Barda-Saad M, Zhang A S, Zipori D, Rozenszajn L A
Department of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel.
Stem Cells. 1997;15(3):229-36. doi: 10.1002/stem.150229.
We recently reported on selective interactions between immature T cell subpopulations and bone marrow (BM) stromal cells. To further study this process, we first examined the efficacy of methods estimating cell-cell adhesion and then investigating the effects of cytokines on thymocyte-stroma associations. Techniques based on the use of the fluorochromes 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 with FDA labeling, there was a rapid cellular efflux. Using calcein-AM, we developed an accurate quantitative fluorometric assay for determining the adherence of thymocytes to a BM stromal cell line (MBA-13). A maximal fraction of about 29% thymocytes was found to adhere to confluent MBA-13 cell layers after four to six h of coculture. Whereas interleukin 1 did not change the rate of adhesion of thymocytes to the stroma, interferon-gamma (IFN-gamma) significantly increased adhesion. Basic fibroblast growth factor (bFGF) had a dose-dependent biphasic effect on thymocyte adhesion, and a greater fraction of double negative thymocytes adhered to stroma pretreated with bFGF. Taken together, these results suggest that IFN-gamma and bFGF modulate T cells-BM stromal cell adhesion.
