1. Complement J. Ochotná

2. Complement  humoral component of nonspecific immunity  helps remove microorganisms and own altered cells (apoptotic cells)  complement proteins are synthesized in the liver, less by tissue macrophages and fibroblasts

3.  system of about 30 serum and membrane proteins  complement components are present in serum in inactive form  complement activation has cascade character Complement

4.  the main complement components: C1-C9 (C3 is the central component)  other complement components: factor B, factor D, factor P  regulatory proteins: C1 - inhibitor, factor I, factor H, C4bp, DAF, MCP, CR1, factor S (vitronectin), CD59 (protektin), anaphylatoxin inactivator Complement

5. Biological significance  Opsonization (C3b, C4b)  Chemotaxis (C3a, C5a)  Osmotic lysis (MAC C5b-C9)  Anaphylatoxins (C3a, C4a, C5a)

6. Riedemann N.C. Pathways of complement activation

7. Alternative complement pathway  C3 component of complement rarely spontaneously breaks into C3b and C3a  C3b can covalently bind on the surface of a particle (own cell, microorganism) or reacts with water, thereby inactivates

8.  to bound C3b joins a factor B, which is cleaved by factor D to Ba and Bb, resulting complex C3bBb is stabilized by factor P (properdin) and functions as an alternative C3 convertase Alternative complement pathway

9.  C3 convertase cleaves C3 to C3a (chemotaxis) and C3b, which binds to the surface of the particles (opsonization), or gives rise to other C3 convertases Alternative complement pathway

10.  some C3 convertases form C3bBbC3b that act as an alternative C5 convertase, which cleaves C5 to C5a (chemotaxis) and C5b (starts terminal lytic phase) Alternative complement pathway

11.  C5b starts terminal lytic phase

12. Classical complement pathway  can be initiated by antibodies IgG ( not by IgG4) and IgM or by pentraxins (CRP, SAP - acute phase proteins)  after binding of antibodies to the bacteria surface, Ab changes its conformation and than can bind C1 protein

13.  C1 changes its conformation and acquires proteolytic activity, than cleaves C4 and C2 proteins Classical complement pathway

14.  fragments C4b and C2a bind to the surface of microorganism and create the classic C3 convertase (C4bC2a), which cleaves C3 to C3a and C3b Classical complement pathway Description of C3 convertase has been modified to C4bC2a

15.  then creates a classic C5 convertase (C4bC2aC3b) that cleaves C5 to C5a and C5b Classical complement pathway

16. Lectin complement pathway  is initiated by serum mannose binding lectin (MBL)  MBL binds to mannose residues on the surface of some microbes, after this binding starts cleave C4 and C2  this way is similar to the classical pathway

17. Lectin complement pathway

18. Terminal (lytic) phase of the complement cascade  C5b fragments creates a complex with C6, C7 and C8, the complex dive into the lipid membrane of the cell and attache into a circle 13-18 molecules of C9 => MAC (membrane attack complex), forms a transmembrane channel, which causes osmotic lysis of the target cell (G-bacteria, protozoans, some viruses).  Most microorganisms are resistant to this lytic effect of complement (protection by cell wall).

19. Terminal phase of the complement cascade

20. Regulation of complement and protection of own cells  Activation of complement cascade is controlled by the plasma and membrane inhibitors. MCP DAFProtectin Anaphylatoxin inactivator

21. Complement regulation  C1 inhibitor (C1-INH) – inhibits C1; if missing→ HAE  factor I with cofactors: MCP (membrane cofactor protein), CR1, factor H – C3b, C4b cleavage  DAF (decay-accelerating protein)-degradation of C3 and C5 convertase

22.  factor S (vitronectin) – inhibits complex C5bC6  CD 59 (protectin) - prevents the polymerization of C9  anaphylatoxin inactivator (CPN) - inactivates anafylatoxins (C3a, C4a, C5a) Complement regulation

23. Complement receptors  Bind fragments of complement components  CR1 - on various cells - promotes C3b, C4b decay - stimulate phagocytosis - erythrocyte transport of immunecomplexes  CR2 - on B lymphocytes and FDC - activation of B cells

