a mixed inflammatory infiltrate including neu-
trophils, eosinophils, lymphocytes, and histiocytes (with or without giant
cells) in which Langerhans cells are identified. Langerhans cells have
eosinophilic to clear cytoplasm and contain oval, grooved, or multilo-
bated nuclei (e-Fig. 48.49); immunohistochemically, they characteristi-
cally express S-1 00 and CDla. Because there is a histologic similarity to
chronic osteomyelitis, lesions thought to be Langerhans cell histiocytosis

should be cultured, and the culture results should be known before for-
mulating a final diagnosis. Langerhans cell histiocytosis has a very good
prognosis except in the disseminated form, which is associated with a
poor outcome (Pediatr Blood Cancer. 2005;45:37).
d. Erdheim-Chester disease is a rare disorder of unknown etiology charac-
terized by the presence of skeletal and extraskeletal foamy histiocytic
infiltrates with associated fibrosis (e-Fig. 48.50). Most patients are older
than 40 years, and there is usually bilateral symmetric or patchy sclerosis
of the medullary cavity of the involved bones (most frequently the long
bones of the extremity). Given the frequent and progressive infiltration
of vital organs (such as the kidney, heart, or lung), most patients succumb
within a few years.
e. Giant cell tumor comprises around 4% to 5% of all primary bone tumors
and has a peak incidence between 20 and 45 years of age; it is rarely
seen in skeletally immature individuals or in patients older than 50 years.
Although there is some controversy as to its exact origin, it is placed in
the histiocytic category because its giant cells are modified histiocytes
and at least some of its stromal cells also express histiocytic markers.
Giant cell tumor typically affects the ends of long bones and extends
to the articular or apophyseal portions of the bone. The pelvis or small
bones of the hand or feet are more rarely affected. Given the often juxta-
articular location of the tumm; joint swelling and limitation of move-
ment are common presenting symptoms. Radiologically, the tumor is
radiolucent and eccentric, may expand the bone, and is well demarcated
(e-Fig. 48.51 ). There is almost never a periosteal reaction associated with
the tumot.
The histologic hallmark of the tumor is the presence of sheets of
round, oval, or elongated mononuclear cells with an open chromatin
pattern, evenly intermixed with numerous osteoclast-like giant cells with
nuclei similar to those of the mononuclear cells (e-Fig. 48.52). The cell
borders are often indistinct, so that on low power the lesion appears
as a syncytium. Mitoses are variable in number; atypical mitoses are
occasionally seen and do not necessarily predict malignant behavior.
The presence of the characteristic mononuclear cells and the even inter-
position of the giant cells are essential for the diagnosis, since giant
cells can be a component of many bone lesions. Other histologic fea-
tures occasionally seen in giant cell tumors include a focal storiform
pattern, which, when abundant foam cells are present, can easily be
confused with benign fibrous histiocytoma. There also may be areas of
fibrosis, as well as secondary cystic areas resembling ABC (see below).
The most important differential diagnosis is the so-called brown tumor
of hyperparathyroidism (see Chap. 49); the lack of radiologic or bio-
chemical evidence of hyperparathyroidism, as well as the absence of
additional bone lesions, can be very helpful in this regard. Another
important differential diagnosis is the so-called giant-cell reparative gran-
uloma that characteristically involves the mandible and is composed
of granuloma-like aggregates of giant cells in a fibrovascular stroma,
but which lacks the mononuclear cells characteristic of giant cell tumor
(e-Fig. 48.53).
Most cases of giant cell tumor behave in a benign fashion; some, how-
ever, are associated with local aggressiveness and occasionally ("'2%)
with distant metastasis. Most of these metastases grow slowly and are
rarely lethal. There are no reliable histologic features that can predict a
malignant outcome.
Chapter 48 • Bone Neoplasms and Other Nonmetabolic Disorders I 811
Malignant giant cell tumor is defined as a sarcomatous lesion arising

within a giant cell tum01; or as a sarcoma that appears in the same area
in which a bona fide giant cell tumor was treated (e-Fig. 48.54). The
former instance is sometimes referred to as a primary malignant giant
cell tumor; the latter as a secondary malignant giant cell tumor. The
prognosis is related to the histology, size, and grade of the malignant
component.
5. Hematolymphoid tumors
a. Solitary plasmacytoma and multiple myeloma. These tumors are malignant
proliferations of plasma cells that account for the majority of tumors
arising primarily in bone. The presentation of myeloma is quite variable
(see Chap. 44 ), but involvement of the skeletal system is usually mani-
fested by bone pain and/or pathologic fractures. Radiologically, plasma-
cytoma and the lesions of multiple myeloma are lytic, well demarcated,
and without a rim of sclerotic bone; however, multiple myeloma may
also present with generalized osteoporosis without any detectable foci
of discrete bone destruction. Histologically, the tumors are composed
of plasma cells and their precursors at various stages of development
(e-Fig. 48.55). The main differential diagnosis is often other hematolym-
phoid tumors, although myeloma cells are occasionally quite anaplas-
tic and can resemble carcinoma or high-grade sarcoma. Accordingly,
immunohistochemical reactivity for CD138 or CD38 (among other
markers; see Chap. 44) can be useful to confirm the diagnosis.
b. Lymphoma. Most bone lymphomas are secondary to disease elsewhere,
but primary bone lymphomas also occur. In general, lymphomas present-
ing in bone are classified as primary provided no lymph node involvement
is present both at the time of presentation and for a long interval after-
ward (the length of this interval varies according to different authors).
Most primary lymphomas of bone represent examples of diffuse large
B-celllymphoma or other high-grade tumor, because most lower-grade
lymphomas and leukemias present with diffuse marrow involvement
rather than as a tumorous mass. Primary Hodgkin lymphoma of bone is
exceedingly rare. Radiologically, lymphomas usually present as radiolu-
cent lesions, sometimes disproportionately destructive when compared
with the patient's clinical symptoms (e-Fig. 48.56). In about 20% of
cases, the lesions present as radiodensities. The histologic findings reca-
pitulate those seen in extraskeletal sites. The prognosis is dependent on
the type of lymphoma and stage.
6. Vascular tumors
a. Hemangioma. Although incidental hemangiomas are relatively common,
clinically symptomatic tumors account for < 1% of primary bone tumors
and tend to present in late adulthood. Hemangioma is most frequently
seen in the vertebrae, followed by the craniofacial skeleton and metaphy-
ses of long bones. It appears as a radiolucent, often expansive lesion in
long bones but as a vertically striate "corduroy pattern" lesion in intact
vertebrae. Histologically, it is composed of capillary sized or cavernous
vessels that permeate the marrow and are lined by bland endothelial cells
(e-Fig. 48.57). As the name indicates, this tumor, as well as the closely
related lymphangioma, is benign with low rates of recurrence following
excision.
b. Angiosarcoma and hemangioendothelioma. These two neoplasms account
for <1% of bone tumors and may present at any age group, although
the peak incidence is in young adulthood. They constitute a spectrum of
lesions ranging from locally destructive but indolent tumors with a good
response to surgery or local radiation, to poorly differentiated malignant
tumors with a high metastatic rate. Most tumors are radiolucent with
poor margination; howeve.t;, a sclerotic rim is occasionally identified.
While angiosarcomas are often solitary, hemangioendotheliomas tend to
present as multifocallesions in the same bone or in the same limb bud
distribution and may be mistaken for skeletal metastases (e-Fig. 48.58).
Angiosarcomas are usually characterized, at least focally, by the pres-
ence of irregularly anastomosing vascular channels that are lined by
highly atypical endothelial cells, but they may largely consist of solid, pat-
ternless aggregates of polyhedral or spindle cells (e-Fig. 48.59). Poorly
differentiated angiosarcomas may fail to express vascular markers. It
should be noted that angiosarcomas are known to express cytokeratins,
which is an important consideration when metastatic carcinoma is in the
differential diagnosis. Most angiosarcomas are associated with a poor
outcome.
Epithelioid hemangioendothelioma is composed of cords, nests, or
sheets of plump cells that, in their attempt to form vessels, are often vac-
uolated, and some of the vacuoles may contain erythrocytes. An extracel-
lular myxoid or hyalinized stroma is characteristic, although not always
identified. Immunohistochemical reactivity with vascular markers such
as CD31, CD34, and Factor VIII-related antigen can be used to con-
firm endothelial differentiation, and may help to exclude a diagnosis
of metastatic signet-ring cell adenocarcinoma. Epithelioid hemangioen-
dothelioma may have an indolent course.
7. Epithelial tumors
a. Adamantinoma comprises <1% of malignant bone tumors. There is a
wide age distribution, with the median age of patients between 25 and
35 years. The tibia is most frequently involved, followed by the fibula,
and both sites synchronously. Radiologically, an intracortical radiolucent
lesion is evident, which may involve the medullary cavity (e-Fig. 48.60).
Histologically, classic adamantinoma is composed of epithelial cells
having a basaloid, tubular, or squamoid appearance; has a predomi-
nantly spindle cell pattern; or consists of a mixture of the two patterns
(e-Fig. 48.61); a storiform fibroblastic proliferation that contains vari-
able amounts of woven or lamellar bone may also be present. Rarely,
the tumor is predominantly composed of the latter component, with
only rare scattered epithelial cells (single or in small nests, sometimes
only detected by immunohistochemistry); such tumors have been termed
as having an "osteofibrous dysplasia-like pattern" or as "differenti-
ated adamantinomas,, and usually present in patients younger than
20 years. Adamantinomas are invariably immunopositive for various
keratins and epithelial membrane antigen, and often also for vimentin.
Classic adamantinomas are indolent tumors with a high local recur-
rence rate and metastasis in only about 20% of patients; differentiated
adamantinomas almost never metastasize.
b. Metastatic carcinoma is the most common tumor affecting the skeleton,
which is the third most common site to be involved by metastatic carci-
noma after the lungs and liver. Primary carcinomas of the breast, lung,
prostate, kidney, and thyroid gland compose >80% of all bone metas-
tases. Radiologically, metastatic deposits can be radiolucent or radio-
dense or can display a mixed pattern (e-Fig. 48.62). Given its high
incidence, metastatic carcinoma should always be in the differential diag-
nosis of solitary or multiple bone lesions in patients older than 40 years.
The histology usually resembles that of the primary carcinoma, if it
is known. It should be noted that a fibroblastic, osteoblastic, or vascular
response to the metastatic tumor may occasionally be quite prominent
and overshadow the tumor cells, which may only be focally evident
(e-Fig. 48.63). Consequently, immunohistochemistry may reveal isolated
subtle tumor cells that are not obvious in small biopsy specimens.
8. Miscellaneous neoplasms
a. EWSJPNET. In addition to its classic presentation in the diaphyses of
long bones, the tumor can involve axial bones such as the pelvis and
ribs, as well as soft tissues (see Chap. 46) and other organs. EWSIPNET
has been described at various ages, but most patients are younger than
20 years. Although patients usually have pain and a mass, they may
also present with fever and leukocytosis suggestive of an infectious pro-
cess. Radiologically, a destructive, permeative lesion is usually evident,
often with an overlying multilayered but discontinuous "onion-skin"
periosteal reaction (e-Fig. 48.64 ). Occasionally, there is a large soft-tissue
mass with no obvious bone destruction on the routine radiographs, but
the intraosseous component becomes evident on aCT scan or MRI.
Histologically, EWS/PNET is the prototype for "small blue round cell
tumors" as it is composed of sheets of such cells, often with glycogen-
containing clear cytoplasm (e-Fig. 48.65). Evidence of neuroectodermal
differentiation may be manifested by extracellular eosinophilic neuropil-
like structures or Homer-Wright rosettes (composed of groups of tumor
cells that surround a central core of eosinophilic extracellular mate-
rial). Immunohistochemically, EWS/PNET shows characteristic strong
cell membrane immunoreactivity for CD99, negativity for CD45, and
variable reactivity for neural markers such as neuron-specific enolase,
synaptophysin, CD57, neurofilament, and S-100 protein. Cytokeratins
may also be positive.
A characteristic feature of this tumor is the presence of a recurrent
balanced reciprocal translocation involving the EWS gene on chromo-
some 22 and a member of the Ets family of genes, the most common of
which (85% of cases) is the FL11 gene on chromosome 11 (Br] Cancer.
1994;70:908). Immunohistochemistry, polymerase chain reaction, and
fluorescent in situ hybridization have all been used to confirm the diag-
nosis by detecting expressed FLI1, the fusion transcript, or the translo-
cation itself, respectively. The prognosis of EWS has greatly improved
with multimodality treatment, and 5-year survival rates now approach
70%.
b. Chordomas are derived from notochordal remnants and account for "'4%
of malignant bone tumors. Most tumors present after 30 years of age
with a peak incidence between 50 and 60 years of age. The midline of
the axial skeleton, particularly the sacrum and the base of the skull, is
usually affected. Radiologically, a lucent lesion is seen with scattered
calcifications; there is often a large associated soft-tissue component
(e-Fig. 48.66). Histologically, chordomas are composed of lobules of
tumor in which sheets, cords, or nests of vacuolated, eosinophilic to
clear cells are embedded in a myxoid matrix (e-Fig. 48.67). Chordo-
mas express cytokeratins, epithelial membrane antigen, and S-100 pro-
tein, an immunohistochemical profile that is helpful in distinguishing
this tumor from chondrosarcoma (which is cytokeratin and epithelial
membrane antigen negative). Chordomas are aggressive tumors that are
most notable for local recurrence when incompletely excised, but also
have a metastatic potential. A controversial entity, "chondroid chor-
doma," characterized by areas of mimicking hyaline cartilage, appears
to have a better prognosis, whereas "dedifferentiated chordoma," with
its high-grade sarcomatous component, is associated with a very poor
outcome.
9. Cystic/cyst-like lesions
a. Simple bone cyst is an intraosseous space-occupying lesion consisting of
an accumulation of fluid. The lesion is usually lined by a thin mem-
brane composed of flattened cells of unknown type that may be involved
in the production of the fluid, which usually appears serous. The base
of the lesion is usually situated at an active growth plate, and the cyst
is more or less maintained by continuous remodeling of the bone around
the area of fluid pressure. The lesion is circumscribed and radiolucent,
and usually involves the proximal humeral, femoral, or tibial region
(e-Fig. 48.68). It does not expand the bone or cause deformity unless
there has been a fracture with displacement before healing, and it tends
to be symmetric. While fluid may not be obvious radiographically,
the contents are demonstrable by MRI. Histologically, the diagnosis is
one of exclusion and depends upon correlation of the imaging, operative
findings, and lack of any other diagnostic tissue. Because most simple
cysts are treated conservatively by injection of steroids or other scleros-
ing agents, it is unusual to see the lining of a simple cyst histologically
unless there has been repeated fracture.
b. lntraosseous ganglion. This is a subarticular defect in the cancellous bone
filled with mucoid fluid. The bone surrounding the defect is remodeled
and sometimes sclerotic (e-Fig. 48.69). The defect is histologically similar
to the subarticular cysts associated with overlying osteoarthritis (geodes)
except that the subarticular plate and articular cartilage are radiographi-
cally intact in patients with intraosseous ganglia. The lesion is presumed
to arise from a microscopic continuity of the subarticular plate with the
joint space that either cannot be detected by imaging studies or that has
healed; this etiology would allow pressurized synovial fluid to come in
contact with the intertrabecular medullary space. There are no diagnos-
tic features histologically as the concentrated fluid is practically acellular
and resembles the contents of tenosynovial ganglia.
c. Aneurysmal bone cyst. This peculiar lesion derives its name from its
expansile character. It is not a true cyst, but rather a collection of
blood-filled spaces that are separated by fibro-osseous tissue septa con-
taining a varying amount of multinucleated giant cells and immature
bone. Most ABCs occur prior to 20 years of age. The lesion usually
arises as an eccentric radiolucent lesion in metaphyses of long bones
(e-Fig. 48.70). When it is central and symmetric it can often be distin-
guished from a simple cyst because it causes the bone to become wider
than the growth plate. The bone destruction associated with ABC is per-
haps the most rapid associated with any osseous lesion. While its inter-
nal edge tends to be well marginated radiographically, ABC sometimes
extends across adjacent bones, particularly if it arises in the spine. Com-
plementary imaging studies, particularly MRI, reveal peculiar fluid-fluid
levels on T2-weighted axial and sagittal views (e-Fig. 48.71); these levels
are caused by the signal differences in erythrocytes and plasma, reflect
erythrocyte sedimentation, and demonstrate that blood in intact ABC is
both unclotted and stagnant or slow moving.
Histologically, the lesion has vascular spaces that progress from very
small capillary spaces to very large sinusoids separated by fibrous septae
and sometimes by bone (e-Fig. 48.72). Within the septae are fibroblasts,
scattered multinucleated giant cells, and osteoblasts associated with the
bone production. Rarely, the lesion is almost entirely solid, although
sometimes the solid variant may demonstrate fluid levels on imaging. In
about half the cases, there is some other lesion associated with the cyst
and admixed with curetted fragments; this has been termed secondary
ABC. The associated lesion is usually benign, although malignant tumors

have also been described in association with ABC. Gene rearrangements
of the USP6 gene on chromosome 17, and/or the CDH11 gene of chro-
mosome 16, have been described in ABC, raising the possibility that
the lesion is truly neoplastic. The genetic abnormalities thus far seem to
apply mainly to the primary variety of this lesion, and have also been
found in the solid variant (Am] Pathol. 2004;165:1773).
1D. Other rare primary bone neoplasms. There are other benign and malignant
soft-tissue neoplasms that can primarily arise in bone, including leiomyoma,
leiomyosarcoma, lipoma, liposarcoma, and schwannoma. All of these are
histologically similar to their soft-tissue counterparts (see Chap. 46).
Metabolic Diseases
of Bone
Deborah Novack
I. NORMAL ANATOMY. Because of its accessibility and composition of both cortical

and trabecular (cancellous) bone, the iliac crest is the site of choice for evaluation
of systemic metabolic bone diseases. Cortex forms the external layer of all bones,
comprises "'80% of bone mass, and supports most of the tissue's mechanical func-
tion. Trabecular bone, the meshwork surrounded by marrow or fat, is much more
metabolically active than cortex. To support both its mechanical and metabolic
functions, bone is dynamically regulated, and the skeleton is replaced completely
every 10 years. The process of replacement, known as remodeling or turnover, is
accomplished by the coordinated action of bone-forming osteoblasts (OBs) and
bone-resorbing osteoclasts (OCs), and is regulated by a variety of systemic factors
including calcium, phosphorus, and parathyroid hormone (PTH). The goal of iliac
crest trochar biopsy is to assess this process.
In either the cortex or the trabeculum, remodeling begins when mononuclear
OC precursors (derived from hematopoietic progenitors) arrive at a bone surface,
fuse, and differentiate into functional polykaryons (Fig. 49.1). Mature OCs polar-
ize and secrete acid and proteases onto an isolated microenvironment of the bone
surface, excavating a pit known as Howship's lacuna. This resorption phase ends
with the OCs' apoptosis, and a reversal phase follows, characterized by activation
of OBs (of mesenchymal origin) to replace the excavated bone; the activity of OCs
and OBs is normally tightly coupled, and the amount of bone synthesized matches
the amount resorbed. Newly secreted matrix called osteoid becomes mineralized to
form mature bone. The remodeling cycle ends when new bone formation is com-
plete, and the OBs are either incorporated into the new bone matrix as osteocytes
or become quiescent surface bone lining cells. The net result of each cycle is the
formation of a new osteon, a packet of bone delineated by a "cement line" in which
the collagen fibers are aligned. These lamellae of bone are easily seen when decalci-
fied hematoxylin and eosin (H&E)-stained sections are examined under polarized
light (e-Fig. 49.1).*
OCs can be identified by their characteristic appearance as multinucleated cells
on the bone surface, with discrete nuclei (in contrast to megakaryocytes, which
have fused nuclei). The most sensitive method of identifying OCs histologically is
expression of tartrate-resistant acid phosphatase (TRAP), which stains OCs bright
red (e-Fig. 49.2), although this is not usually necessary for diagnosis. OBs appear
as cuboidal cells on the bone surface, often in rows, with abundant cytoplasm
and an eccentric nucleus (e-Fig. 49.3). However, the strength of the undecalci-
fied biopsy is in the evaluation of the function of these cells rather than their
morphology.
OBs secrete matrix proteins onto the bone surface, but several days are required
for mineral deposition. Therefore, the extent of bone surface covered by osteoid
(osteoid surface) is one indicator of OB activity. The thickness of the osteoid seams
reflects the rate of mineral apposition, because mineralization converts osteoid to
816
Chapter 49 • Metabolic Diseases of Bone I 81 7
bone
0 OC precursor @] 08
eoc £2.:::>
"'-.(
bone lin ing cell
~ apoptotic OC ..::::::::.._~- osteocyte

;-
Figure 49.1 The bone remodeling cycle. A: OC precursors are recruited to the bone surface,
where they fuse and differentiate into mature polykaryons. B: The OCs resorb both organic and
inorganic matrix of bone. C: The resorption phase ends with OC apoptosis. D: During the reversal
phase, OBs differentiate from mesenchymal precursors under the influence of factors from OCs
(cunted a"ow) and secrete new bone matrix known as osteoid. E: At the end of the remodeling
cycle, some OBs have been incorporated into the bone and become osteocytes, whereas others
remain on the surface as synthetically quiescent bone-lining cells.
bone. Decalcification required for standard paraffin processing removes the dis-
tinction between newly synthesized osteoid and mature calcified bone. In contrast,
undecalcified plastic sections can be stained in several ways to demonstrate osteoid.
The von Kossa stain, a silver-based stain used with a basic fuchsin counterstain,
shows calcified bone matrix as dark brown or black, whereas the unmineralized
matrix (osteoid) appears pink-red (e-Fig. 49.4A). A trichrome stain, either Goldner
(e-Fig. 49.3) or modified Masson (e-Fig. 49.4B ), also distinguishes mineralized bone
from osteoid. These latter stains allow easier interpretation of cellular morphology
than the von Kossa, and also highlight peritrabecular or marrow fibrosis.
A second critical marker of OB function is tetracycline labeling. Tetracycline
family antibiotics are calcium-chelating fluorochromes that bind to actively min-
eralizing bone surfaces, can be taken orally, and are well-tolerated. They are given
in two courses, separated by 2 weeks (see below). If bone formation is active dur-
ing both intervals, examination of unstained sections by fluorescence microscopy
demonstrates two bright bands of labeling (a double label) (e-Fig. 49.5). Similarly,
active bone formation during only one of the labeling periods yields a single tetra-
cycline label (e-Fig. 49.5). Combination of the extent of labeled trabecular bone
surface and the distance between labels provides the mineral apposition rate and
bone formation rate. In a normal subject, most surfaces with osteoid, as seen on
trichrome or von Kossa stains, have single or double labels.
During normal endochondral bone development, cartilage formed at the growth
plate is replaced by bone in the primary spongiosa through the action of OCs.
Toluidine blue stains cartilage purple, and cartilage may be found within trabeculae
near the growth plate in children (e-Fig. 49.6). However, a finding of entrapped
cartilage in an iliac crest bone biopsy in an adult, or > 1 em from the growth plate
in a child, is indicative of OC dysfunction such as in osteopetrosis.
II. INDICATIONS FOR BIOPSY, TISSUE SAMPLING, AND PREPARATION
A. Indications for biopsy. The most common indications for metabolic bone biopsy
are end-stage renal disease (ESRD) and unexplained hypercalcemia or hyper-
phosphatemia, osteoporosis unresponsive to therapy, or suspected osteomala-
cia. Patients who have multiple or unexplained fractures (particularly if they
are failing to heal), unexplained bone pain, or an elevation in serum alkaline
phosphatase may also be candidates.
B. Biopsy procedure. A critical component of evaluation of metabolic bone biop-
sies is in vivo fluorochrome labeling of bone via use of a regimen of tetracycline
(250 mg orally four times a day [PO qid]) for 3 days, followed by a 14-day inter-
val, then 3 more days of therapy (250 mg PO qid). Biopsy is performed on the
third day after the last dose (biopsy interpretation may therefore be confounded
by recent therapeutic use of antibiotic drugs in the tetracycline family; similarly,
inadequate labeling can be caused by malabsorption syndromes or by taking
tetracycline with meals, dairy products, iron-containing medications, antacids,
or calcium supplements).
Biopsy is performed as an outpatient procedure; the most accessible site for
biopsy is the anterior iliac crest. When obtained, the specimen should be placed
directly into 70% ethanol.
C. Sample preparation. The specimen should be fixed in 70% ethanol for at least
48 hours, and this solution is suitable for shipping and long-term storage at room
temperature. Following dehydration with xylene, the specimen is mounted in
methyl methacrylate, and the tissue core is sectioned parallel to its long axis at
5 to 7 J.£-m thickness using a tungsten blade. Undecalcified sections are stained
with von Kossa, Goldner or modified Masson trichrome, and toluidine blue.
One section is decalcified and stained with H&E. Thicker 10-J.£-m sections are
coverslipped without staining for examination under fluorescence.
D. Quantitative versus qualitative evaluation. The American Society for Bone and
Mineral Research has described a nomenclature for a basic set of structural
and kinetic features identified by nondecalcified bone biopsy (] Bone Miner
Res. 1987;2:595), and there are two commercially available systems that allow
quantitative analysis based on these standards (OsteoMeasure, OsteoMetries,
Inc. and Bioquant Osteo II, BIOQUANT Image Analysis Corporation). Refer-
ence values, based on somewhat limited populations, have been published (Engl
I Med. 1988;319:1698; I Bone Miner Res. 1988;3:133; Bone. 2000;26:103;
I Bone Miner Res. 2004;19:1628). Although some laboratories perform quan-
titative analysis on all specimens, qualitative assessments are often adequate
for diagnosis of individual patients. Quantitative analysis is most useful in the
setting of research studies.
Ill. DIAGNOSTIC FEATURES OF METABOLIC BONE DISORDERS
A. Osteoporosis. In the setting of osteoporosis or osteopenia, biopsy establishes
the rate of bone remodeling (turnover), degree of mineralization, architectural
integrity, and effects of treatment. lliac crest bone biopsy is a poor indicator
of bone mass, as bone volume/tissue volume (BV!TV) is variable within this
region. However, trabecular connectivity, which describes the intactness of the
trabecular meshwork, correlates with bone mass. In osteoporosis/osteopenia,
trabeculae are very small and often appear as isolated islands (low connectiv-
ity), rather than as an interconnected grid (good connectivity) (e-Fig. 49.7).
In high-turnover osteoporosis, osteoid surface is enhanced (e-Fig. 49.8A), with
normal or increased numbers of OCs and OBs. The extent of double tetracycline-
labeled trabecular bone surface is also increased, although the distance between
the double labels is usually normal (e-Fig. 49.8B). In low-turnover osteoporosis
Chapter 49 • Metabolic Diseases of Bone I 8 19
or osteopenia, there is little osteoid, few OCs or OBs, and rare or absent tra-
becular double tetracycline labels (e-Fig. 49.9). Even in low-turnover states,
double labeling in the cortex is usually present, and is a good positive control
for adequate tetracycline dosing and specimen processing.
Many patients with osteoporosis have been treated with bisphosphonates,
often for several years. Although biopsy studies have shown that normal
turnover is intact in most patients (]AMA. 2006;296:2927), some cases of
severely suppressed bone turnover have been reported and are associated
with increased fractures (] Clin Endocrinol Metab. 2005;90:1294; N Engl]
Med. 2006;355:2048). Bisphosphonates target the OC, decreasing resorption
and enhancing apoptosis, producing changes that can readily be seen in tis-
sue sections with OCs appearing either hyperchromatic with pyknotic nuclei
(e-Fig. 49.10A) or round and unpolarized (e-Fig. 49.10B).
B. Osteomalacia. In osteomalacia, newly formed organic bone matrix fails to min-
eralize normally, and the result is wide osteoid seams, often greatly increased
in extent along the trabecular bone surface (e-Fig. 49.11A). Some osteoid may
be completely unlabeled by tetracycline (e-Fig. 49.11B), whereas other surfaces
may show irregular and diffuse fluorescence (e-Fig. 49.11C); double tetracycline
labels are rare. Florid osteomalacia due to nutritional rickets (vitamin D defi-
ciency) is rare, but milder cases may be found unexpectedly and bone biopsy is
the only definitive diagnostic tool. Osteomalacia is also seen in fluorosis, usually
caused by excessive fluoride in drinking water (e-Fig. 49.110 and E). The degree
of osteomalacia in hypophosphatasia, a rare deficiency of alkaline phosphatase
activity, can be quite severe (e-Fig. 49.11F).
Tumor-induced osteomalacia (110, also known as oncogenic osteomalacia)
is a rare form of osteomalacia that can occur in both adults and children. lliac
crest bone biopsy in TIO shows the typical features of osteomalacia described
above (e-Fig. 49.12A). The bone manifestations of TIO are caused by mes-
enchymal tumors that secrete the phosphatonin FGF-23, leading to hypophos-
phatemia due to renal phosphate wasting. The tumors are typically small and
slow-growing, and can be difficult to detect. If not visible by CT or MRI, ses-
tamibi or octreotide scans may be useful, as well as targeted venous sampling
for FGF-23 (Expert Rev Endocrinol Metab. 2009;4:435). Histologically, the
majority of the tumors in cases of TIO are best characterized as phosphaturic
mesenchymal tumor (mixed connective tissue variant) and have low cellularity,
bland spindled cells, distinctive "grungy" calcified matrix, and an incomplete
rim of membranous ossification and/or formation of an osteoid-like matrix at
least focally (Am J Surg Pathol. 2004;28: 1) (e-Fig. 49.12B). They may be locally
infiltrative into muscle or fat (e-Fig. 49.12C), have prominent vessels reminiscent
of hemangiopericytoma, myxoid change, hemorrhage, or OCs. TIO tumors typ-
ically stain for FGF23 but not S100, CD34, or smooth muscle actin. Resection
of the solitary tumor mass results in complete resolution of hypophosphatemia
and clinical symptoms in the majority of cases.
C. Renal osteodystrophy. Chronic kidney disease-metabolic bone disease (CKD-
MBD) has been defined as a systemic disorder of bone and mineral metabolism
due to CKD manifested by one or more of the following: (i) abnormali-
ties of calcium, phosphorus, PTH, or vitamin D metabolism; (ii) abnormal-
ities of bone turnover, mineralization, volume, linear growth, or strength;
(iii) vascular or other soft tissue calcification (Kidney Int. 2006;69:1945). CKD-
MBD is associated with increased morbidity and mortality. Three predomi-
nant patterns in bone biopsy have been described: high bone turnover with
osteitis fibrosa (hyperparathyroid bone disease), low bone turnover (includ-
ing low-turnover osteomalacia and adynamic bone disease), and mixed uremic
osteodystrophy.
1. The most important role of biopsy is in the setting of hypercalcemia, in which

a finding of PTH-driven high turnover indicates a need for parathyroidec-
tomy, whereas a finding of low turnover is a contraindication for this surgery.
Bone changes can occur relatively early in CKD, and low turnover correlates
with increased vascular calcification.
2. In high-turnover renal osteodystrophy, elevated PTH levels drive bone resorp-
tion by many OCs; peritrabecular fibrosis, also known as osteitis fibrosa,
may also be present (e-Fig. 49.13A). Bone formation is accelerated, often
with formation of disorganized woven bone (seen on polarization; e-Fig.
49.13B) and increased tetracycline double labels (e-Fig. 49.13C). Trabeculae
may be irregular in shape, and the cortex may be porous due to increased
resorption.
3. In either form of low-turnover osteodystrophy, trabecular surfaces appear
quiescent, with few OBs or OCs. Trabecular connectivity may also be
decreased, and this parameter is more useful than the overall BV!fV (as
mentioned earlier in the discussion of osteoporosis). In the osteomalacic form
of renal osteodystrophy, mineralization is delayed and wide osteoid seams
identified by von Kossa or trichrome staining (e-Fig. 49.14A) are not tetracy-
cline labeled (e-Fig. 49.14B). In adynamic bone disease, both osteoid (e-Fig.
49.14C) and tetracycline labels (e-Fig. 49.14D) are minimal or absent. In the
past, aluminum toxicity from dialysis was often the cause of low-turnover
renal osteodystrophy, but the etiology of the adynamic changes seen more
recently is not known.
4. Mixed uremic osteodystrophy, as the name implies, shows features of increased
PTH and defective mineralization. Additionally, the appearance may vary
considerably within the specimen (e-Fig. 49.15A), with areas of increased
turnover adjacent to more quiescent regions with poor mineralization. Some
areas may show abundant osteoid and robust tetracycline double labels
(e-Fig. 49.15B and C) adjacent to broad single labels or unlabeled osteoid
(e-Fig. 49.15D).
D. Glucocorticoid-induced osteoporosis. Because long-term glucocorticoid therapy
is widely used in chronic inflammatory diseases and in organ transplantation,
there is a high prevalence of glucocorticoid-induced bone disease. Early in treat-
ment, glucocorticoids increase bone resorption, but more prolonged therapy
eventuates in adynamic bone in which the number of OCs and OBs is decreased,
as is tetracycline double labeling.
E. Primary hyperparathyroidism. Diagnosis of most cases of primary hyperparathy-
roidism is based on elevated serum calcium and PTH levels. However, in nor-
mocalcemic patients with variable or borderline PTH levels, bone biopsy can
be useful in making the diagnosis because histologic findings represent the net
effect of PTH over time. As in secondary hyperparathyroidism, such as in high-
turnover renal osteodystrophy, sections typically show increased osteoid sur-
faces (some with woven bone), increased tetracycline labeling, and elevated
numbers of OCs and OBs. Osteitis fibrosa cystica, the formation of cystic bone
loss due to elevated OC activity associated with marrow fibrosis, is found only
in severe cases and due to routine testing of serum calcium is rarely seen. How-
ever, peritrabecular fibrosis may be present. Another effect of PTH is to increase
the porosity of the cortex, which may be difficult to distinguish from trabecular
bone.
Focal well-demarcated osteolytic lesions known as brown tumors are also
associated with hyperparathyroidism (e-Fig 49.16). Brown tumors consist of
clusters of multinucleated giant cells in a fibrotic stroma, and the associated
islands of bone are often woven rather than lamellar. Brown tumors can occur
in any bone as a single lesion, or be multifocal. They generally resolve when
PTH levels are restored to normal.
Chapter 49 • Metabolic Diseases of Bone I 821
SUGGESTED READINGS
Monier-Faugere MC, Langub MC, et al. Bone biopsies: a modern approach. In: Avioli LV, Krane
SM, eds. Metabolic Bone Disease and Clinically Related Disorders. San Diego, CA: Academic
Press; 1998:237-273.
Recker RR, Barger-Lux MJ. Bone biopsy and histomorphometry in clinical practice. In: Favus M,
ed. Primer on the Metabolic Bone Diseases and Disorders of Mineral Metabolism. Washington,
DC: American Society for Bone and Mineral Research; 2006:161-169.
Joints and Synovium
Peter A. Humphrey
I. NORMAL ANATOMY. Joints are composed of the ends of contiguous bones and the
associated soft tissue elements, including cartilage, ligaments, tendons, and syn-
ovium. Diarthrodial movable joints, which are the most common type, are usually
covered by hyaline cartilage. Histologically, this articular cartilage is hypocellular
with a glassy extracellular matrix composed mainly of collagen, proteoglycan, and
water. Embedded within the matrix are chondrocytes within surrounding spaces
(lacunae). There are four zones in articular cartilage-superficial, intermediate,
deep, and calcified (e-Fig. 50.1)"' and chondrocytes have a different appearance
depending on their location. Those near the surface of the articular cartilage are
small and flattened; in the middle zones, the chondrocytes are more rounded and
arranged in columns. The deep and calcified layers of articular cartilage are sepa-
rated by a thin, basophilic line known as the tidemark, which represents the miner-
alized front. The calcified cartilage base interdigitates with underlying subchondral
bone.
Ligaments, which join two adjacent bones, are formed mainly of collagen. At the
insertion site onto bone the ligamentous tissue is calcified. Tendons are connective
tissue structures connecting muscle to bone. Microscopically, scant fibroblasts are
found within parallel collagen bundles.
Synovium is a glistening white membrane with delicate villous projections
that lines the inner surface of the joint capsule (Am j Clin Pathol. 2000;14:773)
(e-Fig. 50.2). The inner lining surface is created by synovial cells, including fibrob-
lastlike cells and histiocytes, which are arranged as a thin two- to three-cell layer
of closely packed cells with elliptical nuclei and abundant cytoplasm. A fibrous or
fibroadipose supporting layer lies beneath the synovial celllayet: Synovium also
lines the flexor tendons of the hand and bursae (subcutaneous and subtendinous
sacs).
II. GROSS EXAMINATION AND TISSUE SAMPLING. Joint or synovial soft tissue is received
as needle core tissue, as fragments from arthroscopic or open synovectomy, as
fragments from revised total joint arthroplasty, or as excisions of soft tissue ten-
don sheath or extra-articular masses. If fragments are received, the numbe.t; colo.t;
shape, and aggregate size of the fragments should be recorded. If meniscus tissue
is submitted, fibrillations or tears should be noted. If chalky white deposits are
identified, some of the tissue should be placed into absolute (100%) alcohol to
preserve the crystals. Decalcification may be required for calcified cartilage, bone,
or soft tissue. Fragments that appear different from normal should be selected for
embedding, along with representative unremarkable-appearing fragments.
A. Revision arthroplasties. The presence or absence of necrosis, purulent exudate,
and foreign material should be recorded. The explanted prosthesis should be
described, including any identification numbers or defects.
B. Soft tissue tumors. For excised soft tissue tendon sheath or juxta-articular masses
size, color, consistency, shape, and nodularity (single vs. multiple) should be
provided. The outer surface of the specimen should then be inked, and cut
sections should be characterized as to color, presence or absence of hemorrhage
and necrosis, and distance of tumor to inked margin. One section per centimeter
822
Chapter 50 • Joints and Synovium I 8 23
of tumor is a useful guide for section submission. Demonstration of tumor in

relation to the closest inked margin(s) and to any recognizable normal tissue is
important.
C. Joint replacement surgeries. Joint tissue may also be removed along with bone
in orthopedic joint replacement surgeries, such as knee and total hip replace-
ment procedures, which are most often performed for osteoarthritis. In such
cases, where there is identifiable bone and attached articular cartilage, gross
examination is particularly important. The overall dimensions and shape of the
submitted bone and soft tissue should be recorded. Articular cartilage presence
or absence, color, thickness, and abnormalities such as loss, cleft, or tuft forma-
tion, crystalline deposits, and bony and cartilaginous overgrowths (osteophytes
or exostosis) should be documented. Synovium col01; thickness, and consistency,
and any nodules or villous projections should be described. One or several sec-
tions of synovium should be submitted for formalin fixation. Any associated
gross bone defects, such as subchondral cyst formation and superficial bony
necrosis, should be noted. Again, if chalky white deposits are identified, tissue
should be placed into absolute (100%) alcohol to preserve the crystals. Sections
of macroscopically abnormal areas should be submitted for histologic examina-
tion as follows. The bone and overlying cartilage should be fixed overnight in
formalin, decalcified, and sectioned into 3- to 5-mm slices; one or two sections
are usually sufficient to document cartilaginous and associated bone abnormal-
ities. Junctions of normal and abnormal cartilage with underlying bone should
be demonstrated by the sections.
D. Synovial fluid. Examination can be extremely useful in the diagnosis of different
types of arthritis, especially infectious arthritis.
Ill. DIAGNOSTIC FEATURES OF COMMON DISEASES OF JOINTS AND SYNOVIUM
A. Osteoarthritis (also known as degenerative joint disease) is defined by the Amer-
ican College of Rheumatology as a "heterogeneous group of conditions that
leads to joint symptoms and signs which are associated with defective integrity
of articular cartilage, in addition to related changes in the underlying bone at
the joint margins" (Semin Arthritis Rheum. 2005;3:1). It is the most common
disease of the joints after the age of 65 years, with a prevalence of about 60%
in men and 70% in women. The etiology of osteoarthritis is multifactorial,
with inflammatory, metabolic, and mechanical causes. Major trauma, repeti-
tive joint use, chronic inflammatory arthritis, and congenital malformations are
major risk factors. The diagnosis is usually based on clinical and radiographic
features. Pathologically, although the name osteoarthritis indicates an inflamma-
tory condition, disruption of the articular cartilage is the fundamental finding.
The joints most commonly affected are the distal and proximal interphalangeal
joints of the hands, the hips and knees, and the cervical and lumbar spine.
Grossly, cartilaginous thinning, disruption, and fibrillation can be seen. In
areas of complete cartilage loss, the underlying bone is exposed; this bone has
a dense polished appearance like marble (known as eburnation). Microscopi-
cally, vertical clefts in the cartilage are characteristic (e-Fig. 50.3). There may be
associated villous hyperplasia and mild chronic inflammation of the synovium
(which should not be confused with rheumatoid arthritis). Papillary masses of
metaplastic cartilage, bone, or adipose tissue may form in the synovial mem-
brane; detachment of the masses results in intra-articular loose bodies (known
as rice bodies). In eburnated areas, the bone may show sclerotic thickened bony
trabeculae, cysts with fluid and fibromyxoid tissue, and superficial bony necro-
sis. It should be noted that in the vast majority of cases, the pathologic findings
are confirmatory of the clinical diagnosis, so it has been suggested that routine
pathologic examination in uncomplicated total hip and total knee arthroplas-
ties may not be necessary. In only a small percentage of cases is the pathologic
diagnosis different from the clinical impression; in these discrepant cases, the
pathologic diagnosis in most cases is avascular necrosis, rheumatoid arthritis,

pseudogout, or pigmented villonodular synovitis (]Arthroplasty. 2000;15:69;
Bone joint Surg Am. 2000;82:1531). Although rare, these cases with unex-
pected findings argue that microscopic examination of all cases of osteoarthritis
is warranted.
B. Rheumatoid anhritis is a chronic multisystem disease of unknown cause. The
hallmark of rheumatoid arthritis is a persistent inflammatory synovitis, typi-
cally involving the peripheral joints in a symmetric distribution. Clinically, the
synovial inflammation causes swelling, tenderness, and limitation of motion.
Histologically, the main attribute is joint destruction. There is hypertrophy and
hyperplasia of the synovium along with a lymphoplasmacytic infiltrate, which
together generate a papillary/polypoid chronic synovitis (e-Fig. 50.4 ). Lymphoid
follicles and acute fibrinous surface exudate (e-Fig. 50.5) can also be seen. Syn-
ovial giant cells and bone and cartilage fragments may be present in the syn-
ovium. This overall histologic picture is not specific for rheumatoid arthritis,
and similar changes may be seen in other arthritides such as systemic lupus ery-
thematosus and psoriasis. Destruction of cartilage and joint fusion (ankylosis)
can occur due to the formation of pannus (inflamed synovium and granulation
tissue) over the surface of the articular cartilage, with invasion into cartilage
and even bone and joint capsule.
Extra-articular manifestations of rheumatoid arthritis, which usually occur
in patients with high titers of rheumatoid factor, include rheumatoid nodules,
vasculitis, pleuropulmonary involvement (such as fibrosis and serositis), and
splenomegaly with neutropenia in Felty syndrome. Rheumatoid nodules develop
in "'25% of patients with rheumatoid arthritis. Although common locations
include the olecranon bursa, proximal ulna, and Achilles tendon, they can
also be found in the heart, lung, pleura, kidney, and meninges. Histologically,
there is a central zone of fibrinoid necrosis surrounded by palisading histiocytes
(e-Fig. 50.6).
C. Synovitis associated with loose large joint anhroplasty can be seen when there is
failure of a total joint replacement, and can be related to a foreign body-type
inflammatory response or infection. The foreign material can be metal, plastid
polyethylene, and/or methyl methacrylate cement. Microscopically, metallic
particles are present as 1- to 3-~m particles within macrophages. Needle-shaped,
polarizable fragments of polyethylene plastic are usually found in foreign body
giant cells. Methyl methacrylate cement is lost upon processing, and conse-
quently the cement causes empty spaces within foreign body-type giant cells
in histologic sections after routine processing. These different types of foreign
material can also elicit an exuberant fibrohistiocytic response (e-Fig. 50.7).
When neutrophils are seen, particularly when they are numerous, the pos-
sibility of infection should be considered. Frozen section of the inflamed tissue
from a failed joint replacement may be requested, and the number of neutrophils
per high-power field (HPF) should be reported. At least 5 HPFs on at least two
sections should be examined. Observation of ~5 neutrophils per HPF in tissue
(not fibrin) is associated with high specificity (of about 95%) for the absence of
infection, whereas the presence of >5 neutrophils per HPF has a sensitivity of up
to 69% in identification of the presence of infection (Mod Pathol. 1998;11:427).
D. In crystal-induced synovitis, deposition of microcrystals in joints and peri-
articular tissues results in gout (urate crystals), pseudogout (also known as
chondrocalcinosis-calcium pyrophosphate dihydrate crystals), and apatite dis-
ease (hydroxyapatite crystals) (Am] Clin Pathol. 2000; 14:773 ). Gout and pseu-
dogout are definitively diagnosed by light microscopic detection of crystals in
joint fluid. Gouty tophi are formed when urate crystals are deposited in subcu-
taneous soft tissue, synovium, bone, or bursae, with a granulomatous response
dominated by histiocytes and foreign body giant cells (e-Fig. 50.8); fixation of
the tissue in alcohol is necessary to preserve the crystals. A de Galantha histo-

chemical stain can be used to highlight the crystals. The crystals of pseudogout
appear rhomboid and purple in nondecalcified H&E-stained slides (e-Fig. 50.9);
with decalcification, the crystals are lost and what is left are rounded pools of
basophilic material surrounding rhomboid areas that mark the prior site of the
crystals. These crystal deposits do not typically elicit a histiocytic or foreign
body giant cell reaction (Semin Diagn Pathol. 2011;28:37).
E. Infectious arthritis is diagnosed by clinical history and joint fluid examination,
including Gram stain of a centrifuged cell pellet and microbiologic culture.
F. Hemosiderotic synovitis follows chronic intra-articular hemorrhage, which can
occur in patients with hemophilia and synovial hemangioma. Microscopically,
there are fine villous projections early in the disease course. Hemosiderin is
present within synoviocytes and macrophages (e-Fig. 50.10). Osteoarthritis
usually ensues.
G. Baker cyst, which can be found in osteoarthritis and rheumatoid arthritis, is
a synovial-lined cyst in the popliteal space that is formed by herniation. In
contrast, ganglia (ganglion cysts), located near a joint capsule or tendon sheath,
are not synovial-lined cysts and do not communicate with the joint cavity.
H. Tissue from a tom meniscus from the knee may be submitted as tissue shavings
or as a fibrocartilaginous loose body. Histologically, there are few changes in
the avascular and collagenized tissue because reparative fibrosis and neovascu-
larization are uncommon.
I. Each intervertebral disc is a type of joint (amphiarthrodial), and tissue frag-
ments from a prolapsed disc may be submitted. Microscopically, fibrous tissue,
fibrocartilage, and cartilaginous tissue can be seen. Chondrocyte necrosis and/or
groups of proliferating chondrocytes may be found. The presence of neovascu-
larization indicates herniation (Hum Pathol. 1988;19:406).
J. Synovial tumors are uncommon and can arise from synovium of the tendon
sheaths, bursae, and joint spaces (Semin Diagn Pathol. 2011;28:37). Regard-
less of their articular or extra-articular location, this family of neoplasms shares
translocations of chromosome 1p 13 (Table 46.1 ), which often also involve 2q35
resulting in formation of a COL6A3-CSF1 fusion gene (Proc Natl Acad Sci
USA. 2006;103:690). It is uncertain whether synovial tumors are best classified
as fibrohistiocytic tumors (as in the WHO classification) or as tumors differen-
tiating toward synovial cells.
1. Giant cell tumor of tendon sheath, localized type (localized tenosynovial tumor,
nodular tenosynovitis) is the most common benign tumor of the tendon
sheath and synovium. It appears to be a neoplasm rather than a reactive
process. These giant cell tumors are usually found in adults on the fingers,
and uncommonly on the ankle and knee (Cancer. 1986;57:875). Grossly,
they are circumscribed lobulated masses a few centimeters in diameter, with
mottled pink-gray cut surfaces that often show flecks of yellow or brown
(representing lipid and hemosiderin, respectively). Microscopically, there is
a vague nodularity at low magnification; at higher powers of magnification,
rounded mononuclear cells, giant cells, foam cells, and collagen are seen in
varying proportions (e-Fig. 50.11). Hemosiderin deposition may be present.
Mitotic figures are uncommon, and when they exceed two figures per 10
HPFs, the likelihood of recurrence is greatet: Immunohistochemistry and elec-
tron microscopy have demonstrated macrophage and synovial cell features
in the lesional cells (and osteoclast attributes in the giant cells), but neither
technique is required for the diagnosis. Similarly, although cytogenetic stud-
ies have demonstrated clonal abnormalities (especially involving 1p11-13)
in the lesional cells, karyotypic analysis is not needed for diagnosis.
Tenosynovial giant cell tumors are benign, with a recurrence rate of
about 10% to 20% related to mitotic activity (as noted earlier), cellularity,
and incomplete excision. Accordingly, margin status and a high mitotic rate
should be reported.
2. Giant cell tumor of tendon sheath, diffuse type (pigmented villonodular synovi-
tis) presents as an intra-articular or extra-articular tumor. The gene expres-
sion profile of the neoplasm resembles that of activated macrophages (Arthri-
tis Rheum. 2006;54:1009), and structural rearrangements of 1p11-12 have
been reported in the lesion; however, these special studies are not needed to
establish the diagnosis.
a. The intra-articular form typically involves the joint space of the knee of
young adults, although ankle, hip, and shoulder involvement have also
been reported. Involvement of the synovium can be localized with the
formation of either nodular or pedunculated masses; the more common
diffuse form affects virtually the entire synovium. Grossly, tissue from the
diffuse form removed by open synovectomy is spongy, diffusely thickened,
and brownish-yellow. Microscopically, subsynovial fibrohistiocytic cells
with uniform round to oval nuclei are seen. Foam cells and iron pigment
are always present, and giant cells are common and tend to be arranged in
groups. Aside from the villous structures, the overall appearance is similar
to nodular tenosynovitis (e-Fig. 50.12).
The localized form has an excellent prognosis and a low recurrence rate
when managed surgically, whereas the diffuse form has a reported recur-
rence rate of up to 46%. Radiation treatment has shown mixed results.
Combined surgical and nonsurgical approaches may be necessary, and in
some patients, total joint arthroplasty may be the only effective treatment
(jAm Acad Orthop Surg. 2006;14:376).
b. The extra-articular form is much less common; the knee, ankle, and foot
are predominant sites of occurrence. Some, but not all, cases are exten-
sions of the intra-articular form; the rare exclusively extra-articular cases
likely arise from the synovium of bursa or tendon sheaths. Grossly, the
tumor is a large, multinodular, and white to yellow-brown mass, with-
out villous projections. Microscopically, there are sheets of rounded to
polygonal mononuclear cells with the formation of clefts and pseudog-
landular spaces. Giant cells are fewer in number compared with the giant
cell tumor of tendor sheath, localized type. Xanthoma cells, spindle cells,
and chronic inflammatory cells are present. Scant data exist on outcome
for these patients, although recurrence rates appear high, at 40% to 50%.
3. Malignant giant cell tumor of tendon sheath is rare type of sarcoma and can
be diagnosed when a histologically benign giant cell tumor of tendon sheath
is admixed with overtly malignant areas, or when malignancy is seen in a
recurrence of the benign tumor. Histologic indicators of malignancy include
diffuse infiltrative growth, scant giant cells, cytologic atypia, necrosis, and a
mitotic count of> 10 per 10 HPFs.
4. Synovial chondromatosis is a benign nodular cartilaginous proliferation aris-
ing in the synovium of joints, bursae, or tendon sheaths. It is usually monoar-
ticular in distribution, involving the knee or hip in adults. Grossly, there
are multiple osteocartilaginous nodules, each measuring <1 mm to several
millimeters in diameter, which may be embedded within synovium and/or
mobile as loose bodies (note that loose bodies can also be seen in epiphyseal
osteonecrosis, osteochondral fracture, and osteochondritis dissecans, so their
presence is not diagnostic). Microscopically, the nodules of hyaline cartilage
show dusters of chondrocytes that can show nuclear atypia with nuclear
enlargement, hyperchromasia, and binucleation (e-Fig. 50.13), findings that
should not be viewed as evidence of malignancy. Clonal chromosomal abnor-
malities have been cited as evidence for a neoplastic process, but cytogenetics
is not used for diagnosis. Local recurrence after excision occurs in ""15% of
cases. Rare cases of chondrosarcoma arising in synovial chondromatosis have

been reported.
5. Synovial chondrosarcoma is extremely rare, and can be classified as primary
or secondary to synovial chondromatosis. Histologic features of malignancy
include loss of the "clustering" growth pattern typical of synovial chon-
dromatosis (e-Fig. 50.14), with replacement by a sheetlike arrangement of
chondrocytes, myxoid change in the matrix, areas of necrosis, and spindling
at the periphery of chondroid lobules (Cancer. 1991;67:155). The prognosis
is poor.
6. Synovial hemangioma is very rare and usually found in the knee of young
adults. Microscopically, most are cavernous hemangiomas; some are capil-
lary or arteriovenous hemangiomas (e-Fig. 50.15).
7. Synovial lipoma is rare. Grossly, the synovium is yellow, thickened, and
exhibits excrescences (lipoma arborescens). Microscopically, the synovium
is infiltrated by mature adipose tissue (e-Fig. 50.16). Recurrence is rare.
8. Rare case reports of other synovial tumors include intra-articular heman-
giopericytoma, intracapsular chondroma, synovial sarcoma, epithelioid sar-
coma, malignant fibrous histiocytoma, and lymphoma.
SECTION XII
Cyto path oIogy
General Principles of
Cytopathology
Brian Collins
I. INTRODUCTION. Cytopathology has developed into a discipline that examines cel-

lular elements from throughout the body collected by a wide variety of methods
and procedures. It shares with other anatomic pathology disciplines a morpho-
logic study of cells utilizing patterns and cellular features to identify specific patho-
logic conditions. Cytopathology provides diagnostic information in a wide variety
of clinical settings, and can thus be used to guide patient management both at
the time of initial diagnosis and during the course of treatment. In broad terms,
there are three main areas of cytopathology, specifically (i) gynecologic/Pap slides,
(ii) nongynecologic (Non-Gyn), and (iii) fine needle aspiration (FNA).
The utility of cytopathology is underscored by the fact that virtually all ancillary
laboratory techniques used in surgical pathology can also be performed on cytology
specimens.
A. Immunocytochemistry. Special stains are commonly applied to cytologic samples,
often to confirm the morphologic diagnosis, to help in determining the possible
primary sites of a tumm; or to determine the presence of specific prognostic
markers. In general, cell block sections are preferred for immunocytochemical
staining because most laboratories have experience in processing the formalin-
fixed, paraffin-embedded (FFPE) tissue sections that are generated from cell
blocks.
B. Flow cytometry. In the presence of a lymphocyte-rich sample, the possibility of a
lymphoproliferative process may need to be addressed by flow cytometry. Body
fluids, particularly effusions, are amenable to this type of analysis.
C. Molecular genetic methods. Cytological preparations, including fluid specimens,
brushings, washings, and scrapings, are suitable substrates for molecular anal-
ysis. Regardless of the method used to process samples (e.g., whether air dried,
ethanol fixed, or used to produce a cell block) it is possible to extract nucleic
acids that can be analyzed by a wide variety of genetic assays, including poly-
merase chain reaction (PCR), DNA sequence analysis, gene expression analysis,
and fluorescence in situ hybridization (FISH).
II. GYNECOLOGIC/PAP SLIDES. The "Pap test" is the most successful cancer screening test
in medicine. After the wide introduction of the Pap smear, the incidence of cervical
carcinoma dropped significantly and it remains the mainstay of screening women
for cervical carcinoma. Standardized terminology and categorizations for the Pap
smear have been developed (The Bethesda System for Reporting Cervical Cytology.
New York, Springer-Verlag, Inc.; 2004). The Bethesda Reporting System forCer-
vical Smears is covered in more detail in the cytopathology section of Chapter 34.
828
Chapter 51 • General Principles of Cytopathology I 829
Ill. NONGYNECOLOGIC CYTOPATHOLOGY encompasses a wide variety of body sites,

organs, and types of specimens. In general, any lesion that can be drained, brushed,
washed, or scraped can be a "non-gyn" cytopathology specimen.
A. Specimen types
1. Fluid. These constitute a major category of non-gyn specimens and include
pleural fluid, ascites/peritoneal fluid, pericardia! fluid, and cerebrospinal
fluid. Any loculated fluid can be drained and submitted for analysis (neck
cyst, hepatic cyst, etc.) including synovial and vitreous fluid. Urine speci-
mens, either voided or catheterized, are also commonly examined.
2. Brush. The endoscope makes it possible to introduce a brush and collect cells
and tissue from a wide variety of internal locations. These include the bron-
chopulmonary tree (bronchial brush), alimentary tract (esophageal brush,
gastric brush, and common/pancreatidhepatic bile duct brushes), peritoneal
cavity, and genitourinary tract (urethra brush, ureter brush, and renal pelvic
brush).
3. Wash. Lesions sampled by a brush are usually accompanied by a "wash"
specimen, which can be collected to sample additional cells. These specimens
commonly include lung (bronchial wash, bronchoalveolar lavage), peritoneal
cavity (pelvic wash), and urinary bladder (bladder wash, ureter wash, and
renal pelvic wash).
4. Scrape. The Tzanck smear of the skin is utilized for the identification of
mucocutaneous herpes virus effect.
B. Preparation and processing
1. Submission. Material can be submitted fresh or fixed. In general, unfixed
material should be refrigerated until it can be delivered to the cytopathology
lab. Even after arrival, it should be refrigerated until processed since unfixed
cells in a fluid environment are in a constant state of degeneration, both in
situ and after being collected. Specimens delayed in processing that have not
properly handled can be uninterpretable.
2. Fixative and stain. Alcohol is the standard fixative. Alcohol fixation can be
utilized at a variety of steps in specimen handling, including at the time of
collection (slides prepared from a brush can be placed in 95% ethanol at the
patient•s bedside, or the brush itself can be placed directly into a liquid fixative
container) or after initial processing of the specimen (after preparation of
cytospin slides from a fluid sample). Regardless, once cellular material is
placed on a slide, it needs to be alcohol fixed as soon as possible. A delay
of more than a few seconds can cause air drying artifact, which renders the
cells indistinct and the nuclei "washed out" when subsequently Pap stained.
Severe air drying artifact can render a slide uninterpretable.
a. The Pap stain highlights nuclear morphologic details including distinct
chromatin and nuclear membrane detail. The cytoplasm tends to have a
blue-green coloration with keratinization showing organophilic decora-
tion.
b. A modified Wright-Giemsa, commonly referred to as Diff-Quik™, is
the other main cytopathology stain. It is performed on slides which are
first air dried. Once completely air dried, slides are placed in a methanol
fixative and then stained (total time to complete the stain can be less than
a minute). The Diff-Quik™ stain highlights lymphoid elements and a
variety of extracellular elements (colloid, matrix, mucin, and so on).
c. The Pap stain and Diff-Quik™ stain are complementary and both are
frequently utilized in combination on non-gyn and FNA specimens.
C. Reporting. Standardized recommendations for reporting the findings in non-gyn
cytopathology specimens have recently been developed (Arch Pathol Lab Med.
2009;133:1743). Uniform reporting enhances patient care and thus implemen-
tation of these guidelines in routine practice is strongly recommended.
830 I SECTION XII: CYTOPATHOLOGY
IV. FNA. One of the most challenging and dynamic areas of cytopathology is FNA.
Virtually any organ or abnormality (either palpable or localized by imaging tech-
niques) can be subjected to FNA. The technique essentially provides a microbiopsy
of cellular material (cells and tissue) for microscopic examination and a wide vari-
ety of ancillary tests. The method permits immediate interpretive evaluation which
makes possible real-time adjustments of the biopsy procedure, appropriate speci-
men triage, and directed ancillary testing.
A. Principles. The procedure is minimally invasive and accurate. At its core, FNA
involves the use of a thin bore needle with a cutting end moved in a piston motion
to obtain cells and tissue. The cellular elements present within the needle are
then processed for diagnosis (e-Fig. 51.1).*
B. Technique. While simple, proficiency and adequacy require an understanding
of the procedure and experience in its use. It is the cutting, bevelled end of
the needle and capillary action of a repetitive "piston-like" motion movement
which provides the cellular elements necessary for diagnosis. The appropri-
ate application of needle movement is critical since insufficient movement will
not adequately sample the tissue, and prolonged movement can lead to hemodi-
lution and entrapment of tissue in clot. The appropriate number of tissue passes
and appropriate negative pressure will vary for each individual case and clinical
presentation.
Negative pressure during the procedure (typically applied by an empty 10- or
20-cc plastic syringe attached to the needle) can be useful but is not always nec-
essary. There are many solid cellular lesions/neoplasms that can be adequately
sampled without aspiration, including lymph nodes and the thyroid. Vascular
organs and neoplasms benefit from a nonaspiration FNA technique since aspi-
ration can lead to bleeding and specimen hemodilution.
Needles utilized can range from 22 to 27 gauge. Larger gauge needles tend to
cause more bleeding and paradoxically tend to be less diagnostic. Needle length
can vary significantly (5/Sth in. up to 8 in.) and will depend on what is required
to reach the lesion.
C. Procedure. In broad terms, the needle is directed to the area of interest by either
palpation or under image guidance.
1. Palpable. This method involves any lesion that can be localized or identified
by manual palpation. It must be stressed that FNA of every location requires
an understanding of the associated local anatomy, especially for areas in
the head and neck, thyroid, chest wall, and various soft tissue locations.
Superficial FNA of palpable lesions is well tolerated, minimally invasive, and
has a very low complication rate.
2. Image guidance. This method involves utilizing an imaging modality to direct
the needle, most commonly, ultrasound and CT imaging. The principles of
the FNA biopsy itself remain the same. In some cases, a stylet within the
needle is helpful; it is used to prevent sampling of organs as the needle moves
toward the lesion, and then removed once the needle tip is in the desired
location. An immediate assessment of the FNA sample makes it possible for
the pathologist to guide the clinician to ensure that a diagnostic sample is
collected and that the appropriate ancillary studies are initiated at the time
of the procedure.
D. Materials
1. Palpable. The materials required are simple and readily available. They can
be conveniently organized in a small basket or box, which can be easily
transported from the laboratory (e-Fig 51.2).
Chapter 51 • General Principles of Cytopathology I 8 31
2. Image guidance. The appropriate type of needle and the FNA approach are
determined by the clinical setting. On-site presence of the cytopathology
team with glass slides, fixative (alcohol), needle rinse tube (normal saline
or RPM!), stains (Diff-QuikTM ), and a microscope, will help optimize the
likelihood that a diagnostic sample is obtained and that appropriate ancillary
studies are initiated.
E. Clinical application
1. Immediate evaluation. Immediate evaluation is the standard of care because it
provides several advantages. First, it guides the procedure and thus ensures
a maximum effort to obtain a diagnosis. By examining the slides during the
procedure, immediate evaluation can be used to help decide if more FNAs
are necessary; guide the needle position placement by determining if lesion
or nonlesional material is present; and if nondiagnostic, support a decision
to move to a potential second site. Second, with immediate evaluation, the
trade-off between the length of the procedure and number of biopsies can be
managed to balance the requirement for diagnostic tissue without unnecessar-
ily extending the length of the procedure. By assuring a diagnostic procedure,
the number of nondiagnostic samples can be minimized and repeat proce-
dures and delays in diagnosis can be avoided. Third, the cytopathologist can
identify those cases where tissue for appropriate ancillary studies needs to be
collected to enhance diagnostic accuracy. These situations include identifying
a lymphoproliferative process where rinse material is sent for flow cytometry
analysis, abscess/granulomatous inflammation where microbiologic cultures
are indicated, poorly differentiated neoplasms where a cell block can provide
material for immunohistochemical evaluation, and lesions where cytogenet-
ics/molecular diagnostics can contribute to diagnosis and treatment.
F. Specimen processing
1. Direct smears are stained by the Papanicolaou and Diff-Quik™ methods.
2. Liquid medium. FNA samples that are collected in a liquid medium provide
a wide variety of options for further analysis. While it is helpful to perform
needle washes during the FNA procedure, the best chance for a sufficient
cell block involves direct (no slides prepared) dedicated FNA samples placed
in the liquid medium (saline or RPM!). Cell block preparation affords a
variety of options and advantages; sections of the cell block contribute to
the morphologic evaluation of the lesion, and also provide cellular material
for immunohistochemical analysis, interphase FISH, and for a wide range of
PCR-based molecular tests.
SUGGESTED READINGS
Crothers BA, Tench WD, Schwartz MR, et al. Guidelines for the reporting of nongynecologic
cytopathology specimens. Arch Pathol Lab Med. 1999;133:1743-1756.
Gupta PK. University of Pennsylvania aspiration cart (Penn-A-Cart): an innovative journey in fine
needle aspiration service. Acta Cytol. 2010a;54:165-168.
Gupta PK. Progression from on-site to point-of-care fine needle aspiration service: opportunities
and challenges. Cyto;ournal. 2010b;7:6.
Wright TC, Massad LS, Dunton C], et al. 2006 consensus guidelines for the management of women
with abnormal cervical cancer screening tests. Am] Obstet Gynecol. 2007;197:346-355.
SECTION XIII
Ancillary Methods
Frozen Sections and

