1. Lecture 4 - Immunology Antibody Lymphoid organs MHC

2. What is an antibody? Produced by Plasma cell (B-lymphocytes producing Ab) Essential part of adaptive immunity Specifically bind a unique antigenic epitope (also called an antigenic determinant) Possesses antigen binding sites Members of the class of proteins called immunoglobulins

3. What does an antibody look like ? 2 identical heavy chains 2 identical light chains Each heavy chain – has a constant and a variable region Each light chain has a constant and a variable region H H LL Constant region Variable region

4. Antibody: structure and function Fab – fragment antigen binding Fc- Fragment constant

5. Antibody: Fab Fab region Variable region of the antibody Tip of the antibody Binds the antigen Specificity of antigen binding determined by V H and V L

6. Antibody: Fc Fc region Constant region Base of the antibody Can bind cell receptors and complement proteins

7. Antibodies occur in 2 forms – Soluble Ag: secreted in blood and tissue – Membrane-bound Ag: found on surface of B-cell, also known as a B-cell receptor (BCR) Antibodies exist in two forms

8. Antibody: 5 classes (isotypes) There are 5 classes/isotypes of antibodies (IgM, IgG, IgD, IgA and IgE). In any given class of antibody, the constant region contains one of five heavy-chain sequences (µ, , , , or  ) called isotypes The heavy-chains determines the class of an antibody (µ, IgM; , IgG; , IgD; , IgA; and , IgE).

9. Immunoglobulin G (IgG) 2 Heavy and 2 Light chains (Monomer) ~75% of serum immunoglobulin. The major antibody of the secondary immune response (re-infection with the same pathogen) Fab Fc VHVH C H1 C H2 C H3 VLVL CLCL

10. Immunoglobulin M (IgM) 10 Heavy and 10 Light chains (Pentamer) <10% of serum immunoglobulin. May be secreted in saliva / milk Single J Chain (15 kDA) – helps in polymerization First Ab to be produced by virgin B cells when stimulated by an antigen; First Ab in primary infection J-Chain

11. Immunoglobulin A (IgA) in serum Monomeric form is present in serum ~15 % of serum immunoglobulins

12. Immunoglobulin A (IgA) in secretions dimeric J-Chain and secretory component Predominant antibody in secretions - saliva /milk Not found in serum; Confers mucosal immunity Secretory Component J-Chain

13. Immunoglobulin E (IgE) Monomeric Trace serum protein; bound to mast cells Associated with allergies (hypersensitivity)

14. IgE in allergy

15. IgE helps eliminate parasites

16. Immunoglobulin D (IgD) Monomeric Serves as a membrane receptor on B lymphocytes

17. Mature B-lymphocytes are coated with IgM and IgD

18. Antibody diversity The B cells produce millions of different types of antibodies If 1 gene encoded 1 immunoglobulin of a given specificity, approximately 10 7 - 10 8 genes would be required to encode JUST antibodies. However, we have only about 35,000 genes in total in the human genome This number of genes cannot encode ALL antibody specificities (~10 7 – 10 8 ) How is such antibody diversity possible ?

19. Antibody diversity https://www.youtube.com/watch?v=hEnvQG m6o00 https://www.youtube.com/watch?v=hEnvQG m6o00

20. VDJ recombination and Ab diversity Heavy chainLight chain V1000300 D15- J44 V x D x JV x J 1000 x 15 x 4300 x 4 = 6 X 10 4 1.2 x 10 3 X = 7.2 x 10 7 One DNA sequence – millions of Abs

21. Lymphatic system

22. What is the lymphatic system ? Part of the circulatory system Removal of interstitial fluid

23. Lymphatic system: What is it made of ? Lymphatic vessels Lymph nodes Lymphatic tissue Organs

24. Primary lymphoid organs Bone marrow: Makes all the blood cells including lymphocytes Removes auto-reactive (B cells that recognize self) B- lymphocytes Thymus: Removal of auto-reactive (T cells that recognize self) T- lymphocytes

