Immunology 911
Step I review approach
• Re-read all of Parham (or Janeway) in meditation • **Review lecture handouts from immunology, with selective review of Parham** • Use a review book only • This lecture only • Count on your eidetic memory
High-Yield (Basic science)*
• Innate immunity – complement – cytokines, (chemokines) – NK cells, phagocytes • Adaptive immunity – macrophages, (other APC) – MHC complex – immunoglobulin (and lymphocyte) diversity and specificity • Antigenic variation, immune system evasion, vaccines *Bhushan et al. First aid for the USMLE Step1. 2001
Innate immunity
• Physical barriers - skin, mucosa, etc. • Innate immune response: recognize certain “pathogenassociated molecular patterns” –bacterial lipopolysaccharides or DNA, mannans –complement, cytokines, chemokines –preprogrammed leukocytes: receptors encoded in germ line phagocytes, macrophages, neutrophils, NK cells
Complement function
• Part of the innate immune system; can also collaborate with adaptive immune system • Functions –binding –opsonization –chemotaxis –activation –lysis
Parham, Fig. 7.29 Commit to memory! Know the sequential steps for classic and alternative activation!
Alternative pathway
• At any given time, some C3 is being spontaneously hydrolyzed • Produces C3a (little piece) and C3b (big piece), an active enzyme
C3
• Activated C3 + factor B or factor D produces a highly unstable complex (half life 0.1 ms) • Binds covalently to nearly any nearby protein or carbohydrate. • Binding stabilizes the complex and can result in activation of the entire complement cascade. • C3b,Bb is the C3 convertase of the alternative pathway
C3
• Deposition of C3b on microbial surfaces is a key step in their annihilation. • Humans and some other animals that use complement for host defense have several inhibitory proteins on cell surfaces • Most pathogens lack this defense system and are quickly “marked for destruction”.
Classical pathway
• Ab can activate complement to assist in an immune response. • IgM best, followed by IgG1 and IgG3, and to a lesser extent IgG2. • IgG4, IgA and IgE do not activate complement. (Why?) • IgM and IgG have an area of their Fc region that can bind C1. In the serum, this doesn’t happen because of 3D structural limitations
Classical pathway
• Ab specifically binds an invading organism • Ab undergoes a structural change that permits C1 binding and activation • Activated C1 then activates the rest of the “classical pathway”
Lectin pathway
• Certain organisms have mannose-containing proteoglycans on their cell surfaces. • Mannose-binding protein, (MBP) in human plasma can bind mannose on one end; it structurally resembles C1q on the other end. • When MBP coats an invader, it can activate the classical pathway.
Chemokines, cytokines
• Certain chemokines and cytokines important in innate immunity. • Enhance innate response, as well as adaptive response
Interferons
• Also part of innate immunity • Virus infected cells produce IFN-a and IFN-b • These help block spread of virus to unaffected cells –resistance to viral replication; induce enzymes to degrade viral mRNA –increased MHC I expression –activate NK cells
NK cells
• Natural killers - large lymphocytes that do their duty without Ag specificity of CD8 cells • Important in viral infections, like Herpes; some tumors • Activated by –infected host cells: IFN-a, IFN-b –macrophages: IL-12, TNFa • Direct killing • Also secrete IFN-g
Phagocytes
Adaptive immunity
• Ag processing, presentation, MHC antigens • APC - T cell interactions • T - B cell interactions
Ag processing
• Antigens taken up by various cells • Processed, presented to T cells • CD8 T cells: cytotoxic; kill cells; immunologic specificity; Ag presented with MHC I • CD4 T cells: helper; Th1 (activate tissue macrophages) or Th2 (B cell antibody response); Ag presented with MHC II • IL-12 production by APCs helps determine Th1 vs. Th2 response
MHC
