Immunological tolerance Presented by Jintana Chataroopwijit, MD. December9, 2016

1. IMMUNOLOGICAL TOLERANCE
JINTANA CHATAROOPWIJIT
9 DECEMBER 2016

2. DEFINITION
• Tolerance : Unresponsiveness to antigen that is
induced by previous exposure to that antigen
• Inherent property of immune system
• Response against foreign antigen (nonself) without
attacking host (self)
• Specific lymphocyte + Antigen --> activated lymphocyte
--> immune responses or tolerance (
inactivation/elimination )

3. DEFINITION
• Tolerogens/Tolerogenic antigens : antigens that
induced tolerance
• Self-tolerance : Tolerance to self antigens
• Autoimmunity : Failure of self-tolerance --> immune
reaction against autologous antigens

4. PHYSIOPATHOLOGY OF IMMUNE
TOLERANCE-RELATED DISEASE
• Complex
• Influenced by
• Genetic susceptibility
• Route of exposure
• Antigen dose
• Time of exposure
• Structural characteristics of allergen and antigen
• Coexposure with stimulators of innate immune response ex. Infections
or commensal bacteria

5. TOLERANCE
1. Central Tolerance
2. Peripheral Tolerance

6. CENTRAL TOLERANCE
• First step of tolerance during maturation in thymus
• Prethymic T cells enter thymus and reach
subcapsular region --> proliferate as large
lymphoblast

7. CENTRAL TOLERANCE
• Maturing cells move deeper into cortex and adhere
to cortical epithelial cells ( increased in expression of
CD3, CD4, CD8 and TCR )
• T cell receptors (TCRs) on thymocytes are exposed
to MHC molecules through these contact

8. CENTRAL
TOLERANCE
• TCRs + autoantigens
(medullary thymic
epithelial cells,
interdigitating cells and
macrophage at
corticomedullary
junction) --> deleted
• Cells expressing CD4 or
CD8 --> periphery

10. FACTORS TO INDUCE NEGATIVE SELECTION
OF SELF REACTIVE THYMOCYTE
(INTRATHYMIC SELECTION)
• Not known
• Possible factors
• Affinity of antigen recognition
• Type of antigen-presenting cells presenting the antigen
• Locally availability of cytokine in thymus
• Presence of antigen in thymus ( local or delivery by blood )
• Affinity of thymocyte T cell receptors (TCRs) to recognize
antigen

11. AIRE
( AUTOIMMUNE RESPONSE ELEMENT)
• Transcription factor controlling expression of some
organ-specific "peripheral antigens" in thymus
• Component of multiple protein complex
• Function : Transcriptional regulator to promote
expression of selected tissue-restricted antigen in
thymus

14. AUTOIMMUNE POLYENDOCRINE SYNDROME TYPE 1
(APS1) OR AUTOIMMUNE POLYENDOCRINOPATHY-
CANDIDIASIS-ECTODERMAL DYSTROPHY/DYSPLASIA
(APECED)
• In mouse model : knockout of AIRE gene
• In mice : low level several proteins in medullary
thymic epithelial cells in peripheral organs ex.
pancreatic insulin
• Characterized by antibody and lymphocyte-
mediated injury to multiple endocrine organs (
parathyroids, adrenals and pancreatic islet )

15. PATHWAY OF APOPTOSIS

17. NECROSIS VS APOPTOSIS

20. TWO SIGNAL MODEL : ANERGY
Signal 1
Recognition by
helper lymphocyte
No Yes
Target lymphocyte
(Naive CD4 T cell)Signal 2
APC + costimulator
molecules
-->
Costimulatory
signal
Signal1 : TcR bind their peptide/class II
MHC complex

23. REGULATION OF T CELL
RESPONSES BY INHIBITORY
RECEPTORS
• Balance between engagement of activating and
inhibitory receptors --> outcome of antigen
recognition by T cells particularly CD4+ cells
• Inhibitory receptors with physiologic self-tolerance :
CTLA-4 and PD-1

24. CTLA-4
• Negative
regulator of
adaptive immune
responses
• Bind to B7 (CD80
and CD86)
costimulatory
molecules on
APCs

