Basophils and mast cells and their importance in immune responses

1. Basophils and mast cells and their importance in immune responses

2. Mast cells • Mucosal mast cells - in the mucous membranes of respiratory and gastrointestinal tract, produce histamine, serotonin, heparin, tryptase, leukotriene C4 ..., participate in parasitosis and allergy • Connective tissue mast cells - the connective tissue, producing tryptase, chymase, prostaglandinD2 ..., are multiplicated in fibrosis, in parasitosis and allergy are not participating

3. Mast cell functions • Defense against parasitic infections • In pathological circumstances, responsible for the early type of hypersensitivity (immunopathological reaction typeI) • Apply during inflammation, in angiogenesis, in tissue remodeling

4. Mast cell activation Mast cells can be stimulated to degranulate by: • cross-linking of IgE Fc receptors • by anafylatoxins (C3a, C4a, C5a) • direct injury, alcohol, some antibiotics

5. Mast cell activation by cross-linking of IgE Fc receptors • Establishing of multivalent antigen (multicellular parasite) to IgE on highaffinnity Fc receptor for IgE (FcRI) • Aggregation of several molecules FcRI • Initiate mast cell degranulation (cytoplasmic granules mergers with the surface membrane and release their contents) • Activation of arachidonic acid metabolism (leukotriene C4, prostaglandin D2) • Start of production of cytokines (TNF, TGF, IL-4, 5,6 ...)

6. Mast cell activation scheme

7. Secretory products of mast cells • Cytoplasmatic granules: hydrolytic enzymes, proteoglycans (heparin, chondroitin sulphate), biogenic amines (histamine, serotonin) Histamine causes vasodilation, increased vascular permeability, erythema, edema, itching, contraction of bronchial smooth muscle, increases intestinal peristalsis, increased mucus secretion of mucosal glands in the respiratory tract and GIT (helps eliminate the parasite) • Arachidonic acid metabolites(leukotriene C4, prostaglandin D2) • Cytokines(TNF, TGF , IL-4, 5,6 ...)

8. The role of mast cells in development of allergy

9. Basophils • Differentiatefrommyeloidprecursor • They are considered to bethecirculatingformof mast • Receptor equipment, containinggranules, themechanismsofstimulation and functions are very similar to mast cells • They are responsibleforthe emergence ofanaphylacticshock • Basophilactivationmarkers: CD 63, (CD 203)

10. Immune mechanisms of inflammation (Local and systemic reactions)

11. Inflammation *Is a summary of physiological responses to breach the integrity of the organism, leading to protection against infection of damaged sites, localization of damage and healing. * The first signals to the development of inflammatory responses come from mast cells, phagocytes, and the substances released from damaged cells and extracellular components of matter.

12. Localbody's response to inflammationManifestations-pain (dolor), fever (calor), redness (rubor), swelling (tumor) and lossoffunction (funkciolaesa)

14. Localinflammation-increased permeability ofbloodvessels (vasoactiveamines, complementcomponents C3a, C5a, leukotrienes ..., swellingatsiteofinflammation) - increasedexpressionofadhesionmolecules on endothelia- activationofcoagulation, fibrinolytic, kinin and complementsystem- influence oflocal nerve endings (prostaglandins, pain) - changes in temperature (IL-1, IL-6, TNF, prostaglandins)

15. Systemic response to inflammation- depends on theextentofdamage and durationoflocalinflammation- fever(proinflammatorycytokines TNF, IL-1, IFN ;stimulatehypothalamic center ofthermoregulation) - mobilizationoftissuemetabolism- inductionofexpressionofHsp (heat-shock-proteins; function as chaperones) - productionofacutephaseproteins (CRP, SAP, C3, C4; opsonization and complementactivation) by liver afterstimulationwithcytokines (TNF-α, IL-1, IL-6)

16. -increased hepatic synthesis of certain serum transport proteins (ceruloplasmin, transferrin) - increased synthesis of protease inhibitors ( macroglobulin) - leukocytosisSeptic shock - the massive penetration of microorganisms into the bloodstream(TNF) Anaphylactic shock- basophil degranulation and complement activation with allergen (histamine)

17. Repair of damaged tissue - elimination of damaged cells with phagocytes - activation of fibroplastic mechanisms - activation of angiogenesis - regeneration and tissue remodeling

