Kuby Immunology: Chapter 13 - Effector Responses: Antibody- and Cell-Mediated Immunity
Antibody-mediated immunity
Basic Structure of Antibodies
- Antibodies are heterodimers.
- Antibodies have a common structure of four peptide chains: two identical light (L) chains and two identical heavy (H) chains.
Basic Structure of Antibodies
- The antibody molecule is divided into two regions: variable region and constant region.
- The antigen binds to the variable region.
- The constant region influences much of the effector function.
Glycosylation:
- Effects antigen binding.
- Effects binding by Fc receptor.
- Increases Ab stability.
Antibody-Mediated Effector Functions
- Two major functions of antibodies:
- Binding foreign antigens encountered by the host.
- Mediating effector functions to neutralize or eliminate foreign invaders.
- Effector functions are responses/actions that result in the removal of the antigen and death of the pathogen.
- The heavy-chain constant region mediates the effector functions of the humoral response.
Antibody-mediated effector functions
- Antibodies mediate the clearance and destruction of pathogens in a variety of ways.
Antibody-Mediated Effector Functions
- Five major effector functions:
- Opsonization
- Complement activation
- Antibody-dependent cell-mediated cytotoxicity (ADCC)
- Transcytosis – movement of macromolecules across the interior of a cell.
- Activation of mast cells
Opsonization
- Opsonin is any molecule that acts as a binding enhancer for the process of phagocytosis.
- Protein molecules called Fc receptors (FcR), which can bind the constant regions of Ig molecules, are present on the surfaces of macrophages and neutrophils.
Complement Activation
- IgM and IgG subclasses can activate the complement system.
- An important byproduct of complement activation is a protein called C3b.
- C3b binds nonspecifically to cell- and antigen-antibody complexes.
- Binding of adherent C3b by macrophages leads to phagocytosis of the cells or molecular complexes that attach to C3b.
- C3b deposition can also lead to the initiation of MAC formation.
Antibody-dependent cell-mediated cytotoxicity (ADCC)
- The antibody acts as a newly acquired receptor enabling the attacking cell to recognize and kill the target cell.
Transcytosis
- Movement of Ig across epithelial layers.
- The capacity to be transported depends on the properties of the constant region.
- In humans, IgA is the major antibody class that undergoes transcytosis.
- Significant amounts of IgG also undergo transcytosis.
- Passive immunization.
Antibody isotypes mediate different effector functions
- The slide exemplifies the various effector functions mediated by different antibody isotypes, including:
- Degranulation (FcεR and IgE)
- Opsonization of bacteria and phagocytosis (FcγR and FcαR and IgG or IgA)
- Maintaining serum levels of antibodies (FcRn and IgG)
- ADCC (FcγR and IgG)
- Transcytosis into secretions (poly-IgR and IgA)
Antibody Classes and Biological Activities
- There are five major classes of antibodies.
- Each class is distinguished by unique amino acid sequences in the heavy-chain constant region that confer class-specific structural and functional properties:
- Order of heavy chain exons: M, D, G3, G1, A1, G2, G4, E, A2
Antibody-mediated effector functions
- Effector functions of IgM Ab:
- First Ab produced in a primary response
- Tend to be lower affinity
- Pentavalent (10 total Ag binding sites)
- Very good at complement fixation leading to MAC formation and target lysis
- Also efficient at forming dense Ab-pathogen complexes that are efficiently engulfed by macrophages
Immunoglobulin M (IgM)
- Accounts for 5 – 10% of the total serum Ig, with an average serum concentration of 1.5 mg/ml.
- The membrane-bound form is monomeric; the secreted form is a pentamer.
- Each pentamer contains an additional Fc-linked polypeptide called the J (joining) chain, which is required for polymerization of the monomers.
Immunoglobulin M (IgM)
- IgM is the first Ig class produced in a primary response to an antigen.
- Has a higher valency than other Ig classes, making it more efficient at binding antigens with repeating epitopes.
- Does not diffuse well; low concentrations in the intercellular tissue fluids.
Antibody-mediated effector functions
- Effector functions of IgG Ab:
- Include several subclasses, each with distinct effector capabilities
- Human IgG1 and IgG3 effective at complement fixation
- Mouse IgG2a and human IgG1 good at mediating ADCC by NK cells
- All variants bind to Fc receptors, enhancing phagocytosis by macrophages
Immunoglobulin G (IgG)
- The most abundant class in serum, constituting ~80%.
- There are four human IgG subclasses:
- The subclasses are encoded by different germ-line CH genes with DNA sequences that are 90 – 95% homologous.
Immunoglobulin G (IgG)
- The structural characteristics that distinguish these subclasses from one another are the size of the hinge region and the number and position of the interchain disulfide bonds between the heavy chains.
- Subtle amino acid changes in the chains affect biological activity.
