Van Voorhis B J, Dunn M S, Snyder G D, Weiner C P
Department of Obstetrics and Gynecology, University of Iowa College of Medicine, Iowa City 52242.
Endocrinology. 1994 Nov;135(5):1799-806. doi: 10.1210/endo.135.5.7525252.
We investigated the presence of nitric oxide (NO) synthase in ovarian follicular cells obtained from women undergoing in vitro fertilization procedures. Endothelial NO synthase messenger RNA was demonstrated by polymerase chain reaction amplification of reverse transcribed RNA. NO synthase was localized to granulosa-luteal cells by immunocytochemistry, using a monoclonal antibody. Ovarian follicular cell NO synthase enzyme activity was confirmed by measuring the conversion of L-arginine to citrulline. To investigate the effect of NO on granulosa-luteal cell steroidogenesis, NO synthase inhibitors and NO donors were added to cell cultures. NG-Monomethyl-L-arginine and N-nitro-arginase methyl ester, selective inhibitors of NO synthase, significantly increased estradiol secretion by granulosa-luteal cells. S-Nitroso-L-acetyl penicillamine (S-NAP) and S-nitroso glutathione, NO donors, caused a dose-dependent decrease in both estradiol and progesterone secretion. The decrease by S-NAP was reversed by hemoglobin, which binds free NO. Although S-NAP increased the concentration of cGMP in granulosa-luteal cells, cGMP analogs had no effect on steroidogenesis in cell cultures. S-NAP and native NO in solution decreased cellular and microsomal aromatase activities. We conclude that NO synthase is present in human granulosa-luteal cells and that NO inhibits estradiol secretion independent of cGMP by directly inhibiting aromatase.
我们研究了从接受体外受精手术的女性获取的卵巢卵泡细胞中一氧化氮（NO）合酶的存在情况。通过逆转录RNA的聚合酶链反应扩增来证明内皮型NO合酶信使RNA的存在。使用单克隆抗体，通过免疫细胞化学方法将NO合酶定位到颗粒黄体细胞。通过测量L-精氨酸向瓜氨酸的转化来确认卵巢卵泡细胞NO合酶的酶活性。为了研究NO对颗粒黄体细胞类固醇生成的影响，将NO合酶抑制剂和NO供体添加到细胞培养物中。NO合酶的选择性抑制剂NG-单甲基-L-精氨酸和N-硝基-精氨酸甲酯显著增加了颗粒黄体细胞的雌二醇分泌。NO供体S-亚硝基-L-乙酰青霉胺（S-NAP）和S-亚硝基谷胱甘肽导致雌二醇和孕酮分泌呈剂量依赖性减少。S-NAP引起的减少可被结合游离NO的血红蛋白逆转。尽管S-NAP增加了颗粒黄体细胞中cGMP的浓度，但cGMP类似物对细胞培养物中的类固醇生成没有影响。S-NAP和溶液中的天然NO降低了细胞和微粒体芳香化酶的活性。我们得出结论，人类颗粒黄体细胞中存在NO合酶，并且NO通过直接抑制芳香化酶来独立于cGMP抑制雌二醇分泌。
