APOPTOSIS AND BCL-2 EXPRESSION IN CULTURED MURINE SPLENIC T-CELLS

To elucidate the mechanism by which bcl-2 affects apoptosis in post-th ymic T cells, we investigated the expression of Bcl-2 protein in prima ry cultures of splenic T cells and in the interleukin-2 (IL-2)-depende nt T-cell line CTLL-2. The overall level of Bcl-2 was determined by im munoblotting, and the variability in Bcl-2 expression was determined b y flow cytometry. For a few days after concanavalin A (Con A) plus IL- 2 activation, the overall level of Bcl-2 in T cells remains unchanged, but it becomes more heterogeneous. By 5 days after activation, the ex pression returns to a more homogeneous distribution, but it is increas ed up to threefold above pre-activation levels, depending upon the dos e of IL-2 supplied. When Con A blasts or CTLL-2 cells are deprived of IL-2 for 34 hr, there is no change in their overall Bcl-2 levels which remain homogeneous even though almost half of the cells are apoptotic . However, when bcl-2 transfected CTLL-2 cells are deprived of IL-2, t hey do not undergo apoptosis, and their endogenous Bcl-2 protein level slowly decreases relative to their total protein. These data document the IL-2-dependent expression of Bcl-2 in activated T cells, confirm the ability of deregulated bcl-2 to inhibit the onset of apoptosis aft er IL-2 withdrawal, but suggest that, after IL-2 withdrawal, a drop in Bcl-2 levels relative to total protein levels does not precede apopto sis.