• HLA I: –presents Ag to CD8 cells –Ags of intracellular origin –gene loci called A, B, C –found on surface of all nucleated cells • HLA II: –presents Ag to CD4 cells –Ags of extracellular origin –gene loci called DP, DQ, DR • HLA typing clinically important
Transduction
• Class switching influenced by CD40-CD40 ligand interaction AND cytokine expression (Th1 vs. Th2) • Th1: IFN-gamma • Th2: IL-4, IL-13
Parts and pieces
• B cell receptor (immunoglobulin) structure and assembly • T cell receptor structure and assembly • Cytokine potpourri
Immunoglobulins
• Soluble form of B cell receptor • Analogous to T cell receptor - assembled much the same way to get diversity • IgG - 4 subclasses • IgA - 2 subclasses • IgM • IgE • IgD
Ig anatomy
• • • •
Constant vs. variable region Heavy vs. light chain Fc vs. Fab (papain) Pepsin cleavage
Constant vs. variable
• • • • •
k locus on chromosome 2 l locus on chromosome 22 Heavy chain locus on chromosome 14 k and l diversity generated by combining V, J, C regions HC diversity by combining VDJC regions
Heavy vs. light
• • • • • Light chain (k or l) does not determine antibody class Heavy chains g, a, m, e, d make the difference Also caused by rearrangement Naïve B cells express IgM and IgD on the surface Ag exposure causes class switching
More Ig words
• Antibody molecules are proteins with epitopes that can be recognized by other antibodies
• Allotype: an Ig epitope that differs among members of the same species • Isotype: an Ig epitope that is specific to a certain class, that is, heavy chain • Idiotype: an Ig epitope located in the hypervariable region - the Ag binding site
Ab function
• Opsonization: Ab coats invader, phagocytic cell recognizes Ab • Neutralization –Ab binds toxin before it does damage –Ab binds invader, preventing adherance • Complement activation –IgM activates complement the best –Enhanced opsonization –Membrane attack complex
Ab function
• IgG: opsonization, fix complement, neutralization, crosses placenta, secondary immune response • IgA: mucosal Ab, does not fix complement, monomer or dimer, secretory component • IgM: fix complement, doesn’t cross placenta, primary immune response • IgE: allergy • IgD: marker for immature B cell; unclear function as soluble Ab
T cells
• T cell receptor looks like an Fab • a chain (chr 14) analogous to Ig light chain (chr 2 or 22) –a diversity generated by combining V, J, C regions • b chain (chr 7) analogous to Ig heavy chain (chr 14) –b diversity by combining VDJC regions
Cytokine soup
• IL-1: produced by macrophages; fever; stimulates T&B cells • TNF-a: produced by macrophages; function similar to IL-1; enhances apoptosis • IL-2: produced by activated CD4 T cells; causes clonal expansion, differentiation • IL-6: causes B cells -> plasma cells; class switch to IgA • IL-3: produced by activated T cells; causes proliferation, differentiation of hematopoietic stem cells
More cytokines
• IL-4: produced by Th2 cells; class switching to IgE • IL-13: similar to IL-4 • IFN-g: produced by Th1 T cells; targets macrophages and lymphocytes; promotes IgG class switching • IL-12: promotes Th1, inhibits Th2 • IL-5: produced by T cells; eosinophil growth, activation
High-Yield (Clinical)*
• • • • • Allergy Immune complex diseases Other immunopathology Immunologic tests Immunodeficiency diseases
*Bhushan et al. First aid for the USMLE Step1. 2001
Immunopathology
• Type 1 (First and Fast): IgE mediated immediate hypersensitivity: rhinitis, asthma, anaphylaxis • Type 2 (Cy-2-toxic): IgG (IgM) mediated cytotoxic reactions: PCN-induced hemolysis • Type 3 (Ag+Ab+C’=3): IgG (IgM) immune complex reactions: serum sickness; Arthus reaction • Type 4 (4th and last): T cell mediated delayed hypersensitivity: contact dermatitis, tuberculin sensitivity
Allergic disease
Sometimes triggered by IgE mechanisms, sometimes not • The atopic diseases (inheritable) – Asthma, rhinitis, atopic dermatitis • Not inherited – Anaphylaxis (insect, drug, food) – Urticaria, pruritus • Not an allergic disease – Contact dermatitis
Allergens
