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Cytokine

Perio Craze
http://drprem.blogspot.com
Introduction
 The immune system
 Generates variety of cells and molecules
capable of specifically recognizing and
eliminating an apparently limitless variety of
foreign invaders.
 Exquisitely adaptable dynamic network
which is complex.
Complex System
Cell “Cross-Talk”
Cytokine
 Group of intercellular signaling low
molecular weight proteins
 Provides a network controlling local and
systemic immune and inflammatory
responses
 Affects wound healing, hematopoiesis and
other biologic process.
Definitions
Encyclopedia of Immunology

Okada and Murakami 1998

Offenbacher 1999
TERM
Classification on Biologic
Activities
Affecting macrophages:
 Migration inhibition factor
 Macrophage activity factor
 Macrophage aggregation factor
 Macrophage chemotactic factor
 Macrophage resistant factor
Affecting lymphocytes:
 Blastogenic or mitogenic factor
 T-cell growth factor
 B-cell growth factor
Affecting granulocytes:
 Chemotactic factor
 Colony stimulating factor
 Inhibition factor
Classification on Biologic
Activities
Affecting cultured cell:
Lymphotoxin
Gamma interferon
TNF
Colony inhibitory factor.
Producing in vitro effects:
Osteoclast activating factor
Transfer factor
Procoagulant factor
Nissengard Newman
Classification
 First group:
 Cytokines which serve as mediators of innate
immunity E.g. α ,β, interferon, TNF, IL-6
 Second group
 Cytokines that regulate the growth &
differentiation of lymphocytes. E.g. IL 2,4 TGF β
 Third group
 Cytokines that regulate hematogenous activity
E.g Gamma CSF, IL-3,7
 Fourth group
 Cytokines that share the common property of
being activation of inflammatory cell function
e.g gamma interferon
Jan Lindhe’s Classification
 Pro inflammatory cytokines
 E.g. IL -1,6, TNF α – stimulates bone
resorption
 Chemotactic cytokines
 E.g. IL-8 ,16
 Lymphocyte signaling cytokines
 Released by Th 1- IL 2, IFN
 Released by Th 2- IL4,5,10,13
Based on Characters of
Cytokines
 Cytokines that regulate lymphocytes
 E.g.: IL 2,4
 2. Cytokines that activate
inflammatory cells
 E.g.: IFN gamma, TNF beta
 3.Cytokines that stimulate
hematopoiesis
 E.g.: IL3,7
 4.Colony stimulating factor
 E.g.: CSF
Biological Properties
Overview of Cytokine
Production
Pleiotropy
Redundancy
Synergism and Antagonism
Cascade Induction
Cytokines Vs Hormones
Cytokines Vs Growth
Factors

Production of growth factors not as


tightly regulated as Cytokines
The major action of growth factors
targeted at non hematopoietic cells
Biological Functions
In the development of cellular and
humoral immune responses
Induction of inflammatory
response
Regulation of hematopoiesis
Control of cellular proliferation
Differentiation and the healing of
wounds.
Cytokines Involved in Innate
Immunity

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IL 2
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Modified from Cell. Mol. d


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Immunol.
Bl ssel
Abbas & Lichtman 5th Ed. e
Mechanism of Action
Engages Cytokine receptors on cells
Induces dimerization or
polymerization of receptor
polypeptides
Activation of intracellular signaling
pathway
Results in active transcriptive factors
Transcription of specific gene
Resulting gene mediate biological
effects
Cytokine Receptors
Immunoglobulin Receptor
Family
Ligands – IL-1, CSF
Class I Cytokine Receptor
Family
Class I Cytokine Receptor
Family
Most receptors belong to Class I
Cytokine family
Class I : consists of four positionally
conserved cysteine residues ( CCCC)
Conserved sequence of trytophan-
serine-(any amino acid)-trytophan-
serine (WSXWS)
Ligands – IL-
2,3,4,5,6,7,9,11,12,13,14,15, etc
Class II Cytokine Receptors
Possess CCCC motifs but lack WSXWS
TNF Receptors
Chemokine Receptor
Signal Transduction
Dimerization of receptors
Receptors – has two polypeptide
chains:
 α chain – for cytokine binding and signaling
 β chain – for signaling, minor role in binding
Differentinactive protein kinases are
associated with the receptor
Janus Kinases (JAK)
STAT
Activated JAK creates docking sites
Transcription Factor - Signal
Transducers and Activators of
Transcription
Phosphorylates key tyrosine
Dissociates from receptor subunits
STAT dimers translocate into nucleus
and induce expression of genes
The resulting gene products then
mediate the various effects typical of
Cytokine Antagonists
Certain proteins inhibit biological
activity of cytokines
They act by either by
 Binding to Cytokine receptor but failing to
activate the cell
 Or they bind directly to the cytokine and
inhibits its activity
Description of each
Cytokine
Interleukin 1 (IL 1 α,IL 1 β)
IL-1
 Up regulated by interferons, TNF,GM
CSF,IL-2,3
 Inhibited by corticosteroids &
prostaglandins
 IL 1 is an important regulator of
hematopoiesis .effect is based on
augmenting
 Induces production of various colony
stimulating factors by bone marrow
stromal cells.
 It is both radio and chemo protectant.
 Restoration of hematopoiesis can also be
observed when IL- 1 is given after
The Perio Relation of IL1
As a marker of Periodontitis
Potent proinflammatory molecules
Previously termed as “osteoclast
activating factor”
IL-1ra binds to the IL -1 receptor
without stimulating host response
Genetic polymorphism of IL-1 and
Periodontitis
Interleukin 2
Interleukin 3
An important growth factor
Secreted by: T h cells,NK cells, mast
cells.
Detected in blood in when massive
activation of T cells occurs – such as
graft vs host
Stimulates mast cell growth and
histamine secretion
Interleukin 4
Interleukin 5
Interleukin 6
Interleukin 7
Interleukin 8
Interleukin 9

