ADHESION OF THYMOCYTES TO BONE-MARROW STROMAL CELLS - REGULATION BY BFGF AND IFN-GAMMA

We recently reported on selective interactions between immature T cell subpopulations and bone marrow (BM) stromal cells. To further study t his process, we first examined the efficacy of methods estimating cell -cell adhesion and then investigated the effects of cytokines on thymo cyte-stroma associations. Techniques based on the use of the fluorochr omes calcein-acetomethylester (calcein-AM) and fluorescein diacetate ( FDA) were studied and compared to regular cell counting methods. With calcein-AM labeling, the retention time was relatively long, while wit h FDA labeling, there was a rapid cellular efflux. Using calcein-AM, w e developed an accurate quantitative fluorometric assay for determinin g the adherence of thymocytes to a BM stromal cell line (MBA-13). A ma ximal fraction of about 29% thymocytes was found to adhere to confluen t MBA-13 cell layers after four to six h of coculture. Whereas interle ukin 1 did not change the rate of adhesion of thymocytes to the stroma , interferon-gamma (IFN-gamma) significantly increased adhesion. Basic fihroblast growth factor (bFGF) had a dose-dependent biphasic effect on thymocyte adhesion, and a greater fraction of double negative thymo cytes adhered to stroma pretreated with bFGP. Taken together, these re sults suggest that IFN-gamma and bFGF modulate T cells-BM stromal cell adhesion.