Overview of the Immune System for Medics

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This is a presentation containing all notes for exams on the topic on immunology. It is mainly useful for Cambridge Medical students but some summaries may also be helpful for others!

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  • This, my dear, is even better than the first one.
    I just made it to page 35 and got tired.
    Thank god I do not have to study this stuff.
    But - really - you should consider writing a book. Like these small ones from O'Reilly 'xx in a nutshell'. This could be 'The Immune System in a Nutshell'. This would be really helpful to sick people that are educated enough to follow the matter. I followed along quite good - at least I think so ;)

    I hope you did well in your courses, at least you deserve an A+ in Immunology.

    Excellent!

    Peter
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Overview of the Immune System for Medics

  1. 1. The immune system Cambridge University, October 2011 Christiane Riedinger
  2. 2. this presentation is not finished but rather than to wait forever to perfect it, I have decided to upload this versionhttp://thecameracollectors.blogspot.co.uk/p/work-in-progress.html
  3. 3. Why does the immune systemseem so frustratingly complex? Organisms have been affected by pathogens all throughout evolution.Initial simple pathogens could be fought with a simple immune response.Then the pathogens evolved, and so did the immune system, or vice versa. The immune system has been refined by a million-year-long foot-race between pathogens and their host. Knowing this helps to relieve the frustration of having to learn about the immune system in detail.
  4. 4. TOCI. Major divisions of the immune system2. Overview of immune system components3. The innate immune system4. Acute inflammation5. The adaptive immune system6. Hypersensitivity, Tolerance and Transplantation
  5. 5. I. Major divisions of the immune system innate adaptive • broad inherent response to • specific acquired response common pathogens afferent efferent • detection of pathogen • destruction of pathogen • lacking innate: uncontrolled infection, adaptive not deployed • lacking adaptive: infection contained but not cleared • my advice: after learning about the innate and adaptive immune system, forget these distinctions again as they confused me more than they helped me.
  6. 6. I. innate adaptive can initiate immune immune system uses some components system• evolutionarily old • evolutionarily newer• present in all multicellular organisms • present only in vertebrates• present on all surfaces (also internal, i.e. epithelium) • present in lymph nodes and spleen (2* lymphoid organs), can migrate to peripheral tissues• components: phagocytes, degranulating cells, complement, interferon... • components: lymphocytes, antibodies• responds fast/immediately. either since components already there or fastly made • responds slow (days) as specific receptors need to be made• uses a few 100 germline-encoded receptors with broad specificity for • each individual cell has sa somatically generated, structurally unique receptor ofcommon pathogen-associated molecular patterns (PAMPS) random but narrow specificity = not inheritable• <30 recognition molecules • many millions of antigen-recognising structures• always the same response for virus, bacteria, helminth • different responses tailor-made to pathogen that can be repeated• no capacity for memory • response stronger and quicker on repeated exposure• recognises extracellular pathogens • recognises extracellular and intracellular pathogens• does not distinguish between subtype of pathogen • very specific response, can distinguish EC from IC pathogen lack innate: uncontrolled infection lack adaptive: infection contained but not cleared
  7. 7. 2. Overview of Immune System Components 2.1. Cells 2.2. Macromolecules cell-derived white blood cells plasma-derived receptors (cell derived)
  8. 8. stems cell in bone marrow erythrocytes ... Cells of the2.1. immune system leukocytes II leukocytes Iphagocytic celldegranulating cell*cell of adaptive immune sys lymphoblast myeloblast NK/T pro-granulocyte ? B T monocyte plasma effector dendritic macro- mast NK neutrophil eosinophil* basophil* cell T cell phage cell* (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  9. 9. monocyte • present in blood 2.1. Leukocyte functions • mature to tissues where they mature and take residence (long lived), e.g. glial cells, myelomonocytic Kupffer cells, osteoclasts lymphocytes B T NK dendritic cell macrophage** mast cell*** • T-helper cells coordinate AB • most effective at• Syntheses specific and cell-mediated adaptive presenting antigen via • constantly present inimmunoglobulins • kills cells without MHC MHCII • recruit neutrophils tissues, activated by responses at the surface (=non-self!) • secrete IL1, TNF and(antibodies), unique to • decide if body is dealing • picks up antigen in direct injury • also kills a cell coated periphery and transports IL8 (a chemokine)individual cell with EC or IC pathogen with AB to lymph node where it • degranulate rapidly• either surface-bound • IC = cytotoxic T-tell presents it to cells of the • cytokine productionor secreted • EC = T-helper cell adaptive system, initiates • AG processing and• subtype: naive, primed, • AG recognition/presentation, adaptive immune presentation via MHCIImature, plasma, memory, • cell killing and regulation of responses • activated by IgET-cell independent immune responses. • activated by interferon • attract eosinophils • each cell has different MHC- • play role in allergies peptide receptor • important in wound r healing ila sim neutrophil eosinophil* basophil* similar platelet •acute inflammation especially • acute inflammation allergy and parasite. • histamine granules • increasing vascular • basic granules • give rise to mast cells? permeability, • eosinophilia: either allergic or chemotaxis 3rd world inhabitantphagocytic cell • digesting ECM • activated by IL3, 8, adaptive granulocytesdegranulating cell*cell of adaptive immune syssentinel cells ** difference between neutrophils and macrophages (both phagocytes):phagocytic cells: EAT neutrophils are short lived, when they die, they themselves are phagocytosed byactivated by TLR-AG binding and complement, internalise and destroy/digest pathogens macrophages. = PUS!degranulating cells*: POISON WHAT IS TO BIG TO EAT, e.g. wormsreleased premade histamine (e.g. to stimulate GI contractions during worm infestation to expel it) and proteolytic enzymesquickly make and release arachidonic acid derivatives: prostaglandins, leukotrienes, thromboxanealso release HOCl, basic protein, cationic protein. (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  10. 10. more on macrophages**2.1. Leukocyte functions • ROI • NO • cytokines: IL12,TNFamore on NK cells • secrete: • FGF, VEGF, metalloproteinases for tissue remodelling • increased MHC molecules and co-• major player of innate immune system stimulators (enhanced ag-• not sentinel but circulate in blood partially activated ready to respond presentation and induction of• migrate to affected tissue and proliferate adaptive immune response)• activating receptors: respond to type 1 interferons IFNa, b - (virally infected cells)• NK cells themselves release type II interferons IFNg• NKR’s: activating and inhibiting normal: bind MHC on self-cell surface and at the same time response inhibited by inhibitory receptor SELF if no MHC present, then bind activating receptor but not inhibitory receptor MISSING SELF if self-cell stressed, then activating receptors upregulated INDUCED SELF KIR - MHC I receptors, inhibitory and activating, very diverse family KIR and MHC/HLA receptor polymorphisms associated with HIV, HCV and autoimmune diseases• effector functions of NK cells kill by degranulation: perforin causes membrane lysis antibody-dependent cytotoxicity via FcR, can induce apoptosis in target cell cytokine release to activate macrophages (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  11. 