25. Removal of auto-reactive B cells in the bone marrow Mature B-cells Immature B-cells Self-antigens Apoptosis (happens to 90% of the B cells)

26. Removal of auto-reactive T cells in the Thymus Mature T-cells Immature T-cells Self-antigens Apoptosis

27. Immunological tolerance Non-responsiveness of the immune system to “self” antigens or substances capable of eliciting an immune response. This is used by the immune system to discriminate “self” from “non-self”

28. What is the function of the lymphatic system? Lymphatic vessels Removal of excess interstitial fluid Transport lymphocytes Primary Lymphoid organs (BM, Thymus) Production of lymphocytes Removal of auto-reactive lymphocytes Secondary Lymphoid organs (L.nodes, tonsils, spleen..) Collection of lymphocytes (Antigen presenting cells / macrophages..) to fight infection

29. MHC (Major histocompatibility complex)

30. What are MHC ? They are glycoproteins that serve as antigen presenting structures on cell surface Essential part of adaptive immune response

31. MHC class I Present on almost all cells Present antigens to CD8+ T cells (cytotoxic T cells) Infected host cell

32. MHC class II Present on cells of the immune system - eg. macrophages, B cells Present antigens to CD4+ T cells (T helper cells)

33. MHC class I vs MHC class II MHC class IMHC class II ExpressionAlmost all cellsOnly some cells of the immune system: macrophages, B cells etc Antigen presentationTo CD8+ T cellsTo CD4+ T cells NomenclatureA, B, C, D, E, FDP, DQ, DR MHC = HLA or human leukocyte antigen

34. MHC Diversity MHCs are highly polymorphic MHCs are isoantigens MHC I - >1300 variants MHC II - > 700 variants

35. MHC-peptide binding is promiscuous binding There are hundreds of thousands of antigens but only a limited number of MHCs To be able to present all/most antigens – MHCs bind promiscuously to peptides  1 peptide binds to many MHCs  1 MHC binds to several peptides

36. What will happen if we all have the same MHC make-up ? Threat to individual’s survival Possible extinction of population

37. MHC diversity and disease outcomes Impact on individual depends on MHC type Population survives

38. T cells and B cells

39. Lymphocyte subsets T HELPER CELLS (T H Cell or CD4+ T cells) Th CYTOTOXIC T LYMPHOCYTES (T c Cell or CD8+ T cells) CTL Produce antibodies PLASMA CELLS PC T B T CELLS B CELLS CLP Common lymphoid precursor

40. Lymphocyte antigen receptors B The B cell antigen receptor is a membrane-bound antibody SURFACE IMMUNOGLOBULIN (BCR = B cell receptor) T The T cell antigen receptor IS NOT membrane bound antibody but a distinct molecule T CELL receptor (TCR) Each antigen receptor binds to a different antigen Each cell has only one antigen specificity

41. B cell and T cell receptors TCRs also undergo VDJ recombination similar to Abs millions of TCRs

42. T helper cell (T H or CD4+ T cell) activation

43. A closer look at T helper cell (T H or CD4+ T cell) activation APC TCR to MHC on APC – “Are you me ?” - YES TCR to Antigen – “Are you me ?” - NO T H Cell to APC – “Do you have B7 co-stimulatory signal ?” - YES Signal 1 Signal 2 Signal 1 + Signal 2 activate T cell

44. What happens when a T helper cell (T H Cell) is activated ? 1 2 3 (1) Proliferation of T H cell (CD4+ T cell) (2) Proliferation of CTL (CD8+ T cell) (3) Activation of B cell to produce antibody

45. What happens when CD8+ cells (Cytotoxic T cells or CTLs) recognize infected cells ? Cytokines Activated T H cell Activated CTL release perforins Killing of host cells