24.  CR3, CR4 - on phagocytes - participation in opsonization, adhesion  C3aR, C5aR – receptors for anaphylatoxins - mast cell activation Complement receptors

26. 4 basic complement functions  Opsonization (C3b, C4b)  Chemotaxis (C3a, C5a)  Osmotic lysis (MAC C5b-C9)  Anafylatoxins (C3a, C4a, C5a)

27. Complement functions

28. Antigens

29. Antigen (immunogen)  substance which provokes specific immune response  usually proteins or polysaccharides (lipids and nucleic acids only in the combination with proteins or polysaccharides)  molecules >5 kDa the (optimal size of the antigen molecules is about 40 kDa)

30. Hapten  small molecules, that are able to induce specific immune response only after the attachment to the macromolecular carrier  separate haptens are not immunogenic  typically drugs (eg penicillin antibiotics, hydralazin)

31. Epitope (antigenic determinant)  part of the antigen which is recognized by immune system (lymphocytes- BCR, TCR, Ig)  cross-reactive antigens - shares one or more identical or similar epitopes

32. Interaction antigen – antibody  Binding site of antibody ( paratop ) form non-covalent complexes with the corresponding part on antigen molecule ( epitope )  participation: the hydrogen bonds, electrostatic and hydrophobic interactions, van der Waals forces  antigen-antibody complex is reversible

33. Antigen  endogenous antigens - autoantigens (self Ag)  exogenous antigens - foreign substances from the environment  allergen is exoantigen that in the susceptible individuals can cause pathological (allergic) immune response

34. Properties of antigen  immunogenicity proteins> carbohydrates> macromolecule complexes ( glycoproteins, nucleoproteins, and glycolipids )> lipids  specificity

35. Factors affecting immunogenicity  Physical: solubility - insoluble Ag more immunogenic molecular weight - ideal 5-40 kDa  Chemical: structure - the number of determinants degradability - "ease" of uncovering the determinants in antigen presenting cells (APC cell)

36.  Biological: biological heterogeneity genetic and physiological disposition of the body Properties of antigen

37. Degree of foreignness  Autogeneic - antigens of the same individual  Syngeneic - antigens of genetically identical individuals (eg twins)  Allogeneic (alloantigens) - antigens genetically different individuals of the same species  Xenogeneic (heterologous) - antigens derived from individuals of different species (eg monkey kidney transplant man)

38. Types of antigens according to antigen presentation  T- dependent antigens  T- independent antigens

39. Thymus dependent antigens  more frequently, mostly protein Ag  for specific humoral immune response to antigen is necessary assistance of T H lymphocytes (or response isn´t enough effective)  assistance implemented in the form of cytokines produced by T H lymphocytes

40. T- independent antigens  can stimulate B cells directly  mainly bacterial polysaccharides, lipopolysaccharides and polymer forms of proteins (e.g. Haemophilus, Str.pneumoniae)

41. T-independent pathway

42. Superantigens  stimulate T cells polyclonaly and massively (massive cytokine release)  massive activation of T cells can cause shock  e.g. bacterial toxins (Staph.aureus, Str.pyogenes, Pseud.aeruginosa)

43. proteins (microbial products) which have 2 binding sites, one interacts with the epitope presented on all MHCgpII, second interacts with other structures presented in many different TCR molecules (connection of T lymphocyte with APC) Superantigens

44. Differcence between antigen and superantigen binding

45. Sequestered antigens  autoantigens that are normally hidden to immune system and therefore unknow (e.g. brain, the lens of the eye, testes)  if they are "uncovered" by demage, exposed to the immune system, are recognized as foreign (one of the theories of autoimmune processes)

46. Immunologically privileged sites  brain, eye, gonads  are protected from potentially damaging inflammatory immune responses  this tissues are far less rejected in allogeneic transplant (cornea)  this privileged position is not absolute

47. Immunologically privileged sites  Mechanisms of protection from the immune system :  isolation from the immune system (blood-brain barrier)  preferences of Th2 and suppression of Th1-response  production of immunosuppressive cytokines ( TGFβ )  FasL expression -active protection against effector T-lymphocytes  increased expression of membrane complement inhibitors

48. Thank you for your attention