11. 2.2. Macromolecules of the immune system / inflammatory mediatorscell-derived★ small molecules • histamine (pre-formed) from mast cells and basophils • serotonin from platelets • NO • oxygen-derived free radicals and other toxic compounds★ peptides / proteins • cytokines - next page ✴ many sub-groups and functions.... • defensins ✴ low mw peptides released during phagocytosis ✴ mainly expressed by neutrophils and specialised epithelial cells in small intestine (paneth ells) ✴ punch holes in pathogen cell wall: intercalate into and disrupt microbial membranes • lysosomal enzymes★ lipid-based • platelet activating factor from inflammatory cells: helps recruit and activate neutrophils and eosinophils • lipid mediators • arachidonic acid metabolites: eicosanoids, prostaglandins, leukotrienes from inflammatory cells
  12. 12. 2.2. Macromolecules of the immune system / inflammatory mediatorscell-derived★ peptides / proteins • cytokines: ✴ like hormones, but not secreted by endocrine organs ✴ peptide messenger molecules, ~25kDa ✴ act locally and distally, paracrine, autocrine (same cell type, back on releasing cell), endocrine ✴ acting over a distance: macrophages secrete colony-stimulating factor G-CSF to increase neutrophil production in bone marrow ✴ all cells of the innate and adaptive immune system secrete and respond to cytokines!!! ✴ intercellular communication ✴ great redundancy and pleiotropism (many molecules same effect, one molecule many effects) ✴ knock out has widespread effects ✴ two main families:haematopoietin and TNF but also structurally distinct molecules ✴ recognition of different pathogens may involve signalling through different receptors, e.g. TNFa by LPS bearing pathogens ✴ cytokine R’s can be Y-kinases • cytokine sub-groups ✴interleukins: initially named so as released by leukocytes IL, affect cell activation and behaviour ✴interferons: antiviral, cell activation ✴ released when cell infected with intracellular pathogen, can be part of innate immune system ✴ dsRNA --> TLR3 --> interferons alpha and beta ✴ activate oligo-adenylate synthase to degrade viral DNA, switch off protein synthesis, stimulate innate and adaptive immune sys ✴ tumour necrosis factors: diverse inflammatory functions ✴ chemokines = cytokines that induce chemotaxis, e.g. CXCL8 (cysteine-X-cysteine in N-terminus, formerly IL-8 all structurally related, receptors are GPCRs, can also be produced by the pathogens themselves ✴ monokines, lymphokines, colony stimulating factors, GFs
  13. 13. 2.2. Macromolecules of the immune system / inflammatory mediatorscell-derived★ peptides / proteins • cytokines: Acute phase response: • Systemic response to infection • Macrophages release IL-1, IL-6 and TNF • liver produces acute-phase serum proteins, e.g. CRP • CRP binds to pathogens and initiates lectin-pathway of complement activation promotes neutrophil • CRP and mannose-binding lectin enhance complement exit from blood fixation • plasma viscosity increases (ESR rises) • body temperature rises • most severe form: septic shock (Again, this figure was from the web but is actually from Janeway’s Immunobiology.)
  14. 14. 2.2. Macromolecules of the immune system / inflammatory mediators • made by B and T IL-8/CXCL8 TNF-b• cytokines: • chemotaxis • apoptosis, anti-inflammatory IFN-a • primes cells • activates RNAse TNF-a • systemic • activated by dsRNA • pyrogen IL-2 neutrophils IFN-b • emigration • early inflammation • T-cells • acute inflammation • inhibits cellular proliferation • systemic • B-cells • inhibits protein synthesis IL-1 • pyrogen • T-reg • activates NK cells • emigration • NK cells • made by mast cell • upreg MHCI production • chemotaxis IL-4 • from NK cell • from NK cell • upreg. TH2-cells IL-12 IFN-g IL-6 • systemic • TH1 response • upreg MHCI+II • B-cells • T-cells IL-10 • TH1 response • TH2-response • most potent macrophage activator! • inhibits IL-2, IF-g • from TH-cell • inhibits TH-1 response IL-17• tissue damage• delayed-type reactions• synergistic with TNFa and IL-1 B, plasma CD4+ T • IL-2 NK macrophage • IF-g • IL-12 neutrophil early response TH1 - IC TH1 Treg • IC path. • IFN-g TH2 • IL-2 • EC path. CD8+ T • TNF-a • IL-4 (early) TH2 - EC • activates • IL-10 CD8+ T • B-cells (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  15. 15. 2.2. macromolecules of the immune system / inflammatory mediatorsplasma-derived★ peptides / proteins • complement - see next page • clotting / coagulation + fibrinolytic makes thrombin which produces insoluble strands of fibrin to make a clot, fibrinolytic to break down the clot • kinins e.g. bradykinin (vasodilator, induces pain), activated by factor XII (hageman factor)cell-surface receptors • Toll-like receptors, part of innate immune system • T-cell receptor, part of adaptive immune system • immunoglobulins (attached as B-cell receptor or free as antibodies), part of adaptive immune system • more in part 3!!!
  16. 16. 2.2. macromolecules of the immune system / inflammatory mediators CRP binds to phosphocholine on• complement spontaneous breakdown of C3 mannose-binding lectin (MBL) lipopolysaccharides, then inhibited by healthy self but not binding to mannose binds C1 which binds to IgM/IgG✴ contained in plasma that complements antibody action by foreign pathogens recruits MASP proteases complexed with AG =AB-dependent✴ nine/~30 liver-derived plasma proteins C1-C9 which are split intoabout 20 cleavage products✴ many involved in proteolytic cascades => amplification!✴ increase vascular permeability, chemotaxis, opsonisation and directlysis of organisms✴ C3 - central protein with internal thioester✴ activated after confo change, cleaved to C3a and b: C3a = smaller, vasc perm and anaphylatoxin C3b = larger, v reactive, fixed to bact. surfaces = opsonin cleaves C5, C5b then causes cell lysis✴ anaphylatoxins C3a and C5a: (wikipedia) Anaphylatoxins are able to trigger degranulation (release of substances) of endothelial cells, mast cells or phagocytes, which produce a local inflammatory response. If the degranulation is widespread, it can cause a shock-like syndrome similar to that of an allergic reaction.Anaphylatoxins indirectly mediate: • smooth muscle cells contraction, for example bronchospasms • increase in the permeability of blood capillaries • chemotaxis — receptor-mediated movement of leukocytes in the direction of the increasing concentration of anaphylatoxins✴ three activation pathways: alternative pathway - acts first lectin pathway - acts second, MBL=collectin (acute phase) classical pathway - CRP (acute phase)✴three functions / outcomes: activation of inflammation: C3a and C5a, increase vascular permeability and are chemotactic for neutrophils and macrophages C1r and C1s = serine proteases of classic complement pathway, cleave C3, opsonisation of microbes: require antigen-antibody complexes for their activation , C1Q surfactant protein A C3b and C3bi, opsonise lysis of target cells: C5b-9 forming MAC (membrane attack complex) causing for process of phagocytosis of opsonised antigen, see 5. cell lysis. C9 polymerases on the C5b678 complex = channel (Again, this figure was from the web but is actually from Janeway’s Immunobiology.) deficiency of any of these components leads to neisseria infections human cells are protected from MAC by CD59 which inhibits C9 lacking CD59: paroxysmal nocturnal haemoglobinuria where red blood cells are lysed by complement.