46. How does B-cell handle a pathogen ?

47. TH cell help to B-cell I can produce antibodies

48. What happens when a B cell is activated ? 2 1 (1) B cells becomes – a plasma cell to produce antibodies (2) B cells becomes – a memory B cell NO Ab Long lived Produces Ab short lived

49. Why do we need memory B cells? Memory B cells Plasma cells Hey. I think I know you. NO Lag time When the body encounters the same pathogen again – it responds very quickly Are there memory T cells – Yes both memory T H cells and memory CTLs( but this will not be discussed) Killing of pathogen in quick time Plasma cells NO Lag time Plasma cells NO Lag time

50. What is the role of antibodies in immunity ? 1)Prevents virus entry into host cell 2)Promotes phagocytosis of pathogens 3)Promotes killing of infected host cells

51. Antibody: (1) Virus neutralization Virus enters the host cell Virus cannot enter the host cell Virus attaches to receptor in host cellAb-bound virus unable to attach to receptor

52. Antibody: (2) opsonisation and phagocytosis Killing of microbe Lysis of microbe by MAC

53. Antibody: (3) antibody-dependent cell- mediated cytotoxicity (ADCC) Killing of microbe

54. Summary: adaptive immune response

55. T regs T regulatory cells (Old term suppressor T cells) – suppress immune response CD4+ (like T helper cells) – but have other surface markers. Prevent autoimmunity Mice without Tregs – die prematurely of autoimmune diseases

56. T regs T regs suppress immune response and prevent auto immune diseases

57. Antibody – class switch T Helper cell B cell activation Plasma cells IgM antibodies Programming switch in B cells – differentiate into plasma cells that make IgG

58. Ab response and class switch Lag phase Log phase Plateau phase Decline phase 1 o Ag 2 o Ag D a y s A f t e r I m m u n i z a t i o n A b T i t e r

59. Ab response and class switch Ab class switch – 1 o - IgM – 2 o - IgG 1 o Ag 2 o Ag Total Ab IgM Ab IgG Ab D a y s A f t e r I m m u n i z a t i o n A b T i t e r IgG antibody circulate in body for years - protect against specific antigens

60. Hypersensitivity INAPPROPRIATE IMMUNE RESPONSE TO AN ANTIGEN

61. Class I hypersensitivity Often treated with anti-histamines

62. Other classes of hypersensitivity Class II-IV Involve IgM, IgG, Ag-Ab complexes, c` activation Not discussed

63. Autoimmunity Immune response against one’s own-self

64. Possible causes of autoimmunity ? 1)Release of Sequestered Antigen 2)Cross-reactive Autoantigens (Molecular Mimicry) 3)Hormones 4)Drugs

65. 1. Sequestered Antigen Some antigens are hidden from the immune system Damage to these organs causing exposure of these sequestered antigens an immune reaction to these antigens may occur. Examples Exposure of Eye lens protein after trauma

66. 2. Cross-reactive or molecular Mimicry Viruses and bacteria may have antigenic determinants that are identical or similar to those of normal host cells e.g. Rheumatic fever - Streptococci and proteins of the heart muscle

67. Molecular mimicry and autoimmunity

68. Ab against streptococci can cause rheumatic heart disease

69. 3. Hormones About 75 percent of autoimmune diseases occur in women, most frequently during the childbearing years. Estrogen (a hormone) is believed to play a role in several autoimmune diseases by modulating immune responses

70. Autoimmune disease affect women more often then men

71. 4. Drugs Drugs = not antigens = no immune response Self-antigens = no immune response Drug + self-antigen = neoantigen = immune response

72. Examples of autoimmune diseases

73. Systemic Lupus Erythematosus (SLE)  AutoAb to nuclei, DNA  Ag-Ab complexes are deposited in kidneys and vascular tissue.  Joint pain, Hemolytic anemia, Kidney dysfunction, skin rashes

74. Multiple Sclerosis MS patients can have autoantibodies which are responsible for the demyelination Mild- numbness of the limbs Severe- Paralysis or loss of vision etc.