1. ADAPTIVE IMMUNE RESPONSE Immune response to infections –1. Innate response Neutrophils, macrophages, NK cells at site of infection –2. Initiation of adaptive response iDCs phagocytose Ag and take to nodes DCs present Ag to T cells –Get specific helper (CD4) T cells »“ help ” B cell activation = humoral immunity »Activate macrophages = CMI –Get specific cytotoxic (CD8) T cells »Kill virally infected cells = CMI

2. ADAPTIVE IMMUNE RESPONSE T CELLS Development: in thymus –Immunogenetic events produce multiple TCRs Similar to B cells TCR gene with multiple V, D, and J segments Recognize 10 15 epitopes –T cells with TCRs that react with self antigens deleted Start with double negative cells: no TCR, CD4 or CD8 Next get double positive cells: TCR, CD4, CD8 Positive selection = weak recognition of MHCI or MHCII Negative selection = if reaction is strong – apoptosis ~2% of cells survive

3. ADAPTIVE IMMUNE RESPONSE T CELLS

4. TCR complex –T cell receptor – similar to Fab of antibodies Alpha and beta Ig-like chains with variable and constant domains Hydrophobic transmembrane regions –CD3 proteins Gamma + epsilon; delta + epsilon, each with one Ig-like domain Transmembrane segment binds to TCR Cytoplasmic domain of each has one ITAM –ITAM = immunoreceptor tyrosine-based activation notif; for signaling –Zeta chains = disulfide-liked homodimer, each protein has 3 ITAM Transmembrane region with aspartate

5. ADAPTIVE IMMUNE RESPONSE T CELLS

6. Co-receptors –CD4 protein binds to MHC-II on APC CD4 T cells = helper T cells MHC-II only on APCs –CD8 protein binds to MHC-I on target cells CD8 T cells = cytotoxic T cells MHC-I on all nucleated cells

7. ADAPTIVE IMMUNE RESPONSE T CELLS Accessory molecules = lymphocyte surface molecule distinct from Ag receptor complex; adhesion or signaling –CD45R, CD28, CD40L Adhesion molecules = tighten interaction with APC –LFA (leukocyte function-associated antigen)-1, CD2 Cytokine receptors: IL-1R, IL-2R, others

8. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Antigen Presentation to T cells –MHC (major histocompatibility complex) 1 all nucleated cells MHC-encoded alpha (hvy) chain and beta 2 (lt) microglobulin Polymorphic alpha 1 and alpha 2 domains for closed binding cleft –agretope Conserved alpha 3 domains = binding site for CD8 Beta 2 interacts noncovalently with alpha 3

9. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Abbas Fig 4-4

10. ADAPTIVE IMMUNE RESPONSE antigen presenting cells –MHC-II = dimer of alpha and beta subunits APCs Both chains MHC encoded Alpha 1 and beta 1 domains variable and form open binding cleft –Most of variability on beta chain –Agretope Alpha 2 and beta 2 folded into Ig domains –B2 Ig domain binds to CD4

11. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Abbas Fig 4-6

12. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Peptide presentation by MHC I molecules –MHC I molecule synthesized in endoplasmic reticulum (ER) –Cytoplasmic protein (including microbial) degraded by proteosome Usually dealing with degraded (misfolded etc) normal proteins –Degraded proteins marked by ubiquitin –To ER via TAP (transporter assoc w Ag processing) –Ag peptide binds to cleft in hvy chain of MHC-I Peptide of 8 or 9 amino acids –MHC-I with peptide to cell membrane via golgi and exocytic vesicle Uninfected cells usually coated with MHC-I with self proteins

13. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Peptide presentation by MHC II molecules –Phagocytized proteins degraded in phagolysosomes by APCs –MHC-II molecule synthesized in ER Invariant chain (Ii) placed in cleft –Ii provides stability and prevents MHC-II from binding with ER peptides –Lysosomal and exocytic vesicles merge –Invariant chain replaced with degraded peptide cleft open at ends; longer peptides (30 aa) presented –Fused vesicle migrates to cell membrane

14. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Antigen processing and presentation – Abbas Fig 5-8

15. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Cross-presentation of Ag –iDCs phagocytise virally infected cells –Some peptides migrate to cytoplasm and processed to MHC I –Also, some cytoplasmic peptides from phagocytised cells processed to MHC II

16. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Cross-presentation of Ag to CD8 T cells. Abbas Fig 5-7

17. ADAPTIVE IMMUNE RESPONSE antigen presenting cells Antigen Presenting Cells (APCs) = Dendritic cells (DCs), B cells, and Macs Dendritic cells = “ only APC that can initiate Ag specific response ” –Presumably from myeloid and lineage –Precursor DCs in blood; differentiate into immature DCs (iDC) –iDC recognized antigen with TLR and other receptors; activated –Pinocytosis/phagocytosis and cytokine production, now DC –DCs can no longer phagocytose; go to T-cell area of lymph nodes where they present Ag to T cells –Langerhan ’ s cells are skin DC