  17. 17. 3. The innate immune system • natural barriers of the body to keep pathogens out: external and internal bodily surfaces, epithelia, mucus, lactic acid and other microbicidal substances • pre-programmed (i.e. inherited, evolved) response to common pathogenes: using bactericidal enzymes, phagocytosis, complement, NK cells (from handout) • brings about inflammatory reaction • deploys adaptive immune sys • components: ✴ physical barriers ✴ macrophages / phagocytes ✴ neutrophils, eosinophils, mast cells, NK cells ✴ complement ✴ cytokines • 5 stages: 1. recognition 2. recruitment of cells (formation of acute inflammatory exudate) 3. elimination 4. resolution 5. (if elimination incomplete) induction of adaptive immunity
  18. 18. 3. Receptors of the innate system • =PRRs, pattern-recognition receptors • bind common non-self structures, i.e. PAMPS • presented on: macrophages, dendritic cells, B cells • since encoded in germline, identical receptors on all cells of given type • if receptor triggered, response immediately • PAMP-recognising domains: leu-rock, Ca2+ dependent lectin domains (recognise CH) collectin: collagenous domain linked to the ca2+-dependent lectin domain = collagen + lectin • types: (1) secreted: bind to microbial cell wall and act as opsonins (2) endocytic: on surface of phagocytes, mediate pathogen delivery to lysosomes (3) signaling: activate signal-transduction pathways, induce expression of response genes, e.g. inflammatory cytokines, e.g. TLRs
  19. 19. 3. Receptors of the innate system • subtype: Toll-like receptors (TLRs) ✴ TLR’s are receptors of the innate immune system ✴ recognise microbial products ✴ each TLR is specific for a different set of microbial products ✴ humans have te///n different TLR genes ✴ TLR4 = receptor for lipopolysaccharide on gram- bacteria, leads to cytokine production ✴ original toll id’d in Drosophila (developmental function) ✴ humans: toll-like receptor ✴ activation leads to NF-kappaB induction or MAPK pathway in sentinel cells ✴ NF-kB induction activates the phagocytic process TLR1 lipopeptides (bacteria) TLR2/6 lipoteichoic acid (gram+) and zymosan (yeast/fungi) TLR3 sense dsRNA from viruses TLR4 + LPS on gram- TLR5 flagellin (motile bacteria) TLR7 ss viral RNA (HIV) TLR8 ss viral RNA (influenza) TLR9 un-methylated CpG-rich DNA (bacteria/viruses) TLR10 unknown (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  20. 20. 3. PRRs recognise pathogen-associated molecular patterns main features of PAMPS • only present on foreign hosts • usually essential for pathogen survival or its pathogenicety • usually an invariant structure (such as bact. lipopolysacharride) • many are carbohydrates • shared by different classes of organisms • molecular characteristics essential for survival • highly conserved • not present in vertebrate hosts • how innate immune system distinguishes self from non-self common PAMPS • bacterial lipopolysaccharide (common to all gram-neg bacteria) • ds RNA • peptidoglycans • lipoteichoic acid • mannans • bacterial dna • glucans
  21. 21. 4.5. Acute inflammation• ideally localised but can become systemic• exact type of response is pathogen-dependent!!!• = rapid coordinated response to infection or tissue injury• injury: necrosis where certain self-proteins e.g. HSPs are recognised as pamps = sterile inflammation!• main features: main events mediated by: rubor calor tumour dolor vascular changes!!! more blood more blood oedema nociceptor activation• distinguish: acute inflammation chronic inflammation • short duration (few mins-few days) • months to years • localised (ideally) can become systemic or chronic • lymphocytes, plasma cells, macrophages • PAMPS recognised • resident macrophages secrete IL-1, TNF, chemokines • increased blood flow / permeability • recruit neutrophils (arrive first) and monocytes => macrophages (arrive later) • pyogenic = pus! • systemic: too much vasodil, BP way down • then AG to nodes, adaptive induced, lymphadenopathy • AB good to terminate infection and prevent re-infection • histologically: white cells and oedema
  22. 22. 4. Acute inflammation★ dilatation of blood vessels (rubor, calor)★ increased permeability of vessel wall fluid and proteins escape into tissues (tumor=oedema!) most important protein: fibrin, which forms insoluble strands providing a web to exudate = filtered fluid from circulatory system into lesions contain infection and area of inflammation transudate = extravascular fluid of low protein content★ leukocyte emigration (see next page: margination - adhesion - diapedesis) exit between endothelial cells to site of injury neutrophils, later other cells, e.g. monocytes, depending on type of injury★ chemotaxis to destination (see later) E vents in innate immunity★ phagocytosis and degranulation (see later)
  23. 23. 4. Acute inflammationmargination - adhesion - diapedesis - chemotaxis - phagocytosis/degranulationLeukocyte recruitment• cell injury recruits neutrophils and macrophages from the bloodstream• used by neutrophils, basophils, eosinophils, macrophages and lymphocytes (erys pass passively)• margination / adhesion: “rolling” / weak tethering leukocytes slow down in blood, line up along endothelium: IL-1+TNF by macrophages cause upregulation of E-selectin expression, thrombin and mast cell products (histamine) cause upregulation of P-selectin Selectins bind to glycoproteins (Sialyl-Lex, Lewis X or A) on leukocyte surface => leuko rolls along endothelium Chemokines causes integrin activation in the leukocytes, conversion to high-affinity state ICAM-1(Ig superfamily) binds LFA-1 (integrin) => leukocyte is arrested main types of adhesion molecules: integrins, IG superfamily, selectin, adherins• emigration / diapedesis endothelial cells contract to make vessel walls more leaky loss of tight junctions by reorganisation of the cytoskeleton proteins also diffuse out and bring water with them ==> oedema main proteins: fibrin=>fibrinogen=web for cells to migrate along, complement
  24. 24. 4. Acute inflammation• emigration / chemotaxis: chemokines stimulate migration through the inter-endothelial space (CXCL8) ICAM-1 binding causes pseudopodia-formation to push between endothelial cells leukos release proteases to digest basement membrane and reach extravascular space now move towards target by chemotaxis (bacterial endotoxins/LPS, C5a, prostaglandins, leukotrienes, IL-8/CXCL8), chemokine binds GPCR on leuko causing actin repol/depolymerisation = movement lymphos also migrate similarly, but to specific tissues, i.e. lymph, as naive cells bound to addressins monocytes arrive later:VCAM-1 is upregulated more slowly and binds to VLA/VAL?-4 on monocytes Neutrophil: PSGL - P-selectin Monocyte: VAL4 - VCAM-1 Lymphocyte: LFA1 - ICAM-1
  25. 25. 4. Acute inflammation - complement Ror Fc R on phagocytes bind opsonised pathogen - R-binding triggers actin assembly and internalisation - pathogen in phagocytic vaculole (phagosome) - phagosome fuses with acidic lysosome to form phagolysosome - pathogen is killed and degraded - difference between neutrophils and macrophages (both phagocytes): Phagocytosis neutrophils are short lived, when they die, they themselves are phagocytosed by macrophages. = PUS! • once at site digest debris and pathogensdocks via complement or Fc receptor opsonised by AB or C3b figure from: http://classes.midlandstech.edu/carterp/Courses/bio225/chap16/lecture3.htm
