IMMUNOLOGY OF PERIODONTAL DISEASES

OUTLINE:
 Introduction
 Lipopolysaccharide
 Natural history of periodontal disease lesions and their histologic findings
 Role of complement system in immunology of periodontal disease:
 Role of various inflammatory mediators in periodontal diseases:

INTRODUCTION
Periodontal diseases are initiated by oral bacteria disturbing epithelial cells, which trigger
innate, inflammatory, and adaptive immune responses. These processes result in the
destruction of the tissues surrounding and supporting the teeth and eventually result in tissue,
bone, and finally tooth loss.

 Periodontal diseases are multi-factorial diseases, thus several factors amplify the
inflammatory process making some individuals more susceptible to periodontal disease
than others.
 One of the most important factors that determine the outcome of periodontal disease is
patient susceptibility. Although bacteria are the primary etiological agents, the host
immune response to these bacteria is of utmost importance.

LIPOPOLYSACCHARIDE (LPS)
LPS is a cell wall constituent of virtually all subgingival gram negative micro-organisms.
They are known to induce; PMN leukocyte infiltration in inflamed periodontal tissues, oedema
and vascular dilatation, absence of osteoblasts, disappearance of osteocytes and to have a
stimulatory effect on osteoclasts and on collagen phagocytosis.
Thus LPS is a potent inducer of inflammatory cytokines synthesis: IL-1, PGE2, TNFα, IL-6 and IL-
8.

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PROGRESSIVE “NATURAL HISTORY” OF PERIODONTAL DISEASE LESIONS:
1. Initial lesion: The initial lesion begins within 2-4 days after deprivation of oral hygiene.
In this stage we can see few neutrophils in the junctional epithelium, which are in the
process of migration into the gingival crevice. There are small numbers of macrophages
and lymphocytes in the connective tissue, and the lymphocytes are primarily T-cells.The
neutrophils infiltrate the junctional epithelium and gingival crevice whereas
lymphocytes and other mononuclear cells are within the subjacent connective tissue.

Main histological findings:

a. Vascular dilation and vasculitis subsequent to the junctional epithelium.
b. Infiltration of polymorphonuclear neutrophils (PMNs) into the junction and
sulcular epithleium.
c. Predominant immune cells are PMNs.
d. Perivascular loss of collagen.
e. Alteration of the coronal part of the junctional epithelium.

2. Early lesion: It progresses from early lesion within 4-7 days and continues up to 14 days.
Histological examination shows a marked increase in lymphocytes infiltrating the
subjacent connective tissue; but also, the junctional and sulcular epithelium continue in
conversion to pocket epithelium. Neutrophils increase in numbers in the periodontal
pocket until about the 12th day. More coronal aspects of the junctional epithelium may
resemble micro-abscesses, with severe leukocyte infiltration. Lymphoid cells dominate
the subjacent connective tissue, comprising up to 75% of the inflammatory cells
infiltrate. Early in this period, the predominating lymphocytic infiltrate are T-cells, but
eventually, B-cells become abundant and dominate later in the lesion. T-cell blast
transformation (ie., proliferation and functional activation) can be observed.

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 Main histological findings:
a. Proliferation of blood vessels
b. Formation of rete peg and atrophic areas in the SE and JE.
c. Predominant immune cells are lymphocytes (75%)
d. Increased collagen loss, 70% of collagen destroyed around the cellular infiltrate.

3. Established lesion: After 14 days, the neutrophil infiltration of the pocket and junctional
epithelium is intense. B-cells undergo blast transformation and conversion to plasma
cells (with local production of antibody) and monocytes derivatives clearly look like
macrophages. The established lesion was could be stable for many years.
Main histological findings:
a. Proliferation of blood vessels and blood stasis.
b. More advanced areas of rete-peg formation and atrophic areas in the SE and JE.
c. Predominant immune cells are plasma cells.
d. Continued collagen destruction.
4. Advanced lesion: The lesion is considered advanced when the destruction of bone is
evident. At this late stage, the pocket epithelium shows ulceration, junctional
epithelium shows great apical extension. Plasma cells continue to dominate the
connective tissues, and neutrophils continue to dominate in the junctional epithelium
and gingival crevice. Many of the plasma cells appeared to be “degenerating” 10,
probably normal apoptosis, in retrospect. One of the proposed mechanisms for the
release of the cytokine, interleukin-1ß, (IL-1ß), may involve apoptosis, since IL-1ß is
activated by the cytosolic enzyme important in apoptosis (caspase-1; aka “interleukin-1ß
converting enzyme [ICE]”).