18. ADAPTIVE IMMUNE RESPONSE antigen presentation to T cells T Lymphocytes –Recognize antigens (on dendritic cells) in peripheral lymphoid organs, resulting in expansion of Ag-specific lymphocyte pool –Differentiate into effector cells and memory cells –Effector lymphocytes become activated when they recognize Ag –Activation of T cells requires recognition of Ag displayed on APCs, costimulators, and cytokines produced by both APC and T cell

19. ADAPTIVE IMMUNE RESPONSE antigen presentation to T cells Activation of CD 4 T Cells –MHC II on APC binds with TCR TCR recognizes foreign protein + MHC self proteins –Interaction strengthened by binding of CD4 to MHC II molecule –Signal transmitted via TCR/CD3 complex zeta proteins –Adhesion molecule interactions LFA-3 on APC with CD2 on T cell ICAM-1 on APC with LFA-1 on T cell –Costimulatory signals also required B7 on APC to CD28 on T cell cytokines Partial activation without adequate costimulatory signal = Anergy or apoptosis; method to eliminate self-reacting cells or induce tolerance

21. ADAPTIVE IMMUNE RESPONSE Helper T cell Function TH1 and TH2 CD4 (helper) T cells –TH0 cells mature into TH1 or TH 2 depending on Nature and concentration of antigen How antigen presented Type of APC Cytokines –TH1 = IgM, IgG, activated Macs –TH2 = humoral response; IgA, IgE

22. ADAPTIVE IMMUNE RESPONSE Cytotoxic T cells Activation of CD8 T cells = cytotoxic T cells (CTLs) –Precursors in nodes bind TCR + CD8 to MHC-1 of APC + costim TCR recognizes foreign protein in self MHC molecule –Specific clone expands by ~100,000 –Activated CTLs bind with target cell –Granulysin, granzymes and perforin released from granules = apoptosis –Also interaction of FasL on CTL with Fas on target = apoptosis Apoptosis –Cell DNA and internal membranes fragment –Shrink to “ apoptotic bodies ” which are easily phagocytosed –“ Clean ” cell death as apposed to necrosis

24. HUMORAL IMMUNE RESPONSE HIR = production of antibodies by plasma (B) cells Some Definitions –Albumin, globulin = serum proteins Gamma-globulin = immunoglobulin –Immunogen, antigen –Epitope (linear, conformational) –Hapten; carrier –Adjuvant (alum, Freunds, liposomes, CW components) –Immune tolerance (dose, co-stimulators, transplants ) –T (cell) independent antigen; T dependent Ag –Route of Ag administration –Immunoglobulin superfamily on next slide

26. HUMORAL IMMUNE RESPONSE Immunoglobulin (antibody) structure

27. HUMORAL IMMUNE RESPONSE Immunoglobulin structure –Heavy chain determines class/subclass IgM, G, A, D, E Mu, gamma, alpha, delta, epsilon –Light chain (kappa, lambda) –Domains – constant and variable One variable domain on each lt chain and one variable domain on each hvy chain Three constant domains on hvy chain of IgG and IgA; four on IgM and IgE One constant domain on lt chains Variable domains on hvy/lt chains have 3 complementary determining regions (CDRs) – complements epitope structure

28. HUMORAL IMMUNE RESPONSE Immunoglobulin structure –Fab portion; variable region –Fc portion; C ’ fixatin; bind to Fc receptors on effector cells Fc of IgG and IgA involved with transfer across placenta and mucosa –Hinge region Papain = 2 Fab + Fc; Pepsin = F(ab) 2 Fc –Membrane-spanning portion –Interchain disulfide bonds –Isotype = IgD, IgM, IgG1, IgG2, IgG3, IgG4, IgA1, IgA2, IgE –Allotype = variation of isotype in different individuals –Idiotype = protein sequences in variable region Interacts with epitope

29. HUMORAL IMMUNE RESPONSE Immunoglobulin Classes

30. HUMORAL IMMUNE RESPONSE Membrane-associated antibodies –All isotypes may be membrane-associated on B cells –All membrane-associated AAs monomeric –Last C H region has 26 AAs with hydrophobic side chains Membrane spanning region Secreted antibodies –Found in blood and other extracellular fluids –IgG and IgE are monomeric –IgD negligible –IgM and IgA are multimeric Have tail pieces on end of hvy chains which are bound to J chains

31. HUMORAL IMMUNE RESPONSE Immunoglobulin Isotypes –IgD = <1% of serum Ig membrane Ig on early B-cells –IgM = 5-10% of serum Ig membrane Ig on early B-cells –IgD and IgM are only isotypes that can be expressed together on same cell 5 day half-life in serum Serum IgM pentameric; 10 Ag-binding sites; tail piece and j chain Complement fixation; opsonin; bacteriolysis First isotype after immunization; then wanes