18. Physiologicalmechanismsofregulationoftheimmunesystem

19. Regulation by antigen • Induce immune responses and extinction • Affinity maturation of B lymphocytes • Maintaining immunological memory • Antigenic competition • Threshold density of the complex MHC II-gp Ag on APC

20. Regulation by antibodies • Antibodies competes with the BCR for antigen (negative regulator of B lymphocyte stimulating) • IgG immune complexes bind to the BCR and FcgR on B cells, resulting in blocking activation of B lymphocytes • Regulation via idiotypic network

21. Regulation by cytokines and cellular contact • Interaction APC - T lymphocyte • Interaction TH1 – macrophages • Interaction TH2 - B lymphocytes • Mutual regulation of activity TH1 versus TH2 • Development of leukocyte subpopulations Negative regulation of effector cells: • CTLA-4 - T cell inhibitory receptor, binds ligands CD80 and CD86 • Inhibitory receptors of NK cells • Self-destruction interaction of the apoptotic receptor Fas with ligand FasL on the surface of activated T lymphocytes

22. Interaction of APC with T lymphocyte T cell:TCR - antigen-specific receptor (signal 1)CD4 or CD8 - coreceptor (MHCgp binding)CD 28 - costimulatory receptor (signal 2, binds CD 80, CD 86)CTLA-4 - inhibitory receptor (binds CD 80, CD 86)CD-40LAPC:MHC gp I + antigenic peptideMHC gp II + antigenic peptideCD 80, CD 86 - costimulatory ligandsCD 40

26. Suppressionmediated by T lymphocytes • Mutual negative interaction TH1 and TH2 cytokine-mediated (TH2 lymphocytesproduce IL-4 and IL-10 thatsuppresstheimmune response based on TH1 cells) • Clonaleliminationoranergyof T lymphocytesaftercontactwith antigen on thesurfaceofothercellsthan APC (lackingcostimulatingsignals) • Regulatory T cells (Tr1 CD 4+) help to maintain tolerance to autoantigens

27. Factors influencing the outcome of the immune response The same antigen can induce an active immune response or an active state of tolerance, the result of response depends on many factors: • State of the immune system • Properties of antigen • Dose of antigen • Route of antigen administration

28. Cytokines (Tissue hormones)

29. Cytokines • Regulatory proteins and glycoproteins produced by leukocytes and other cells • Essential regulators of the immune system • Apply also outside the immune system (angiogenesis, tissue regeneration, carcinogenesis, treatment of many brain functions, embryonic development ...) • Cytokines - secreted      - membrane (CD 80, CD86, CD40L, FasL ..)

30. Pleiotropic effect • Operates in a cascade • Cytokine Network • Cytokine system is redundant • Effects of cytokines - autocrine - paracrine - endocrine • Are known as interleukins (exception: TNF, lymphotoxin, TGF, interferons, CSF and growth factors)

31. B cells communicate via cytokines with other inflammatory cells, such as T cells and macrophages

32. Distributionofcytokinesaccordingtheirfunction • Proinflammatorycytokines (IL-1, IL-6,IL- 8,IL- 12,IL- 18, TNF) • Antiinflammatorycytokines (IL-1Ra, IL-4, IL-10, TGF) • Cytokineswiththeactivityofhematopoieticcellsgrowthfactor (IL-2, 3, 4, 5, 6, 7, 9, 11, 14, 15, CSF, SCF, LIF, EPO) • Cytokinesapplying in TH2 humoralimmunity (IL-4, 5, 9, 13) • Cytokinesapplying in the cell-mediatedimmunity TH1 (IL-2, 12, IFN, GM-CSF, lymphotoxin) • Cytokineswith anti-virus effect (IFN-, IFN-, IFN- )

33. Cytokine receptors • Consisting of 2 or 3 subunits • One subunit binds cytokine, other are associated with cytoplasmic signaling molecules (protein kinases) • Signaling subunit is shared by several different cytokine receptors - called receptor family • Signaling through these receptors may lead to proliferation, differentiation, activation of effector mechanisms or blocking the cell cycle and induction of apoptosis