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 SUMMARY:
 While the advanced lesion represents periodontitis the others are responsible
for gingivitis.
 Neutrophils are the first leukocytes to arrive and in pathogenesis of periodontal
disease, neutrophils are always the dominant cell type within the junctional
epithelium and gingival crevice.
 Periodontitis usually progresses from gingivitis and this transition occurs after B
cell changes to plasma cell.

The pathogens in periodontal disease and their products, as well as inflammatory

mediators produced during the diseases process are actually responsible for the
connective tissue destruction.

PROGRESSION FROM THE INITIAL LESION TO FORM AN ESTABLISHED LESION

a) Early reaction to plaque accumulation (Initial lesion):

The bacteria present in the plaque produce various substances which can initiate vascular
events of acute inflammation. These products are metabolic acids, bacterial lipopolysaccharides
(LPS), FMLP (N- Formyl- Metheonyl-Leucyl-phenylalanine), volatile sulphur compounds,
extracellular enzymes and fatty acids.
1. These mediators stimulate the cells of junctional epithelium to produce various inflammatory
mediators like IL-8, TNF-α, PGE2, IL1-α and MMP.
2. Neuropeptides and histamine produced by free nerve endings causes vascular effects in that
area. These mediators cause increased vascular permeability.
3. Perivascular mast cells produce histamine which causes the endothelium to release IL-8
which leads to the PMN recruitment.
4. Because of alteration in vasculature in that area plasmin which is a part of fibrinolytic system
gets involved in the inflammatory response. It causes degradation of fibrin. Degradation
products of fibrin further promote vascular permeability.

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5. The above changes lead to the alteration in gingival vascular plexus. The vessels proliferate,
and a distinct loop formation develops in the small vessels directly below the basement
membrane of the sulcular epithelium.

b) Initiation of the cellular response (Early lesion):

1. Initiation of the host immune reaction against bacteria and their products lead to the
activation of complement system.
2. The products of complement system like C5a are chemoattractant for neutrophils. This leads
to the recruitment of leukocytes and monocytes in that area.
3. Cellular response is characterized by the presence of neutrophils which are hallmark of acute
inflammatory reaction. Because of increase in vascular permeability PMN’s emigrate from
blood vessels into the connective tissue by a process called trans-endothelial migration.
4. Activated macrophages produce various chemical mediators which further intensify the
inflammatory reaction. These include IL-1β, IL-6, IL-10, IL-12, TNF-α, MMP, INF-γ, PGE2.
5. They also release chemotaxins like monocytes chemo-attractive protein (MCP), macrophage
inflammatory proteins (MIP), RANTES (regulated on activation, normal T-cell expressed and
secreted).

c) Initiation of connective tissue breakdown (Established lesions)

1. As duration of inflammatory response lengthens, the inflammatory infiltrate is dominated by
lymphocytes. Subsets of T-cells i.e. helper T-cells (Th) and cytotoxic T-cells (Tc) are involved in
regulating the immune responses. They also help in initiation of B cell response that leads to
formation of antibodies.
2. T-cell mediated response leads to release of various interleukins, TNF-α, TGF-β, INF-γ.
Cytokines released by T-cells can activate macrophages. Macrophage is the cell that is
classically present in chronic inflammation. They have important role in antigen presentation,
and as the duration of inflammatory lesion increases, the number of macrophages increases.
3. Plasma cells synthesize antibodies and are responsible for humoral response. Mature plasma
cells may be seen in chronically inflamed tissue.

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4. Activated PMN’s synthesize diverse cytokines, leukotrienes and MMP’s. These are
responsible for progressive breakdown of connective tissue.
5. Activated fibroblasts also produce MMP’s and TIMP’s which further leads to connective
tissue alterations.

d) Initial attachment loss (Advanced lesion):

1. This stage is dominated by plasma cells.
2. Connective tissue infiltrated by various immunologically active cells lead to its degradation.
The activated macrophages and other cells further elaborate the immune response.
3. Immunocompetent cells produce various cytokines, matrix metalloproteinases, PGE2 and
tissue inhibitors of matrix metalloproteinases.
4. These all events lead to the disturbed tissue homeostasis and destruction of connective
tissue matrix, collagen fibres and bone. This ultimately leads to the pocket formation and
consequently periodontitis.