Other Intraoperative
Consultations
Michael E. Hull, Peter A. Humphrey,
and John D. Pfeifer
I. INTRODUCTION. Intraoperative consultations fall into two general categories. Micro-

scopic consultations, usually performed as frozen sections, are undertaken to estab-
lish a tissue diagnosis, determine the nature of a lesion that may require ancil-
lary testing, establish that sufficient diagnostic tissue has been obtained, identify
metastatic disease, and assess surgical margins or extent of disease. Microscopic
consultation can also be performed using touch preparations, a practice that has the
advantage of preserving valuable tissue. Nonmicroscopic consultations are gross
examinations of a specimen that provide the surgeon with real-time information
on tissue margins and the anatomic extent of disease processes. They also facilitate
the triage of fresh tissue for ancillary diagnostic studies or research protocols.
II. FROZEN SECTIONS. High-quality frozen sections can be performed with remarkable
speed if equipment is kept in optimum working condition and if the operator is well
versed in the technique. In experienced hands, the entire consultation can often be
performed in 10 to 15 minutes from the time of the arrival of the specimen in the
frozen section room to the notification of the surgeon of the diagnosis. For larger tis-
sue specimens, proper interpretation requires a thorough gross examination of the
tissue before sectioning. Good communication with the surgeon regarding opera-
tive findings and knowledge of pertinent clinical history are also absolutely essential
for optimization of the process.
A. Indications. Frozen sections are indicated to establish a tissue diagnosis (such as
the presence of malignancy, which will guide intraoperative patient management
and extent of surgery); for tissue identification (e.g., to confirm the presence of
parathyroid tissue in a parathyroidectomy specimen); to determine the nature of
a lesion that may require ancillary testing that requires special fixatives or media
(e.g., RPMI for flow cytometry or glutaraldehyde for electron microscopy); to
establish that sufficient diagnostic tissue has been obtained; to identify metastatic
disease; and to assess surgical margins or extent of disease.
Frozen sections should not be used merely to satisfy a surgeon's curiosity,
to compensate for inadequate preoperative evaluation, or as a mechanism to
communicate information more quickly to the patient or the patient's family.
B. The frozen section procedure. Frozen sections are performed by freezing the tissue
in a block of specialized embedding medium, followed by cutting thin (usually
5 11-m) sections from the block using a cryostat (refrigerated microtome). The sec-
tions are adhered to glass slides, fixed in ethanol, and stained with hematoxylin
832
Chapter 52 • Frozen Sections and Other Intraoperative Consultations I 8 33
and eosin (H&E). Small specimens may be completely utilized for frozen section
slide preparation, but if possible, a portion of the tissue should be preserved for
routine handling to avoid freezing artifacts that can compromise interpretation
of the permanent sections (e.g., in the case of brain biopsies). For larger tissue
samples, judgment must be exercised in gross sampling so that the area(s) of
highest diagnostic yield is selected for frozen section. Cytological imprints from
tissues can be an important adjunct in diagnosis, especially for hematolymphoid
abnormalities, lymph node biopsies, and thyroid lesions.
C. Interpretation. The interpretation of frozen sections requires integration of the
histologic morphology in the H&E-stained sections; the gross features of the
specimen; information from the surgeon regarding the origin of the tissue,
the indication for the consultation (including the clinical history, radiological
findings, and intraoperative observations); and the ways in which the frozen sec-
tion diagnosis will affect the operative strategy. In some difficult cases, it may be
necessary to request additional tissue for frozen section analysis. If additional
tissue cannot be obtained, deferral of a definitive diagnosis pending examination
of formalin-fixed paraffin-embedded sections is acceptable. In routine clinical
practice, such deferrals are employed in <5% of frozen section diagnoses.
D. Communication of findings. Clear communication of a concise diagnosis to the
surgeon is the last step of the intraoperative consultation. Usually, there is a
narrowly defined clinical problem that frozen section is to solve; this should
be specifically and unambiguously addressed in the diagnosis. The diagnosis is
written and signed, with the date and time, by all interpreting pathologists and
made part of the final pathology report. If the written diagnosis is verbally trans-
mitted, the pathologist should first confirm with the recipient of the information
the identity of the patient (using two identifiers) and the surgeon. The operating
room staff member taking the diagnosis should repeat it back to the pathologist
to confirm accurate communication.
Certain phraseologies tend to be resistant to misinterpretation. For instance,
"negative for malignancy" or "positive for malignancy" are generally under-
stood well. Since some diagnosis can easily be incompletely heard or misunder-
stood in a busy operating room, the diagnosis should always be repeated back
as a safeguard.
E. Accuracy of frozen sections. The accuracy of frozen sections will vary from insti-
tution to institution on the basis of the types of surgical cases evaluated and the
experience of the involved pathologists. Table 52.1 highlights the fact that the
accuracy of frozen section diagnosis is dependent on the anatomic site. Regular
self-audits of the frozen section service are desirable so that surgeons and pathol-
ogists are aware of the performance characteristics of the modality in their own
hands. Such audits of single institutions, and pooled data across hundreds of
institutions, show that accurate diagnoses are made overall in >95% of cases,
while discordance with the final diagnosis occurs in 1% to 2% of cases. Deferral
of the diagnosis until permanent section diagnosis occurs in 1% to 4% of cases.
F. Sources of error in frozen sections. Errors can be divided into errors of interpre-
tation and errors of sampling; both usually result in false-negative diagnoses.
False-positive diagnoses are rarer, likely because experienced pathologists tend
to appropriately defer to permanent section rather than making an incorrect
diagnosis on substandard material.
Misinterpretation accounts for about 40% of errors overall (Arch Pathol
Lab Med. 1996;120:804; Arch Pathol Lab Med. 1996;120:19). Interpretations
of frozen sections are more prone to this error than interpretation of permanent
sections due to the presence of artifacts that are not encountered in routinely
fixed, paraffin-embedded sections. Some tissues are more likely to show signifi-
cant artifact, especially those with high fat content, such as most pelvic lymph
nodes.
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Sampling errors occur in two ways. The first is sectioning error. Diagnos-
tic tissue may be present in the frozen block, but the block may not be faced
sufficiently for the lesion to be present on the actual frozen section slides;
the diagnostic material may then be found in routine permanent sections of the
residual frozen block, a scenario that accounts for 10% to 15% of errors. The
second type of sampling error is gross sampling error. This accounts for about
45% of errors overall, and is seen when the diagnostic tissue is not in the portion
of the specimen sampled by the frozen section; good gross pathology skills will
minimize, but never eliminate, this problem. Furthermore, it is not feasible to
completely sample larger lesions by frozen section, and so sampling errors will
always exist for large lesions with heterogeneous composition.
Discrepancies between a frozen section and the final (permanent section)
diagnosis should be documented in the final surgical pathology report, along
with the reason for the discrepancy. If the discrepancy is of clinical significance,
the pathologist should immediately alert the surgeon to the change in diagnosis.
G. Anatomic sites deserving special mention. The anatomic sites with the most dis-
crepancies between frozen and permanent sections are skin, breast, lymph nodes
for metastatic disease, the female genital tract, and thyroid. Frozen sections do
have a role in the surgical management of disease in these sites, but since loss of
diagnostic material during the performance of frozen sections is unavoidable,
each case must be critically evaluated as to whether the frozen section diagnosis
will add enough value to warrant this loss.
1. Skin. Frozen sections for margin assessment in the excisions of large non-
melanoma skin cancers such as basal cell carcinoma and squamous cell car-
cinoma are indicated if the lesion has vague infiltrative borders or is in a
location in which wide excision is not possible, as in the case of tumors
of the eyelid, nose, or ear (Arch Pathol Lab Med. 2005;129:1536). If the
borders of a lesion are well defined, frozen sections are not as necessary.
It is widely agreed that the intraoperative primary diagnosis of pigmented
lesions is ill-advised. The determination of prognosis and subsequent man-
agement of melanoma requires an accurate assessment of Breslow thickness,
and frozen artifact may cause so much specimen distortion as to make a
depth of invasion assessment meaningless in both permanent and frozen sec-
tions. Freezing artifact compounded by actinic damage, frequent in patients
with pigmented lesions, also conspires to obscure histology. Subtle changes,
such as intraepidermal spread by single melanocytes, are very difficult to
appreciate in frozen sections, making the method a poor choice for margin
assessment as well. For example, the false-negative rate for lentiginous spread
of melanoma by frozen section evaluation may be as high as 50%.
2. Breast. Once commonplace, the initial pathologic diagnosis of breast tumors
is now very rarely made during surgery, since methods for detection of smaller
tumors and more sophisticated treatment algorithms (as opposed to mastec-
tomy only) now usually lead to diagnosis on the basis of needle core biopsy or
aspiration cytology. Margin evaluation is sometimes requested during breast-
conserving procedures, but the high fat content of breast tissue makes frozen
specimens technically difficult to section and prone to freezing artifact. Eval-
uation of margins by imprint histology is an alternative to frozen section,
but it is insensitive unless the margins are grossly involved, in which case the
examination is unnecessary (Arch Pathol Lab Med. 2005;129:1565). As a
rule, an intraoperative gross examination of a lumpectomy specimen is used
to determine whether or not there will be additional excisions.
3. Lymph nodes for metastatic disease (including axillary sentinel lymph nodes for
breast carcinoma)
a. Sentinel lymph nodes. Although current diagnostic modalities have reduced
the need for breast frozen sections, requests for frozen sections of breast
836 I SECTION XIII: ANCILLARY METHODS
sentinel lymph nodes have increased (e-Fig. 52.1A and B).* Unfortunately,
frozen sections on sentinel nodes consume considerable tissue; if no metas-
tases are identified, further evaluation of the node has therefore been sig-
nificantly hampered (e-Fig. 52.2).
Numerous studies have indicated that the sensitivity of frozen section
evaluation of sentinel nodes is about 60%, although the specificity is near
100% (Mod Path. 2005; 18:58). It has been argued that the metastases
that are typically missed by frozen section are submicroscopic (<0.2 mm in
greatest dimension) or are detected by cytokeratin immunohistochemistry
alone, and that since the significance of these classes of metastases is not yet
cleat; the low sensitivity of the approach underestimates its clinical utility.
A sensible middle ground seems to be to perform frozen sections when the
clinical history and gross examination of the node give a high degree of
suspicion for metastasis. Imprint cytology, as a routine measure, is another
possible approach to intraoperative sentinel lymph node evaluation.
b. Other lymph node frozen sections. The intraoperative evaluation of lymph
nodes can be performed by frozen section or by imprint cytology, and if
indicated, tissue should be allocated for flow cytometric analysis, routine
histology, and molecular diagnostics.
4. Thyroid. The widespread use of fine needle aspiration has altered the approach
to frozen section evaluation of the thyroid. Frozen section evaluation of
thyroid excision specimens is best performed in conjunction with imprint
cytology, the latter of which is more sensitive for the nuclear grooves, inclu-
sions, and chromatin-clearing characteristic of papillary carcinoma. Since
the distinction between follicular adenoma and carcinoma by frozen section
(e-Fig. 52.3) would require thorough sampling of the tumor and capsule for
invasion and angioinvasion, frozen section is poorly suited to this application
(Can I Surg. 2004;47:29).
5. Female genital tract. Of all the neoplastic processes of the female genital tract,
ovarian neoplasms are probably the best suited to intraoperative frozen sec-
tion diagnosis (Arch Pathol Lab Med. 2005;129:1544; lnt I Gynecol Cancer.
2005;15:192). Even so, borderline mucinous neoplasms of the ovary have a
significant rate of discordance between the frozen section and permanent
section diagnoses, primarily due to sampling error. The diagnosis of cervical
dysplasia is fraught with difficulties due to frozen section artifact and low
concordance with permanent section diagnosis.
Ill. OTHER INTRAOPERATIVE CONSULTATIONS. Intraoperative nonmicroscopic consulta-
tions are often required even though no frozen section is needed. Indications include
gross diagnosis (such as benign simple ovarian cyst or leiomyoma of uterus); gross
confirmation of the presence of a lesion or mass; identification of a margin or
region of interest that requires special sampling for permanent sections; specimen
orientation; triage of tissue for ancillary testing modalities that require special pro-
cessing or fixatives (in which case good judgment is required to ensure a balance
between the need for tissue for routine histopathologic evaluation and the need for
tissue for specialized testing); and collection of tissue for tumor banking or research
studies.
A. Opening of a viscus organ for gross examination and fixation. Gastrointestinal
specimens commonly require opening in the operating room so that the surgeon
can see whether or not lesional tissue has been excised. In partial intestinal
excisions for inflammatory disease, in which the risk for malignancy is increased,
a careful intraoperative examination may inform the surgeon of a previously
undetected tumor, which may have implications for additional surgical therapy.
Similarly, gross examination and opening of the adnexa and/or uterus can
aid in determining the extent of surgery that is required. Such gross examination
often leads to frozen sections when ovarian surface papillary excrescences are
identified, or when solid or complex architecture features are discovered in an
ovarian cyst.
B. Tumor for banking must be chosen so that the remaining lesional tissue material
will still be suitable for a complete diagnostic evaluation. The banked tissue
should not include a surgical margin, or have an important relationship to an
anatomic structure that would impact staging or the need for adjuvant therapy.
In cases of small specimens, it may be necessary to defer banking for the sake
of a thorough diagnostic evaluation.
SUGGESTED READINGS
Lechago J. Frozen section examination of liver, gallbladder, and pancreas. Arch Pathol Lab Med.
2005;129:1610.
Marchevsky, AM, Changsri C, Gupta I, et al. Frozen section diagnoses of small pulmonary nodules:
accuracy and clinical implications. Ann Thorac Surg. 2004;78:1755.
Young MP, Kirby RS, O'Donoghue EP, et al. Accuracy and cost of intraoperative lymph node
frozen sections at radical prostatectomy.] Clin Pathol. 1999;52:925.
Electron Microscopy
Ashima Agarwal and Frances V. White
I. INTRODUCTION. Transmission electron microscopy (EM) has been used by sur-

gical pathologists over the past 50 years for the diagnosis of a wide range of
diseases in various organ systems (Table 53.1). The method allows for the visu-
alization of subcellular morphology, with appreciation of disease processes and
structural abnormalities that cannot be resolved by light microscopy. EM is an
essential part of the work-up of medical renal diseases, peripheral nerve diseases,
muscle diseases, and primary ciliary dyskinesia. It is also useful in evaluating
metabolic and inherited diseases, providing an initial differential diagnosis, or
ruling in or out a specific disease process. The role of EM in the diagnosis of neo-
plasms has decreased since the advent of immunohistochemical and molecular
techniques, but it is still an ancillary tool for the evaluation of atypical tumors or
when other techniques yield indeterminate results. Although not usually needed,
EM is occasionally used to demonstrate infectious agents or evidence of drug
toxicity.
In recent years, EM has been combined with immunohistochemical and in
situ hybridization methods, allowing antigen detection and localization at the
subcellular level. These combined methods require special fixation and processing
protocols. Although immunoelectron microscopy is currently used primarily in
research laboratories, the method is now considered to be a promising diagnostic
technique in oncologic surgical pathology, in particular, for the identification and
localization of targets for gene therapy.
II. METHODOLOGY. Tissue for EM must be immediately fixed. A thin slice of tissue
should be immersed in a cold fixative such as buffered glutaraldehyde 2% to 4%,
or buffered glutaraldehyde plus paraformaldehyde, and then diced into 1-mm
cubes using a sharp, clean scalpel blade. Specimens are usually postfixed in
osmium tetroxide, dehydrated in ethanol, and then embedded in an epoxy resin
or plastic. Semithin (1-~m-thick) sections are cut from the blocks and stained
with toluidine blue or methylene blue, and light microscopic examination of the
semithin sections is used to select the blocks from which thin sections are cut and
placed on grids. The thin sections are usually stained with uranyl acetate and lead
citrate; other stains, howeve.t; may be selected to enhance electron contrast of spe-
cific particles or structures depending on the tissue type and diagnostic question.
Tissue processing typically takes a couple of days, but with microwave techniques,
grids can be ready within 5 hours postfixation.
For some disease processes, if glutaraldehyde-fixed tissue is not available, EM
can be performed on formalin-fixed wet tissue or paraffin-embedded tissue. Wet
tissue is preferable to paraffin-embedded tissue, but previous prompt fixation
in formalin is essential. Autopsy material, whether fixed in glutaraldehyde or
formalin, is often unsatisfactory due to the prolonged postmortem interval prior
to fixation.
A focused differential diagnosis based on integration of the clinical history
and light microscopic findings is essential for the correct interpretation of ultra-
structural findings. Except for the most routine specimens, the pathologist should
personally review the semithin sections by light microscopy and select the blocks
for further processing. In addition, the pathologist should communicate his differ-
ential diagnosis to the electron microscopist and specify the cell type and subcel-
lular structures of interest. Obviously, sampling error is minimized and the most
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is presented in detail elsewhere in this manual. Because EM makes it possible

to visualize the individual components of the glomerular capillary wall, includ-
ing endothelium, glomerular basement membrane, and visceral epithelial cells,
it is used for identification and localization of discrete electron-dense deposits
in glomeruli, either of immunoglobulins or amyloid and amyloid-like proteins.
The glomerular basement membrane can also be evaluated for abnormal thick-
ening, thinning, and/or splitting and for the presence of electron lucent, granu-
lar, or other deposits. Tubular basement membranes, arterioles, and the intersti-
tium can also be evaluated by EM for pathogenic changes, for example, non-
immune deposits such as amyloid, light chain dense deposits, and cryoglobu-
lins. Chapter 19 describes in detail the ultrastructural findings in specific renal
diseases.
IV. NEOPLASMS. The role of EM in the diagnosis of neoplasms has decreased since the
advent of immunohistochemical and molecular markers. EM, however, is still a
useful ancillary method in selected cases and for poorly differentiated tumors for
which immunohistochemical and molecular studies are inconclusive. In general,
a differential diagnosis is first developed on the basis of light microscopic find-
ings, and EM is then used to look for evidence of cellular differentiation toward
the tumors in the differential diagnosis (e-Fig. 53.1).* For example, desmosomes,
tonofilaments, melanosomes, and premelanosomes may be sought in the differen-
tial diagnosis of carcinoma versus melanoma. EM does not differentiate reactive,
benign, neoplastic, and malignant processes.
V. INHERITED METABOLIC DISEASES. Initial studies in the work-up of inherited
metabolic diseases often include a biopsy of affected organs (such as liver, muscle,
or peripheral nerve). For select diseases, EM findings may be pathognomonic. In
most cases, however, light microscopy and EM findings are useful in narrowing
the differential diagnosis and providing direction for further laboratory studies.
Definitive diagnosis typically requires enzyme studies of fibroblast cultures and/or
molecular studies.
A. Prenatal studies. EM can be performed on amniotic cells and chorionic villous
tissue obtained for prenatal diagnosis. Ultrastructural studies on noncultured
amniotic cells can yield a rapid diagnosis or differential for certain metabolic
diseases, including type 2 glycogen storage disease, lysosomal storage diseases,
and peroxisomal disorders.
B. Liver. Inherited metabolic diseases often result in hepatocellular dysfunction,
either as part of a systemic disorder or as part of disease limited to the liver.
In the work-up of hepatitis and cholestatic liver disease in infancy and child-
hood, a portion of the liver biopsy is routinely placed in glutaraldehyde for
possible ultrastructural studies. In cases where obstruction and infection have
been ruled out, EM is performed to look for evidence of primary metabolic dis-
ease. Certain metabolic diseases have pathognomonic or near-pathognomonic
findings, such as type 2 and 4 glycogen storage diseases, alpha-1-antitrypsin
disease, and Wilson disease (e-Fig. 53.2). Abnormalities in the number and
structure of mitochondria and peroxisomes are characteristic of other spe-
cific diseases, as are lysosomal inclusions. Ultrastructural findings are always
interpreted in conjunction with light microscopic findings, clinical history, bio-
chemical assays, and other laboratory studies.
C. Lung. The protocol for lung biopsy in the work-up of interstitial lung disease in
infancy includes placing a portion of the specimen in EM fixative so that tissue
is available for ultrastructural analysis if needed, based on the light microscopic
findings. Diseases that have characteristic ultrastructural findings (e-Fig. 53.3)
Chapter 53 • EIectron M icroscopy I 84 1
include pulmonary interstitial glycogenosis and surfactant processing abnor-

malities (surfactant protein B [SPB] deficiency and adenosine 5'-triphosphate
[ATP]-binding cassette transporter A3 [ABCA3] mutations).
D. Heart. Glycogen storage disease type 2 (Pompe disease) can be diagnosed on
the basis of ultrastructural findings in cardiac biopsies. Adriamycin toxicity
involving the heart results in characteristic cytoplasmic changes that can be
seen by light microscopy in semithin sections prepared for EM.
VI. NEUROPATHOLOGY SPECIMENS. EM is critical for the evaluation of peripheral nerve
biopsies and muscle biopsies (Can] Vet Res. 1990;54:1) and is also useful in the
evaluation of selected neuro-oncologic specimens (] Neuropathol Exp Neurol.
2002;61:1027). EM examination of skin and conjunctival biopsies is used in the
diagnosis of neuronal ceroid lipofuscinosis, infantile neuroaxonal dystrophy, and
cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoen-
cephalopathy (CADASIL) (e-Fig. 53.4).
VII. PRIMARY CILIARY DYSKINESIA. At present, EM is the method of choice for the diag-
nosis of primary ciliary dyskinesia (Acta Otorhinolaryngol Belg. 2000;54:309 and
Nat Rev Mol Cell Biol. 2007;8:880) using ciliary biopsies and brushings obtained
from the nasal mucosa and lower airway. The most frequent ultrastructural abnor-
malities are decreased numbers or absence of outer and/or inner dynein arms, some
of which are associated with specific genetic abnormalities that have an autosomal
recessive inheritance pattern (Respiration. 2007;74:252) (e-Fig. 53.5). Ultrastruc-
tural abnormalities in other ciliary components have also been reported, but not
in a consistent manner. Difficulties in interpretation result from significant overlap
of EM findings in chronic inflammatory processes and an inherent difficulty in
the visualization of inner dynein arms.
VIII. MICROVILLUS INCLUSION DISEASE. Microvillus inclusion disease presents as
intractable secretory diarrhea in the neonate. Characteristic light microscopic
findings include villous atrophy and loss of the brush border, but EM is required
for diagnosis. Pathognomonic ultrastructural findings include absent or decreased
numbers of stubby microvilli on the apical cytoplasmic membrane of enterocytes,
along with cytoplasmic membrane-bound inclusions with microvillus projections.
Morphologic variants have been reported.
SUGGESTED READINGS
D'Agati VD, Jennette JC, Silva FG. Non-neoplastic Kidney Disease. Atlas of Nontumor Pathology
(First series, Fascicle 4). Washington, DC: American Registry of Pathology Press; 2005.
Iancu TC. The ultrastructural spectrum of lysosomal storage diseases. Ultrastruct Pathol.
1992;16:231-244.
Sherman PM, Mitchell DJ, Cutz E. Neonatal enteropathies: defining the causes of protracted diar-
rhea of infancy.] Pediatr Gastroenterol Nutr. 2004;38:16-26.
Histology and
Histochemical Stains
Kevin Selle
I. RECEIPT, ACCESSIONING, AND GROSS DISSECTION. Most, if not all, biopsies and
large tissue specimens are routed to the pathology laboratory. Under normal cir-
cumstances, the specimens are received in 10% neutral buffered formalin (formalin
begins the fixation process and prevents autolysis and decomposition). The speci-
men is first logged into the surgical pathology computer system and given a unique
identifying number, referred to as an accession number or case number. Once acces-
sioned, the specimen is taken to the gross dissection room; depending on the prac-
tice setting, residents, fellows, pathologist assistants, and/or trained technicians are
responsible for gross processing of the specimen under the supervision of an attend-
ing pathologist. Gross processing entails describing the specimen by its size, shape,
color, and overall general appearance, followed by placing samples of the tissue
in processing cassettes (for biopsies, the entire tissue specimen is placed in a cas-
sette; for larger specimens, regions of tissue are sampled according to established
protocols). Each cassette is labeled with the accession number as well as a part
designator and number; this numbering scheme is designed to allow the location
of a particular section of tissue within the context of the whole specimen.
II. PROCESSING. The loaded cassettes are stored in 10% neutral buffered formalin
until automated tissue processing. The normal processing cycle is ""'8 hours long
and in general is designed to remove the water from the specimen and replace it
with paraffin. Automated, closed-system tissue processors utilize agitation, vac-
uum, and increased temperature to optimize the process. In general terms, the
process is as follows. First, the tissue is subjected to 10% neutral buffered formalin
to ensure complete fixation. Complete fixation aids in the dehydration steps and
prevents tissue shrinkage and other artifacts caused by excessive or rapid dehydra-
tion; chemically, formalin fixation produces methylene cross-links between nucleic
acids and/or proteins. Once the tissue is well fixed, it is subjected to several changes
of graduated alcohols in a gradient starting at 70% and ending at 100%, a process
that removes water from the tissue at a slow controlled rate designed to prevent
excessive shrinkage and disruption of the architecture and cellular components.
After complete dehydration of the tissue has been accomplished, a clearing agent
is used to remove the alcohol and allow tissue infiltration by paraffin; this clearing
agent must therefore be miscible in both alcohol and paraffin. Xylene is most often
used for this purpose, although commercial xylene substitutes are available. In the
next step of processing, heated paraffin infiltrates into the tissue. Paraffin is a solid
at room temperature but has a relatively low melting point, and so is a good choice
as an infiltration and embedding media. While pure paraffin wax was used in the
past, current commercially available paraffins are formulated with various plastic
polymers to allow better infiltration and a more rigid crystalline structure, both of
which aid subsequent microtomy.
Ill. EMBEDDING. Properly fixed and processed tissue sections are embedded in molds to
prepare them for microtomy. The tissue is removed from the cassette and oriented
in the base of a mold that is of a size to allow paraffin to surround the tissue
section. During embedding, the tissue is oriented with the understanding that the
surface placed down in the mold will become the face of the tissue block, and
will thus be the surface cut into first by the microtome blade. Attention must be
842
Chapter 54 • Histology and Histochemical Stains I 843
given to tissues requiring specific orientation such as tubular structures requiring

complete cross sections (e.g., ureteral margins). Orientation is critical to proper
tissue representation on the finished slide and leads to proper pathologic diagnosis,
and so the importance of proper embedding cannot be overstated. After proper
orientation, the mold and tissue are touched to a cold plate to begin to solidify
the paraffin so that the tissue is held in place as the mold is filled with paraffin.
The empty cassette is placed on top of the mold so that it becomes the back of the
tissue block, conveniently retaining the identification of that tissue sample. Finally,
the mold is allowed to cool so that the paraffin block containing the oriented tissue
can be easily removed.
IV. MICROTOMY. Proper microtomy requires a well-trained and highly skilled micro-
tomist, usually a trained histotechnician or histotechnologist. The microtome
instrument is designed to hold the paraffin tissue block firmly in place as it is
cyclically presented to a stationary microtome blade. Each turn of the microtome
handle (each cycle) advances the tissue block a set distance, so microtomes must
be kept clean and in good working order. Most tissues are sectioned at 4 to 5 Jtm
thick; however, some tissues are cut thinner at 3 Jtm (e.g., kidney biopsies and
lymph node biopsies), and others thicker at 5 to 6 Jtm (e.g., bone and brain).
In practice, the paraffin block is first "faced in" to reach a level within the tissue
where there is a representative tissue section plane; the block is then cooled on
wet ice to further harden the paraffin and aid microtomy. If proper care is taken,
individual sections come off of the microtome blade connected to each other, a
string of tissue sections called a "ribbon." The tissue ribbon is then floated on
a warm water bath at a temperature 6oC to soc below the melting point of the
paraffin to make the paraffin very pliable. This aids in mounting the sections on a
glass microscope slide labeled with the corresponding accession number and part
identifier for that particular block (in addition, the convection currents formed in
the water bath help gently stretch the tissue sections, removing any wrinkles).
V. HEMATOXYLIN AND EOSIN (H&E} STAINING. Slides that have been sectioned are stained
to reveal their histologic detail. The primary stain used for pathologic diagnosis is
the H&E stain. Hematoxylin is derived from the log wood tree, and has long been
in use in the pathology laboratory. By itself it is not a dye, but once it is oxidized
to hematein and combined with a metallic mordant, it acquires a strong affinity
for nuclear chromatin. Eosin is a dye that at a pH of approximately 4.6 to 5.0 is a
strong anion and thus has an affinity for positively charged, cationic, tissue protein
groups. At the proper pH, eosin combines at different rates to tissue proteins and
thus produces a graduation of distinct shades from light pink to pinkish red.
There are several methods for performing the H&E staining that can be used
to achieve slight variations in the end stain to match the preference of the pathol-
ogist. The two general variations of H&E staining in common use are progressive
and regressive methods. Progressive methods involve staining slides for a desig-
nated period of time, and then stopping the reaction as soon as optimal staining
has occurred; every tissue stains slightly differently with hematoxylin on the basis
of its type, fixation, and prior decalcification, and therefore the length of time
in hematoxylin is critical using the progressive method. Regressive methods over-
stain the tissue sections with hematoxylin, and then differentiate the hematoxylin
using acid alcohol; by overstaining and then differentiating the hematoxylin by the
regressive method, a darker, crisper stain can be achieved with the assurance that
all hematoxylin-positive elements are represented.
The routine regressive staining protocol for the H&E stain is briefly as follows.
The tissue sections are dried completely in an oven since water left on or under
the tissue sections can allow the sections to fall off of the slide during the staining
process. The slides are then deparaffinized by soaking in xylene, the xylene is
removed by alcohol, and the slides are rehydrated by 95% alcohol and then water
before staining with hematoxylin. Excess hematoxylin is then removed with a water
rinse, the slides are differentiated using acid alcohol, rinsed, and the hematoxylin
is "blued" by immersion in a weak ammonia water solution. The slides are rinsed
again, placed in 80% alcohol, and stained with eosin. Excess eosin is removed by
alcohol rinses, and the slide is prepared for mounting with a coverslip and resinous
media by removal of the alcohol using xylene rinses.
VI. OTHER FREQUENTLY USED HISTOCHEMICAL STAINS. Tissue stains range from very
simple to complex in methodology, and can be used to demonstrate most major
tissue elements relevant to pathologic diagnosis. They are based on the chem-
istry of various dyes and metals, and most were developed prior to the advent of
immunohistochemistry. In general, a histochemical stain consists of the main chem-
ical reaction that demonstrates the specific tissue element of interest, followed by
chemical reactions that provide staining of the background uninvolved tissue ele-
ments, often including nuclear detail. Histochemical stains are usually grouped by
the tissue element they stain.
A. Carbohydrates. In humans, carbohydrates exist as various sugars and polymers
linked to proteins. Simple sugars cannot be detected by standard histochemical
procedures because they are water soluble and thus removed during process-
ing; however, polymers such as glycogen can be detected. Naturally occurring
polysaccharides can be classified into four groups on the basis of their histochem-
ical staining differences: neutral polysaccharides (Group I), acid mucopolysac-
charides (Group II), glycoproteins (Group III), and glycolipids (Group IV).
Amyloid must also be included because, even though it is not a carbohydrate,
its histochemical staining properties are similar to those of polysaccharides. The
histochemical stains most often used to detect carbohydrates and differentiate
various types of carbohydrates are Aldan blue, colloidal iron, mucicarmine, the
periodic acid-Schiff (PAS) reaction, Congo red, and Thioflavin T.
1. Mucicarmine. The mucicarmine method is used to detect tissue mucins and
utilizes the tissue dye carmine. When carmine is reacted with aluminum, it
forms a compound that has a net positive charge and is attracted to the
negative acid groups of epithelial mucins. Metanil Yellow and Weigert's
hematoxylin are used as counterstains and produce yellow staining of the
background tissue elements and blue-black nuclear staining (e-Fig. 54.1).*
2. Alcian blue, a phthalocyanine basic dye, forms salt bridges with the acid
groups in mucopolysaccharides. Staining tissue sections in an Aldan blue
solution at pH 1.0 produces staining of only sulfated mucopolysaccha-
rides, while staining at pH 2.5 produces staining of all mucopolysaccharides.
These two methods make it possible to differentiate sulfated from carboxy-
lated mucopolysaccharides; further differentiation between mucosubstances
of connective tissue origin and that of epithelial origin can be achieved by
the addition of hyaluronidase digestion (e-Fig. 54.2).
3. Colloidal iron. The colloidal iron stain is based on the chemical principle that
at low pH colloidal ferric ions can be absorbed by both carboxylated and
sulfated mucopolysaccharides, as well as other glycoproteins. The absorbed
ferric ions are detected by use of the Prussian blue reaction (see below).
4. Congo red. Congo red reacts with cellulose and amyloid. The dye is a lin-
ear molecule that attaches to amyloid in a sheet-like fashion resulting in
so-called apple green birefringence when subjected to polarized light. This
"apple green" birefringence is considered specific for amyloid in Congo red-
stained tissue sections (e-Fig. 54.3).
5. Thioftavin T. Thioflavin Tis a fluorescent tissue dye that has an affinity for
amyloid. Thioflavin T fluoresces yellow to yellow-green when the tissue
section is viewed by fluorescent microscopy, but the dye is not as specific

for amyloid as Congo red.
6. PAS. The PAS reaction is invaluable in histochemistry because of its versatility.
In this reaction, the glycol groups of polysaccharides, mucosubstances, and
basement membranes are subjected to oxidation by a solution of periodic
acid. The oxidation of the glycols results in the formation of dialdehydes.
The dialdehydes are then reacted with Schiff's reagent, a colorless solution
created by reducing basic fuchsin in the presence of sulfurous acid. When
reacted with the previously oxidized tissue, Schiff's reagent is bound to the
dialdehyde groups and gains a red color (e-Fig. 54.4).
The differentiation of glycogen from other mucosubstances can be
achieved using the PAS stain as follows: Two identical tissue sections are
cut. The first section is treated (or digested) with either amylase or diastase
to remove any glycogen in the tissue, and then both sections are stained fol-
lowing the PAS procedure. If the substance in question is glycogen, it will be
present in the undigested section but not in the digested one (e-Fig. 54.5).
B. Connective tissues. Connective tissue is made up of three elements in varying
amounts, namely cells, a variety of protein fibers, and so-called ground sub-
stance. The commonly used connective tissue stains are used to demonstrate cells
and various protein fibers, and include the reticulin stain, trichrome stain, Jones'
methenamine stain, phosphotungstic acid hematoxylin (PTAH) stain, Verhoef£
Van Gieson (VVG) stain, and Oil red 0 stain. Each of these stains has many
different modifications on the basis of the preferences of the lab and pathologists
involved. Since ground substance is principally composed of mucopolysaccha-
rides, it can be demonstrated by the carbohydrate stains mentioned previously.
1. Reticulin. The reticulin stain is similar to the PAS stain in that glycols are first
reduced to dialdehydes by the use of an acid. The tissue is next sensitized to
accept metallic silver ions with an ammoniacal silver solution, and the silver
ions that are attached on and around the dialdehyde groups are then reduced
to metallic silver. Finally, the tissue sections are toned from brown to black
by replacing the metallic silver with metallic gold; sodium thiosulfate is used
to remove any remaining unreacted silver in the tissue to prevent darkening
of the slide over time. The end result is that reticulin fibers are stained black
against a clear background (e-Fig. 54.6).
2. Jones' methenamine silver (JMS). The JMS stain uses methenamine to form a
complex with silver, which is then reacted with dialdehyde groups formed by
the reduction of glycol units in the basement membrane in a process similar
to that of the reticulin stain (e-Fig. 54.7).
3. Trichrome. There are a number of variations of the trichrome stain; in general,
they all use three dyes with affinities for different connective tissue elements.
The first step in the trichrome stain involves mordanting the tissue with a
heavy metal fixative such as Bouin's Fixative. The tissue is then dyed with a
nuclear stain, most often an iron hematoxylin. Next, an acidic dye such as
acid fuchsin or Biebrich scarlet is used to stain the cytoplasm of cells, collagen,
and muscle; either phosphotungstic acid or phosphomolybdic acid is then
used to remove the acid dye from the collagen (since the cytoplasm of cells
is less permeable than collagen, with proper timing the dye can be removed
from collagen without complete removal from other tissue elements). Finally,
collagen is stained using aniline blue. As the name suggests, the method
produces tissue sections that are stained in three colors: black cell nuclei; red
cell cytoplasm and muscle; and blue collagen and mucus (e-Fig. 54.8).
4. VVG. The VVG is another compound stain. The tissue section is first over-
stained with an iron hematoxylin solution, and then the hematoxylin is differ-
entiated by removal with ferric chloride; since elastic fibers have the greatest
affinity for iron hematoxylin, they are the last to decolorize, and so it is
possible to halt the differentiation at the point when the elastic fibers are the
only tissue elements still stained. The tissue section is then treated with van
Gieson's solution, which contains the dye acid fuchsin; in a very strong acid
solution the dye selectively stains only collagen. Picric acid, used to maintain
the proper pH during staining, stains the rest of the tissue elements yellow.
The VVG stain demonstrates red-stained collagen, black elastic fibers, and a
yellow background (e-Fig. 54.9).
5. PTAH. The PTAH stain requires mordanting of the tissue section in Zenker's
fixative before staining. Because phosphotungstic acid is present in the stain-
ing solution in excess over hematoxylin, all of the hematoxylin is bound into
a tungsten-hematein lake, which selectively binds to cell nuclei, fibrin, and
cross-striations in muscle fibers. The excess unreacted phosphotungstic acid
stains the remaining tissue elements red to red-brown.
6. Pentachrome. The pentachrome stain is a compound stain that essentially
combines the elastic fiber staining of a modified VVG stain with a modified
trichrome stain. Alcian blue is first used to stain mucosubstances, and then
iron hematoxylin is used to stain elastic fibers. Following the differentiation
of the iron hematoxylin, a combined crocein scarlet and acid fuchsin solution
is used to stain muscle, cellular cytoplasm, amorphous ground substance,
and collagen red. Phosphotungstic acid in solution is then used to decolorize
the collagen and amorphous ground substance, which is then subsequently
stained yellow using a saturated alcoholic safran solution.
7. Oil red 0. This stain is used to demonstrate fat in tissue sections, or lipid
droplets in cell cytoplasm. The dye Oil red 0 is highly soluble in lipids, and
when used in solution with isopropanol is actually more soluble in fat than
in alcohol.
This stain requires the use of frozen section tissues because the alcohol
and xylene steps in standard paraffin processing remove virtually all lipids
from the tissue. The staining itself is fairly straightforward: frozen tissue
sections are cut, fixed with formaldehyde, and then stained in the Oil red 0
solution. The sections are next rinsed free from any excess stain and then
counterstained using hematoxylin. The stain results in blue cell nuclei and
bright red staining of fat droplets.
C. Microorganisms. There are many different stains that can be performed to
demonstrate microorganisms, specifically bacteria and fungi, in tissue sections.
The most commonly used stains are the acid-fast bacteria (AFB), the Fite modi-
fication of the AFB, Gram, Grocott methenamine silver (GMS), Warthin-Starry,
and PAS.
1. Gram stain. The tissue Gram stain is not much different from the standard
Gram stain performed in the microbiology lab. In the tissue Gram stain, how-
evet; after the use of crystal violet to demonstrate gram-positive bacteria by
a blue colot; basic fuchsin (a red dye) is used to demonstrate gram-negative
organisms as well as cell nuclei. Differentiation of gram-positive and gram-
negative bacteria is still a critical step; overdifferentiation is a common stain-
ing error. The final step in the Brown-Hopps Gram stain involves treating
the tissue with a picric acid solution that renders the background yellow.
In addition to identification of bacteria, the stains can be used to demon-
strate some cases of actinomyces, Nocardia infections, coccidioidomycosis,
blastomycosis, cryptococcosis, aspergillosis, rhinosporidiosis, and amebiasis
(e-Fig. 54.10).
2. AFB. The tissue AFB stain is simply a modification of the standard Ziehl-
Neelsen and Kinyoun stains that are based on the fact that carbol-fuchsin,
a solution created by reacting basic fuchsin with phenol in alcohol, is sol-
uble in lipids. Tissue sections are first treated with the carbol-fuchsin solu-
tion and then differentiated using acid alcohol; bacteria that have waxy,
lipid-containing cell walls resist decolorization with acid alcohol and are said
to be "acid fast." A methylene blue counterstain is used to highlight other
tissue elements and provide a background to highlight the red microorgan-
isms (e-Fig. 54.11). A slight modification of this procedure can be used to
specifically stain for Nocardia species in tissue sections. Another modifica-
tion of this stain, known as the Fite AFB stain, is used when Mycobacterium
leprae is suspected (e-Fig. 54.12).
3. GMS. This stain utilizes most of the same chemical reactions and principles as
the JMS stain. In the GMS stain, however, a stronger oxidizer, chromic acid,
is used instead of the weaker periodic acid. Since the cellular walls of fungi are
very thick and contain much more carbohydrate than basement membranes
and reticulin fibers of the surrounding tissue, the stronger oxidizer allows for
creation of dialdehyde groups from the carbohydrates of the fungi cell walls
with overoxidation and subsequent destruction in basement membranes and
other carbohydrate structures in the tissue section. The GMS stain utilizes a
light green counterstain, resulting in fungus cell walls that are various shades
of black to taupe in a light green background (e-Fig. 54.13).
4. Warthin-Starry. The Warthin-Starry method is used primarily for the demon-
stration of spirochetes, but other bacteria are also stained. The procedure is
based on the principle that bacteria in general and spirochetes in particular
have the ability to bind silver ions.
The staining procedure therefore involves impregnation of the spirochetes
in the tissue with silver ions, with subsequent reduction of these ions to metal-
lic silver using a developer containing hydroquinone. The stain demonstrates
black spirochetes against a yellow to pale brown background. The spiral mor-
phology of this form of bacteria can be fully appreciated by the use of this
method (e-Figs. 54.14 and 54.15).
5. PAS. This stain is often used for the demonstration of fungi in tissue, but is
most helpful when a counterstain of light green is applied. The method used in
stains for microorganisms is no different than when used for carbohydrates.
D. Nervous system. Most of the stains used on tissues from the nervous system are
for demonstration of either nerve fibers or myelin sheath. Two commonly used
stains for central nervous system tissues are the Bielschowsky and the Luxol fast
blue.
1. Bielschowsky. The Bielschowsky and all of its modifications are silver stains
that follow the principles and general steps of the reticulin stain. In the
Bielschowsky technique, the tissue sections are impregnated with a 20%
silver nitrate solution and then treated with an ammoniacal silver solution to
which formaldehyde has been added. Nerve endings, neurofibrils, neurofib-
rillary tangles, and neuritic plaques are all stained black.
2. Luxol fast blue. Luxol fast blue, a phthalocyanine dye that is soluble in alcohol,
is attracted to bases found in the lipoproteins of the myelin sheath. For this
stain, tissue sections are treated with Luxol fast blue over an extended period
of time (usually overnight) and then differentiated with a lithium carbonate
solution. Since Luxol fast blue has a strong affinity for the lipoproteins of the
myelin sheath, it remains bound to these lipoproteins even after removal from
other tissue elements. The myelin sheath is stained blue against a colorless
background.
E. Pigments and minerals. Pigments are substances deposited in the interstitium of
tissues, or as inclusions or granules in the cytoplasm of cells. Pigments can be
derived from minerals such as iron and calcium, or can be endogenous such as
melanin. The following staining techniques are used for the demonstration of
the most commonly encountered pigments.
1. Iron. The Prussian blue reaction is the most common staining technique for
the demonstration of iron in tissue sections. Prussian blue stains only weakly
bound iron. Strongly bound iron, such as iron in hemoglobin, will not stain.
The principle of this stain is simple: when treated with potassium ferro-
cyanide in an acidic solution, ferrous ions in tissue react to form an insoluble
blue pigment. A nuclear fast red counterstain is usually applied to demon-
strate the background tissue morphology (e-Fig. 54.16).
2. Urates. A modified GMS stain can be used to demonstrate uric acid crystals.
No oxidation of the tissue sections is performed; instead, the sections are
reacted in the methenamine solution for an extended period at an elevated
temperature. Silver ions deposit on the uric acid crystals which in turn reduce
the silver ions to metallic silvet; so no toning is necessary. A light green
counterstain is usually applied, and the resulting stained section demonstrates
black uric acid crystals in a green background.
This stain requires the use of alcohol-fixed tissues because uric acid is
soluble in water.
3. Calcium (von Kassa method). The von Kossa method to stain for calcium is
very simple. Tissue sections are incubated in a 5% silver nitrate solution
under a very strong light source. The silver ions deposit on the calcium and
are reduced to metallic silver by the strong light, in much the same pro-
cess as occurs in photographic film. The stained section is rinsed free of
any unreacted silver ions by a sodium thiosulfate wash, and then counter-
stained with nuclear fast red to highlight the background tissue morphology
(e-Fig. 54.17).
4. Copper (Rhodanine method}. Copper can be demonstrated in tissues using
several methods, but the most sensitive method employs rhodanine. Tis-
sue sections are subjected to a saturated solution of 5-(p-dimethylamino-
benzylidene) rhodanine in aqueous solution. The rhodanine reacts with
proteins that have bound copper rather than directly with the copper itself.
The excess stain is rinsed from the sections, and the sections are counter-
stained with Mayer's hematoxylin, an aqueous hematoxylin that will not
overstain the rhodanine reaction. This tissue stain demonstrates bound cop-
per as a granular red pigment with pale blue cell nuclei (e-Fig. 54.18).
5. Argyrophil staining. Argyrophil substances within a cell bind silver ions. They
are "silver loving" but do not reduce silver to its visible metallic form. There
are several different techniques for the demonstration of argyrophil sub-
stances, all of which are chemically similar to the Warthin-Starry technique.
A solution of silver nitrate is used to impregnate the argyrophilic substances
in the tissue, and a reducing solution containing hydroquinone is then used
to reduce the bound silver ions to metallic silver. Nuclear fast red is often
used as a counterstain. By this approach, argyrophilic substances are stained
black.
6. Argentaffin staining (Fontana-Masson method). Argentaffin substances not only
bind silver ions like argyrophilic substances, but also reduce bound ionic sil-
ver to metallic silver without the use of a developer or other reducing agent.
This property of argentaffin substances, which include melanin, underlies
the Fontana-Masson stain. An ammoniacal silver solution is used to treat
tissue sections, and the argentaffin substances within the tissue not only
bind the silver ions in the solution, but also reduce them to metallic silver.
Gold chloride is used as in the reticulin stain to tone the metallic silver from
brown to black. Nuclear fast red is the counterstain of choice for this stain
(e-Fig. 54.19).
7. Bile pigments. Bile pigment stains are used on liver sections to distinguish
bile pigments from lipofuchsin. Fouchet's reaction demonstrates biliverdin,
bilirubin, and most other bile pigments. The tissue sections are treated with an
aqueous solution of trichloroacetic acid and ferric chloride, which renders an
emerald green precipitate, and von Gieson's solution is used as a counterstain
..,.
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rigors of formalin fixation and paraffin processing but not acid decalcification;
therefore, application of the stain to bone marrow specimens requires the use of
nonacid decalcification). In the Leder method, tissue is treated with a solution
of naphthol-chloroacetate and pararosaniline, and reaction with the cellular
chloroacetate esterase forms a red precipitate. Hematoxylin is used as the coun-
terstain to demonstrate nuclear detail (e-Fig. 54.20).
G. Staining. Table 54.1 presents tissue stains listed by their most common name,
the tissue element that they demonstrate, and the resulting coloration or specific
result.
SUGGESTED READINGS
Bancroft JD, Gamble M, eds. Theory and Practice of Histological Techniques. 5th ed. Edinburgh,
UK: Churchill Livingston; 2002.
Carson FL. Histotechnology: A Self-Instructional Text. 2nd ed. Chicago: ASCP Press; 1997.
Sheehan DC, Hrapchak BB, eds. Theory and Practice of Histotechnology. 2nd ed. Columbus, OH:
Battelle Press; 1980.
Immuno histochemistry
Peter A. Humphrey
I. INTRODUCTION. Immunohistochemistry is one of the most powerful and widely

used ancillary methods in surgical pathology. The technique makes it possible to
simultaneously visualize cell type and differentiation markers in standard tissue
sections by light microscopy, and has revolutionized diagnostic surgical pathology.
Antigens in tissue sections were first detected using antibodies via immunoflu-
orescence performed on frozen sections (Proc Soc Exp Bioi Med. 1941;47:200).
Although immunofluorescence is still used in the evaluation of medical kidney
biopsies (see Chap. 19), currently the most common approach for diagnostic
detection of antigens uses formalin-fixed paraffin-embedded (FFPE) tissue sec-
tions and immunoperoxidase methodology. This enzymatic labeling technique
has evolved from simple direct peroxidase conjugation of the primary antibody, to
the use of multistep peroxidase-antiperoxidase (PAP), avidin-biotin (and related)
conjugate methods, which, along with amplification techniques such as tyramide
and polymer-based labeling, allow for much greater sensitivity in antigen detec-
tion.
The laboratory utilization of immunohistochemistry (also known as immuno-
histology and immunostaining) requires appropriate test selection, specimen
acquisition and management, methodology, validation, reporting, and interpre-
tation. This chapter provides a concise overview of these elements. Guidelines
pertaining to these elements were published in 2011 by the Clinical and Labo-
ratory Standards Institute (CLSI) (www.clsi.org) in a comprehensive document
entitled "Quality Assurance for Design Control and Implementation of Immuno-
histochemistry Assays; Approved Guideline-Second Edition."
II. TEST SELECTION (PREANALYTICAL PHASE). Immunohistochemical stains are usu-
ally ordered after examination of hematoxylin and eosin (H&E)-stained sections.
Common indications for immunohistochemistry are the diagnosis and character-
ization of neoplasms, but there are other indications as well, such as detection of
infectious organisms and evaluation of prognostic and/or predictive factors.
The use of specific immunostains is driven by the clinical and morphological
context of each individual case. Panels of antibodies are often used, and these pan-
els should be devised on the basis of the anticipated value added to the clinical,
radiographic, and pathological differential diagnosis. Panels of antibodies should
thus be directed toward a specific question. Various approaches have been used to
help construct appropriate immunostain panels, including algorithmic approaches
and tabular approaches. Web sites with information on the specificity and sensi-
tivity of various immunostains, and on construction of immunostain panels based
on differential diagnosis of specific neoplasms, also exist (see below).
Ill. SPECIMEN TYPE AND TISSUE MANAGEMENT. Immunostains can also be performed
on cytological specimens, although they are usually performed on standard his-
tological tissue sections. Whereas the use of FFPE tissue sections offers obvious
logistical advantages, some antigens require the use of fresh tissue or tissue pre-
served with ethanol-based fixatives. The discussion here focuses on tissues fixed
in 10% neutral buffered formalin, because this is the tissue type most commonly
available for analysis in routine clinical practice.
Immediate fixation in neutral pH formalin for 12 to 48 hours at room tem-
perature is desirable. However, it must be noted that formalin induces cross-
links that may mask some epitopes, resulting in loss of immunoreactivity. Acid
851
decalcification of bone samples can also cause loss of immunoreactivity. "Unmask-

ing" of some epitopes from FFPE tissue (and tissue treated with acid decalcifica-
tion) can be accomplished by antigen retrieval techniques. Enzyme digestion was
used for this purpose in the past, but now simple heat treatment (heat-induced
antigen retrieval) is the most commonly used approach to optimize antigen detec-
tion. Unstained tissue sections cut onto charged slides or poly-L-lysine-coated
slides (or gelatin- or albumin-coated slides) are typically used for immunohisto-
chemistry, but it is possible to perform immunostains on sections that have already
been stained with H&E (Am] Clin Pathol. 2005;124:708); because success with
such restaining protocols is variable, unstained tissue sections remain the best
resource for immunostains. Immunostaining should be performed on freshly cut
sections from the paraffin block, because unstained sections exposed to air may
lose antigen immunoreactivity over the course of days to weeks.
IV. METHODOLOGY
A. The primary antibody is an immunoglobulin molecule that binds to the target
antigen in the tissue sections. The primary antibody may be either a mono-
clonal antibody derived via the hybridoma technique, or a polyclonal antibody
from an antiserum. In general, polyclonal antibodies tend to be more sensitive
but less specific than monoclonal antibodies. Unlike monoclonal antibodies,
polyclonal antibodies are not uniform reagents of unlimited supply; different
batches of antisera may result in polyclonal antibody heterogeneity.
Each antibody, whether polyclonal or monoclonal in origin, needs to be
tested for sensitivity and specificity in target antigen detection, and the reac-
tion conditions for its use need to be optimized. Titration experiments must be
performed to achieve a working dilution of the primary antibody that yields the
greatest contrast between specific staining and nonspecific staining. If predi-
luted reagents and kits are used, it is recommended that the manufacturer
protocol be followed because validation was performed with those reaction
conditions.
B. Background staining results from nonspecific antibody binding and from
endogenous enzymes that nonspecifically interact with the chromogenic sub-
strate. Nonspecific antibody binding is more likely to occur with poly-
clonal antibodies. Endogenous enzymes that cause background staining are
found in normal cells including erythrocytes, neutrophils, eosinophils, hepa-
tocytes, plasma cells, and neoplastic cells; their activity can often be blocked
(e.g., endogenous peroxidase can be blocked by incubation with hydrogen
peroxide).
c. Detection systems
1. Direct conjugate-labeled antibody method. In this method, the label-such
as peroxidase or fluorescein-is directly chemically linked to the primary
antibody. Disadvantages of this approach include a requirement for a large
amount of primary antibody for labeling and a lack of signal amplification.
2. Indirect or sandwich method. The primary antibody is unlabeled, and a
secondary antibody, reactive against the primary antibody, carries the
label.
3. Unlabeled antibody method. Also known as the PAP method. This procedure
uses an unlabeled primary antibody, an unlabeled bridge antibody, and
a complex of an antiperoxidase antibody and the peroxidase molecule
itself. The bridge antibody, directed against both the primary antibody
and the anti peroxidase antibody, links the primary antibody-tissue antigen
reaction to the signal generated by the peroxidase. This approach has
largely been replaced by the more sensitive approaches below.
4. Avidin-biotin or streptavidin--biotin conjugate method. A biotinylated sec-
ondary antibody is used to recognize the primary antibody; avidin or
streptavidin complexed with biotinylated peroxidase is then bound to the
Chapter 55 • Immunohistochemistry I 8 53
secondary antibody (both avidin and streptavidin have extremely high