  26. 26. 4. Acute inflammationPhagocytosis: Opsonins • IgG • C3b • mannose binding lectin • fibronectin • fibrinogen • CRPPhagocytosis: Chemical killing of pathogens • superoxide, hydroxyl radical, hydrogenperoxide • hypochlorous acid, NO2 • lysozyme • lactoferrin (iron-binding protein, inhibits growth as bacteria like iron-rich media) • major basic protein (MBP), cationic from eisinophils • bactericidal permeability increasing protein • low pH • hydrolytic and proteolytic enzymes, phospholipase A2 and plasminogen activator • neutrophil granules: specific and azurophilic (myeloperoxidase)
  27. 27. 4. Acute inflammationAcute phase response: Sepsis:• Systemic response to infection • blood clotting and TNF-a prevent• Macrophages release IL-1, IL-6 and pathogens from entering blood but can failTNF • sepsis = if pathogens enter the blood• liver produces acute-phase serum streamproteins, e.g. CRP • in that case TNFa can trigger catastrophic• CRP binds to pathogens and initiates vasodilatation => septic shocklectin-pathway of complement activation • clotting in capillaries can lead to organ• CRP and mannose-binding lectin failureenhance complement fixation • low bp leads to vessel collapse• plasma viscosity increases (ESR rises)• body temperature rises• most severe form: septic shock
  28. 28. 4. Acute inflammation - Resolution • important switch from damage to repair towards anti-inflammatory mediators • e.g lipoxygenase pathway • macrophages crucial, phagocytose debris • fibroblasts, recruited by macrophage FGF, increase collagen syn and ECM => scar • angiogenesis (pathological state in adults except female reproductive tract) by macrophage VEGF:endothelial cells break of from BM of pre-existing vessels, migrate to site of injury and repair, proliferate, differentiate to form lumen and acquire supporting pericytes • induction of adaptive response!!!! (if innate cannot control the infection) activation of T and B-cells delivery of pathogen to 2* lymphoid organs by dendritic cells direct type of response required (EC pathogen - B-cells, IC pathogens - T-cells)
  29. 29. 5. The adaptive immune system 5.1. Overview of cells of the adaptive system: lymphocytes = T-cells and B-cells 5.2. Receptors of the adaptive immune system: BCR and TCR 5.3. Maturation and activation of lymphocytes
  30. 30. 5.5.1. Overview T-cells B-cells ✴ T-helper ✴ naive ✴ cytotoxic T ✴ primed ✴ TH1 ✴ mature ✴ TH2 ✴ plasma ✴ ... ✴ memory ✴ T-cell independent• adaptive system responds with increasing specificity and speed with each encounter of the pathogen• first encounter: 1* response• 2nd encounter: 2* response, more rapid and powerful• main cells: lymphocytes = T and B cells• generated in bone marrow from haematopietic stem cells via common lymphoid progenitor• B and T-cells are produced, then selected/matured, then activated and able to differentiate further• during maturation they first express their antigen receptors, but as long as they have not yetencountered antigen, they are called “naive” (or “mature”)• T-cells mature in Thymus (1*/central lymphoid organ)• B-cells mature in Bone marrow (1*/central lymphoid organ)• nodes and spleen are peripheral or 2* lymphoid organs• lymphocytes enter nodes from blood through high endothelial venules = specialised endothelium• in 2* lymphoid organs, lymphocytes meet professional antigen presenting cells (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  31. 31. 5.5.1. Overview T-cells • control both antibody and cell-mediated adaptive responses • made in bone marrow but mature in thymus till adolescence • then proliferation in periphery • initially T-cells express both CD4 and CD8, whatever binds first stays and gives survival signal, the other is down-regulated • selection: must recognise either class I via CD8 or class II via CD4 • must not recognise self-peptides ==> negative selection in medulla • encounter dendritic cells that display self-peptides, if bind ==> apoptosis • cortex of thymus: T-cells that bind AG in the absence of danger signal => anergic = permanently dormant • 90% die!!!! • thymus: naive T-helper cells => node where they meet dendritic cell and danger signal => activation and proliferation (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  32. 32. 5.5.1. Overview - B cells • generated in and matured in bone marrow • negative selection if recognise self • naive cell has not yet encountered AG • activated if binds AG and T-helper cells and IL2,4,5,6 as co-stimulatory signal • B and T-cells both need to recognise AG! • then B becomes either plasma or memory B-cells • mature B-cells + AB => somatic hypermutation! (binds less well => apoptosis, better => affinity maturation • plasma B: ER activated, release of AB • memory B: long-lived, survive in germinal centres, can produce rapid 2* AB response with T-cell help • T-cell independent B cells against protein-free (e.g. polysaccharide) pathogen (bacteria with sugar capsules do not cause T-cell response!), do not recombine, cross reaction possible, live in spleen (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  33. 33. 5.2. Receptors of adaptive system 5. • T-cell receptors • immunoglobulins (attached as B-cell receptor or free as antibodies) * from the Cambridge University Immunology handout. • if receptor triggered, response occurs after proliferation • each cell has a different receptor • receptors bind to antigens regardless of their origin: bacterial, environmental or self • when linked to surface linked to kinase cascades • T and B-cell receptors are the only genes to undergo recombination (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  34. 34. 5.2. B-cell receptors and antibodies 5. • B-cell receptors (BCR): recognise parts of entire proteins in their natural conformation • upon activation, B-cells differentiate into plasma cells and secrete AB (soluble version) • recognise 10E11 different antigens (cp. to 10E08 by TCRs) • AB titre = lowest dil that can still bind AG = c and Kd dependent • structure: variable region ✴ Fab = fragment antigen binding hinge region ✴ Fc = fragment crystallisable (constant region) ✴ 4 variable domains contain 3 complimentary determining regions each, bind AG ✴ CDR1, 2, 3 on each variable domain, in total each antibody has 12 CDRs ✴ 10 constant domains ✴ 2 light chains, 2 heavy chains ✴ light chain 1V and C ✴ heavy chain 1V and 3C or 4C if IgM and IgE ✴ different heavy chains determine type of AG, alpha, delta, epsilon, mu ✴ heavy chain determines the function of the ab ✴ hinge region for flexibility ✴ alpha, gamma, delta, epsilon, mu = constant region --> different stalk but same Fab constant region ✴ constant Fc fragment can be recognised by Fc receptors on macrophages, neutrophils, basophils and mast cells and can interact with complement ✴ variation in variable domains due tohttp://www.abcam.com/index.html?pageconfig=resource&rid=11258&pid=11287 • bind epitopes, can be called linear epitopes if they bind consecutive amino acids
