Aetiology & Pathogenesis of Periodontal

Disease

Dr Luan Ngo BDSc Hons (Melb)

 Aetiology & Pathogenesis
Periodontal disease
- bacterial mediated inflammatory disease
- destruction of alveolar bone, tooth mobility
and eventually tooth loss
- complex interplay between bacterial
virulence factors and host response, often
modified by behavioural factors
 Clinical and Histopathologic Features of
Periodontal Disease
• The progression of periodontal disease can be roughly
divided into the following stages:
– Pristine Gingiva = no plaque, clinically healthy
gingiva
– Initial Lesion = plaque, clinically healthy gingiva
– Early Lesion = Gingivitis
– Established Lesion = Gingivitis
– Advanced Lesion = Periodontitis

• See Lindhe textbook for pictures used in this lecture

Pristine Gingiva (Healthy)
Clinical:
• No plaque seen
• No inflammation seen

Histology:
• Continuous sparse neutrophil migration into
coronal part of junctional epithelium and crevice.
Some or no GCF flow
• Shallow gingival sulcus
• JE firmly attached to root, sulcular epithelium and
CT
Initial lesion
Clinical: 24 hours to 4 days (no OH)
• Difficult to differentiate from pristine
• 24hrs-4days of plaque accumulation

Histolopathology:
• Capillaries dilate, increasing blood flow
• Perivascular collagen loss
• GCF flow starts to increase
• Increased neutrophil migration from CT into
gingival crevice
• Lymphocytes begin to accumulate
Early lesion (Early Gingivitis)
Clinical: 4-7 days (no OH)
• Erythema (red) + Oedema (swollen)
(deeper crevice favours more anaerobic organisms)

Histopathology:
• Proliferation of capillaries
• 70% perivascular collagen lost
• Gingival fibre groups starting to break down
• Fibroblasts = less collagen formation
• JE & sulcus full of neutrophils
• More immune cells moving into CT beneath JE  mainly
lymphocytes (75% of these are T cells)
Established gingivitis
Clinical: 14-21 days (no OH)
• Reddish ± bluish hue
• Moderate/Severely inflamed
gingiva
• BOP
• Deepening of gingival sulcus
• no loss of attachment
Established gingivitis II
Histopathology:
• Blood vessels engorged and congested
• Blood flow sluggish
• Intense chronic inflammatory reaction
• Increasing proportion of plasma cells
– Page and Schroeder described this lesion as plasma cell
dominated
• JE develops rete pegs or ridges
• Epithelium not attached to tooth surface
• In CT, Collagen breaks down to make room for influx
of inflammatory cells (inflammatory infitrate)
 Progression from Gingivitis
• Established lesion can go either of two ways:
– Remain stable and does not progress for
months or years
– Become more active and convert to
progressive destructive lesion
(Advanced lesion/Periodontitis)
– To date, we cannot predict which patients
or sites are going to progress from the
established lesion to the advanced lesion
 remember that NOT ALL patients with
gingivitis progress to periodontitis
 Advanced Lesion (Periodontitis)
• Lesion extends into alveolar
bone
• Damage now irreversible
• CT attachment lost
• Apical extension of pocket,
which favours more
anaerobic organisms
• Inflammatory infiltrate
spreads laterally and further
apically
• Associated with pocketing,
bleeding on probing and
sometimes suppuration

