414/514 Immunology Principles 415/515 Immunology Principles and Practice

Antibody Response

Primary Response

• APCs capture, process and present antigen to T helper (Th) cells
  • Dendritic cells and macrophages are much more important numerically as initial antigen-processers than are B cells (which must use their BCRs to interact with antigen before they can process it . . . and there are very few B cells specific for any native antigenic determinant at the onset of an antibody response)
  • ACRs of APCs present antigen fragments bound to class II MHC molecules and B7 on their surfaces
• T cell TCRs bind antigen fragments and MHC-II alpha1- and beta1-chain contact residues, while CD4 binds beta2-domain of MHC-II
• T cells proliferate and differentiate to form large numbers of T helper (Th) and Th memory (Thm) cells
  • Th cells secrete cytokines to regulate antibody responses
  • Thm cells are very long-lived Th cells (half-life of ~40 years in humans) that can respond to processed antigen to help provide a much quicker secondary antibody response
• B cells bind native antigen with their BCRs, then engulf and process it
• B cells interact with Th cells by presenting processed antigen to them
  • Th cell TCRs bind processed antigen fragments bound to class II MHC molecules on B cell surfaces (CD4 also binds MHC-II)
  • Th cells activate genes for CD40 ligand (CD40L), various cytokines (especially IL-2 and IL-4) and their receptors
  • B cells upregulate genes for cytokine receptors (IL-2, IL-4, etc.) as a result of the signal generated by interaction of their constitutively-produced CD40 with Th cells newly-induced CD40L
• T cells secrete cytokines, which trigger B cell proliferation and differentiation into:
  • plasma cells, which produce antibody
    • IgM antibody is transiently produced at first
    • IgA, IgG and IgE antibodies are produced later (depends on the cytokines produced by the Th cells)
  • B memory cells, which are relatively long-lived cells (half-life of ~2 years in humans) that possess IgG, IgA or IgE BCRs can respond to native antigen to provide a much quicker secondary antibody response

Secondary Response

• APCs capture, process and present antigen
  • Dendritic cells are still important APCs in this response
  • B cells are numerically much more significant antigen processers now that their populations have been expanded as a result of the primary response
• T cells interact with APCs, especially in T cell rich areas of lymph nodes (paracortex) and spleen (T cell/marginal zone region of PALs) where they are activated to proliferate and differentiate into cytokine-producing Th cells
• B cells bind native antigen with their BCRs, then engulf and process it
  • BCRs bind native antigen, providing signal 1, which potentiates B cell differentiation
  • B cell co-receptors enhance signal 1 generation when CD21 (CR2) binds C3d attached to antigen that is bound to BCR paratopes
• Activated Th cells interact with B cells that have processed antigen
  • Th cell TCRs bind processed antigen fragments bound to class II MHC molecules on B cell surfaces (CD4 also binds MHC-II)
  • Th cells activate genes for CD40 ligand (CD40L), various cytokines (including IL-2, IL-4, IL-5, IL-6, IL-10, IL-13, interferon-gamma [IFN-gamma], and transforming growth factor-beta [TFG-beta]) as well as activating the genes for receptors that specifically bind these cytokines (especially IL-2 and IL-4)
  • B cells upregulate genes for cytokine receptors (IL-2, IL-4, etc.) and begin to proliferate (become primary B blasts)
• Antibody response maturation occurs as primary B blasts migrate (together with Th cells) into lymphoid follicles, lose their BCRs (become centroblasts), proliferate very rapidly and simultaneously undergo two processes:
  • Isotype (class) switching occurs in the basal dark zone of the lymphoid follicle (which is in the process of developing into a germinal center)
    • Cytokines produced by Th cells trigger Ig constant-region gene rearrangement
      • IgM is produced when IL-4 and IL-5 predominate
      • IgG is produced when IL-4, IL-5, IL-6 and IFN-gamma predominate
      • IgE is produced when IL-4 predominates
      • IgA is produced when IL-5 and TGF-beta predominate
    • Switch sequences (SS) are recognized by RAG1 and RAG2
    • Stem-loop regions are generated, then excised at the SS sites
    • New Ig gene transcriptional units are thus generated
      • Excision of C-mu and C-delta genes results in activation of a transcriptional unit that generates IgG mRNA
      • Excision of C-mu, C-delta and C-gamma genes results in activation of a transcriptional unit that generates IgE mRNA (remember, there are four C-gamma genes in humans, so one or more C-gamma genes must be excised to allow production of the various IgG subclasses)
      • Excision of C-mu, C-delta, C-gamma and C-epsilon genes results in activation of a transcriptional unit that generates IgA mRNA
  • Somatic hypermutation occurs in the basal dark zone of the developing germinal center
    • High-frequency mutation occurs in hypervariable regions of Ig genes
      • heavy and light chain genes both participate
      • generates new paratopes that may bind antigen better, worse or the same as the original paratope
    • Selection of B cells that produce higher-affinity antibodies occurs in the basal light zone of the developing germinal center
      • follicular dendritic cells (FDCs) capture antigen on their surfaces as a result of binding of antigen-antibody complexes to their FcRs
      • centrocytes generated from centroblasts as they stop proliferating and begin expressing their new BCRs can now interact with this antigen, either on the surface of FDCs or in the form of iccosomes ("chunks" of membrane with FcR-associated immune complexes of antigen and antibody)
      • bcl-2 produced due to signal 1 generated when centrocytes bind antigen blocks the apoptosis (programmed cell death) they would otherwise experience (tingible-body macrophages engulf and destroy the apoptotic centrocytes are seen in germinal centers of lymphoid follicles)
      • high-affinity B cells are thus selected for during the maturation phase of the antibody response
    • Ig gene rearrangement can occur in B cells whose somatic hypermutation results in production of antibodies that cannot compete successfully to bind antigen, thus allowing another avenue for competition for antigen leading to selection
• Secondary B blasts
  • Develop as centrocytes migrate into the apical light zone of the developing germinal center
  • Proliferation of these cells leads to clonal expansion of the new high-affinity B cells, assuring that large amounts of antibody (and Bm cells) will be produced
• Two differentiated B cells generate two different versions of Ig heavy chains for different purposes
  • Plasma cells secrete antibody molecules and migrate to medullary region of lymph nodes (or marginal zone of spleenic PALs)
    • heavy and light chain genes are activated to transcribe their respective mRNAs
    • heavy chains are generated by translation of heavy-chain gene mRNA
    • assembled into Ig molecules by association with light chains in the endoplasmic reticulum
    • glycosylated in the Golgi apparatus, then incorporated into vesicles
    • transported to the cytoplasmic membrane
    • secreted by after fusion of the vesicles with the cytoplasmic membrane
  • B memory (Bm) cells generate BCRs by insertion of specialized antibody molecules into their cytoplasmic membranes and migrate to follicular mantle
    • heavy chains are generated by translation of heavy-chain gene mRNA that contains information for transmembrane and cytoplasmic tail portions
    • assembled into Ig molecules by association with light chains in the endoplasmic reticulum
    • glycosylated in the Golgi apparatus, then incorporated into vesicles
    • transported to the cytoplasmic membrane
    • inserted into cytoplasmic membrane as BCRs during membrane fusion of the vesicles
    • Bm cells generally have IgG, IgA or IgE heavy chains in their BCRs, although a few do have IgM heavy chains