Immunoglobulin G (IgG)
- IgG1, IgG3, IgG4 – readily cross the placenta
- IgG3 – most effective complement activator
- IgG1 and IgG3 – bind with high affinity to Fc receptors on phagocytic cells
Antibody-mediated effector functions
- Effector functions of IgA Ab:
- Major isotype found in secretions
- Mucus in the gut
- Milk from mammary glands
- Tears
- Saliva
- Effective at neutralizing toxins and pathogens
- Does not fix complement, so does not drive inflammation
- Long half-life in secretions due to protease-resistant amino acid sequence in Fc region X.
Immunoglobulin A (IgA)
- Constitutes 10 – 15% of the total Ig in serum.
- Predominant Ig class in external secretions such as breast milk, saliva, tears, etc.
- Exists primarily as a monomer; polymeric forms are seen.
- Secretory IgA – IgA found in external secretions; consists of a dimer or tetramer, a J-chain polypeptide, and a secretory component. X
Immunoglobulin A (IgA)
- The secretory component is derived from the receptor that is responsible for transporting polymeric IgA across cell membranes.
- The secretory component consists of five Ig-like domains that bind to the Fc region domains of the IgA dimer.
- Poly-Ig receptor – binds polymer IgA and facilitates passage across the epithelium. X
Immunoglobulin A (IgA)
- The secretory component masks sites susceptible to protease cleavage in the hinge region of secretory IgA, allowing the polymeric molecule to exist longer in the protease-rich mucosal environment.
- The Poly-Ig receptor interacts with the J chain of both polymeric IgA and IgM antibodies. X
Immunoglobulin A (IgA)
- IgA can cross-link large antigens with multiple epitopes.
- Binding of secretory IgA to bacterial and viral surface antigens prevents attachment of the pathogens to the mucosal cells, thus inhibiting viral infection and bacterial colonization.
Immunoglobulin A (IgA)
- Complexes of secretory IgA and antigen are easily entrapped in mucus.
- Secretory IgA has been shown to provide an important line of defense against bacteria such as Salmonella, Vibrio cholerae, and Neisseria gonorrhoeae and viruses such as polio, influenza, and reovirus.
- These are pathogens that typically gain entry across mucus membranes (e.g., respiratory, digestive, urogenital)
Immunoglobulin A (IgA)
- The daily production of secretory IgA is greater than that of any other Ig class.
- IgA-secreting plasma cells are concentrated along mucus membrane surfaces. X
Antibody-mediated effector functions
- Effector functions of IgE Ab:
- Best known for role in allergy and asthma
- May also play a role in protection against parasitic helminths (worms) and protozoa
- Made in very small quantities but induce potent effects
- Degranulation of eosinophils and basophils
- Release of molecules such as histamine to damage large pathogens X
Immunoglobulin E (IgE)
- Extremely low average serum concentration of 0.3ug/ml.
- Mediate the immediate hypersensitivity reactions that are responsible for the symptoms of hay fever, asthma, hives, and anaphylactic shock. X
Immunoglobulin E (IgE)
- IgE binds to Fc receptors on the membranes of blood basophils and tissue mast cells.
- Cross-linkage of receptor-bound IgE molecules by antigen (allergen) induces basophils and mast cells to translocate their granules to the plasma membrane and release their contents to the extracellular environment, or degranulation. X
Immunoglobulin E (IgE)
- Localized mast cell degranulation induced by IgE also may release mediators that facilitate a buildup of various cells necessary for an ADCC anti-parasitic defense. X
Immunoglobulin D (IgD)
- First discovered in a patient with multiple myeloma.
- Serum concentration of 30 ug/ml and constitutes about 0.2% of the total Ig in serum.
- IgD, along with IgM, is the major membrane-bound Ig expressed by mature B cells.
- Function is to activate the B cell. X
Antibody-mediated effector functions
- Visualizing antibody and FcR effector responses
- B cells encounter Ag, bind it, and become activated.
- After receiving T-cell help, they differentiate into Ab-secreting plasma cells.
- Plasma cell-secreted Ab is carried to various body sites for isotype-specific effector functions.
- Opsonizing pathogens for phagocytosis
- Activating complement cascades for pathogen lysis
- Enhancing inflammatory activity of neutrophils
- Recruiting cytotoxic cells
- Recruiting and activating NK cells for ADCC killing
Antibody-mediated effector functions
- Fc receptors mediate many effector functions of antibodies
- FcR signaling
- FcγRs
- FcαR
- FcεR
- pIgR
- FcRn X
Antibody-mediated effector functions
- FcR signaling
- Multiple FcRs need to be cross-linked to initiate a signal.
- A signal may be positive (enhancing effector function) or negative (inhibiting effector function).
