Overview of Ig functions

1. Overview of Ig functions Shawn Babiuk

2. Overview of Ig Functions • Neutralization • Opsonization • ADCC • Complement • Mucosal Immunity • Immune Evasion • Review through questions and discussion

3. Humoral Immunity • Antibodies (secreted proteins produced in B cells) • Extracellular • Mediate several immunological functions • X gammaglobulinemia illustrates the importance of antibodies

4. Roitt, Brostoff, Male Fourth EditionImmunology

5. Antigen recognition • Affinity the strength of the binding between an antibodies binding site and a single epitope • Avidity the functional affinity depends on the number of binding sites on the antibody and their ability to react with multiple epitopes

6. Antibody Isotypes • IgG Neutralization, opsinization, classical complement pathway, ADCC, passive immunity • IgM Pentamer activation of the classical complement pathway • IgA Dimer Mucosal immunity • IgE Mast cell degranulation and parasite killing by eosinophils

7. Toxin neutralization • Binding of antibody to a toxin preventing the toxin from receptors • Examples tetanus toxin Abbas and Lichtman 2nd addition Basic Immunology

8. Antibody Function Neutralization Abbas and Lichtman 2nd addition Basic Immunology

9. Neutralization • Not all antibodies can neutralize viruses • There is a difference in binding and neutralization • Example: GP120 HIV antibodies can bind but not neutralize HIV • Why? There are certain epitopes that are neutralizing

10. Methods to detect different antibodies functions • ELISA assays detect antibody antigen binding • Virus neutralization assays detect antibodies that prevent viral infection • These two assays can give different results from identical serum samples

11. Opsinization • Antibodies coat microbes and promote their ingestion by phagocytes • Splenectomy leads to increased susceptability to disseminated infections by encapsulated bacteria • Reason: The spleen contains phagocytes and is the major place where phagocytic clearance of opsinized bacteria take place

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13. Opsinization Abbas and Lichtman 2nd addition Basic Immunology

14. Fc Receptors • FcγRI on Macrophages, neutraphils and eosinophils used for phagocytosis and activation of phagocytosis • FcγRIIA on Macrophages, neutraphils and eosinophils and platlets used for phagocytosis • FcγRIIB on B lymphocytes used for feedback inhibition • FcγRIIIA on NK cells used for ADCC • FcεRI on Mast cells, basophils and eosinophils used for cell activation

15. ADCC Abbas and Lichtman 2nd addition Basic Immunology

16. IgE Mediated killing of parasites Abbas and Lichtman 2nd addition Basic Immunology

17. Complement • A collection of proteins that help in host defence • A sequential sequence of proteolytic cleavage of these proteins lead to the elimination of microbes • Part of the innate immune system, with evolved mechanisms of self/non-self discrimination • Three pathways • Alternative • Classical • Lectin

18. Abbas and Lichtman 2nd addition Basic Immunology

19. Proteins in the early stages of complement (alternative pathway) • C3 -Serum protein in plasma • Low levels of C3 are spontaneously hydrolyzed in plasma to C3b and C3a, where its products are unstable and break down • C3a stimulates inflammation • Factor B broken down to Bb fragment forming C3bBb complex (C3 convertase) • Factor D plasma serine protease that cleaves factor B • Properdin stabilizes the C3bBb convertase • C3bBbC3b is a C5 convertase

20. Proteins in the early stages of complement (classical pathway) • C1(C1q, C1r, C1s) initiates the classical pathway, binds the Fc antibody region protease for C4 and C2 • C4 C4b binds to the surface where antibody is bound, C4a stimuates inflammation • C2 binds to C4 where C2a is cleaved by C1 and forms C4b2a (C3 convertase) • C4b2aC3b is a C5 convertase

21. Late stages of complement activation Abbas and Lichtman 2nd addition Basic Immunology

22. Proteins in late stages of complement activation • C5 C5b initiates assembly of the membrane attack complex (MAC) C5a stimulates inflammation • C6 binds C5b and accepts C7 • C7 component of the MAC inserts in to lipid membranes • C8 component of the MAC initiates binding and polymerization of C9 • C9 component of the MAC polymerizes to form membrane pores

