Architecture of the primary immune response: Phenotypic and molecular genetic changes in antigen-driven B cell populations

Abstract

Much of our understanding of B cells and the immune response is based primarily upon the study of dissociated lymphocytes. If the observations made in vitro are to be related to events occurring in vivo, it is crucial to study antigen-activated cells in their architectural context. To this end, I have investigated the clonally restricted immune response of {dollar}Igh\sp{lcub}b{rcub}{dollar} mice to immunogenic forms of the hapten, (4-hydroxy-3-nitrophenyl)acetyl (NP) as it occurs in vivo, by analyzing serial spleen sections. Following immunization, two populations of antigen-activated B cells are seen in the spleen; foci and germinal centers (GC). Foci and GC are discrete areas of intense B cell proliferation and are oligoclonal; each founded on average by 1-3 precursor cells. Using reagents specific for the hallmarks of this response and DNA sequence analysis of individual NP-specific foci and GC B cells, I have attempted to detail the architecture, clonal dynamics and somatic diversification of an immune response. Even though foci and GC arise as a common consequence of immunization, these two B cell populations are distinct; they differ in their anatomical location, growth kinetics and phenotype. Foci appear in the T cell-rich periarteriolar lymphoid sheaths in the white pulp, prior to the appearance of GC in the follicles. B cells in foci do not bind PNA; while GC B cells avidly bind this lectin. While isotype switching (IgM {dollar}\to{dollar} IgG) occurs in both foci and GC, somatic mutation is restricted to the GC microenvironment. Somatic mutation is initiated in GC B cells at days 7-8 postimmunization and mutations are introduced into the IgV genes at random in a stepwise manner. Thus foci represent populations of antigen-driven selection among unmutated B cells while GC are populations of selection among mutated sister B cells which presumably give rise to memory B cells. Interestingly, B cells in adjacent foci and GC have a common clonal origin based upon comparison of the third complementarity determining region (CDR) of the H chain suggesting that GC are most likely founded by antigen-activated B cells in neighboring focus. Thus antigen-activated B cells appear to have the potential to follow different differentiation pathways and microenvironmental influences determine the fate of B cells in vivo.