32. HUMORAL IMMUNE RESPONSE Immunoglobulin Isotypes –IgG = 85% of serum Ig 4 subclasses; all monomeric 23 day half-life T cells required Complement fixation; opsonin Maternal IgG transported across placenta and neonatal intestinal epithelium Second isotype after immunization; persists –Also anamnestic (booster) response

33. HUMORAL IMMUNE RESPONSE Immunoglobulin Isotypes –IgA = 5-15% of serum Ig 2 subclasses; both dimer, trimer? with tail piece and J chain IgA transported across epithelial cells has additional secretory component –IgE = <1% of serum Ig Bind to Fc receptors on Mast cells, basophils and eosinophils Immediate hypersensitivity Parasite infections

34. Transport of IgA through Epithelial Cells

35. HUMORAL IMMUNE RESPONSE Neonatal Immunity: Neonates lack ability to mount effective immune response. Therefore, use maternal IgG and IgA –IgG transported across placenta into fetal circulation –IgG and IgA in breast milk ingested IgG and IgA neutralize organisms in gut IgG transported across gut epithelium into circulation –Neonatal Fc receptor for IgG on placta and gut epithelium cells

37. HUMORAL IMMUNE RESPONSE B cell development

38. HUMORAL IMMUNE RESPONSE Immunogenetics –Chromosomes 2, 22, 14 have genes for kappa, lambda, hvy chains –Variable region genes upstream from constant region genes –Genetic recombination events to form variable region of hvy and lt chains: V (variable) and J (joining) segments for light chains –~ 300 V gene segments and ~5 J segments = ~ 1500 possible V, D (diversity), and J segments for hvy chains –300 - 1000 V genes, 12 D genes, and 6 J genes = <100,000 possible –Association of hvy and light chain = > 1,000,000 possible

39. HUMORAL IMMUNE RESPONSE Immunogenetics (continued) Class switching –Genes for constant region of hvy chain in set order IgM, D, G 3, G 1, G 2, G 4, E, A 1, A 2 –Different constant region genes deleted in response to T cell cytokines

40. HUMORAL IMMUNE RESPONSE Production of antibodies by B cell –B cell receptor = Ig molecule specific for non-self antigens Cells with receptors for self antigens removed during development in bone marrow Ig = IgD and IgM on naive B cells –B cell receptor complex IgM or IgD + Ig alpha/Ig beta with immunoreceptor tyrosine- based activity motif (ITAM) Signal transducing receptor initiates activation signal via Ig ITAM –Ultimately activates factors that induce transcription of genes

41. B Cell Antigen Receptor Complex

42. HUMORAL IMMUNE RESPONSE Antibody response to T (cell) independent antigens Surface Ig receptors recognize repeating sites on Ag –Long polysaccharides Get cross-linking of many Ig receptors, activating signal cascade Clonal expansion and plasma cell development of specific B cells Disadvantages of T independent response –No T cell cytokines for class switching; IgM only –Ab has lower affinity for Ag (limited affinity maturation?) –No response in young (<2yr) children

43. HUMORAL IMMUNE RESPONSE Antibody response to T (cell) dependent antigens –Ig receptors on B cell recognize Ag but cross-linking inadequate to activate cell –Therefore need second signal from T helper cell; thus –1) Ag binds to Ig receptor on B cell as above –2) Some bound Ag internalized, processed and presented in MHC-II surface molecule to T cell (which has receptor for this Ag); i.e., B cell functions as APC. T cell, so activated, secretes cytokines which are received by and stimulate the B cell –3) The two signals stimulate the B cell to replicate, clonal expansion A) Plasma cells which produce and secrete the Ig of the receptor –In nodes and bone marrow B) Memory cells with the same receptor –Anamnestic (booster) response

44. HUMORAL IMMUNE RESPONSE Activation of B cells: continued –A second signal may be provided by C ’ receptor –Interaction of B cell with T cell via CD40 (B cell) – CD40L (T cell) –Class switching from influence of helper T cell cytokines –Initial = IgM –INFg = IgG –IL-4 = IgA, IgE –IL-5 = IgA (also induces eosinophil production) –Somatic mutation = increased affinity –Primary and Secondary responses Primary immune response = strong IgM Secondary immune response (second exposure to Ag) = faster, more IgG

45. C ’ in B Cell Activation

46. T Cell - Mediated B Cell Activation

47. HUMORAL IMMUNE RESPONSE Antibodies inactivate microorganisms by –Agglutination –Neutralization Antibody to toxins Antibody to microbial surface molecules that bind to host cells –Opsonization Natural killer cells have receptors for IgG Eosinophils have receptors for IgG, IgA, and IgE –Complement fixation Neutrophils, macrophages have receptors for C3b Gram negative organisms susceptible to MAC