Selective involvement of ceramide in cytokine-induced apoptosis. Ceramide inhibits phorbol ester activation of nuclear factor kappaB.

Among its diverse biologic effects, the cytokine tumor necrosis factor alpha causes the rapid nuclear translocation of the transcription factor, nuclear factor kappaB (NF-kappaB). The p55 tumor necrosis factor (TNF) receptor shares with the related APO-1/Fas antigen the ability to initiate apoptosis. We investigated the role of the sphingolipid mediator ceramide in the cytokine-induced signaling mechanisms leading to NF-kappaB activation and cell death. Several lines of evidence presented here suggest that ceramide generated in response to TNFalpha or Fas activation is not involved in NF-kappaB activation. (i) Cell-permeable ceramides and exogenous sphingomyelinase failed to induce either nuclear translocation of NF-kappaB or degradation of its cytosolic inhibitor, I-kappaB, in Jurkat T cells. (ii) Ceramide treatment of cells inhibited phorbol ester-induced activation of NF-kappaB. (iii) TNFalpha potently activated NF-kappaB in a cell line deficient in acid sphingomyelinase. (iv) TNFalpha activated NF-kappaB within minutes without altering ceramide levels. (v) Treatment of Jurkat cells with cross-linking antibodies to APO-1/Fas induced large scale increases in ceramide and apoptosis without affecting NF-kappaB. (vi) Ceramide generation in response to Fas activation was inhibited by N-acetyltyrosinylvalinylalanylaspartyl chloromethyl ketone, a peptide inhibitor of interleukin-1beta-converting enzyme-like proteases, whereas TNFalpha-induced NF-kappaB activation was unaffected by the inhibitor. These results show that ceramide accumulation belongs selectively to the apoptotic pathway(s) induced by cytokines, and, if anything, ceramide may participate in negative feedback regulation of NF-kappaB.