JE migrating
apically, PDL Bone loss
attachment lost
How do Pockets Form?
Changes to the Gingival Sulcus:
• Inflammation & degeneration of CT wall
• Collagen fibres just apical to JE destroyed and
replaced by inflammatory cells and oedema
• Apical cells of JE proliferate along the root
• Coronal portion detaches as apical portion migrates
• Tissue detaches from tooth surface as more and
more neutrophils invade the gingival sulcus
• With continued inflammation, JE continues to
migrate along the root and separate from it
How do Pockets Form?
Changes to the Lateral Soft Tissue Wall
• dilated, engorged bv’s
• CT degeneration
• Epithelium here shows both proliferative
and degenerative changes
• Buds or cords of epithelium project into
inflamed CT
• Progressive degeneration and necrosis
lead to ulceration of the wall
• Can have superimposed acute
inflammation
How do Pockets Form?
Changes to the Lateral Soft tissue wall II
• But remember – the host: parasite balance
is constantly changing
• Therefore the pocket wall is constantly
changing
• Histologically, there are areas of:
– quiescence
– bacterial accumulation & emergence of leukocytes
– leukocyte-bacteria interaction
– intense epithelial desquamation
– Ulceration & haemorrhage
How Attachment Loss Occur?
• Periodontal ligament broken down by host
proteases (matrix metalloproteinases)
derived from fibroblasts, keratinocytes,
neutrophils (increase in MMPs or
decrease in TIMPs)
• Alveolar bone resorbed by osteoclasts
• Both processes are host mediated (via
inflammatory cytokines e.g. PGE2, IL-1, IL-
6) and occur at a small distance from the
actual site of infection
 Host Defences available to fight
Plaque
• Innate (Natural) immunity

• Acquired (Adaptive) immunity

There is a great deal of interaction and

communication between all the components.
And many of which are common to more than one
pathway.
Innate Immunity
1. Barriers:
– Skin, Mucous Membranes, Connective Tissue
2. Flushing Action:
– Peristalsis, Cilia, Flow of Body Fluids, Saliva,
Cough/Sneeze Reflex
3. (Bio)chemical:
– Enzymes, Gastric Acid, Bile, Sweat, Fatty Acids
4. Biological:
– Inflammation, Complement, Phagocytosis,
Natural Killer Cells, Acute-phase response,
Cytokines, Competition with Natural Flora
Innate Immunity - Oral Cavity
• Oral mucosa
• JE = permeable barrier
• Oral lymphoid tissues:
– lymph nodes, tonsils, salivary glands (sIgA) and
gingival tissues
• Saliva (supragingival action)
– Salivary peroxidase
– Lysozyme - degrades cell walls
– Lactoferrin - binds iron, important for bacterial
growth
• Gingival Crevicular Fluid (subgingival action)
 Acute Inflammation
• Quick response to tissue injury
• Classic signs: redness, swelling, heat, loss of
function
• Vascular changes:
–  blood flow
–  permeability   exudate
• White cell events (PMN’s/Macrophages)
– margination & pavementing
– emigration
– chemotaxis
– phagocytosis
Chronic inflammation
• Persistent injurious stimuli
• causing mononuclear cell infiltration and
fibroblast proliferation
• Bursts of destruction and repair
• Involves macrophages, T/B lymphocytes and
plasma cells
• Can result in a specific immune response
The Immune Response
Innate Immunity Acquired Immunity
–rapid –develops & adapts to
–relatively non-specific pathogens
–soluble & cellular effectors –memory
–cytokines important –highly specific
regulators –soluble & cellular effectors
–bystander damage likely –cytokines important
–less bystander damage??
Basic Steps in Local Inflammation in the
Gingiva
1. Insult to local tissue
2. Signalling the endothelium (via cytokines e.g. IL-
8)
3. Expression of adhesion molecules
4. Adhesion to vessel wall
5. Passage through endothelium
6. Chemotaxis to point of insult:
1. Initially neutrophils
2. Later macrophages
The Immune Response
Toolkit
• For the host: • Against the
– Neutrophils host:
– Macrophages
– T/B cells
Plaque bacteria
– Plasma Inappropriate Host
cells/Antibodies Responses
– Pro-inflammatory
molecules:
• Cytokines
• MMP’s
• Prostaglandins
– Complement
Neutrophils (PMN’s, polymorphs)

• First leucocyte recruited in inflam

response
• Associated with acute inflammation –
first response
• Short life span, most common WBC in
blood
• Has receptors for complement and
antibodies
• Kills bacteria by:
– Phagocytosis – ingests bacterium
– Oxidative and non-oxidative enzymes
– Phagosome is fused with oxidative
enzymes inside the neutrophil
Disease
Neutrophil defects
Defect Oral manifestations