• All allergens are antigens that are recognized by IgE, and most are immunogens • Many allergens can also be recognized by other classes (notably IgG) • An allergenic substance may have several allergens • Major allergen: recognized by most people in large population • Minor allergen: recognized by fewer people (but still important)
IgE
• IgE is homocytotropic • FceRII (“low affinity” receptor) –Ag specific capture, processing, presentation of allergens –B cells, eosinophils, platelets, dendritic cells • FceRI (“high affinity” receptor) –Ag receptor for certain effector cells –Mast cells, basophils
Firing the gun
• Allergic mast cell activation • Release of histamine, leukotrienes • Also tryptase • Recruitment, activation of eosinophils
Preformed in mast cell granule
Synthesized after MC activation
Figure 10.5
Histamine
• Preformed in mast cell, basophil • Via H1 receptor, histamine accounts for most symptoms of acute allergic reactions • Itching, sneezing, dripping • Smooth muscle contraction: bronchospasm • Increased vascular permeability: edema, wheal of ST reaction • Antihistamine = specific H1 receptor antagonist
Leukotrienes
• Once called SRS: synthesized on mast cell activation • Via CysLT1 receptor: inflammation, smooth muscle contraction, mucus secretion • Antileukotrienes = specific CysLT1 receptor antagonists • Inhibitor also available
Eosinophils
• Major effector cells of allergic response • Eosinophilia characteristic of asthma • Also many nonallergic disorders • Many toxic mediators released on activation • Responsible for chronic inflammation of asthma and allergic rhinitis
Preformed
Synthesized
Figure 10.9
Type 2
• Antigens bind surface of RBC or platelet • Specific IgM and IgG antibody bind antigen on the cell surface • Complement activation produces cell death either by lysis or phagocytosis • Often are hemolytic reactions caused by drugs • Classic disease is hemolytic disease of the newborn (Rh incompatibility), PCN hemolytic anemia
Type 3
• Circulating AgAb immune complexes • Complement activation causes tissue damage via several mechanisms • Immune complex size regulated by relative Ag and Ab concentrations • Arthus reaction: Type III reaction in the skin • Serum sickness: Type III reaction in the circulation
Immune complexes
• Immune complexes constantly formed • Range in size from a single protein + single Ab to millions of components • Big immune complexes fix complement and get cleared from the circulation • Smaller immune complexes can get trapped in small blood vessels and activate complement, causing vasculitis or mast cell activation • Immune complex size depends on the amount of Ag and Ab present, as well as whether the Ag is multivalent
Type 4
• A Th1 response mediated by effector T cells - NOT ALLERGY • Processed antigen peptides presented by MHC Class II • Symptoms due to lymphocyte cytokine release, activating inflammatory mechanisms • Occur 1-3 days after antigen contact • Clinical examples: tuberculin reactivity, poison ivy reactions and other types of contact dermatitis, glutin enteropathy
Immunologic tests
• ELISA • Coombs testing • HLA typing
ELISA
• Enzyme-linked immunosorbent assay • Assay for antibody –Ag attached to a carrier –Calibration standards or unknowns added –Excess washed away –Add special detection antibody conjugated to an enzyme, wash away excess –Add substrate - measure color change, fluorescence, etc. • Assay for Ag
Autoantibodies
• ANA (antinuclear antibodies): SLE – Anti-ds DNA: SLE • Anti-histone: drug-induced SLE • Anti-IgM (rheumatoid factor): part of RA • Anti-neutrophil: vasculitis • Anti-centromere: CREST - scleroderma • Anti-mitochondrial: primary biliary cirrhosis • Anti-basement membrane: Goodpasture’s (renal, lung) • Anti-epithelial cell: pemphigus vulgaris • Anti-gliadin (not an autoantibody): celiac disease, dermatitis herpetiformis
Coombs testing
• Detection of antibodies to blood groups • Also called the “antiglobulin test” • Direct method tests for Ab already bound to the surface of an antigen-positive RBC • Indirect method checks either for RBC antigens or serum antibody depending on how it is set up.