ROLE OF COMPLEMENT SYSTEM IN IMMUNOLOGY OF PERIODONTAL DISEASE:

The complement system is a series of proteins circulating in the blood and bathing the fluids
surrounding tissues. It is a major component of innate immune system involved in defending
against all the foreign pathogens through complement fragments that participate in
opsonization, chemotaxis, and activation of leukocytes and through cytolysis by C5b-9
membrane attack complex.
Complement proteins circulate in the blood in an inactive form.
Complement system and proteins can be activated by immune complexes and immunologic
molecules like endotoxins to form active enzymes.
The activation of complements occurs by 3 mechanisms:
a. Classical Pathway.
b. Alternate Pathway.
c. Lectin Pathway.

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For example, the relative quantity of C3 fragments has been shown to correlates with the
severity of inflammation and the amount of supragingival plaque.
In addition, C4 cleavage is rarely seen except in the case of LJP.

ROLE OF VARIOUS INFLAMMATORY MEDIATORS IN PERIODONTAL DISEASES:

Various inflammatory mediators play a crucial role in acute and chronic inflammation of
periodontal tissue.
These include a range of interacting molecules like the cytokine system, proteinases, proteinase
activators, thrombin, histamine, prostaglandins, leukotrienes, tissue and blood factors such as
Hageman’s factor, complement and clotting factors.
These mediators provoke the initiation and eventually termination of an inflammatory response
to insult and they are present in high concentrations in inflamed gingiva and gingival crevicular
fluid of diseased sites. Their concentration decrease following successful periodontal
treatment.
In some individuals the elevated amount of these mediators is released from cells like
monocytes which cause rapid destruction of periodontal connective tissue.

a) Role of Cytokines in periodontal disease:

The term ‘‘cytokine’’ means cell movement. They can be classified into following catagories:
a. Pro-inflammatory cytokines.
b. Cytokines with predominant Immunoregulatory Functions.
c. Cytokines that regulate lymphocyte growth, activation and differentiation.
d. Cytokines that help in haematopoiesis.
e. Chemokines.

Interleukin-1α (IL-1α), interleukin-1β (IL-1β), interleukin 6 (IL-6), interleukin-8 (IL-8), and tumour
necrosis factor-α (TNF-α) can be detected in gingival crevicular fluid (GCF) in periodontal
disease and that the cytokine levels in GCF are closely associated with the severity of
gingival inflammation and/or periodontal tissue destruction.
Interleukin-1 (IL-1):

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IL-I is a polypeptide having important roles in immunity, inflammation, tissue breakdown, and
tissue homeostasis.
Functions: IL-α, IL-β, and tumor necrosis factor-alpha (TNF-α) stimulate bone resorption and
inhibit bone formation.
In addition, IL-1 synergizes the bone-resorptive actions of TNF-α.
IL-β is major mediator of tissue destruction in periodontal disease.
IL-β is a potent stimulator of matrix metalloproteinase (MMP’s) production (collagenases,
gelatinases, stromelysins) thus indirectly involved in degradation of extracellular collagenous
matrix.
Interleukin-6 (IL-6):
IL-6 is responsible for the induction of the final maturation of B-cells into plasma cells that
secrete immunoglobulins.
The expansion of B-cells/plasma cells seen in periodontitis lesions may result from an increased
production of IL-6 at diseased sites.
Interleukin-8 (IL-8):
Secreted by monocytes, fibroblasts, lymphocytes, and endothelial cells.
IL-8 is a potent chemotactic factor for leukocytes.
IL-8 induces neutrophil extravasation and plays a very important role in accumulation of
leukocytes at the site of inflammation.
IL-8 may also attract T-cells and induce their motility in inflamed gingiva.
TNF-α:
It stimulates secretion of collagenase by fibroblasts causing resorption of cartilage and bone,
and it has been implicated in the destruction of periodontal tissue in periodontitis.
Lipopolysaccharide (LPS) is a strong inducer of production of TNF-α by peripheral blood
monocytes, which in turn leads not only to alveolar bone resorption but also to the enhanced
synthesis of collagenase by human gingival fibroblasts. As it is potent inducer of collagenase
secretion by fibroblasts,

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PGE2:
PGE2 is the product of arachidonic acid metabolism. As it is one of the important mediators of
inflammation, PGE2 levels are elevated in periodontitis compared to gingivitis. PGE2 is detected
in higher levels in inflamed gingival tissue and GCF proportional to the severity of periodontal
disease.