affinity for biotin). These reactions deliver several peroxidase molecules
to the primary antibody binding site and so boost sensitivity. Streptavidin
has several advantages over avidin, including decreased background stain-
mg.
5. Tyramine amplification {catalyzed signal amplification [CSA]) methods. With
these methods, there is increased sensitivity due to greater accumulation
of biotin at the antigen-primary antibody reaction site as a result of the
catalytic activity of peroxidase on biotinylated tyramine.
6. Polymer-based labels. This approach uses dextran chain polymers to local-
ize numerous enzyme molecules to the antigen site by linking multi-
ple antibody and enzyme molecules together along the polymer chain
(Fig. 55.1). This method avoids problems due to endogenous biotin.
7. Alkaline phosphatase can be used instead of peroxidase when the target
antigen is in tissues rich in myeloid cells that contain high levels of endoge-
nous peroxidases, such as bone marrow.
8. Chromogens are the color-producing reactants in the detection system.
Methods using peroxidase, diaminobenzidine (DAB, produces a brown
color), and 3-amino-9-ethyl carbazole (AEC, produces a red color) are
commonly used.
9. Counterstaining is most often accomplished using the nuclear stain hema-
toxylin. Care must be taken not to overstain the tissue sections, especially
when the target antigen is located in the nucleus.
10. Antibody cocktails can be used to detect two or more antigens at the same
time. Single-color (Am I Surg Pathol. 2005;29:579) or two-color (Am I
Clin Pathol. 2005;123:231) approaches can be used.
11. Automation. Automated immunostaining devices are in routine use in many
laboratories and can improve standardization, throughput, and repro-
ducibility of immunohistochemical procedures.
12. Quantitative immunohistochemistry. In the past, immunohistochemical reac-
tions have been manually scored in a semiquantitative manner by patholo-
gists, via analysis of the staining intensity and estimates of the percentage
of cells stained in the area of interest. Such scoring has been particu-
larly important for a few markers, for example, detection of HER2/neu
immunoreactivity in breast carcinoma to determine the eligibility of
breast cancer patients for trastuzumab (Herceptin) therapy(] Clin Oncol.
2007;25:118). Because image analysis improves the consistency of quan-
titative immunohistochemical scoring, it is likely that digital microscopy
(see Chap. 63) with image analysis will be increasingly used in this context
in the future.
V. VALIDATION. Positive and negative controls should be included in every sample run
and reviewed along with the test immunohistochemical reaction. A positive tissue
or cell control known to express the antigen under investigation should be used,
and should be subjected to the same reaction conditions in the same analytical
run as the test tissues or cells. Some laboratories place a positive control tissue
section on the same slide as the test tissue section; for some immunostains, there
may be an internal positive control in the test tissue. A negative control can
be generated using a tissue known to lack the antigen of interest or by replacing
the primary antibody with an irrelevant nonimmune antibody or antiserum; a
search should also be made in the test tissues for negative internal controls.
VI. REPORTING. Immunohistochemical stain reports should include specific content
elements (Table 55.1).
VII. INTERPRETATION. Interpretation of immunostains, including their significance,
should be integrated with the interpretation of the clinical, radiographic, gross,
and histopathologic findings of the case, as well as the results of any additional
8!4 I SECTION XIII: ANCILLARY METHODS
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method. This technique allows linkage of numerous molecules of
enzyme (either peroxidase or alkaline phosphatase) to one (B) or
more (A) molecules of secondary antibody. Delivery of a large
number of enzyme molecules to the antigen-primary antibody
reaction site yields high sensitivity. (Modified from Taylor CR,
Cote RJ, eds. Immunamicroscopy. A Diagnostic Tool for the Sur-
gical Pathologist, 3rd ed. Philadelphia: Elseviet; 2006.)
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Immunofluorescence
Anne C. Lind, Joseph P. Gaut, and Rosa M. Davila
I. INTRODUCTION. Immunofluorescence studies are used to support fixed tissue diag-

noses and to provide additional diagnostic and prognostic information as it relates
to autoimmune diseases, vesiculobullous diseases, transplantation, and glomeru-
lar disease. Immunofluorescence requires fresh tissue submitted in a preservative
nonfixative transport medium such as Michel's medium, or fresh frozen tissue; cur-
rently, immunofluorescence is not routinely performed on formalin-fixed paraffin-
embedded (FFPE) tissue. For fresh tissue, the transport medium should be held at
room temperature; temperature extremes should be avoided. A specialized micro-
scope and a room where the majority of ambient light can be extinguished are
required for either direct or indirect immunofluorescence examination.
II. DIRECT IMMUNOFLUORESCENCE
A. Skinlmucosa. Cutaneous/mucosal biopsies for immunofluorescence are stained
with fluorescein-labeled antibodies to immunoglobulin (lgG, IgA, IgM), comple-
ment (C'3), and collagen IY. The patterns of staining that correlate with specific
diseases are discussed in more detail with the corresponding diagnoses in the
chapter on inflammatory disorders of the skin (Chap. 38). Patterns of stain-
ing include basement membrane (e-Fig. 56.1)* and intercellular (e-Fig. 56.2)
positivity.
1. Bullous pemphigoid. A biopsy of perilesional tissue to include the edge of a
blister is optimal. Some studies have reported a high false-negative rate for
tissue from the lower extremity; if possible, tissue for direct immunofluores-
cence should be obtained from above the knee. Direct immunofluorescence
is positive in approximately 85% to 90% of cases of bullous pemphigoid.
2. Pemphigus vulgaris. A biopsy of perilesional tissue, without including the
edge of a bliste.t;, is optimal. Direct immunofluorescence is positive in approx-
imately 90% to 95% of cases of pemphigus vulgaris.
3. Dermatitis herpetiformis. A biopsy of perilesional tissue, avoiding excoriated
areas, is optimal. Direct immunofluorescence is positive in ""80% of cases of
dermatitis herpetiformis.
4. Vasculitis. A biopsy of lesional tissue is required. Sources vary on recommen-
dations regarding age of lesion; howeve.t;, the majority favors a lesion that
has been present for <48 hours.
5. Lupus. A biopsy of an established lesion that has been present for at least
8 weeks, preferably 12, is required to detect immunofluorescence positivity
in discoid lupus. In the past, prognostic information regarding disease activity
was associated with immunofluorescence positivity in sun-protected, nonle-
sional skin in systemic lupus.
B. Kidney. It is the current standard of practice to obtain a minimum of two tis-
sue cores to distribute for light microscopy, immunofluorescence, and electron
microscopic analysis (Kidney Int. 1999;55:713; Mod Pathol. 2004;17:1555).
Renal biopsies are usually performed under ultrasound guidance, and the tissue
cores are evaluated in the ultrasound suite with a dissecting microscope. Since
it is important to avoid air drying of the tissue, the biopsy is placed in a petri
856
Chapter 56 • Immunofluorescence I 8 57
dish with a balanced salt solution while it is examined with the dissecting micro-
scope. Because glomerular diseases are a common indication for renal biopsy,
the tissue is distributed in such a way that approximately two or more glomeruli
are examined by electron microscopy, two or more by immunofluorescence, and
10 or more by light microscopy. The tissue assigned to electron microscopy is
placed in glutaraldehyde; tissue for immunofluorescence is frozen, and tissue
for light microscopy is placed in formalin. When a biopsy needs to be trans-
ported to the laboratory from a remote site, it should be placed in a container
with transport medium such as Michel's medium. Although it has been shown
that immunofluorescence can be performed on FFPE tissue for the evaluation
of renal biopsies (Kidney Int. 2006;70:2148), FFPE tissue is not routinely used
for immunofluorescence evaluation.
Light microscopic evaluation of the renal biopsy is performed using hema-
toxylin and eosin, periodic acid-Schiff (PAS), trichrome, and methenamine
silver-stained sections. The PAS and silver stains facilitate examination of the
basement membranes, and the trichrome stain highlights areas of interstitial
fibrosis. Evaluation with fluorescein isothiocyanate (FITC)--conjugated antibod-
ies against IgG, IgM, IgA, C'3, C1 q, fibrinogen, albumin, K. and }.. are performed
by direct immunofluorescence. Transplant kidney biopsies are also stained for
C4d using an indirect immunofluorescence technique. C4d is used to assess for
humoral rejection, usually seen as immunopositivity of the peritubular capillar-
ies. Addition of a 3% Evans Blue counterstain (Health Scientific, Saint Louis,
MO) is helpful to decrease the FITC background and to enhance visualization
of the tissue (Am] Transplant. 2009;9:812) (e-Fig. 56.3).
C. Lung. In the event of suspected humoral rejection after pulmonary transplan-
tation, biopsy of pulmonary parenchymal tissue acquired via transbronchial
biopsy can be submitted in Michel's medium for evaluation of complement
(C4d) by direct immunofluorescence. It is worth noting that evaluation for the
presence of C4d can also be accomplished using an immunoperoxidase method
on formalin-fixed tissue.
Ill. INDIRECT IMMUNOFLUORESCENCE. Blood (5 to 10 mL) drawn into a tube without
anticoagulant is required for all indirect immunofluorescence studies. The serum
is removed and is applied to an epithelial substrate. The substrate varies with the
clinical diagnosis, and the clinical diagnosis should guide the decision to pursue the
appropriate indirect immunofluorescence study. For indirect immunofluorescence,
serial dilutions (1:10 to 1:1280) of serum are inoculated onto the tissue substrate
together with fluorescein-labeled anti-lgG.
A. Pemphigus vulgaris. The primary utility for indirect immunofluorescence is for
the diagnosis of pemphigus vulgaris and to follow response to therapy. Serial
dilutions are performed and the end point of positivity of intercellular IgG is
reported (e-Fig. 56.2). Commercially prepared slides using guinea pig or mon-
key esophagus are used. As in other serologic tests, it is possible to get a pro-
zone effect in patients with pemphigus vulgaris, so additional dilutions may be
required to avoid a false-negative result.
B. Paraneoplastic pemphigus. The substrate for evaluation of paraneoplastic pem-
phigus is murine/rat bladder epithelium. Because this test is not commonly
ordered, it is usually only performed at reference laboratories.
C. Bullous and/or cicatricial pemphigoid. Circulating antibodies that produce
a linear basement membrane zone positivity (e-Fig. 56.1) are detected in
fewer than half of the patients with documented pemphigoid, making ancil-
lary testing by indirect immunofluorescence minimally useful for this disease
process.
D. Dermatitis herpetiformis. Circulating antibodies are not detectable in the serum of
patients with dermatitis herpetiformis; therefore, indirect immunofluorescence
is not indicated.
E. Reference laboratories. Reference laboratories are a useful resource for indi-

rect immunofluorescence testing for rare diseases. Evaluation of epidermolysis
bullosa is performed by Beutner Laboratories (Buffalo, NY). Testing for para-
neoplastic pemphigus is performed by Mayo Clinical Laboratories (Rochestet;
MN). Some research laboratories also perform specialized testing for rare vesicu-
lobullous dermatoses; however, testing in this setting may not be approved for
clinical use.
Flow Cytometry
Friederike Kreisel
I. BASIC PRINCIPLE. Flow cytometry simultaneously measures and analyzes multiple

physical and/or chemical characteristics of single particles, usually cells, as they
flow in a fluid stream through a beam of light. With this technique, any suspended
microparticle, ranging in size from 0.2 to 150 ~m, can be analyzed. Peripheral
blood or bone marrow aspirate specimens already represent a suspension of single
cells but they must be prevented from clotting by using collection tubes contain-
ing disodium ethylenediaminetetraacetic acid (EDTA), sodium citrate, or heparin.
Enrichment of leukocytes can be achieved by lysis of accompanying red blood
cells with ammonium chloride buffer or use of density-gradient separation. Many
protocols also exist for producing cell suspensions from solid tissue suspensions.
II. FLOW CYTOMETER. The flow cytometer is composed of three main systems
(Fig. 57.1).
A. The flow system. The sample is injected into a stream of sheath fluid within the
flow chamber. Through the principle of hydrodynamic focusing, the particles
are forced into the center of the stream and transported through a laser beam
for analysis, one particle or cell at a time. A higher flow rate is generally used for
the immunophenotyping of cells. A lower flow rate is important in applications
where greater resolution is needed, such as DNA analysis.
B. The optical system. Lasers illuminate the particles in the sample stream and
optical mirrors and filters route the different wavelengths of the generated light
scatter and fluorescent signals to the appropriate photodetectors.
1. Light scatter. Light scattering occurs when a particle or cell deflects laser
light. Forward-scattered (FSC) light is in line with the laser light beam and
represents a measurement of the cell surface size. Side-scattered (SSC) light is
collected perpendicular to the laser light beam and analyzes the granularity
or internal complexity of a cell. Leukocytes can be separated into different
subpopulations using FSC and SSC. For example, lymphocytes will show
both a low forward scatter and a low side scatter due to the small size and
lack of cytoplasmic granulation. In contrast, neutrophils are larger in size
and show granular cytoplasm as well as a complex nucleus, and therefore
will show both a high forward and a side scatter (e-Fig. 57.1). •
2. Fluorescence. Another way to identify particular subpopulations is to con-
jugate fluorescent dyes to monoclonal antibodies directed toward antigens
on a particular cell subset. The staining procedure can be carried out in a
direct or indirect staining process. The direct staining procedure involves a
single staining incubation, followed by several washes to remove nonspecifi-
cally bound antibodies. The indirect staining procedure involves the incuba-
tion of cells with a nonfluorescent monoclonal antibody directed toward the
specific antigen. After washing to remove nonspecifically bound antibody,
there is a second incubation with a fluorescent antibody directed against the
monoclonal antibody. Although more time-consuming, the indirect staining
procedure is less expensive.
Argon ion lasers are the most common lasers used in flow cytome-
try because the 488-nm light emitted can be absorbed by more than one
859
Electronic System
..---
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Figure 57.1 The How cytometer is composed of a How system, optical system, and electronic
system. The flow system transports cells in a stream to the laser beam for analysis. The optical
system consists of lasers to illuminate the cells in the sample stream and optical filters to direct
the resulting light signals to the appropriate dete<:tors. The electronic system converts light signals
into electronic signals that are processed by the computer.
fluorochrome. Examples of fluorochromes that are conjugated to antibodies

are fluorescein isothiocyanate (FITC) and phycoerythrin (PE). FITC absorbs
light in the range of 460 to 510 nm and then fluoresces in the range of 510 to
560 run, with a peak at,....525 nm, giving a green fluorescent color. PE absorbs
light in the range of 480 to 565 run and fluoresces at ,....570 nm, giving a red
fluorescent color. Although both fluorochromes absorb light at "-'488 nm,
the resulting different peak emission wavelengths can be detected by differ-
ent detectors, which make it possible to use more than one fluorochrome in
one sample to simultaneously collect information on the expression of sev-
eral markers in a specific cell. Combined with the FSC and SSC data, the
staining pattern of each subpopulation will aid in delineating which cells are
present in a sample and in what percentage they are present (e-Fig. 57.2).
C. The electronic system. Photodetectors convert the generated FSC, SSC, and flu-
orescent light signals into electrical impulses.
Chapter 57 • Flow Cytometry I 8 61
1. Photodetectors. Generally, two types of photodetectors are used in flow

cytometry: photomultiplier tubes (PMTs) and photodiodes. PMTs amplify
the electrical current generated from the light signals, and are mostly used to
detect weaker signals generated by SSC and fluorescence. Photodiodes are less
sensitive to light signals and are used to detect stronger FSC signals. Ampli-
fication of a signal detected by a photodetector can be achieved by means of
log amplification or linear amplification. Log amplifiers are usually used to
separate negative from dim positive signals, and are commonly used for sig-
nals from cells stained with fluorochrome-labeled antibodies because these
cells often exhibit a great range of fluorescence intensities. Linear amplifiers
are generally used to analyze forward and side scatter signals.
2. Conversion into a digital value. The intensity of the electronic impulses derived
from the photodetectors is assigned a digital value by means of an analog
to digital converter (ADC). The role of the ADC is to analyze a continuous
distribution of signals falling into a channel of a certain light intensity range
and to organize these signals into a data plot.
An electronic threshold is used to limit the number of events to the popu-
lation of interest. For example, the threshold can be set on FSC to eliminate
events that represent debris smaller than the threshold channel number. Aher
the acquired data are saved, the cell populations can be displayed by several
different types of data plots. A single parameter such as FSC or FITC (FL1)
can be displayed as a single-parameter histogram, on which the horizontal
axis represents the signal intensity (expressed as the parameter's signal value
in channel numbers) and the vertical axis represents the number of events per
channel (e-Fig. 57.3). Two parameters such as FITC (FL1) and PE (FL2) can
be displayed simultaneously in a dot plot in which one parameter is displayed
on the x-axis and the other on they-axis (e-Fig. 57.4).
Finally, a subset of data can be defined through a gate. On the basis of
FSC and SSC, an electronic gate can be set on a selected population of interest
and analysis restricted to only that subset.
Ill. USES
A. Cell markers. Flow cytometry has become a valuable ancillary method for clas-
sifying acute leukemias and lymphomas. Monoclonal antibody technology has
provided flow cytometry with a large variety of antibodies specific to nuclear,
cytoplasmic, and surface antigens characteristic of particular cell subsets. These
are organized as clusters of differentiation (CD) antigens, which help differen-
tiate cells into the different subpopulations of the hematopoietic and lymphoid
system. Selected CD markers that are used in the diagnosis of acute leukemias
and lymphomas are discussed in Chapters 42 and 43. Flow cytometric scat-
tergrams of an example of precursor B-lymphoblastic leukemia are shown in
e-Figure 57.5. Typical flow cytometric findings of chronic lymphocytic leukemia
(CLL) are shown in e-Figures 57.6 and 57.7.
B. DNA content. Besides analyzing surface or cytoplasmic properties of a cell, flow
cytometry can also be used to analyze the DNA content of a cell. Several types
of fluorescent stains are available that target different DNA bases within the
double helix, most of which require the use of laser with significant ultraviolet
output to be specific. For example, HOECHST 33342 or 4',6-diamidino-2-
phenylindole (DAPI) are specific for the adenine/thymine base pairs, whereas
mithramycin and chromomycin A3 preferentially target guanine/cytosine base
pairs. Propidium iodide is not very specific because it stains all double-stranded
nucleic acids, but it can be included as a DNA stain in conventional cytometers
with low-power argon lasers because it absorbs light at 488 nm.
DNA staining is generally performed to assess the amount of DNA in the
nucleus of a cell or to analyze cell division. Because most normal cells contain
the same amount of DNA (diploid or euploid), measurement of DNA content
DNA Histogram
G1
DNA Content
The Cell Cycle

Figure 57.2 Histogram of the DNA distribution in a cycling population
of cells that illustrate the distribution of nuclear DNA content present at
a particular moment in the population of cells.
of a cell will differentiate normal cells from aneuploid malignant cells. The
histograms from malignant tumors will show abnormal peaks corresponding to
more (hyperdiploid) or less (hypodiploid) DNA than nonnal cells.
C. Cell cycle analysis. The cell cycle is composed of four phases. Cells in Gl are
recovering from division or are preparing for division; cells in S phase are in
the process of synthesizing new DNA. Cells in G2 phase have finished DNA
synthesis and therefore have double the normal amount of DNA (tetraploid).
M phase encompasses division into two similar daughter cells. (Note that cells
in GO phase are not cycling at all). The cell cycle can be plotted as a histogram
with the number of cells per channel on the y-axis and the fluorescence intensity
of cells stained for DNA content on the x-axis (Fig. 57.2).
SUGGESTED READINGS
Keren DFt McCoy 1Pt Carey JLt eds. Plow Cytometry in Clinical Diagnosis. 3rd ed. Chicago: ASCP
Press; 2001:31-65.
Givan AL. Flow Cytometry, First Principles. New York: Wtley-Liss; 1.9.92:75-102.
Ormerod MG, ed. Flow Cytometry. 3rd ed. Oxford: Oxford University Press; 2005:23-33.
Cytogenetics
Shashikant Kulkarni, Hussam AI-Kateb,
and Catherine Cottrell
I. INTRODUCTION. The three significant milestones in the history of clinical cytoge-

netics are the preparation of chromosome spreads from peripheral blood cultures
(Exp Cell Res. 1960;20:613), the development of hypotonic methods to obtain
enhanced chromosome spreads (Cancer Res. 1960;20:462), and the discovery that
fluorescent quinacrine compounds could be used to demonstrate a unique banding
pattern for each human chromosome pair (Hereditas. 1971;67:89). The remark-
able advancement of the field of human cytogenetics is emphasized by the fact that
it has been only 50 years since the correct number of human chromosomes was
established. The various banding methods in current use not only permit identifi-
cation of each chromosome, but also make it possible to detect specific alterations
associated with hereditary syndromes and neoplasms.
II. TRADITIONAL CYTOGENETIC ANALYSIS. While cytogenetic analysis is commonly used
in the evaluation of congenital disorders (specifically, to diagnose syndromes asso-
ciated with abnormalities of chromosomal number or structure, to establish the
chromosomal sex in cases of sexual ambiguity, and to screen for karyotypic abnor-
malities in patients with multiple birth defects) and for prenatal diagnosis, the
technique•s primary application in surgical pathology is in the evaluation of neo-
plastic disorders. The utility of the technique in surgical pathology rests on the fact
that specific cytogenetic abnormalities have been recognized that are closely, and
sometimes uniquely, associated with morphologically and clinically distinct sub-
sets of lymphoma and leukemia, or with soft tissue neoplasms. Cancer cytogenetic
studies have greatly aided targeted therapy, prognosis, and risk-based stratification
of intensity of therapy.
A. Advantages. The power of conventional cytogenetics lies in its ability to provide
simultaneous analysis of the entire genome without any foreknowledge of the
chromosomal regions involved in the disease process. In most cases, the type
and location of an identified chromosomal abnormality is either directly diag-
nostic or can be used to direct additional testing. Contrary to some predictions,
the advent of technologies such as array comparative genomic hybridization
(aCGH) has not diminished the importance of traditional cytogenetics; in fact,
these novel molecular techniques achieve some of their greatest utility when
they are utilized in conjunction with traditional clinical cytogenetics.
B. Limitations. The clinical utility of traditional cytogenetic analysis is restricted by
two general features of the method. From a technical standpoint, analysis can
only be performed on viable tissue specimens that contain proliferating cells
(discussed in more detail below). From a sensitivity standpoint, analysis has
resolution of only about 3 to 4Mb at an 850-band level, and only about 7 to 8
Mb at a 400-band level. Traditional cytogenetic analysis is therefore only suited
for detection of numerical abnormalities and gross structural rearrangements.
The method does not have the sensitivity to detect mutations such as small
deletions and amplifications, single base pair substitutions, and so on.
C. Basic laboratory procedures. Chromosomes that can be individually distin-
guished by light microscopy can only be obtained during cell division, and
so the fundamental requirement for traditional cytogenetic analysis is a tissue
specimen that contains actively proliferating cells, or cells that can be induced
to proliferate in vitro. The basic method for production of metaphase chromo-
somes for cytogenetic analysis is shown in Figure 58.1.
863
draw blood-
mix with
anticoagulant
Blood in culture
medium with mitogen Add colcemid to
for 72 hrs at 37"C arrest cell division
at metaphase
Add hypotonic solution

to rupture red blood
cells and swell white
blood cells
Figure 58.1 Overall scheme for the production of metaphase chromosomes for traditional cyto~
genetic analysis.
1. Culture initiation. Different specimen types have different sample and han~
dling requirements (Table 58.1). Inappropriate handling, as well as delay
between specimen collection and culture initiation, can markedly decrease
the likelihood that the sample will grow in vitro, so communication and
coordination with the cytogenetics laboratory are essential.
In vitro culture relies on a sterile microenvironment, and so specimens
should be collected under sterile conditions. In practice, sterility is most
difficult to achieve when sampling solid tissues; in this setting, clean instru~
ments and a clean cutting surface, together with transport of the specimen
in medium supplemented with broad~spectrum antibiotics, can be used to
minimize contamination.
( ii.;i:)!Jj;ll Specimen Requirements

nssuetype sample collection
Peripheral blood Preservative-free sodium heparin; transport refrigerated or at room
temperature
Bone marrow aspirate Preservative-free sodium heparin; the first several milliliters of the
aspirate usually contains the greatest proportion of cells and so is
the optimal sam pie for cytogenetic analysis; transport at room
temperature
Solid tissue Collect and transport in sterile culture medium containing
broad-spectrum antibiotics; carefully select maximally viable
tumor for analysis; transport on ice to minimize autolysis and
microbial overgrowth
Chapter 58 • Cytogenetics I 8 65
Bone marrow and solid tissue neoplasms consist of cell types that pro-
liferate spontaneously in culture, although often at a low rate. Lymph
nodes are composed of cells that have a low intrinsic proliferative rate
but that can be induced to divide much more rapidly by the addi-
tion of mitogens. Phytohemagglutinin (PHA) stimulates proliferation of
T-lymphocytes. Lipopolysaccharide (LPS), protein A, 12-0-tetradecanoly-
phorbol-13-acetate (TPA), Epstein-Barr virus, synthetic oligonucleotides,
and pokeweed mitogen induce proliferation of B-lymphocytes, and are also
required for successful culture of some leukemias and lymphomas of B-cell
origin.
2. Culture maintenance. The length of in vitro culture depends on cell type.
Since bone marrow cultures contain spontaneously proliferating cells, they
can be harvested after only a 24- to 48-hour culture interval, if not
directly after specimen collection. Peripheral blood cultures usually require a
72-hour culture interval. The growth rate of solid tissue specimens is difficult
to predict; some solid tumors require culture periods of 2 weeks or longer.
3. Cell harvest. Colcemid, a synthetic analogue of colchicine (an alkaloid from
the bulb of the Mediterranean plant Colchicum), prevents separation of
sister chromatids and is used to block the proliferating cells in metaphase,
thus allowing an accumulation of cells at metaphase stage. A hypotonic
solution is then used to swell the cells so that, after fixation, the chromosomes
are adequately spread for microscopic analysis.
Since cells in culture do not proceed through the cell cycle in synchrony,
chemical synchronization of cell division is often required to obtain an
acceptable mitotic index. A common chemical approach involves addition
of excess thymidine, which stalls cells at the S-phase of the cell cycle by
decreasing the amount of dCTP available for DNA synthesis. When the
excess thymidine is removed (or the effect of excess thymidine is elimi-
nated by the addition of deoxycytidine), normal DNA replication resumes,
and the collective release of the cells from S-phase produces a transiently
high mitotic index. Alternatively, 5-fluorodeoxyuridine (which inhibits the
enzyme thymidylate synthetase) can be used to stall cells at the G liS bound-
ary; in this method, addition of thymidine releases the block.
4. Banding. The different techniques that can be used to stain metaphase chro-
mosomes can be divided into two general categories: methods that produce
specific alternating white and dark regions (bands) along the length of each
chromosome and methods that stain only a defined region of specific chro-
mosomes (Table 58.2). In general, the dark bands are gene-poor AT-rich
regions, whereas the light bands are composed of gene-rich GC-rich regions.
The quality of staining depends on several technical factors, including suffi-
cient separation of the chromosomes in the metaphase spread to allow clear
visualization. Although there are no internationally accepted standards for
banding resolution, ideograms are used as reference points (e-Fig. 58.1).*
Many countries, including the United States, Canada, UK, France, Japan,
and Australia have established standards that specify the minimum require-
ments for the number and quality of cells that must be processed for chro-
mosome analysis depending on sample type, although many cases require
even more detailed analysis.
5. Microscopic analysis. The method used to stain the chromosomes dictates
whether bright-field microscopy or fluorescence microscopy is used to visu-
alize the chromosomes. Conventional photography has traditionally been
used to produce high-resolution prints of the stained chromosomes, but
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Cytogenetic analysis of solid tumors highlights a number of these intrin-

sic technical limitations. First, since benign solid tumors contain few mitotic
cells, cultures are susceptible to overgrowth by nonneoplastic cells. Second,
even high-grade malignant solid tumors often grow poorly in vitro, especially
if grown without the appropriate culture medium and growth factor supple-
mentation. Third, the number of neoplastic cells in a solid tumor sample can
be difficult to determine on the basis of gross examination, and the material
submitted to the cytogenetics laboratory may consist primarily of stromal cells
and inflammatory cells. Fourth, the viability of the neoplastic cells is often
uncertain; even tumor samples that are not grossly necrotic may contain pre-
dominantly nonviable tumor cells. Fifth, in vitro culture selects for subclones
within the neoplastic population that have a growth advantage, and so the
karyotype may not be representative of the entire neoplasm. Sixth, contami-
nation is often unavoidable for samples collected in the frozen section area or
gross room, or from specimens arising from anatomic sites normally colonized
by bacteria, such as the oral cavity, gastrointestinal tract, and skin.
The overall failure rate of conventional cytogenetic analysis is difficult to
quantify for many tissue types, neoplasms, and diseases, and so it is difficult to
provide objective statements regarding the utility of analysis in routine surgical
pathology. In studies that specifically address this issue for hematolymphoid
neoplasms, cytogenetic analysis has a success rate for detecting characteristic
chromosomal aberrations that varies from 33% to 100% depending on the
specific diagnosis, but is about 70% overall (Am] Clin Pathol. 2004;121:826).
For solid tumors, the success rate of analysis is less certain; reports describ-
ing cytogenetic abnormalities of many solid tumors often do not provide data
on failed analyses or negative cases. Objective measures (including sensitiv-
ity, specificity, predictive value of a positive or negative result, etc.) of tradi-
tional cytogenetic analysis as an ancillary testing methodology in routine clinical
practice are therefore often unknown.
Ill. METAPHASE FLUORESCENCE IN SITU HYBRIDIZATION (FISH). Virtually, all metaphase
chromosome in situ hybridization analysis is performed using probes that are
directly or indirectly labeled with fluorophores. Guidelines for the use of metaphase
FISH in clinical laboratory testing have been developed by the American College of
Medical Genetics (http://www.acmg.net/StaticContent/SGs/Section_E_201 0. pdf),
and standardized nomenclature for reporting results has been developed (discussed
in more detail below).
Metaphase FISH is essentially a modified Southern blot in which the target
DNA consists of chromosomes rather than membrane-bound DNA. Technically,
the method has four steps: the probe and metaphase target are denatured by a high
temperature and formamide, the probe is hybridized to the chromosomal target,
unbound probe is removed by posthybridization washes, and finally, the bound
probe is detected by fluorescence microscopy. A fluorochrome-based counterstain
is virtually always used to help detect the chromosomes during microscopic exam-
ination; the use of 4,6-diamidino-2-phenylindole (DAPI) as a counterstain makes
it possible to localize the position of the bound probe to specific chromosomal
bands.
A. Probes. A variety of fluorophores can be incorporated into metaphase FISH
probes either directly or indirectly. The choice of labels is largely governed by
practical issues, such as the excitation and emission filters on the microscope
that will be used to view the chromosome spreads.
A few probe kits have been cleared by the United States Food and Drug
Administration (FDA) for in vitro diagnostic testing, although many probes
for metaphase FISH are classified as analyte-specific reagents (ASRs) and so
are exempt from FDA approval. Standards and guidelines for clinical use of
ASRs have been established by the American College of Medical Genetics,
as have recommendations for interpretation of a metaphase FISH result (see

http://www.acmg.net/StaticContent/SGs/Section_E...2010.pdf).
1. Repetitive sequence probes. The most widely used repetitive sequence probes
bind to a-satellite sequences of centromeres; these probes produce strong
signals since a-satellite sequences are present in hundreds of thousands of
copies. Chromosome-specific centromere-specific probes have been devel-
oped for most human chromosomes on the basis of differences in a-satellite
sequences, and are particularly useful for demonstrating aneuploidy. These
FISH probes can be used on both metaphase and interphase preparations,
and simultaneous analysis of more than one locus is possible when a cocktail
of differentially labeled probes is used in the same hybridization. Other repet-
itive sequence probes include probes that recognize .B-satellite sequences
(located on the short arms of acrocentric chromosomes), and probes that
recognize the telomeric repeat sequence TTAGGG.
2. Unique sequence probes. Probes of this type are used to detect sequences
that are present only once on a given chromosome homologue. They are
usually derived from genomic clones, but can also be produced from eDNA
or by polymerase chain reaction (PCR). Different cloning vectors are used to
produce unique sequence probes of different length, including plasmids for
probes 1 to 10 kb long, bacteriophage). for probes up to 25 kb long, bacterial
artificial chromosomes (BACs) for probes up to about 300 kb long, and yeast
artificial chromosomes (YACs) for probes from 100 kb to 2Mb long. The
availability of mapped BAC libraries, originally developed as part of the
Human Genome Project, has greatly simplified the production of probes
for any locus under study (http://genome.ucsc.edu/cgi-bin/hgGateway and
http://bacpac.chori.org/).
Unique sequence probes (also known as locus-specific identifier [LSI]
probes) are used primarily to detect changes in the copy number of a spe-
cific locus, to confirm the presence of rearrangements involving a specific
locus, or to detect so-called cryptic rearrangements that cannot be identi-
fied by examination of chromosomes stained by routine banding methods
(e-Fig. 58.3). The advantages and disadvantages of metaphase FISH analy-
sis using unique sequence probes directly parallel those of interphase FISH
(as discussed in Chap. 59).
3. Whole chromosome probes (WCPs}. WCPs, also known as chromosome paint-
ing probes or chromosome libraries, consist of thousands of overlapping
probes that recognize unique and moderately repetitive sequences along
the entire length of individual chromosomes. They are isolated through
flow sorting of specific chromosomes, microdissection of specific chromo-
somes accompanied by PCR amplification, or via production of somatic
cell hybrids. WCPs are used to identify rearrangements that are not evident
by routine banding methods, to confirm the interpretation of aberrations
identified by routine banding methods, or to establish the chromosomal
origin of rearrangements that are difficult to evaluate by other approaches.
These probes are designed for use with metaphase chromosome preparations
because hybridization to the decondensed chromatin in interphase nuclei
gives a splotchy, undefined hybridization pattern. WCPs for each human
chromosome are commercially available.
IV. MULTIPLEX METAPHASE FISH. Multiplex FISH (also known as multicolor FISH) and
spectral karyotyping (SKY) are related techniques in which metaphase chromo-
some spreads are hybridized with a combination of probes labeled with different
fluorophores. Since N different fluorophores can produce (2N-1) different color
combinations, five different fluorophores yield sufficient different color combina-
tions to uniquely label WCPs so that all24 different human chromosomes can be
identified in one hybridization (1-22 autosomes, X andY chromosomes).
Chapter 58 • Cytogenetics I 869
For both multiplex FISH and SKY, a cocktail consisting of labeled probes for
each of the 24 chromosomes is hybridized to metaphase chromosome spreads,
and the fluorescent emissions are measured by computerized imaging systems.
Specialized software is used to determine the combination of fluorophores present
along the length of each chromosome, which makes it possible to assemble a
karyotype.
A. Advantages. Multiplex FISH and SKY are used to detect aneuploidy, detect
interchromosomal rearrangements, and identify marker chromosomes (extra-
chromsomal material of unknown origin). In many cases, multiplex-FISH or
SKY make it possible to establish the chromosomal origin of rearrangements
that cannot be defined on the basis of routine cytogenetic analysis. A web-
based database has been developed to facilitate identification of chromosomal
aberrations detected by multiplex FISH (http://www.ncbi.nlm.nih.gov/sky/), a
database that contains links to other websites that can be used to integrate the
cytogenetic map with physical and sequence maps.
B. Disadvantages. The lower limit of the size of individual DNA chromosomal frag-
ments that can be visualized by either technique is in the range of 1 to 2Mb,
although neither technique provides direct information on the involved chro-
mosomal bands. Similarly, multiplex FISH and SKY will only reveal intrachro-
mosomal deletions and duplications that are large enough to result in a change
in size of the affected chromosome; neither technique is designed to detect intra-
chromosomal rearrangements such as inversions, and neither is informative in
regions with repetitive DNA.
C. Modifications of multiplex FISH and SKY. Mixtures of so-called partial chromo-
some paints, each of which hybridizes to only a band or subband of an indi-
vidual chromosome, can be used to produce a pseudocolor-banded karyotype
at a resolution of about 550 bands (Cytogenet Cell Genet. 84:156, 1999). The
use of partial chromosome paints makes it possible to employ multiplex FISH
and SKY methodology to identify translocation breakpoints and to detect inter-
chromosomal rearrangements.
V. COMPARATIVE GENOMIC HYBRIDIZATION (CGH). While CGH often has a higher sen-
sitivity than conventional cytogenetic analysis, of even greater significance is the
fact that CGH can be performed using DNA extracted from fixed as well as fresh
tumor samples. The technique therefore makes it possible to perform a genome-
wide scan for structural alterations even on those cases for which conventional
cytogenetic analysis is not feasible or is unsuccessful. CGH essentially opens the
entire formalin-fixed tissue archive to at least limited cytogenetic analysis.
For a typical CGH test, genomic DNA from a tumor sample is labeled with a
red fluorophore, and genomic DNA from a paired normal tissue sample is labeled
with a green fluorophore. The green and red probes are mixed and used in a single
hybridization.
A. Metaphase CGH. This technique is basically a variation of metaphase FISH used
to survey the entire genome for chromosomal deletions and amplifications (Sci-
ence. 1992;258:818; Trends Genet. 1997;13:405). The labeled probe mixture
is used in a hybridization to metaphase chromosomes prepared from normal
cells, and the ratio of the green to red fluorescent signals is measured along the
length of each chromosome. Areas where the ratio deviates significantly from
the expected one-to-one relationship indicate a change in DNA copy number in
the tumor; areas where the red to green ratio is significantly> 1 are areas of chro-
mosomal gain (usually amplifications), and areas where the red to green ratio is
significantly <1 are areas of chromosomal loss (deletions). The smallest chro-
mosomal alterations that can be reproducibly detected are about 3Mb long.
B. Array CGH (aCGH). This approach utilizes a microarray consisting of an ordered
arrangement of DNA molecules (features) linked to a solid matrix support. The
labeled probe mixture is hybridized to the micro array, and the ratio of the green
Patient's DNA labeled Control DNA labeled

with Cy5 with Cy3
000000
000000
Cy5/Cy3 ratio>1 eooooo Cy5/Cy3 ratio< 1

Duplication 000000 Deletion
~··,<;,~<W &fl:ffi],ruJ
Figure 58.2 Array CGH methodology. Top: Method. A patienfs DNA is labeled
with a red dye and a control genomic DNA preparation is labeled with green
dye. The DNA preparations are mixed and co hybridized to an array of BACs or
oligonucleotides on a glass slide. The DNA bound to each spot (known as a fea-
ture) of the array is quantified using a laser scanner. In the patient's DNA, normal
regions will be indicated by a yellow balanced color; regions of duplication will be
identified as red, and regions of deletion will be identified as green. Bottom: Data
presentation. The data from each feature of the array (represented by circles) are
plotted in relation to the features' positions along the chromosome and a balanced
copy number (horizontal line). In this illustratiofit a duster of adjacent features
that falls significandy below a balanced copy number result (oval) indicates the
presence of a deletion. The resolution of array CGH is in theory limited only
by the number of features in the array; commercially available arrays currently
provide a resolution of <10 kb. (Adapted from Beaudet et al. Annu Rev Med.
2008;59:113.)
to red fluorescent signals is measured for each feature (Fig. 58.2). Because each
DNA feature has been mapped to a specific region of the genome, the ratio of
the green to red fluorescent signal for each feature provides information on the
gain or loss of the corresponding chromosomal region. The resolution of aCGH
is in theory limited only by the number of features in the array; commercially
available arrays currently provide a resolution of< 10 kb. Genomic microarrays
(aCGH and related microarray-based methods) are currently clinically applied
to detect genomic copy number changes as well as copy neutral changes (uni-
parental disomy [UPD]Iloss of heterozygosity [LOH]).
VI. MICROARRAY ANALYSIS. Since the advent of the use of genomic microarrays in the
clinical laboratory, the technology has rapidly become the standard of care to eval-
uate patients for genomic imbalance, especially in diagnostic testing for patients
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duplication, and more generally, consanguinity and UPD. Ultimately, SNP
detection helps maximize the potential for detecting disease-associated abnor-
malities and also offers mechanistic evidence for the molecular basis of the
disease. A comparison of SNP and oligoarrays is shown in Table 58.4.
The technique of allelic discrimination by SNP analysis differs between
commercially available platforms, but nonetheless involves hybridization of
fragmented single-stranded DNA (derived from the patient sample) to arrays
that contain unique nucleotide probe sequences (two million or more). One
widespread commercial approach is based on allele-specific hybridization to
probes representing the possible alleles; signal intensities that correspond to
the level of binding (Fig. 58.3) are measured by scanning technology. The other
Affymetrix I!lumina
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Figure 58.3 SNP array procedures. In the Affymetri:x platform (Affymetrix; Santa Clara, CA),
genomic DNA is digesred with the Nspl restriction enzyme and the resulred DNA fragments
are ligated to adaptors and subsequently amplified; the amplification products are fragmented,
end-labeled, and hybridized to the array. In the Illumina platform (IDumina, Inc., San Diego,
CA), the entire genome is amplified and then hybridized to a bead array; allelic discrimination is
achieved by a single base extension reaction. In both platforms, the probe inrensity is measured and
compared with an in silico reference to evaluate DNA copy number. (Adapred from Schoumans
and Ruivenkamp. Methods Mol Bioi. 2010;628:53.)
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(a summary of the more common symbols and abbreviations is shown in Table

58.5). For metaphase chromosome in situ hybridization, the results of con-
ventional cytogenetic analysis (if performed) are listed first, followed by the
results of in situ hybridization analysis. Ideally, loci are designated according
to the HUGO Gene Nomenclature Committee (http://www.genenames.org/);
when HUGO designations are unavailable, probe names are used.
SUGGESTED READINGS
Beaudet AL, Belmont ]w. Array-based DNA diagnostics: let the revolution begin. Annu Rev Med.
2008;59: 113-129.
Gersen SL, Keagle MB, eds. The Principles of Clinical Cytogenetics. 2nd ed. Totowa: Humana
Press; 2005.
Kallioniemi OP, Kallioniemi A, Sudar D, et al. Comparative genomic hybridization: a rapid
new method for detecting and mapping DNA amplification in tumors. Semin Cancer Bioi.
1993;4:41-46.
LaFramboise T. Single nucleotide polymorphism arrays: a decade of biological, computational and
technological advances. Nucleic Acids Res. 2009;37:4181-193.
Miller DT, Adam .MP, Aradhya S, et al. Consensus statement: chromosomal microarray is a first-tier
clinical diagnostic test for individuals with developmental disabilities or congenital anomalies.
Am] Hum Genet. 2010;86:749-764.
Rooney DE, ed. Human Cytogenetics: Constitutional Analysis: A Praaical Approach. 3rd ed.
Oxford: Oxford University Press; 2001.
Schoumans J, Ruivenkamp C. Laboratory methods for the detection of chromosomal abnormalities.
Methods Mol Bioi. 2010;628:53-73.
Shaffer LG Slovak ML, Campbell LJ, eds. ISCN 2009: An International System for Human Cyto-
genetic Nomenclature. Basel: S Karger; 2009.
Fluorescence in Situ
Hybridization
Tu Dong Nguyen, Arie Perry, and Anj um Hassan
Fluorescence in situ hybridization (FISH) utilizes tagged probes that bind to

chromosome-specific DNA sequences of interest, thereby allowing for the identification
of both structural and numeric aberrations characteristic of certain hematopoietic and
nonhematopoietic malignancies. While FISH can be performed on dividing (metaphase)
cells, it has several major advantages over conventional cytogenetics in that it can be
applied in many clinical settings (Table 59.1 ), can be performed on nondividing (inter-
phase) cells, can be performed on air-dried or formalin fixed specimens, can facilitate
detection of molecular abnormalities in neoplasms with low proliferation rates such
as multiple myeloma, and can facilitate detection of numeric abnormalities. In sur-
gical pathology, the technique is used primarily to detect somatic cancer-associated
alterations with known diagnostic, prognostic, or therapeutic implications.
FISH provides insight into intranuclear target DNA localization and copy num-
ber. Therefore, using locus-specific probes (with the exception of XY sex chromosome
determinations in males), two signals per nucleus are expected and so four common
alterations are readily detectable: aneusomy (gain or loss of a chromosome), gene
deletion, gene amplification, and translocation (e-Figs. 59.1 through 59.4).* Sex chro-
mosome determinations can be useful in patients with sex mismatched bone marrow
or organ transplants (e-Fig. 59.5), in order to monitor engraftment success or failure.
I. ADVANTAGES AND LIMITATIONS OF FISH

A. Specimens, retained morphology, and combined FISH/immunohistochemistry. FISH
is applicable to a variety of specimen types, including fresh or frozen tissue, cyto-
logic preparations, and formalin-fixed paraffin-embedded (FFPE) tissue (Table
59.2). The latter provides a particularly rich source of archival material.
In clinical diagnostic testing, morphologic preservation is one of the principal
advantages of FISH, particularly for studies on heterogeneous tissue samples
in that it eliminates the need for microdissection (e-Fig. 59.6). An extension
of this morphologic advantage comes from the possibility of combining FISH
with immunohistochemistry, wherein separate assessments can be rendered in
immunopositive and immunonegative cellular populations.
B. FISH versus other cytogenetic and molecular techniques. When compared with
metaphase cytogenetics (see Chap. 58), interphase FISH has several clear advan-
tages. One advantage is the lack of a requirement for mitotically active cells via
cell culture, which removes potential artifacts due to in vitro growth selection
biases such as overgrowth of nonneoplastic stromal elements. On the other hand,
FISH is not a genomic screening tool; it provides a targeted approach for alter-
ations that have been initially identified by more global molecular techniques,
such as classic cytogenetics, loss of heterozygosity (LOH) screening, compara-
tive genomic hybridization (CGH), array CGH (aCGH), array single nucleotide
polymorphism (SNP) analysis, and gene expression profiling.
In terms of resolution, FISH is more sensitive than conventional karyotypic
analysis and CGH (both of which are limited to alterations of several Mb in size)
876
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to the traditional whole slide approaches (Adv Anat Pathol. 2001;8:14), and
although complications due to tumor heterogeneity can be problematic, ade-
quate sampling can be optimized by incorporating multiple cores from each
specimen. TMA-FISH is an excellent method for new probe validation, profi-
ciency testing, interlaboratory comparisons, and quality assurance/quality con-
trol(] Histochem Cytochem. 2004;52:501).
D. Disadvantages and pitfalls of FISH. Although recent technical advances have
greatly enhanced the clinical applicability of FISH, a number of limitations
remain. Signal fading is one of the main disadvantages. Clinical labs typically
circumvent this pitfall by capturing digital images as a permanent record of
each case; a permanent record is not otherwise possible unless chromogenic
detection (CISH) is used. Unfortunately, multicolor CISH is not as simple as
multicolor FISH; currently available chromogens lack the spectral versatility,
sensitivity, and spatial resolution attainable with fluorochromes. Some com-
mercial CISH applications bypass this problem by providing the test and ref-
erence probes separately, so that in place of one dual-color FISH assay, two
single-color CISH experiments are performed. Recently developed photostable
quantum dots offer a potential alternative for permanent fluorescent signals
(] Histochem Cytochem. 2003;51:981). Other limitations include a variety of
artifacts, particularly common in paraffin sections, that make correct interpre-
tation of FISH results dependent on significant experience.
1. Truncation artifact. This artifact is due to the underestimation of copy number
because of an incomplete DNA complement within transected nuclei, and it
is therefore important to assess controls cut at the same thickness.
2. Aneuploidy and polyploidy. Artifacts due to aneuploidy and polyploidy can
result in confusing signal counts and are a particularly common finding in
malignant and even some benign neoplasms. Although the simplest approach
is to interpret absolute losses ( <2 copies) and gains (>2 copies), "relative"
losses and gains can also be delineated on the basis of a reference ploidy,
obtained either by flow cytometry or the assessment of multiple chromosomes
by FISH. For example, cells with four chromosomes, nine centromeres, and
two copies of the p16 gene region on 9p21 would be interpreted as having
polysomy 9 and a hemizygous p16 deletion (e-Fig. 59.7A); a similar tumor
with no p 16 signals would be interpreted as polysomy 9 with homozygous
p16 deletion (e-Fig. 59.7B).
3. Autofluorescence. This is a particularly common problem in FFPE tissue sec-
tions. Although autofluorescent tissue fragments are usually larger and more
irregular than true signals, some fragments have just the right size to mimic
true nuclear signals. The use of multiple filters is helpful, since autofluores-
cence will often appear at several wavelengths of light, whereas true signals
only fluoresce at one wavelength.
4. Partial hybridization failure. This issue is most problematic when combining a
highly robust probe (e.g., centromere) with a comparatively weak probe (e.g.,
small locus specific probe). This artifact can be minimized by counting only
in regions where the majority of cells have discernible signals. Signals from
both probes should be seen in normal cells (e.g., endothelial cells) within the
region for the counts to be considered reliable.
E. Additional technical considerations. Many different FISH protocols are available;
they vary depending on individual preferences and specimen type. Simple pro-
tocols are generally better, requiring less "hands on" time, fewer opportunities
for error, and fewer troubleshooting requirements. Automated instruments are
now available to minimize hands on time, though they are expensive. In general,
the basic steps of the protocols are similar to those of immunohistochemistry
and include deparaffinization, pretreatment/target retrieval, probe and target
Chapter 59 • Fluorescence in Situ Hybridization I 879
DNA denaturation, hybridization (a few hours to overnight), posthybridization

washes, detection, and microscopic interpretation/imaging. FISH is therefore
typically a 2-day assay, although same day assays are possible if the probes are
particularly robust (e.g., centromere probes).
Similar to immunohistochemistry, microwave or heat-induced target
retrieval often enhances hybridization more effectively than chemical forms of
pretreatment (Anal Cell Pathol. 1994;6:319). Nonetheless, optimal pretreat-
ment and digestion varies from specimen to specimen and depends on a number
of variables, including method of fixation and processing. Some hybridization
buffers are also significantly more efficient and may lower probe concentration
requirements considerably, which can be particularly beneficial with expensive
commercial probes. Lastly, a variety of amplification steps are available for
enhancing weak signals, although such steps are rarely necessary with robust
commercial probes. One exciting advancement made possible by high-level sig-
nal amplification techniques is the potential use of smaller probes, down to the
level of 1 kb or less (Biotechniques. 1999;27:608).
II. FISH PROBES AND PROBE DEVELOPMENT
A. Centromere enumerating probes (CEPs) were among the first types of probes devel-
oped and remain ideal for detecting whole chromosome gains and losses, such as
monosomy, trisomy, and other polysomies. CEPs target highly repetitive 171 bp
sequences of a-satellite DNA, and so are associated with excellent hybridization
efficiencies and typically produce large, bright signals. Unfortunately, sequence
similarities in some pericentromeric regions result in cross-hybridization arti-
facts with the potential for overestimating signal counts. Another artifact is
caused by the interesting phenomenon observed in nonneoplastic brain speci-
mens in which certain chromosomes in interphase nuclei are packaged such that
paired centromeres are in close proximity, a process known as somatic pairing
(Hum Genet. 1989;83:231; Cytogenet Cell Genet. 1991;56:214); because of the
close proximity, FISH yields an unexpected fraction of cells harboring a single
large signal rather than two smaller ones, potentially leading to overinterpreta-
tion of monosomy. Despite these technical limitations, CEPs remain extremely
useful for detecting aneusomies and are still among the best FISH probes avail-
able. The presence of repetitive DNA sequences in subtelomeric regions has led
to the development of commercially available probes for each chromosomal
arm as well.
B. A whole chromosome paint (WCP) probe consists of a cocktail of DNA frag-
ments that targets all the nonrepetitive DNA sequences of an entire chromo-
some. Because a WCP covers such a large region, it produces a diffuse signal in
interphase nuclei (although some of the smaller acrocentric chromosomes yield
sufficiently discrete signals for enumeration, even in interphase nuclei). For this
reason, WCPs are not often used in interphase FISH, but instead are primarily
utilized in advanced cytogenetic applications (see Chap. 58).
C. Currently, the most versatile FISH probes are locus-specific identifier probes
(also known as LSI, or gene-specific, probes). These probes target distinct chro-
mosomal regions of interest and utilize single copy rather than repetitive DNA
sequences. In order to yield signals of sufficient size in interphase FFPE nuclei, the
probe typically needs to be at least 20 kb long; the largest LSI probes are > 1 Mb
long, although most fall into the 100 to 300 kb range. The assortment of LSI
probes available commercially has expanded greatly over the last few years.
Additionally, cloning vectors, such as cosmids, bacterial artificial chromosomes
(BACs), P1 artificial chromosomes (PACs), and yeast artificial chromosomes
(YACs) are excellent sources for developing analyte specific (i.e., homemade)
FISH probes. In the past, development of LSis required a rather lengthy and
tedious process of screening vector libraries, but the BAC libraries generated
as part of the human genome project have made it possible to rapidly iden-
tify vectors that contain sequences of interest, gene names, or physical maps
of individual chromosomes (http://www.genome.ucsc.edu). Similarly, mapped
BAC clones spread throughout the human genome at 1-Mb intervals have
also become available (http://mp.invitrogen.com). However, regardless of how
a probe is obtained, it is important to verify its identity, either by screen-
ing for the DNA sequence of interest by PCR or by performing metaphase
FISH to determine that the probe localizes to the appropriate cytogenetic band
(e-Fig. 59.8).
Ill. CLINICAL APPLICATIONS. FISH testing is clinically useful when a cytogenetic alter-
ation (deletion, gain, amplification, and translocation) is sensitive and specific for
a single tumor type, either as a diagnostic biomarker (as in many hematopoi-
etic malignancies) or as a prognostic biomarker helping to predict which tumors
will be aggressive or indolent (HER-2/neu amplification testing in breast cancer).
Some translocation and deletions detected by FISH are also helpful in predicting
response to a specific therapy; examples include detection of t(11;18) in a sub-
set of mucosa-associated lymphoid tissue (MALT) lymphomas (which tend to be
resistant to conventional therapy), detection of FIP1L1-PDGFRA fusion formed
as a cryptic deletion at 4q12 in chronic eosinophilic leukemia which is sensitive
to imatinib therapy, and detection of del(13q) and/or t(4;14) in a subset of cases
of multiple myeloma (which tend to have a worse prognosis). The most common
clinical applications of FISH testing currently include HER-2/neu amplification
testing for breast cancer, UroVysion testing in urine cytology specimens, lp/19q
deletion testing in gliomas, and testing for signature translocations associated with
specific hematologic, soft tissue, and/or pediatric malignancies. Clinically relevant
examples for each alteration type detectable by FISH are summarized in Table 59.3.
A. Aneusomies and deletions. Aneusomies represent gains and losses of whole chro-
mosomes. Deletions are losses of distinct chromosomal regions, varying in size
from loss of a specific gene or portion of a gene to an entire chromosomal arm.
Aneusomies and deletions are amongst the most common alterations detected in
neoplasms by FISH (Table 59.3), although it is sometimes difficult to distinguish
specific tumor-associated polysomies and monosomies from nonspecific gains
and losses that are secondary changes due to the genomic instability that is char-
acteristic of many malignancies. The use of reference probes helps to distinguish
such chromosomal gains or high-level polysomies from true gene amplification
(e-Fig. 59.3).
One of the most common FISH applications is testing for deletions of 1 p
and 19q in diffuse gliomas. The presence of lp and 19q codeletion (typically,
loss of the entire arm of each) has diagnostic, prognostic, and predictive value in
that this genetic signature (e-Fig. 59.2A and B) is associated predominantly with
pure oligodendrogliomas with enhanced therapeutic responsiveness and overall
survival time(/ Neuropathol Exp Neurol. 2003;62:111). There is no current
consensus for the optimal way to enumerate signals for chromosomal losses and
gains, though in clinical cases a common approach is to use two individuals who
count signals in 100 cells each; when the counts are concordant, they are simply
added for a total enumeration of 200 cells, but when there is a discrepancy or the
counts are borderline for an alteration, then either the same individuals count
additional cells or a third enumerator is utilized. Despite the clinical utility of 1 p
and 19q testing, the precise gene targets on these chromosomes remain unclear.
In other tumor types, a specific tumor suppressor is known to be targeted
by various chromosomal deletions (Table 59.3). Notable examples include the
INI1/hSNF5 gene on 22q11.2 in malignant rhabdoid tumors and atypical ter-
atoid/rhabdoid tumors, the NF2 gene at 22q12 in meningiomas, the RB1 gene
at 13q14, the TP 53 gene at 17p13.1 in multiple myeloma, and the NF1 gene at
17q11.2 in malignant peripheral nerve sheath tumors.
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In terms of tumor-specific chromosomal gains and losses, Vysis {http://www.