Chediak-Higashi syndrome Migration, degranulation and Aggressive periodontitis, oral ulcerations

phagocytosis defects (defective
intracellular organelles)
Chronic granulomatous Impaired intracellular killing (defective Periodontitis, recurrent infections
NADPH oxidase)
Cyclic neutropenia Periodic absolute neutrophil count <1500 Gingivitis / periodontitis, periodic oral
ulcers
Chronic neutropenia Persisitent absolute neutrophil count Recurrent oral ulcers, exhuberant
<1500 gingivitis
Leukocyte adhesion deficiency Impaired rolling / adherence to Severe periodontitis
endothelium, phagocytosis
Papillon-LeFevre syndrome Chemotaxis, phagocytosis and Severe aggressive periodontitis
intracellular killing
(mutation in Cathepsin C gene)
Kostmann syndrome Absolute neutrophil count <1500 (treated with GMCSF) Severe
Deficient antimicrobial peptides periodontitis

Lazy leukocyte syndrome Neutropenia, depressed chemotaxis Periodontitis

Down’s syndrome Variable – chemotaxis, phagocytosis and More severe gingivitis and periodontitis
killing
 Macrophages
• Monocytes become macrophages once
they leave the blood stream & differentiate
• live longer than PMN’s – come out later in the
inflammatory response
• Communicate with lymphocytes and other
surrounding cells  often by cytokines
• Present antigens to T-cells
• Associated with Chronic Inflammation
• Receptors for complement, antibodies and MHC
class II
Lymphocytes: T cells
• Required for a specific immune
response
• Interact with antigen-presenting cells
to become activated
• Recognise antigen in conjunction
with MHC molecules
• Produce pro-inflammatory molecules
upon activation
• Responsible for Cell-mediated
immunity
• Various subsets
T-cell subsets

• T-helper cells: secrete cytokines that

promote B cells to produce antibody. Express
CD4+ receptors.

• T-suppressor cells: suppress the immune

response (T-helper cells). Allow tolerance of
“self”. Express CD4+ receptors.

• Cytotoxic T-cells: kill other cells esp. those

infected by viruses. Express CD8+ receptors.
 Helper T-cell subsets
– Th1 – these are important for cell mediated
immunity – controlling intracellular pathogens such
as viruses, Listeria, P. gingivalis?
• IL-12
• Secretes TNF-β and IFN-β

– Th2 – these are essential for antibody-mediated

immunity, providing help for B cells
• IL-4, IL-5, IL-10 and IL-13

– Th17 – these may be important in protecting

surfaces (e.g. skin, lining of intestine) from
extracellular bacteria
• Secrete IL-17 and IL-22
 Th1 / Th2 Paradigm
• In the past
– Th1  inflammatory diseases
 early / stable periodontal lesions
– Th2  allergy reactions
 late lesion / disease progression

Th2 responses can provide the cytokines for

polyclonal B-cell activation (elevated levels
of low-affinity antibody)
 Th17 cells
• IL-17 regulates MMPs and inflammatory
cytokines in gingival fibroblasts
• Th17 cells can inhibit Th1 but favour
Th2 development (Harris et al. 2002)
 Th cell modulation
• Bacteria may modulate the Th response
– Via antigen presenting cells (APCs)

– APCs can recognise bacterial patterns through

toll-like receptors (TLRs)
• TLR4 activation favours Th1
• TLR2 activation favours Th2

• P. gingivalis LPS triggers TLR2 (Pulendran et al. 2001)

B cells
• Targets extracellular antigens such as
bacteria

• Once a particular antigen is recognised, B

cells clonally expand to become antibody-
secreting plasma cells

• Some antigen-activated cells become

memory cells (stored until the same antigen
is encountered again)

