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Many of the stromal-derived signals and factors that regulate B lymphopoiesis have been identified. We review recent evidence from our laboratory that shows that there are at least three phases during B-cell development when cells direct their own maturation, independent of stromal cells. Following the expression of the preB-cell receptor (preBCR), cells acquire the ability to proliferate in low levels of interleukin-7 (IL-7), which acts as a self-selecting mechanism to expand cells that have successfully expressed a preBCR in environments that are non-permissive to preBCR- cells. Second, the preBCR is required for a contact-mediated event between B-cell progenitors. Disruption at this stage prevents the further maturation of progenitors to the lipopolysaccharide (LPS)-responsive stage. Finally, the transition from IL-7 receptor to mature antigen receptor-based signaling is enhanced by a novel member of the tachykinin family, hemokinin-1. This series of maturation, survival, and differentiation signals is generated by B-lineage cells as they progress through developmental checkpoints on the way to becoming functionally mature cells.

B lymphocyte development is regulated at multiple checkpoints, mediated by signals originating both inside and outside the cell. Two signaling pathways known to be essential in this process are interleukin-7 (IL-7) and the pre-B cell receptor (pBCR). We have shown previously that these signaling pathways intersect functionally. Specifically, response to low concentrations of IL-7 requires pBCR expression. In this report, we identify the ERK/MAP kinase pathway as a key regulatory component of this response. We propose a molecular mechanism for the selective expansion of pBCR(+) precursors and for the culling of inappropriately rearranged pro-B cells.

Numerous studies have demonstrated that B lymphopoiesis is dependent upon a stromal cell microenvironment. Many of the stromal cell-derived factors and cell surface interactions that regulate B cell development have been identified; however, little consideration has been paid to the intimate interactions known to occur among B cell precursors themselves in both the fetal liver and marrow microenvironments. In this study we show that homotypic interactions between B cell precursors play an important role in promoting the development of mature B cells. We used an in vitro assay system to demonstrate that the function of stromal cells can be replaced by culturing B cell precursors in proximity. B cell precursors isolated from bcl-2 transgenic mice were used to rule out the possibility that improved survival, hypothesized to result from culturing precursors in proximity, solely accounted for the observed increase in B cell maturation. The putative maturation signal(s) were shown to be dependent upon direct contact between precursors rather than the release of soluble factors from nearby cells. Upon examination of the potential role of several known cell surface proteins, we found that blocking mu heavy chains with monovalent Fab antibody fragments dramatically inhibited maturation, in a stage-specific manner. Together these results suggest that a major function of stromal cells in vivo may be to act as a docking site to promote critical preB-preB homotypic interactions and ensuing signals. Further, the antibody blocking experiments raise the interesting possibility that interactions between B cell precursors themselves may promote and/or regulate preB cell receptor-driven signals.

Considerable progress has been made in defining intermediate stages in the process leading from stem cells to mature B cells. Cell-bound and secreted molecules direct the progression through these stages and regulate the selection of clones from which the immune repertoire emerges. In fact, a myriad of signals derived from B-cell progenitors themselves and the microenvironment in which they develop direct the differentiation process. These signals are provided by B-cell antigen receptors (BCR) and their surrogates, and by adhesion and cytokine receptors. The co-operation of these receptors to control survival, expansion, and differentiation of early B-cell progenitors is the topic of this review. Specifically, we will summarize recent findings from our laboratory demonstrating that preBCR expression lowers the threshold for interleukin (IL)-7 responsiveness. How signals initiated by these receptors may intersect at this critical point of B-cell selection will be discussed. At the stage following IL-7 responsiveness we have shown that interactions between B-cell progenitors themselves promote their differentiation to immunoglobulin-secreting B cells. We propose that one function of stromal cells, known to be central to B lymphopoiesis, is to promote critical preB-preB homotypic interactions and ensuing signals.

The IL-7R and the pre-B cell receptor (pre-BCR) each provide critical signals during differentiation of B cell precursors. In this study we examine the interplay between signals dependent upon these receptors. We demonstrate that pre-BCR-deficient pro-B cells differ significantly from controls in their ability to use the IL-7R. We show that this difference, characterized by a failure to proliferate in response to IL-7, is narrowly restricted to IL-7 concentrations in the picogram per milliliter range and can be overcome with increasing amounts of IL-7. Restoration of Ig heavy chain to recombinase-activating gene-2-deficient pro-B cells leads to a restored response to picogram per milliliter levels of IL-7, providing strong evidence that modulation of the IL-7 dose-response threshold is dependent on pre-BCR. Culture of normal pro-B cells under low IL-7 conditions leads to selective outgrowth of cells expressing mu heavy chain, suggesting that modulation of IL-7 dose-response thresholds can allow for selective expansion of pre-BCR+ cells under conditions where IL-7 is limiting. We also provide evidence that expression of pre-BCR on pro-B cells limits the duration of IL-7 responsiveness by causing differentiation to an IL-7-unresponsive pre-B cell stage. Thus, the pre-BCR-dependent modulation of IL-7 responsiveness affects both the dose-response threshold and the duration of IL-7-induced clonal expansion. Our results suggest that positive selection of pre-BCR+ pro-B cells may be achieved through the fine tuning of IL-7 responses.