Molecular wt :30,000 to 40,000


Secreted by Th cells
Acts on Th cells as a mitogen
,supporting proliferation in absence of
antigen.
Interleukin 10
Interleukin 11
Secretedby bone marrow stromal cells
Promotes differentiation of progenitor B
cells
Promotes differentiation of
Megakaryocytes
Interleukin 12
Interleukin 13
Interleukin 14
Molecular wt : 50 to 60 kd
IL14 thought to play a role in
development of B cell memory .
It has been characterized only in
human being produced by follicular
dentriticcells and activated T cells.
It enhances the proliferation of
activated B cells and inhibit synthesis
of Ig.
Interleukin 15
Stimulate intestinal epithelium
proliferation
IL 15 widely expressed in platelets,
skeletal muscle, kidney, lung, liver,
heart, bone marrow
Mitogenfor activated B cells
Interleukin 16
Interleukin 17
Secreted by T cells

Acts on macrophages and thus


initiates & maintains inflammation
Interleukin 18
Secreted by activated macrophages

Induces T-cells for IFN gamma


production

Enhances NK cell cytotoxicity.


Immuno Regulatory
Cytokines

Immuno Regulatory
Cytokines
TGF Beta
TGF beta
Humans expresses 3 forms: TGFβ
1,2,3
These are the products of separate
genes, but they all bind to 5 types of
high affinity cell surface receptors
 Type I,II - receptors transducesignals

 Type III,IV,V - receptor roles not clear.


Suppressesproduction of lymphokines
and monokines
TNF
TNF alpha and beta
TNF alpha
 Cytotoxic effect on Tumor cells
 induces cytokine secretion and is responsible
for extreme weight loss(cachexia) associated
with chronic inflammation
TNF beta
 Cytotoxic effect on Tumor cells
 Enhances phagocyticactivities of neutrophils
and macrophages.
Interferons
Classification
Type I:
 IFNs( anti viral activity)E.g. interferon α,β,ω .

Type II :
 IFNs ( immune interferon) E.g.IFN γ
Interferon α,β
IFN α – Secreted by lymphocytes
 Inhibits viral replication in uninfected cells
IFN β – Secreted by fibroblasts
 Also inhibits viral replication in uninfected
cells
Interferon Gamma
Secreted by T H1 ,T C ,NK Cells
Inhibits viral replication in uninfected
cells
Enhances activity of macrophages
Oncostatin
Pleitropic cytokine that belongs to the
Interleukin 6 group of cytokines
Secreted by macrophages, T cells
Inhibits growth of tumor cells
Induces hepatocytes to synthesis
acute phase protein
Stimulates growth of Kaposi’s sarcoma
RANTES
 Regulated upon Activation Normal T cell
Expressed and Secreted.
 Source: T Lymphocytes, Platelets
 Chemo attracted cells: monocytes,T-
lymphocytes,basophils, eosinophils, mast
cells.
 Other activities
 Macrophage activation
 Integrin expression by T cells
 Chronic inflammation
 Basophil degranulation
Cytokine Assays

Cytokine Assays
ELISA
 Coat well of micro titreof place with anti cytokine
antibody
 Block un occupied sites with protein
 Add cytokine (the cytokine will be captured by the
antibody)
 Add enzyme conjugated anti cytokine antibody
(secondary) It forms a sandwich with the captured
cytokine .
 Add chromogenic substrate .the enzyme generates
a colour whose integrity is proportional to the
amount of cytokine bound to the captured
antibody.
 Optimal density of colour is measured
Immuno Radiometric
Assays
Bead coated with anti- cytokine
antibody
Add cytokine
Add iodinated anti cytokine
monoclonal antibody
It is sandwiched to capture cytokine
and because it is radio-labelled it is
detected radio metrically
Regulation of Cytokines
 Receptor antagonists bind to a specific receptor
but do
not transmit a signal (competitive inhibition)
 The extra cellular domains of cytokine receptors
can be
shed. They bend their cytokine in the fluid phase
and to stop the cytokine from reaching receptor on
cell membranes.
 After the cytokine bind to receptors another
cytokine bind to the other receptor of cell and
produce opposite effects on the cell.
 Some cells can express cytokine binding molecules
(deceptors) which specifically bind ligand but do
The Perio View