• Primary antibody made against a new

antigen = IgM

• Secondary responses to that antigen

(based on memory T cells) = IgG & IgA
Antibodies

• Five different classes: IgA, IgD, IgE, IgG

& IgM
• Functions:
1. deactivate some antigens
2. prevent binding eg bacteria to host (sIgA)
3. clump cells together to make phagocytosis easier
4. Opsonisation - tag bacteria for later ingestion by
phagocytes
5. activate various cells types
Cytokines & Prostaglandins
Cytokines:
• Chemical messengers
• released from cells during inflammation & immune
responses
• Possess a range of biological activities
– Eg bone formation/ resorption, T&B cell proliferation, fibroblast
proliferation
– Production of cytokines can be induced by bacterial components
• Examples: Interleukins (IL-1, IL-2, IL-8), IFN-, TNF…etc
Prostaglandins:
• Produced by macrophages and fibroblasts, associated
with inflammation
• Up-regulated by IL-1, TNF- & LPS
• Induce MMP and bone resorption
Are what NSAIDs reduce to control pain & inflammation e.g.
Nurofen
Matrix Metalloproteinases (MMP’s)
• proteolytic enzymes used to remodel
or degrade connective tissue
• Important for normal tissue turnover
and repair
• Released from fibroblasts &
macrophages E.g. Collagenases
Matrix Metalloproteinases (MMP’s)

• Levels of MMPs decrease with

clinical improvements following SRP
and maintenance (Kinane et al. 2003)

• MMPs can be inhibited by low-dose

doxycycline (Ryan and Golub 2000)
 Complement
• Important for both innate and adaptive immunity
• Series of plasma proteins
• can lyse Gram -ve bacteria, enhance phagocytosis
and contribute to the inflammatory reaction
• Classic & Alternative pathways
• Classic  forms the Membrane attack complex
(MAC)
– punches a hole through G- bacterial cell membrane
• Alternative  aggregation of Igs, endotoxin, some
cell walls & viruses directly activate of C3  then
triggers the rest of the classic pathway
Periodontal Microbiology
Periodontal Microbiology
 What Is a Biofilm?
• Biofilms are composed of microbial communities that
are attached to an environmental surface.
• These organisms usually encase themselves in an
extracellular polysaccharide or slime matrix.
• Bioflims may be found on any environmental surface
where sufficient moisture and nutrients are present.
• Their development is most rapid in flowing systems
where sufficient nutrients are present.

From www.biofilmsonline.com
Background
• Over 500 different species have been
found in the oral cavity
• Bacteria are found in the mouth from the
time of birth
• Any individual can harbour 150 or more
species
• Positive relationship between amount of
bacterial plaque and severity of gingivitis
• Periodontitis not as clear relationship
 How Do Biofilms Form?

Initial Irreversibl Maturation Complex Dispersion

attachment e community
attachment

From: Looking for Chinks in the Armor of Bacterial Biofilms Monroe D PLoS Biology Vol. 5, No. 11, e307, 2007
 Features Of Biofilms