vysis.com) markets a number of multicolor probe cocktails for clinical and
translational studies, each with different recommendations for minimum num-
ber of nuclei counted and cutoffs for alterations. For example, the UroVys-
ion {e-Fig. 59.9) and LAVysion probe sets have been shown to increase diag-
nostic sensitivities in body fluid cytology specimens for urothelial carcinoma
(J Urol. 2003;169:2101) and lung carcinoma (Am] ClinPathol. 2005;123:516),
respectively. Similarly, the CLL and ProVysion FISH assays have been shown to
identify prognostically relevant subsets of chronic lymphocytic leukemia {Can-
cer Gen Cytogenet. 2005;158:88) and prostatic adenocarcinoma (Genes Chrom
Cancer. 2002;34:363) patients, respectively. Likewise, the detection of specific
aneusomies and deletions by FISH has been clinically useful in identifying diag-
nostically challenging cases of glioblastoma (particularly the small cell variant),
medulloblastoma (especially the anaplastidlarge cell variant), and prognosti-
cally relevant subsets of leukemialmyelodysplastic syndrome.
B. Gene amplifications. High level gene amplifications typically occur in one of
two patterns. If the amplified gene is present on small extra-chromosomal seg-
ments known as double minutes , FISH will show numerous individual signals
(e-Fig. 59.3A and C). If the gene amplification consists of contiguously arranged
gene copies within a single chromosomal region manifested as a homogenously
stained region on chromosomal banding, FISH will show regions of hybridiza-
tion so dose together that they coalesce into abnormally large linear or globular
signals (e-Fig. 59.3B); rough estimates are made regarding how many signals
are contained within the coalescent signals on the basis of their overall size. In
both settings, CEP probes are often used as references for chromosome number
in order to distinguish polysomies (i.e., gains of the entire chromosome; e-Fig.
59.3D) from true gene amplification.
Of the current clinical applications of FISH in surgical pathology, the assess-
ment of HER-2/neu amplification status in breast cancer is one of the most
common applications (Cancer. 2003;98:2547). Although there is agreement that
HER-2/neu assessment provides clinically useful information, the optimal diag-
nostic approach is still much debated. Recently, the American Society of Clin-
ical Oncology and College of American Pathologist recommended guidelines
for testing of HER-2/neu amplification in breast cancer (Arch Pathol Lab Med.
2007;131:18). HER-2/neugene amplification is presentin20% to 35% of breast
carcinomas (e-Figs. 59.3C and 59.10) and provides both prognostic and predic-
tive information with the following associations for positive tumors: reduced
patient survival, especially in lymph node positive cases; increased responsive-
ness to Adriamycin-based therapeutic regimens; increased responsiveness to
Herceptin (trastuzumab), which specifically targets the overexpressed surface
protein; and decreased responsiveness to radiation therapy, cyclophosphamide,
methotrexate, 5-FU, hormonal therapy, and Taxol. Additionally, due to the sig-
nificant risk of cardiotoxicity with combined Adriamycin and Herceptin therapy,
testing is justified to identify patients with a low probability of response. New
methods using chromogenic in situ hybridization methods for HER-2/neu gene
status assessment in breast cancer are also being explored (Am] Clin Pathol.
2009;131:490). Since trastuzumab-based therapies have also been shown to be
beneficial in HER-2/neu, positive patients with advanced gastric cancers, guide-
lines for testing for HER-2/neu amplification are also being developed for gastric
carcinomas (Virchows Arch. 2010;457:299; Adv Anat Pathol. 2011;18:53).
In pediatric pathology, testing of neuroblastomas for MYCN amplification
has similarly become standard of care, with positive cases typically showing
an aggressive biology (] Pathol 2002;198:83). A similar pattern is encoun-
tered in a subset of the CNS counterpart medulloblastoma; MYCN and CMYC
Chapter 59 • Fluorescence in Situ Hybridization I 8 83
amplifications are particularly common in the highly aggressive anaplastic/large

cell variant.
EGFR amplification, often in combination with monosomy 10 or chromo-
some 10q deletion, helps to distinguish the clinically aggressive and therapeu-
tically refractory small cell variant of glioblastoma from the more biologically
favorable and chemotherapeutically responsive look-alike anaplastic oligoden-
droglioma.
C. Translocations. The list of known tumor-associated chromosomal translocations
is already extensive and continues to grow. Translocation analysis by FISH is par-
ticularly useful as an ancillary diagnostic aid in primitive-appearing hematopoi-
etic and soft tissue malignancies (Table 59.3 ). Chromosomal translocation anal-
yses are unique among FISH assays in that interpretation relies on the spatial
relationships of the signals rather than their number. For optimal probe design,
detailed knowledge of the breakpoints is needed, though reliable probes are
commercially available for most of the common translocations.
1. Fusion FISH. Also known as FISH-F, this strategy employs two locus specific
probes with different fluors (color tags), targeting two different partners in
a translocation (e.g., BCR on 22q and ABL on 9q). Separated or "split"
signals (e.g., two green, two red signals; e-Fig. 59.4A) are therefore present
in translocation-negative cells, but "fusion" yellow or red-green signals are
present in translocation-positive cells due to the juxtaposition of the target
loci by the translocation (e.g., one fusion, one green, one red signal; e-Fig.
59.4B). FISH-F results must be scored carefully to avoid overinterpreting
small cellular populations where green and red signals overlap purely by
chance. On the basis of signal proximities in normal controls, typical con-
servative cutoffs for positive test results require the presence of fused signals
in >30% cells for FISH-F (Mod Pathol. 2006;19:1).
2. Break apart FISH. Also known as FISH-BA, this strategy utilizes two probes
localizing just proximal and distal to one of the two breakpoints of interest.
The two probes are therefore in close proximity to one another in normal
cells, resulting in fusion signals (e.g., two fusion signals; e-Fig. 59.4C). Cells
harboring the translocation will contain at least one pair of split signals (e.g.,
one fusion, one green, one red signal; e-Fig. 59.4D). The advantages ofFISH-
BA are that split signals don•t occur purely by chance, the test yields a positive
result even when the translocation can involve multiple partner genes (e.g.,
C-MYC at chromosome 8q24 can fuse with multiple partner genes in Burkitt
lymphomas including JGH at 14q32, or less commonly the light chain loci
on 2q11 or 22q11), and commercial break apart probes yield large easily
interpretable signals (e-Fig. 59.11 through e-Fig. 59.14). Disadvantages of
FISH-BA include that it provides no information regarding the identity of
the fusion partner. On the basis of signal proximities in normal controls,
typical conservative cutoffs for positive test results require the presence of
split signals in> 15% of cells for FISH-BA (Mod Pathol. 2006;19:1).
3. Other approaches. A strategy that has been used to increase the sensitivity of
FISH-F is to include one particularly large FISH probe that spans a break-
point region. In the presence of a translocation, the large DNA probe is
split, leading to an extra signal (hence the name FISH-ES) smaller than the
remaining nonsplit, nonfused signals (e.g., one fusion, one normal green, one
normal red, and one ES red signal). Since it is unlikely that individual cells
will contain both a fusion signal and an extra signal, smaller populations of
tumor cells can be confidently identified in heterogeneous specimens (e.g.,
minimal residual disease in CML).
An even more reliable method to increase the sensitivity of FISH-F is to
use two large probes that span both breakpoint regions. By this approach, a
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Direct and Indirect
Methods for DNA
Sequence Analysis
TuDong Nguyen, Barbara Zehnbauer,
and John D. Pfeifer
I. INTRODUCTION. Clinical molecular diagnostic methods have been integrated into

many laboratory disciplines, and guidelines and recommendations from both pro-
fessional societies and regulatory agencies have been developed to assist in the devel-
opment and performance of clinical molecular pathology testing (Table 60.1 ). Most
molecular tests performed in surgical pathology focus on somatic or acquired DNA
variations in the cells of the disease process that provide information that aids in
diagnosis, identifies prognostic indicators, stratifies patients into effective treatment
options, helps monitor treatment response, and/or identifies patients at increased
risk of disease. Because polymerase chain reaction (PCR)-based approaches are
quick, reliable, and sensitive, PCR has become a central technology for much of
clinical molecular genetic testing.
II. SPECIMEN REQUIREMENTS, HANDLING, AND PROCESSING. Clinical PCR-based molec-
ular testing in the setting of surgical pathology requires the same attention to detail
regarding efficient specimen collection, identification, preparation, and routing as
any other pathology test.
A. Specimens. Specimen requirements are dictated by the disease process, including
the type of tissue, amount of tissue, type of sample (fresh or frozen tissue,
formalin-fixed paraffin-embedded tissue [FFPE], cytology specimen, and so on),
and the extent of the disease in the sample. The amount of tissue required for
PCR-based testing is relatively small, which contributes to the clinical utility of
the technique.
Regardless of specimen type, two general features of the tissue sample influ-
ence molecular assays. First, there must be a sufficient quantity of the specific
target cell (and therefore target DNA or RNA) in the sample. Second, the size
or integrity of the nucleic acid molecules after isolation from the tissue can
dramatically affect the sensitivity of the detection of specific alterations, thus
nucleic acid degradation (whether due to fixation, or enzymatic, heat, pH, or
mechanical forces) can reduce the sensitivity of testing.
1. Tissue type. Fresh peripheral blood, bone marrow, solid tissue biopsies, cytol-
ogy specimens, enriched cell populations (e.g., from flow cytometry), and
FFPE tissue sections are all sources of nucleic acids for molecular analysis.
Specimens should be collected and transported to the molecular pathology
laboratory using aseptic techniques, if possible. Transport on ice reduces cell
lysis, minimizes nuclease activity, and reduces nucleic acid degradation.
2. Tissue quality
a. Fresh tissue and cell suspensions are the optimal templates for PCR.
The preferred method of preservation of fresh tissue prior to isolation
of nucleic acids is ultra-low-temperature frozen storage at -70°C, which
permits indefinite preservation with virtually no effect on the quality of the
extracted nucleic acids. Low-temperature frozen storage around -20°C
can adequately preserve DNA and RNA for several months.
Cell suspensions (including hematologic specimens such as periph-
eral blood and bone marrow) should be collected in the presence of an
887
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Chapter 60 • Direct and Indirect Methods for DNA Sequence Analysis I 889
anticoagulant, preferably ethylenediaminetetraacetic acid (EDTA) or acid

citrate dextrose (ACD); heparin should be avoided because heparin carry-
over after nucleic acid isolation may inhibit subsequent PCR steps. Freez-
ing of hematologic specimens presents distinct obstacles to the preparation
of good quality nucleic acid and should generally be avoided.
b. Nucleic acids extracted from fixed tissue can also be used in PCR, although
the type of fixative and length of fixation both have a profound effect
on their recovery. Non-cross-linking fixatives such as ethanol provide
the most consistent preservation of amplifiable DNA, with more vari-
ability from tissues fixed with formalin, Zamboni's and Clark's fixatives,
paraformaldehyde, and formalin-alcohol-acetic acid. Tissues processed
in Carnoy's, Zenker's, Bouin's and B-5 fixatives are poor substrates for
PCR testing since little amplifiable DNA can be recovered from them.
The effects of formalin fixation on PCR-based testing have been evalu-
ated in some detail, not surprising given that most surgical specimens are
fixed in formalin. Formaldehyde reacts with nucleic acids and proteins
to form a mixture of end products that are covalently linked by methy-
lene bridges. Thus, the quality of DNA isolated from formalin-fixed tissue
is critically dependent on the length of fixation, with a deterioration of
PCR signal with increasing fixation time. In general, tissue fixed in neu-
tral buffered formalin for <8 hours contains DNA and RNA from which
PCR products > 600 bp in length can be reliably amplified, but fixation
extended for greater than 8 to 12 hours decreases the length of PCR prod-
ucts that can consistently be amplified.
3. Tissue quantity. Minimum sample requirements are determined by the assay
methodology and extent of target cell involvement in the tissue. A typical
PCR-based assay requires only 20 to 200 ng of DNA (about 103 to 104
cells), although multiplexed PCR may require a bit more DNA in order to
equally represent all targets. The sensitivity of PCR for detection of a few
tarr,et molecules in a large background of unaltered DNA molecules (1 in
10 ) is one of the principle strengths of the methodology.
Ill. ANALYTIC/TECHNICAL VERSUS DIAGNOSTIC AND OPERATIONAL ASPECTS OF TESTING.
The familiar probabilistic model used to define the likelihood that a particular
patient is correctly classified on the basis of a test result (Table 60.2) can be applied
to molecular genetic tests as with any other laboratory test. However, it is important
to recognize that the quantitative performance of a lab test can be evaluated at four
different levels (Table 60.3 ), and that test performance at one of the four levels does
not necessarily predict performance at the other levels. Differences between these
four levels of analysis are often overlooked even though they account for many
of the confusing or seemingly conflicting results regarding the utility of molecular
genetic testing in surgical pathology.
IV. BASIC PCR METHODOLOGY
A. Amplification. Selective amplification of the target sequence is achieved through
the use of oligonucleotide primers that hybridize to the 5' and 3' ends ofthe DNA
target sequence (Fig. 60.1). In addition to the two primers and input (template)
DNA, the reaction mixture also includes the four deoxynucleotide triphosphates
(dATP, dCTP, dGTP, dTIP) and a heat-stable (thermostable) DNA polymerase.
The first step of the PCR itself involves heating the mixture to a high temperature
to denature the target DNA; in the second step, the reaction is cooled to allow
the primers to anneal to their complementary sequence in the target DNA; in
the third step, the reaction is heated to the temperature at which the heat-stable
DNA polymerase has optimal activity. As a result of this three step denaturation,
annealing, and polymerization cycle, the two primers will initiate synthesis of
new DNA molecules from opposite strands of the input DNA heteroduplex.
With each repetition of the three-step cycle, the newly synthesized DNA strands
110 I SECTION X.lllt ANCILLARY t.IETHOD5
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Chapter 60 • Direct and Indirect Methods for DNA Sequence Analysis I 8 91
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7
Figure 60.1 Schematic diagram of PCR. Each cycle consists of three sreps: The reaction mix
is heated to denature the double-stranded DNA template, the reaction mix is cooled to permit
annealing of oligonucleotide primers to sequences that flank the targetregioD.t and then the reaction
mix is warmed to permit the heat~stable polymerase to synthesize new DNA strands. Each newly
synthesized DNA strand then acts as a template in subsequent three~step cycles of denaturation,
annealing, and DNA synthesis, producing exponential amplification of the target region.
b. PCR has hi&h sensitivity and specificity. When optimized, PCR can detect
one abnormal cell in a background of 105 normal cells, and can even be
used to analyze single copy genes from individual cells (Methods Enzymol.
2002;356:295, 334). PCR can also be used to detect a broad range of
generic abnormalities ranging from gross structural alterations such as
translocations to single base-pair changes.
c. PCR products are easily labeled for detection. For primer-mediated labeling,
a labeled chemical group (usually a fluorophore) is attached to the 5' end of
either or both oligonucleotide primers. Alternatively, the PCR product can
be directly labeled by including one or more labeled nucleotide precursors
into the PCR mix.
d. Phenotype-genotype correlations are possible. When performed on tissue
sections, PCR provides only an indirect correlation of morphology with
underlying genetic abnormalities. Microdissection, in which the region of
interest is carved out of the FFPE tissue block, scraped from tissue sections
or cytology slides, or collected more precisely with a micromanipulator
apparatus, provides some enrichment for morphologic-genetic correla-
tions. More precise phenotypic-genotypic analysis is achieved by collect-
ing individual cells by laser capture microdissection, by flow cytometry, or
even by immunomagnetic methods. In situ PCR performed on histologic
tissue sections themselves is perhaps the ultimate method for providing
morphologic localization of genotypic expression; however, the technique
is so technically demanding that it has limited use in clinical laboratories.
2. Limitations of PCR
a. PCR only analyzes the target region. Testing only provides information on
the target segment amplified by the specific primer set employed.
b. PCR only amplifies intact target regions. Mutations that damage a primer
binding site (including insertions, deletions, and even point mutations)
preclude amplification of the target region by PCR and can easily lead to
errors in test interpretation. Similarly, mutations that alter the structure
of the target region in ways not accounted for during primer set design
(e.g., large insertions, deletions, inversions, or translocations) may pre-
dude amplification.
c. Amplification bias. PCR bias refers to the fact that some DNA templates
are preferentially amplified versus other templates within the same reac-
tion. PCR bias can be caused by differences in template length, random
variations in template number (especially with very low target abundance,
producing an artifact known as allele dropout), and random variations in
PCR efficiency with each cycle. Amplification bias can even result from dif-
ferences in the target sequence itself as small as a single base substitution.
PCR bias can cause over tenfold differences in amplification efficiency in
some settings, a difference that can influence quantitative PCR (Q-PCR)
test results and loss of heterozygosity analysis. PCR bias can be a partic-
ularly troublesome problem in multiplex PCR.
d. Technical factors. There are several technical factors that can lower the
sensitivity and specificity of PCR in routine clinical practice below that
obtained in optimized research settings. Nonspecific inhibitors ofPCR are
sometimes present in patient samples, including heparin and uncharacter-
ized components of CSF, urine, and sputum. With the extreme sensitivity
of PCR, strict attention to the physical organization and methodologies
of the laboratory are required to avoid cross-contamination of specimens.
However, the most important technical limitations are introduced
when fixed rather than fresh tissue specimens are used for testing due
to the degradation of DNA and mRNA that occurs prior to and dur-
ing fixation, as noted earlier. Test sensitivity and specificity are compro-
mised by degradation since it makes it necessary to amplify shorter target
sequences or employ a nested PCR approach, both of which increase the
risk of amplification of nonspecific sequences and cross-contamination
(Am] Surg Pathol. 2002;26:965).
C. Factors that affect testing on a diagnostic level. The intrinsic biologic variability
of disease has the greatest impact on the diagnostic sensitivity and specificity
a...,«). Oi'«t tftd fl\tlil'ld Ultltledt fer DNA S&qi./8/'W!i& AII•!Jti:a I I ••
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loa IU"Olt of die fact that tbo ttA!•lq"" it 10 IICIII!IIvc tbat It am:Pll6co Wl<t
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both, lll'lectico hias !also ..n..i...,;&ation bial, poo<li<'" rm.r.a1 hiu,arulwcrk·
up lriu) i& iDtmclaa:d ill"' die-. Dioal:poDt Alllllym !also .,....., u 4iJ.:cr.
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a. The relative merit of the molecular testing should focus on the ability of
the test findings to improve patient care. For clinical utility, the molecular
diagnostic test must provide an improvement in the standard of patient
care by providing new or refined information with the potential for clini-
cal stratification of disease subtypes, prognostic categories, treatment reg-
imens, gene-targeted therapies, survival statistics, or disease progression.
The test results should complement the findings of established tests, such
as cytogenetics, immunohistochemistry, and cell surface marker analysis.
In the context of surgical pathology, the test results must be correlated
with the histopathologic features of the case.
b. Routine clinical use of molecular tests must consider practical aspects
of clinical prevalence (the disease should represent a significant health
problem or diagnostic dilemma), test run frequency, clinically relevant
turn-around-time, sensitivity, and specificity. Testing for diseases which
are common in many populations (e.g., cancer, microbial infections, and
genetic predispositions) and have well-defined molecular markers will be
performed in many laboratories. Molecular genetic testing for rare disor-
ders will be routinely available only at selected laboratories with specific
clinical programs or areas of institutional focus and expertise.
c. Testing must include steps to validate the result, and will include positive
and negative assay controls, definition of the details and limits of interpre-
tation of test results, and provisions for proficiency testing of the analytic
method, competency of the technologists, and interpretive expertise of the
laboratory director.
d. Results must be reported in a context that explains the molecular assay
data and integrates the findings with other pathology results to avoid
seemingly contradictory reports in comparison with other laboratory tests
that possess different levels of resolution or detection.
e. Biosafety, legal, ethical, and privacy issues must consistently be observed.
2. Discordant cases. Cases will arise in which there is a lack of concordance
between the diagnosis suggested by the molecular test results and the mor-
phologic diagnosis. The debate over the best approach to resolve the ambi-
guity presented by these cases reflects the fundament impact of molecular
genetics on the classification of disease as well as the status of morphology
as the historical standard of diagnosis by which new methods are measured.
Rather than arbitrarily assuming that genetic testing or morphology is supe-
rior in all cases, the most reasonable way to handle discordant cases is to
acknowledge the presence of the discrepancy, and then reappraise the clin-
ical data, pathological findings, and therapeutic implications of all the test
results.
For those cases in which the diagnosis suggested by morphology and
genetic testing are different, prospective clinical trials are required to assess
whether stage, prognosis, and response to treatment are more accurately pre-
dicted by the molecular test results than by the morphologic findings on which
most staging and treatment protocols are based. Epidemiologically, there is a
distinction between diagnostic testing and prognostic testing, with different
study designs required to assess the performance of tests in these different
settings(/ Clin Epidemiol. 2002;55:1178; Ann Intern Med. 2003;139:950).
V. VARIATIONS OF PCR
A. Nested PCR. In this technique, two consecutive PCRs are performed on the same
DNA sample; an initial amplification of a longer target sequence followed by a
second amplification of a shorter sequence contained within the first amplicon.
The second PCR may involve two internal primers (fully nested) or one internal
primer and one of the original primers (seminested). Nested PCR provides a
marked increase in sensitivity compared with traditional PCR, and is desirable
when the target sequence is present at an extremely low copy number, such
as when the mutation is present in only a small subset of the cell population
under study, when the nucleic acids have been degraded as a result of tissue
fixation, or, for reverse transcriptase-PCR (RT-PCR) as discussed below, when
the target mRNA is expressed at an extremely low level. Since the increased
sensitivity carries an increased risk of cross-contamination, reproducible nested
PCR results require strict attention to laboratory technique, rigorous use of
controls, and confirmation of product identity.
B. RT-PCR. RT-PCR makes it possible to amplify RNA extracted from a tissue
sample; a complementary DNA (eDNA) strand is synthesized from the RNA
template using the enzyme RT, and the eDNA is then amplified by conventional
PCR. Fresh (or fresh frozen) tissue is the preferred source of RNA for RT-
PCR. RNA from fixed tissue is an acceptable substrate for testing (e-Fig. 60.1),*
even though it always suffers some degree of degradation depending on the
prefixation interval, the type of fixative, the length of fixation, and the method
used to isolate the RNA.
1. Advantages of RT-PCR. RT-PCR permits direct amplification of multiexon
sequences by eliminating the intervening introns, and thus greatly simpli-
fies mutation scanning methods. Similarly, RT-PCR makes it much simpler
to demonstrate the presence of translocations that create fusion genes by
making it possible to directly detect the fusion transcripts encoded by the
translocations (e-Fig. 60.2). RT-PCR can also be used to detect changes in
mRNA structure that result from alternative splicing, to demonstrate aber-
rant splicing due to mutations, and to evaluate the level of gene expression
through the quantitative methods discussed below.
2. Limitations of RT-PCR. RNA is a more technically demanding substrate with
less stability than DNA. Tissue samples must be processed rapidly (ideally,
within 20 minutes) to avoid mRNA degradation, especially since many muta-
tions render transcripts more susceptible to cellular mechanisms that clear
abnormal transcripts from the cell and result in unstable mRNA. A nested
PCR approach is often necessary when the target RNA is present at very low
levels, but RT-PCR carries an increased risk of contamination and amplifica-
tion of nonspecific sequences because the transfer of the first PCR product to
a separate tube for the nested PCR entails transmission of a highly amplified
DNA preparation.
C. Q-PCR. An ideal PCR would generate a perfect twofold increase in the number
of copies of the amplicon in each cycle of the reaction. In reality, inhibitors of
the reaction, accumulation of pyrophosphate molecules, decreasing polymerase
activity, and reagent consumption all contribute to a plateau phase in the later
stages of the reaction during which the amplicon is no longer accumulating at an
exponential rate (Clin Chem Lab Med. 2000;38:833). Reliable quantitation of
PCR therefore involves more than simple measurement of the amount of product
DNA present at the end of 30 to 40 cycles of the reaction. Real-time PCR, also
referred to as Q-PCR, employs real-time measurements of DNA accumulation
(usually via fluorescence-based approaches) during the early exponential phases
of PCR progress to provide precise estimates of the initial concentration of the
target sequence(s).
A wide variety of different chemistries for Q-PCR are in routine use, includ-
ing the so-called TaqMan (also known as 5' exonuclease or hydrolysis real-time
PCR), molecular beacon (which can be designed to distinguish targets differing
by only a single nucleotide), scorpion (also known as self-probing amplicons),
hybridization probe, and intercalating dye methods. Regardless of the chem-

istry, changes in fluorescence that result from target amplification are measured
by a detector for each cycle of the reaction, and used by a computer to construct
an amplification plot of fluorescence versus the cycle number to quantify the
concentration of the input target DNA sequence.
1. Advantages of Q-PCR. The method can be applied to fresh as well as FFPE tis-
sue, and phenotype-genotype correlations are possible through analysis of
specific cell populations collected via microdissection, laser capture microdis-
section, and so on. Q-RT-PCRis also a robust analytic approach (e-Fig. 60.3).
2. Disadvantages of Q-PCR. Even for optimized assays, testing can be compli-
cated by amplification bias, which in the context of Q-PCR has two major
sources; PCR drift due to random fluctuations in amplification efficiency
in the early cycles of the reaction when the templates are present at very
low concentration, and PCR selection due to mechanisms that systemati-
cally favor amplification of some particular target(s). For Q-RT-PCR, the
reverse transcription reaction can introduce additional variables into the
analysis.
3. Use of Q-PCR in nonquantitative seHings. Since the probes used in Q-PCR
(and Q-RT-PCR) have specificity for the target amplicon, the amplification
plot confirms not only the presence of the DNA product, but also its iden-
tity. Intercalating dyes can also provide confirmation of both the presence
and the identity of the DNA product when coupled with subsequent melting
curve analysis. Because Q-PCR eliminates the need for gel electrophoresis to
demonstrate successful amplification while simultaneously confirming prod-
uct identity, Q-PCR is often used as a "one-step" alternative to conventional
PCR or RT-PCR even when quantitation is not required.
D. Multiplex PCR. Multiplex PCR is the simultaneous amplification of multiple
target sequences in a single reaction through the simultaneous use of multi-
ple primer pairs. The technique saves time and money, is ideal for conserving
templates that are in short supply, and has been successfully applied to many
amplification approaches including nested PCR and Q-PCR. However, even in
optimized reactions, multiplex PCR may be complicated by amplification bias
due to PCR drift and PCR selection. Rigorous optimization of primer design
and careful titration of the relative primer concentration among separate primer
pairs are essential for robust, reproducible multiplex PCR.
E. Methylation-specific PCR. Methylation of CpG sites in human DNA has been
associated with transcriptional inactivation of imprinted genes, is important
for X chromosome inactivation, and is an important mechanism for develop-
mentally regulated and tissue-specific gene regulation. An altered pattern of
methylation is also characteristic of many human diseases (e-Figs. 60.4 and
60.5). Changes in the CpG methylation pattern in some malignancies have been
associated with differences in response to specific chemotherapeutic agents and
overall survival.
Recently developed methylation-specific PCR techniques exploit the
sequence differences produced when methylated CpG (meCpG) and unmethy-
lated CpG are treated with sodium bisulfite (Proc Natl Acad Sci USA.
1996;93:9821). This chemical modification will not alter methyl cytosine but
will depurinate cytosine to produce a transversion, which results in replacement
by thymidine in subsequent DNA synthesis during PCR. Since the two strands
of genomic DNA are no longer complimentary after sodium bisulfite treatment,
PCR with specifically designed primers for meC and T substituted sequences
makes it possible to infer the methylation status of the original untreated DNA.
Methylation-specific PCR can be applied to DNA extracted from fresh tissue,
FFPE tissue, and even archival cytology specimens.
F. Telomerase repeat amplification protocol (TRAP). The hexanucleotide TTAGGG

repeat sequence of human telomeres is essential for the maintenance of chro-
mosome stability and integrity, and abnormalities of telomere length have been
associated with a number of developmental abnormalities and malignancies.
The PCR-based TRAP assay is a simple, sensitive, and reproducible method for
measurement of telomerase activity as an adjunct in early diagnosis, for prog-
nostic testing, or as a means to identify drugs that inhibit telomerase function.
The technique has limitations; the protein extract used for testing can only
be prepared from fresh cells or tissue samples, false-negative and false-positive
reactions are commonly encountered, and heterogeneity within a tumor can lead
to significant variability in the test result. Although early studies suggested that
altered telomerase activity is a characteristic finding in a variety of neoplasms,
a growing body of evidence suggests that telomerase activity demonstrated by
the TRAP assay is neither a consistent feature of malignancy nor specific for a
malignant phenotype (Hum Pathol. 2004;5:393). Attempts to correlate a malig-
nant phenotype with telomerase activity measured by other techniques (such as
RT-PCR-based measurement of the level of expression of mRNA encoding the
hTERT catalytic subunit of telomerase, or immunohistochemical analysis of
hTERT) have likewise produced mixed results.
VI. DNA SEQUENCE ANALYSIS
A. Direct DNA sequence analysis
1. The dye terminator cycle sequencing method for direct DNA sequencing is cur-
rently used for virtually all routine DNA sequence analyses. This technique
(e-Fig. 60.6) utilizes synthetic oligonucleotide primers complimentary to a
known sequence of the template strand to be analyzed, and is greatly simpli-
fied by the use of fluorescently labeled, chain-terminating dideoxynucleotide
triphosphates. As initially described, enzymatic extension of the primer was
performed only once per sequencing reaction, but the utility of the method
is greatly increased by the modification known as cycle sequencing (or linear
amplification sequencing). Cycle sequencing is similar to conventional PCR
in that it employs a thermostable DNA polymerase and a temperature cycling
format for DNA denaturation, annealing, and enzymatic DNA synthesis, but
only one primer (the sequencing primer) is added to the reaction mixture.
2. Next-generation sequencing (NGS). Recent developments in the technology of
so-called next-generation DNA sequencing (also known as deep sequenc-
ing) using several different platforms can generate millions of short sequence
reads per run, making it possible to sequence gene panels, all expressed genes
(known as the exome), or even the entire human genome at an extremely
low cost (Brie(Bioinform. 2010;2:484;Protein Cell. 2010;1:520). The many
genetic variations that can be detected by NGS include single-nucleotide poly-
morphisms (SNPs), indels (small insertions and deletions), structural variants
(SVs), and copy number variations (CNVs). Massive sequencing of eDNA
libraries, also known as RNA-Seq, is an application of NGS technologies
focused on the transcribed portions of the human genome (known as the tran-
scriptome), which provides a comprehensive readout of gene expression that
exceeds the sensitivity of microarray-based methods. And through the use of
sodium bisulfite treatment of DNA, NGS technologies also offer the potential
to study genome-wide DNA methylation patterns (epigenomic analysis).
Complete whole genome sequences have already been published from
several human cancers, including acute myeloid leukemia, breast cancer,
melanoma, lung cancet; and glioblastoma. Recent demonstration of whole
genome sequencing of a proband with Charcot-Marie tooth syndrome
demonstrates that NGS also has significant diagnostic potential in clinical
settings besides cancer biology, such as the diagnosis of inherited diseases.
As with all other molecular diagnostic tests, significant attention must be paid
to NGS results to ensure data quality, including the incorporation of con-
trols that allow the identification of sample contaminations, library chimeras,
sample mix-ups, tumor-normal switches, and variable run quality.
B. Indirect DNA sequence analysis. Indirect identification of normal and mutant
alleles at a specific locus, which correlate with the presence of disease can be
of clinical utility and substitute for direct determination of specific nucleotide
sequences. Virtually all of the indirect methods are based on PCR and can be
applied to a broad range of clinical specimens. Examples of indirect methods
include the following.
1. Allelic discrimination by size. Alleles that vary by small insertions or deletions
can be distinguished on the basis of the size of the PCR product after gel
electrophoresis, perhaps the most straightforward method for indirect DNA
sequence analysis (e-Figs. 60.4, 60.7, and 60.8).
2. Allelic discrimination based on susceptibility to a restriction enzyme. Using
the technique known as restriction fragment length polymorphism (RFLP)
analysis, mutations that either create or destroy a restriction endonuclease
site can easily be distinguished by a two-step process that involves DNA
digestion with the restriction endonuclease followed by gel electrophoresis
to size fractionate the digested DNA. Virtually all RFLP analysis is performed
on PCR product DNA (e-Figs. 60.7, 60.9, and 60.10).
3. Allele-specific PCR. Allele-specific PCR (also known as the amplifica-
tion refractory mutation system [ARMS]) employs oligonucleotide primers
designed to discriminate between normal and mutant target DNA sequences
that may differ by a single base (] Mol Diagn. 2007;9:272). Simultane-
ous analysis of multiple loci via a multiplex PCR format is possible if the
amplicons from different loci are of different sizes. PCR conditions must be
optimized with sufficient stringency so that amplification only occurs when
complete DNA sequence complementarity exists between primer and target
molecules.
4. Single-strand conformational polymorphism (SSCP) analysis. SSCP is one of
the most simple and widely used techniques used as a mutation scanning
system. Single-stranded DNA molecules fold into complex three-dimensional
structures, stabilized primarily by intrastrand base-pairing hydrogen bond
formation, that alter the mobility of the molecule during nondenaturing gel
electrophoresis. SSCP has limited resolution of DNA fragment sizes (100 to
400 bp long is optimal), and while a base sequence may be indicated by the
altered mobility, SSCP does not provide information about either the location
of the base change within the DNA fragment or the chemical identity of the
base change. In practice, the DNA fragments evaluated by SSCP are generated
byPCR.
5. Melting curve analysis. Every DNA duplex has a characteristic melting tem-
perature (dependent on sequence and duplex length) and though small, the
differences in melting temperature between the duplexes can be reliably
detected by high-resolution melting analysis. Differences in the melting tran-
sition of PCR products can therefore be used to infer sequence variations
such as SNPs, small deletions, and small insertions. The most straightforward
methods melt unlabeled PCR products in the presence of DNA-binding dyes
such as SYBR Green that differentiate double-stranded from single-stranded
DNA during melting by changes in fluorescence intensity (Nature Protocols.
2007;2:59).
C. Clonality assays. Demonstration that the cells in a lesion share a common genetic
alteration can be used to support classification as a neoplasm rather than as a
polydonal reactive process, although it is important to emphasize that clonal
neoplasms are not necessarily malignant.
1. Assays based on immunoglobulin and T-cell receptor genes. Most PCR clon-
ality assays performed clinically are used to assess lymphoid infiltrates on
the basis of evaluation of immunoglobulin gene or T-cell receptor gene rear-
rangements. PCR primer design is an important component of these assays;
since generation of immunoglobulin and T-cell receptors involves deletions,
template-independent nucleotide additions, and single base-pair changes,
consensus primers are designed to bind to conserved sequence regions, and
multiple sets of primers are used in order to insure that a broad range of
rearrangements can be detected. Demonstration of a monoclonal or oligo-
clonal population of cells within an infiltrate is very often, but not always,
indicative of malignancy since oligoclonal or monoclonal gene rearrange-
ments may characterize reactive lymphoid proliferations.
2. Clonality assays based on specific gene mutations. This class of assays focuses
on detection of specific mutations in individual genes, including single base-
pair changes and larger-scale structural changes (such as deletions or inser-
tions of viral genomes). This type of analysis can be useful when attempting to
show that two neoplasms represent independent synchronous tumors rather
than one tumor with metastases.
D. Microsatellite instability (MSI) assays. Defects in the DNA mismatch repair sys-
tem produce a characteristic pattern of mutations known as MSI. Direct anal-
ysis of the genes responsible for mismatch repair is not desirable in routine
clinical practice because it requires complete DNA sequencing of (at least) four
causative genes; there are no specific mutation "hot spots," and the genes may
be inactive as a result of epigenetic silencing rather than mutation. PCR-based
analysis offers a more efficient, though indirect, method to identify defects in the
DNA mismatch repair system via detection of the short increases or decreases
in the length of the short tandem repeat (STR, or microsatellite) sequences that
are the characteristic feature of MSI. Mutations in the genes which normally
monitor the fidelity of DNA replication of these repeated sequences is defective
and allows for the generation of the variable lengths of repeats in the tumor
cells.
Laboratory testing regimens for MSI have only been formally addressed in
the context of colo rectal cancer (Cancer Res. 1998;58:5248 ). For all other tumor
types, MSI testing is not yet standardized in terms of the number and identity
of microsatellite loci that must be analyzed, or in terms of the number of loci
that need to show length alterations to be considered indicative of MSI.
DNA derived from either fresh or FFPE tissue can be analyzed in the MSI
assay. Comparison of the size of the PCR products from the target STR loci in
the neoplasm versus normal tissue is used to detect changes in the length of the
microsatellite sequences indicative of MSI. In most cases, the profile of PCR-
amplified sequences permits straightforward classification as indicative of MSI
or not; however, there are no uniform criteria for interpretation of marginal test
results, although standards have been proposed (Mutat Res. 2001;461:249).
Sources of variation in MSI analysis include the presence of contaminating non-
neoplastic tissue (which can limit test reliability because demonstration of MSI
in even the most sensitive testing regimens requires that neoplastic cells comprise
at least 10% of the total population), the identity of the microsatellite markers
used in the analysis (the susceptibility of a given microsatellite to instability is
highly dependent on both the number of repeats and the length of the repeat
units), and the potential for biased amplification of some alleles.
E. Infectious disease testing. PCR-based molecular genetic approaches frequently
have higher sensitivity than standard special stains, and can often provide
information not typically available from special stains such as species-specific
identification and drug sensitivity. Molecular methods are also a useful way to
detect organisms that cannot be cultured (e.g., human papilloma virus [HPV])
or that are notorious for their slow growth in culture (e.g., Mycobacterium
tuberculosis).
1. Bacteria
a. Mycobacteria. In most assays, PCR targets the highly conserved gene that
encodes the 65 kDa heat shock protein, but other target loci include the
genes encoding 16S rRNA or the repetitive insertion element IS611 0. PCR
testing has been successfully applied to FFPE tissue from a wide variety of
sites, including the respiratory tract, GI tract, GU tract, skin, bone, liver,
and lymph nodes, and also to cytology specimens. Alternative isother-
mal amplification approaches originally developed for use in the clinical
microbiology laboratory have also been adapted for use with processed
tissue specimens (Expert Rev Mol Diagn. 2004;4:251).
b. Helicobacter pylori. Although the genome of H. pylori is remarkable for
the polymorphism between different clinical isolates, PCR-based methods
have nonetheless been developed that permit successful detection of vir-
tually all reported forms of H. pylori by PCR from either fresh or FFPE
tissue, including the nonculturable coccoid form. The most common target
loci in PCR assays include the 16S rRNA gene, urease gene, or arbitrary
regions chosen empirically based on their utility. Recently developed real-
time Q-PCR methods that target the 23S rRNA gene permit simultaneous
detection of H. pylori and antibiotic-resistant testing.
c. Other bacterial pathogens. PCR has been used to detect Bacillus anthracis
organisms in patient specimens associated with bioterrorism or acciden-
tal environmental release from bioweapons facilities (] Clin Microbial.
2002;40:4360).
2. Fungi. Most PCR assays target sequences within the fungal rRNA genes.
PCR can be performed using universal primers that bind to highly conserved
sequences in the region, followed by direct or indirect DNA sequence analysis
of the PCR product to identify the specific fungal pathogen. Alternatively,
sequence differences in the 18S rRNA gene can be used to design primers
that are specific for individual fungal pathogens. Sequence polymorphisms
of the mitochondrial large subunit rRNA gene have also been used as a target
in PCR tests to identify fungal pathogens in tissue specimens, and Q-PCR
methods have also been developed for detection of fungal pathogens in tissue
specrmens.
3. Viruses
a. HPV. Most PCR protocols for HPV testing make use of consensus primers
targeted to the viral Lt gene that are potentially capable of detecting
all HPV types that affect the anogenital region. Following amplification
using consensus primers, the HPV type can be determined by either DNA
sequence analysis or membrane hybridization with type-specific probes.
However, both approaches are labor intensive and difficult to automate,
which makes them poorly suited for screening a large volume of patient
specimens. For this reason, HPV testing of cytology specimens is usually
performed using liquid-based methodologies (as is discussed in more detail
in the cytopathology section of Chap. 34 ).
b. Hepatitis C virus (HCV). RT-PCR methods used to detect HCV in liver
biopsy specimens focus on the 5' noncoding region of the virus that is
highly conserved between the (at least) six genotypes and more than 90
subtypes of HCV that have been described worldwide. Maximal RT-PCR
test sensitivity can only be achieved via a nested RT-PCR approach, or
when the PCR products are evaluated by Southern blot hybridization.
c. Epstein-Barr virus (EBV). A Q-PCR methodology has been described that
targets five highly conserved segments of the EBV genome(] Mol Diagn.
Chapter 60 • Direct and Indirect Methods for DNA Sequence Analysis I 90 1
2004;6:378). The method can be applied to FFPE tissue, but maximum

test sensitivity is only obtained when analysis involves all five marker loci.
d. Respiratory viruses. Although PCR assays have been developed for many
individual viruses, a multiplex RT-PCR assay for seven common respira-
tory viruses (specifically, adenovirus, influenza types A and B, respiratory
syncytial virus, and parainfluenza types 1 to 3) is one of the most effi-
cient molecular approaches thus far described(] MolDiagn. 2004;6:125).
The multiplex assay not only has a specificity of 100% for each viral
pathogen, but also has sensitivity for each virus that is superior to either
direct immunofluorescent antibody staining alone or antibody staining
combined with viral culture.
A nested RT-PCR assay has been described for detection of the coro-
navirus responsible for severe acute respiratory syndrome (SARS) (Am I
Clin Pathol. 2004;121:574). The method can be applied to FFPE tissue
from both open-lung biopsies and necropsy specimens, a feature of the
assay that is important given the virulence of the pathogen. The rapidity
and simplicity of the approach make it ideally suited for diagnosis given
the epidemiology of SARS outbreaks.
4. Protozoans
a. Toxoplasmosis. Several different PCR-based assays (for use with either
fresh or FFPE tissue) have been developed for detection of Toxoplasma
gondii, the etiologic agent of toxoplasmosis. The assays have clinical utility
because serologic diagnosis of active infection is unreliable since IgM levels
do not correlate with recent infection, and since reactivation of disease is
not always accompanied by changes in antibody levels.
b. Leishmaniasis. A number of different loci serve as targets in PCR-based
tests for leishmaniasis, including repetitive nuclear DNA sequences, genes
encoding rRNA, and kinetoplast DNA. In fact, PCR utilizing primers that
amplify a 120-bp fragment of kinetoplast DNA has a higher sensitivity
than all other diagnostic methods that have been evaluated, including
serologic testing, microbiologic culture, routine histopathologic evalua-
tion, and immunohistochemical staining.
c. Intestinal parasites. A nested PCR that targets the gene encoding the small
subunit rRNA gene of microsporidia has been described that can be used
with fresh tissue for species-specific detection of four different pathogenic
microsporidia; since different pathogenic microsporidia can have different
sensitivities to antimicrobial agents, testing provides an opportunity to
establish the diagnosis as well as direct therapy. PCR targeted to genes
encoding 18S rRNA or oocyst wall proteins can be used to document
infection with Cryptosporidium.
F. Identity determination. Although the advantages of DNA-based identification
analysis have been most widely publicized in forensics and parentage studies,
this testing also has a role in the routine practice of surgical pathology including
resolution of specimen identity issues (Am I Clin Pathol. 2011;135:132), differ-
entiation of synchronous and metachronous tumors from metastases, evalua-
tion of tumors in transplant recipients, evaluation of bone marrow engraftment,
diagnosis of hydatidiform moles (e-Fig. 60.11), and demonstration of natural
chimerism.
PCR-based approaches for DNA typing have greatly expanded the range of
testing because they require such small amounts of DNA and can be performed
on fresh, fixed, or even partially degraded specimens. Virtually all DNA typing
is currently performed on the basis of a core set of STR loci chosen by the Fed-
eral Bureau of Investigation of the United States for use in a national database
of convicted felons known as the Combined DNA Index System (CODIS).
Commercial kits for either monoplex or multiplex PCR amplification of COD IS

loci have greatly simplified STR typing and made the method accessible to most
molecular genetic laboratories.
SUGGESTED READINGS
Leonard DGB. Molecular Pathology in Clinical Practice. New York: Springer; 2006.
Pfeifer JD. Molecular Genetic Testing in Surgical Pathology. Philadelphia: Lippincott Williams &
Wilkins; 2006.
Rudin N, Inman K, eds. An Introduction to Forensic DNA Analysis. 2nd ed. Boca Raton: CRC
Press; 2002.
Strachan T, Read AP. Human Molecular Genetics. 4th ed. London: Garland Science; 2010.
Microarrays
Mark Watson
I. INTRODUCTION. Nucleic acid microarrays are an ordered arrangement of DNA

molecules (probes or features) on a solid surface. A sample of DNA or RNA derived
from cells or tissue (target) is then hybridized to the array to quantify the level of
nucleic acid corresponding to each probe. Micro arrays can be utilized for a number
of different experimental and clinical applications (Fig. 61.1).
A. Array comparative genomic hybridization (aCGH). This microarray-based assay is
used to compare genome copy numbers between biospecimens. Often, a patient
tumor DNA sample is directly compared with a corresponding nonmalignant or
germline DNA sample from the same individual to assess quantitative changes in
a tumor genome. This approach is routinely used in clinical molecular diagnos-
tics and provides a higher-resolution complement to more traditional cytogenetic
and fluorescent in situ hybridization (FISH) assays.
B. Genotyping. Microarray technology can be used to assay single nucleotide poly-
morphism (SNP) genotypes or copy number variation (CNV) across 1 to
2 million genomic loci in a single DNA sample. This approach is useful for
identifying correlations between phenotype and genotype in familial linkage
or genome-wide association (population-based) studies and can be used clini-
cally to identify CNV polymorphisms associated with inherited or constitutional
traits.
C. Sequencing. Sequencing by hybridization utilizes microarray technology to
determine the complete nucleotide sequence of a DNA target, usually 100 to
300 kilobases in length. Unlike conventional sequencing chemistry using cap-
illary gel electrophoresis, the target sequence need not be a contiguous stretch
of DNA, but may consist of a set of regions of diagnostic relevance distributed
throughout the genome.
D. Genome tiling. Very high density microarray designs contain nucleotide probes
with 10 to 35 nucleotides spaced across the entire genome. These high-resolution
probe arrays allow for identification of methylation and DNA-protein binding
patterns at the single nucleotide level.
E. Gene expression. By hybridizing cellular RNA to microarrays with probes
directed to mRNA or microRNA (miRNA) targets, it is possible to perform
qualitative (i.e., detection of alternative splicing) and quantitative gene expres-
sion analysis simultaneously on 30,000 to 50,000 genes from a single-RNA
specimen. Measurement of transcript abundance (i.e., gene expression profil-
ing) has been the most common use of microarray technology in investigative
and diagnostic pathology to date.
II. MICROARRAY TECHNOLOGY. Nucleic acids microarrays are fabricated using several
different technologies and probe types, depending upon the intended application
(Fig. 61.1).
A. Bacterial anificial chromosome (BAC} arrays. BAC-cloned DNA is deposited onto
a solid surface (usually a glass microscope slide) by a robotic, mechanical spot-
ting device. Each BAC clone represents a large stretch of genomic DNA from
a specific chromosomal region, usually several hundred kilobases in length.
BAC arrays have been particularly useful in assessing genome-wide DNA copy
number changes in tumor specimens, and when employed in a CGH have
enhanced the resolution of traditional cytogenetic studies. Validated BAC arrays
903
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Figure 61.1 Microarray applications and technology. Microarray technology can be used for many different
applications, depending upon the nucleic acid composition of the array (probes) and the test material (target) that
is hybridized to it. A: For aCGH assays, fragments of genomic DNA (i.e., BACs) are spotted as probes onto the
microarray surface using robotics. Genomic DNA from the patient's tissue and germ line are then cohybridized to
the array to identify regions of chromosomal gain or loss. B: Alternatively, genomic DNA sequence can be used to
computationally design oligonucleotide probes that detect specific alterations in DNA samples (either germ line
or tissue) for either SNP genotyping or DNA resequencing studies. C: eDNA clones generated from cellular RNA
also can be spotted as probes onto the microarray and patient tissue RNA subsequently hybridized to quantify
relative mRNA transcript abundance for gene expression profiling experiments. D: Similarly, oligonucleotides can
be designed based upon known gene exon structure and spotted on the array surface to perform quantitative or
qualitative assessment of mRNA or miRNA expression.
Chapter 61 • Microarrays I 9 05
are used in a number of clinical laboratories, although compared with newer,

oligonucleotide-based platforms, they suffer from poor resolution and require a
sufficient laboratory infrastructure to grow and maintain BAC libraries which
are used to generate the DNA probes. BAC arrays have been largely supplanted
by oligonucleotide arrays for these reasons.
B. eDNA microarrays. Some of the earliest microarray designs utilized eDNA probes.
Messenger RNA (mRNA) from a defined tissue or cell source is converted into
a double stranded eDNA clone library. Plasmid DNA from each clone is then
spotted onto the microarray surface. Genome sequence information is not nec-
essarily required for microarray design, a particular advantage for studying the
few remaining experimental organisms where genome sequence information is
not available. However, because eDNA probes correspond to relatively long
stretches of transcribed mRNA, cross-hybridization and lack of specificity can
often limit the accuracy of eDNA microarray results. Like BAC arrays, the effort
and infrastructure necessary to store, grow, purify, monitor quality control, and
track individual eDNA clones are considerable and not easily standardized. This
last constraint has and will continue to limit the use of eDNA microarrays as
clinical diagnostic tools.
C. Oligonucleotide microarrays. The availability of the completely sequenced and
annotated human genome, coupled with improved synthesis chemistries, has
shifted the fabrication of nucleic acid microarrays toward the use of synthetic
oligonucleotide probes, usually 60 to 75 nucleotides in length. Probe sequences
may be customized for specific genes, gene transcripts, or gene transcript seg-
ments using defined nucleic acid sequences. Sophisticated bioinformatics pro-
grams can select optimized oligonucleotide sequences for any gene or transcript
of interest while minimizing cross reactivity with other sequences, and at the
same time standardizing hybridization properties such as melting temperature
and G/C sequence content. This level of customization has provided a new level
of standardization and flexibility to the design of sequence content on nucleic
acid microarrays that is aptly suited for clinical diagnostic assays.
D. In situ synthesized microarrays. Another strategy for microarray design involves
simultaneously synthesizing specific oligonucleotide probes in situ using com-
binatorial photochemistry. Affymetrix GeneChip® microarrays use a series of
micron-scale "masks" to direct light to specific locations on the microarray
surface. Photoreactive nucleotides (A,C,G,T) are sequentially passed over the
array surface in the presence of each mask. Depending upon the mask pattern,
a specific nucleotide is added to the growing chain of oligonucleotides at a
specific position. In this combinatorial method, the use of 25 different masks
sets (A, C G, T) in 100 sequential nucleotide addition steps can result in 425
(1 x 1015) different sequences that are simultaneously created on the array
surface.
Alternate methods use a "maskless" approach for in situ probe synthesis
in which a sheet composed of micron-scale electronic mirror is programmed
to direct light to specific areas of the microarray during sequential steps of
photochemical oligonucleotide synthesis. The method is similar to GeneChip®
fabrication, but does not rely on the creation of fixed lithographic masks and
therefore allows for flexible design on a single array basis.
E. Bead arrays. An alternate approach to traditional microarray design involves
the use of a beaded microarray. In this approach, micron-sized beads, each
containing a unique oligonucleotide gene sequence in tandem with a unique
nucleotide address sequence, are allowed to randomly assemble onto a solid
surface. By repeated interrogation of each bead address sequence, the identity
of each bead at each position is deduced. The "decoded" array can then be used
for its intended hybridization assay.
Ill. MICRDARRAY ASSAYS. Microarray assays are complex (e-Fig. 61.1)*, both because
of the amount of data generated and the exacting specimen requirements that are
necessary to produce high quality data.
A. Study design. To date, most microarray-based studies have been designed as
biomarker discovery experiments, with the aim of defining a panel of multiple
biomarkers that can then be transitioned into a more conventional clinical assay.
Microarray experiments generally fall into several classes.
1. Class discovery. In such studies, experimental specimens are classified based
solely upon their microarray data values, and the results of the classification
are reviewed to identify new, previously unappreciated clinical or pathologic
classifications. Perhaps the most elegant illustration of this approach has
been the reclassification of breast adenocarcinoma based upon microarray-
generated gene expression profiles (Clin Cancer Res. 2005;11:5678). Similar
studies have effectively identified other molecular subtypes of tumors as well
(] Clin Oncol. 2006;24:5079; N Engl] Med. 2003;348:1777).
2. Class distinction. This type of study is designed to identify novel predictive
biomarkers, patterns of gene expression, CNVs, or sequence alterations that
demonstrate a correlation to an already known parameter such as clinical
outcome or treatment response(] Clin Oncol. 2009;27:1160).
3. Single sample classification. Ultimately, to achieve clinical utility, it is neces-
sary to create a robust molecular signature that can be prospectively applied
to individual patient specimens to accurately predict clinical phenotype. Typ-
ically, a specific subset of probes on a microarray (sometimes a customized
array designed for a specific diagnostic purpose) is examined and a weighted
discriminate index is calculated. The resulting index provides a probability
measure that a given specimen falls into a specific, predefined diagnostic cat-
egory. Studies which independently validate a previously identified signature
are relatively rare to date, but are obviously a critical step in transitioning
any assay into routine clinical use (Clin Cancer Res. 2010;16:5222).
B. Statistical considerations. In principle, microarray data analysis is no differ-
ent than evaluating whether a single biomarker demonstrates a statistically
significant difference between defined sample classes using traditional statis-
tics. By definition, a traditional significance threshold of p = 0.05 allows for
a 5% false-positive (false discovery) rate. Therefore, when analyzing 50,000
to 2,000,000 independent biomarker values obtained by a microarray assay, as
many as 100,000 values will appear to be "significant" by chance alone. To con-
tend with this problem of multiple testing, several methods have been applied to
calculate a true significance threshold when analyzing thousands of variables in
relatively few numbers of samples (Genome Biol. 2003;4:210). Although these
approaches minimize false-positive results for a given sample set, they can in no
way substitute for data validation using multiple, independent sets of samples
across different technology platforms and laboratories.
C. Specimen requirements. Because of the inherent complexity of microarray-based
assays, specimen quality assurance is essential.
1. Specimen collection. Careful consideration must be given to specimen collec-
tion for microarray studies. While DNA and DNA methylation patterns are
relatively stable and probably less sensitive to environmental conditions, the
same is not true for mRNA and miRNA when targeted in microarray-based
gene expression profile assays. Global changes in gene expression can occur
in tissue biospecimens as a result of tissue warm ischemia time (] Clin Oncol.
2006;24:3763 ), creating artificial differences in gene expression patterns seen
between specimens based on collection procedures rather than important
clinical differences. For peripheral blood and bone marrow specimens, the
method in which a specimen is collected and processed can also influence gene
expression signatures (Physiol Genomics 2004;19:247). Finally, most tissue
specimens are inherently heterogeneous collections of many cell types. Vari-
able cellular composition between tissue specimens may lead to differences
in genomic and transcriptional profiles generated from microarray assays.
For example, two prostate tumor samples, one of which contains 5% neo-
plastic cellularity and a second which contains 70% neoplastic cellularity,
may demonstrate two different gene expression signatures based simply on
the content of neoplastic epithelial cells present in the tissue. Similarly, mea-
surement of a tumor-associated change in DNA copy number will vary con-
siderably depending upon the content of neoplastic epithelial cells. For this
reason, many investigators use techniques such as laser microdissection to
isolate more homogeneous cell populations for both gene expression and
DNA copy number microarray analysis (see Chap. 62).
2. Specimen processing. Generally, diagnostic surgical pathology tissue spec-
imens are subjected to formalin fixation and paraffin embedding, a pro-
cess that results in chemical cross-linking and degradation of nucleic
acids. DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tis-
sue may be suitable for some microarray-based DNA analyses (Methods
Mol Biol. 2011;724:127). However, most investigators have found that
RNA derived from FFPE tissue is unsuitable for traditional gene expres-
sion microarray-based assays, although novel molecular amplification pro-
cedures and microarray platforms suggest that this may no longer be true
(]Mol Diagn. 2011;13:48; BMC Cancer. 2011;11:253). Nonetheless, freshly
procured, snap frozen biospecimens remain the "gold standard" for microar-
ray analysis, particularly for RNA-based gene expression assays. Since many
clinical centers do not have access to resources needed for the processing and
storage of frozen samples, a number of solutions have been proposed to cir-
cumvent the limited availability of fresh-frozen biospecimens for microarray
analysis(] Mol Diagn. 2006;8:31). Such advances enable routine prospective
analysis of clinical specimens for RNA- or DNA-based microarray assays.
D. Target preparation. To prepare samples for microarray assays, RNA or DNA
derived from a tissue or cell specimen is converted into a synthetic target in
the presence of labeled deoxynucleotides. In order to analyze clinical specimens
containing small amounts of cellular material, such as diagnostic core or needle
aspiration biopsies, most protocols for microarray target synthesis employ some
method of molecular amplification including PCR, isothermal DNA polymer-
ization, or in vitro transcription (Br] Cancer. 2004;90:1111).
IV. DATA ANALYSIS. Data analysis is by far the most complicated aspect of any microar-
ray study (Nat Rev Genet. 2001;2:418; BMC Bioinform. 2005;6:115). The prin-
cipal steps in microarray data analysis involve the following:
A. Image analysis. Current microarray technology allows for laser scanning of sev-
eral square centimeters of microarray surface at the resolution of micron-sized
image elements. The result is a primary image data file that can be hundreds of
megabytes in size. While a number of software solutions and data repositories
have been developed to hold and distribute experimental microarray data, reg-
ulatory issues related to storage and transfer of data in a clinical setting have
yet to be fully addressed.
The first step in micro array analysis involves the conversion of these raw, pix-
ilated images into numerical values that relate to hybridization signal intensity
at each feature (probe). For two color arrays, the fluorescence intensity must
be sequentially captured and analyzed for each emission spectrum. In other
microarray platforms such as the Affymetrix GeneChip, multiple probes are
used to assay for a single transcript or genomic locus and the signal from the
multiple different probes must be integrated to create a single, averaged intensity

value. Due to the widespread use of the Affymetrix platform, several different
algorithms have been developed to translate raw hybridization data into gene
expression values. Using standardized data sets, the sensitivity and specificity
of each of these algorithms to detect known changes in copy number between
samples have been evaluated at length.
B. Normalization. Once image data have been converted into numerical values for
each probe or probe set represented on the array, the composite set of data
is usually normalized to a reference point to allow for comparison between
sets of array data (e-Fig. 61.2). For example, inter-array normalization algo-
rithms compensate for global differences in the signal intensity between arrays.
Inter-gene normalization algorithms are useful for identifying common patterns
of gene expression between study samples, even when absolute gene expression
values are considerably different. Other types of data transformation techniques,
such as log transformation of two-color signal ratios, may also be appropriate
depending upon the nature of the primary microarray data set.
C. Visualization and data reduction. After normalization, data must be visualized
and statistically analyzed to address a specific research question (e-Fig. 61.2).
There are a number of relatively standard methods in which voluminous and
multidimensional microarray data may be visualized.
1. Hierarchical clustering. Samples are organized based upon their similarity
in gene marker values, and genes are organized based upon their similar-
ity across samples. Several different measures of similarity can be used and
several different algorithms can be applied to perform the clustering, and
no particular algorithm can be considered to be the gold standard. In fact,
while hierarchical clustering is a useful tool to provide a manageable view of
immense data sets, it does not necessarily impart any underlying "truth" to
microarray data.
2. Heat maps. A colorimetric representation of numerical data, usually presented
in combination with hierarchical clustering. This visualization scheme pro-
vides a convenient method to identify patterns or blocks of similarity between
gene markers and/or samples.
3. k-means clustering and principal components analysis {PCA}. These data reduc-
tion methods are particularly useful for reducing the level of microarray data
complexity. A large number of variables (i.e., microarray probe values) are
placed into a finite number of "bins" based upon their similarity of values
across a much smaller number of observations (i.e., target samples). The
number of bins created (the "k" in k-means) can be adjusted to create a
much smaller set of similar, collective values which can then be used as the
basis for further analysis.
In PCA, samples are plotted in "gene marker space" where the distance
between samples in this space is related to their similarity based upon gene
marker values. However, for a 47,000-element microarray, gene marker
space is represented in 47,000 different dimensions. Therefore, the goal
of principal component analysis is to reduce 47,000 dimensions into two
to three principal components. Then, relatedness between samples can be
plotted.
D. Annotation. A final significant challenge in data analysis is to determine how
patterns of gene expression can be related to biologically meaningful results.
As annotation and understanding of the human genome continue to improve,
investigators have been able to classify a large number of human genes into
ontologies based on function, cellular location, and structural determinants.
Several software programs are available that can map lists of biomarkers to
these ontologies, which often results in a clearer view of altered biologic pro-
cesses associated with differential gene expression. For single cellular and simple
Chapter 61 • Microarrays I 909
multicellular organisms, this approach has led to sophisticated models for cell
signaling and transcriptional regulatory networks, whereas for humans this type
of analysis is still evolving.
E. Validation. Like any other diagnostic test, the results of a microarray assay must
be validated through independent testing. Validation of microarray assays is
particularly problematic for many reasons. First, microarray assays are relatively
expensive ($200 to $800 per sample), which creates financial constraints on the
number of samples that can be analyzed. Second, given the stringent specimen
requirements needed to perform most microarray assays, availability of suitable
specimens is often limiting. Finally, as discussed above, the large number of
variables associated with a microarray data set requires that a relatively large
number of observations (e.g., samples) must be analyzed to create any degree
of statistical confidence. To perform data validation for microarray results in
the face of these limitations, investigators have devised a number of approaches
(Expert Rev Mol Diagn. 2003;3:587).
1. Cross validation. One of the most popular approaches for data validation is
sequential sampling or "leave-one-out" cross validation analysis in a single
sample set. In an analysis of N study samples, N- 1 samples are used for the
initial statistical analysis to identify groups of signature genes. The ability
of these genes to correctly classify the Nth sample is then calculated and the
gene list modified, discarding biomarkers that perform poorly and solidifying
those with the best performance. This process is repeated, removing all N
samples, one at a time, until a list of genes with the best class prediction score
is created. The advantage of this method is that no additional data or exper-
imentation is needed for validation. Howevet; because the cross-validation
is still applied to a single set of samples (i.e., the "test" and "validation" sets
are one in the same), the ability to generalize conclusions to independent or
larger sample sets may still be limited.
2. Sample set splitting. If an initial microarray sample set is large enough, it
is also possible to divide the experiment into independent sets of test data
and validation data. In this scheme, patterns of "significant" gene expression
are identified using the first set of samples, and patterns are validated in a
second set of arrays. Although this approach utilizes two truly independent
data sets, it necessarily limits the number of independent samples available
for the discovery phase and validation phase. The desire to split a limited
number of samples into test and validation sets raises the question of sample
size requirement for performing microarray analyses with sufficient statis-
tical power (Physiol Genomics. 2003;16:24). While multiple methods have
been proposed to calculate required sample sizes, the number of samples
required will ultimately depend upon the expected biologic effect. For exam-
ple, relatively few study samples may be necessary to identify fundamental
genomic differences between acute myelogenous leukemia {AML) and acute
lymphocytic leukemia (ALL), as these tumor cell types are biologically very
distinct. On the other hand, a considerably larger study set may be required
to identify reliable differences in molecular signatures associated with clin-
ical outcome within ALL patients if the intrinsic biologic basis for patient
outcome is more subtle (BMC Med Genomics. 2011;4:31).
3. Meta-analyses. Microarray data results can be validated using multiple, inde-
pendent study data sets. As an increasing number of microarray studies are
published and corresponding data sets are made publicly available in micro ar-
ray data repositories, it has become increasingly possible to validate pat-
terns of gene expression identified in one experiment using other microarray
experiments in the published literature (PLoS Med. 2008;5:e184). In fact,
meta-analyses of microarray data are becoming more frequent, and while
some studies have shown that significant patterns of gene expression can be
validated in experiments conducted by independent investigators (N Engl