  35. 35. 5.2. B-cell receptors and antibodies 5.Generation of antibody diversity •10E11 different specificities! • too much for it to be encoded in the genome • V-regions encoded in gene segments which are assembled at random by somatic DNA recombination to form gene rearrangement • gene rearrangement is common to both B and T-cell receptors (= V(D)J or somatic recombination) • B-cell receptors then undergo 2* modifications to generate even more diversity = somatic hypermutation • Class switching = the B-cell can then switch between different forms of antibody1. Gene rearrangement / V(D)J recombination • Variable, Diverse and Joining gene segments make up the repertoire from which the variable domain can be generated • heavy chain locus has 65 variable gene segments for the variable domain, responsible for variation in CDR1 and CDR2 (located at chromosome 14) * from the Cambridge University • there are two light chain loci kappa and lambda (located at chromosomes 2 and 22) with a total of 70 V segments , responsible for variation in CDR1 and CDR 2 Immunology handout. • the light chain loci contain no diversity segments • the sequences of CDR1 and 2 are inherited • CDR3 contains the most variation and is formed by joining of V-D-J segments, the final result is achieved by combination • order of events: 1st DJ joining, followed by V-DJ joining resulting in the DNA sequence of the variable domain • then the constant domain has to be added • IgM is always made 1st since c(mu) = constant region closest to D and J (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  36. 36. 5.2. B-cell receptors and antibodies 5. 1. Gene rearrangement / V(D)J recombination • recombination is carried out by recombinases • first, the D and J segments are brought together and a piece of DNA between the segments is deleted • then a DNA segment between the DJ and V elements is removed • the CDR3 sequence is generated from the joining of VDJ segments and therefore has the highest variability • CDR1 and 2 segments are upstream and are just made of different combinations of variable segments • the primary RNA transcript will contain VDJ-cmu-cdelta but alternative splicing selects for the mu constant chain • light chain: since there is no D-segment, the V and J segments are joined • the constant region is introduced on the RNA level by splicing • the protein is then directed to the cell’s excretory pathways for transport to the surface and combination with the heavy chain http://en.wikipedia.org/wiki/V%28D%29J_recombination Junctional diversity: variable addition and loss of nucleotides and VDJ junctions - addition of nucleotides by deoxynucleotide transferase TdT (N-nucleotide addition) - addition doe to the recombination mechanism (P-nucleotide addition)allelic exclusion: - deletion of nucleotidesone chromosome will manage to create the intact heavy chain DNA firstand the recombination in the 2nd chromosome is shut down. (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  37. 37. 5.2. B-cell receptors and antibodies 5. 2. Somatic hypermutation and affinity maturation • point mutations are introducted into the heavy and light chain variable regions • antibodies with increased affinity are selected by affinity maturation • B-cell R binds AB and internalises it to display it on the surface via MHC II • this is recognised by T-helper cells which can activate B-cells to produce antibody • only B-cells with the highest affinity for antigen will continue to capture and present antigen to T-helper cells • these cells will receive T-cell help to proliferate 3. Isotope or class switching • B-cells initially produce IgM, then switch to IgG (T-cell CD4/IL-4 induced), then IgA (IL-4/ TGF-b induced), then IgE (IL-/IL13 induced) = CLASS SWITCHING (each class different function) • this is generated by alternative splicing on the RNA level • each heavy chain confers class specific functional and structural properties • some isotypes can be split into subclasses IgG and IgA (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  38. 38. 5.2. B-cell receptors and antibodies 5. IgA ✴ produced later ✴ mucosal antibody ✴ dimer to prevent breakdown ✴ present in breast milk IgD ✴ transient * from the Cambridge University ✴ not made by mature B-cells Immunology handout. IgM IgG IgE ✴ MAIN antibody in blood ✴ 1st AB made: ✴ binds well to cell receptors FcR, ✴ made by immature B-cell opsonin ✴ pentamer ✴ binds to Fc on NK cells ✴ present in blood and intercellular fluid ✴ can diffuse into tissues and across ✴ binds to mast cells ✴ very good at complement activation placenta ✴ if IgE cross-linked by AG (likes many Fc’s close together) ✴ activates phagocytes => degranulation of mast cell ✴ recruits innate immune system, ✴ worm activates mast cells ✴ esp phagocytes and complement ✴ stim. mucus, contraction of gut (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  39. 39. 5.2. B-cell receptors and antibodies 5.• three ways in which antibodies can work: 1. block biological activity of target, e.g. neutralise toxin 2. opsonise pathogens for phagocytosis 3. activate complement which may cause direct lysis or further opsonisation • antibody diversity is generated by several processes 1. different heavy and light chain combinations 2. selection of different VDJ segments in heavy chains 3. selection of different VJ segments in light chains 4. variable addition and loss of nucleotides and VDJ junctions = junctional diversity addition of nucleotides by deoxynucleotide transferase TdT (N-nucleotide addition) addition doe to the recombination mechanism (P-nucleotide addition) deletion of nucleotides 5. somatic hypermutation 6. class switching? • antibody affinity: strength of interaction of single AB and its epitope • antibody avidity: strength of interaction of AB with multiple epitopes, e.g. IgM (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  40. 40. 5.2. Receptors for antibodies 5. • Fc receptors bind the constant region (Fc=stalk) antibodies • they are located on NK cells, macrophages, neutriphils, mast cells • by binding an AB that has AG bound, they link AG to molecules or cells that cause its destruction • they are named by the constant region they bind to, i.e. FceR binds IgE, FcgR binds IgG etc. • there are high (e.g. FcgRI) and low affinity receptors (e.g.FcgRII and III) • high affinity receptors bind monovalent AB-AG complexes • low affinity receptors bind multivalent AB-AG complexes • if a multivalent AB-AG complex draws together several Fc receptors on the surface of an NK cell, degranulation is triggered = antibody dependent cell-mediated cytotoxicity ADCC analogous: IgE binding to helminths and degranulation by eosinophils analogous: IgE and G crosslinking on mast cells and basophils leads to degranulation and cytokine /inflammatory mediator release • receptor binding (also works for complement receptor binding) induces phagocytosis • IgM has no Fc receptor but is recognised by the C3b receptor on red blood cells, if bound, complex is delivered to liver or spleen for removal by macrophages • Fc receptors can also serve to transport antibodies to their destinations, e.g. to mucosal surfaces or across epithelial layers (IgA), or across the placenta (IgM) into the fetal bloodstream * from the Cambridge University Immunology handout.