• Transfer of genetic material

• Different genes switched on c.f. planktonic bacteria
• Complex microbial interactions
• Quorum sensing
• Stable population
• Glycocalcyx
• Detachment of cells
• Migration
Antibiotic Resistance
 Perio Microbiology Basics
• Formation of supra- and subgingival plaque,
calculus  Biofilms
• Main bacteria associated with periodontitis:
– Aggregatibacter actinomycetemcomitans (Aa)
– Porphyromonas gingivalis (Pg)
– Tannerella forsythia
– Treponema denticola (Td) & spirochaetes
Actinobacillus
actinomycetemcomitans
• Facultative Gram -ve small non-motile
short/straight/curved rod
• 5 serotypes (B + Y4 + Jp2)
• Increased prevalence in LAgP, decreased in health
• Causes an elevated serum antibody response
• Elimination/suppression associated with successful
therapy
• Leukotoxin, LPS, enzymes, invasive
• Causes disease in animals
 Porphyromonas gingivalis
• Gram -ve anaerobic non-motile short rod
• Forms black pigmented colonies on blood agar
plates
• Elevated in CP, active lesions and recurrent
lesions
• Lower in health and gingivitis
• Elimination associated with successful therapy
• Elevated serum antibody response
• Large array of enzymes esp. proteases, LPS,
vesicles, degrade Ig’s and invasive
• Causes disease in animals
Tannerella forsythia
• Difficult to grow
• Frequent finding in sites breaking down
• Can be most common species recovered from perio
pockets
• Infrequently detected in health
• Produces AB response
• Virulence factors?? Invasive
• Now called Tannerella forsythensus
Spirochaetes - Treponema
denticola
• Seen by microscope but difficult to culture
• Found in high no. in disease though differentiating
species is difficult
• At least 15 species….Td seems to be one of the main
ones
• Decreased no. in health and successful Rx
• Produces AB response
• Tissue invasive
• Much more work needed
Plaque Hypotheses
Non-specific plaque hypothesis (1950-
1960’s)
• Bulk of bacteria produce enough irritants to
result in inflammation causing periodontal
destruction.
Specific Plaque Hypothesis (1970’s – now)
• differences in composition of bacterial plaque
(subject to subject/site to site).
• elimination of certain bacteria following
treatment coincides with return to health.
• Pathogens cause periodontitis.
Bacteria & Tissue Destruction
DIRECT DAMAGE:
– endotoxins
– exotoxins
– metabolic waste products
– enzymes
– tissue invasion

INDIRECT DAMAGE:
Pathogenic plaque bacteria can switch on the
host’s own immune system to cause tissue
damage
Bacteria and CP
• Numbers game – bacteria present overwhelm
host response
• Associated with complexes of bacteria – “red
complex”
• Once the biofilm is removed, the host stops
being overwhelmed and can activate
appropriate measures to heal
– Periodontal healing after treatment is associated
with reduction/ suppression of red complex
bacteria
Bacteria and AgP
• The underlying host response can be
defective in some way
• And/ or the bacteria are invasive and
actively cause tissue destruction
• Aa most commonly related to LAgP
• Need aggressive treatment and
frequent review.
• Often require antibiotics ± surgery
Necrotising Periodontitis
• Associated specifically with Prevotella
intermedia and spirochaetes
• Also associated with acute stress
• The body sets up conditions for bacteria
(already present) to invade the periodontal
tissues
• Associations:
– Smoking
– Poor nutrition
– Extreme stress
– Lack of sleep
– Poor oral hygiene
Necrotising Perio II
• Signs & Symptoms:
– Red, bleeding, grey
pseudomembrane, slough
– Acute pain, halitosis, rapid onset

• Treatment:
– Debridement under local anaesthesia
– Chlorhexidine rinses
– Metronidazole if systemic signs and
symptoms
Polymorphisms……
Genetic Susceptibility
 Polymorphisms
• Cytokines
– Signalling proteins
– Involved in cellular communication
– Critical for both innate and adaptive
immune response

– Often associated with antagonists

• E.g. IL-1 with IL-1RA
 What is a Single Nucleotide
Polymorphism?

From Chapter 17, Lindhe 2003

 Periodontally relevant SNP’s

But also IL-4, IL-6, IL-10, FcR, VDR, TLR, TCR,

RSVP…the list is endless
From Chapter 17, Lindhe 2003
Frequency of IL1α -889 genotype
 But what does it all mean?

From Chapter 17, Lindhe 2003

Bringing it all
together …
Inadequate Biofilm
OH Biofilm
maturation

Increase GCF
Inflammation
Periodontal Increase proteins
Formation of a
pathogens Anaerobic
environment periodontal pocket
 Inadequate Biofilm
Biofilm
OH maturation

Inflammation
HOST Deepened pocket
Immunoglobulins
Neutrophils
Inflammatory Antimicrobial peptides

Polymorphisms
mediators
Periodontal
Autoimmune pathogens
complexes Virulence factors
Stress
Diabetes
Smoking
Repair

Resorption of Bone
Activation of osteoclasts
Release of host collagenases Destruction of PDL
Thank you to Drs Melinda Newnham and Ivan
Darby for some of these slides
Questions???