J Med. 2006;355:560), other such analyses have demonstrated clear differ-
ences between studies (/ Natl Cancer Inst. 2005;97:927).
V. CLINICAL APPLICATIONS. Microarray technology was initially utilized primarily
for basic science studies that involved whole genome analyses to identify novel
biomarkers or elucidate biologic signaling pathways. Howevet; as this technology
has evolved, nucleic acid microarrays are now being utilized as a clinical diagnostic
platform.
A. Gene expression profiling as an ancillary diagnostic tool for the pathologist. Perhaps
one of the most useful applications of gene expression microarray technology
has been its use in the diagnosis of histologically ambiguous tumor specimens.
Gene expression profiles can molecularly define the cell lineage of metastases
of unknown origin (particularly adenocarcinoma) with 80% to 90% accuracy
based upon thorough retrospective analysis of clinical data (Expert Rev Mol
Diagn. 2010;10:17). Gene expression data can also be used to discriminate his-
tologic "look-alikes" with distinct cell origins, such as small round blue cell
tumors (Nat Med. 2001;7:673). Of greater interest are results demonstrating
that molecular profiling can subclassify histologically indistinguishable tumors
into biologically relevant subtypes. The first study to define this principle was
conducted in diffuse large B-cell lymphoma, where gene expression patterns
clearly segregate tumors with aggressive and indolent molecular profiles that
also have significance for patient survival (N Engl] Med. 2003;348:1777).
Gene expression profiles of invasive ductal breast carcinoma can also clearly
define "basal-cell" and "luminal cell" tumor types (among others) that have
unique biologic characteristics and therapeutic response profiles (Clin Can-
cer Res. 2005;11:5678). An even more intriguing application of microarray-
based molecular profiling involves complete tumor reclassification based not on
anatomic site or histopathologic features, but by patterns or modules of gene
expression (Cancer Res. 2008;68:369). Given that the phenotypic behavior of
tumor cells and their response to molecular therapies may be more dependent
upon molecular profiles than organ site of origin, this approach has the poten-
tial to significantly modify the role of pathology and the practice of clinical
oncology.
B. Predicting disease behavior. Gene expression profiles of primary tumor samples
have been used to develop predictive signatures of local and distant metas-
tasis, for both specific and more global tumor types (Cancer Genomics Pro-
teomics. 2007;4:211). Further clinical validation of such signatures could, for
example, define a new diagnostic category of "lymph node potential positive"
tumors which might play an equal if not more important role than traditional
histopathology for staging cancer patients and therapeutic decision making.
C. Predicting survival. Several large clinical studies have identified gene expression
signatures that predict patient survival in breast cancet; lung cancer, leukemia,
lymphoma, and many other tumor types. While results from these studies are
usually individually validated using statistical methodology or through indepen-
dent training and test patient populations, an increasingly worrisome finding is
that clinically predictive gene expression signatures are not reproducible across
multiple studies of the same tumor type (PLoS Med. 2008;5:e34). Proposed
explanations for these discordant findings include the use of differing and non-
standardized microarray platforms, the examination of patient cohorts that are
not matched for clinical and treatment parameters, and the relatively complex
phenotype of survival which is dependent upon many variables in addition to
tumor molecular signatures.
D. Predicting therapeutic response. In the context of neoadjuvant chemotherapy
trials, gene expression microarray analyses of pretreatment tumor biopsy spec-
imens have been used to develop predictive signatures of treatment response
(Clin Cancer Res. 2005;11:5678). To date, these studies have been small and
retrospective. However, the ability to use tumor gene expression data to prospec-
tively manage patient treatment will be an important step toward the concept
of personalized medicine.
E. Nontumor pathology. Microarray technology has been applied to biomarker dis-
covery in other fields of pathology and clinical medicine such as neuropathology
(Alzheimer disease, Parkinson disease, epilepsy, schizophrenia), immunopathol-
ogy (systemic lupus, multiple sclerosis), organ transplantation, reproductive
endocrinology, trauma and sepsis, and cardiovascular disease. However, in mul-
tiorgan disease processes, the appropriate target cell population for study is
often not obvious, or often difficult to obtain from a large number of patients.
Therefore, many microarray studies focusing on noncancer disease processes
have been limited to very small sample sizes. Known and unknown variability
within these disease processes and between patient participants makes it diffi-
cult to establish definitive associations between patterns of gene expression and
disease phenotype.
F. Genotyping. DNA sequence alterations, either single nucleotide polymorphisms
(SNPs) present in germline DNA or somatic point mutations which occur in
tumor cells, are important diagnostic markers for disease predisposition, diag-
nosis, and treatment. Microarray technology is a high-throughput method for
genotyping individuals at as many as one million loci across the genome, and
is beginning to have enormous implications in clinical genetics. For example,
familial linkage studies designed to identify inherited disease genes associated
with tumor syndromes have greatly benefited from this technology. Microarray-
based genotyping has also been used in genetic association studies to define loci
associated with disease predisposition and clinical phenotype. Although most
applications of genotyping microarrays have been used to discover a single dis-
ease locus of interest, it is likely that complex multigenic diseases will require
genotyping at multiple loci in order to accurately classify a genetic phenotype.
G. Genome copy number. Another major application of microarray technology
involves assessment of genome copy number (Nature. 2008;452:553). Tumor
cells experience a wide variety of chromosomal gains and losses, and while these
events have been traditionally measured using cytogenetic techniques, microar-
rays are becoming an increasingly useful method to correlate gene copy number
changes with clinical and pathologic features of tumors. Microarray-based gene
copy number assessment has been used to define critical regions, and even sin-
gle genes, whose gain or loss has previously been measured only on the scale
of chromosome arm losses or gains, which has led to the rapid identification
of new oncogenes and tumor suppressor genes. Microarray analysis of gene
copy number has also identified genomic alterations that are characteristic of
particular tumor types and which can be associated with clinical outcome.
Microarray analysis of gene copy number has also demonstrated that the
germline genome of normal populations contains a large number of copy num-
ber polymorphisms as well (Nat Genet. 2007;39:516). This normal variability
in locus copy number has known significance for disease predisposition and
pharmacogenomics, making the use of copy number-based microarray analysis
a useful tool for patient management.
H. Methylation. Methylation of CpG dinucleotides occurs frequently in tumor
genomes and is associated with transcriptional silencing of key tumor suppres-
sor genes. Microarray measurement of methylation patterns has many potential
applications; for example, it has allowed for whole-genome methylation pro-
filing to distinguish subtypes of leukemia, and has made it possible to identify
patterns of methylation associated with treatment response.
I. DNA resequencing. A final, but perhaps most promising, application of microar-
rays involves their use in gene resequencing. Oligonucleotide microarrays can
be constructed to interrogate individual bases of DNA sequence, thus providing

a method for sequencing clinical samples by hybridization. Microarrays have
been designed to sequence the genomes of the SARS and HIV virus for the pur-
poses of strain classification and predicting response to antiviral agents, respec-
tively. In another application, investigators have developed a "MitoChip," a
microarray designed to sequence the entire mitochondrial genome since specific
mitochondrial mutations occur frequently in human cancer cells and therefore
have become intriguing candidates for tumor biomarkers. As a final example,
a microarray designed to resequence the entire p53 tumor suppressor gene has
been used to predict clinical outcome and therapeutic response in patients, based
on the spectrum of somatic p53 sequence alterations in their primary tumors;
the sensitivity of this approach is >90% and, with the exception of detecting
nucleotide insertions and deletions, is comparable to that of traditional dideoxy
sequencing (Breast Cancer Res Treat. 2011;128).
Now that the cancer genome is being characterized in detail using whole
genome sequencing technologies (see Chap. 60), there is compelling evidence
that specific gene sequence alterations in tumors will predict vulnerability and
resistance to specific therapies such as the epidermal growth factor receptor
inhibitor, gefitinib (J Clin Oncol. 2007;25:587). As more and more clinically
relevant somatic sequence alterations are identified in viruses, tumor cells, and
other pathologic specimens, the ability to perform rapid resequencing of multi-
ple genetic loci on a single clinical specimen will be an important requirement for
the clinical pathology laboratory. In this respect, as compared with traditional
sequencing methodologies, microarrays and the sequencing by hybridization
approach could become routine clinical molecular diagnostic tools that supple-
ment many traditional histopathologic and immunohistochemical approaches
currently used to evaluate pathology specimens. Alternatively, as the cost of
next-generation sequencing technology continues to fall, assays that employ
quantitative RNA-sequencing or targeted genome sequencing could eventually
obviate the use of any microarray technology in the clinical laboratory alto-
gether.
VI. USE OF MICROARRAYS IN THE CLINICAL LABORATORY. While nucleic acid microarrays
have been used for numerous research studies with potential clinical significance,
the routine use of this technology platform in the clinical laboratory still requires
several significant advancements.
A. Array content. Since the microarray format is well suited to measure multiple
markers from a single sample, it is ideal for the clinical laboratory that per-
forms prospective analysis on single clinical specimens. For discovery experi-
ments, thousands of individual genetic markers can be simultaneously assayed,
but in doing so, each marker is optimized for neither sensitivity nor specificity.
In clinical testing, this may be problematic for measuring low levels or subtle
alterations of gene expression, or for detecting genomic alterations that may be
present in only a subset of cell populations. By designing disease-specific marker
sets, optimizing hybridization conditions for each marker and placing redun-
dant probes for each marker on the same array, it may be possible to increase
the sensitivity and precision of microarray assays for clinical use, although this
has only recently become a micro array design consideration. A microarray used
for discovery purposes may assay 10,000 genes using a single set of probes;
a similarly sized array could assay 100 genes using 100 different probe mea-
surements. Thus, the ability to create large number of redundant probe sets and
control sequences could provide a level of precision not currently available from
most research-based microarrays.
B. Automation and standardization. Another potential advantage of microarray tech-
nology is that it can be highly automated and regulated, attributes that are
critical for any clinical laboratory test. Newer sample preparation methods
utilize single tube, "hands-off" chemistries that facilitate routine use in the
clinical laboratory. Self-contained microarray cassettes, such as those manufac-
tured by Affymetrix and other vendors, also provide an acceptable format for
regulated clinical tests. Finally, as diagnostic biomarkers become more estab-
lished, microarray platforms will need to evolve into clinical diagnostic tools
which meet all regulatory requirements of standard clinical tests, including addi-
tional standardization and inter-laboratory assay validation, which are both
now receiving appropriate attention (Nat Biotechnol. 2010;28:827).
C. Assay complexity. Microarrays are indisputably high complexity assays, and
although streamlined protocols, automation, and standardized array manu-
facturing can mitigate some inherent technical complexity, microarray assays
still remain in the domain of specialized clinical laboratories. Like most
hybridization-based assays, the time required to perform microarray analy-
sis even in its most automated format may still be three to four days. Assay
turnaround time could be problematic when assay results are needed to guide
immediate therapy. Finally, because microarray assays are based on signal detec-
tion by hybridization, there is an inherent limit to sensitivity as compared with
amplification-based PCR detection assays.
D. Sample requirements. Another technical limitation of microarray assays is their
requirement for high sample quality. Although exact specimen requirements
depend upon the analyte measured (i.e., mRNA, genomic DNA, methylated
DNA), the use of formalin-fixed tissue or other specimens that have been col-
lected under routine hospital conditions currently limits the use of microarray
technology in a number of clinical settings. Methods are available for amplify-
ing and labeling nanogram quantities of nucleic acids samples (both DNA and
RNA), so the ability to utilize small numbers of cells obtained from minimally
invasive procedures such as fine needle aspiration, swabs, or lavages allows
microarray-based assays to be used for diagnostics in an increasing number of
different clinical scenarios. Refinements in the use of routine pathology tissue
specimens that have been fixed and embedded in paraffin will allow full inte-
gration of this technology with routine histopathologic assessment.
E. Clinical relevance. The biggest challenge for implementing microarray technol-
ogy in the clinical laboratory, however, is not array technology itself but rather
the clinical validity of the array biomarker content. To date, the majority of
microarray platforms have been designed as whole genome discovery tools.
Studies have used a relatively small number of observations (patients) and a
large number of variables (genes), an approach which has consistently led to
high false-positive rates that are unacceptable for clinical assay validation. A
considerable amount of microarray data generated and reported in the liter-
ature (particularly gene expression data) has not been reproducible in inde-
pendent samples sets (PLoS Med. 2008;5:e184). Therefore, while microarray
technology itself still holds promise for use in a clinical laboratory, there is
a need for many larger, prospective correlative studies to support the use of
microarray-based biomarker panels in routine diagnostic pathology.
Biospecimen Banking
Mark Watson
I. SIGNIFICANCE TO PATHOLOGY AND TRANSLATIONAL RESEARCH. National initia-

tives such as The Cancer Genome Atlas (http://cancergenome.nih.gov) and the
1,000 Genomes Project (http://www.1000genomes.orgl) have greatly enhanced the
resources available to better understand the molecular and genetic basis of human
disease. Technologies such as gene expression profiling, methylation scanning, com-
parative genome hybridization, and high-throughput resequencing have also made
it possible to harness genomic information to identify disease-specific molecular
alterations that occur in human tissues. However, to ultimately understand the
significance of basic biological findings toward the improved diagnosis and treat-
ment of human disease, primary human biospecimens (i.e., tissue and body fluids)
must be available for clinico-genomic correlative studies. Six important principles
embody the rationale for biospecimen banks.
A. Availability of large biospecimen cohorts. The number of specimens available
must be relatively large to generate statistically significant findings. For example,
because of the large number of variables (e.g., gene expression values) generated
from DNA microarray experiments compared to the much smaller number of
observations (patients), traditional statistical approaches generate a large num-
ber of false positive results, that is, genes whose expression appears to classify
subpopulations of patients but only does so by mere chance. Validating find-
ings from such whole genome approaches requires large numbers of patient
specimens.
B. Accurately diagnosed biospecimens. Several anecdotal examples have illus-
trated how gene expression profiling can correctly classify specimens that
were histologically misidentified by routine clinical pathology (Am I Pathol.
2001;159:1231). However, initial diagnostic mislabeling can seriously impede
supervised learning approaches to identify new biomarkers. In addition to issues
of diagnostic accuracy, knowledge of the precise cellular makeup of specimens
(particularly for heterogeneous tissues) is important to properly interpret both
gene expression profile and DNA sequencing data. Techniques such as laser cap-
ture microdissection (LCM) can be used to isolate homogeneous cell populations
(Acta Histochem. 2007;109:171), but requires considerable technical expertise.
An added level of complexity in longitudinal studies is that multiple specimens
collected from the same patient need to be accurately coded and tracked to
ensure that the intended specimen (e.g., initial presentation vs. relapse) is used
for downstream analyses.
C. Availability of specimens with high molecular integrity. Although genomic DNA is
relatively stable and unaffected by variables in clinical specimen processing and
storage, the same is not true for cellular RNA and protein. Cellular RNA and
protein may be degraded in cells ex vivo, if clinical specimens are not rapidly
and properly processed and stored (Am I Clin Pathol. 2002;118:733). More
insidiously, the mRNA and protein complement of cells may rapidly change
as a function of processing time and methodology(] Clin Oncol. 2006;10;24:
3763).
D. Maximal utilization of diverse biospecimens. Because of the number and diver-
sity of potential biomarkers that can be evaluated from banked biospeci-
mens, repositories should contain a wide variety of specimens in a format
that can be distributed as widely as possible. For example, preoperative serum
914
Chapter 62 • Biospecimen Banking I 9 15
from cancer patients may be useful for identifying tumor-associated serum

markers for early detection, while nucleated cell fractions from bone mar-
row, peripheral blood, or cavity washings from the same patient may be used
to evaluate markers for tumor metastasis detection (Figure 62-1 ). Collected
specimens may be rare or small in size, so strategies to efficiently consoli-
date specimen resources (such as tissue microarrays [TMAs]) or to molec-
ularly amplify limiting amounts of genomic material from needle biopsies,
tissue touch preps, or washings are critical to providing a useful specimen
resource.
E. Biospecimen annotation. Given the cost and effort associated with next gen-
eration sequence analyses of clinical specimens (or proteomic or metabolomic
analysis), molecular data associated with human biospecimens is arguably more
valuable than the specimen itself. A data system that can accurately track indi-
vidual samples within large clinical specimen sets and that allows for accurate
integration and correlation of sequence, expression, biomarke.r; and clinical data
is obviously crucial. A large and viable specimen resource is only useful if it is
linked to complete and accurate clinical data that can be used to substantiate
or refute clinical hypotheses. At the same time, increasing concerns for medical
record privacy and, in particula.r; genetic data privacy require proper measures
to protect patient confidentiality.
F. Biospecimen custodianship. There have been increasing concerns regarding own-
ership of tissue specimens (Clin Chem. 2010;56:1675) and the intellectual
property that is generated from their use. There is also a continuing need to
appropriately honor the intent and privacy of patient donors. This requires
the establishment of an unbiased agent who can judiciously administrate and
regulate the collection and distribution of specimens.
The biospecimen bank is such an integral part of medical research, and is
becoming such an integral part of clinical medicine, that several national and
international agencies, such as the National Cancer Institute (NCI), have cre-
ated dedicated offices and guidelines for bank operation (http://biospecimens.
cancer.gov). The principles of biospecimen banking obviously relate to the gen-
eral practice of pathology and, therefore, it is almost exclusively the pathologist
and the pathology department that have sufficient expertise to govern these activ-
ities. The development and maintenance of a biospecimen resource involves many
policies and processes which are outlined in Figure 62-2, summarized in this chap-
te.r; and described in more detail elsewhere (Br j Cancer. 2004;90:1115; Methods
Mol Bioi. 2010;576:1).
II. BIOSPECIMEN COLLECTION. The types of biospecimens collected and the methods
used to process and store them depends greatly on their projected use, although
for many specimens the intended use is unknown. Since the cost associated with
biospecimen procurement and storage is not insignificant ($20 to $150 per speci-
men), careful consideration should be given to the scope and focus of the bank's
operation (Clin Trans/ Sci. 2009;2:172).
A. Defining the scope of operation. The type and number of participants from
whom biospecimens are collected depends upon the stated mission and avail-
able resources of the biospecimen bank. In some cases, small banks may collect
only defined specimens from participants enrolled in specific clinical trials. In
other cases, the bank rna y be disease-based and seek to generically collect all
available tissue specimens for a given disease type. When resources or institu-
tional barriers do not permit the creation of a large centralized facility, several
small banks may elect to create a federated system where each bank operates
independently, but is linked by a common informatics network (Adv Exp Med
Biol. 2006;587:65). A biospecimen bank need not be a "bank" at all, but may
simply serve to collect, process, deidentify, and immediately distribute biospec-
imens; the Cooperative Human Tissue Network frequently operates as such a
....
C.D
O'l
I >
I >
~
r==-:>
,[l
•> ..
~
I > u
Figure 62.1 From a single participant, a biospecimen resource may collect multiple specimen types and corresponding data to enable
a wide variety of translational research projects. A corresponding informatics system is often required to track and maintain these data.
q ---+
q - .__
q I
~ ~ ~ ~ ~
co
....
...,
Figure 62.2 Flow diagram of the many processes and resources required to operate a typical full service biospecimen resource
tissue broker (Cancer Epidemiol Biomarkers Prev. 2009;18:1676). A biospeci-

men bank may also simply exist as a more formal representation of diagnostic
paraffin block specimens already available in the pathology department.
B. Regulatory requirements. Biospecimen banking is going to be a necessary ancillary
activity to provide tissue for emerging genomic, proteomic, and metabolomic
testing methods. Several regulatory agencies have therefore produced guidelines
for clinical biospecimen banks (e.g., www.cap.org) to support direct patient
care.
Howevet; many aspects of biospecimen banking are considered a research
activity and must conform to regulatory requirements that are different than for
a clinical laboratory. Policies have evolved significantly at the national level over
the past 5 years, and also vary greatly between states and institutions. Generally,
the following points need to be considered with regard to biospecimen-based
human subjects' research.
1. Institutional Review Board (IRB} review. The intended purpose, scope, and
policies of the biospecimen bank must be reviewed by the IRB. In some cases,
a Certificate of Confidentiality, a document that asserts the right of the tissue
bank director (or honest broker) to protect the confidentiality of biospecimen
data even under court ordet; may be required (Genet Test. 2004:8:209).
2. Participant consent. In most cases of prospective biospecimen collection, some
form of informed participant consent is required. Explicit informed consent
may be waived if it is impossible or impractical to obtain and the risk to
the participant is minimal. Rarely, some institutions have ruled that generic
language present in a hospital admissions document or surgical consent form
provides sufficient consent for biospecimen collection, assuming that the
specimens are distributed and utilized in a de-identified or anonymous man-
ner. Generally, howevet; explicit written consent for biospecimen banking
should be obtained from the participant. Many generic templates for the lan-
guage used in such a document are available (] Clin Pathol. 2006;59:335).
Since the main risk to an individual participating in a biospecimen banking
program is loss of confidentiality, measures used to protect such confidential-
ity and the risks associated with this loss are the main risks to convey to the
participant. Howevet; when human biospecimens are being used for genomic
and disease-predisposition studies, and when research results from biospec-
imen use generate patentable biomarker inventions, properly documented
more specific informed participant consent may be critical.
3. Investigator agreements. In addition to IRB requirements that must be satisfied
by the biospecimen bank for the initial collection, coding, and storage of
biospecimens, investigators seeking to utilize biospecimens from the bank
may need to meet additional regulatory requirements, which usually requires
approval of an independent IRB protocol. In addition, investigators may
be expected to sign a Data Use Agreement or other investigator agreement
that establishes what will or will not be done with distributed biospecimens.
If specimens are to be distributed to an investigator outside of the bank's
institution, a material transfer agreement (MTA) may also be necessary.
c. Specimen types
1. Tissues. Collected tissues may be snap-frozen or fixed in a variety of cross-
linking or precipitating fixatives. Snap frozen tissue provides the highest qual-
ity protein and nucleic acid derivatives, and is often required for genomic
or proteomic studies. Howevet; the logistics of frozen tissue collection are
difficult and proper storage is relatively expensive. Formalin-fixed, paraf-
fin embedded (FFPE) tissue blocks are easy to obtain, easy to store, and
very familiar to any pathology service. The quality of molecular derivatives
from such tissues is vastly inferiot; although several recent technical advances
allow FFPE tissue to be used for the generation of DNA sequence and RNA
expression data (PLoS One. 2011;11;6:e17163; J Mol Diagn. 2011;13:

325). Fixation of tissue in precipitating fixatives such as ethanol, acetone, and
others (Mod Pathol. 2001;14:116; Diagn Mol Pathol. 2011;20:52), followed
by embedding in low temperature polymers(] Mod Diagn. 1999;1:17), pro-
vides tissue with excellent histologic detail and molecular material that is
superior to that of traditional FFPE tissue (e-Fig. 62.1).* However, RNA and
protein quality still do not match that of material derived from frozen tissue.
2. Blood components. Serum and plasma (often collected at multiple time points
throughout a patient's clinical course) are often banked. They are the pre-
ferred biospecimen for proteomic analysis, but when properly aliquoted from
multiple patients from multiple time points, can occupy a large amount of
storage space. Peripheral blood leukocytes are the ideal source for germline
DNA, and can be stored or processed without the need for additional cell
separation protocols such as mononuclear cell isolation. Bone marrow may
also be banked, particularly for analysis involving hematologic malignancies,
other bone marrow dyscrasias, or occult tumor cell detection.
3. Other body fluids. Urine, sputum, cavity lavages, and effusions serve as useful
analytes. However, large fluid volume specimens (such as urine) may be diffi-
cult and expensive to process and store for long periods of time, particularly
for a future unknown use. Furthermore, analysis of such biospecimens may
require only a small amount of material per assay, which then necessitates that
collections be excessively aliquoted to prevent multiple freeze/thaw cycles of
the same specimen. As an alternative, the cellular component of these fluids
may be isolated by centrifugation and stored as cell pellets.
D. Collection methods. Depending on the biospecimen type and its intended appli-
cation, there are a number of important considerations involved in collecting
specimens for a biospecimen bank.
1. Specimen acquisition. For most general tissue bank activities, tissue specimens
are collected in the course of routine treatment, such as a surgical resection
of a solid tumor, a therapeutic needle aspiration to remove joint or body cav-
ity fluid, or a diagnostic bone marrow biopsy. In these cases, it is important
that biospecimen collection for research purposes do not compromise the
diagnostic utility of the specimen. In many cases, such as the resection of a
small breast tumor or malignant prostate gland, it is important that biospec-
imen procurement occur under the direct guidance of a pathologist who can
supervise and assure that diagnostic integrity (e.g., an intact tumor margin)
is not compromised. In cases where the diagnostic tissue specimen is limit-
ing, properly supervised ex vivo sampling by core needle biopsy instrument
or skin punch may be used to acquire sufficient material for research with-
out compromising the diagnostic specimen. Other protocols may allow for
redundant tissue sampling at the time of a diagnostic procedure; for example,
for a patient with a suspicious liver mass, standard of care may dictate that
two specimens from a CT-guided needle biopsy be submitted for diagnostic
evaluation, while two additional needle cores may be taken for the research
tissue bank. In certain cases, the research bank may receive tissue samples
that harbor clinically relevant diagnostic findings that are not present in the
routinely processed tissue samples; for this reason, every pathology depart-
ment will need to develop policies regarding clinical versus research specimen
collection, which may include holding all research specimens in escrow until
a final clinical diagnosis is rendered. Finally, an IRB-approved study pro-
tocol may require the collection of biospecimens above and beyond what is
needed for routine standard of care; for example, a tissue biopsy or peripheral
blood specimen collected before and at multiple time points after a course of
therapy.
2. Biospecimen preservation. For molecular and genomic analyses, proper tissue
preservation is critical. A specimen's molecular profile (i.e., gene expression,
protein phosphorylation) can rapidly change from the time it is first col-
lected until the time when it is ultimately preserved by freezing or fixation
(J Clin Oncol. 2006;10;24:3763) due to so-called "warm ischemia time".
Although few evidence-based guidelines exist, warm ischemia time should
be limited and at least documented so as not to introduce additional prean-
alytical variability. Although snap-frozen tissue is the preferred substrate for
many studies, several commercial and "home-brew" tissue preservatives are
available that provide acceptable biospecimen preservation (BMC Genomics.
2004;5:88). Delays in blood processing can affect proteomic biomarker pro-
files (Expert Rev Proteomics. 2006;3:409); unlike tissue, whole blood cannot
be immediately snap frozen, but must be centrifuged to remove the spe-
cific plasma or serum component which then, in turn, must be aliquoted
and rapidly frozen. As with tissue, a number of commercial products are
available to preserve blood cell components at ambient temperature without
freezing, although these products are relatively expensive and require special-
ized downstream isolation procedures (Cytometry A. 2004;59:191; Physiol
Genomics. 2004;19:247).
3. Remote site collection. For multi-institutional research protocols, it may be
necessary to collect biospecimens from remote sites, often from health cen-
ters or medical offices which have little experience in dealing with intricate
biospecimen requirements such as snap-frozen tissue. Proper and efficient
collection of biospecimens from such sites may require preassembled spec-
imen procurement kits (e-Fig. 62.2) and personnel training to ensure the
uniform collection of biospecimens.
4. Specimen quantity. New molecular amplification strategies such as PCR,
whole genome amplification, and transcript amplification allow investigators
to use nanogram quantities of DNA and RNA for analysis. Instrumentation
for proteomic analysis has also become considerably more advanced and,
although it is not possible to amplify protein analytes from biospecimens,
tissue requirements have been minimized even for these technologies. There-
fore, even limited size biospecimens such as needle aspirates, tissue touch
preps, and core biopsies are valuable and frequently amenable to molecular
analyses. Obviously, however, such specimens must be judiciously distributed
to maximize their research potential. Conversely, the collection of large tissue
specimens is not necessarily desirable; for example, large resected carcinoma
specimens are often not properly preserved when fixed or frozen in their
entirety. For such cases, tissue specimens should be divided into samples no
> 1 cm3 for fixation and/or freezing.
5. Associated data collection. Clinical and pathologic data corresponding to
the patient and specimen (for clinical applications), and the participant and
specimen (for research applications), are as important as the specimen itself.
Without these associated data, the utility of the specimens for clinical or
correlative studies is lost. The scope of the data that are collected and stored
with the biospecimen will depend upon the mission of the biospecimen bank.
It is usually impractical to gather detailed clinical information represented
as written notes in a clinical chart or report; instead, it may be more efficient
to rely on electronic sources of data. Although data are usually represented
in text format, surgical pathology reports provide an accurate and detailed
source of pathology information for tissue specimens that are collected and
diagnosed as part of routine clinical care. In some centers, an electronic
medical record may provide access to basic patient demographics and clinical
diagnostic data. For cancer patients and their specimens, the hospital tumor
registry is often a reasonably detailed and standardized source for cancer-
related clinical and pathology data. Except for specific studies, it is generally
advisable to collect a limited but standardized data set corresponding to each
collected case, rather than an exhaustive (and inevitably incomplete) data set
for every specimen.
E. Biospecimen data management. As depicted in Figure 62.1, biospecimen banks
may house a complex array of specimens and associated data. It is critical that
specimens be accurately tracked and annotated if they are to be useful (Cancer
Inform. 2008;6:127). While smaller banks may rely on written log books and
electronic spreadsheets to track specimen information, these methods become
rapidly constraining as the bank becomes larger and more diversified. Basic data
types that may be required in a biospecimen bank information system include
the following:
1. Receipt, study association, and consent tracking. An accurate record must
be maintained of when and from where biospecimens were received, and
whether they are intended for clinical or research use. For the lattet; the
intended protocol must also be recorded, as well as the consent under which
the specimens were collected, since the consent likely regulates how the spec-
imens may be used.
2. Inventory and storage. It is important to accurately maintain the availability
and storage location of every specimen so that it can be rapidly retrieved on
demand.
3. Quality assurance. Quality assurance measures such as tissue histology review,
warm ischemia time, details of specimen processing, and nucleic acid quality
should be maintained for each specimen.
4. Clinical annotation. As discussed above, a minimal set of clinical and pathol-
ogy annotation data should be associated with each specimen.
5. Distribution and utilization. The efficacy of a biospecimen bank is judged
almost entirely upon its distribution of biospecimens for clinical testing or
productive translational research. Therefore, detailed data concerning spec-
imen distribution is essential in justifying the activity and operation of the
bank.
Several commercial software packages are available to assist in the man-
agement of biospecimen banks. More frequently, individual centers develop
their own data systems, often creating rather arbitrary and custom data
schemes and data definitions. While such applications may serve the imme-
diate needs of the bank, they present a significant obstacle to collaboration
among different biospecimen resource centers. To address this challenge, the
NCI's Cancer Biomedical Informatics Grid (caBIG) program has fostered
a group of pathologists, biologists, and informatics specialists working to
develop data standards and tools for pathology and biospecimen resources
(https://cabig.nci.nih.gov/workspacesffBPT). Among these tools is the caTis-
sue software application, a freely available web-based tool for managing
biospecimen inventory (https://cabig.nci.nih.gov/tools/catissuesuite).
Ill. BIOSPECIMEN PROCESSING. The same principles of specimen processing used in
the clinical laboratory apply to biospecimens banked for clinical and/or research
purposes. Howevet; since specimens collected for research purposes may be used
for experimental studies that are not a part of traditional diagnostic histopathology,
special specimen processing needs must be considered.
A. Frozen tissue. Although the histologic quality of frozen tissue is inferior to that of
FFPE tissue, most molecular studies require high grade DNA, RNA, and protein
that can only be obtained from frozen specimens. Tissue may be frozen in liquid
nitrogen, an isopentane cryobath (such as is available in most pathology frozen
section rooms), or a make-shift dry ice ethanol bath. Tissue specimens should
be no > 1 cm3 to ensure rapid and consistent freezing throughout the specimen.
If the specimen is to be used for histologic sectioning for quality review or
laser microdissection (LM), it may first be embedded in freezing media such as
optimal cutting temperature (OCT) compound. Since OCT material does not
interfere with nucleic acid isolation, but may have an effect on proteomic studies,
it may be advisable to freeze both embedded and nonembedded tissue for the
widest range of downstream uses. Once frozen, tissue may be stored in cassettes
in -80° ultralow freezers or liquid nitrogen (LN2) vapor inventory systems.
While the latter is more expensive and difficult to maintain, specimens stored
in LN2 are immune to the power failures and mechanical breakdowns that can
occur with electric freezers. Any frozen storage system should be equipped with
a temperature recording device to document appropriate storage conditions and
a remote alarm system that can contact the laboratory supervisor in the event
of machine failure or power loss.
B. Fixed tissue. FFPE tissue specimens are not as well suited for molecular and
genomic analyses as frozen tissue. However, the quality of fixed tissue specimens
can be improved using several quality control measures.
1. Control and documentation of fixation and processing times. By minimizing the
time for which tissues are fixed, over-fixation (which leads to excessive anti-
gen crosslinking and molecular degradation) can be avoided. Documentation
of processing times for each specimen can remove this preanalytical variable
from tissue specimens collected over different time periods or from different
locations.
2. Use of 11 mDiecularly friendly" fixatives. Several commercially available and
"home brew" precipitating fixatives preserve tissue and tissue histology while
minimizing the damaging effects to protein antigens and nucleic acids, caused
by cross-linking fixatives such as formalin (Mod Pathol. 2001;14:116; Diagn
Mol Pathol. 2011;20:52).
3. Vacuum storage. Although there are no convincing data to suggest an
added benefit, many biospecimen repositories recommend that paraffin tissue
blocks and, more importantly, cut unstained sections, be vacuumed, sealed,
and stored at 4o. The premise of this storage approach is that antigenicity
and nucleic acids are protected from further degradation by both removal of
oxygen and reduced temperature.
C. Blood components. For isolation and storage of serum and plasma, whole blood
must be immediately spun and the appropriate liquid component removed from
cellular material, aliquoted, and frozen. Processing must be performed rapidly
before cell lysis and protein degradation occurs. Consortia of proteomics inves-
tigators have published recommendations on blood processing for proteomic
studies (Expert Rev Proteomics. 2006;3:409) and while some of these guide-
lines are evidence based, robust guidelines have not been firmly established.
Many institutions freeze whole blood at the site of collection as a convenient
way to store blood for future genomic DNA isolation; however, freezing blood
in glass Vacutainer tubes presents a safety hazard, requires considerably more
freezer space to store, and inevitably causes cell lysis which results in DNA that
is frequently lower in yield, degraded, and contaminated with heme products
that can effect downstream assays. Although more labor intensive, immediate
spinning of whole blood followed by selective removal, washing, aliquoting, and
freezing of the huffy coat creates a high-quality specimen that is easy to store
and results in higher DNA yield and quality. In some cases, it may be desirable
to isolate the peripheral blood mononuclear cell (PBMC) fraction and preserve
these cells; this approach has the advantage of removing peripheral blood gran-
ulocytes and preserving the monocyte fraction which can be manipulated for
future uses such as flow cytometry and cell immortalization. However, viable
preservation of PBMC is expensive, requires additional expertise, and is seldom
necessary for simple DNA isolation. When PBMC isolation is not a considera-
tion, whole blood may be stored at 4° for as long as 72 hours with little loss in
PBMC viability (Ann Epidemiol. 2000;10:538).
D. Laser microdissection. For some types of genomic analysis, it may be desirable to
have material derived from homogeneous cell populations. Laser microdissec-
tion (LM) is a method that makes it possible to use a laser to either "capture" or
cut the desired cells away from surrounding tissue under direct microscopic visu-
alization (e-Fig. 62.3). The isolated cells may then be used for DNA, RNA, or
protein isolation for molecular analysis (Acta Histochem. 2007;109:171; Meth-
ods Mol Bioi. 2005;293:187). Although LM instrumentation is expensive, and
the technique is time-consuming and requires expertise, it has become routine
in many biospecimen banks.
E. Tissue microarrays. TMAs provide another format to maximize utilization of
limiting tissue specimens (Methods Mod Med. 103:89, 2005). TMAs are con-
structed by sampling one to three 0.8 mm (up to as large as 3 mm) tissue cores
from a donor paraffin tissue block, and assembling them into an array of as
many as 300 cores in a recipient block (e-Fig. 62.4). The resulting recipient
block can then be sectioned as a traditional tissue block, making it possible to
perform simultaneous immunohistochemistry or fluorescence in situ hybridiza-
tion on each of the 300 cores on a single slide. TMAs allow researchers to
easily validate patterns of biomarker expression in a large sample series using
relatively inexpensive technology. Few special reagents are required and basic
TMA instrumentation is relatively inexpensive, but expert review and informat-
ics tracking of the 300 cores per slides is required. A recognized disadvantage
of the TMA format is that it is prone to sampling error as only a small sample
from each tissue block is incorporated into the array.
F. Nucleic acid. Genomic technologies have shifted requirements from fixed tissue
sections to derivative nucleic acids, and so many biospecimen banks now pro-
duce DNA and RNA for research investigators. Nucleic acid can be effectively
derived from frozen and fixed tissue using several standard protocols, although
the quality from the later is often inferim; as discussed above. In some cases, it
may be desirable to isolate nucleic acid directly from collected specimens, such
as DNA from peripheral blood. Nucleic acids (RNA and DNA) are more stable
once isolated and can be more conveniently stored as compared to the parent
tissue or fluid. However, the initial cost and effort to generate DNA and/or
RNA from every banked specimen as it is received is great, and so prospective
preparation of nucleic acids should only be considered when material will be of
immediate use.
IV. QUALITY ASSURANCE. A biospecimen resource should only provide material that is
subject to rigorous quality control. In fact, one of the main reasons for a pathologist-
supervised biospecimen resource is that it is pathologists who best understand the
approach to and importance of clinical specimen quality control.
A. Specimen identification. Just as in any clinical laboratory, proper specimen label-
ing and identification are critical, because a biospecimen bank may store biospec-
imens for prolonged periods of time; maintenance of proper specimen identifi-
cation is important. As biospecimen banks become subject to regulatory review
as with any clinical laboratory (as noted above), they will need to implement
standard clinical laboratory practices to prevent sample mislabeling including
use of preprinted barcode labels, double data entry, and routine data auditing.
B. Representative tissue sampling. In most cases, biospecimens that are submitted
for clinical diagnosis and those that are submitted to a bank are not identical.
They may be collected at slightly different times or locations, or may involve
sampling bias. For example, in a heterogeneous prostate tumor, a tissue biopsy
banked for research may contain no neoplastic cells although the routinely
processed tissue for diagnosis shows adenocarcinoma; distribution of the banked
specimen under the assumption that it truly reflects the diagnostic material may
result in a compromised research study. Consequently, it is important that indi-
vidual biospecimens received by the bank undergo individual diagnostic review,
either prospectively or prior to distribution. Such a review of tissue or cellu-
lar samples may involve confirmation of a histopathologic diagnosis as well
as notation of general cellular features such as histologic preservation, tissue
cellularity, and necrosis. A special circumstance arises when a novel finding or
diagnosis is observed in a banked research specimen that was not reported from
the material for diagnosis; resolution of such a problematic circumstance will
depend upon the policies of both the IRB and pathology service, and should be
documented as part of the bank's standard operating procedures.
C. Tissue preservation and molecular integrity. When the bank will be responsible
for generating and distributing nucleic acids (DNA and RNA), it is important
that molecular integrity is verified. As discussed above, the quality of nucleic
acid is directly related to the manner in which the biospecimen was collected and
preserved. Fixation and delays in specimen processing (warm ischemia time) can
compromise nucleic acid quality as can repeated freeze/thaw cycles (e-Fig. 62.5).
Inherent necrosis and tissue cellularity in the specimen can also affect the quality
of nucleic acids. Documentation of molecular specimen quality can be accom-
plished by readily available (albeit often expensive) instrumentation designed for
quality review of small amounts of nucleic acid, including fiberoptic spectropho-
tometers and capillary microelectrophoresis systems. Metrics have been devel-
oped which relate quantitative results from these instruments to expected per-
formance in downstream applications and assays (BMC Mol Biol. 2007;22:8).
V. BIOSPECIMEN UTILIZATION. The success of any biospecimen bank is measured by the
utilization of collected biospecimens. Distribution of biospecimens for patient care
or funded research projects is also an important revenue source to support the larger
biospecimen banking effort. Although there is often a 3 to 5 year lag phase before
a new biospecimen bank is routinely utilized, while the bank is developing and
maturing its collection, a fundamental principle of biospecimen banking is that
the collection should be biased in favor of those biospecimens that will be used
for funded research projects. Measures which can help promote the productive
utilization of a biospecimen resource include the following.
A. Advertisement Df available resources. This is best accomplished through a web-
based catalog of available sources.
B. Streamlined administrative processes. Often, clinics and researchers are unfa-
miliar with procedures for requesting specimens, obtaining appropriate IRB
approval, and creating necessary MTAs. Having a standardized and facilitated
process for dealing with regulatory steps necessary for biospecimen banking
and subsequent distribution will greatly enhance utilization of a biospecimen
resource.
C. High quality annotated specimens. Clinical testing and translational research can
only be accelerated when the biospecimens distributed are properly quality con-
trolled and "assay ready." Minimal but sufficient clinical and/or pathology data
should be supplied with each specimen in order to easily enable the correlative
analysis.
D. Facilitated and judicious distribution. The process of case selection, specimen
retrieval and processing, specimen annotation, and distribution can be time-
consuming and expensive. As a biospecimen bank matures, increasing requests
may overtax the bank's available resources and can result in significant delays
in providing specimens. Competing interests for limited biospecimens may also
become problematic as bank utilization increases. Therefore, a biospecimen
bank should establish a utilization review committee so that the finite effort
and physical resources of the bank can be fairly allocated to the clinical and
research committees. These committees should be an unbiased representation of
pathologists and scientists, with allocation decisions based on clinical necessity,

scientific merit of proposed projects, investigator track record with previous
requests, funding status, and likelihood that distributed biospecimens will mean-
ingfully contribute to patient care or grant or scientific publication development.
VI. SUPPORT. The biospecimen bank must find its own means of financial support.
Frequent sources of support include the following:
A. Clinical activities. Biospecimen banking to support direct patient care (i.e., for
diagnosis and to direct therapy) should be compensated in a way consistent with
its value. Possible mechanisms include the following:
1. Hospitals. The costs can be borne by hospitals, since clinical biobanking is
part of the general hospital infrastructure required to support best patient
care.
2. Specific billing codes. Clinical tissue banking can be compensated via estab-
lishment of specific billing codes that reflect the technical components (spec-
imen handling, quality assurance, informatics, and storage) and professional
components (selection of appropriate tissue, microscopic review of banked
specimens) of biospecimen banking. Such codes currently do not exist.
B. Research activities
1. Institutional support. Academic institutions or departments often support the
initial development and operation of a biospecimen bank, which is not sur-
prising considering the central importance of the bank in the mission of trans-
lational research. In return, the institution expects a return on its investment
in terms of the number of biospecimen-based projects, publications, and
research grants that can be enabled through such a resource.
2. Extramural funding. Unlike traditional, hypothesis-based research grants,
there are a few extramural (noninstitutional) funding mechanisms to specif-
ically support biospecimen banks. A bank or tissue procurement shared
resource facility may, howevet; be a key funded component of larger initia-
tives such as NIH program project grants, center grants, or disease-focused
programs (e.g., Specialized Programs of Research Excellence, or SPOREs).
3. Investigator fees. A viable specimen resource will need to develop investigator
fees for the services it provides, including biospecimen collection, storage, and
processing. These fees should be calculated realistically and take into account
all operations and resources that are required from the initial collection of
a biospecimen to its final distribution to an investigator. Such fees should
be structured to also allow for bank growth and further development. At
the same time, fees should not be so burdensome as to deter the conduct of
sound research, particularly for pilot translational studies where investigator
funding may be limited, or where a unique opportunity exists to collect
biospecimens (perhaps in the context of a clinical trial), store them, and
utilize them at a later date once additional research funding is obtained.
Obviously, the greater a bank's participation in funded research, the easier it
will be to maintain long-term support for the operation, and the more likely
it will be that the bank will significantly contribute to translational research.
SUGGESTED READINGS
Handbook of Human Tissue Source: A National Resource of Human Tissue Samples. Elisa Eise-
man and Susanne B. Haga. RAND Corporation. ISBN: 0-8330-2766-2. http://www.rand.org/
pubs/monograph· reports/MR954/.
NCI Office of Biorepositories and Biospecimen Research. http://biospecimens.cancer.gov.
NCI Specimen Resource Locator. http://pluto3.nci.nih.gov/tissue/expediter.cfm.
International Society for Biological and Environmental Repositories. http://www.isbe~:orgl.
Imaging Technologies
in Surgical Pathology:
Virtual Microscopy and
Telepathology
Jochen K. Lennerz, Michael Isaacs, Erika Crouch,
and John D. Pfeifer
I. CONVENTIONAL LIGHT MICROSCOPY remains the core tool in diagnostic surgical

pathology. The future of surgical pathology will, however, include electronic modes
of image presentation to support diagnoses rendered by viewing the cells or tissue
on a computer screen rather than with an optical microscope, since digitized images
of glass slides increase the clinically useful information that can be obtained from
a pathologic specimen, and permit modes of analysis including electronic consul-
tations, morphometry, and slide storage that go beyond the capabilities of current
microscopy-based diagnostic pathology.
II. VIRTUAL MICROSCOPY is the technique whereby entire glass slides or selected areas
of slides are scanned and converted to digital data files (also known as virtual
slides), which can then be viewed on a computer screen. The characteristics of the
displayed image, in terms of resolution and range of magnification, are primarily
determined by the optical features of the scanning system and are overall compara-
ble with those of glass slide microscopy. However, in contrast to conventional light
microscopy where the magnification, focus, and condenser setting can be adjusted
at any time, for digital scanning, the "scanning depth" must be determined prior to
image acquisition. The scanning depth includes the region of interest, the scanning
power, and the number of horizontal levels to be obtained in the plane of the tissue
section (scanning power includes the physical magnification [e.g., 20x] and reso-
lution [e.g., 2048 x 4800 pixels], while the number of horizontal levels [so-called
z-stacking] is dependent on the section thickness and the need to be able to focus up
and down through the tissue and cells in the final virtual image). Depth of focus is a
requirement for interpretation of specimens that rely on a three-dimensional assess-
ment of cellular morphology, for example, cytology specimens (Cancer Cytopathol.
2007;111:203).
Average scanning time for a single horizontal level of a slide (non-z-stacked) is
about 5 minutes and generates a data file that is enormous (ranging from about 200
MB to 1.5 GB). Unfortunately, no uniform (open-source) virtual slide file format
is currently available, although interfaces are available that can convert image files
between the different virtual slide formats.
Each virtual file consists of multiple parts, called file segments, which include an
identification tag, a scanned barcode linked to the laboratory information system
(LIS), a low-power overview, and the images at the selected power. While image
acquisition methods vary (linear, meander, array), currently all available systems
produce tiled images, that is, small images that are retrieved and stitched together
to form the final image on the computer screen at the selected powe.t Most inter-
faces allow electronic zooming (i.e., additional magnification) beyond the original
scanned powe.t
Resolution and functionality of virtual slides have achieved levels that are com-
parable with conventional light microscopes. Scanning time, on the other hand, is
still time-consuming, since scanning time is mainly dependent on computational
926
Chapter 63 • Imaging Technologies in Surgical Pathology I 9 27
speed and optical physics, the latter of which is determined by the magnification
of the scanning objective. The actual image acquisition step occurs via a charge-
coupled device (CCD), which consists of several hundred thousand individual pic-
ture elements (pixels) that transform the optical image into a virtual image; whole
slide images are therefore acquired via a single optical lens that moves over the
slide. Some scanners shorten scanning time by using a meander rather than a linear
scanning pattern to acquire images, but both the meander and the linear meth-
ods have inherent physical limitations that become most apparent when acquiring
multiple images within each plane of section (i.e., when z-stacking). An emerging
technology, so-called lens array microscopy that uses arrays of detectors (miniatur-
ized lenses) to simultaneously capture information from larger areas of the tissue
section, may be able to markedly shorten scanning times but still meet the high
standards of diagnostic pathology (Hum Pathol. 2004;35:1303).
A. Setting up vinual microscopy in the surgical pathology laboratory requires an
electronic and organizational infrastructure as well as a slide scanning instru-
ment. The infrastructure requires an efficient interaction between information
technology (IT) and LIS personnel, and dedicated and trained technical person-
nel (an image technologist) are required to load and maintain the scanning
instrument, perform initial screening (and rescanning if necessary), monitor
scan quality, and distribute the virtual files in a manner that can be conve-
niently viewed by the pathologist (]. Pathol Inform. 2011;2:39). A file server
with the necessary storage capability (upgradeable tera- to petabyte range or
beyond), databases for the virtual files (linked and updated to the LIS), and
the necessary maintenance and updates must be managed by a knowledgeable
IT person (Human Pathol. 2003;34:968). For optimal use in routine patient
care, high-quality LCD monitors of sufficient size (at least 53 em diagonal)
for peripheral vision are required (Hum Pathol. 2006;37:1543), with extended
desktop computer functionality to be able to simultaneously view the LIS, the
gross description and gross images of the specimen, and the virtual slide viewer.
B. Diagnostic and clinical applications of virtual microscopy include diagnostic con-
sultations, archiving, primary diagnosis, and quality assurance (QA).
1. The use of virtual microscopy for diagnostic consultation (so-called
e-consultation) is one of the most useful applications of virtual slide
microscopy. In addition to convenience (no packing or mailing of slides is
required), there are several advantages to e-consultations. Electronic distri-
bution is virtually instantaneous and allows consulting pathologists to review
the scanned slides at any time and location. Online, real-time conferencing
and simultaneous viewing by two or more pathologists is easily achieved. In
addition, there are no risks of losing the primary data (i.e., tissue blocks or
glass slides).
2. Scanning of selected cases for archiving. Virtual slide creation of selected
slides sent in consultation (the medicolegal climate in the United States dic-
tates return of all diagnostic materials to the referring institution) makes
it possible for the consulting institution to have a permanent record of the
diagnostic slides. Such an archive enhances patient care by providing an
immediately available permanent record of the slides to guide frozen section
diagnosis, or final diagnosis at the time of definitive excision. In addition,
the virtual slides are available for subsequent clinicopathologic conferences
(Hum Pathol. 2010;41;751).
3. Primary diagnosis based on review of virtual slides has been shown to have the
same accuracy as primary diagnosis based on routine light microscopy (Arch
Path Lab Med. 2011;135;372), although the use of virtual slides for primary
diagnosis requires an extensive validation procedure. While the use of virtual
slides has dear applications for improved pathology services to underserved
areas (Natl Med] India. 2002;15:363; Ethiop Med ]. 2005;43:51), there is
little evidence that digitizing all slides in routine practice results in cost or
time savings (]Path Inform. 2011;2:39; Anal Cell Pathol. 2011;34:1).
4. QA programs may be enhanced by review of virtual slides from cases with diag-
nostic discrepancies. Files can be flagged for subsequent review, and trends
in diagnostic errors easily identified (Dis Mark. 2007;23:459).
C. Education. Virtual microscopy is already an indispensable tool for medical educa-
tion in pathology, including medical student teaching, teaching of pathology resi-
dents and fellows, and continuing medical education for practicing pathologists.
Many national pathology conferences and slide seminars now post virtual slides
on websites before the meeting, and some organizations are building a collection
ofteaching cases (e.g., see the United States and Canadian Academy of Pathology
[USCAP] Virtual Slide Box at www.uscap.org). Other examples of educational
activities based on virtual slides include teaching of histology to medical students
(Anat Rec. 2006;289B:128), a tutorial on Gleason grading of prostatic adeno-
carcinoma (Hum Pathol. 2005;36:3 81), didactic presentations using a combina-
tion of text and virtual microscopy (Ann Diagn Pathol. 2003;7:67), and online
virtual atlases (e.g., of breast pathology at www.webmicroscope.net).
Education and experience with interpretation of virtual slides is critical
for pathology residents and fellows; at the very least, it prepares trainees for
the American Board of Pathology virtual microscopy practice examination
(www.abpath.org/VMinstr.htm).
D. Research applications for virtual microscopy in surgical pathology include quan-
titative image analysis, and imaging of tissue micro arrays used to study patterns
of gene and protein expression. Virtual images can also provide documenta-
tion of specimens or tissue microarrays retained in tissue banks (Eur] Can-
cer. 2006;42:3110), or for patients enrolled in clinical trials. As for diagnostic
pathology, virtual slides make it possible to easily share specimens among inves-
tigators at different sites, anytime. Virtual slides also allow for the electronic
publication of whole slides rather than selected fields of slides, which facilitate
the transfer of new information to practicing pathologists.
Image analysis can be performed on any of the virtual slide file formats pro-
duced by the current generation of slide scanners, and widely accepted freeware
and a variety of subprograms are available to support clinical applications. The
most versatile solution for image processing and analysis is the Nlli freeware
named Image] (Biophot Int. 2004;11:36, available at http://rsb.info.nih.gov/ij/).
The versatility of this program derives from its open architecture that
allows users to implement small and typically customized subprograms (called
"plug-ins"). Over the years, many plug-ins (mostly research-derived) have been
developed; for example, 'Image J for microscopy' (www.macbiophotonics.ca/
imagej) includes numerous tools such as intensity and time analysis, particle
analysis, colocalization analysis, intensity processing, color processing, stack-
slice manipulation, z-functions, t-functions, deconvolution, and annotation.
Ill. TELEPATHOLOGY is the practice whereby pathologists render diagnoses from a dis-
tance by viewing electronically transmitted images rather than by examination of
the glass slides themselves by light microscopy. Electronic Images can be transmitted
by ordinary telephone lines, high-speed digital lines, or satellites, but increasingly
the images are transmitted via the Internet. With some overlap, three systems are
currently available: dynamic, static, and virtual.
A. Dynamic telepathology systems. In these systems, pathologists view images in real
time by electronic control of a distant robotic microscope that has motorized
optics and a motorized stage. Dynamic-robotic telepathology has been primarily
used to provide intraoperative frozen section diagnoses to hospitals without on-
site pathologists (Hum Pathol. 2007;38:1330). Typically, in less than a minute,
a digital overview of the slide is created; virtual controls enable the pathologist
Chapter 63 • Imaging Technologies in Surgical Pathology I 929
to remotely control the movement of the slide on the microscope. Reported

diagnostic accuracy is comparable to that of conventional light microscopy.
A variation of the dynamic method is the submission of a live ("streaming")
image from the remotely located microscope either with or without robotic
control; in the latter paradigm, manipulation of the slide and magnification are
simply controlled via instructions to the person using the microscope.
One important disadvantage of either of the dynamic telepathology method
is the extra time needed to review the virtual slides compared with standard
slides. In one study (j Neuropathol Exp Neurol. 2007;66:750), the lack of an
on-site presence affected every stage of the intraoperative consultation including
the gathering of patient information, gross specimen examination and handling,
frozen section and smear preparation, communication between various parties
involved, and documentation of the consultation process. Thus, implementation
of telepathology requires substantial planning, communication, and training of
both pathologists and support personnel.
B. Static telepathology systems use images that have been selected, stored, and
forwarded to the pathologist. Static systems are mainly used to obtain second
opinions on difficult cases. Overall, the diagnostic accuracy approaches conven-
tional glass slide optical microscopy. Problems leading to discordance include
field selection and poor image quality (Hum Pathol. 2003;34: 1228). For large or
complex specimens, the handicaps of preselected images are more pronounced.
C. Virtual slide telepathology. The use of virtual microscopy in telepathology has
been shown to have clinical utility, even in time-sensitive settings such as intra-
operative frozen section diagnosis (Hum Pathol. 2009;40:1070). However, the
scanning time to produce a whole slide image, the required infrastructure, and
the large file sizes may limit the usefulness of virtual slide telepathology in many
practice settings.
IV. EMERGING IMAGING-RELATED TECHNOLOGIES include spectral imaging (Cytometry A.
2006;69:735), nonconventional optical techniques, computer-aided detection (e.g.,
visual field tracking, pattern recognition, image segmentation), and the application
of artificial intelligence to image analysis. However, these techniques remain largely
experimental and have yet to be tested in routine surgical pathology.
V. OBSTACLES. Significant issues must be addressed to allow the routine implemen-
tation of virtual microscopy and telepathology in diagnostic surgical pathology.
First, there are unresolved licensure and reimbursement issues concerning diag-
nostic surgical pathology practice from a distance. Second, there is debate as to
whether virtual microscopy instrumentation is subject to governmental approval,
and whether users must be certified prior to rendering diagnoses using these tech-
nologies. Third, validation and incorporation of digital pathology images into the
electronic medical record are haphazard and not standardized (]. Biomed Opt.
2007;12:051801). Despite these uncertainties, there is little question that recent
advances in imaging technologies provide unique opportunities to improve patient
care.
SUGGESTED READINGS
European Virtual Microscopy network at http://www. webmicroscope.net
Pathology informatics at http://www. pathologyinfromatics .org
Index
Abetalipoproteinemia, 219 minimal deviation, 543
Abscess, 665, 782 mixed,526
Acantholytic dyskeratosis, 593 mucinous, 501, 543
Acanthoma, 595 non-dear cell, 401
Acanthosis, 191, 580 pancreas cytopathology, 29 5
Accessory spleen, 732 pancreatic ductal adenocarcinoma (PDA),
Accessory tragic congenital anomaly, 98 287-290
Accreta, 572 paratesticular tumors, 464
Acid fast bacteria (AFB) stain, 846-847 polymorphous low-grade (PLGA), 88-89
Acinar adenocarcinoma prostatic, histologic grading of, 479-480
prostate cancer, 474 renal pelvis cancer, 376
variants, 477-478 salivary glands, 88-89
Acinar cell serosal membranes, 188
carcinoma, 89, 96, 291, 296 serous,500,526,544
cystadenoma, 287 sinonasal region, 43-44
Acquired adrenal hyperplasia, 431-432 stomach neoplasm, 206, 208, 212
Acquired cystic kidney disease, 346, 364-365 ureter cancer, 376
Acral arteriovenous tumor, 602 urethra malignant neoplasms, 401
Acral myxoinflammatory fibroblastic sarcoma, urinary bladder, 393, 398
756 uterus epithelial malignancies, 526
Acrochordon,606-607 well-differentiated villoglandular, 544
Acrospiroma, 600 Adenocarcinoma in situ (AIS), 543, 549
ACTH-producing adenoma, 449 Adenofibroma, 352, 366, 531
Actinic keratosis, 595 Adenoid basal carcinoma, 90, 96, 544-545
Actinomyces, 5, 548, 551 Adenoid cystic carcinoma (ACC), 140
Acute disseminated encephalomyelitis Adenoma
(ADEM), 661 adult renal neoplasms, 365-366
Acute inflammatory demyelinating adrenal cortical, 432-433
polyneuropathy (AIDP), 678 adrenal glands, 445
Acute interstitial pneumonia (AlP), 107 atypical pituitary, 450
Acute motor and sensory axonal neuropathy basal cell, 86, 95
(AMSAN), 678 bile duct, 268
Acute motor axonal neuropathy (AMAN), 678 black (pigmented), 433
Acute tubular necrosis (AlN), 337 canalicular, 86-87
Acute vasculitis (inflammatory dermatoses), carcinoma ex pleomorphic, 91
587 congenital pleomorphic, 93
Adamantinoma, 658-659, 812 oorticotroph (ACTH-producing), 449
Addison disease, 430 ectopic adrenal cortical, 433
Adenoameloblastoma, 56 fibroadenoma, breast pathology, 324-325
Adenocarcinoma gallbladder and extrahepatic biliary system,
acinar, 474, 477-478 275
appendix, 241 gonadotroph, 44 9
biliary, 278 hepatocellular (HCA), 265, 272
cervical glandular neoplasms, 543-544 hyalinizing trabecular, 420
dear cell, 401, 526, 544, 556 large intestinal neoplasms, 233, 235
ductal, 327, 478 liver, 265-266, 2 72
endometrioid, 502, 522-526, 543 metanephric, 352,366
esophagus, 197-198,200 neoplasms of middle ear, 101
extrahepatic bile ducts, 278 nephrogenic, 400
fetal (of lung), 139 papillary, 365
gallbladder, 278, 280 parathyroid, 424-425
invasive, 543 pleomorphic (PA), 84, 95
large intestinal neoplasms, 236, 238 pituitary, 447-450
lobular, breast cytology, 327 salivary glands, 84, 86, 91, 95
lung, 128-132, 147 sebaceous, 600
mesonephric, 544 sessile serrated, 233, 241
microinvasive, 543 small intestinal neoplasms, 220
931
932 INDEX
Adenoma (Contd.) Adriamycin (doxorubicin) toxicity, 156-157

somatotroph (GH cell), 449 Adult fibrosarcoma (AFS), 755, 786
stomach neoplasm, 205 Adult granulosa cell tumors (AGCTs), 503
thyroid neoplasms, 409-410 Adult renal neoplasms, 357, 369. See also
urethra metaplasia, 400 Pediatric kidney neoplasms
urinary bladder, 383, 393 benign epithelial tumors, 365-366
Adenoma malignum, 543 carcinoma
Adenoma sebaceum, 606 Birt-Hogg-Dube syndrome, 360
Adenomatoid tumors, 181, 443, 463, 513, dear cell papillary RCC, 365
533-534 clinical diagnosis, 360
Adenomyoma of ampulla vater, 220 collecting duct carcinoma (CDC), 363
Adenomyosis, 531 constitutional chromosome 3
Adenosarcoma, 531, 545 translocations, 360
Adenosis, 325, 552 hereditary leiomyomatosis, 360
Adenovirus, 118, 252 histologic typing, 360
Adipocytic tumors mucinous tubular carcinoma, 364
benign renal carcinomas associated with Xp11.2
angiolipoma, 746 translocations, 364
chondroid lipoma, 746 renal cell cancer, 360-365
diffuse lipomatosis, 746 renal epithelial malignancies diagnosis,
hibemoma, 747 360
HIV-lipodystrophy, 747 renal medullary carcinoma, 364
lipoblastoma, 747 risk factors, 359-360
lipomas, 745-747 spindle cell carcinoma, 364
lipomatosis, 746-747 succinate dehydrogenase germline
myolipoma, 746 mutations, 360
nevus lipomatosus, 747 thyroid-like follicular carcinoma, 365
pelvic lipomatosis, 746 tubulocystic carcinoma, 364
pleomorphic lipomas, 746 unclassified RCC, 364
spindle cell lipoma, 746 Von Hippel-Lindau {VHL) disease, 359
steroid lipomatosis, 746 cystic nephroma, 368
symmetric lipomatosis, 746 germ cell tumors, 368
intermediate (locally aggressive), 747-748 gross examination and tissue sampling,
malignant 357-359
dediffl:rentiated LPS, 748 hematopoietic tumors, 36 8
myxoid LPS/round cell LPS, 748 histologic grading of RCC, 368
pleomorphic LPS, 748-749 lymphoid tumors, 368
retroperitoneum, 783-785 mesenchymal tumors, 366-367
soft tissue, 745-749 metanephric tumors, 366
Adnexal tumors, 601 metastatic tumors, 368
Adrenal cortical rests, 4 53 mixed epithelial and stromal tumor, 368
Adrenal glands nonneoplastic tumorous conditions
cortical renal malakoplakia, 370
hypofunction,430-431 renal tuberculosis, 371
neoplasms, 432-435 xanthogranulomatous pyelonephritis,
cysts, 444 370
cytology pathologic staging, 369
adrenal cortical neoplasms, 444-445 reporting, 369-370
fine needle aspiration (FNA), 444 Adult rhabdomyoma, 762
metastatic malignancy, 445 AIDS
myelolipoma, 444 bone marrow benign diseases, 712
pheochromocytoma, 445 infectious neuropathies, 681
extraadrenal paraganglioma lesions, penile infections in, 485
441-443 related cysts, 83
gross examination, 428-429 Airway rejection, 124
medullary lesions, 436-441 Alcian blue, 844
metastatic neoplasms, 444 Alcoholic liver disease (ALD), 253
normal anatomy and histology, 428 ALK mutations, 131
parenchymal lesions, 443 Allelic discrimination, 898
Adrenectomy, 429 Allergic bronchopulmonary aspergillosis
Adrenogenital syndrome, 433 {ABPA), 115
INDEX 933
Allograft kidney pathology. See under Kidney Angioma