34. HLA system (MHC glycoproteins)

35. MHC glycoproteins class I (Major histocompatibility complex) • ThefunctionofMHCgpIispresentationof peptide fragmentsfrominsidethe cell (which are produced by cell, includingviralpeptidesif are present) on the cell surface to T lymphocytes (cytotoxic CD8+) • Present on allnuclearcellsoftheorganism • 3 isotypesofclassicalhuman MHC gp.(HLA - A,-B,-C) • 3 isotypesofnonclassical MHC gp.(HLA - E,-F,-G; molecule CD1)

36. Structure of MHC gp I • MHC gpclass I consistsoftransmembranechaina and non-covalentlyassociatedb2mikroglobulin • a chain has 3 domains, 2 N-terminal (a1, a2 - bindingsiteforpeptides) and 1 C-terminaldomain (a3 - anchored in thecytoplasmicmembrane, a structuresimilar to imunoglobulin domain) • Bindingof peptide isnecessaryfor a stableconformationofMHCgp and thusensureits long presentation on the cell surface

37. Peptidesbinding to MHCgpI • MHC gp I bindpeptideswith a lengthof 8 to 10 aminoacides • Certain MHC gpmoleculebindspeptidessharingcommonstructuralfeatures - couplingmotif (critical are aminoacidesnearthe end of peptide) • Thebindingofendogenouspeptidesoccurs in theendoplasmicreticulumduringbiosynthesisof MHC gp

38. Peptidesbinding to MHCgpI • After a stringa and b2mikroglobulin create in the ER, foldingintothecorrectconformation and themutualassociation and theassociationofanappropriate peptide, thecomplexisfurtherprocessed in the Golgi apparatus and thenispresented on the cell surface • Linkedpeptidesderivedfromproteinsdegradedproteasome, whichcleavescytoplasmicproteinsfordestruction (labeledwithubiquitin), peptide fragments are transportedintothe ER by specificmembrane pump

39. Peptides binding to MHC gp I

40. Peptides binding to MHC gp I

41. Non-classical MHC gpI • HLA - E,-F,-G; CD1 molecules • Structurally similar to classical MHC gp • Are less polymorphic • There are only on some cells • They specialize in binding of specific ligands

42. HLA-E and HLA-G - occurs on the trophoblast cells • Complexes of HLA-E and HLA-G with peptides are recognized by inhibiting receptors of NK cells and contribute to the tolerance of the fetus in utero • CD1 molecules - bind glycolipids (recognized by NK-T lymphocytes)

43. MHC glycoproteins class II • The function of MHC gpII is the presentation of peptide fragments from protein whitch were engulfed by antigen presenting cell on the cell surface to T lymphocytes (auxiliary CD4) • Occur on the APC (dendritic cells, monocytes, macrophages, B lymphocytes) • 3 isotypes of MHC gpII (DR, DQ, DP)

44. Structure of MHC gp II • MHC gp II consist of 2 non-covalently associated transmembrane subunits a and b • The peptide binding site consists of N-terminal domains a1 and b1 • Binding of peptide is necessary for a stable MHC gp conformation and thus ensure its long presentation on the cell surface

45. Bindingofpeptides to MHC gp II • MHC gpIIbindpeptideswith a lengthof 15 to 35 aminoacides (but possiblylonger - becausethe peptide bindingsiteis open atbothends) • Certain MHC gpmoleculebindspeptidessharingcommonstructuralfeatures - couplingmotif

46. Bindingofpeptides to MHC gp II • After a stringa and b are created in ER, foldintothecorrectconformation and themutualassociated are connectedwithanothertransmembranechaincalled invariant chain, whichblocksthebindingsiteforthe peptide, thiscomplexisfurtherprocessed in the Golgi apparatus, secretoryvesiclesisolatedfrom GA mergewithendosomes, then split the invariant chain and peptide fragmentsfrom cell absorbedproteinsbindintobindingsiteof MHC gp and thecomplexisthenpresented on cell surface

47. Peptides binding to MHC gp II

48. HLA system – genetic background • HLA complexislocalizedon chromosome 6 • Codominant inheritance of HLA ( Individual has 3 cell surfaceisotypesof HLA molecules (HLA-A,-B,-C) mostlyin 2 differentalelicforms )

49. Polymorphismof MHC glycoproteins • For MHC gpistypicalhighpolymorphism (exceptthe non-classical MHC gp) • Polymorphism has a protectivesignificanceatindividual and populationlevel • Ppolymorphism MHC gpcausescomplications in transplantation

50. Carry outduringthetestingbeforetransplantation and in determinationof paternity