The goal of this proposal is to characterize early B cell precursors and to investigate the regulation of primary B cell differentiation by bone marrow stromal cells.
The first aim will assess the proliferative and differentiative potential of cells that express the early B cell marker, BP-1/6C3, by measuring their potential to reconstitute and maintain B cells following their injection into SCID mice. This study will elucidate if these cells can maintain steady state B lymphopoiesis or if input from a more immature cell compartment is necessary.
The second aim i s to further characterize defined stromal cell derived factors and identify additional mediators with effects on B lymphopoiesis. Specific experiments will examine if early B cell precursors that have not yet expressed the B220 antigen are IL-7 responsive, determine if IL-7 is involved in the maturation of pre-B cells into B lymphocytes, and explore potential synergistic/additive effects of IL-7 with other cytokines. Finally, a cDNA library prepared from the S17 stromal cell line will be screened using expression vector strategies in order to identify a non-IL-7 factor from the S17 stromal cell line that potentiates the formation of clonal, pre-B cell colonies in semi-solid medium. The cells in these colonies express cytoplasmic u heavy chain protein but not kappa. The objective of the third aim is to determine the status of light chain gene rearrangements of cells in S17 supported colonies in order to define the precise molecular stage at which the S17 factor acts. An inverted PCR approach will be used for this analysis. Cytokines play a key role in B lymphopoiesis, but contact between developing B cells and stromal cells also occurs.
The fourth aim will use anti-stromal cell monoclonal antibodies to identify and biochemically characterize cell surface determinants important in stromal cell-lymphocyte interactions. Taken together, the data from these studies will further elucidate a fundamental developmental process and be of value in understanding abnormalities of B lymphopoiesis.
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