Nitric oxide sensitizes ovarian tumor cells to Fas-induced apoptosis

Fas-mediated apoptosis represents one major mechanism by which tumor cells can be eliminated by activated cytotoxic immune lymphocytes. Previously, we have reported that interferon-gamma (IFN-gamma) sensitizes human ovarian carcinoma cell lines to Fas-mediated apoptosis. Furthermore, IFN-gamma, together with many other proinflammatory cytokines (TNF-alpha, IL-1beta, LPS, etc.), can stimulate the induction of inducible nitric oxide synthase (iNOS) and the generation of nitric oxide (NO). In this study, we examined whether nitric oxide is a mediator of IFN-gamma-induced sensitization of human ovarian carcinoma cell lines (A2780 and AD10) to Fas-mediated apoptosis and whether NO regulates the expression of the Fas receptor. Treatment of quiescent A2780 and AD10 ovarian carcinoma cells with IFN-gamma alone induced the expression of iNOS mRNA as examined by RT-PCR. There was accumulation of nitrite in the culture medium of IFN-gamma-treated cells, suggesting the generation of NOx. Like IFN-gamma, the use of exogenous sources of NO (S-nitroso-N-acetylpenicillamine (SNAP)) mimicked the sensitization of both cell lines to anti-Fas cytotoxic antibody (CH11) by IFN-gamma. Endogenously produced NO, by IFN-gamma pretreatment or exogenous nitrodonors, resulted in the upregulation of Fas receptor mRNA and protein expression. Blocking iNOS activity by NG-monomethyl-l-arginine (l-NMA) significantly reduced the sensitization, Fas mRNA, and protein expression observed with IFN-gamma pretreatment of the tumor cells. These findings demonstrate that sensitization of human ovarian carcinoma cell lines to Fas-mediated apoptosis by IFN-gamma can be due, in part, to the induction of iNOS and the subsequent upregulation of Fas gene expression by reactive nitrogen intermediates. Thus, the sensitivity of tumor cells to Fas-L-mediated cytotoxic immune lymphocytes can be regulated by the induction of NO or intermediates.