25. CTLA-
4

26. CTLA-4
• Knockout mice lacking CTLA-4 --> uncontrolled
lymphocyte activation with massive enlarge lymph
node and spleen and fatal multiorgan lymphatic
infiltration : autoimmunity
• Failure of peripheral tolerance and severe T cell
mediated disease

27. CTLA-4
• Animal model : blocking of CTL-4 with antibodies -->
autoimmune disease ex. encephalomyelitis
• In human : Polymorphism in CTLA-4 gene : type 1
diabetes and Graves' disease

28. ACTION OF CTLA-4
• Low on most T cells until cells are activated by
antigen
• Inhibiting activation of naive T cells
• Express on regulatory T cells

29. THERAPEUTIC APPLICATION
• Blocking CTLA-4 --> increased immune responses
to tumor
• Anti-CTLA-4 antibody : approved for advance
melanomas

30. PD-1
• Programmed cell death 1
• Immunoreceptor tyrosine-based inhibitory motif-
containing receptor
• Express on T cell activation

31. PD-1
• Recognized 2 ligands
1. PD-L1 : expressed on APCs and many other tissue
cells
2. PD-L2 : expressed mainly on APCs

32. PD-1
• Important in terminating peripheral responses of
effector T cell esp. CD8+ cell
• May not be required for function of regulatory T cells

33. PD-1
• Engagement with ligand -->
• Inactivation of T cells
• Inhibit IL-2 production
• May play role in suppressive function of Treg
cells

34. PD-1
• In mice : PD-1 knocked out --> autoimmune
diseases ex. Lupus-like kidney disease and arthritis
• Autoimmune disorder in Pd-1 knockout mice less
severe than CTLA-4 knockouts

35. A. Direct deletion of immune effector :
expression of death-inducing ligand
B. Direct tolerization effector T cells :
suppressive cytokines released by
tissue cells
C. Suppression effector T cells by
regulatory T cells
D. Tolerization of host T cells by
tolerizing dendritic cells
E. Ignorance : spatial seperation of T
cells and tissue cells ex. Basement
membranes
F. Immune priviledge

36. IMMUNE PRIVILEDGE
• Certain site in body tolerate induction of antigen
without eliciting inflammatory immune response
• Maybe for protect vital structures from potentially
damaging effects
• Sites : brain, anterior chamber of eyes, placenta,
fetus, testes

37. IMMUNE PRIVILEDGE : FETUS
• Express MHC derived from both parents
• Peripheral tolerance of mother to fetus --> fetal
survival
• Cells of villous trophoblast lack expression of MHC
class I

38. IMMUNE PRIVILEDGE : FETUS
• Increase expression of non-classical MHC molecule
ex. HLA-G : inhibitory receptor
• Immune deviation to Th2
• Increase expression of FasL at placenta

39. IMMUNE PRIVILEDGE : EYE AND
BRIAN
• Limit capacity for regeneration
• Immune response in these area could have
devastating effect on individual
• Low or no expression of classical MHC class Ia
protein on cell
• "Sympathetic ophthalmia"

40. IMMUNE PRIVILEDGE :
POSSIBLE MECHANISM
• Limit lymphatic drainage
• Cytokine : inhibition of inflammation
• TNF-beta and MIF(migration inhibitory factor) :
inhibition of NK cell mediated cytolytic activity

41. NATURALLY OCCURING
CD4+CD25+FOXP3+ REGULATORY T
CELLS
• in healthy : <5% of CD3+CD4+ population
• Expression of high levels of alpha chain of CD25, IL-2
receptor
• Hypothesis for generation of Treg cells
1. From thymus : specific for self-peptides
2. From naive T cells in periphery : required for
environmental antigen/allergen specific T cells

42. NATURALLY OCCURING
CD4+CD25+FOXP3+ REGULATORY T
CELLS
• TNFRSF18/GITR(glucocorticoid-induced tumor
necrosis factor receptor family related-gene
• Expressed by activated Treg cells
• Be trigger : role in resistance to Treg cell
mediated-suppresion