The Perio View


Cells producing Cytokines in
Periodontitis

Monocytes / Macrophages
 Macrophages constitute 5 to30% of
infiltrating cells in inflamed periodontal
lesions.
 They produce cytokines IL
1,6,10,12,13,TNF ALPHA,IFNα
 They produce a central role in production
of IL1 in inflamed sites.
 Studies have shown that
periodontopathic bacteria such as P.g,
A.a,F.n induce expression of IL1 in mono
nuclear Cells.
Lymphocytes
Activated T cells with a helper CD 4
phenotype secrete a variety of
cytokines such as IL
2,3,6,9,10,13,15,TNF.
Cytokines are produced in large
numbers due to the numerous
lymphocytes in inflammatory
periodontal lesions produced
Gingival Fibroblasts
Abundant cells In periodontal tissues
Secretes IL 1,alpha,beta ,IL 6,TNF
alpha.
mRNA expression
Gingival fibroblasts lymphocyte
interaction may facilitate the cytokine
network in inflamed gingival tissues.
Gingival Epithelial Cells
 Gingival epithelial cells play an important
role as the first barrier against offending
bacterial agents in periodontal pockets.
 There is increased expression of IL 1
beta,6,8,TNF alpha in the activated
gingival epithelial cells lines.
 So it has been speculated that epithelial
cells confronting periodontopathic
bacteria may initiate the recruitment of
immuno competent cells by secreting
these cytokines.
Endothelial Cells

Endothelial cells have important role


in extravasion of leukocytes and
cytokine expression other than the
routine role to act as a non
thrombogenic substrate for blood
Role in Periodontal Disease
 Periodontitis – interaction of host defense
with micro organisms
 Loss of connective tissue support for the
dentition represents a defensive reaction to
the bacteria in the teeth.
 The host attempts to protect itself by
epithelial proliferation apically and by fibrosis
to wall of the lesion and reject the toxic root
surface of the tooth.
 T Lymphocytes predominatedin stable lesion
and B cells and plasma cells in Progressive
lesion.
 “Double edged Sword”
Cytokine and Perio
 IL1 alpha, beta, IL6,IL8,TNF alpha:Pro-
inflammatory cytokines,
 Involved in the resorption of alveolar
bone
 Cytokines that play important role in
inflammatory responses are also
prominent regulators of normal tissue
homeostasis.
 (e.g.) IL1 in periodontitis it cause fever,
bone resoption,collagenase and
plasminogen activation, has physiological
role – proliferation of keratinocytes,
fibroblasts, endothelial cells synthesis of
Cytokine and Perio
 Pilon et al – low levels of IL2 in GCF of
periodontitis sites
 Fyihashi et al – gingival mononuclear cells
from adult periodontitis patients produce
IL4,IL5 but not IL2 .
 Seymour et al – IL2 activity in peripheral blood
mononuclear cell cultures stimulated by two
putative periodontal pathogens, P.g, F.n was
found to be less than in unstimulated cultures.
 IL4 is the predominant cytokine in
periodontally affected tissues compared
with IL2,IFN gamma, IL6.
Cytokine and Perio
 Reinhardet al showed that concentration of IgG4
was many times higher in sites of active
periodontitis than in serum as well as significant
elevated conc. compared with stable lesions also
suggests the role for IL4 and increased TH2
responses in periodontitis
 Analysis of IL2: IL4 ratios revealed significant
lower ratios for cells derived from periodontitis
tissues compared with cells from gingivitis
lesions.
 Thesedata support the hypothesis that TH1 cells
are associated with stable lesion, where as TH2
may be important in pathogenesis of progressive
periodontal lesion
Cytokine Expression and PDL
Disease

Masada et al ( JPR 90)


Seymour et al(JPR 93)
Shinohara et al ( JPR 93 )
Reinhard et al ( JCP 03)
Cytokine Expression and PDL
Disease

Aslanidis et al ( JPR 2003 )


Kurdowska et al ( JPR 2003)
Faizuddin et al ( JPR 2003)
Ishihara et al ( JPR 2003 )
Lopes et al (JP 2005)
Cytokine Related Diseases
Bacterial
Septic Shock
Lymphoid and myeloid cancers
Chagas disease
Cytokine Therapy
Over production of cytokines may be
inhibited by cytokines suppressing
anti inflammatory drugs .
Defective or reduced cytokines and
their receptors can be replaced
directly to reconstitute a reduced
immune system or used to stimulate
further the immune system in case of
overwhelming infection or neoplasias
Conclusion
Signaling molecules
Found in GCF and Periodontal tissues
Used for treatment purposes
Further studies required to define role
of cytokine in periodontal pathology
Thank You

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