Garbán H J, Bonavida B
Department of Microbiology, Immunology, and Molecular Genetics, and Jonsson Comprehensive Cancer Center, UCLA School of Medicine, 10833 Le Conte Avenue, Los Angeles, California 90095-1747, USA.
Gynecol Oncol. 1999 May;73(2):257-64. doi: 10.1006/gyno.1999.5374.
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.
Fas介导的细胞凋亡是活化的细胞毒性免疫淋巴细胞清除肿瘤细胞的一种主要机制。此前，我们曾报道干扰素-γ（IFN-γ）可使人卵巢癌细胞系对Fas介导的细胞凋亡敏感。此外，IFN-γ与许多其他促炎细胞因子（TNF-α、IL-1β、LPS等）一起，可刺激诱导型一氧化氮合酶（iNOS）的诱导及一氧化氮（NO）的生成。在本研究中，我们检测了一氧化氮是否是IFN-γ诱导人卵巢癌细胞系（A2780和AD10）对Fas介导的细胞凋亡敏感的介质，以及NO是否调节Fas受体的表达。用RT-PCR检测，单独用IFN-γ处理静止的A2780和AD10卵巢癌细胞可诱导iNOS mRNA的表达。IFN-γ处理的细胞培养基中出现亚硝酸盐积累，提示有NOx生成。与IFN-γ一样，使用外源性NO（S-亚硝基-N-乙酰青霉胺（SNAP））可模拟IFN-γ使两种细胞系对抗Fas细胞毒性抗体（CH11）敏感的作用。通过IFN-γ预处理或外源性硝基供体产生的内源性NO导致Fas受体mRNA和蛋白表达上调。用NG-单甲基-L-精氨酸（L-NMA）阻断iNOS活性可显著降低肿瘤细胞经IFN-γ预处理后观察到的敏感性、Fas mRNA和蛋白表达。这些发现表明，IFN-γ使人卵巢癌细胞系对Fas介导的细胞凋亡敏感，部分原因可能是iNOS的诱导以及随后活性氮中间体对Fas基因表达的上调。因此，肿瘤细胞对Fas-L介导的细胞毒性免疫淋巴细胞的敏感性可通过NO或中间体的诱导来调节。
