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The Journal of Immunology, Vol 154, Issue 4 1634-1643, Copyright © 1995 by American Association of Immunologists
ARTICLES |
J Lomo, HK Blomhoff, K Beiske, T Stokke and EB Smeland
Department of Immunology, Norwegian Radium Hospital, Montebello, Oslo, Norway.
TGF-beta and agents that elevate intracellular cAMP levels are potent inhibitors of B cell activation in vitro and have been shown to arrest stimulated B cells in the G1 phase of the cell cycle. We tested the effects of TGF-beta 1 and the cAMP-inducing agent, forskolin, on the viability of resting B cells from human peripheral blood, and found that both agents caused a significant, dose-dependent increase in cell death relative to spontaneous death in medium alone, as measured by vital dye staining with propidium iodide. Apoptosis was shown to be the overall mode of death by demonstrating DNA fragmentation using DNA nick end labeling and by verifying the characteristic morphologic changes. In contrast with TGF-beta 1 and forskolin, various B cell activation stimuli generally inhibited spontaneous apoptosis of resting cells. The most potent effects were observed with IL-4 and the phorbol ester, O- tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C. IL-4 also partly inhibited TGF-beta 1 and forskolin-induced apoptosis. In contrast, TPA completely reversed cell death in forskolin- treated cultures, but had no effect on TGF-beta 1-induced apoptosis, indicating that TGF-beta 1 and forskolin promote apoptosis by different mechanisms. The relative protein expression of bcl-2, a proto-oncogene that inhibits apoptosis, was unaltered by the apoptotic as well as the survival stimuli tested, suggesting that apoptosis was regulated by a bcl-2-independent mechanism. We conclude that apoptosis is a regulated phenomenon in resting human B cells. Furthermore, TGF-beta and cAMP may inhibit B cell responses not only by blocking cell cycle progression in activated cells, but also by inducing apoptosis in resting cells.
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