23. Functions of the Complement System • Important in elimination of microbes during innate and adaptive immune responses • Opsinization • Direct killing of microbes • Chemotactic attraction • Processing of immune complexes • Augmentation of localization of antigen to B cells and APCs

24. Abbas and Lichtman 2nd addition Basic Immunology

25. Roitt, Brostoff, Male Fourth EditionImmunology

26. Inherited deficiencies of complement • C3 deficient susceptible to infections early in life usually fatal • C2 and C4 deficient are normal, indicating that deficiency in the classical and lectin pathways can be overcome by the alternative pathway • C9 MAC deficiency increased Neisseria infections

27. Regulation of Complement • Why? To prevent uncontrolled and harmful activation • Host cells express regulatory membrane proteins • Decay accelerating factor (DAF) • Membrane co-factor protein (MCP) • Type 1 complement receptor (CR1)

28. Complement regulatory proteins Abbas and Lichtman 2nd addition Basic Immunology

29. Host plasma complement regulatory proteins • C1 Inhibitor inhibits C1r and C1s serine protease activity • Factor I cleaves C3b and C4b • Factor H causes dissociation of alternative pathway C3 convertase subunits co-factor for factor I • C4 binding protein causes dissociation of classical pathway C3 convertase subunits co-factor for factor I

30. Regulation of Complement Abbas and Lichtman 2nd addition Basic Immunology

31. Complement control • Host cells can be overwhelmed by complement activation • For virus neutralization assays it is therefore important to heat inactivate sera to destroy complement activity

32. Inherited deficiencies of complement regulatory proteins • Deficiency in C1 inhibitor causes hereditary angioneurotic edema a disease characterized by excessive C1 activation causing edema • Deficiency in glycolipid anchor synthesis results in deficiencies in decay accelerating factor and membrane cofactor protein results in uncontrolled complement activation and lysis of erythrocytes

33. Mucosal Immunity • The vast majority of pathogens invade at mucosal surfaces • Blocking pathogens at the site of infection the mucosal surfaces can prevent infection • IgA can be actively transported to and secreted from mucosal surfaces • IgA can be transported from the lamina propria by the poly-Ig receptor on the basal surface of epithelial cells where it is endocytosed and secreted into the lumen where the poly-Ig receptor is cleaved by a protease

34. Neonate immunity • In humans maternal antibodies are actively transported across the placenta (Passive immunity) • In some other animals (cattle) maternal antibodies do not cross the placenta • Antibodies are passively transferred in milk • The presence of maternal antibodies can inhibit immune responses to vaccines

35. Evasion of humoral immunity by microbes • Antigenic variation • Inhibition of complement activation - Neisseria miningitides sialic acid expression inhibits c3 and c5 convertases -Streptococcus M protein blocks C3 binding and C3b binding to complement receptors • Staphylococcal protein A binding FC of ABs • Streptocoocal protein G binding FC of ABs • Resistance to phagocytosis Pneumococcus

36. Discussion Questions • What regions of antibodies are involved in the functions of antibodies?

37. Discussion Questions • How do heavy chain class switching and affinity maturation improve the abilities of antibodies to combat infectious pathogens?

38. Discussion Questions • In what situations does the ability of antibodies to neutralize microbes protect the host from infections?

39. Discussion Questions • How is the complement system activated, and why is it effective against microbes but does not react against host cells and tissues?

40. Discussion Questions • What are the functions of the complement system, and what components of complement mediate these functions?

41. Discussion Questions • How do antibodies prevent infections by ingested and inhaled microbes?

42. Discussion Questions • How do neonatal animals develop the capacity to protect themselves from infections even before their immune systems have reached maturity?

43. Discussion Questions • Why are there so many numerous pathways for Ig function?