  41. 41. 5.5.2. T-cell receptor structure and production• recognise MHC complexes on cells bound to antigenic peptides• need to recognise one of 20E08 possible antigens presented on </= 12 different MHC molecules• closely related to the Fab fragment with immunoglobulin fold-like domains (part of immunoglobulin superfamily)• however: monovalent, membrane bound, NO SOMATIC HYPERMUTATION, no effector function (only ag recognition)• made of alpha and beta chains, variable + constant, 2x2 domains: V-alpha - C-alpha, V-beta - C-beta,• constant domain is attached to surfaceT-cell receptor rearrangement• also happens by VDJ rearrangement in variable regions• constant region: 1 gene per alpha and beta chain, no genetic variation• variable regions: 2 genes for alpha, 3 genes for beta --> 10E12 different combinations!• beta chain is made by VDJ recombination (D to J, then V to DJ)• surface expression occurs with surrogate alpha chain pTa, which reaches the surface• the cell then expresses both CD4 and CD8• then the alpha chain is made by V to J recombination and replaces the pTa chain at the surface• now selection can occur in the thymus• enzymes used for recombination: RAG1 and RAG2 http://pathmicro.med.sc.edu/bowers/mhc.htm• non-template P and N nucleotides are added to the VDJ and VJ junctions• primary RNA transcripts undergo splicing to give mRNA• again, CDR3 is the most variable region and makes major contact with the peptide/MHC complex• alpha: 50V, 70J• beta: 57V, 13J, 2D• joined by V(D)J recombinase and terminal deoxynucleutide transferase (possible frameshift)= 3E06 variants===> 1.56x10E13 TCR vs 2.6x10E10 peptide antigens (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  42. 42. 5.2. Different T-cell receptors and their ligands 5.• T-cell receptors (TCR): related to immunoglobulins but recognise peptides in complex with major histocompatibility complex (MHC) displayed at surface of a cell presenting an antigen MCH (immunohistocompatibility complex) = “hotdog”, peptide = “bun”: see next page picks up peptides in cytoplasm and presents them at cell surface• the type of MHC indicates if origin of antigen IC (MHC I) or EC (MHC II) but not if self/non-self T-helper cell: • coordinate the immune response • TCR + CD4 recognise MHC class II on professional antigen- presenting cells • MHC bound to peptides from exogenous proteins, e.g. bacteria that macrophages have eaten: from phagosome to Golgi where they are loaded onto the MHC (more see next page) • peptides are displayed on the surface via MHC • downstream effects: help naive B-cells activate macrophages help CD8 T-cells secrete cytokines Cytotoxic T-cell: • TCR + CD8 recognise MHC class I present on almost every cell of the body • bound to naturally degraded peptides that are displayed on the surface: host protein digested by proteaseome to ER loaded onto MHC class 1 • if cell is infected, e.g. by virus, pathogen peptides are displayed too and cell is killed by cytotoxic T-cell. cytokine release (IFNg) • downstream effects inhibit viral replication activate macrophages kill virus infected cells (cytotoxin), kill tumour cells (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  43. 43. 5.2. Different T-cell receptors and their ligands 5. MCH (immunohistocompatibility complex) / HLAWikipedia: there are two classical pathways by which proteins can be processed and displayed.Phagocytic cells such as macrophages and immature dendritic cells engulf pathogens in a process known as phagocytosis. Pathogen-containing endosomes then fusewith lysosomes and mediate their destruction. Degraded particles are then loaded onto MHC Class II molecules and trafficked to the cell surface.[4]All nucleated cells can also present cytosolic peptides on their class I MHC. This pathway is particularly important during infection by a bacterium or virus, or if thecell was cancerous. In a normal cell, class I MHC continually present self-peptides derived from basal protein turnover and defective ribosomal products.However, during a viral infection or tumor development, peptides loaded onto class I MHC will include those of processed pathogens degraded in the proteasome.MHC class I: large 3-domain alpha and small 1-domain beta chain (b2 microglobulin)3 genes of alpha for class I. binding groove beta sheet and two alpha helicesclass I groove binds N and C terminus of peptides which put restriction on length: 9-10aaMHC class II: alpha and beta chains of same length (2x2 domains)3 genes of alpha and beta for class II.co-dominant --> 12 in total expressedclass II groove open ended 13-25 aathese genes underlie many polymorphisms --> population as a whole can survive a new pathogen if even if not every individual can• MHC binds broad range of peptides•T-cell recognises a combination of the peptide and the MHC, i.e. the entire hotdog• MHC protein is generated from multiple independent loci class I (HLA-A, B, C) and class II (HLA-DP, DQ, DR)• large numbers of variants at each locus ( can be >250!!!), different in 1-50 aa,• allelic polymorphism is pathogen driven and unique to MHC/HLA• MHC loci are inherited as sets of alleles as haplotypes• polymorphisms are concentrated in the peptide binding domain• genes are co-dominantly expressed so each cell will have MHC molecules from the paternal and maternal allele• the more different they are, the more they are of benefit to the individual in terms of recognising pathogenic peptides• the whole population has lots of variants of the HLA genes, this ensures survival at a population level• each MHC molecule can bind a series of peptides with certain conserved features (“anchor residues” = peptide motifs)• HLA A3 XIFXXXXXK, HLA B7 APRXXXXXL, HLA B8 XXKXKXXXI, HLA A68 XTXXXXXXR (1/400k peptides)• high stringency: MHC recognises very few peptides and very small chance that viral peptide will be displayed• low stringency: MHC recognises more peptides, higher chance that viral protein will be displayed• MHC I pathway: proteins are degraded by the proteasome, passed into ER via Transporter associated with AG processing (TAP), in ER loaded onto partially folded MHC, passed through Golgiand follow secretory pathway membrane.• MHC II pathway: MHC associates with invariant chain in the ER to chaperone, block groove and target to endocytic pathway. Then passed through Golgi and follow endocytic pathway (duringwhich invariant chain Ii is replaced by CLIP peptide). In MIIC compartment, CLIP peptides are removed and replaced with antigen peptides derived from endosome. (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  44. 44. 5.2. Different T-cell receptors and their ligands 5. peptide loading compartments known as MIIC compartments from Janeway Immunobiology (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  45. 45. 5.3. Selection, maturation and activation of lymphocytes 5.thymus • central organ for T-cell development and selection • bone marrow precursors enter outer sub-capsular region • progress towards the medulla through cortex where they meet cortical epithelial cells to carry out positive selection (next page) • cells are still in the process of re-arranging their alpha chains • in the medulla, they meet macrophages and dendritic cells to trigger negative selection • macrophages are also involved in removing those thymocytes that don’t survive • mature naive T-cells can not be activated by professional APC cells T-cell selection in thymus • designed to eliminate the harmful and reject the useless • only 1-2% survive!!! 1. by successful b-chain rearrangement 2. by positive selection: ensures that T-cells can recognise MHC unique to T-cells cells tested against MHC/self peptides expressed on cortical epithelial cells in thymus moderate affinity results in selection no affinity results in neglect (=major cause of death) cells become CD4+ if react against MHC II or CD8+ if react against MHC I 3. by negative selection: prevents autoimmunity by deleting autoreactive cells high affinity for MHC results in apoptosis as MHC-peptide complexes in thymus are limited, negative selection continues in periphery carried out by macrophages and dendritic cells (by AIRE the autoimmune regulator inducing apoptosis) B-cell selection in bone marrow • only negative selection if recognise self (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  46. 46. 5.3. Selection, maturation and activation of lymphocytes 5. T-cell activation• two signal hypothesis: 1st signal is TCR engagement and 2nd signal is engagement of co-stimulatory surfacemolecules• if only TCR engagement and no co-stimulation: peripheral tolerance• co-stimulatory signal: CD28 and B7.1(CD80) and B7.2(CD86) on the APC, CD40L and CD40• APC’s meet antigen in the periphery (innate immune system)• they upregulate B7.1 and B7.2 and MHCII and migrate to the nodes where they present the AG to the T-cells• T-cells get to nodes via high endothelial venules (if not activated they return to the blood and cycle over and over again)• if they meet matching peptide-MHCII they proliferate and differentiate in to armed effector T-helper cells• in this way, T-cells with CD8 or CD4 can be activated B-cell activation• naive B-cells bind antigen on follicular dendritic cell and internalise it (1st signal)• follicular dendritic cells are only present in follicles where B-cells form germinal centres• then they present processed antigen via their MHCII complex on the surface• this AG-MHCII complex is similar to the one presented by the APC and can be recognised by armed effector T-helper cell• CD28 and CD40L on the T-cell bind B7 and CD40 on the B-cell (2nd activation signal for the B-cell!)• this also activates the T-cell = B-T cell cooperation• now the B-cell can proliferate and undergo class switching• thymus independent antigens can stimulate naive B-cells without T-cell help, e.g. microbial products (polysaccharides,lipopolysaccharides that can cross-link receptors in the membrane) (The mini-images of the immune cells are part of a larger picture from Janeway’s Immunobiology, 7th edition. I had found the figure by itself on the web at the time.)