Allograft livers, 261 cherry, 602, 767
Alport syndrome, 336 intramuscular, 767
Alveolar proteinosis, 114 littoral cell
Alveolar rhabdomyosarcoma (ARMS), 47, sclerosing angiomatoid nodular
764 transformation (SANT)
Alveolar soft part sarcoma, 777 soft tissue, 766-767
Alzheimer disease (AD), 632--670 spleen neoplastic disorders, 737-738
Amalgam tattoo, 8 tufted, 766
Amebic encephalitis (AE), 664-665 Angiomatoid fibrous histiocytoma (AFH), 774
Ameloblastoma, 54-57 Angiomatosis, 766, 768
Amiodarone, 110-111 Angiomyofibroblastoma, 554, 751
Amnion nodosum, 567-568 Angiomyolipoma, 356, 366-367, 371
Amniotic bands, 568 Angiomyxoma, 554
Amplification, 889 Angiopathy, 164, 668
Amplification bias, 892 Angiosarcoma
Ampullary carcinoma, 220 bone, 811-812
Amyloid cardiac, 158
angiopathy, 164, 668 epithelioid, 271
dermal deposits, 592 liver, 271-272
neuropathies, 680 skin, 603
Amyloidosis soft tissue, 769-770
cardiac, 156 spleen, 738
kidney, 335-336 Anorchism, 4 53
larynx, 23 Antibody, 853
liver, 255 cytopathology, 852-853
nodular pulmonary, 115 mediated rejection and cardiac transplants,
pulmonary, 115 159
tracheobronchial, 115 neuropathy, anti-Hu, 679
urinary bladder, 384 Antiglycolipid, anti.sulfatide, and
Anal biopsy, 217 antiganglioside neuropathies, 679
Anal canal Anti-Hu antibody neuropathy, 679
anal tag, 245 Antimyelin associated glycoprotein (MAG)
condyloma acuminatum, 245 neuropathy, 679
extramucosal (perianal) carcinoma, Antineutrophil cytoplasmic antibodies
246 (ANCA), 163-164
hemorrhoids, 245 a1-Antitrypsin deficiency, 255
hidradenoma papilliferum, 245 Antral vascular ectasia, 204
inflammatory doacogenic polyp, 245 Aortic valves disorders, 153
melanoma, 246 Aortitis, 338
nonneoplastic and neoplastic conditions, Aphakic bullous keratopathy, 68
244-246 Aplastic anemia, 711
normal anatomy, 215 Apocrine
Paget's disease, 246 changes, breast pathology, 324
sec, 245 cutaneous appendages tumors, 600
Anal squamous intraepithelial neoplasia cystadenoma, 600
(ASIN), 245 Apocrine tumors, 599--601
Anaplasia, 350 Apoplexy, 44 7
Androgen deprivation therapy, hormonal, Apoptosis, 631
479 Appendages (cutaneous) tumors
Anemia, 710-711 benign, 596
Aneuploidy, 878 apocrine tumors, 600--601
Aneurysmal bone cyst, 54, 814-815 eccrine tumors, 599-600
Aneusomies, 880 hair follicle tumors, 596-599
Angiectatic tumor, 774 sebaceous adenoma, 600
Angiitis, 668 sebaceous hyperplasia, 600
Angiocentric glioma (AG), 64 7 malignant, 601
Angioendothelioma, 769 Appendectomy, 217
Angiokeratoma, 602 Appendicitis, 241
Angioleiomyoma, 607,759 Appendix
Angiolipoma, 607, 746 normal anatomy, 215
934 I INDEX
Appendix neoplasms, 241 Aural sinuses, 98

adenocarcinoma, 241 Autoimmune hepatitis (Alli), 256
cystic fibrosis, 241 Autoimmune pancreatitis (AlP), 284
hyperplastic polyps, 241 Autosomal dominant polycystic kidney disease
neuroendocrine neoplasms, 241-244 {ADPKD), 345
sessile serrated adenoma/polyp, 241 Autosomal recessive polycystic kidney disease
signet-ring cell carcinoma, 241 (ARPKD), 345
Aranasal sinus mucoceles, 3 8 Autotransplantation, 424
Argentaffin staining, 848 Avidin-biotin method, 852
Argyrophil staining, 848 Axons
Arias-Stella reaction, 539 degeneration, 674
Array comparative genomic hybridization injury, 631
(aCGH), 869-870, 903 neuropathy, 678, 680
Arrhythmogenic right ventricular
cardiomyopathy, 156 Bacterial artificial chromosome (BAC) arrays,
Arteriopathy 903-905
cerebral autosomal dominant, 668-669 Bacterial infections
microscopic, 157 cervicitis, 537
transplant, 161 disease testing, 900
Arteriovenous hemangioma, 602, 767 intestinal, 219, 232
Arteriovenous malformations (AVMs), 667 liver, 253
Arteritis, 162-163 lung, 118-119, 146-147
Arthritis, 823-825 oral cavity and oropharynx, 5
Asbestosis, 121 Baker cyst, 825
Askin tumor, 142 Balanitis, 484
Aspergillosis, 115 Balanitis circinata, 484
Aspergillus, 119-120 Balanitis xerotica obliterans (BXO), 484, 593
Assays, 898-899. See also DNA sequence Balanoposthitis, 484
analysis Banked tissue, 837, 914, 918. See also
Astroblastoma (AB), 647 Biospecimen banking
Astrocytes, 632-633 Barrett esophagus, 194-195, 197, 200
Astrocytoma Bartholin
anaplastic (AA), 634, 638 abscess, 558
circumscribed, 642-643 cysts, 552, 560
desmoplastic infantile, 648 Basal cell carcinomas
diffuse (DAs), 634 epidennis, 596
Asymmetric neuropathies, 677 salivary glands, 86, 95
Asymptomatic inflammatory prostatitis, 4 70 scrotal, 489
Atypical teratoid/Rhabdoid tumors (AT/RT), Basal cell hyperplasia, 4 72
653 B-celllymphoma of mucosa-associated
Atopic dermatitis, 585 lymphoid tissue {MALT) type, 211-212
Atrophy B-celllymphomas
cervix, 538 incidence and epidemiology, 685
prostate, 4 70-471 lymph nodes cytopathology, 707
testis, 455 lymphoblastic leukemia {bone marrow),
uterus, 520 721
Atypia markers helpful in subclassifying mature
cytologic, 521 B-celllymphomas, 691
flat urotheliallesions with, 385 mediastinal large, 174
focal glandular (ASAP), 473 pathophysiology, 685-687
prostatic, 4 73 primarily B-cell associated markers, 691
reactive urothelial, 383 splenic marginal zone (SMZL), 735
thyroid cytopathology, 419 Bead microarrays, 905
Atypical adenomatous hyperplasia adenosis, Benign acquired melanosis (BAM), 64
472 Bergmann gliosis, 632
Atypical alveolar hyperplasia (AAH), 132 Bethesda System, 548
Atypical carcinoid Bielschowsky, 847
cervical, 545 Bile duct
lung, 135-136 adenoma, liver biliary origin tumors, 268
Atypical ductal hyperplasia (ADH), 322-323 gross examination, 274
Atypical lobular hyperplasia (ALH), 323 Bile pigment stains, 848
INDEX 935
Biliary intraepithelial neoplasia (BiliN), 269, aplastic anemia, 711

275 benign lymphoid aggregates, 712
Biliary origin tumors Chediak-Higashi syndrome, 713
benign, 268 erythrocytosis, 711
malignant, 269 Gaucher disease, 713
premalignant, 269 granulocytic left shift, 71 0
Bilobed placenta, 573 granulomas, 712
Binudeation, 631 hemophagocytic syndrome, 713
Biomarkers (breast cancer), 319-322 Hermansky-Pudlak syndrome, 713
Biopsy. See Nerve biopsy lipid storage disorders, 713
Biospecimen banking, 914 megaloblastic anemia, 710
accurately diagnosed biospecimens, 914 mucopolysaccharidoses, 713
annotation aspeects, 915 Niemann-Pick disease, 713
biospecimen data management, 921 serous degeneration, 711
biospecimen processing, 921-923 thrombocytosis, 711
biospecimen utilization, 924 gross examination and tissue sampling,
collection 709-710
methods, 919-920 malignant diseases
regulatory requirements, 918 CUJSLL and related disorders, 726
scope of operation, defining of, 915 eosinophilia neoplasms, 719
custodianship, 915 essential thrombocythemia, 718
financial support aspects, 925 fibroblast growth factor 1 receptor, 719
large biospecimen cohorts, availability of, leukemia, 717, 719, 721, 726, 728
914 myelodysplastic syndromes, 713-714, 716
maximal utilization of diverse biospecimens, myeloproliferative neoplasms, 716-719
914-915 platelet-derived growth factor receptors
quality assurance, 923-924 abnormalities, 719
significance to pathology and translational polycythemia vera, 717-718
research, 914 primary myelofibrosis, 718-719
specimen types systemic mastocytosis, 719
blood components, 919 normal gross and microscopic anatomy,
body fluids, 919 709
tissues, 918 Bone neoplasms
specimens availability with high molecular chondrosarcoma, 803-804
integrity, 914 iatrogenic disorders, 798
Biphasic pulmonary blastoma, 139 infectious disorders, 798
Birt-Hogg-Dube syndrome, 360 neoplastic and tumor-like lesions, 798
Black (pigmented) adenoma, 433 adamantinoma, 812
Bladder cuff, 374. See also Urinary bladder aneurysmal bone cyst, 814-815
Blastomycosis, 120 angiosarcoma, 811-812
Blood components, 919,922 benign fibrous histiocytoma, 809
Blue nevus, 539, 617 bone-forming tumors, 805-807
Bone cartilage-forming tumors, 798-799,
bone-forming osteoblasts, 816-817 802-804
bone-forming tumors, 805-807 chondroblastoma, 803
bone-resorbing osteoclasts (OCs), 816-817 chondromas, 799, 802
cysts, 54 chondromyxoid fibroma, 803
decalcification, 817 chondrosarcoma, 803-804
normal anatomy, 790, 816 chordomas, 813
Bone disorders, 793-794 cystidcyst-like lesions, 814-815
circulatory disturbances, 796-797 desmoplastic fibroma, 808-809
congenital and developmental diseases, enchondromas, 802
794 epithelial tumors, 812
infectious disorders, 797 Erdheim--Chester disease, 810
metabolic disorders. See Metabolic bone EWS/PNET, 813
disorders fibrosarcoma, 809
Paget disease, 797 fibrous tumors, 807-809
traumatic disorders, 794, 796 giant cell tumor, 810-811
Bone marrow hemangioendothelioma, 812
benign diseases hemangioma, 811
AIDS, 712 hematolymphoid tumors, 811
936 INDEX
Bone neoplasms (Contd.) radial scar/complex sclerosing lesions,

histiocytic and fibrohistiocytic tumors, 325-326
809-810 sclerosing adenosis, 325
intraosseous ganglion, 814 biomarkers in breast cancer, 319-322
Langerhans cell histiocytosis, 809-810 intraductal proliferative lesions, 322-323
lymphoma, 811 invasive breast. See Invasive breast cancer
malignant fibrous histiocytoma, 809 lymph node status, 326
metastatic carcinoma, 812 noninvasive carcinoma, 316-319
multiple myeloma, 811 normal breast, 304-305
nonossifying fibroma, 808 reporting template example, 304
osteoblastoma, 805 specimen processing, 299-303
osteochondroma, 798 Brenner tumors, 502
osteofibrous dysplasia, 808 Breslow thickness, 621, 623
osteoid osteoma, 805 Bronchiolitis, 117
osteoma, 805 Bronchiolitis obliterans-organizing pneumonia
osteosarcoma, 805--807 (BOOP), 107-108
PNET, 813 Bronchogenic cysts, 175, 782
solitary plasmacytoma, 811 Brown bowel syndrome, 232
vascular tumors, 811-812 Brown tumors, 62, 820
specimen processing, 791-793 Brunner's gland hyperplasia, 220
Borderline tumors, 187, 499-501 Buccal exostosis, 6
Botryoides, sarcoma, 556 Bulla,580
Bowel. See also Inflammatory bowel disease Bullosa acquisita, epidermolysis, 586
resection, 215-217 Bullous pemphigoid, 585, 856-857
syndrome, 232 Burkitt lymphoma, 707,721
Bowen disease, 596 Burnt out (regressed) germ cell tumors, 461
Bowenoid papulosis, 484
Brain C3 glomerulopathy, 332
abscess, 665 Calcifying odontogenic tumor, 52, 56
herniation,633--634 Calcifying epithelioma of Malherbe, 598
invasion, 656 Calcifying panniculitis, 589
Branchial deft anomalies, first, 98 Calcineurin inhibitor (CNI) toxicity, 343
Bronchioloalveolar carcinoma (BAC), Calcinosis, 204, 485, 591
131-132 Calcium, 848
Break apart fluorescent in situ hybridization Calymmatobacterium granulomatis, 485
(FISH), 883 Campbell de Morgan spot, 602
Breast Canalicular adenoma, 86--87
atypical vascular lesion, 768 Candida albicans, 4
carcinoma, cutaneous metastasis, 612 Candida infections
cytopathology, 835 cervix cytopathology, 548
Breast cytology esophagitis, 193, 199
diagnostic categories, 327 lung infection, 119
ductal adenocarcinoma, 327 vulva infection, 559
fibroadenoma, 327 Candidiasis, 4, 551
fibrocystic changes, 327 Capillaritis, pigmented purpuric, 588
gynecomastia, 32 7 Capillary hemangioma, 601, 765-766
lobular adenocarcinoma, 327 Carbohydrates, 844
special studies, 327-328 Carcinoid
specimen adequacy, 327 appendiceal, 242, 244
specimen procurement methods, cervica~ 545
326-327 colon neoplasms (nonepithelial), 239
subareolar abscess, 32 7 heart disease, 158-159
Breast pathology, 298 lung, 147
benign abnormalities tumorlets, 135
apocrine changes, 324 tumors
duct ectasia, 324 adult renal, 368
fat necrosis, 324 atypical, 135-136
fibroadenoma,324-325 chemodectomas, 135
intraductal papillomas, 325 lung, 134-136
microcysts, 324 neuroendocrine, 368
phyllodes tumor, 325 ovarian,506
INDEX 937
paraganglioma, 135 sella and/or suprasellar region,

stomach, 212 657-658
Carcinosarcoma neurodegenerative disorders, 669-671
fallopian tubes, 516 neurons pathology
ovarian neoplasms, 503 apoptosis, 631
uterus epithelial malignancies, 526-52 7 axonal injury, 631
Cardiomyopathy binucleation, 631
arrhythrnogenic right ventricular, 156 bunina bodies, 632
cardiac amyloidosis, 156 ferrugination, 631
dilated, 155 granulovacuolar degeneration, 632
drug-/radiation-induced, 156-157 Hirano bodies, 632
hereditary hemochromatosis, 156 intraneuronal inclusion bodies, 631
hypertrophic, 155 Lewy bodies, 631
infiltrative, 156 rnarinesco bodies, 632
ischemic, 155 necrosis, 631
restrictive (obliterative), 156 neurofibrillary tangles, 632
Cardiovascular system. See Heart; Vessels neuronal cytoplasmic inclusions, lDP-43,
Caroli's disease, 259 632
Cartilage-forming tumors, 798-799, 802-804 pick bodies, 631
Caruncle, 400 seizure disorders, 666
Cast nephropathy, 338 vascular disorders, 667-669
Castleman disease, 174, 700-701, 789 Central neurocytoma, 648-649
fi-catenin-activated adenomas, 265 Central osteosarcomas, 806
Catheterized urine, 396 Cerebral amyloid angiopathy, 668
Cavernous hemangioma, 270, 602, 767 Cerebral cavernous malformation (CCM),
CD30 + lyrnphoproliferative disorders, 667
primary cutaneous, 609, 611 Cerebral edema, 633
eDNA microarrays, 905 Cerebral infarcts, 669
Celiac disease, 217-218 Cerebral malaria, 664
Cernentoblastorna, 57 Cerebrospinal fluid (CSF), 671-672
Cernento-osseous dysplasia, 61 Ceruminous gland tumors, 99-100
Cementa-ossifying fibroma, 57-58 Cervical intraepithelial neoplasia (CIN), 541
Central giant cell granuloma of jaw, 61-62 Cervicitis, 537-538
Central nervous system (CNS) Cervicofacial actinomycosis, 5
anatomy and histology, 625-626 Cervix
astrocytes, 632-633 adenoid carcinoma, 544-545
brain herniation and, 633-634 adenosquarnous carcinoma, 544
cerebral edema, 633 Arias-Stella reaction, 539
cytology, 671-672 atrophy, 538
Duret (secondary brain stern) hemorrhage, cytopathology
634 adequacy statement, 548
hydrocephalus, 633 ancillary testing, 5 50
inflammatory and infectious disorders, Bethesda System, 548
660-665 epithelial cell abnormality, 549
intracranial pressure, 633-634 exfoliative cytology, 547-548
intraoperative evaluation and gross glandular cells abnormality, 549
examination,626-630 negative for intraepitheliallesion or
microglia, 633 rnalignanc~548-549
neoplasms specimen types, 547
choroid plexus tumors, 646-647 squamous cell abnormality, 549
embryonal, 651-653 decidual change, 539
ependymoma, 645-646 ectocervix, 5 35
germ cell, 658-659 endocennx,535,539
gliomas, 634, 638-645 endometriosis, 539
hemangioblastoma, 657 glandular neoplasms (adenocarcinoma),
lymphomas and histiocytic tumors, 543-544
659-660 glassy cell carcinoma, 544
meninges, 653-657 gross anatomy, 5 35
metastatic tumors, 660 gross examination and tissue sampling,
neuronal and glioneuronal, 647-650 536-537
pineal parenchymal, 650-651 hernatolyrnphoid neoplasms, 54 7
938 INDEX
Cervix (Contd.) Chronic inflammatory demyelinating

hyperplasia, 538 polyradiculoneuropathy (CIDP),
inclusion cysts, 539 678-679
inflammation and infection, 537-538 Chronic kidney disease-metabolic bone disease
melanoma, 547 (CKDMBD), 819
mesenchymal neoplasms, 545 Chronic myelogenous leukemia (CML),
mesonephric remnants, 539 717-718, 736
metaplasia, 538 Chronic rejection, 260, 341
microscopic anatomy, 535-536 Churg-5trauss syndrome, 116
mixed epithelial and mesenchymal Cicatricial pemphigoid, 3, 63, 857
neoplasms, 545-547 Ciliary dyskinesia, 841
mucoepidermoid carcinoma (MEC), 544 Ciliated cell metaplasia, 520
neoplasia, 539, 541-543 Ciliated foregut cyst, 268
neuroendocrine neoplasms, 545 Circulatory disorders
pathologic and clinical staging of bones disorders, 796-797
malignancies, 547 placental, 569-572
postoperative spindle-cell nodule, 539 Circumscribed astrocytomas, 642-643
radical hysterectomy, 537 Cirrhosis, 257-258
secondary malignancies, 547 Clark nevus, 618
Chagas disease, 155 Classical Hodgkin lymphoma (CHL), 702
Chancroid, 485 Clear cell adenocarcinoma
Charcot-Marie-Tooth disease (CMT1), cervica~ 544
hypertrophic, 679 epithelial malignancies, 526, 556
Chediak-Higashi syndrome, 713 urethral, 4 01
Chemodectomas, 135. See also Paraganglioma Clear cell metaplasia, 520
Cherry angioma, 602, 767 Clear cell tumors
Cherubism, 62 acanthoma, benign tumor of epidermis,
Chlamydia trachomatis, 485 595
Cholangiocarcinoma, 267, 273, 268, 281 chondrosarcoma, bone, 804
Cholangitis, 260, 275 endometrium, 520
Cholecystectomy, 2 74 hidradenoma, cutaneous appendages
Cholecystitis, 27 5 tumors, 600
Choledochal cysts, 2 75 lung, 133
Cholesteatoma, 100 ovarian, 502
Cholesterol granulomas, 100 pulmonary, 142
Cholesterolosis, 2 75 RCC, 353, 360-362, 365, 372
Chondroblastoma, 803 sarcoma, 352-353, 777
Chondrodermatitis nodularis helicis, 98 Clonality assays, 898-899
Chondroma, 770, 799, 802 Clustering, hierarchical, 908
Chondromatosis, synovial, 826 Coccidiomycosis, 120-121
Chondromyxoid fibroma, 803 Coiling, umbilical cord, 569
Chondro-osseous tumors, 770 Colitis, 228-232
Chondrosarcoma Colitis cystica profunda, 240
bone, 803-804 Collagen disorder, 593
clear cell, 804 Collagenous gastritis, 203
chondro-osseous tumors, 770 Collagenous sprue, 218
dedifferentiated, 804 Collapsing focal segmental glomerulosclerosis
extraskeletal myxoid, 777 (FSGS), 330, 332
mesenchymal, 770, 804 Collecting duct
synovial, 827 carcinoma (CDC), 363
Chorangiomas, 572 testis tumors, 463
Chorangiosis, 572 Colloid carcinoma, 290
Chordoid glioma of third ventricle (CG), 647 Colloidal iron, 844
Chordomas, 813 Colon
Choriocarcinoma, 173, 507, 533 diseases, 239-241
Choristoma, 100,449 normal anatomy, 214
Choroid plexus, 646-647 Columnar cell hyperplasia (CCH), 323
Chromogens, 853 Columnar epithelium, 535
Chromophobe renal cell carcinoma, 363, 372 Combined hepatocellular-cholangiocarcinoma,
Chromosome, 873-875. See also Cytogenetics 267-268,273
Chronic eosinophilic pneumonia (CEP), 114 Common variable immunodeficiency, 219
INDEX 939
Comparative genomic hybridization {CGH), Cylindroma, 599

869-870, 903 Cystadenocarcinoma, 269,285
Conduction system, 149 Cystadenofibroma, 499
Condyloma acuminatum Cystectomy, 373, 3 79-380
anal canal, 245 Cystic acinar cell cystadenoma, 287
cutaneous infections, 590 Cystic fibrosis, 241, 256
oral cavity and oropharynx, 9 Cystic kidney diseases {CKD)
urethral, 400 acquired, 346
vaginal, 554 medullary {MCKD), 346
Congenital infantile fibrosarcoma {OF), nonneoplastic, 345-346
754-755 Cystic nephroma {CN), 350, 368
Congestive cardiomyopathy, 15 5 Cystica profunda, 240
Congo red, 844 Cystoprostatectomy, 373, 380, 469
Congophilic {or cerebral) amyloid angiopathy Cysts
{CAA), 668 acquired cystic disease-associated RCC,
Conjunctiva diseases. See also Eye 364-365
degenerative, 63 adrenal glands, 444
inflammatory, 63 Baker, 825
neoplasms, 63-67, 72 Bartholin gland cysts, 552
Connective tissues biliary, 268
cytopathology, 845 bone,814-815
disorders, 110 breast pathology, 324
Copper, 848 bronchogenic, 175
Cor bovinum, 149 cervix, 539
Cornea diseases. See also Eye ciliated foregut cyst, 268
Fuchs endothelial dystrophy, 67 digital mucous, dermal deposits, 591
intraocular neoplasms, 71-73 epithelial, adrenal glands, 444
keratoconus, 69 epithelial inclusion cysts, 5 52
pseudophakic bullous keratopathy, 68-69 fallopian tubes, 512
trauma, 71 jaws, 51-54
ulcerative keratitis and, 69 keratinous, 592
vascular and infectious diseases, 70-71 larynx, 23
chronic retinal detachment, 70 lymphoepithelial, 285
intraocular inflammation and infection, mature cystic teratoma, 173
70-71 mediastinum, 175
phthisis bulbi, 71 mesonephric cysts, 5 52
secondary glaucoma, 70 mucinous cystic neoplasm {liver), 269
Corpora amylacea, 632 Miillerian,552
Corpus luteum cysts, 494. See also Uterus multicystic mesothelioma, 187
Cortical dysplasia, 666 multicystic peritoneal inclusion cyst, 181
Cortical nodules. See also Adrenal glands oral cavity and oropharynx, 6, 7
congenital abnormalities, 429-430 ovarian,494-495
cortical hypofunction, 430-431 pancreas, 284-285, 296
hyperplasia, 431-432 parasitic, 444
incidental, 430 parathyroid, 423
neoplasms, 432-435 penile, 486
Corticotroph {ACTH-producing) adenoma, pseudocysts, 444
449 retroperitoneum, 782-78 3
Craniopharyngiomas, 657-659 salivary glands, 83-84
Crescentic glomerulonephritis, 334 serosal membranes, 180
Creutzfeldt-Jakob Disease (CJD), 670-671 tubulocystic carcinoma, 364
Cribriform hyperplasia, 472 urinary bladder cystitis, 381-382
Crohn's disease, 228, 552, 5 59 vaginal, 5 52
Cryptococcus lung infection, 120 vascular, 444
Cryptorchidism, 453 vulva, 560
Cryptosporidiosis, 219 Cytogenetics. See also Cytopathology
Crystal-induced synovitis, 824-825 comparative genomic hybridization (CGH),
Cushing syndrome, 433 869-870
Cutanea tarda, porphyria, 586 FISH, 867-869
Cyclophosphamide toxicity, 157 human chromosome nomenclature, 873-875
Cylindrical cell carcinoma, 43 microarray analysis, 870-873
940 INDEX
Cytogenetics (Contd.) Dematiaceous fungus, 591

traditional cytogenetic analysis, 863 Demyelinating viral infections, 661-662
assay failure, 866-867 Demyelination, 674-675
banding, 865 Dendritic cell tumors, 737
basic laboratory procedures, 863-866 Dermal deposits, 591-592
cell harvest, 865 Dermal duct tumors, 599-600
culture initiation, 864 Dermal melanocytosis, 617
culture maintenance, 865 Dermatitis herpetiformis, 586, 856-857
limitations, 863 Dermatofibroma, 606
microscopic analysis, 865 Dermatofibrosarcoma protuberans,
Cytologic atypia, 521 606
Cytomegalovirus (CMV) infections Dermatophytosis, 590-591
esophagitis, 200 Dermatosis, 587-588
hepatitis, 252 Dermis, 579
infectious esophagitis, 194 Dermoid cysts, 7
kidney, tubulointerstitial diseases, 33 7 Desmoid fibromatosis, 752, 786
lung infection, 11 7-118 Desmoplastic
placental, 576 fibroma, 808-809
Cytopathology. See also Biospecimen banking; infantile
Cytogenetics; DNA sequence analysis; astrocytoma, 648
Microarrays ganglioglioma, 648
anatomic sites deserving special mention mesothelioma, 184
breast, 835 neurotropic type melanoma, 621
lymph nodes for metastatic disease, Desmoplastic small round cell tumor (DSRCT)
835-836 mesenteric, 24 7
skin, 835 paratesticular tumors, 464
communication of findings, 833 pediatric kidney neoplasms, 356
electron microscopy (EM), 838-841 serosal membranes, 186-188
FISH, 876-886 soft tissue tumor, 776
flow cytometry, 859-862 Desquamative interstitial pneumonia {DIP),
frozen sections, 832-835 109-110
histology and histochemical stains, Developmental odontogenic cysts, 51-52
842-850 Diabetes
immunofluorescence, 85 6-858 mellitus {DM), 334-335
immunohistochemistry, 851-855 metabolic neuropathies, 676-677
intraoperative consultations, 832-833 placenta, maternal disease, 573-574
opening of viscus organ for gross Diabetic autonomic neuropathy, 677
examination and fixation, 836 Diff-Quik stain, 829
tumor for banking, 837 Diffuse {infiltrating) astrocytomas {DAs), 634
telepathology, 928-929 Diffuse alveolar damage (DAD), 106-107
Cytopathology principles, 828 Diffuse large B-celllymphoma {DLBCL), 212
fine needle aspiration (FNA), 830-831 Diffuse malignant mesothelioma {DMM),
flow cytometry, 828 181-184, 187
gynecologic/PAP slides, 828 Digital fibrokeratoma, 606
immunocytochemistry, 828 Digital mucous cyst, 591
molecular genetic methods, 828 Digital myxoma, 773
nongynecologic, 829 Dilated cardiomyopathy, 155
preparation and processing, 829 Dilated pore of Winer, 598
specimen types, 829 Dirofilariasis, 121
Disordered proliferative endometrium, 521
De novo diseases, 261, 342 Disseminated peritonealleiomyomatosis, 179
De Quervain thyroiditis, 406, 418 Distal pancreatectomy, 282-283
Decalcification, 792, 817 Diversion colitis, 230
Decidual change, 539 Diverticula, 384, 399
Deep aggressive angiomyxoma, 554, 773 Diverticulosis, 231
Deep benign fibrous histiocytoma, 757 DNA resequencing, 911-912
Deep lymphangioma, 602 DNA sequence analysis
Deep penetrating nevus, 618 analytidtechnical versus diagnostic and
Deep-seated leiomyoma, 785 operational aspects of testing, 889
Degenerative conjunctiva diseases, 63 clonality assays, 898-899
Deletions, 880 direct
INDEX 941
dye terminator cycle sequencing method, middle, 97

897 neoplasms, 101-102
next-generation sequencing (NGS), nonneoplastic diseases, 100
897-898 neoplasms of external ear, 98
identity determination aspects, 901 benign ceruminous gland tumors, 99
indirect malignant ceruminous gland tumors, 100
allele-specific PCR, 898 normal anatomy, 97
allelic discrimination, 898 Eccrine
melting curve analysis, 898 poroma, 599-600
single-strand conformational tumors, 599-600
polymorphism (SSCP) analysis, 898 Ectasia, 204, 324
infectious disease testing, 899-901 Ectomesenchymoma, 765
microsatellite instability (MSI) assays, 899 Ectopic adrenal cortical adenoma, 433
PCR Ectopic decidual reaction, 179
methodology, 893-896 Ectopic pregnancy, fallopian tubes and, 513
testing affecting factors, 890-893 Ectopic tissue, 192, 384, 539
specimen requirements, 887-889 Eczema, 585
Doxorubicin toxicity, 156-157 Effusion, 157, 185
Drug-induced Elastic fibers, 580
cardiomyopathy, 156-157 Elastin, 593
colitis, 232 Elastofibroma, 750
pulmonary changes, 11 0-111 Elastosis, solar, 593
Drug-induced liver injury (DILl), 256 Electron microscopy (EM), 838
Ductal carcinoma CNS intraoperative evaluation and gross
adenocarcinoma examination, 627
breast cytology, 327 heart,841
pancreatic (PDA), 287-291 inherited metabolic diseases, 840
prostate cancer, 478 kidney, 839, 840
in situ (DCIS), 316-318 liver, 840
invasive, 311-312 lung, 840
Ductal hyperplasia, 322-323 methodology, 838
Ductal lavage, 327 microvillus inclusion disease, 841
Ductal plate malformation (DPM), 259 neoplasms, 840
Duret (secondary brain stem) hemorrhage, 634 neuropathology specimens, 841
Dye terminator cycle sequencing method, 897 prenatal studies, 840
Dysembryoplastic neuroepithdial tumor primary ciliary dyskinesia, 841
(DNT), 649 renal biopsy, 329
Dyskeratosis, 580, 593 Elephantiasis, 486
Dysplasia Embryonal carcinoma, 459-460, 507,
cortical, 666 651-653
fibrous, 807-808 Embryonal rhabdomyosarcoma (ERMS),
large intestinal inflammatory bowel disease, 762-764
229-230 Emphysematous bulla, 180
multicystic, 344 Encapsulated neuroma, 603
osteofibrous, 808 Encephalitis
renal, 344 amebic (AE), 664-665
stomach neoplasm, 205 brain abscess, 665
urinary bladder, 385 cerebral malaria, 664
urothelial, 385, 374 herpes, 663-664
Dysplastic cerebellar gangliocytoma (DCG), HN, 664
648 neurocysticeroosis, 664
Dysplastic nevus, 618 parasitic, 664-665
Dystrophies, muscular, 761 rocky mountain spotted fever, 664
toxoplasmosis, 665
Ear varicella zoster virus, 664
external, 97 viral, 663-664
neoplasms, 98-100 West Nile Virus, 663
nonneoplastic diseases, 98 Encephalomyelitis, 661
inner, 97 Enchondromas, 802
diseases, 102-103 Endocardial fibroelastosis, 153
neoplasms, 102-103 Endocardium disorders, 151, 153
942 INDEX
Endocervical glandular hyperplasia, 538 Epithelial neoplasms

Endocervical polyps, 539 bone, 812
Endocervicosis, 179, 384 conjunctiva diseases, 63--64
Endocervix, 549 esophagus, 194-195, 197-198
Endodermal sinus tumor (EST), 173, 355, 460 kidney, 365-366, 368
Endometrial hyperplasia, 521 liver, 262, 264-267
Endometrial sarcoma, 527-528 oral cavity and oropharynx, 9-13
Endometrial stromal tumors, 527 penis and scrotum, 486
Endometrioid adenocarcinoma, 502, serosal membranes, 187
522-526,543 stromal tumors, 499-503
Endometrioid tumors, 501-502 surface ovarian neoplasms, 499-503
Endometriosis ureter, 374
cervix, 539 urethral, 400
fallopian tubes, 513 vaginal, 552,554-556
ovarian, 494 Epithelioid cell nevus, 616
serosal membranes, 178-1 79 Epithelioid hemangioendothelioma (EH)
urinary bladder, 383 bone, 812
uterus, 533 liver, 271-272
vaginal, 552 lung, 141
Endometriotic cysts, 494-495 serosal membranes, 184-185
Endometritis, 519-520 soft tissue, 769
Endometrium vascular tumor of skin, 603
benign diseases Epithelium
atrophy, 520 cervix microscopic anatomy, 535
disordered proliferative endometrium, columnar, 535
521 squamous, 535
endometritis, 519-520 squamous metaplastic, 535-536
metaplasia, 520 Epstein-Barr virus (EBV)
polyps, 520-521 hepatitis, 252
dating, 518 infectious disease testing, 900
endometrial intraepithdial carcinoma (EIC), oral cavity and oropharynx infections, 5
522 Erdheim-Chester disease, 810
endometrial intraepithdial neoplasia (EIN), Erythema induratum, 588
521-522 Erythema multiforme (EM), 583-584
exogenous hormone therapy, 519 Erythema nodosum (septal), 588
hyperplasia, 521 Esophagitis
pregnancy and, 518-519 eosinophilic, 193
Endomucosal resection (EMR), 217 herpetic, 199
Endosalpingiosis, 179, 384 infectious
Endothdial hyperplasia, 765 Candida, 193, 199
Enteric duplication cyst (EDC), 783 cytomegalovirus (CMV), 194, 200
Enterocolitis, neonatal necrotizing, 227 herpes simplex virus (HSV), 194
Enteropathy, 218, 225 pill, 193
Enzymes stains, 849-850 reflux, 192
Eosin staining, 843 Esophagus
Eosinophilic granular bodies (EGBs), 633 benign nonneoplastic changes
Eosinophilic metaplasia, 471, 520 ectopic tissue, 192
Eosinophils, 396 gastric heterotopias, 191
Ependymoma, 645-646 glycogenic acanthosis, 191
Epidermal growth factor receptors (EGFR), cytopathology, 199-200
130-131 endoscopic biopsy, 190
Epidermal inclusion cysts, 486 endoscopic mucosal resection, 190
Epidermal maturation disorders, 593 epithelial neoplasms and preneoplasms
Epidermis, 579, 595-596 adenocarcinoma, 197-198
Epidermoid cysts, 7 Barrett esophagus, 194-195, 197
Epidermolysis bullosa acquisita, 586 neuroendocrine neoplasms, 198
Epidermotropism, 580 salivary gland-type neoplasms, 198
Epididymitis, 454 sec, 194
Epithelial hyperplasia, 383, 471-472 squamous dysplasia, 194
Epithelial inclusion cysts, 5 52 esophagectomy, 191
Epithelial membrane antigen (EMA), 629 lymphoma, 199
INDEX 943
melanoma, 199 cystic lesions, 512

mesenchymal neoplasms, 198-199 ectopic pregnancy and, 513
metastasis, 199 endometriosis, 513
mucosal injury gross examination and tissue sampling,
chemical injury, 192 511
eosinophilic esophagitis, 193 benign specimens, 511
graft versus host disease (GVHD), 193 neoplastic specimens, 511
infectious esophagitis, 193-194 prophylactic excision specimens, 511
pill esophagitis, 193 inflammatory lesions
radiation injury, 193 acute salpingitis, 512-513
reflux esophagitis, 192 chronic salpingitis, 513
normal anatomy, 190 granulomatous salpingitis, 513
polyps, 194 salpingitis isthmica nodosum, 513
Essential thrombocythemia, 718 malignant tumors
Esrerogen,320,519 carcinosarcoma, 516
Ewing sarcoma (EWS), 355-356, n5-n6, leiomyosarcoma, 516
813 metastatic tumors, 516
Exocrine pancreas solid tumors, 287-293 primary fallopian tube carcinoma, 516
Exocytosis, 580 reporting, 516
Exogenous hormone therapy, 519 normal anatomy, 511
Exophytic papilloma, 38 Familial adenomatous polyposis (FAP),
Extraadrenal paraganglioma lesions 235
extraadrenalparaganglioma,441-443 Fasciitis, 749
normal anatomy and histology, 441 Fat necrosis, 324, 589
retroperitoneal, 787-788 Fatty and muscular tumors, 607
Extrachorialis, 572 Fetal membranes disorders, 567-568.
Extrahepatic bile ducts See also Placenta
adenocarcinomas, 278 Fetus papyraceous, 568
adenoma, 275 Fibrin deposition, 570
biliary atresia, 275 Fibroadenoma, 324-325, 327
biliary intraepithelial neoplasia (BiliN), Fibroblast growth factor 1 receptor
275 abnormalities, 719
intraductal papillary neoplasms (IPMNs), Fibroblastidmyofibroblastic tumors.
278 See also Fibrohistiocytic tumors
intraluminal papillary neoplasms, 277 benign, 749-751
mucinous cystic neoplasm (MCN), 278 inrermediate
normal anatomy, 274 locally aggressive, 752-753
Extrahepatic biliary sysrem rarely metastasizing, 753-755
cytopathology malignant, 755-756
inflammatory lesions, 280-281 retroperitoneum, 786
lithiasis, 280 soft tissue, 749-756
PSC,281 Fibroepithelial polyps
reactive, 280--281 skin, 606
nonneoplastic diseases urethral congenital anomalies, 400
biliary atresia, 275 urinary bladder, 385
cholangiocarcinoma, 281 vaginal epithelial neoplasms, 552
PSC, 275 vulvar, 560
Eye Fibrofolliculoma, 598-599
cornea diseases, 67, 71-73 Fibrohistiocytic tumors. See also Fibroblastid
conjunctiva diseases, 63-67 myofibroblastic tumors
eyelid diseases, n, 79 benign, 757-758
lacrimal gland diseases, 73, 75 bone,809-810
lacrimal sac diseases, 75 inrermediate (rarely metastasizing), 758
orbit diseases, 77 malignant, 758-759
retroperitoneum, 787
Faciale, granuloma, 587 skin mesenchymal tumors, 605--607
Fallopian tubes soft tissue, 757-759
benign tumors Fibroid polyps, 240
adenofibromas, 516 Fibrokeratoma, digital, 606
adenomatoid tumors, 513 Fibrolamellar hepatic cell carcinoma (HCC),
epithelial papillomas, 513 267,272
944 I INDEX
Fibromatosis Focal nodular hyperplasia (FNH), 262, 264,

desmoid, 786 272
desmoid-type, 752 Focal segmental glomerulosclerosis (FSGS),
infantile subcutaneous, 752 330,332
mesenteric, 24 7 Fontana-Masson method, 848
retroperitoneum, 786 Fordyce granules, 6
Fibromyxoid tumor, ossifying, n4 Foreskin resection, 483
Fibro-osseous pseudotumor of digits, 749 Foveolar hyperplasia, 204
Fibrosarcoma Frozen sections. See also Biospecimen banking
adult (AFS), 755 786 cytopathological aspects
bone, 809 accuracy, 833
congenital infantile (CIF), 754-755 indications, 832
fibroblastidmyofibroblastic tumors, interpretation, 833
754-755 procedure, 832-833
sclerosing epithelioid (SEF), 755 sources of error in, 833, 835
Fibrosing cholestatic hepatitis (FCH), 262 gross sampling error, 835
Fibrosis sampling errors, 835
congenital hepatic, 259 sectioning error, 835
idiopathic retroperitoneal, 782 tissue, 921-922
serosal membranes, 179-18 0 Fuchs endothelial dystrophy, 67
Fibrous hamartoma of infancy (FI-ll), 750 Functional endoscopic sinus surgery, 35
Fine needle aspiration (FNA), 326, 830-831 Fundic gland polyps, 205
First branchial deft anomalies, 98 Fungal cervicitis, 538
Fissural cysts, 53 Fungal infections
Flat urotheliallesions with atypia, 385 cutaneous infections, 590-591
Florid cemento-osseous dysplasia, 61 disease testing, 900
Flow cytometry liver, 253
cytopathology principles, 828 lung, 119-121, 146
electronic system, 860-861 oral cavity and oropharynx, 4
flow system, 859 sinonasal region, 36-38
optical system, 859-860 Fungiform papilloma, 38
principle, 859 Fusion fluorescent in situ hybridization
uses, 861-862 (FISH), 883
Fluorescence, 859-860. See also Cytogenetics
Fluorescence in situ hybridization (FISH), 876 Gallbladder
advantages and limitations adenocarcinoma, 278
aneuploidy and polyploidy, 878 adenoma, 275
autofluorescence, 878 biliary adenocarcinomas, 278
disadvantages and pitfalls, 878 cytopathology, 2 79-280
partial hybridization failure, 878 gross examination, 2 74
specimens, retained morphology, and intracystic papillary neoplasms, 277
combined FISH/immunohistochemistry, intraluminal papillary neoplasms, 277
876 mucinous cystic neoplasm (MCN), 278
tissue microarray-FISH, 877-878 nonneoplastic disease, 27 5
truncation artifact, 878 normal anatomy, 274
versus other cytogenetic and molecular Gangliocytic paraganglioma, 222,442-443
techniques, 876-8n Gangliocytoma, 64 7-648
clinical applications, 880 Ganglioglioma, desmoplastic infantile, 648
aneusomies and deletions, 880, 882 Ganglion cell tumor (GG and gangliocytoma),
break apart FISH, 883 647-648
fusion FISH, 883 Ganglion, intraosseous, 814
gene amplifications, 882-883 Ganglioneuroma (GN), 175,240, 441
translocations, 883 Gangrene, 485
metaphase, 867-868 Gardner-associated fibroma, 751
multiplex metaphase, 868-869 Gardnerella vaginalis, 551
nomenclature, 885-886 Gastrectomy, 201-202
probes and probe development, 879-880 Gastritis, 202-204
technical considerations, 878-879 Gastroenteritis, 219
test development for clinical use, 884-885 Gastrointestinal stromal tumors (GIST), 199,
Focal anaplasia, 350 208-212,222,240,786
Focal glandular atypia (ASAP), 473 Gastropathy, 204
INDEX 945
Gaucher disease, 713 Glycoprotein (MAG) neuropathy, antimyelin

Gene expression profiling, 910 associated, 679
Genetic neuropathies, 679-680 GMS stain, 84 7
Genital rhabdomyoma, 554,762 Goblet cell carcinoid, appendiceal, 242, 244
Genome copy number, 911 Goiter, multinodular, 408-409
Genome tiling, 903 Gonadoblastoma, ovarian, 507-508
Geno~pillg,903,911 Gonadotroph adenomas, 449
Germ cell tumors (GCTs) Gonorrhea, 485
adult renal neoplasms, 368 Gout, 592
CNS, 658-659 Graft versus host disease (GVHD), 193
mediastinal, 173-174 acute, liver transplantation pathology, 262
ovarian,505-507 esophagus mucosal injury, 193
retroperitoneum, 788-789 inflammatory dermatoses, 584
serosal membranes, 189 large intestinal nonneoplastic conditions,
testiscular, 455-462 232
Gestation nonneoplastic conditions of stomach and,
multiple, 567 204
diamniotic dichorionic, 574 Granular cell tumors (GCT)
diamniotic monochorionic placentation, adult (AGCTs), 503
574 esophagus, 198, 200
monoamniotic placentation, 574 juvenile GGCTs), 503-504
TITS, 575 larynx, 25
trophoblastic diseases, 531-533 oral cavity and oropharynx, 15
Giant axonal neuropathy, 680 ovarian neoplasms, 503-504
Giant cell tumors pituitary, 449
bone, 810-811 skin, 604
invasive urothelial neoplasms variant, 391 soft tissue peripheral nerve tumors, 772
lung, 133 testis tumors, 462-463
malignant fibrous histiocytoma (MFH), vulvar, 562
759 Granular parakeratosis, 594
diffuse-~pe, 757, 826 Granulocytic left shift, 710
synovial tumors, 825-826 Granuloma annulare, 586
soft tissue, 758 Granuloma faciale, 587
of tendon sheath (GCTIS), 757 Granuloma inguinale, 485
Giardiasis, 219 Granulomas
Gingival cyst of adult, 52 bone marrow, 712
Glandular cells, 548-549 cholesterol, 100
Glandular neoplasms, 393, 543-544 inflammatory dermatoses, 586-587
Glandular odontogenic cyst, 52 liver, 259
Glandularis, 400 ovary, 494
Glassy cell carcinoma, 544 spleen, 733
Glaucoma, 70 Granulovacuolar degeneration (GVD), 632
Glial markers, 629 Graves disease, 407-408
Glioblastoma multiforme (GBM), 638, Gross sampling error, 835
641-642 Guillain-Barre syndrome (GBS), 678
Gliomas, 634, 638, 641-645, 647 Gynecologic/PAP slides, 828
Gliomatosis cerebri, 642 Gynecomastia, 32 7
Gliosarcoma (GS), 642
Globulomaxillary cysts, 53 Hair cyst, 592. See also Keratinous cysts
Glomangioma, 602 Hair follicle tumors, 596-599
Glomangiopericytoma, 4 7 Hairy cell leukemia (HCL), 726, 728, 736
Glomerular diseases. See under Kidney Halo nevus, 618
Glomeruloid hemangioma, 765 Hamartoma
Glomerulonephritis (GN), 332-334 fibrous hamartoma of infancy (FI-ll), 750
Glomerulopathy, C3, 332 mesenchymal liver, 270
Glomus tumors, 602, 760-761. See also peribiliary gland, 268
Paraganglioma pulmonary, 142
Glycogen storage diseases, 255 respiratory epithelial adenomatoid, 38
Glycogenoses, 255 small intestinal neoplasms, 220
Glycogen-rich (dear-cell) variant of invasive Hamartomatous polyps, 205, 235
urothelial neoplasms, 392 Hashimoto thyroiditis, 406-407, 418
946 INDEX
Heart. See also Vessels Hematolymphoid neoplasms

cardiac bone, 811
transplants, 159 cervix, 547
valves, 151 penile cancer, 489
electron microscopy (EM), 841 prostate cancer, 479
endocardium disorders, 151, 153 renal pelvis cancer, 376
endomyocardial biopsies, 149 testis tumors, 463
heart explant specimens, 151 ureter cancer, 376
myocardium disorders, 153-157 Hematomas, 569,571
myomectomy specimens, 151 Hematopoietic malignancies
neoplasms, 157 adult renal neoplasms, 368
angiosarcoma, 158 lung cytopathology, 148
carcinoid heart disease, 158-159 ovarian, 508
cardiac myxoma, 158 urethra malignant neoplasms, 403
intramural cardiac fibroma, 158 Hematoxylin and eosin (H&E) staining,
lipomas, 158 843-844
papillary fibroelastoma, 158 Hemithyroidectomy, 404
rhabdomyoma, 158 Hemochromatosis, hereditary, 156, 254
normal anatomy, 149 Hemophagocytic syndrome, 255, 713
pericardium disorders, 157 Hemorrhage
valves disorders, 153 cystitis, 3 82
Helicobacter pylori, 900 Duret (secondary brain stem), 634
Hemangioblastoma (HB), 657 Hemorrhoids, 245
Hemangioendothelioma Hemosiderin deposition, 567
bone, 812 Hemosiderosis, 147
composite, 768 Hemosiderotic synovitis, 825
epithelioid (EH) Hepatectomy, 249
liver, 271-272 Hepatic fibrosis, congenital, 259
lung, 141 Hepatitis, 251-252
serosal membranes, 184-185 Hepatitis C virus (HCV), 900
soft tissue malignant tumors, 769 Hepatoblastoma, 268
vascular tumor of skin, 603 Hepatocellular adenoma (HCA), 264-266,
Kaposiform, 788 272
littoral cell, 738 Hepatocellular carcinoma (HCC), 266-268,
polymorphous, 768 272
pseudomyogenic, 768 Hepatopathy, 25 4
retiform, 769 Hepatosplenic T-celllymphoma (HSTL), 735
Hemangiomas Hereditary neuropathy with pressure palsies
arteriovenous,602, 767 (HNPP), 679
bone, 811 Hereditary sensory and autonomic
capillary, 766 neuropathies (HSANs), 680
cavernous, 270, 767 Hermansky-Pudlak syndrome, 713
congenital, 766 Herniation, brain, 633-634
epithelioid, 767 Herpes simplex virus (HSV)
glomeruloid, 765 cutaneous infections, 590
infantile (IH), 270 infectious esophagitis, 194
liver, 270, 272 lung infection, 118
lobular capillary (LCH), 765-766 oral cavity and oropharynx infection, 4
Masson vegetant, 765 penis and scrotum infection, 484
mediastinal soft tissue, 176 placental infection, 576
mesenchymal, 367 vaginal infection, 551
oral cavity and oropharynx, 14 vulva infection, 559
soft tissue vascular, 765-767 Herpes virus hepatitis, 252
spindle cell, 767 Herpes zoster infections, 681
spleen, 737 Heterotopias, 191
synovial, 767, 827 Heterotopic
venous, 767 congenital intestinal anomalies
verrucous, 767 gastric mucosa, 217
Hemangiopericytoma (HPC), 47, 656-657, heterotopic pancreas, 217
753-754, 786 epithelial elements, breast pathology, 326
Hematolymphoid lesions, 144 Hibemoma, 747
INDEX 947
Hidradenitis suppurativa, 485, 558 penis and scrotum inflammation and