HLA Typing
• HLA types were originally defined serologically, and serological reagents routinely used for many years for HLA typing • Many tissue typing laboratories are now using molecular techniques, such as the polymerase chain reaction.
Clinical Relevance
• An individual inherits 2 copies of chromosome 6, one from each parent, and can express as many as 6 different Class I proteins and 6 different Class II proteins. • Expression is codominant. The probability that 2 unrelated individuals will express identical HLA proteins is extremely low.
Clinical relevance
• Strong associations between HLA type and rheumatic, allergic, infectious, and neoplastic diseases • Transplant rejection • Paternity testing, other forensic studies - review how to work through a “mixed up baby” question
Disease associations
• Celiac disease – HLA-DQ2 – (Molecularly: DQA1*0501, DQB1*0201) • Narcolepsy – HLA-DR2, HLA-Dw2 haplotype – (Molecularly: HLA-DQB1*0602 has stronger association) • Ankylosing spondylitis, Reiter’s syndrome – HLA-B27 – (Molecularly: HLA-B*2702, *2704, *2705 have strongest associations; no association with *2706) • Diabetes type 1 – HLA-DR3, HLA-DR4 heterozygote – DQA1*0102-DQB1*0602 haplotype is protective
Transplant rejection
• • • •
Hyperacute: entirely antibody mediated Acute Graft-versus-host Chronic
Hyperacute rejection
• Recipient antibody source –Ab against MHC Class I Ag from previous transplant or blood transfusion –ABO mismatch - recipient isohemagglutinins against endothelial A or B Ag in donor organ • Recipient Ab binds donor Ag, forming immune complexes, activating complement and coagulation pathways • Occurs immediately; not affected by routine (T cell) immunosuppressive therapy
Acute rejection
• Direct recognition –Donor APCs migrate to recipient’s regional lymph nodes and stimulate the recipient’s T cell system • Indirect recognition –Donor antigens (e.g., HLA Ag) taken up by recipient’s APCs, processed and presented to recipient’s T cells • Cytotoxic T cell (CD8+) activation…
And…
• No man is an island… • Nor is any piece of the immune system • Tc’s lead the way, but other parts of immune system also activated • Antibody produced by the recipient (as part of this immune response) can cause a local vasculitis, and… • Persist for a long time, causing a hyperacute rejection episode in a subsequent transplant
GVH
• Donor immunocompetent, recipient immunocompromised • Thus, donor Tc cells attack recipient tissue • Can be big problem in bone marrow transplants • HLA-DR mismatch, or “minor” transplantation antigens
GVH
• Acute (day 6-28) –Rash, diarrhea,jaundice, fever; begins 6-28 days after transplant –Prevented by immunosuppressive therapy (e.g., cyclosporine, methotrexate); T cell depletion of donor marrow • Chronic (after day 100) –Presents like autoimmune (rheumatologic) disorder –Can result in severe immunodeficiency –Treated with prednisone, other immunosuppressives
Chronic rejection
• Months to years after transplantation • Progressive narrowing of arterial lumens - due to growth of endothelial cells • Initially reversible; fibrosis is not treated
Immunodeficiency diseases
• • • • • • • • • • • SCID: severe combined immunodeficiency Thymic aplasia: DiGeorge anomaly Chronic mucocutaneous candidiasis XLA: Bruton’s agammaglobulinemia Common variable immunodeficiency Selective IgA deficiency Wiskott-Aldrich syndrome Ataxia-telangiectasia Chronic granulomatous disease Chédiak-Higashi syndrome Hyper-IgE syndrome (Job’s syndrome)