Kodaman P H, Aten R F, Behrman H R
Department of Obstetrics and Gynecology, Yale University School of Medicine, New Haven, Connecticut 06520-8063.
Endocrinology. 1994 Dec;135(6):2723-30. doi: 10.1210/endo.135.6.7988463.
At functional luteolysis, the rat corpus luteum generates hydrogen peroxide (H2O2), which is known to rapidly inhibit gonadotropin-sensitive cAMP and progesterone production in isolated luteal cells. Lipid peroxides also increase markedly in the rat corpus luteum with the onset of functional luteolysis, and H2O2 is a potent inducer of lipid peroxidation. However, the actions of lipid peroxides on cell function are unknown. The objective of this study was to investigate the impact of typical lipid peroxides, cumene hydroperoxide (CuOOH) and 15(S)-hydroperoxyeicosatetraenoic acid, on rat luteal cells. CuOOH inhibited both LH-sensitive cAMP accumulation (ED50, 25 microM) and progesterone production (ED50, 20 microM). 15(S)-hydroperoxyeicosatetraenoic acid also dose dependently inhibited steroidogenesis. A significant reduction of LH-stimulated progesterone production was evident within 5 min of treatment with CuOOH, whereas inhibition of cAMP accumulation was not evident until 60 min. 8-Bromo-cAMP and 22-hydroxycholesterol caused partial and complete reversal of CuOOH-inhibited progesterone secretion, respectively. Preincubation of cells with o-phenanthroline completely reversed the inhibitory effects of CuOOH on cAMP accumulation and partially reversed its effects on progesterone production. Incorporation of radiolabeled amino acids into luteal proteins was significantly inhibited by CuOOH (25 microM) within 2 min of treatment and was reduced to 40 +/- 14% of control levels at 60 min. CuOOH (25 microM) maximally stimulated PGE2 production within 30 min of treatment (180 +/- 30% of control), a response that was completely blocked by aristolochic acid (100 microM), a phospholipase-A2 inhibitor, and indomethacin (1 microgram/ml), a prostaglandin (PG) synthesis inhibitor. The present results suggest that the acute inhibitory action of lipid peroxides on LH-stimulated progesterone production occurs down-stream of cAMP synthesis and appears to be due to impaired cholesterol utilization for steroidogenesis, possibly through inhibition of protein synthesis. The stimulation of PGE2 production by CuOOH appears to involve the activation of phospholipase-A2, which is a rate-limiting step in PG synthesis. Lipid peroxides as well as H2O2 may serve as mediators of functional luteolysis.
在功能性黄体溶解时，大鼠黄体产生过氧化氢（H₂O₂），已知其能迅速抑制分离的黄体细胞中促性腺激素敏感的环磷酸腺苷（cAMP）和孕酮的产生。随着功能性黄体溶解的开始，大鼠黄体中的脂质过氧化物也显著增加，且H₂O₂是脂质过氧化的有效诱导剂。然而，脂质过氧化物对细胞功能的作用尚不清楚。本研究的目的是探讨典型的脂质过氧化物，即氢过氧化异丙苯（CuOOH）和15（S）-氢过氧化二十碳四烯酸对大鼠黄体细胞的影响。CuOOH抑制促黄体生成素（LH）敏感的cAMP积累（半数有效剂量[ED₅₀]，25微摩尔）和孕酮产生（ED₅₀，20微摩尔）。15（S）-氢过氧化二十碳四烯酸也呈剂量依赖性地抑制类固醇生成。用CuOOH处理5分钟内，LH刺激的孕酮产生明显显著降低，而cAMP积累的抑制直到60分钟才明显。8-溴-cAMP和22-羟基胆固醇分别部分和完全逆转了CuOOH抑制的孕酮分泌。用邻菲罗啉预孵育细胞完全逆转了CuOOH对cAMP积累的抑制作用，并部分逆转了其对孕酮产生的作用。用CuOOH（25微摩尔）处理2分钟内，放射性标记氨基酸掺入黄体蛋白的过程即受到显著抑制，60分钟时降至对照水平的40±14%。用CuOOH（25微摩尔）处理30分钟内最大程度地刺激了前列腺素E₂（PGE₂）的产生（为对照的180±30%），这一反应被磷脂酶A₂抑制剂马兜铃酸（100微摩尔）和前列腺素（PG）合成抑制剂吲哚美辛（1微克/毫升）完全阻断。目前的结果表明，脂质过氧化物对LH刺激的孕酮产生的急性抑制作用发生在cAMP合成的下游，似乎是由于类固醇生成中胆固醇利用受损，可能是通过抑制蛋白质合成。CuOOH对PGE₂产生的刺激似乎涉及磷脂酶A₂的激活，这是PG合成中的限速步骤。脂质过氧化物以及H₂O₂可能作为功能性黄体溶解的介质。