Hidradenoma, 560, 562, 600 infection, 484
Hidradenoma papilliferum, 245 vulvar infection, 559
Hidroacanthoma simplex, 599-600 Human polyoma virus, 397
Hidrocystoma, 600 Hiirthle cells, 419
High grade dysplasia (HGD - adenocarcinoma Hyaline-vascular Casdeman disease, 701
in situ), 236 Hyalinizing angiectatic tumor, 774
High-grade squamous intraepitheliallesion Hyalinizing trabecular adenoma, 420
(HSIL), 549 Hydrocephalus, 633
Hilus cell Hyperacute rejection, 123
hyperplasia, 495 Hyperaldosteronism, 432-433
tumors, 505 Hypercalcemic small cell carcinoma, 508
Hippocampal (mesial temporal) sclerosis (HS), Hypercellularity, 332
666 Hypergranulosis, 580
Hirano bodies, 632 Hyperkeratosis, 580
Hirschsprung's disease, 225-226 Hyperparathyroidism, 62, 820
Histiocytic disorders Hyperplasia
Langerhans cell histiocytosis (LCH), adrenal cortical, 431-432
659-660 adrenal medullary (AMH), 436-437
non-Langerhans cell histiocytosis (LCH), atypical adenomatous hyperplasia adenosis,
660 472
spleen neoplastic disorders, 737 atypical alveolar (AAH), 132
tumors atypical ductal (ADH), 322-323
bone,809-810 aypicallobular (ALH), 323
CNS neoplasms, 659-660 basal cell, 4 72
Histiocytoma Brunner•s gland, 220
benign fibrous, 757, 809 columnar cell (CCH), breast pathology, 323
malignant fibrous (MFH), 758-759, cribriform, 4 72
809 diffuse (paracortical), 684
Histiocytosis, Langerhans cell, 809-810 diffuse laminar endocervical glandular, 538
Histology and histochemical stains. See under endometrial, 521
Cytopathology epithelial predominant BPH nodules, 472
Histoplasmosis, 120 fibrous gingival, 6
HIV follicular, 684
encephalitis, 664 foveolar, 204
lipodystrophy, 747 idiopathic myointimal, 247
placental infection, 577 inflammatory papillary, 6
HNF1a mutation, 265 lobular endocervical glandular, 538
Hodgkin lymphoma (HL). See also mesonephric remnant, 4 72
Non-Hodgkin lymphomas mesothelial, 178
classical (CHL), 702 microglandular, 538
cytopathology, 708 mixed epithelial, 4 72
immunophenotype studies, 703-704 neuroendocrine cell, 134
lymph nodes cytopathology, 708 nodular, 408-409, 471
mediastinal neoplasms, 174 ovarian,495
nodular lymphocyte predominant Hodgkin papillary endothelial (PEH), 765
lymphoma (NLPHL), 702 parathyroid, 423-424
pathophysiology, 702-703 parietal cell, 204
retroperitoneum, 789 pituitary, 447
splenic, 735 pseudocarcinomatous epithelial, 383
staging, 704, 706 pseudoepitheliomatous, 7
Hori nevus, 617 sebaceous, 600
Hormonal androgen deprivation therapy, 479 sinus, 684
Human chromosome nomenclature, 873-875. squamous, 538
See also Cytogenetics squamous papilloma, 538
Human epidermal growth factor receptor 2 stromal, 4 71-4 72
(HER2), 321 urethral, 40 0
Human papilloma virus (HPV) urothelial, 383
cervical infections, 541, 5 50 usual ductal (UDH), 323
infectious disease testing, 900 usual nodular epithelial, 4 71
oral cavity and oropharynx, 4 verumontanum gland, 4 72
948 INDEX
Hyperplastic perilobar nephroblastomatosis, Immunophenotype, 700

diffuse, 350 Immunoproliferative small intestinal disease
Hyperplastic polyps, 205, 233, 241 (IPSID), 222
Hyperreactio luteinalis, 495 Implantation disorders. See under Placenta
Hypersensitivity myocarditis, 154 Inclusion cysts, 539
Hypersensitivity pneumonitis (HP), 111-112 Infancy, fibrous hamartoma of (FHI), 750
Hypersplenism, 730--731 Infantile digital fibroma-fibromatosis, 752
Hypertension, 258, 339 Infantile fibrosarcoma, congenital (CIF),
Hyperthecosis, 495 754-755
Hypertrophic cardiomyopathy, 155 Infantile hemangioma (IH), 270
Hypertrophic Charcot-Marie-Tooth disease Infarcts
(CMT1), 679 bone, 796-797
Hypertrophic (onion -bulb) neuropathies, 679 cerebral, 669
Hypophysitis, 447 maternal floor, 570
Hyposplenism, 731 placental circulatory disorder, 569
Hysterectomy, 517, 537 spleen, 733
subcortical, 668--669
Iatrogenic disorders, 798 Infections. See also Infectious disease testing;
Idiopathic adulthood ductopenia (lAD), 258 Infectious diseases
Idiopathic interstitial pneumonitis, 108-110 bacterial
Idiopathic myointimal hyperplasia, 247 lungs, 118-119
Idiopathic posttransplant chronic hepatitis oral cavity and oropharynx, 5
(IPTH), 261 fungal
Image analysis, 907, 928 lungs, 119-121
Immune-mediated mechanism, neuropathies oral cavity and oropharynx, 4
with, 678-679 viral
Immunocytochemistry, 828 lungs, 117-118
Immunofluorescence oral cavity and oropharynx, 4-5
direct, 85 6-857 varicella zoster (VZV), 118
indirect, 857-858 Infectious disease testing
renal biopsy, 329 bacteria, 900
Immunoglobulin mesangial hypercellularity, DNA sequence analysis, 899
332 fungi, 900
Immunohistochemistry, 851 protozoans, 901
CNS intraoperative evaluation and gross viral, 900
examination, 62 7 Infectious diseases
data sources, 855 bones, 797-798
interpretation, 853, 855 cardiac transplants and, 161
methodology CNS,660-665
alkaline phosphatase, 853 cornea diseases, 70-71
antibody cocktails, 853 intestinal, 219-220
automation, 853 kidney, 337, 343
avidin-biotin method, 852 larynx, 22
background staining, 852 liver, 251-253
chromogens, 853 lungs, 117-121, 146-147
counterstaining, 853 ovarian, 493-494
detection systems, 852 penis and scrotum, 484-485
direct conjugate-labeled antibody method, placental, 575-578
852 prostate, 469
indirect or sandwich method, 852 salivary glands, 82
polymer-based labels, 853 sinonasal region, 35
primary antibody, 852 syphilis, 5 59
quantitative immunohistochemistry, 853 testis, 454-455
streptavidin-biotin conjugate method, 852 urinary bladder cytology, 397
tyramine amplification, 853 urethral inflammation and infection, 400
unlabeled antibody method, 852 vagina, 551
reporting, 853 vulvar, 559
specimen type and tissue management, Infertility, testis and, 453-454
851-852 Infiltrates. See Cutaneous lymphoid infiltrates
test selection (preanalytical phase), 851 Infiltrative cardiomyopathy, 156
validation,853 Inflammation
INDEX 949
acute, 397 subcutaneous fat necrosis of newborn, 589

cervical, 53 8 toxic epidermal necrolysis (TEN), 583
CNS, 665 urticaria, 588
fallopian tubes, 512-513 vasa-occlusive vasculopathies, 587
granulomatous, 665 Inflammatory diseases
intraoculax; 70--71 conjunctiva diseases, 63
larynx, 22 lesions
mediastinum, 167 extrahepatic biliary tree, 280-281
oral cavity and oropharynx, 2-3 gallbladder, 2 79
penis and scrotum, 484 ovary, 493-494
prostate, 469-470 polyps, 36, 194, 240
salivary glands, 82-83 retroperitoneum, 781-782
sinonasal region, 35 vaginal, 551-552
stomach, 212 Inflammatory myofibroblastic tumor (IMT) ,
testis, 454-455 143, 247, 754, 786
thyroid, 406-408 Inflammatory prostatitis, 4 70
urethral, 400 Inflammatory venoocclusive disease, 247
urinary bladder, 397, 380, 382 Influenza, 118
vulva, 558-559 Infundibular type keratinous cysts, 592
Inflammatory adenoma (IA), 265-266 Inherited metabolic diseases, 840
Inflammatory bowel disease (IBD), 227 INI11BAF47 deletions, 630
bowel resection, 216 Intestine
Crohn's disease, 228 anal biopsy, 217
dysplasia, 229-230 anal canal
indeterminate colitis, 229 nonneoplastic and neoplastic conditions,
pouchitis, 230 244-246
ulcerative colitis, 228-229 normal anatomy, 215
Inflammatory cap polyps, 240 appendectomy, 217
Inflammatory dermatoses appendix
acute vasculitis, 587 nonneoplastic and neoplastic conditions,
bullous pemphigoid, 585 241-244
calcifying panniculitis (lobular), 589 normal anatomy, 215
dermatitis herpetiformis, 586 bowel resection
eczema/atopic dermatitis/allergic or irritant inflammatory bowel disease, 216
contact dermatitis, 585 neoplastic, 215-216
eosinophilic spongiosis, 585 nontumor, 216-217
epidermolysis bullosa acquisita, 586 polyposis, 216
erythema induratum, 588 colon
erythema multiforme (EM), 583 colitis cystica profunda, 240
erythema nodosum (septal), 588 GIST, 240
graft versus host disease (GVHD), 584 inflammatory cap polyps, 240
granuloma annulare, 586 inflammatory fibroid polyps, 240
granuloma faciale, 587 inflammatory polyps, 240
lichen planus, 583 leiomyoma, 240
lichen planus-like keratosis, 583 mucosal folds, 240
lichen simplex chronicus, 5 85 mucosal ganglioneuroma, 240
lupus erythematosus, 584 mucosal prolapse, 240
lymphocytic vasculitis, 587 neoplasms {nonepithelial), 239-241
membranous lipodystrophy (lobular), 589 neuroendocrine neoplasms, 239
necrobiosis lipoidica, 586 normal anatomy, 214
necrobiotic granuloma, 586 pneumatosis coli, 241
neutrophilic dermatosis, 587-588 polypoid lesions, 241
noninflammatory purpura, 588 submucosal lipoma, 240
pemphigus vulgaris, 586 endoscopic biopsy, 215
pigmented purpuric dermatosis/capillaritis, endoscopic mucosal resections, 215
588 large intestine
pityriasis rosea, 585 neoplasms, 233-238
porphyria cutanea tarda, 586 nonneoplastic conditions, 225-232
psoriasis, 584 normal anatomy, 214
sarcoidal granuloma, 5 87 mesenteric fibromatosis, 247
Stevens-Johnson syndrome (SJS), 583 mesentery, 247
950 INDEX
Intestine (Contd.) NETs, 241-242

neoplasms of colon (nonepithelial) neuroendocrine neoplasms, 241-244
colitis cystica profunda, 240 pseudomyxoma peritonei, 241
GIST, 240 sessile serrated adenoma/polyp, 241
inflammatory cap polyps, 240 signet-ring cell carcinoma, 241
inflammatory fibroid polyps, 240 tubular carcinoid, 244
inflammatory polyps, 240 nonneoplastic conditions of large
leiomyoma, 240 Brown bowel syndrome, 232
mixed adenoneuroendocrine carcinomas, diversion colitis, 230
240 diverticulosis, 231
mucosal folds, 240 drug-induced colitis, 232
mucosal ganglioneuroma, 240 GVI-ID,232
mucosal prolapse, 240 Hirschsprung's disease, 225-226
NET (carcinoid), 239 infectious colitis, 232
neuroendocrine carcinoma, 23 9 inflammatory bowel disease, 227-230
neuroendocrine neoplasms, 239-240 irritable bowel syndrome (IBS), 232
pneumatosis coli, 241 ischemic bowel disease, 226-227
polypoid lesions, 241 melanosis coli, 232
submucosal lipoma, 240 microscopic colitis, 231
neoplasms of large neuromuscular disorders, 225-226
adenocarcinoma, 236, 238 pseudo-obstruction syndrome, 226
adenoma, 233,235 radiation colitis, 231
FAP, 235 nonneoplastic conditions of small
hamartomatous polyps, 235 abetalipo proteinemia, 219
HGD (adenocarcinoma in situ), 236 ampullary carcinoma, 220
hyperplastic polyps, 233 autoimmune enteropathy, 218
lynch syndrome, 235 celiac disease, 217-218
mixed hyperplastic and adenomatous collagenoussprue,218
polyp, 235 common variable immunodeficiency, 219
sessile serrated adenoma, 233 congenital anomalies, 217
sessile serrated polyp, 233 cryptosporidiosis, 219
neoplasms of small eosinophilic gastroenteritis, 219
adenomas, 220 giardiasis, 219
adenomyoma of ampulla vater, 220 heterotopic gastric mucosa, 217
Brunner's gland hyperplasia, 220 heterotopic pancreas, 217
enteropathy-type T-celllymphoma, 225 infectious diseases, 219-220
gangliocytic paraganglioma, 222 lymphangiectasia, 219
GIST, 222 malabsorptive disorders, 217-219
hamartoma, 220 malrotation, stenosis, atresia, duplication,
immunoproliferative small intestinal and defects of musculature, 217
disease (IPSID), 222 Meckel's diverticulum, 217
neuroendocrine tumor, 220 microvillus inclusion disease, 219
nonneoplastic and neoplastic conditions of polyps and neoplasms, 220
anal canal, 244 refractory sprue, 218
anal squamous intraepithelial neoplasia strongyloidiasis, 220
(ASIN), 245 tropical sprue and bacterial overgrowth,
anal tag, 245 219
extramucosal (perianal) carcinoma, 246 polypectomy specimens, 215
hemorrhoids, 245 polyps of small intestine, 220
melanoma, 246 small intestine
Paget's disease, 246 neoplasms, 220, 222, 225
sec, 245 nonneoplastic conditions, 217-220
nonneoplastic and neoplastic conditions of normal anatomy, 214
appendix polyps, 220
acute appendicitis, 241 suction biopsy, 215
adenocarcinoma, 241 Intracranial pressure (ICP), 633-634
cystic fibrosis, 241 Intracystic papillary carcinoma (IPC), 318
goblet cell carcinoid, 242-244 Intracystic papillary neoplasms, 277
hyperplastic polyps, 241 Intradermal melanocytic nevus, 615
mixed carcinoid-adenocarcinoma Intraductal papillary neoplasms (IPN), 269,
(MANEC), 244 278,286,296
INDEX 951
Intraductal papillomas, 325 Isocitrate dehydrogenase (IDH1/IDH2)

Intraductal proliferative lesions, 322-323 mutations, 630
Intraductal tubulopapillary neoplasm (ITPN),
287 Jaws
Intrahepatic cholangiocarcinoma (ICC), 269 benign nonodontogenic tumors, 59
Intralobar nephrogenic rests, 350 cemento-osseous dysplasia, 61
Intraoperative consultations, 832-833, fibrous dysplasia, 60
836-837 juvenile ossifying fibroma, 60-61
Intraosseous ganglion, 814 epithelial odontogenic tumors
Intratubular germ cell neoplasia of unclassified adenoameloblastoma, 56
type (IGCNU), 458 ameloblastic carcinoma, 56
Intrauterine device (IUD), 548 ameloblastoma, 54-55
Invasive adenocarcinoma, 236, 543 calcifying, 56
Invasive breast cancer, 305 malignant ameloblastoma, 56-57
histologic grade, 310 squamous odontogenic tumors, 56
histologic subtype, 305-306, 308-311 giant cell lesions
IDC, 311-312 brown tumor of hyperparathyroidism, 62
inflammatory carcinoma, 314-315 central giant cell granuloma, 61-62
invasive lobular carcinoma (ILC), 312 cherubism, 62
invasive micropapillary carcinoma, 314 mesenchymal odontogenic tumors
invasive papillary carcinoma, 314 benign cementoblastoma, 57
lymphvascular space invasion, 311 cemento-ossifying fibroma, 57-58
medullary carcinoma, 313 odontogenic myxoma, 57
metaplastic carcinomas, 314 mixed odontogenic tumors
microinvasive carcinoma, 315 ameloblastic fibroma, 58
mucinous carcinoma, 313 ameloblastic fibrosarcoma, 59
pathology of treated, 315-316 odontogenic carcinosarcoma, 59
prognostic and predictive factors, 305 odontomas, 58
skin and nipple involvement, 311 nonodontogenic cysts, 53-54
surgical margin, 311 aneurysmal bone cyst, 54
tubular carcinoma, 313 epithelial-lined, 53
tumor size, 308, 310 fissural cysts, 53
Invasive squamous cell carcinoma (SCC) globulomaxillary cyst, 53
cervical intraepithelial, 542 incisive canal cyst, 53
skin, 596 median palatal cysts, 53
Invasive urothelial neoplasms, 386, 388-389 nasopalatine duct, 53
giant cell variant, 391 non-epithelial-lined, 54
with glandular differentiation, 389 simple bone cysts, 54
glycogen-rich (clear-cell) variant, 392 normal anatomy, 51
lipid-rich variant, 392 odontogenic cysts, 51-53
lymphoepithelioma-like variant, 391 calcifying cystic odontogenic tumor, 52
lymphoma-like variant, 391 dentigerous cyst, 51
microcystic variant, 391 developmental, 51-53
micropapillary variant, 391 gingival cyst, 52
nested variant, 390-391 glandular, 52-53
plasmacytoid variant, 391 inflammatory, 53
sarcomatoid variant, 390 keratocyst cyst, 51-52
with squamous differentiation, 389 lateral periodontal cyst, 52
with trophoblastic differentiation, 389, 390 Joints
Inverted papillomas, 40-41, 386 diseases
Iron infectious arthritis, 824-825
colloidal, 844 loose large joint arthroplasty associated
histology and histochemical stains, 84 7 synovitis, 824
overload (IO), 254 osteoarthritis, 823-824
Irritable bowel syndrome (IBS), 232 rheumatoid arthritis, 824
Irritant contact dermatitis, 585 gross examination and tissue sampling,
Irritation fibroma, 5 822
Ischemic bowel disease, 226-22 7 normal anatomy, 822
Ischemic cardiomyopathy, 155 replacement surgeries, 823
Ischemic heart disease, 157 Jones' methenamine silver (]MS), 845
Ischemic neuropathies, 675-676 Junctional melanocytic nevus, 614-615
952 INDEX
Juvenile granulosa cell tumors (JGCfs), membranoproliferative glomerulonephritis

503-504 (MPGN), 333-334
Juvenile nasopharyngeal fibroma, 753 membranous GN, 332-333
Juvenile ossifying fibroma, 60--61 mesangial proliferative
Juvenile papillomas, 23 glomerulonephritis, 332
Juxtaarticular myxoma, n3 postinfectious glomerulonephritis (PI GN),
Juxtaglomerular cell tumors, 356, 367 333
thin basement membrane disease,
Kaposi sarcoma (KS) 336-337
epithelioid, 271 immunofluorescence, 856-857
liver, 271 nonneoplastic cystic diseases
lung, 141 acquired cystic kidney disease, 346
oral cavity and oropharynx, 14-15 autosomal dominant PKD, 345
soft tissue, 769 autosomal recessive PKD, 345
spleen, 738 malformation syndromes associated cysts,
vascular tumor of skin, 602-603 346
Kaposiform hemangioendothelioma (KHE), medullary CKD, 346
768, 788 medullary sponge kidney, 34 5
Karyotypes, 874 nephronophthisis, 346
Kawasaki disease, 163 simple renal cysts, 346
Keloid, 605 normal anatomy, 344
Keratinization disorders, 10, 43, 542, pediatric, 347. See also Pediatric kidney
593-594 neoplasms
Keratinocytes, 579 renal biopsy, 329
Keratinous cysts tubulointerstitial diseases, 337-338
skin, 592 vascular diseases
vulva, 560 systemic HTN, 339
Keratoconus, 69 Takayasu's aortitis, 338
Keratocysts, 51 vasculitides, 338
Keratocystic odontogenic tumor, 51 Kidney neoplasms cytopathology. See also
Keratopathy, pseudophakic bullous, 68--69 Adult renal neoplasms; Pediatric kidney
Keratosis, 583, 595 neoplasms
Ki-67 biomarkers in breast cancer, 321 benign neoplasms, 3 71
Kidney angiomyolipoma, 371
allograft kidney pathology, 339 oncocytoma, 371
acute humoral (antibody-mediated) malignant neoplasms
rejection, 341 chromophobe renal cell carcinoma, 372
acute T-cell-mediated rejection, 339-341 clear cell RCC, 372
calcineurin inhibitor (CNI) toxicity, 343 metastatic tumors, 3 72
chronic antibody-mediated rejection, 341 papillary RCC, 372
chronic rejection, 341 urothelial carcinoma, 372
De novo disease, 342 Knots, umbilical cord, 569
infection, 343
posttransplant lymphoproliferative disease Lacrimal gland diseases, 73, 7 5
(PlLD), 342 Lacrimal sac diseases, 7 5
recurrent glomerular disease, 341 Langerhans cells, 579-580
renal allograft biopsies processing, 339 Langerhans cells histiocytosis (LCH),
transplant rejection, 339-341 659-660. See also Pulmonary
development abnormalities eosinophilic granuloma
multicystic dysplasia, 344 bone,809-810
renal dysplasia, 344 Large B-celllymphoma, mediastinal, 174
electron microscopy (EM), 839-840 Large cell carcinoma
glomerular diseases, 329 cervical neuroendocrine neoplasms, 545
Alport syndrome, 336 lung, 133, 147
amyloidosis, 335-336 Large cell lymphoma
crescentic glomerulonephritis, 334 anaplastic, 708
diabetes mellitus (DM), 334-335 lung, 144
focal segmental glomerulosclerosis, 330, lymph nodes cytopathology, 707-708
332 Large cell neuroendocrine carcinoma
lupus nephritis (LN), 334 (LCNEC), 136
MCD, 329-330 Large cell neuroendocrine tumor, 147
INDEX 953
Large intestine Lentigo simplex, 614

neoplasms, 233, 235-236, 238 Leprosy, 681
nonneoplastic conditions, 225-232 Leukemia
normal anatomy, 214 acute, 719, 721, 726, 736-737
Large vessel vasculitides, 162-163 B-lymphoblastic, 721
Laryngeal amyloidosis, 23 bone marrow, 719, 721, 726
Laryngocele, 23 chronic myelogenous (CML), 717, 736
Larynx hairy cell (HCL), 726, 728, 736
amyloidosis, 23 liver, 255
cysts, 23 lung, 144
endoscopic biopsies, 19 splenic, 736-737
frozen sections, 21 T-lymphoblastic leukemia/lymphoma, 726
inflammation and infection, 22 Leukoencephalopathy, 668-669
laryngeal carcinoma staging, 32 Leukoplakia, 5
mesenchymal tumors, 30 Lewy bodies, 631
neoplastic lesions Lewy body disease (LBD), 670
benign, 23, 25 Leydig cell tumors, 462, 505
malignant, 26-30 Lhermitte-Dudos disease, 648
premalignant, 26 Libman-5acks endocarditis, 151
neuroendocrine carcinomas, 29-30 Lichen planus, 3, 583
non-neoplastic lesions Lichen planus-like keratosis, 583
metabolic, 23 Lichen sclerosus, 560, 593
traumatic, 22-23 Lichen simplex chronicus, 560, 585
normal anatomy Light microscopy, 329, 926
macroscopic/gross, 19 Lingual thyroid, 406
microscopic, 19 Lipid
resections, 19-21 rich variant of invasive urothelial
sec, 26-29 neoplasms, 392
Leiomyomas storage disorders, 713
benign metastasizing, 529 Lipoblastoma, 747
cellular, 529 Lipodystrophy, 589
colon neoplasms (nonepithelial), 240 Lipofibromatosis, 752
deep soft tissue, 759-760 Lipoidica necrobiosis, 586
deep-seated, 785 Lipoleiomyoma, 529
epithelioid, 529 Lipoma
esophagus, 198 adipocytic tumors, 745-747
mesenchymal, 367 angiolipoma, 746
myxoid, 529 angiomyolipoma
pilar, 607 kidney neoplasms cytopathology, 371
skin, 607 mesenchymal, 366-367
symplastic, 529 pediatric kidney neoplasms, 356
uterus, 528-529 cardiac, 158
vaginal mesenchymal neoplasms, 554 chondroid, 746
vessels, 164 colon, 240
vulva, 562-563 epithelioid angiomyolipoma, 367
Leiomyomatosis inner ear, 103
hereditary, 360 mediastinal soft tissue neoplasms, 176
intravascular, 529 myelolipoma, 443
peritoneal disseminated, 179 myolipoma, 746
Leiomyosarcoma (LMS) pleomorphic, 746
adult renal neoplasms, 366 skin, 607
mesenchymal malignant neoplasms, spindle cell, 746
556-557 submucosal, 240
retroperitoneum, 785 synovial, 827
skin, 607-608 Lipomatosis, 746-747
uterus, 529-530 Liposarcoma (LPS)
vaginal, 556-557 atypical lipomatous tumor, 747-748
Leiomyosarcomafallopian tubes, 516 dedifferentiated, 748, 784
Leishmaniasis, 901 myxoid/round cell, 748, 785
Lentiginous type melanoma, 620 pleomorphic, 748-749, 785
Lentigo maligna melanoma, 620 well-differentiated, 747-748, 784
954 I INDEX
Listeria monocytogenes, 577 bacterial infections, 253

Lithiasis, 2 80 ductal plate malformation (DPM),
Littoral cell angioma, 737 259
Littoral cell hemangioendothelioma, 738 fungal infections, 253
Liver granulomas, 259
alcoholic liver disease (ALD), 253 metabolic and toxic liver diseases,
amyloidosis, 255-256 253-256
antitrypsin deficiency, a 1, 25 5 parasitic infections, 25 3
ascending cholangitis, 260 pregnancy and, 259
autoimmune hepatitis (AIH) disorders, 256 vascular disorders, 258-259
biliary origin tumors viral hepatitis, 251-252
benign, 268 normal anatomy, 248
malignant, 269 overlap syndrome, 257
premalignant, 269 partial hepatectomy, 249
Caroli's disease, 259 PBC disorders, 256
{J-catenin-activated adenomas, 265 polycystic liver disease, 259
congenital hepatic fibrosis, 259 primary sclerosing cholangitis (PSC)
cystic fibrosis, 256 disorders, 256-257
cytopathology, 271 Reye's syndrome, 255
angiosarcoma, 2 72 secondary biliary cirrhosis, 257-258
benign non -neoplastic masses, 271-272 segmentectomy, 249
cholangiocarcinoma, 273 total hepatectomy (explant), 249
combined hepatocellular- total parenteral nutrition (TPN), 25 5
cholangiocarcinoma, 2 73 transplantation pathology, 260--262
epithelioid hemangioendothelioma, 272 vascular disorders, 258-259
hemangioma, 272 wedge biopsy, 24 9
hepatocellular adenoma, 2 72 Wilson's disease, 255
hepatocellular carcinoma, 272 Lobular
metastatic malignancy, 2 73 adenocarcinoma, breast cytology, 327
nodular hyperplasia (NH), 272 inflammatory dermatoses, 588-589
vascular lesions, 2 72 Lobular capillary hemangioma (LCH), 601,
drug-induced liver injury (DILl), 256 765-766
electron microscopy (EM), 840 Lobular carcinoma, invasive, 312
epithelial tumors Lobular carcinoma in situ (LCIS), 319
benign hepatocellular tumors, 262-266 Lobular endocervical glandular hyperplasia,
DN, 266 538
fibrolamellar hepatocellular carcinoma Lobular hyperplasia, 323
(HCC), 267 Lobular neoplasia, 319
focal nodular hyperplasia (FNH), 262, 264 Loeffler endocarditis, 153
hepatocellular adenoma (HCA), 264-266 Loeffler syndrome, 114
hepatocellular carcinoma (HCC), 266-267 Loop electrosurgical excision procedure
malignant tumors, 266-267 (LEEP), 536
premalignant tumors, 266 Low-grade appendiceal mucinous neoplasm
glycogenic hepatopathy, 254 {LAMN), 241
glycogenoses, 255 Low-grade fibromyxoid sarcoma (LGFMS),
hematologic disorders, 255 756
hepatoblastoma, 268 Low-grade squamous intraepitheliallesion
HNF1a mutation, 265 (LSIL), 549
idiopathic adulthood ductopenia (lAD), 258 Lumpy jaw, 5
inflammatory adenoma (IA), 265-266 Lung
iron overload (IO), 254 adjunct information, 105
lobectomy, 249 allergic diseases, 114-115
lysosomal storage diseases, 255 bronchiolitis, 117
mesenchymal origin tumors, 270-271 cytopathology
mixed tumors, 267-268 adenocarcinoma, 147
needle core biopsy, 248-249 atypical cytology, 145
nonalchoholic fatty liver disease (NAFLD), carcinoid tumor, 14 7
253 endoluminal samples exfoliation, 145
nonneoplastic conditions, 250 hematopoietic malignancies, 148
autoimmune hepatitis (AIH) and bile duct hemosiderosis, 147
disorders, 256-258 large cell neuroendocrine tumor, 14 7
INDEX 955
mesothelioma, 148 lymphangioleiomyomatosis (LAM),

metastases, 148 113-114
negative for malignancy, 145 pulmonary eosinophilic granuloma, 113
neoplasms, 147-148 sarcoidosis, 112-113
neuroendocrine carcinoma, 147 normal anatomy, 104
non-neoplastic bacterial infection pneumoconiosis, 121-122
conditions, 146-147 pulmonary transplantation, 122-125
non-neoplastic fungal infection conditions, resections, 105
146 samples, 104
non-neoplastic viral infection conditions, specimen handling and reporting, 104-106
146 vasculitis and related diseases of, 115-116
positive for malignancy, 146 Lung neoplasms, 125
pulmonary alveolar proteinosis (PAP), adenoid cystic carcinoma (ACC), 140
147 biphasic pulmonary blastoma, 139
sample type, 145 fetal adenocarcinoma, 139
sarcoidosis, 147 hematolymphoid lesions, 143-144
sec, 147 inflammatory myofibroblastic tumor (IMT),
small cell carcinoma, 147 143
specimen adequacy and preparation, 145 metastatic tumors, 144-145
suspicious for malignancy, 146 mucoepidermoid carcinoma, 140
electron microscopy (EM), 840 neuroendocrine
endoscopic biopsies, 104 atypical carcinoid tumor, 135-136
immunofluorescence, direct, 857 carcinoid tumors, 134-135
infectious diseases chemodectomas, 135
adenovirus, 118 large cell neuroendocrine carcinoma, 136
Aspergillus, 119-120 neuroendocrine cell hyperplasia, 134
atypical mycobacterial infection, 119 paraganglioma, 135
bacterial, 118-119 small cell lung carcinomas (SCLC),
blastomycosis, 120 137-138
Candida infection, 119 non -small cell carcinomas
coccidiomycosis, 120-121 adenocarcinoma, 128-132
Cryptococcus, 120 adenosquamous carcinoma, 133
cytomegalovirus (CMV), 117-118 dear cell carcinoma, 133
dirofilariasis, 121 giant cell carcinoma, 133
fungal, 119-121 large cell carcinoma, 133
herpesvirus (HSV), 118 sec, 132-133
histoplasmosis, 120 pathologic reporting, 125-128
measles virus, 118 pleuropulmonary blastoma (PPB), 139-140
mucormycosis, 119 pulmonary dear cell tumor, 142
mycobacterial infections, 119 pulmonary hamartoma, 142
mycoplasma, 118 salivary gland tumors and, 140
parainfluenza and influenza, 118 sarcomas, 141-142
parasite, 121 sarcomatoid carcinoma, 138
Pneumocystis carinii pneumonia (PCP), Lupus, 338, 856
121 erythematosus, 584
respiratory syncytial virus (RSV), 118 nephritis (LN), 334
tuberculosis (TB), 119 Luteinalis, 495
varicella zoster (VZV), 118 Luteoma, 495, 505
viral, 117-118 Luxol fast blue, 847
microscopic description, 105-106 Lyme disease, 682
neoplasms. See Lung neoplasms Lymph nodes, 483
non-neoplastic disease benign diseases
acute lung injury patterns, 106-108 acute lymphadenitis, 685
alveolar proteinosis, 114 diffuse (paracortical) hyperplasia, 684
connective tissue disorders, 110 follicular hyperplasia, 684
drug-induced pulmonary changes, granulomatous lymphadenopathy, 684
110-111 sinus hyperplasia, 684
hypersensitivity pneumonitis (HP), breast pathology, 303, 326
111-112 Castleman disease, 700
idiopathic interstitial pneumonitis, hyaline-vascular, 701
108-110 plasma cell type, 701
956 INDEX
Lymph nodes (Contd.) deep, 602

cervical, 53 7 mediastinal soft tissue, 176
cytopathology, 706 mesenchymal, 367
anaplastic large cell lymphoma, 708 oral cavity and oropharynx, 14
Burkitt lymphoma, 707 retroperitoneum, 782
diffuse large B-celllymphoma (DLBCL), superficial, 602
707 vascular, 602, 768
female genital tract, 836 Lymphocytosis, 710
follicular lymphoma, 707 Lymphoepithelioma-like variant of invasive
Hodgkin lymphoma, 708 urothelial neoplasms, 391
intermediate cell lymphoma, 707 Lymphogranuloma venereum, 485
large cell lymphoma, 707 Lymphoid infiltrates (cutaneous), 608-611
lymphoblastic lymphoma, 707 Lymphoid interstitial pneumonitis (LIP), 110
lymphoplasmacytic lymphoma, 707 Lymphomas
mande cell lymphoma, 707 adrenal glands, 44 3
metastatic malignancy, 708 adult renal neoplasms, 368
myeloid sarcoma, 708 bone, 811
nodal marginal zone B-celllymphoma, 707 Burkitt, 707, 721
peripheral T-celllymphoma, unspecified CNS,659-660
type, 708 conjunctiva disease, 67
reactive lymphadenopathy, 706 esophagus, 199
sentinel lymph nodes, 835-836 follicular, 707
small lymphocytic lymphoma, 707 general lymphoma workup approach,
small-cell lymphoma, 707 690-692
thyroid lymph nodes, 836 hematolymphoid lesions, 144
general B-cell workup approach hepatosplenic marginal zone T-cell
conventional karyotyping, 692 lymphoma (HSlL), 735
cytogenetic analysis, 692 Hodgkin, 702-704, 706, 708, 734
flow cytometry, 691 intermediate cell, 707
leukocyte marker, 691 large cell, 707-708
primarily T cell- and NK cell-associated liver, 255
markers, 691 lymphoblastic, 707
general lymphoma workup approach, 690 mantle cell lymphoma (MCL), 736
FISH, 692 mediastinal, 174
leukocyte marker, 690 non-Hodgkin, 735
markers helpful in subclassifying mature parenchymal lesions, 443
B-celllymphomas, 691 primary CNS lymphomas (PCNSLs), 659
markers of clonality, 691 primary effusion, 185
markers of immaturity, 690 pyothorax-associated, 185
PCR, 692 retroperitoneum, 789
primarily B-cell associated markers, 691 serosal membranes, 185
primarily T cell- and NK cell-associated small cell, 707
markers, 691 splenic, 733-736
southern blot analysis, 691 stomach, 213
gross examination and tissue sampling, thyroid, 420
683-684 T-cell,225,685,687-688, 734
Hodgkin lymphoma, 702-706 T-lymphoblastic, 726
non-Hodgkin lymphomas urothelial, 391
incidence and epidemiology, 685 uterus, 534
pathophysiology, 685-688 Lymphomatoid granulomatosis (LYG), 144
non-neoplastic salivary glands conditions, Lymphomatoid papulosis (LyP), 611
95 Lymphoplasmacytic lymphoma, 707
normal anatomy, 6 83 Lymphoproliferative disorders
penis,483,484 cardiac transplants and, 161
plasma cell neoplasms, 692, 699-700 post-transplant (PlLD), 124-125, 342
soft tissue tumors, 766 primary cutaneous CD30+, 609, 611
staging, 706 Lynch syndrome, 235
Lymphangioleiomyomatosis (LAM), 113-114 Lysosomal storage diseases, 255
Lymphangioma
circumscriptum, 602 Malabsorptive disorders
cystic, 782 abetalipoproteinemia, 219
INDEX 957
autoimmune enteropathy, 218 Medullary carcinoma

celiac disease, 217-218 adrenal glands, 436-441
collagenoussprue,218 breast pathology, 313
common variable immunodeficiency, 219 pancreatic ductal adenocarcinoma (PDA)
eosinophilic gastroenteritis, 219 variant, 290
lymphangiectasia, 219 renal cell carcinoma (RCCs)
microvillus inclusion disease, 219 adult, 364
refractory sprue, 218 pediatric, 355
Malakoplakia thyroid cytopathology, 420
renal, 370 Medullary cystic kidney disease (MCKD),
retroperitoneum, 781 346
urethral, 400 Medullary sponge kidney, 345
urinary bladder, 3 82 Medullary thyroid carcinoma (MTC),
Malformation of cortical development 416-417
(MCD), 666 Medulloblastoma, 651-652
Malherbe, calcifying epithelioma of, 598 Megaloblastic anemia, 710, 710
Malignant fibrous histiocytoma (MFH) Melanin, 579
bone,809 Melanocytes, 579, 613
giant cell, 759 Melanocytic lesion, 614
inflammatory oral cavity and oropharynx, 13
fibrohistiocytic tumors, malignant, 759 skin (benign proliferations), 615-619.
retroperitoneum, 787 See also Nevus
Malignant mixed MUllerian tumor (MMMT), Melanocytic neoplasms
526-527,545-547 conjunctiva diseases, 64-65
Malignant peripheral nerve sheath tumor cornea diseases, 72-73
(MPNST), 787 Melanoma
MALT type (mucosa-associated lymphoid anal canal, 246
tissue type), 211-212 cervical, 547
MALToma, 143 esophagus, 199
Mammary implants, 303 sinonasal region, 44
Mantle cell lymphoma (MCL), 707, 736 skin, 619-621
Mast cell disease (MSD), 736-737 urethral, 403
Mastectomy specimens, 302-303 of uvea, 72-73
Mastocytosis, 719, 736-737 vaginal, 5 57
Maternal disease, 573-574 vulvar, 564
Maternal floor infarct, 570 Melanosis coli, 232
Maternal thrombophilic disorders, 574 Melting curve analysis, 898
Mature cystic teratoma, 173 Membranoproliferative glomerulonephritis
Measles virus, 118 (MPGN), 333-334
Meckel's diverticulum, 217 Membranous glomerulonephritis, 332-333
Meconium Membranous lipodystrophy (lobular), 589
placental fetal membranes disorders, 567 Menetrier's disease, 204
peritonitis, 178 Meninges tumors, 653--657
Median palatal cysts, 53 Meningiomas,653-656
Median raphe cyst, 486 Meningitis, 663
Mediastinal large B-celllymphoma, 174 Merkel cell carcinoma, 604
Mediastinitis, 167 Mesangial hypercellularity, 332
Mediastinum Mesangial proliferative glomerulonephritis,
biopsy, 165-166 332
cysts, 175 Mesenchymal chondrosarcoma
fine needle aspiration (FNA) biopsy, 165-166 bone,804
gross anatomy, 165 soft tissue, 770
neoplasms, 168 Mesenchymal hamartoma, 270
ganglioneuroma, 175 Mesenchymal neoplasms. See also
germ cell tumors, 173-174 Mesenchymal tumors
lymphomas, 174 cervical, 545
neuroblastoma, 175 esophagus, 198-199
neurogenic and neuroblastic, 174 leiomyosarcoma, 556-557
soft tissue, 176 oral cavity and oropharynx, 15, 18
thymic, 168-173 penile cancer, 489
resection, 166 prostate cancer, 4 79
958 INDEX
Mesenchymal neoplasms (Contd.) endocervicosis, 179

renal plevis cancer, 376 endometriosis, 178-179
serosal membranes, 184-185 endometrium, 520
ureter, 376 endosalpingiosis, 179
urethral, 400 eosinophilic, 471, 520
urinary bladder, 394, 395 intestinal, 383
vaginal, 554-556 mucinous, 471,520
Mesenchymal tumors. See also Mesenchymal nephrogenic, 383
neoplasms prostate, 471
adrenal glands, 44 3 squamous
adult renal neoplasms, 366-367 endometrium, 520
larynx, 30 placental, 568
liver, 270-271 urinary bladder, 382
odontogenic tumors of jaw, 57-58 squamous cell, 471
ovarian, 508 transitional cell, 471
sinonasal region, 45, 47 tubal, 520
skin, 601-607 urinary bladder, 383
Mesenteric inflammatory myofibroblastic urothelial, 4 71
tumor (IMT), 24 7 uterus, 520
Mesenteric inflammatory venoocclusive Walthard nests, 179
disease (MIVD), 247 Metaplastic carcinomas, 314
Mesenteritis, 24 7 Metaplastic epithelium, squamous, 535-536
Mesoblastic nephroma (MN), 352 Metastasis. See also Metastatic tumors
Mesonephric remnant cutaneous (from visceral carcinoma), 612
cervical, 53 9 esophagus, 199
hyperplasia of prostate, 472 neoplasms of middle ear, 102
Mesothelioma salivary glands, 93
biphasic, 184 vaginal, 55 7
desmoplastic, 184 Metastasizing tumors, 91, 529
diffuse malignant, 181-182, 184, 187 Metastatic tumors. See also under Metastasis
epithelioid, 182 adult renal neoplasms, 368
localized malignant mesothelioma, 184 adrenal glands, 444
lung cytopathology, 148 bone, 812
malignant paratesticular tumors, 463 CNS neoplasms, 660
multicystic peritoneal neoplasms, 187 fallopian tubes, 516
sarcomatoid, 184 kidney neoplasms cytopathology, 372
well-differentiated papillary, 184, 186 lung, 144, 145
Meta-analyses, 909 malignancy
Metabolic bone disorders, 816-817 adrenal glands cytology, 445
biopsy, 818 esophagus cytopathology, 200
chronic kidney disease- (CKDMBD), 819 liver cytopathology, 273
glucocorticoid-induced osteoporosis, 820 lung cytopathology, 148
mixed uremic osteodystrophy, 820 lymph nodes cytopathology, 708
osteomalacia, 819 ovarian neoplasms, 508
osteoporosis, 818-819 secondary malignancies, 376
primary hyperparathyroidism, 820 thyroid cytopathology, 420
renal osteodystrophy, 819-820 ureter cancer, 376
tissue sampling, and preparation, 818 parathyroid, 426
Metabolic diseases retroperitoneum, 789
bone. See Metabolic bone disorders spleen, 738
electron microscopy (EM), 840 uterus, 534
metabolic neuropathies, 676-677 Methotrexate therapy, 111
non-neoplastic lesions of larynx, 23 Methylation, 896-897, 911
Metanephric adenofibromas (MAFs), 352, 366 Microarray assays
Metanephric stromal tumors (MSTs), 351-352 specimen requirements, 906-907
Metaplasia statistical considerations, 906
cervix, 538 study design
ciliated cell, 520 class discovery, 906
clear cell, 520 class distinction, 906
disseminated peritonealleiomyomatosis, 179 single sample classification, 906
ectopic decidual reaction, 179 target preparation, 907
INDEX 959
Micro arrays Mixed uremic osteodystrophy, 820

analysis, 870 Moderately differentiated adenocarcinoma,
advantages, 871-872 295
limitations, 873 Mole. See Hydatidiform mole
bacterial artificial chromosome (BAC) Molluscum contagiosum, 485, 559, 589
arrays, 903, 905 Mongolian spot, 617
bead,905 Monoamniotic placentation, 574
eDNA, 905 Monoclonal gammopathy of undetennined
clinical applications, 910 significance (MGUS), 692
disease behavior prediction, 910 Monoclonal immunoglobulin deposition
DNA resequencing, 911-912 disease (MIDD), 336, 699-700
gene expression profiling, 910 Morphea, 593
genome copy number, 911 Morphometry, 674
genotyping, 911 Motor axonal neuropathy, 678
methylation, 911 Mucicarmine, 844
nontumor pathology, 911 Mucinous cystic neoplasm (MCN)
survival prediction, 910 gallbladder and extrahepatic biliary system,
therapeutic response prediction, 910-911 278
data analysis, 907 liver, 269
annotation, 908 pancreatic,285-286,296
cross validation, 909 Mucinous tumors
heat maps, 908 appendiceal, 241
hierarchical clustering, 908 borderline,500-501
image analysis, 907 breast pathology, 313
kappa-means clustering, 908 cervical, 543
meta-analyses, 909 cystadenomas, 500
normalization, 908 with intraepithelial carcinoma, 501
principal components analysis (PCA), 908 ovarian,500-501
sample set splitting, 909 tubular, 364
validation,909 Mucoceles, 84
visualization and data reduction, 908 Mucoepidermoid carcinoma {MEC)
in situ synthesized, 905 cervical, 544
oligonucleotide, 905 lung, 140
tissue, 923 salivary glands, 87-88, 96
uses in clinical laboratory Mucoid cysts, 486
array content, 912 Mucoid impaction of bronchi {MIB),
assay complexity, 913 115
automation and standardization, 912-913 Mucopolysaccharidoses, 713
clinical relevance, 913 Mucorales, 38
sample requirements, 913 Mucormycosis, 119
Microcysts, 324, 391 Mucosa-associated lymphoid tissue (MALT)
Microglandular hyperplasia, 538 type, 211-212
Microglia, 633 Mucosal diseases
Microinvasive carcinoma, 315, 542-543 cysts, 560, 591
Microorganisms, 846 esophagus injury, 192-194
Microsatellite instability (MSI) assays, 899 neuroma, 771
Microscopy, 926-928 prolapse, 240
Microvillus inclusion disease, 219, 841 Mucous impaction, 38
Minimal change disease (MCD), 329-330 Mucous membrane pemphigoid, 3
Mixed adenoneuroendocrine carcinomas, 240 MUllerian cysts, 552, 782
Mixed carcinoid-adenocarcinoma (MANEC), MUllerian tumors
244 retroperitoneum, 789
Mixed epithelial hyperplasia, 472 vaginal, 554-555
Mixed hyperplastic polyp, 235 Multicystic dysplasia, 344
Mixed oligoastrocytoma (MOA), 645 Multicystic mesotheliomas, 187
Mixed tumors Multicystic peritoneal inclusion cysts, 181,
liver, 267-268 782
odontogenic tumors, 58-59 Multilocular cystic RCC, 362
retroperitoneum benign fatty tumors, 784 Multinodular goiter, 408-409
salivary glands, 90-91 Multiple gestations, 567,574-575.
testis, 461 See also Placenta
960 INDEX
Multiple myeloma Myxoma

bone, 811 cardiac, 158
staging, 706 deep aggressi~e angiomyxoma, 554,773
Multiple sclerosis (MS), 66()-661 dermal depos1ts, 591
Multiplex metaphase FISH 868-869 digital, 773
Multiplex PCR, 896 ' intramuscular, 773
Mumps orchitis, 454 juxtaarticular, 773
Muscle. See Skeletal muscle; Smooth muscle odontogenic, 57
neoplasms ovarian, 508
Muscular dystrophies, 761 Myxopapillary ependymoma, 646
Mycobacterial infections, 119, 900
Nabothian cysts, 539
Mycobacterium avium intracellulare (MAl)
119 ' NAFLD liver, 253
Nasal vestibule, 41
Mycobacterium tuberculosis, 119
Nasolabial cysts, 7
Mycoplasma, 118
Nasopalatine duct cyst, 53
Mycosis fungoides, 608-609
Nasopharynx
Myelodysplastic syndromes, 713-714 716
' carcinoma (NPC), 42
Myelofibrosis, primary, 718-719
fibroma
Myeloid neoplasms, 736-737
angiofibroma, 45
Myeloid sarcoma, 708
juvenile, 753
Myelolipoma (adrenal glands), 443-444
normal anatomy, 34
Myeloma
Necrobiosis lipoidica, 586
multiple, 811
Necrobiotic granuloma, 586
plasma cell, 699-700
Necrosis
Myeloproliferative neoplasms, 716-719
acute tubular (ATN), 337
Myocardial ischemia, 157
bone, 796
Myocarditis, 153-154
CNS neurons pathology, 631
cardiac sarcoidosis and, 155
fat, 324
Chagas disease, 155
Necrotizing enterocolitis (NEC), 227
chronic active, 154
Necrotizing otitis externa, 98
eosinophilic, 154
Necrotizing sialometaplasia, 83
fulminant, 154
Neisseria gonorrhoeae, 485
granulomatous, 155
Neonatal necrotizing enterocolitis (NEC), 227
idiopathic giant cell, 154
Neoplasms
primary viral, 154
secondary, 155 anal canal, 244-246
appendix, 241
Myocardium disorders
benign epithelial, 374
cardiomyopathy, 155-157
bones. See Bone neoplasms
myocardial ischemia, 157
colon, 239-241
myocarditis, 153-155
conjunctiva diseases, 63-67
Myoepitheliomas, 87
cornea diseases, 71-73
Myofibroblastic tumors. See also
ear
Fibroblastic/myofibroblastic tumors
external, 98-100
inflammatory (IMT), 143, 247 754
urinary bladder, 394 ' inner, 102-103
middle ear, 101-102
Myofibroma-myofibromatosis 750-751
electron microscopy (EM), 840
Myointimal hyperplasia 247 '
esophagus
Myolipoma, 746, 783 '
cytopathology, 200
Myomectomy specimens, 151
epithelial, 194-198
Myopathies, 761
lymphoma, 199
Myopericytoma, 761
melanoma, 199
Myositis, 749-750
mesenchymal, 198-199
Myrmecialdeep palmoplantar wart, 590
metastasis, 199
Myxo~brosarcoma (MYFS), 756
gallbladder and extrahepatic biliary system
Myx01d 275-278, 280 ,
adrenal cortical neoplasms, 433
heart, 157-159
chondrosarcoma, 777
intestinal
dermal deposits, 591
large, 233, 235-236, 238
leiomyoma, 529
small, 220, 222, 225
liposarcoma, 748, 785
liver
valves disorders and, 153
biliary origin tumors, 268-269
INDEX 961
epithelial tumors, 262-267 plexiform neurofibromas, 771

mesenchymal origin tumors, 2 70-271 Schwannoma, 771-772
mixed tumors, 267-268 traumatic neuroma, 770
lung. See Lung neoplasms Nervous system
malignant urethral, 400 central. See Central nervous system (CNS)
mediastinum, 168-175 cytopathology, histology and histochemical
pericardium disorders, 157 stains, 847
plasma cell, 692, 699-700 Nested PCR, 894-895
salivary glands, 84 Nested variant of invasive urothelial
benign, 84-87, 9 5 neoplasms, 390-391
malignant, 87-93, 96 Neural neoplastic lesions, 15
serosal Neural tumors
benign, 181 ovarian, 508
malignant, 186-189 retroperitoneum, 787-788
stomach, 208-212 skin, 603--604
ureter, 374 Neurilemmoma, 604
urethra, 400 Neuroblastic neoplasms, 174
vessels, 164 Neuroblastoma (NB)
Neoplastic bowel resection, 215-216 adrenal glands, 4 3 8-441
Neoplastic lesions ganglioneuroblastoma (GNB), 440-441
extrahepatic biliary tree cytopathology, 281 mediastinum, 175
larynx, 23-30 olfactory, 45
oral cavity and oropharynx, 8-15 RCC associated with, 364
sinonasal region retroperitoneum, 787
benign, 38-41 sinonasal region, 45
malignant, 41-4 7 specimens, adrenal glands, 429
Nephrectomy, 358-359, 370, 428 undifferentiated, 355-356
Nephritis. See also Glomerulonephritis (GN) Neurocysticercosis, 664
allergic interstitial, 338 Neurocytoma, central, 648--649
lupus (LN), 334 N eurodegenerative disorders
Nephroblastoma, 348, 350 Alzheimer disease (AD), 669--670
Nephroblastomatosis, 350 Creutzfeldt-Jakob Disease (CJD), 670--671
Nephrogenic metaplasia, 383 Lewy body disease (LBD), 670
Nephrogenic rest, 348, 350 Neuroectodermal neoplastic lesions, 15
Nephroma Neuroendocrine neoplasms
cystic, 350, 368 adult renal neoplasms, 368
mesoblastic (MN), 352 appendix,241-244
Nephronophthisis, 346 atypical carcinoid tumor, 135-136
Nephropathy, 338 carcinoid tumors, 134-135
Nephroureterectomy, 374 cervical, 545
Nerve biopsy, 673 chemodectomas, 135
amyloid and related neuropathies, 680 colon neoplasms (nonepithelial), 239
genetic neuropathies, 679--680 esophagus, 198
infectious neuropathies, 681-682 large cell neuroendocrine carcinoma, 136
ischemic neuropathies, 675--676 larynx, 29-30
metabolic neuropathies, 676--677 lung cytopathology, 147
neuropathies with immune-mediated neuroendocrine cell hyperplasia, 134
mechanism, 678-679 oral cavity and oropharynx, 13-14
peripheral nerve, 673-674 pancreatic, 287, 293-294
pathologic mechanisms, 674-675 paraganglioma, 135
toxin-induced neuropathies, 675 sinonasal region, 44
traumatic neuropathy, 682 skin, 603--604
Nerve tumors small cell lung carcinomas (SCLC), 137-138
diffuse neurofibromas, 771 small intestinal neoplasms, 220
granular cell tumor, 772 thymic, 173
localized neurofibromas, 771 urinary bladder, 393-394
malignant, 772-773 Neuroepithelial tumor, dysembryoplastic, 649
mucosal neuroma, 771 Neurofibrillary tangles (NFTs), 632
neurofibromas, 771 Neurofibromas, 604,771
neurothekeoma, 772 Neurofilament (NF) protein, 629
perineurioma, n2 Neurohypophysis, 449
962 INDEX
Neuroma Nodular lymphocyte predominant Hodgkin

encapsulated, 603 lymphoma (NLPHL), 702
mucosal, n1 Nodular regnerative hyperplasia (NRH), 258
palisaded, 603 Nodular vasculitis (lobular), 588
skin, 603 Nodules
solitary circumscribed, 603 endometrial stromal, 527
traumatic, 603, 770 postoperative spindle cell, 554
Neuromuscular disorders, 225-226 vaginal, 554
Neuronal cytoplasmic inclusions (NCis), 632 Non-dear cell adenocarcinoma, 401
Neurons pathology. See under Central nervous Non-Hodgkin lymphomas. See also Hodgkin
system (CNS) lymphomas
Neuropathy pediatric kidney neoplasms, 356
acute motor axonal (AMAN), 678 retroperitoneum, 789
amyloid and related, 680 splenic, 734
antiglycolipid, antisulfatide, and N onkeratinizing tumors
antiganglioside neuropathies, 679 cervica~ 542
anti-Hu antibody, 679 oral cavity and oropharynx, 11
antimyelin associated glycoprotein (MAG), sinonasal region, 43
679 Non-Langerhans cell histiocytosis (LCH), 660
autoimmune autonomic, 679 Nonmelanocytic pigmented lesions, 624
hereditary, 679-680 Nonneoplastic conditions
hypertrophic (onion-bulb) neuropathies, 679 cystic diseases
immune-mediated mechanism, 678-679 kindey, 345-346
infectious, 681, 682 pancreas, 284-285
ischemic, 675--676 ear
metabolic, 676-677 external, 98
specimens, electron microscopy (EM), 841 middle, 100
toxin-induced, 675 esophagus, 191-194
traumatic, 682 large intestinal, 225-232
Neurothekeoma, n2 larynx, 22-23
Neutrophilic dermatosis, 587-588 liver, 250-253
Nevoid type melanoma, 621 lung, 106-114, 146-147
Nevus oral cavity and oropharynx, 5-8
ambiguous, 619 salivary glands conditions, 95
with architectural disorder, 618-619 serosal membranes, 177-180
blue, 617 sinonasal region, 38, 40
cell aggregates, breast pathology, 326 stomach, 202-205
cellular blue, 617 N onossifying fibroma, 808
Clark, 618 Non-specific interstitial pneumonitis (NSIP),
combined melanocytic, 619 109
compound melanocytic, 615 Nontumor bowel resection, 216-217
congenital and congenital-pattern, 617--618 Normal pressure hydrocephalus (NPH), 633
deep penetrating, 618 Normalization, 908
dysplastic, 618 Nuchal-type fibroma, 751-752
epithelioid cell, 616 Nucleic acid, 923
halo, 618
Hori, 617 0 bliterative bronchiolitis, 117
intradermal melanocytic, 615 Obliterative cardiomyopathy, 156
of Ito, 617 Ocular cicatricial pemphigoid, 63
junctional melanocytic, 614 Odontogenic carcinosarcoma, 59
lipomatosus, 74 7 Odontogenic cysts, 51-53
of Ota, 617 Odontogenic tumors of jaw, 54-59
pigmented spindle cell nevus of Reed, 617 Odontomas, 58
sebaceous of Jadassohn, 601 Oil red 0 stain, 846
on special sites, 619 Olfactory neuroblastoma, 45
spindle cell, 616 Oligoastrocytoma, 645
Spitz, 616--617 Oligodendrogliomas, 644
New variant CJD (nvCJD), 671 Oligonucleotide microarrays, 905
Next-generation sequencing (NGS), 897-898 Oilier disease, 802
Niemann-Pick disease, 713 Oncocytoma
Nodal marginal zone B-celllymphoma, 707 adult renal neoplasms, 365-366
INDEX 963
conjunctiva disease, 67 Otosclerosis, 100

kidney neoplasms cytopathology, 3 71 Ovarian neoplasms, 495
Oophoritis, autoimmune, 494 germ cell tumors, 505-507
Opportunistic infections, 122-123 gonadoblastoma, 507-508
Oral cavity and oropharynx hypercalcemic small cell carcinoma, 508
benign diseases mesenchymal origin, 508
infections, 4-5 metastases, 508
inflammation, 2-3 primary hematopoietic malignancies, 508
neoplastic lesions, 11 reporting, 510
non-neoplastic lesions, 5-8 sex cord-stromal tumors
fibrous lesions adult (AGCTs), 503
amalgam tattoo, 8 fibromas, 504
buccal exostosis, 6 Hilus cell tumors, 505
cysts, 6-7 juvenile UGCTs), 503-504
fibrous gingival hyperplasia, 6 Leydig cell tumors, 505
Fordyce granules, 6 sclerosing stromal tumors, 505
inflammatory papillary hyperplasia, 6 Sertoli cell tumors, 504
irritation fibroma, 5 Sertoli-Leydig cell tumors (SLCT), 504,
pseudoepitheliomatous hyperplasia, 7 505
torus mandibularis, 6 sex cord tumors with annular tubules
torus palatinus, 6 (SCTAT), 505
frozen sections, 2 steroid cell tumors, 505
neoplastic lesions, 8 stromal luteomas, 505
benign epithelial, 9-13 thecomas, 504
melanocytic, 13 staging, 510
mesenchymal, 15, 18 surface epithelial-stromal tumors
neural/neuroectodermal, 15 benign Brenner tumors, 502
neuroendocrine carcinomas, 14 benign endometrioid tumors, 501
vascular, 14-15 borderline Brenner tumors, 502
normal anatomy, 1 borderline endometrioid tumors, 501
open and endoscopic biopsies, 2 borderline mucinous tumors, 500-501
pathologic reporting aspects carcinosarcoma, 503
staging, 18 clear cell tumors, 502
resections, 2 endometrioid adenocarcinomas, 502
Oral hairy leukoplakia, 5 endometrioid tumors, 501-502
Orbit diseases, 77 malignant Brenner tumors, 502
Orchiectomy, 452-453 mucinous tumors, 500-501
Orchitis, 454 serous tumors, 499-500
Organoid nevus, 601 transitional cell carcinomas (TCC), 502
Oropharynx, 1. See also Oral cavity and Ovary
oropharynx autoimmune oophoritis, 494
Orthokeratosis, 581 cysts, 494-495
Osler-Weber-Rendu syndrome, 259 endometriosis, 494
Ossifying fibromyxoid tumor, 774 gross examination and tissue sampling, 493
Ossifying renal tumor of infancy, 356 hyperplasia, 495
Osteoarthritis, 823-824 infection, 493-494
Osteoblastoma, 805 inflammatory diseases, 493-494
Osteoblasts, bone-forming, 816-817 noninfectious granulomas, 494
Osteochondroma, 798 normal gross and microscopic anatomy, 493
Osteoclast-like giant cells, 291, 296 pregnancy changes, 495
Osteoclasts, bone-resorbing, 816-817 torsion, 495
Osteodystrophy, 819-820 Overlap syndrome, 257
Osteofibrous dysplasia, 808 Oxophilic cystadenoma. See Oncocytoma
Osteoid osteoma, 805
Osteoma, 805 Paget's disease, 246, 318, 489, 564, 797
Osteoma cutis, 591-592 Palisaded neuroma, 603
Osteomalacia, 819 Palmoplantar wart, myrmecia/deep, 590
Osteoporosis, 818-820 Pancreas
Osteosarcoma, 770, 805-807 biopsy, 282
Otitis extema, 98 brushing cytology specimens, 282
Otitis media, 100 distal pancreatectomy, 282-283
964 I INDEX
Pancreas (Contd.) Papillary carcinoma. See also Papillary

exocrine pancreas solid tumors, 287-293 neoplasms
fine needle aspiration, 282 breast pathology, 314, 317-318
heterotopic, 217 renal cell (RCC)
neoplastic cystic lesions, 287 adult renal neoplasms, 363
cystic acinar cell cystadenoma, 287 clear cell, 365
cystic pancreatic neuroendocrine tumors, hereditary, 360
287 kidney neoplasms cytopathology, 372
intraductal tubular adenoma, pyloric pediatric, 353
gland type, 287 squamous cell (SCC)
IPMN,286 cervical, 542
ITPN, 287 oral cavity and oropharynx, 13
MCN, 285-286 thyroid neoplasms, 412-413,415
serous cystadenocarcinoma, 285 Papillary cystadenoma
serous cystadenoma, 285 epididymis, 463
neuroendocrine neoplasms, 293-294 lymphomatosum, 84, 86, 95
nonneoplastic cystic lesions, 284 Papillary endothelial hyperplasia (PEH), 765
ductal retention cyst, 285 Papillary glioneuronal tumor (PGNT), WHO
lymphoepithelial cyst, 285 grade I, 649
paraampullary duodenal wall cyst, 285 Papillary neoplasms, 277, 386. See also
pseudocyst, 284 Papillary carcinoma
normal anatomy, 282 Papillary thyroid carcinoma (PTC), 420
pancreatic ductal adenocarcinoma (PDA) Papilloma
variants, 290-291 choroid plexus papilloma, WHO grade I,
Whipple procedure, 283 646
Pancreas cytopathology, 294-295 intraductal,breast pathology, 325
chronic pancreatitis, 295 fallopian tubes, 513
cystic lesions MUllerian,554-555
IPMN,296 sinonasal region, 38, 40, 41
MCN,296 squamous
pseudocysts, 296 esophagus, 194
serous cystadenoma, 296 urinary bladder, 392
neoplasms solid lesions vaginal, 5 52
acinar cell carcinomas, 296 urinary bladder, 386
adenocarcinoma, 295 vaginal mesenchymal neoplasms, 554-555
moderately differentiated Paraganglioma
adenocarcinoma, 295 extraadrenal, 441-443, 787-788
mucinous noncystic carcinoma, 295 gangliocytic, small intestinal neoplasms, 222
pancreatic endocrine neoplasms (PEN), larynx, 25
296 lung, 135
poorly differentiated adenocarcinoma, neoplasms of middle ear, 101
295 urinary bladder, 393
signet-ring adenocarcinoma, 295 Parainfluenza, 118
solid-pseudopapillary neoplasm, 296 Parakeratosis, 581, 594
undifferentiated carcinoma, 295-296 Paranasal sinuses, 34
well-differentiated adenocarcinoma, 29 5 Paraneoplastic pemphigus, 3, 857
Pancreatectomy, distal, 282-283 Paraovarian cysts, 494
Pancreatic ductal adenocarcinoma (PDA), Paraparesis, tropical spastic (TSP), 662
287-291 Paraphimosis, 484
Pancreatic intraepithelial neoplasia (PaniN), Parasitic cervicitis, 538
292-293 Parasitic cysts, 444
Pancreatitis, 283-284, 295 Parasitic encephalitis, 664-665
Pancreatoblastoma, 291 Parasitic infections
Panniculitis, 588-589 liver, 253
Pap smear test, 536, 828. See also Cervix lung, 121
Papillary Parasympathetic paragangliomas, 441-44 2
adenoma, adult renal neoplasms, 365 Parathyroid, 422
larynx, 28 autotransplantation for hyperplasia, 424
tumors cytopathology, 427
middle ear, 101-102 gross examination and tissue sampling, 422
of pineal region (PTPR), 650-651 neoplasms
INDEX 965
adenoma, 424 mesenchymal neoplasms, 489

carcinoma, 425-426 pathologic staging of, 490
histologic grading, staging, and reporting, reporting, 490, 492
426 risk factors, 486
metastatic tumors, 426 secondary malignancies, 489
nonneoplastic diseases squamous cell carcinoma, 487-489
abnormal development, 423 Penile cysts, 486
cysts, 423 Penile intraepithelial neoplasia (PelN), 486
hyperplasia, 423-424 Penis
parathyroiditis, 423 scrotum gross examination and tissue
normal anatomy, 422 sampling, 483
Parathyroid hormone (PTH), 422 inflammation and infection, 485
Parathyroiditis, 423 penile intraepithelial neoplasia (PeiN), 486
Parathyromatosis, 424 Penis and scrotum, 485
Parenchymal lesions benign epithelial neoplasms, 486
adrenal glands, 443 gross examination and tissue sampling, 483
CNS neoplasms, 650--651 foreskin resection, 483
Parvovirus B19 infections, 576-577 lymph nodes, 484
Pearly penile papules, 606 partial penectomy specimens, 483
Pediatric kidney, 347 punch and shave biopsies, 483
Pediatric kidney neoplasms, 351. See also total penectomy specimens, 483
Adult renal neoplasms inflammation and infection
angiomyolipomas, 356 balanitis, 484
dear cell sarcoma of kidney (CCSK), balanoposthitis, 484
352-353 chancroid, 485
desmoplastic small round cell tumor, 356 elephantiasis, 486
Ewing sarcoma/primitive neuroectodermal gangrene, 485
tumor, 355-356 gonorrhea, 485
gross examination and tissue sampling, granuloma inguinale, 485
347-348 herpes simplex virus, 484
juxtaglomerular cell tumors, 356 human papillomavirus (HPV), 484
mesoblastic nephroma (MN), 352 idiopathic scrotal calcinosis, 485
metanephric tumors, 351-352 lipogranulomas, 485
nephroblastoma, 348 lymphogranuloma venereum, 485
nephroblastomas, 350 molluscum contagiosum, 485
nephrogenic rests, 350 paraphimosis, 484
non-Hodgkin lymphoma, 356 pediculosis pubis, 485
ossifying renal tumor of infancy, 356 penile infections in AIDS, 485
pathologic staging, 35 6 phimosis, 484
pediatric renal cell carcinoma (RCCs), plasma cell balanitis, 484
353-355 scabies, 484
reporting, 357 syphilis, 485
rhabdoid tumor of kidney (RTK), 353 normal anatomy, 483
rhabdomyosarcoma (RMS), 356 Pentachrome stain, 846
synovial sarcomas, 356 Periapical cemento-osseous dysplasia, 61
teratoid Wilms tumor, 350 Peribiliary gland hamartoma, 268
undifferentiated neuroblastoma, 355-356 Pericardia!
Wilms tumor, 350 cysts, 180
Pediculosis pubis, 485 disorders, 157, 188-189
Peliosis, 737 Pericarditis, 157, 177
Pemphigoid Pericytic (perivascular) tumors
bullous inflammatory dermatoses, 585 benign, 760-761
immunofluorescence, 856-857 malignant, 761
ocular cicatricial, 63 Perilobar nephrogenic rests, 350
Pemphigus vulgaris, 3, 586, 856-857 Perineurioma, 772
Penectomy, 483 Periodic acid-schiff (PAS), 845, 847
Penile cancer. See also Penis and scrotum Periodontal cysts, 52
clinical diagnosis, 486 Peripheral nerve tumors, 770-773,787
hematolymphoid neoplasms, 489 Peritoneal inclusion cysts
histologic grading of, 489 multicystic, 181, 7 82
histologic typing and diagnosis, 486-487 serosal membranes, 180
966 INDEX
Peritonitis, 177-178, 180 massive subchorial thrombosis, 571