  47. 47. 5.3. Selection, maturation and activation of lymphocytes 5. Activation of naive (but mature) lymphocytes in 2* lymphoid system T-cell activated by AG-presentation by professional AG-presenting cell maturation in Thymus ? naive T-cell immature T-cell maturation in affinity maturation Bone marrow naive B-cell immature B-cell B-cells with receptors specific B-cell activated by for antigen meet follicular dendritic cells that have T helper cell trapped antigen in the form of cytokine production AG-C3d-AB complexes for screening by the B-cells (from flesh and bones of immunology) affinity maturation: cells with the highest affinity for antigen are preferentially induced to proliterate
  48. 48. 5.3. Selection, maturation and activation of lymphocytes 5. T-cell activation... there’s even more! • Naive CD4Th0 cells can differentiate into TH1 or TH2 cells depending on the environment during proliferation: The differentiation into THI or TH2 cells is guided by signals of the innate immune system (cytokines from dendritic cells, macrophages and NK cells. Abundant AG and high affinity AG binding and IL12 => TH1, little AG and weak affinity and IL4 => TH2 TH1 produce cytokines that support inflammation and cell-mediated responses, activate macrophages, NK and CTLs, => IC pathogens, cell mediated response, produce IgG2a AB in mice TH2 activate B-cells and AB-dependent immune responses, => EC pathogens, humoral response, IL-4, IL-6, IgG1 AB (IL-4), IgA (IL-5) and IgE (IL-4) in mice, mast cells, eosinophils http://en.wikipedia.org/wiki/Th1_cell#Determination_of_the_effector_T_cell_response
  49. 49. 5.3. Selection, maturation and activation of lymphocytes 5. T-cell activation http://en.wikipedia.org/wiki/File:Lymphocyte_activation.png * from the Cambridge University Immunology handout.
  50. 50. 5.3. Selection, maturation and activation of lymphocytes 5. lymph nodeshttp://php.med.unsw.edu.au/embryology/index.php?title=File:Lymph_node_cartoon_02.jpg
  51. 51. 5.3. Summary of events C.Riedinger The lives of lymphocytes B-cell T-cell Generation Bone marrow Bone marrow Maturation Bone marrow Thymus Gene rearrangement Bone marrow Thymus of receptor Bone marrow Thymus Selection -ve selection only +ve and –ve Result Leave marrow as naïve B-cells Leave thymus as naïve T-cells Lymph nodes: Lymph nodes: • In germinal centre 1. TCR engagement: 1. naïve binds AG presented on TCR/CD4 - MHCII** follicular dendritic cell* 2. Engagement of co-stim molecules: • Then internalise AG CD28 - B7.1/2 (CD80/86) • Present AG on MHCII CD40L - CD40 Activation 2. Non-naïve B-cell meets T-cell: MHCII - TCR B7.1/2 - CD28 CD40 - CD40L Result Activated B-cell Activated CD4 TH0 cell Proliferation • Can now give help to CD8 accompanied by cytotoxic T-cell somatic hypermutation Class-switching • Abundant AG • Little AG • High affinity • Weak affinity AG binding AG binding • IL-12 • IL-4 Downstream functions and differentiation Differentiation to plasma cell, TH1-cell TH2-cell memory B-cell… • Cell-mediated • Humoral • termination of B-cell response response response -ve feedback • IC pathogen • EC pathogen • IFN!, TNF" • IL10 B-cell binds AG with immunoglobulin and FcgRIIB this terminates the B-cell response *Follicular dendritic cell: • NOT derived from bone marrow haematopoetic stem cells, but mesenchymal precursors • This can be downloaded separately on the • LACK MHC class II and express FEW pattern-recognising receptors • Do express complement receptors and Fc!RIIb website!! • They trap antigen opsonised by complement or antibodies • How is the antigen displayed on the surface of the FDCs? ** any professional antigen-presenting cell
  52. 52. 6. Tolerance * from the Cambridge University• response to the host has to be avoided => immune system develops tolerance• danger hypothesis: immune system discriminates dangerous from harmless Immunology handout.• adjuvant: give vaccine along with a substance that increases the host’s response to the antigen, otherwise tolerance may develop?• central tolerance: occurs during lymphocyte development neglect of T-cells with no affinity for MHC/peptides positive selection of T-cells with moderate affinity for MHC/peptides negative selection of T-cells with very high affinity for MHC/peptides (by AIRE the autoimmune regulator inducing apoptosis) B-cells that react to abundant antigens on self cells are eliminated as they develop• peripheral tolerance: occurs after leaving the 1* organs backup as many self-antigens are not present in the thymus or bone marrow, even though transcription factor AIRE turns on many peripheral genes in the thymus 1. ignorance: potentially self-reactive T-cells are not activated, as they have no access to antigens in immunologically privileged sites such as the brain, eye and testis 2. split tolerance: if T-cells have established tolerance then B-cells don’t need to as they won’t get T-cell help. Common for serum proteins. 3. anergy: is a state of non-responsiveness. Induced in T-cells if the 2nd activation signal is absent. The cell then becomes unresponsive. Can also happen to B-cells if AG is not cross-linked at the surface. 4. suppression/regulation: T-reg cells (CD25+ and ILR+, express Fox3p TF = marker!!) suppress the proliferation of naive T-cells responding to autoantigens. Requires cell contact and non-inflammatory cytokines such as IL4, IL10 and TGF-beta T-reg cells are either natural, i.e. educated in thymic selection, or inducible Lack of FoxP3 = profound systemic autimmunity: IPEX syndrome =immune dysregulation, polyendocrinopathy, entropathy, x-linked• factors affecting tolerance: timing, dose of AG, costimulation, location• chimerism: inject bone marrow cells at birth, establish tolerance in thymus, if injected later then T-cells in periphery will kill them before they can get to thymus• special case of tolerance: pregnancy physical barrier to mother’s T-cells trophoblast lacks MHC I expression immunosuppressive factors such as alpha-fetal protein or IDO (indoleamine 2,3, dioxygenase, a tryptophan catabolising enzyme)• experimental tolerance: aerosolised oral MHC peptides lead to tolerance (peptide sniffing), monoclonal ABs against co-receptors (co-receptor blockade)• pathogens evading the immune response: immunologically inert coatings (e.g. hydrophobin by fungal spores), varying surface antigens (e.g. strep pneumoniae, trypanosomes, antigenic drift or shift problem: if pathogen changes antigens during infection with the host or in 2* infection, then immune system does not respond efficiently to new epitopes latency by herpes viruses mechanisms to avoid MHCI presentation: EPB virus protein inhibits its proteasomal degradation (MHCI loss also important in carcinogenesis) superantigens created by staphylococci which bridge MHCII and TCR and avoid specific response
