Co-ligation of surface IgM and CD40 on naive B lymphocytes generates a blast population with an ambiguous extrafollicular/germinal centre cell phenotype

We have previously shown that dual occupancy of sIgM and CD40 - essential receptors in T-dependent B cell responses - by antibodies held on CD32-L cells results in the rapid proliferation of resting human B lymphocytes in a cytokine-independent manner. Here we report the detailed phenotype of the blast population emerging in such cultures. By 3 days the levels of CD19 and CD20 have increased 4- and P-fold respectively: such high level expression of these two pan-B markers is characteristic of cells of germinal centre (GC) origin, B cells co-stimulated via sIgM and CD40 express low level CD23 and almost half became CD5(+); they also acquire CD38 and - importantly - CD77, both of these being selective markers of GC B cells, Expression of sIgM and IgD is down-regulated on these cells and a minor, but significant, population of IgG(+) cells appears, In marked contrast to GC B cells, the population proliferating in response to dual occupancy of sIgM and CD40 has up-regulated and strongly expresses CD44, Morphologically, the cells are heterogeneous but there is a dominant blastic cell type with relatively scanty cytoplasm and having multiple nucleoli, both of which are characteristic of centroblasts; nevertheless, these cells remain morphologically distinct from freshly isolated GC B cells and do not show hallmark features of centrocytes, Although there is substantial cell death occurring by days 6-7 in these cultures, there is no morphological evidence for apoptosis, Thus, the proliferating population that emerges from the dual engagement of antigen receptor and CD40 on resting B cells appears to be bestowed with some features of GC B cells but has others which are incompatible with that particular stage of differentiation, The possibility that it might represent (i) a blast stage that is transitional between activation in T zones and entry into the follicle or (ii) a precursor population that colonizes the primary follicle prior to GC formation is discussed.