- The outcome depends on whether the receptor is associated with ITAM or ITIM. X
Antibody-mediated effector functions
- FcγRs
- Most diverse group of FcRs; four families total
- Main mediators of Ab functions in the body
- Expressed by a wide range of cells
- Most are activating receptors (three activating, one inhibiting families)
- Will induce phagocytosis if expressed by macrophages
- Will induce degranulation if expressed by cytotoxic cells X
Antibody-mediated effector functions
- FcαR
- Expressed by myeloid cells
- Monocytes/macrophages
- Granulocytes
- Dendritic cells
- Contributes to pathogen destruction by triggering ADCC and phagocytosis
- Stimulates myeloid cells to release inflammatory cytokines and generate superoxide-free radicals to help kill internalized pathogens X
Antibody-mediated effector functions
- FcεR
- Expressed by granulocytes
- Mast cells/basophils
- Eosinophils
- Two types
- High-affinity FcεRI
- Low-affinity FcεRII (B cells growth and differentiation)
- Triggers a signaling cascade that releases histamines, proteases, and other inflammatory mediators
- Most often associated with allergy symptoms X
Antibody-mediated effector functions
- pIgR
- Polymeric immunoglobulin receptor
- Expressed by epithelial cells
- Initiates transport of IgA and IgM from blood to the lumen (inside) of multiple tissues
- Gastrointestinal tract
- Respiratory tract
- Reproductive tract
- Responsible for carrying Ab into tears and milk and populating gut mucosa with IgA Ab to protect against ingested microbes and toxins X
Antibody-mediated effector functions
- FcRn
- Neonatal Fc receptor
- Related to MHC class I
- Expressed on many different cell types early in an organism’s lifespan
- Epithelial/endothelial cells
- Helps to carry Ab ingested in milk across epithelial cells of the intestine into the bloodstream
- Transfers IgG across the syncytiotrophoblast of the chorionic villi.
- In adults, can help recycle IgG taken up through endothelial cell pinocytosis processes back into the blood
- Cytotoxic effector cells include three subsets
- Each has slightly different killing mechanism triggers, but each induces apoptosis in targets
- Can eliminate infected cells and abnormal tumor cells
- CTLs
- NK T cells
- NK cells
- Cytotoxic T lymphocytes recognize and kill infected or tumor cells via T-cell receptor activation.
- The importance of cross-presentation in CTL activation.
- The best CTL activation is achieved when the APC used can present peptides on both types of MHC molecules.
- Not all cells can do that, though.
- Cross-presentation allows dendritic cells to present antigens on both types of MHC molecules.
- The cell does not have to wait to become infected; infected cells have impaired function.
- Cytotoxic T lymphocytes recognize and kill infected or tumor cells via T-cell receptor activation.
- TC1 and TC2: Two types of effector CTLs
- TC1―secrete IFN-γ, but no IL-4
- Can use perforin and Fas-mediated death induction
- TC2―differentiate in the presence of IL-4; secrete IL-4 and IL-5
- Appear to only use perforin death-induction strategies
- Relatively similar in concept to TH1/TH2 subsets
- Still under investigation
- How CTLs kill cells
- Fas/FasL pathways (a)
- Perforin/granzyme pathways (b)
- Know the function of: Fas, FasL, FADD, caspase-8, caspase-9, caspase-3, granzyme B, perforin, bid, cytochrome c, Apaf-1
- NK cells
- Make up 5–10% of circulating lymphocytes
- Lack specific Ag receptors (no TCR)
- Help to regulate innate/adaptive immunity by cytokine secretion
- Recognize and destroy pathogen-infected cells and abnormal tumor cells
- Proliferate earlier in infection than CTLs
- Phenotype of NK cells
- Lymphoid cells derived from CLPs in the bone marrow
- The thymus is not required for NK development
- Do not undergo receptor gene rearrangements
- Mouse NK cells typically express CD122, NK1.1, CD2, FcγRIII, and CD49b
- Human NK cells lack NK1.1 but express CD56
- The defining trait is the expression of a set of activating and inhibiting NK receptors
- These receptors are used to determine whether to kill a target or not
- NK cells recognize and kill infected and tumor cells by their absence of MHC class I
- How NK cells recognize targets: The missing self-model
- Normal cells present a ligand for the activating (killing) receptor on NK cells AND a ligand for the inhibitory receptor (class I MHC serves as this second ligand)
- When viruses infect cells, some may inhibit MHC class I expression to evade detection and elimination by CTLs
- This makes them a prime target for elimination by NK cells
- How NK cells induce apoptosis of their targets
- Once activating signal molecules are engaged, NK cells use mechanisms very similar to CTLs to induce target cell death
- Release of perforins/granzymes at the junction of two cells
- NK activity “licensing” and regulation
- NK cells don’t automatically possess killing potential
- It seems they’re “licensed to kill” by a prior interaction with a healthy cell through MHC class I/inhibitory receptor interactions
- Important since not all NK cells generated have the ability to recognize MHC Class I.
- This gives the “license” only to those NK cells that can exhibit restraint when encountering a healthy, normal cell
- Still an area of active research for further clarification
- NKT cells bridge innate/adaptive immune systems
- Not NK cells, not quite T cells
- Possess a TCR, but it is invariant
- This TCR recognizes glycolipids presented by nonpolymorphic CD1d
- Can act as helper cells (secreting cytokines) or killer cells
- Killing seems dependent on Fas-FasL interactions
- Include both CD4+ and CD4– cell types
- Don’t form memory cells
- Possess NK surface proteins rather than T-cell varieties