Perivascular tumors, 760-761, 775 maternal floor infarct, 570
Perivenous encephalomyelitis, 661 retroplacental thrombohematomas,
Peutz-Jeghers polyp, 220 570
Peyronie disease, 486 subamniotic hematomas, 571
Pharynx, 34. See also Nasopharynx subchorionic fibrin deposition, 570
Pheochromocytoma, 437-438, 445 fetal membranes disorders
Phimosis, 484 amnion nodosum, 567-568
Photodetectors, 861 amniotic bands, 568
Phthisis bulbi, 71 fetus papyraceous, 568
Phyllodes tumor, 325 hemosiderin deposition, 567
Pigmented adenoma, 433 meconium, 567
Pigmented purpuric capillaritis, 588 squamous metaplasia, 568
Pigmented purpuric dermatosis, 588 gross examination and tissue sampling,
Pigmented spindle cell nevus of Reed, 617 566
Pigmented villonodular synovitis, 826 disk, 567
Pigments and minerals, 847 fetal membranes, 566
Pilar leiomyoma, 607 multiple gestation, 5 67
Pill esophagitis, 193 umbilical cord, 566
Pilocytic astrocytoma (PA), 642--643 implantation disorders
Pilomatrixoma, 598 bilobed placenta, 573
Pilomyxoid astrocytoma (PMA), 643 extrachorialis, 572
Pindborg tumor, 56 placenta accreta, 572
Pineal parenchymal tumors, 650-651 shape abnormalities, 572-573
Pineoblastoma, 650 succenturiate (accessory) lobe, 573
Pineocytoma, 650 infection, 575-578
Pituicytoma, 449 maternal disease, 573
Pituitary gland, 446 diabetes, 573-574
atypical adenomas, 450 pre-eclampsia, 573
benign neoplasms sickle cell disease, 574
adenoma, 44 7 thrombophilic disorders, 574
adenomas, 448 multiple gestations
corticotroph (ACTH-producing) diamniotic dichorionic, 574
adenoma, 449 diamniotic monochorionic placentation,
GCT of neurohypophysis, 449 574
gonadotroph adenomas, 449 monoamniotic placentation, 574
pituicytoma, 449 TITS,575
prolactinoma, 448 normal anatomy, 5 66
somatotroph (GH cell) adenoma, 449 trophoblastic tumor, 533
high-grade neoplasms, 450 umbilical cord disorders
intraoperative evaluation and tissue abnormal cord insertion, 568
handling, 446 coiling, 569
low-grade neoplasms, 450 hematomas, 569
nonneoplastic lesions length abnormalities, 568
granulomatous hypophysitis, 447 marginal insertion, 568
lymphocytic hypophysitis, 447 single umbilical artery (SUA), 568
pituitary apoplexy, 447 stricture, 569
pituitary hyperplasia, 44 7 umbilical cord knots, 569
Rathke's cleft cyst, 44 7 velamentous insertion, 568
normal anatomy and histology, 446 Plasma cell
Pityriasis rosea, 585. See also Inflammatory balanitis, 484
dermatoses myeloma
Placenta genetics, 700
circulatory disorders, 569 staging, 706
chorangiomas, 572 neoplasms, 692, 699-700,706
chorangiosis, 572 type Castleman disease, 70 1
fetal thrombotic vasculopathy, 571 Plasmacytoid variant of invasive urothelial
infarcts, 569 neoplasms, 391
intervillous thrombohematomas, 571 Plasmacytomas, 699, 811
marginal hematomas, 571 Platelet-derived growth factors, 719
massive perivillous fibrin deposition, 570 Pleomorphic adenoma (PA), 84, 95
INDEX 967
Pleomorphic lobular carcinoma in situ (LCIS), endometrial, 520-521

319 esophagus, 194
Pleomorphic malignant fibrous histiocytoma fibroepithelial
(MFH), 758-759 vaginal, 552
Pleomorphic sarcoma, 758-759, 787 vulva, 560
Pleomorphic xanthoastrocytoma (PXA), 643 gastric, 205
Pleural fibrosis, 179-180 hamartomatous, 235
Pleural neoplasms, 181-186 hyperplastic
Pleural plaques, 179-180 appendix, 241
Pleuritis, 177 large intestinal neoplasms, 233
Pleuropulmonary blastoma (PPB), 139-140 inflammatory (colon neoplasms
Plexiform fibrohistiocytic tumor (PFH), 758 (nonepithelial), 240
Plexiform neurofibromas, 771 cap,240
Plexus tumors, 646-647 fibroid, 240
Pneumatosis coli, 241 large intestinal neoplasms, 235
Pneumoconiosis, 121-122 larynx, 22
Pneumocystis infection, 114 See also Lung mixed hyperplastic and adenomatous, 235
Pneumonia Peutz-Jeghers, 220
acute interstitial (AlP), 107 sessile serrated
bacterial, 147 appendix, 241
bronchiolitis obliterans-organizing (BOOP), large intestinal neoplasms, 233
107-108 small intestinal, 220
desquamative interstitial (DIP), 109-110 urethral, 400
eosinophilic, 114 urinary bladder, 385
pneumocystis carinii (PCP), 121 uterus, 520--521
Pneumonitis, 10 8-112 Polyradiculoneuropathy, 678-679
POEMS, 700 Poorly differentiated carcinoma
Polyarteritis, 116, 163, 338 insular
Polycystic diseases malignant cytopathology, 420
kidney, 345 thyroid neoplasms, 411-412
liver, 259 pancreatic, 294-295
ovarian syndrome, 494 Porokeratosis, 594
Polycythemia vera, 717-718 Poroma group eccrine tumors, 599-600
Polyembryoma, 507 Porphyria cutanea tarda, 586
Polymerase chain reaction (PCR) Portal vein thrombosis (PVf), 259
allele-specific PCR, 898 Postinfectious glomerulonephritis (PIGN),
implications for clinical testing, 893-894 333
methodology Postneuroblastoma renal cell carcinoma
advantages, 890-892 (RCCs), 355
amplification, 889-890 Postoperative spindle cell nodule
limitations, 892 cervical, 539
testing affecting factors uterus, 531
analytic/technical level, 890-892 vaginal mesenchymal neoplasms, 554
diagnostic level, 892-893 Posttransplant lymphoproliferative disorder
operational level, 893 (PTill), 124-125, 342
variations Pouchitis, 230
methylation-specific PCR, 896-897 Pregnancy
multiplex PCR, 896 changes
nested, 894-895 hyperreactio luteinalis, 495
quantitative PCR, 895-896 pregnancy luteoma, 495
RT-PCR, 895 solitary luteinized follicle cysts, 495
Polymer-based labels, 853 complete molar pregnancies, 531-532
Polymorphous low-grade adenocarcinoma ectopic, 513
(PLGA), 88-89 endometrium and, 518-519
Polyneuropathy, 676-678 invasive molar pregnancies, 533
Polyorchidism, 453 liver diseases and, 259
Polyploidy, 878 partial molar pregnancies, 533
Polypoid, 241, 382 Prenatal studies, 840
Polyposis bowel resection, 216 Primary acquired melanosis (PAM), 65
Polyps Primary angiitis of CNS (PACNS), 668
anal canal, 245 Primary CNS lymphomas (PCNSLs), 659
968 INDEX
Primary pigmented nodular adrenocortical mixed epithelial, 4 72

disease (PPNAD), 431 nodular stromal, 4 71
Primary sclerosing cholangitis (PSC), 257, stromal, 4 71-4 72
275,281 usual nodular epithelial, 471
Primitive neuroectodermal tumor (PNET) verumontanum gland, 4 72
adult renal neoplasms, 368 inflammation and infection, 469-470
bone, 813 metaplasia, 471
CNS-PNET, WHO grade IV, 652 normal anatomy, 467
pediatric kidney neoplasms, 355-356 pathologic staging, 480
soft tissue tumor, 775-776 prostatic intraepithelial neoplasia (PIN),
Principal components analysis (PCA), 908 472-473
Profunda colitis cystica, 240 seminal vesicles and, 479
Progesterone, 320, 519 urethra {prostatic), 479
Progressive multifocalleukoencephalopathy Prostatectomy, 468
(PML), 662 Prostatic intraepithelial neoplasia (PIN),
Prolactinoma, 448 472-473
Prolapse, mucosal, 240 Prostatitis, 4 70
Prominent lymphoid follicles, 240 Protozoans infectious disease testing, 901
Prostate Psammomatoid juvenile ossifying fibroma,
atrophy, 470-471 60-61
cancer, 474 Pseudocarcinomatous epithelia! hyperplasia,
acinar adenocarcinoma, 474 383
acinar adenocarcinoma variants, 477-478 Pseudocysts
adenocarcinoma, histologic grading of, adrenal glands, 444
479-480 panoreatic,284,296
clinical diagnosis, 4 74 Pseudoepitheliomatous hyperplasia, 7
ductal adenocarcinoma, 4 78 Pseudolymphoma, lung, 144
gross diagnosis, 474 Pseudomelanosis, 204
hematolymphoid neoplasms, 479 Pseudomembranous colitis, 232
histologic typing and diagnosis, 474 Pseudomyogenic hemangioendothelioma, 768
hormonal androgen deprivation therapy, Pseudomyxoma peritonei, 188, 241
479 Pseudoneoplasms, 737-73 8
immunohistochemical studies, 477 Pseudoneoplastic lesions, 73 8
mesenchymal neoplasms, 4 79 Pseudo-obstruction syndrome, 226
microscopic diagnosis, 474, 476-477 Pseudopapillary neoplasms, 292, 296
molecular studies, 4 77 Pseudophakic bullous keratopathy, 68-69
pathologic staging, 480 Pseudoxanthoma elasticum, 593
radiation therapy, 479 Psoriasis, 584
rare types of, 4 78-4 79 PTAH stain, 846
reporting, 481-482 Pubis, pediculosis, 485
risk factors, 474 Pulmonary alveolar proteinosis (PAP), 147
secondary malignancy, 4 79 Pulmonary transplantation
treatment effects, 479 biopsies, microscopic features of
focal glandular atypia (ASAP), 4 73 acute rejection, 123
gross examination and tissue sampling, 467 chronic airway rejection, 124
cystoprostatectomy, 469 humoral rejection, 123
needle cores, 467 opportunistic infections, 122-123
open suprapubic or retropubic simple post-transplant lymphoproliferative
prostatectomy, 468 disorder {PlLD), 124-125
pelvic lymph nodes, 468 preservation injury, 122
prostate gland and seminal vesicles, recurrent disease, 125
468-469 gross processing of transplant specimens,
radical prostatectomy, 468 122
TURP chips, 468 Pulmonary valves disorders, 153
hyperplasia Pulp
atypical adenomatous hyperplasia red, 729
adenosis, 472 white, 729
basal cell, 4 72 Punch biopsies, 581-582
cribriform, 4 72 Purpura, 588
epithelial predominant BPH nodules, 4 72 Pyelonephritis, xanthogranulomatous (XGP),
mesonephric remnant, 4 72 370
INDEX 969
Pyogenic granulomas, 14, 601 malignant mesenchymal neoplasms, 376

Pyothorax-associated lymphoma, 185 pathologic staging, 376, 377
reporting, 377-378
Quantitative PCR (Q-PCR), 895-896 sec, 376
Quilty effect, 159 small cell carcinomas, 376
urothelial carcinoma, 3 75
Racial melanosis, 64 gross examination and tissue sampling, 373
Radial scar/complex sclerosing lesions, 325-326 needle core biopsies, 3 73
Radiation pyeloplasty specimens, 373
colitis, 231 radical nephroureterectomy with bladder
esophagus mucosal injury, 193 cuff, 374
induced cardiomyopathy, 157 ureteroscopic biopsies, 373
therapy for prostate cancer, 479 normal anatomy, 3 73
Radical cystectomy, 3 73 ureteropelvic junction (UPJ) abnormalities,
Radical hysterectomycervix, 537 374
Radical nephrectomy, 428 obstruction (UPJO), 374
Radical nephroureterectomy, 374 Renal tuberculosis, 3 71
Radical prostatectomy, 468 Renomedullary interstitial cell tumors, 367
Rathke's deft cyst, 44 7 Renovascular hypertension, 339
Reactive astrocytosis, 632 Resequencing, DNA, 911-912
Reactive endocervical cells, 548 Residual cysts, 53
Reactive gastropathy, 204 Respiratory epithelial adenomatoid
Reactive lymphadenopathy, 706 hamartoma (REAH), 38
Reactive peritoneal fibrosis, 180 Respiratory syncytial virus (RSV), 118
Reactive pleural fibrosis, 179 Respiratory viruses, 90 1
Reactive splenic disorders, 733 Restrictive (obliterative) cardiomyopathy, 156
Reactive urothelial atypia, 383 Rete, testis tumors, 463
Reactive vascular proliferations, 765 Retention cysts, ductal, 285
Recurrent diseases Reticulin stain, 845
glomerular, 341 Retiform hemangioendothelioma, 769
pulmonary transplantation biopsies, Retinal anlage tumor, 463
microscopic features of, 125 Retinal detachment, chronic, 70
Reed, pigmented spindle cell nevus of, 617 Retinal vascular disease, 70
Reflux esophagitis, 192 Retinoblastoma, 73
Refractory sprue, 218 Retroperitoneal abscesses, 782
Rejection Retroperitoneal teratoma, 788-789
kidney transplantation, 339-341 Retroperitoneum
liver transplantation, 260 cystic lesions, 782-783
Renal biopsy. See under Kidney fibroblastic tumors, 786
Renal carcinoid tumors, 36 8 germ cell tumors, 788-789
Renal cell carcinoma (RCCs) inflammatory diseases, 781-782
acquired cystic disease-associated, 364-365 lymphomas, 789
adult renal neoplasms, 360, 362-365, 368 neoplasms, 783-789
associated with Xp11.2 translocations, 364 nonneoplastic diseases
chromophobe, 363 cystic lesions, 782-783
dear cell, 360-362, 372 idiopathic retroperitoneal fibrosis, 782
dear cell papillary RCC, 365 inflammatory, 781-782
cutaneous metastasis, 612 normal anatomy, 781
hereditary papillary, 360 neural tumors, 787-788
multilocular cystic, 362 soft tissue tumors, 783-789
neuroblastoma associated, 364 Retroplacental thrombohematomas, 570
papillary, 363, 372 Reverse transcriptase PCR (RT-PCR), 895
pediatric, 353-355. See also Pediatric kidney Reye's syndrome, 255
neoplasms Rhabdoid tumor, 353, 777-778
unclassified, 364 Rhabdomyoma, 158, 554,762
ureter cancer, 375-376 Rhabdomyosarcoma (RMS)
Renal pelvis alveolar (ARMS), 47, 764
cancer, 374-378 pediatric kidney neoplasms, 35 6
adenocarcinoma, 376 pleomorphic, 764
hematolymphoid neoplasms, 376 retroperitoneum, 785
histologic grading, 376 sclerosing, 765
970 INDEX
Rhabdomyosarcoma (RMS) (Contd.) Salpingitis, 512-513

skeletal muscle tumor, malignant, 764-765 Salpingitis isthmica nodosum, 513
urinary bladder, mesenchymal neoplasms, Sampling errors, 835
395 Sarcoidosis
Rheumatic heart disease (RHD), 153 cardiac, 15 5
Rheumatoid arthritis, 338, 824 inflammatory conjunctiva diseases, 63
Rhinosinusitis, 35 inflammatory dermatoses, 587
Rhodanine method, 848 lung, 112-113, 147
Riedel thyroiditis, 408, 419 Sarcomas. See also Adenosarcoma;
Rocky mountain spotted fever, 664 Angiosarcoma
Rosenthal fibers (RFs), 632 alveolar soft part, 777
Rosette-forming glioneuronal tumor (RGNT), aortic intimal, 164
649-650 botryoides, 55 6
Round cell liposarcoma, 748, 785 clear cell, 777
clear cell sarcoma of kidney (CCSK),
S-1 00 protein, 629 352-353
Saccular cysts, 23 endometrial stromal, 527
Salivary anlage tumor, 93 epithelioid, 607, 778
Salivary duct carcinoma, 91-92 Ewing, 355-356
Salivary glands extraskeletal myxoid chondrosarcoma, 777
benign neoplasms Kaposi, 271, 769
basal cell adenoma, 86, 96 leiomyosarcoma, 164
canalicular adenoma, 86-87 lung, 141-142
myoepitheliomas, 87 mesenchymal, 366
pleomorphic adenoma (PA), 84,95 myeloid, 708
Warthin's tumor, 84, 86, 95 pleomorphic, 758-759,787
biopsies, 81 rhabdomyosarcoma. See
cysts, 83 Rhabdomyosarcoma (RMS)
inflammation and infection soft tissue (STS), 744-745
autoimmune sialadenitis, 82 scrotal cancer, 489
bacterial sialadenitis, 82 synovial
chronic sialadenitis, 83 pediatric kidney neoplasms, 356
necrotizing sialometaplasia, 83 retroperitoneum, 788
viral sialadenitis, 82 serosal membranes, 186
malignant neoplasms soft tissue tumor, 776-777
acinic cell carcinoma, 89-90, 96 undifferentiated
adenoid cystic carcinoma, 90, 96 endometrial, 528
epithelial-myoepithelial carcinoma, 92 pleomorphic, 758-759
malignant counterparts to benign salivary undifferentiated-poorly differentiated,
gland tumors, 93 778-779
malignant mixed tumor, 91 Scabies, 484
metastasis, 93 Schneiderian papillomas, 38-41
mucoepidermoid carcinoma, 87-88, 96 Schwannoma
pediatric tumors, 93 cutaneous, 604
polymorphous low-grade adenocarcinoma mediastinal neoplasms, 176
(PLGA), 88-89 retroperitoneum, 787
salivary duct carcinoma, 91-92 soft tissue, peripheral nerve tumors,
sec, 92 771-772
neoplasms, 84 vestibular neoplasms of inner ear, 102
non-neoplastic conditions, 95 Scleroderma, 338, 593
chronic sialadenitis, 95 Sclerosing adenosis, 325
lymph nodes, 95 Sclerosing angiomatoid nodular
normal anatomy, 81 transformation (SANT), 738
pathologic features, 95 Sclerosing cholangitis, 2 75
resections, 82 Sclerosing epithelioid fibrosarcoma (SEF),
tumors 755
lung and, 140 Sclerosing in mesenteritis, 247
staging, 95 Sclerosing peritonitis, 180
type neoplasms Sclerosing stromal tumors, 505
esophagus, 198 Sclerosis
sinonasal region, 43 hepatoportal, 258
INDEX 971
hippocampal (mesial temporal), 666 fibrosis, 1 79-180

multiple (MS), 660-661 granulomatous serositis, 177-178
Scrotal cancer, 489 mesothelial hyperplasia, 178
Scrotum meta plasias, 178-179
leiomyomas, 607 splenosis, 180
normal anatomy, 483 tunica vaginalis testis neoplasms, 189
Sebaceous adenoma, 600 uterus, 533-534
Sebaceous hyperplasia, 600 Serositis, 177-178
Seborrheic keratosis, 595 Serous
Sectioning error, 835. See also Cytopathology adenocarcinoma
Segmental cervical glandular, 544
demyelination, 674 ovarian, 500
ureterectomy, 373 uterus epithelial, 526
Seizure disorders, 666 cystadenocarcinoma, 285
Sella and/or suprasellar region, 657-658 cystadenoma
Seminal vesicles, 468-469, 479 ovarian neoplasms, 499
Seminoma,174,458-459 pancreatic,285,296
Senile angioma, 602 degeneration, 711
Sensory neuropathies, 678, 680 tumors, 499-500
Sentinel lymph nodes, 303, 623, 835-836 Serrated polyposis, 233
Septal erythema nodosum, 588 Sertoli cell tumors, 462, 504
Sequence analysis. See DNA sequence analysis Sertoli-Leydig cell tumors (SLCf), 504-505
Sequencing, 903 Sessile serrated
Serosal membranes, 177 adenoma, 233, 241
benign serosal neoplasms polyps, 233, 241
adenomatoid tumor, 181 Sex cord stromal tumors
multicystic peritoneal inclusion cyst, 181 fibromas, 504
biopsy, 177 granulosa cell tumors (GCTs), 462-463
epithelial tumors, 187 adult (AGCTs), 503
excision, 177 juvenile OGCTs), 503-504
malignant pericardia! neoplasms Hilus cell tumors, 505
germ cell tumors, 189 Leydig cell tumors, 462, 505
mesothelial, 188 sclerosing stromal tumors, 505
secondary neoplasms, 189 Sertoli cell tumors, 462, 504
teratomas, 189 Sertoli-Leydig cell tumors (SLCT), 504, 505
malignant peritoneal neoplasms sex cord tumors with annular tubules
adenocarcinomas, 188 (SCTAT), 505
carcinomas, 188 steroid cell tumors, 505
desmoplastic small round cell tumor, stromal luteomas, 505
187-188 thecomas, 504
epithelial, 187 Sex hormone production, 4 33
mesothelial, 186-187 Shave biopsies, 581
pseudomyxoma peritonei, 188 Shingles, 681
secondary neoplasms, 188 Sialadenitis, 82-83, 95
malignant pleural neoplasms Sialoblastoma, 93
desmoplastic small round cell tumor, 186 Sialometaplasia, necrotizing, 83
lymphoid, 185 Sickle cell disease, 574
mesenchymal, 184-185 Signet-ring cell carcinoma, 241, 290, 295
mesothelial, 181-184 Silicosis, 122
secondary neoplasms, 186 Single strand conformational polymorphism
synovial sarcoma, 186 (SSCP) analysis, 898
mesothelial tumors Single umbilical artery (SUA), 568
diffuse malignant mesothelioma (DMM), Sinonasal region, 34
187 endoscopic biopsies, 34
multicystic mesothelioma, 187 frozen sections, 35
well-differentiated papillary functional endoscopic sinus surgery, 35
mesothelioma, 186 inflammation and infection, 35-38
nonneoplastic lesions malignancies staging, 49
acute serositis, 177 neoplastic lesions
cysts, 180 adenocarcinoma, 43-44
eosinophilic peritonitis, 180 alveolar rhabdomyosarcoma, 4 7
972 INDEX
Sinonasal region (Contd. ) mycosis fungoides, 608-609

angiofibroma, 45 primary cutaneous CD30+
cylindrical cell carcinoma, 43 lymphoproliferative disorders, 609, 611
glomangiopericytoma, 47 cutaneous metastasis (from visceral
hemangiopericytoma, 4 7 carcinoma), 611
intestinal-type adenocarcinomas, 43 metastatic breast carcinoma, 612
malignant, 41-47 metastatic renal cell carcinoma, 612
melanoma, 44 Sister Mary Joseph nodule, 612
mesenchymal tumors, 45, 47 cytopathology, 835
nasopharyngeal angiofibroma, 45 dermal deposits
nasopharyngeal carcinoma (NPC), 42 amyloid, 592
neuroendocrine carcinomas, 44 calcinosis cutis, 591
nonintestinal-type adenocarcinomas, 44 digital mucous cyst, 591
olfactory neuroblastoma, 45 gout, 592
salivary gland-type tumors, 43 myxoid, 591
sec, 43 myxoma, 591
Schneiderian papillomas, 38-41 osteoma cutis, 591-592
sinonasal undifferentiated carcinoma dermatoses. See Inflammatory dermatoses
(SNUC), 41 epidermal maturation and keratinization
verruca vulgaris (nasal vestibule), 41 disorders
nonneoplastic lesions acantholytic dyskeratosis, 593
mucous impaction, 38 granular parakeratosis, 594
paranasal sinus mucoceles, 38 porokeratosis, 594
respiratory epithelial adenomatoid fatty and muscular tumors, 607-608
hamartoma, 38 fibrous and fibrohistiocytic tumors, 605-607
normal anatomy fungal infections, 590-591
nasal cavity, 34 gross examination and tissue sampling,
nasopharynx, 34 581-583
paranasal sinuses, 34 infections
resections, 35 fungal, 590-591
Sinonasal undifferentiated carcinoma (SNUC), viral, 589-590
41 keratinous cysts, 592
Sinosoidal onstruction syndrome (SOS), 258 melanocytes, 613
Sinuses melanocytic lesions
congenital anomaly of external ear, 98 benign melanocytic proliferations,
congenital aural, 98 614-619
hyperplasia, 684 blue nevus, 617
paranasal, 34 cellular blue nevus, 617
Sister Mary Joseph nodule, 612 combined melanocytic nevus, 619
Skeletal muscle. See also Soft tissue compound melanocytic nevus, 615
biopsies preparation, 761 congenital and congenital-pattern nevi,
disorders, 761 617-618
Skeletal muscle tumors deep penetrating nevus, 618
retroperitoneum, 785 dermal melanocytosis, 617
rhabdomyoma, 762-765 halo nevus, 618
Skin intradermal melanocytic nevus, 615
benign tumors of epidermis junctional melanocytic nevus, 614-615
clear cell acanthoma, 595 lentigo simplex, 614
seborrheic keratosis, 595 melanoma, 619-621
collagen and elastin disorders melanoma microscopic staging of, 621,
lichen sclerosus et atrophicus, 593 623
morphea/localized scleroderma, 593 nevi on special sites, 619
pseudoxanthoma elasticum, 593 nevus with architectural disorder, 618-619
solar elastosis, 593 pigmented spindle cell nevus of Reed, 617
cutaneous appendages tumors recurrent melanocytic proliferations, 619
apocrine tumors, 600--601 Spitz nevus, 616-617
eccrine tumors, 599-600 melanoma, 619-621
hair follicle tumors, 596-599 melanoma, microscopic staging of, 623
sebaceous adenoma, 600 Breslow thickness, 621, 623
sebaceous hyperplasia, 600 mitotic rate, 623
cutaneous lymphoid infiltrates, 608 satellite and/or in-transit metastasis, 623
INDEX 973
sentinel lymph node examination, 623 joints and synovium, 822

ulceration, 623 mediastinum tumors, 176
mesenchymal tumors, 601--607 neoplasms
neural and neuroendocrine tumors, 603--604 acral myxoinflammatory fibroblastic
nonmelanocytic pigmented lesions, 624 sarcoma, 756
normal microanatomy, 579-580 adipocytic tumors, 745-749
dermis, 579-580 adult fibrosarcoma (AFS), 755
epidermis, 579-580 adult rhabdomyoma, 762
subcutis, 579 alveolar rhabdomyosarcoma (ARMS), 764
premalignant and malignant tumors of angioleiomyoma, 759
epidermis angiolipoma, 746
actinic keratosis, 595 angiomyofibroblastoma, 751
basal cell carcinoma, 596 atypical lipomatous tumor/
invasive sec, 596 well-differentiated LPS, 747-748
squamous cell carcinoma in situ, 596 benign, 741
vascular tumors cellular angiofibroma, 751
angiokeratoma, 602 chondroid lipoma, 746
angiosarcoma, 603 congenital infantile fibrosarcoma (CIF),
arteriovenous hemangioma, 602 754-755
cherry angioma, 602 dedifferentiated LPS, 748
epithelioid hemangioendothelioma, 603 deep benign fibrous histiocytoma, 7 57
glomus tumor and glomangioma, 602 desmoid-type fibromatosis, 752
Kaposi sarcoma, 602-603 diffuse lipomatosis, 746
lobular capillary hemangioma, 601 ectomesenchymoma, 765
lymphangioma, 602 elastofibroma, 750
viral infections, 589-590 ERMS, 762-764
Skin tag, 606 fetal rhabdomyoma, 762
Small cell carcinoma fibroblastidmyofibroblastic tumors,
hypercalcemic, 508 749-756
lung cytopathology, 147 fibrohistiocytic tumors, 757-759
neuroendocrine neoplasms, 545 fibrous hamartoma of infancy (FHI), 750
renal pelvis cancer, 376 Gardner-associated fibroma, 751
thoracopulmonary, 142 genital rhabdomyoma, 762
ureter cancer, 376 giant cell angiofibroma, 751
urinary bladder, 393-394 giant cell malignant fibrous histiocytoma
Small cell glioblastoma multiforme (scGBM), (MFH), 759
641--642 giant cell tumor of soft tissue, 758
Small cell lung carcinomas (SCLC), 137-138 glomus tumor, 760-761
Small cell osteosarcomas, 806 hemangiopericytoma (HPC), 7 53-754
Small duct PSC, 257 hibemoma, 747
Small intestine HN-lipodystrophy, 747
neoplasms, 220, 222, 225 infantile digital fibroma-fibromatosis, 752
nonneoplastic conditions, 217-220 infantile subcutaneous fibromatosis, 752
normal anatomy, 214 inflammatory malignant fibrous
polyps, 220 histiocytoma (MFH), 759
Small lymphocytic lymphoma, 707 inflammatory myofibroblastic tumor
Small vessel vasculitides, 163-164 (IMT), 754
Smoking-related interstitial fibrosis (SRIF), intermediate (locally aggressive), 741
110 intermediate (rarely metastasizing), 741
Smooth muscle neoplasms ischemic fasciitis, 749
angioleiomyoma, 759 juvenile nasopharyngeal fibroma, 753
benign, 759 leiomyoma of deep soft tissue, 759-760
leiomyoma of deep soft tissue, 759-760 LGFMS, 756
malignant, 760 lipoblastoma, 747
retroperitoneum, 785 lipofibromatosis, 752
urinary bladder mesenchymal neoplasms, lipomas, 745-747
394 lipomatosis, 746-747
uterus, 528-530 LMS, 760
Soft fibroma of skin, 606 malignant, 744
Soft tissue. See also Skeletal muscle myofibroma-myofibromatosis, 7 50-751
chondro-osseous tumors, 770 myolipoma, 746
974 I INDEX
Soft tissue (Contd.) deep aggressive angiomyxoma, 773

myopericytoma, 761 desmoplastic small round cell tumor
myositis ossificans and fibro-osseous (DSRCT), 776
pseudotumor of digits, 749-750 digital myxoma, 773
myxofibrosarcoma (MYFS), 756 epithelioid sarcoma, 778
myxoid LPS/round cell LPS, 748 EWS/PNET, 775-776
nevus lipomatosus, 747 extrarenal malignant rhabdoid tumor,
nodular fasciitis, 749 777-778
Nuchal-type fibroma, 751-752 extraskeletal myxoid chondrosarcoma,
pelvic lipomatosis, 746 777
pericytic (perivascular) tumors, 760-761 giant cell fibroblastoma, 775
pleomorphic lipomas, 746 intermediate (rarely metastasizing), 774,
pleomorphic LPS, 748-749 775
pleomorphic malignant fibrous intramuscular myxoma, 773
histiocytoma (MFH), 758-759 juxtaarticular myxoma, 773
pleomorphic rhabdomyosarcoma, 764 malignant, 775-779
plexiform fibrohistiocytic tumor (PFH), myoepithelial neoplasms, 774
758 ossifying fibromyxoid tumor, 774
proliferative fasciitis, 749 perivascular epithelioid cell tumors
proliferative myositis, 749 (PEComas ), 775
rhabdomyoma, 762 pleomorphic hyalinizing angiectatic tumor,
sclerosing epithelioid fibrosarcoma (SEF), 774
755 synovial sarcoma, 776-777
sclerosing rhabdomyosarcoma, 765 undifferentiated-poorly differentiated
skeletal muscle, 762-765 sarcoma, 778-779
smooth muscle tumors, 759-760 vascular tumors
soft tissue sarcomas (STS), 744-745 angiomatosis, 768
solitary fibrous tumor (SFT), 753 angiosarcoma, 769-770
spindle cell lipoma, 746 arteriovenous hemangioma, 767
steroid lipomatosis, 746 atypical vascular lesion of breast, 768
superficial fibromatoses, 752 bacillary angiomatosis, 766
symmetric lipomatosis, 746 benign, 766-768
synovial tendon-based tumors, 757 capillary hemangioma, 766
undifferentiated high-grade pleomorphic cavernous hemangioma, 767
sarcoma, 758-759 cherry angioma, 767
undifferentiated pleomorphic sarcoma congenital hemangioma, 766
with giant cells, 759 epithelioid hemangioendothelioma, 769
vascular tumors, 765 epithelioid hemangioma, 767
nonneoplastic disorders of skeletal muscle glomeruloid hemangioma hemangioma,
mitochondrial myopathies, 761 765
muscular dystrophies, 761 hemangioma, 766
peripheral nerve tumors inflammatory lesions, 765
diffuse neurofibromas, 771 intermediate (locally aggressive), 768
granular cell tumor, 772 intermediate (rarely metastasizing), 769
localized neurofibromas, 771 intramuscular angioma, 767
malignant, 772, 773 Kaposi sarcoma, 769
mucosal neuroma, 771 KHE, 768
neurofibromas, 771 lobular capillary hemangioma (LCH),
neurothekeoma, 772 765-766
perineurioma, 772 lymphangioma, 768
plexiform neurofibromas, 771 malignant, 769-770
Schwannoma, 771-772 papillary endothelial hyperplasia (PEH,
traumatic neuroma, 770 Masson vegetant hemangioma), 765
processing, 739 papillary intralymphatic
reporting of tumors, 779 angioendothelioma, 769
retroperitoneum tumors, 783-788 pseudomyog-enic hemangioendothelioma,
tumors of uncertain differentiation 768
AFH, 774 reactive vascular proliferations, 765
alveolar soft part sarcoma, 777 retiform hemangioendothelioma, 769
benign, 773 spindle cell hemangioma, 767
clear cell sarcoma, 777 synovial hemangioma, 767
INDEX 975
tufted angioma, 766 hemangiomas, 737

vascular transformation of lymph nodes, histiocytic, 737
766 Kaposi sarcoma, 738
venous hemangioma, 767 littoral cell angioma, 737
verrucous hemangioma, 767 littoral cell hemangioendothelioma, 738
Soft tissue sarcomas (STS), 744-745 lymphoid neoplasms, 733-736
Solar elastosis, 593 malignant lesions, 73 8
Solitary circumscribed neuroma, 603 metastatic tumors, 738
Solitary fibrous tumor (SFf), 185, 656-657, myeloid neoplasms, 736-737
753 nonhematopoietic neoplasms and
Solitary plasmacytoma, bone, 811 pseudoneoplasms, 737-73 8
Somatotroph (GH cell) adenoma, 449 peliosis, 737
Spectral karyotyping (SKY), 868-869 pseudoneoplastic lesions, 738
Spermatocytic seminoma, 459 sclerosing angiomatoid nodular
Spindle cell transformation (SANT), 738
carcinoma stromal lesions, 73 7
adult renal, 364 vascular lesions, 737-738
larynx, 2 7, 28 normal gross and microscopic anatomy, 729
metaplastic, 314 red pulp, 729
oral cavity and oropharynx, 12 white pulp, 729
urinary bladder, 395 reactive splenic disorders, 733
hemangioma, 767 Splenectomy, 729
lipoma, 746 Spleniculi, 732
nevus, 616 Splenomegaly, 730, 734
nevus of Reed, pigmented, 617 Splenosis, 180, 732
nodule Sponge kidney, 345
uterus, 531 Spongiosis, eosinophilic, 585
vaginal, 554 Sprue, 218-219
Spiradenoma, 599 Squamous cell, 547, 549
Spirochetosis, 232 Squamous cell carcinoma (SCC)
Spitz nevus, 616--617 anal canal, 245
Spitzoid type type melanoma, 621 basaloid, 13
Spleen cervical, 542, 549
focal reactive disorders, 733 epidermis, 596
general considerations esophagus, 194,200
accessory spleen, 732 in situ, 596
hypersplenism, 730-731 invasive, 542
hyposplenism, 731 larynx, 26-29
splenomegaly, 730 lung, 132-133, 147
splenosis, 732 microinvasive, 542
gross examination and tissue sampling middle ear, 102
biopsy and FNA cytology, 729 oral cavity and oropharynx, 10-11, 13
processing, 729 penile cancer, 487-489
splenectomy, 729 renal, 376
lymphoid neoplasms, 733-736 salivary glands, 92
hairy cell leukemia (HCL), 736 scrotal, 489
hepatosplenic marginal zone T-cell, 734 sinonasal region, 43
Hodgkin, 734 skin, 596
mantle cell, 736 ureter, 376
non-Hodgkin, 734 urethral, 401
primary, 733 urinary bladder, 392-393, 398
secondary, 734 uterus, 527
splenic marginal zone B-cell, 734 vaginal, 556
myeloid neoplasms verrucous, 393
acute leukemia, 736 vulvar, 564
chronic myelogenous leukemia, 736 Squamous metaplasia
mast cell disease, 736-737 endometrium, 520
systemic mastocytosis, 736-737 placental, 568
neoplastic disorders, 733 prostate, 4 71
angiosarcomas, 738 urethral, 400
dendritic cell tumors, 737 urinary bladder, 3 82
976 INDEX
Squamous metaplastic epithelium, 535-536 Subependymal giant cell astrocytoma (SEGA),

Squamous papilloma 643
cervix, 538 Subependymoma, 646
esophagus, 194 Superficial
larynx, 23 fibromatoses, 752
oral cavity and oropharynx, 9 lymphangioma, 602
urinary bladder, 392 spreading melanoma, 620
vaginal, 552 Sweet's syndrome, 587-588
Steatocystoma, 592 Sympathetic paragangliomas, 442
Stenosis, 552 Synaptophysin {SYN), 629
Steroid cell tumors, 505 Synovial fluid examination, 823
Steroid lipomatosis, 746 Synovial sarcoma
Stevens-Johnson syndrome (SJS), 583 lung, 141
Stomach mediastinal soft tissue neoplasms, 176
cytopathology, 212 pediatric kidney neoplasms, 356
endoscopic biopsies, 201 retroperitoneum, 788
gastrectomy, 201-202 serosal membranes, 186
gastric polyps, 205 soft tissue tumor, 776-777
neoplasms Synovial soft tissue, 822
adenocarcinoma, 206, 208, 212 Synovial tumors, 757, 825-827
adenoma, 205 Synovitis, 824-825
carcinoid tumors, 212 Synovium, 822, 824
dysplasia, 205 Syphilis, 485, 559, 577
gastrointestinal stromal tumors (GIST), Syphilitic orchitis, 45 5
208-212 Syringocystadenoma papilliferum, 600-601
malignant lymphoma, 213 Syringoma, 599
marginal zone B-celllymphoma of Systemic l-ITN, 339
mucosa-associated lymphoid tissue Systemic mastocytosis {SM), 719, 736-737
(MALT) type, 211-212
nonneoplastic conditions Takayasu's aortitis, 338
foveolar hyperplasia, 204 Takayasu's arteritis, 163
gastric antral vascular ectasia, 204 Tamoxifen therapy, 519
gastritis, 202-204 T-celllymphomas
gastropathy, 204 enteropathy-type, 225
Menetrier's disease, 204 hepatosplenic marginal zone {HSTL), 735
mucosal calcinosis, 204 incidence and epidemiology, 685
pseudomelanosis, 204 pathophysiology, 685-688
xanthelasma, 205 peripheral, 708
xanthoma, 205 primarily T cell- and NK cell-associated
normal anatomy, 201 markers, 691
Strawberry gallbladder, 275 IDP-43 neuronal cytoplasmic inclusions, 632
Streptavidin-biotin conjugate method, 852 Telangiectatic osteosarcomas, 806
Stricture, 569 Telepathology, 928-929
Stromal hyperplasia, 471-472, 495 Temporal arteritis, 162
Stromal luteomas, 505 Tendon
Stromal tumors. See also Gastrointestinal sheath tumors, 825-826
stromal tumors (GIST) synovial tendon-based tumors, 757
cervical, 53 9 Teratoid Wilms tumor, 350
endometrial, 52 7 Teratomas
metanephric (MSTs), 351-352 cystic, 173
mixed epithelial and adult renal, 368 germ cell, 173, 460-461
ovarian, 499-503 immature, 506
sclerosing ovarian, 505 mature, 173,505-506
sex cord. See Sex cord stromal tumors monodermal, 506
Strongyloidiasis, 220 ovarian, 505-506
Subacute sclerosing panencephalitis (SSPE), retroperitoneal, 78 8-78 9
662 serosal membranes, 189
Subchorial thrombosis, 571 testis, 460-461
Subcutaneous fat necrosis of newborn, Tertiary adrenal cortical insufficiency, 431
589 Testis
Subcutis, 579 atrophy, 455
INDEX 977
congenital abnormalities Thyroid, 404

adrenal cortical rests, 453 developmental diseases
anorchism, 453 lingual thyroid, 406
cryptorchidism, 453 pyramidal lobe, 405
polyorchidism, 4 53 thyroglossal duct cyst, 405
testicular-splenic fusion, 453 gross examination and tissue sampling, 404
germ cell tumors, 455-462 biopsy, 404
burnt out (regressed) germ cell tumors, resection, 404-405
461 inflammation
embryonal carcinoma, 459-460 acute thyroiditis, 406
histologic typing and diagnosis, 457 chronic thyroiditis, 406
histopathologic diagnosis, 457 de Quervain thyroiditis, 406
intratubular germ cell neoplasia of focal lymphocytic thyroiditis, 406
unclassified type (IGCNU), 458 Graves disease, 407-408
mixed malignant germ cell tumor, 461 Hashimoto thyroiditis, 406-407
molecular genetics, 462 Riedel thyroiditis, 408
new immunophenotypic markers, subacute thyroiditis, 406
461-462 neoplasms
seminoma, 458-459 adenoma, 409-410
spermatocytic seminoma, 459 folliculru; 409-411
teratomas, 460-461 medullary, 416-417
trophoblastic tumors, 460 papillary, 412-413,415
yolk sac tumor (endodermal sinus tumor), pathologic reporting, 417
460 poorly differentiated, 411-412
gross examination and tissue sampling undifferentiated, 415-416
bilateral orchiectomy specimens, 453 nodular hyperplasia, 408, 409
fine needle aspiration biopsy, 452 normal anatomy, 404
radical orchiectomy, 452 Thyroid cytopathology
retroperitoneal lymph node dissection, 453 benign
testicular biopsies, 452 acute thyroiditis, 418
unilateral simple orchiectomy, 452 chronic thyroiditis, 418
infertility and, 453-454 de Quervain thyroiditis, 418
inflammation and infection follicular nodules, 418
epididymitis, 454 Hashimoto thyroiditis, 418
mumps orchitis, 454 Riedel thyroiditis, 419
syphilitic orchitis, 455 subacute granulomatous thyroiditis, 418
tuberculous orchitis, 454 cyst fluid only, 418
normal anatomy, 451 follicular neoplasm, 419
pathologic staging, 464, 465 malignant, 420
tumors anaplastic carcinoma, 420
collecting ducts and rete, 463 hyalinizing trabecular adenoma, 420
germ cell, 455-462,465-466 lymphoma, 420
hematolymphoid, 463 medullary carcinoma, 420
paratesticular. See Paratesticular tumors metastatic malignancy, 420
sex cord/gonadal stromal, 462-463 papillary thyroid carcinoma (PTC), 420
tunica vaginalis, 189 poorly differentiated (insular) carcinoma,
vascular disorders 420
systemic vasculitis, 455 nondiagnostic/unsatisfactory, 418
torsion and infarction, 455 suspicious for follicular neoplasm, Hiirthle
varicocele, 455 cells type, 419
Thecomas, 504 suspicious for malignancy (SFM), 419-420
Thin basement membrane disease, 336-337 thyroiditis, 418
Thioflavin T, 844 Thyroid-like follicular carcinoma, 365
Thrombocythemia, essential, 718 Tissue microarrays, 877-878, 923
Thrombocytosis, 711 Tropical spastic paraparesis (TSP), 662
Thrombohematomas, 570-571 Torsion,ovarian,495
Thrombosis, 571 Torus mandibularis, 6
Thrombotic endocarditis, 151 Torus palatinus, 6
Thrombotic vasculopathy, 571. See also Total cystectomy, 380
Placenta Total parenteral nutrition (TPN), 255
Thyroglossal duct cysts, 405 Toxic epidermal necrolysis (TEN), 583
978 INDEX
Toxin-induced liver injury (DILl), 256 radical nephroureterectomy with bladder

Toxoplasmosis, 577, 665, 901 cuff, 374
Trabecular juvenile ossifying fibroma, 60 segmental ureterectomy, 3 73
Tracheobronchial amyloidosis, 115 ureteroscopic biopsies, 3 73
Transitional cell carcinomas (TCC), 502 normal anatomy, 3 73
Transitional cell metaplasia, 4 71 urothelial dysplasia, 374
Translocation renal cell carcinoma (RCCs), Ureterectomy, segmental, 373
354 Ureteropelvic junction (UPJ) abnormalities,
Translocations, 360, 364 374
Transplantation Urethra
cardiac, 159, 161 congenital anomalies
liver, 260-262 diverticula, 399
for parathyroid hyperplasia, 424 fibroepithelial polyp, 400
pulmonary. See Pulmonary transplantation urethral valves, 399
Traumatic neuropathy, 682 gross examination and tissue sampling, 399
Traumatically implanted fungus, 591 histologic grading of carcinoma, 403
Treponema pallidum, 485 hyperplasia, 400
Trichilemmal type keratinous cysts, 592 inflammation and infection, 400
Trichoadenoma (of Nikolowski), 598 metaplasia, 400
Trichodiscoma, 598-599 neoplasms
Trichoepithelioma, 598 benign, 400
Trichofolliculoma, 596, 598 malignant, 400-401, 403
Trichomonas, 397 normal anatomy, 399
Trichomonas vaginalis, 548, 551 pathologic staging of carcinoma, 403
Trichomoniasis, 551 prostatic, 4 79
Trichrome, 845 reporting of carcinoma, 403
Tricuspid valves disorders, 153 Urethritis, 400
Trophoblastic tumors, 460, 531-533 Urinary bladder
Tropical sprue, 219 adenoma, 383
True cementoma, 57 amyloidosis, 384
Truncation artifact, 878 biopsy specimens, 379
Tuberculosis (TB), 119, 371 congenital malformations
Tubular carcinoma, 244, 313, 364 exstrophy, 380
Tubulocystic carcinoma, 364 urachal abnormalities, 380
Tubulointerstitial diseases, 337-338 cystoprostatectomy, 380
Tumor-induced osteomalacia (110), 819 cytopathology, 396-398
Tunica vaginalis testis neoplasms, 189 diverticula, 384
Twin-twin transfusion syndrome (TITS), 575 ectopic prostatic tissue, 384
Tyramine amplification, 853 endocervicosis, 384
endometriosis, 383
Ulcerative endosalpingiosis, 384
colitis, 228-229 fibroepithelial polyp, 385
keratitis, 69 glandular neoplasms, 393
Ultrastructure, 673 gross examination and tissue sampling, 379
Umbilical cord disorders. See under Placenta inflammatory conditions, 380, 382
Unclassified renal cell carcinoma (RCC), 364 intestinal metaplasia, 383
Urachal abnormalities, 380 mesenchymal neoplasms, 394-395
Urates, 848 neoplastic urotheliallesions, 385-386,
Uremia, 677 388-392
Uremic osteodystrophy, mixed, 820 nephrogenic metaplasia, 383
Ureter neuroendocrine neoplasms, 393-394
benign normal microscopic anatomy, 379
conditions, 374 partial cystectomy specimens, 379-380
epithelial neoplasms, 374 pathologic staging, 395
nonepithelial neoplasms, 374 pseudocarcinomatous epithelial hyperplasia,
cancer, 374-377 383
gross examination and tissue sampling, 373 reactive urothelial atypia, 383
cystoprostatectomy, 373 reporting, carcinoma, 395
needle core biopsies, 373 squamous metaplasia, 382
pyeloplasty specimens, 373 squamous neoplasms, 392-393
radical cystectomy, 373 total cystectomy specimens, 380
INDEX 979
transurethral resection, 3 79 HSV infection, 551

urothelial hyperplasia, 383 infectious diseases, 551
Urine, 396 inflammatory diseases, 551-552
Urothelial carcinoma, 375 mesonephric cysts, 552
gross diagnosis, 3 75 Miillerian cysts, 5 52
immunohistochemical studies, 376 stenosis, 552
in situ, 385 trichomoniasis, 551
malignant neoplasms, 372,401 vulvovaginal candidiasis, 551
microscopic aspects, 375-376 neoplasms
molecular studies, 376 benign,552,554-555
urinary bladder cytology, 397 malignant, 555-557
Urothelial dysplasia, 374, 385 metastasis, 5 57
Urothelial hyperplasia, 383, 400 normal anatomy, 5 51
Urothelial metaplasia, 4 71 Vaginal intraepithelial neoplasia (VAIN),
Urothelial neoplasms, 386-392 555-556
Urothelial papilloma, 386 Vaginitis, 5 51
Urticaria, 588 Vaginosis, 551
Usual interstitial pneumonitis (UIP), 108-109 Validation, 909
Usual nodular epithelial hyperplasia, 471 Valves. See also Heart
Uterus. See also Cervix cardiac, 151
adenofibroma, 531 disorders, 153
adenomyosis, 531 Varicella zoster virus (VZV), 118, 590, 664
adenosarcoma, 531 Varicocele, 455
endometrial stromal tumors, 527 Vascular ectasia, 204
endometrium, 518-522 Vasculitides, 161-163
epithelial malignancies, 522-527 kidney, 338
gestationa~ 531-533 large vessel, 162-163
gross examination and tissue sampling, 517 medium vessel, 163
lymphoid neoplasms, 534 small vessel, 163
metastatic tumors, 534 Vasculitis, 115, 587-588
normal anatomy, 517 cervical, 53 8
postoperative spindle cell nodule, 531 immunofluorescence, direct, 856
radical hysterectomy, 537 systemic, 455
serosal tumors, 533-534 Vasculopathies
smooth muscle neoplasms cardiac allograft, 161
benign metastasizing leiomyoma, 529 fetal thrombotic, 571
cellular leiomyomas, 529 inflammatory dermatoses, 587-588
epithelioid leiomyomas, 529 Verhoeff Van Gieson (VVG) stain, 845-846
intravascular leiomyomatosis, 529 Verruca plana, 589
leiomyoma, 528-529 Verruca vulgaris, 9-10, 41, 589
leiomyosarcoma, 529-530 Verruciform xanthoma, 9-10
lipoleiomyoma, 529 Vessels. See also Heart
myxoid, 529 amyloid angiopathy, 164
symplastic leiomyoma, 529 neoplasms, 164
tumors of uncertain malignant potential normal anatomy, 161
(STUMP), 530 temporal artery biopsies, 161
trophoblastic tumors, 531-533 vasculitides, 161-163
undifferentiated endometrial sarcoma, 528 Vestibular schwannoma, 102
Villitis of unknown etiology (VUE), 577-578
Vacuolar change, 581 Viral hepatitis, 251-252
Vagina Viral infections
benign conditions CNS, 661-662
actinomyces-like organisms, 551 cutaneous infections, 589-590
adenosis, 552 demyelinating, 661-662
atrophic vaginitis, 551 disease testing, 900-901
bacterial vaginosis, 551 lung, 117-118, 146
Bartholin gland cysts, 552 oral cavity and oropharynx, 4-5
Crohn•s disease, 552 Virtual microscopy, 926-929
cysts, 552 Virtual slide telepathology, 929
endometriosis, 5 52 Von Hippel-Lindau (VHL) disease, 359
epithelial inclusion cysts, 5 52 Von Kossa method, 848
980 INDEX
Von Meyenburg complex, 268 West Nile virus encephalitis, 663

Vulva Whipple
cystic lesions, 5 60 disease, 219
gross examination and tissue sampling, 558 procedure, 283
infection, 559 White matter damage, 662
inflammation, 558-559 White pulp, 729
neoplasms Wilms tumor, 348
benign, 560-563 Wilson's disease, 255
malignant, 563-564
noninfectious squamous lesions, 560 Xanthelasma, 205
normal anatomy, 558 Xanthoastrocytoma, 643
Vulvar intraepithelial neoplasia (VIN), 563 Xanthogranulomatous pyelonephritis {XGP),
Vulvovaginal candidiasis, 551 370
Xanthoma, 205
Walthard nests, 179 Xp 11.2 translocations, 364
Warts, 542, 590 Yolk sac tumor, 460, 507
Warthin's tumor, 84, 86, 95
Warthin-Starry method, 847 Zellballen, 25
Wegener granulomatosis, 36, 116, 338 Zoon balanitis, 484

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The Lauren V.

Ackerman Laboratory of Surgical Pathology

© 2012 by Department o£ Pathology and Immunology, Washington University School of Medicine

Library o£ Congress Cataloging-in-Publication Data

Hussam Al-Kateb, MSc, PhD Yumei Chen, MD, PhD

Louis P. Dehner, MD Johann D. Hertel, MD, MA

Jeffery M. Klco, MD, PhD Jochen K. Lennerz, MD, PhD

Michael J. Klein, MD James S. Lewis, Jr., MD

Julie Elizabeth Kunkel, MD Mitra Mehrad, MD

Kathryn M. Law, MD ILKe Nalbantoglu, MD

TuDong Nguyen, MD, PhD Jon H. Ritter, MD

Samuel J. Pruden, II, MD, FACS Nathan C. Walk, MD

Hanlin L. Wang, MD, PhD Frances V. White, MD

Joshua I. Warrick, MD Heather N. Wright, MD

Peter A. Humphrey, MD, PhD

I HEAD AND NECK 1

Tumors and Cysts of the Jaws

Ell The Eye 63

Ell Cardiovascular System 149

lli1 Mediastinum 165

Serosal Membranes 177

Ill Gl TRACT 190

IE1 The Stomach 201

Cytopathology of the Stomach 212

The Intestines, Appendix, and Anus 214

1m The Liver 248

Cytopathology of the Liver 271

IT:I The Gallbladder and Extrahepatic

Cytopathology of the Gallbladder and Extrahepatic Biliary Tree 279

IR The Pancreas 282

Cytopathology of the Pancreas 294

Cytopathology of the Breast 326

V URl NARY TRACT 329

~~~ Surgical Diseases of the Kidney 344

Maldevelopment and Nonneoplastic

OJ Renal Pelvis and Ureter 373

r;J;J The Urinary Bladder 379

&1 Urethra 399

VI ENDOCRINE SYSTEM 404

rwt Parathyroid Glands 422

r;r:t The Adrenal Gland and Paraganglia 428

DJ Pituitary Gland 446

VII REPRODUCTIVE TRACT 451

Ell The Ovary 493

EJ;J Fallopian Tube 511

Ell Uterine Cervix 535

ET:I Vu Iva 558

Em Nonmelanocytic Tumors of the Skin 595

twi1 Skin: Melanocytic Lesions 613

R;J Nerve Biopsies 673

X HEMATOPOIETIC SYSTEM 683

Ill Bone Marrow Pathology 709

XI SOFT TISSUE AND BONE 739

1m Bone Neoplasms and Other Nonmetabolic

1m Metabolic Diseases of Bone 816

er.1 Joints and Synovium 822

XII CYTOPATHOLOGY 828

ft1 Electron Microscopy 838

ell Histology and Histochemical Stains 842

6Et Immunohistochemistry 851