43. NATURALLY OCCURING
CD4+CD25+FOXP3+ REGULATORY T
CELLS
• CD103 (alphaEbeta7)
• CD122 (beta chain of IL-2 receptor)
• Both highly expressed on Treg cells
• Correlated with their suppressive activity

44. NATURALLY OCCURING
CD4+CD25+FOXP3+ REGULATORY T
CELLS
• Other proposed markers
• Certain chemokine receptors
• TLRS
• Membrane-bound TGF-beta
• Neuropilin 1 ( NRP1 )
• Lymphocyte activation gene ( LAG3 )
• Granzyme

45. NATURALLY OCCURING
CD4+CD25+FOXP3+ REGULATORY T
CELLS
• Additional marker from gene arrays
• GPR83 : G protein-coupled receptor 83
• ECM1 : extracellular maxtrix 1
• IKZF2 : IKAROS family zinc finger 2-helios

46. REGULATORY T CELL
GENERATION
• Augmented by
• FOXP3+ T reg cells
• Low doses of pathogen-derived molecules :
filamentous, hemagglutinin
• Exogenous signals : histamine, adenosine,
vitamin D3 metabolites
• Retinoic acid

47. REGULATORY T CELL
GENERATION : RETINOIC ACID
• Balance of inflammatory Th17 cells and suppressive
Treg cells by Th17 cells
• Enhancing expression of FOXP3 through
STAT3/STAT5 independent signaling pathway

48. REGULATORY T LYMPHOCYTE
• Mostly express high levels of IL-2 receptor alpha
chain (CD25)
• Transcription factor for development and function :
FOXP3

49. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• In mice : expressed by naturally occuring Treg cells
• In humans : upregulation in all activated T cells
• Required for development and function of naturally
occuring Treg cells

50. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• Directly interact with RUNX1( runt-related
transcription factor 1)
• RUNX1( runt-related transcription factor 1)
• Impair expression of IL-2 and IFN-gamma
• Exert suppressive activity
• In murine : maintain high level of FOXP3
expression

51. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• Induction of RUNX1 and RUNX3 by TGF-beta :
generation and suppresive function of induced Treg
cells
• RUNX1 and RUNX3 bind to FOXP3 promotor :
expressing functional Treg cells

52. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• Leucine-rich repeat-containing 32 receptor (LRRC32
or GARP) : key receptor to control FOXP3 levels in
naturally occurring Treg cells through positive-
feedback loop

53. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• Cytokines : IL-2, IL-10, TGF-beta
• Surface markers : CD25, CTLA4, CD103, GITR,
NRP1, latency-associated peptide (LAP)

54. FORKHEAD WINGED
TRANSCRIPTION FACTOR :
FOXP3
• ICOS+FOXP3+ Treg cells use IL-10 and TGF-beta
to suppress dendritric cells and T cells function
• ICOS-FOXP3+ Treg cells express TGF-beta
• Marker to differentiate between human regulatory
and activated effector T cell : alpha chain of IL-7R
(CD127)

55. MUTATION : DIMINISH
FUNCTION
• X-linked autoimmune and allergic dysregulation
syndrome (XLAAD)
• Immune dysregulation, polyendocrinapathy,
enteropathy, X-linked syndrome (IPEX)

56. IMMUNE DYSREGULATION,
POLYENDOCRINAPATHY, ENTEROPATHY, X-
LINKED SYNDROME : IPEX
• Rare autoimmune disease
• Mutation of FOXP3 gene
• Associated with deficiency of regulatory T cells
• Significant skewing of T lymphocyte toward Th2
phenotype

57. IMMUNE DYSREGULATION,
POLYENDOCRINAPATHY, ENTEROPATHY, X-
LINKED SYNDROME : IPEX
• Autoimmunity
• Severe atopy : eczema, food allergy
• Eosinophillic inflammation

59. REGULATORY T CELL
GENERATION
• Immune response of memory T cells : essential for
inflammation and immune regulation processes in
diseases ex. allergic rhinitis, asthma, atopic
dermatitis