  53. 53. 6. Autoimmunity • immune system attacks host components causing damage • 5% of population suffer from autoimmune diseases • failure of self tolerance • autoimmune T-cells are directed against autoantigens • animal models: spontaneous = result of deliberate inbreeding / genetic susceptibility, induced = external trigger of autoimmunity • immunoregulation: immune system can return to a tolerant state, CD4+ T-cells can protect, or immunosuppressive drugs, or anti TNFalpha antibodies (new) • also involved: newly discovered TH17 cells 1. characteristics of autoimmune diseases - spectrum from systemic (lupus, scleroderma, rheumatoid arthritis) to organ specific (hashimoto’s, grave’s, stomach = pernicious anaemia, TIIDB) 2. mechanisms of autoimmune pathology (3) - no autoimmune diseases mediated by IgE (~type I hypersensitivity) 2.1. direct antibody mediated effects (~type II hypersensitivity): Grave’s disease: activating ABs to TSH R Myestenia gravis: ABs to AchR causing downregulation Rheumatic fever: ABs to tissue Autoimmune haemolytic anaemia: ABs to Rh blood AGs => RBC destruction Goodpasture’s syndrome: ABs to type IV collagen => glomerulonephritis 2.2. Immune-complex mediated effects (~type III hypersensitivity): SLE (systemic lupus erythematosus): anti-cytoplasmic and anti-nuclear (DNA, histones, ribosomes, snRNP, scRNP) auto-ABs. Butterfly rash. Complement depletion. Glomerulonephritis,Vasculitis, arthritis. vasculitis subacute bacterial endocarditis: bacterial antigen causing complex formation and glomerulonephritis immune complexes which are normally cleared in the liver are not removed successfully 2.3. T-cell mediated (cellular immune) effects: (~type IV hypersensitivity): - T1DM: T-cells against beta pancreatic cells - RA: T-cells against synovial joint antigens causing inflammation and destruction - MS: T-cells against myelin basic protein proteolipoprotein leading to degeneration - T-cell mediated datamage, no AB, e.g. CD8+, TNH, macrophages, apoptosis via Fas ligand - as proven by re-injection of T-cells which cause the disease in other animals 3. predisposing factors - MHC/HLA allotypes DR2+ve = 16x increased risk for Goodpasture’s syndrome, betaMHC A57V/S/A = negative to hydrophobic swap can cause pathology - multiple genetic loci = multigenic - endocrine factors: females vs males, thyroid problems more common in females, ankylosing spondylitis in males - environment: twin concordance 20-40%, environment 50%, MHC25%, other 25% 4. initiation of disease - released of sequestered antigen (autoimmune sympathetic ophthalmia) - bypass of T-cell tolerance a) modification: tolerated self-protein is modified by neoAG and no longer tolerated (penicillin binding to RBCs) b) inflammation: activates anergic T-cells c) molecular mimicry: T-cells specific for pathogen cross-react with self-protein
  54. 54. 6. Hypersensitivity • damaging immune responses, overreactions of the immune system1. Type I hypersensitivity (2-30mins) • IgE mediated • antigen to which host has pre-existing IgE • hayfever, eczema, asthma • activation of mast cells via FceRI high affinity receptors leading to degranulation and inflammatory mediator production • IgE can be pre-bound by mast cell and cross-linked by AG • common allergens: pollens, foods, drugs, insect products, animal hair, often the antigens are proteases • 20-30% of the population, raised IgE levels in atopic individuals with multiple allergies • genetic component • can protect against parasites, but can also lead to anaphylaxis • desensitisation: convert TH2 to TH1 responses2. Type II hypersensitivity (5-8h) • IgM or IgG mediated response to modified previously tolerated antigens • haemolytic anaemia, thrombocytopenia • penicillin binds to RBC/platelet surface and renders it antigenic • spleen macrophages pick it up via Fcg receptors or complement lysis, antigen-dependent cellular toxicity • blood transfusion: ABO blood group is the only histocompatibility alloantigen for which pre-existing ABs exist group A (40%) has anti B, group B (11%) has anti A, group AB (4%) has no ABs, group 0 (40%) has anti A and anti B 0 = unmodified antigen to which sugars (A=N-acetylgalactosamine and/or B=terminal galactose) are attached 0 = universal donor, AB = universal recipient • Rhesus reaction: rhesus - mother can develop AGs against rhesus + child, give anti Rh antibody to prevent so that child’s cells in mother circulation are destroyed before humoral response is created.3. Type III hypersensitivity (2-8h) • caused by AB-containing immunocomplexes which cannot be cleared • created by persistant inhalation or infection • if AG is soluble in high quantities, immune complexes with IgG form and are deposited in tissues which are recognised by mast cells via the FcgRIII (low affinity) • result: local inflammatory response causing arthritis or glomerulonephritis • arthus reaction: type III hypersensitivity reaction after subdermal injection • serum sickness: transient immune-complex mediated syndrome caused by passive immunisation, i.e. serum injection into the blood4. Type IV hypersensitivity (24-72h) • T-cell mediated (delayed type hypersensitivity, 24-72h) • TB mantoux test • 10-100x more AG needed than for AB-mediated hypersensitivity • contact hypersensitivity: cutaneous reponses to haptens, e.g. poison ivy, TH1-cell mediated damage via macrophages • A hapten (Halbantigen) is a small molecule that can elicit an immune response only when attached to a large carrier such as a protein; the carrier may be one that also does not elicit an immune response by itself. (In general, only large molecules, infectious agents, or insoluble foreign matter can elicit an immune response in the body.) Once the body has generated antibodies to a hapten- carrier adduct, the small-molecule hapten may also be able to bind to the antibody, but it will usually not initiate an immune response; usually only the hapten-carrier adduct can do this.
  55. 55. 6. Transplantation• interaction between the recipient immune system and the donor MHC results in rejection• there are no universal donors• recognition can take place via ABs or T-cells (remember since this is a mammalian transplant there are no pamps!)• the antigenic substances are either the donor MHC (direct recognition) or donor peptides on self MHC (indirect recognition)• autologous: self to self• syngeneic: between genetically identical organisms• allogeneic: between members of the same species, rejection is faster if exposure to allograft repeated• xenogeneic: between members of different species• rejection mechanisms: 1. Hyperacute rejection - occurs rapidly, within minutes or hours - due to pre-existing AB - damage due to complement activation, fluid leakage, platelet aggregation (cutting off the blood supply in the microvasculature) - takes place in (discordant) xenotransplants, but different antigenic surface CHs can be engineered out - decay accelerating factor DAF activates complement 2. Acute rejection - allotransplants - T-cell recognition of the implanted tissue 2.1. direct recognition of allo MHC - as MHC is highly polymorphic, and will be antigenic irrespective of the loaded peptide - allogeneic cells migrate from donor organ into host lymph nodes where host T-cells are activated and then go back and attack the graft 2.2. indirect recognition of allo-peptide on self-MHC - peptides displayed by self-APCs - peptides called minor/H-antigens, e.g. polymorphic proteins in the population, non-same sex proteins - minor antigen rejection is slower although many can combine to give rapid rejection - “allophylogeneic” transplant rejection: can be a problem in haemopoietic stem cell transplants 3. Chronic rejection - many years after transplant - kidney damage can cause loss by non-immune mechanism• Types of transplants: A. transplants at privileged sites: e.g. cornea, no immune reaction B. vascularised solid organs: kidney, heart, long, liver, pancreas C. haemopoietic stem cell transplants: (formerly bone marrow transplants) from blood, marrow, cord blood. if autologous no immunological problems. in this case graft may reject the host! But this can be useful in leukaemia (GvH rxn)• HLA matching: - stringency depends on which tissue is transplanted - high stringency: haematopoietic stem cells, stringency: kidney, heart, not so important: liver - degree of match has a correlation with long-term survival - precise match is uncommon as there are six polymorphic genes• Immunosuppression: - essential follow up treatment - steroids, cytotoxic drugs (azathipine), immunosuppressive drugs (cyclosporin), others (rapamycin, FK506) - kidney transplant 3 different drugs with immunosuppressive drugs continued indefinitely - novel treatments: ABs blocking cell surface molecules, e.g. anti-CD40L on T-cells, tolerise with Treg cells - very few patients forget to take their immunosuppressive drugs and still survive

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