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Naive and memory B cells were isolated from human tonsils and examined for expression of APO-1/Fas and for their sensitivity to the APO-1-dependent apoptosis. APO-1 was found to be constitutively expressed on memory but not on naive B cells. The susceptibility of both cell types to the APO-1 apoptotic pathway was acquired upon CD40 triggering and was correlated with increased expression of the APO-1 receptor. Both naive and memory B cells were protected from the APO-1-mediated death signal after dual ligation of the Ag receptor adn CD40. Our findings suggest that the APO-1 pathway controls the specificity of B cell responses to T-dependent Ags and that occupancy of the Ag receptor dictates the outcome of APO-1-ligation on B cell survival.

It is generally accepted that memory B cells can be defined by their ability to produce, upon antigenic challenge, somatically mutated antibody molecules characterized by an increased affinity and by the expression of a downstream heavy chain isotype. However, the inability to isolate this particular B cell compartment has precluded the study of memory B lymphocyte physiology in man. We previously reported on the identification of an IgD- B cell subset in human tonsils that we defined as CD38- B cells, whose phenotype is highly reminiscent of that of memory B lymphocytes from the splenic marginal zone of rodents. In the present study, we developed a model of the measles virus (MV)-specific secondary antibody response in vitro to assess the presence of memory B lymphocytes in different B cell subsets isolated from human tonsils and explore the activation requirements of human memory B cells. Our findings show that the memory B cell pool resides in the CD38- B cell subpopulation and that the differentiation of MV-activated memory B cells into antibody-secreting cells can be achieved upon co-stimulation with interleukin (IL)-2 and IL-10, but does not require engagement of CD40. Interestingly, the CD40-mediated signal was found to synergize with Ig-cross-linking agents for the proliferation of memory B cells, but strongly suppressed their capacity to differentiate along the plasmacytoid pathway. Collectively, our results suggest that the CD40 signaling pathway is instrumental for the clonal expansion of the memory B cell pool, but does not operate in the later phase of the response, which allows their maturation into antibody-secreting cells.

Two major B cell subpopulations were identified in the IgD- compartment of tonsils and subsequently isolated. They displayed the following phenotypes: CD10+CD38+CD44- (CD38+ B cells) and CD10-CD38-CD44+ (CD38- B cells). Of the CD38- B cells, 70% also expressed CD24 and CD39, whereas CD77 was specifically distributed on 40% of CD38+ B cells, suggesting an additional level of heterogeneity in the cellular composition of these two B cell types. Whereas the majority of CD38+ B cells were in cycle, most CD38- B cells were quiescent. Conversely, Bcl-2 was expressed in CD38- B cells but was not detected in CD38+ B cells. Of the CD38- B cells, 30% bore the homing receptor Leu-8/Mel-14, whereas CD38+ B cells lacked this marker. Thus, CD38- B cells have both survival capacity and migratory competence. Both subsets expressed surface (s) Igs which were mainly of the IgG class, implying that most of these cells have already undergone isotype switching. CD38- B cells proliferated vigorously and produced large amounts of IgG in response to cytokines, following ligation of slgs or CD40. In contrast, CD38+ B cells were only stimulated for DNA synthesis by a combination of IL-4 and anti-CD40 antibodies, and failed to differentiate into Ig-secreting cells regardless of the stimulus applied. We propose that CD38- B cells represent an extra-follicular mature B cell population which has been positively selected and rescued from apoptosis, whereas the CD38+ B cell subset is composed of germinal centre B cells.