63. GENERATION AND MAINTAINANCE
OF REGULATORY T CELLS
• Regulatory T cell
1. Thymic / natural regulatory T cells
2. Peripheral / adaptive / inducible

64. SUBSETS OF REGULATORY T
CELLS

65. MECHANISM OF ACTION OF
REGULATORY T CELLS
• Directly suppress B cell activation
• Inhibit proliferation and differentiation of NK cells
• Production of immunosuppresive cytokine ( IL-10
and TGF-beta )
• Reduced ability of APCs to stimulate T cells
• Consumption of IL-2

66. TRANSFORMING GROWTH
FACTOR-BETA (TGF-BETA)
• Tumor product
• Promote survival of tumor cells in vitro
• Type : 1-3
• Mostly : TGF-beta1

67. TRANSFORMING GROWTH
FACTOR-BETA1 (TGF-BETA1)
• Produced by CD4+ regulatory T cells, activated
macrophages and other cells
• Synthesized as inactive precursor
• Proteolytically cleaved in Golgi complex and form
homodimer
• Mature form is secreted in latent form in associated
with other polypeptides

68. TRANSFORMING GROWTH
FACTOR-BETA 1 (TGF-BETA1)
• In mice : Suppress airway disease
• Target cell : T cells
• Association with airway inflammatiom

69. • Maintain tolerance esp. Oral tolerance
• Inhibit proliferation , differentiation and survival of B
and T lymphocyte
• Inhibit immunoglobulin isotype switching and
promote differentiation of IgA secreting plasma
ROLE OF TRANSFORMING
GROWTH FACTOR-BETA (TGF-
BETA)

70. • Promote differentiation of Langerhan cells and DCs
with immature phenotype
• In mast cell : promote chemotaxis but inhibit
expression of high-affinity receptor for Fc fragment
of IgE
• Inhibit human Th2 response in vitro
ROLE OF TRANSFORMING
GROWTH FACTOR-BETA (TGF-
BETA)

71. ROLE OF TRANSFORMING
GROWTH FACTOR-BETA (TGF-
BETA)
• Regulate differentiation of functionally distinct
subsets of T cells
• Promote tissue repair after local immune and
inflammatory reaction subside

72. INTERLEUKIN-10
• Control of allergy and asthma
• Synthesized by B cells, monocyte, DCs NK cells and
T cells
• Inhibit proinflammatory cytokine production TH1 and
Th2 cell activation

73. INTERLEUKIN-10
• In T cells : IL-10 receptor-associated tyrosine kinase
2 (TYK2) - constitutive reservoir for protein tyrosine
phosphatase nonreceptor 6 (PTPN6/SHP1)
• PRPN6 rapidly binds to CD28 and ICOS
costimulatory receptors and dephosphorylates them
within minutes

74. INTERLEUKIN-10
• Member of family of heterodimeric cytokines : IL-22,
IL-27 and others
• Consist of two chains
• IL-10 receptor belongs to type II cytokine receptor
family

75. INTERLEUKIN-10
• Associate with JAK1 and TYK2 Janus family kinase
and activate STAT3
• Produced by activated macrophage and dendritic
cells, regulatory T cell, TH1 and TH2 cells and some
B lymphocyte

76. ROLE OF INTERLEUKIN-10
• Inhibit expression of costimulators and class II
molecule on dendritic cells and macrophage -->
inactivation
• Control of innate immune reactions and cell-
mediated immunity
• Inhibit production of IL-12 by activated dendritic cells
and macrophage

77. MUTATION INTERLEUKIN-10
RECEPTOR
• Rare inherited autoimmune disease
• Severe colitis

79. FACTORS FOR DETERMINE
IMMUNOGENICITY AND TOLEROGENICITY
OF PROTEIN ANTIGEN

80. B LYMPHOCYTE TOLERANCE
• Maintain unresponsiveness to thymus independent
self antigens (polysaccharide and lipid)
• Prevent antibody responses to protein antigens

81. CENTRAL TOLERANCE IN B
CELLS

83. PERIPHERAL TOLERANCE IN B
CELLS

86. “Thank you.”