Nitric Oxide Sensitizes Ovarian Tumor Cells to Fas-Induced Apoptosis

Abstract

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-γ (IFN-γ) sensitizes human ovarian carcinoma cell lines to Fas-mediated apoptosis. Furthermore, IFN-γ, together with many other proinflammatory cytokines (TNF-α, IL-1β, 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-γ-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-γ alone induced the expression of iNOS mRNA as examined by RT-PCR. There was accumulation of nitrite in the culture medium of IFN-γ-treated cells, suggesting the generation of NOx. Like IFN-γ, 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-γ. Endogenously produced NO, by IFN-γ pretreatment or exogenous nitrodonors, resulted in the upregulation of Fas receptor mRNA and protein expression. Blocking iNOS activity byNG-monomethyl-l-arginine (l-NMA) significantly reduced the sensitization, Fas mRNA, and protein expression observed with IFN-γ pretreatment of the tumor cells. These findings demonstrate that sensitization of human ovarian carcinoma cell lines to Fas-mediated apoptosis by IFN-γ 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.