Petrino T, Schuetz A W
Biol Reprod. 1987 Jun;36(5):1219-28. doi: 10.1095/biolreprod36.5.1219.
Treatment of isolated amphibian ovarian follicles with frog pituitary homogenate (FPH) increases follicular progesterone levels, which, in turn, initiate oocyte maturation. Recent studies have demonstrated that follicular progesterone production requires concomitant protein synthesis at some stage preceding pregnenolone formation. Experiments were carried out to determine whether cholesterol metabolism plays a role in mediating these biochemical and physiological processes. Aminoglutethimide (AGI, and inhibitor of P450 side-chain cleavage enzyme) inhibited FPH-induced intrafollicular progesterone accumulation and oocyte maturation (or germinal vesicle breakdown, GVBD) in a dose-dependent manner. Follicular progesterone accumulation and GVBD were both stimulated, in the absence of FPH, after addition of 25-OH-cholesterol, but not cholesterol, to the culture medium. Higher levels of progesterone were present in defolliculated oocytes as compared to intact ovarian follicles after incubation with 25-OH-cholesterol. The results indicate that the surface epithelium and theca layer in the follicle wall retard 25-OH-cholesterol access to steroid-producing follicle cells. AGI blocked 25-OH-cholesterol-induced accumulation of progesterone and GVBD in defolliculated oocytes, suggesting that 25-OH-cholesterol does not directly induce GVBD and is metabolized by the follicle cells. The capacity of follicles to accumulate progesterone following preincubation with FPH or 25-OH-cholesterol along with AGI was compared. Intrafollicular levels of progesterone increased after AGI- and 25-OH-cholesterol-treated follicles were washed. In contrast, progesterone levels decreased in follicles pretreated with AGI and FPH after washing. The results indicate that considerable 25-OH-cholesterol, but not endogenous cholesterol (FPH stimulation), remains available for steroidogenesis after removal of AGI. A significant, but incomplete, inhibition of progesterone accumulation occurred when follicles were incubated in the presence of 25-OH-cholesterol and cycloheximide. This partial blockage produced by the protein synthesis inhibitor indicates that some basal protein synthesis is required for progesterone accumulation from exogenous 25-OH-cholesterol. We conclude that intracellular cholesterol stores in the follicle wall are utilized to mediate FPH induction of progesterone accumulation and oocyte maturation in amphibian follicles.
用青蛙垂体匀浆（FPH）处理分离的两栖类卵巢卵泡可提高卵泡孕酮水平，进而启动卵母细胞成熟。最近的研究表明，卵泡孕酮的产生在孕烯醇酮形成之前的某个阶段需要伴随蛋白质合成。进行实验以确定胆固醇代谢是否在介导这些生化和生理过程中起作用。氨鲁米特（AGI，一种P450侧链裂解酶抑制剂）以剂量依赖的方式抑制FPH诱导的卵泡内孕酮积累和卵母细胞成熟（或生发泡破裂，GVBD）。在培养基中添加25-羟基胆固醇而非胆固醇后，在没有FPH的情况下，卵泡孕酮积累和GVBD均受到刺激。与完整卵巢卵泡相比，用25-羟基胆固醇孵育后，去卵泡卵母细胞中的孕酮水平更高。结果表明，卵泡壁中的表面上皮和卵泡膜层阻碍了25-羟基胆固醇进入产生类固醇的卵泡细胞[1]。AGI阻断了25-羟基胆固醇诱导的去卵泡卵母细胞中孕酮的积累和GVBD，这表明25-羟基胆固醇不会直接诱导GVBD，而是由卵泡细胞代谢。比较了用FPH或25-羟基胆固醇与AGI预孵育后卵泡积累孕酮的能力。用AGI和25-羟基胆固醇处理过的卵泡冲洗后，卵泡内孕酮水平升高。相比之下，用AGI和FPH预处理过的卵泡冲洗后，孕酮水平下降。结果表明，去除AGI后，大量的25-羟基胆固醇（而非内源性胆固醇，FPH刺激）仍可用于类固醇生成。当卵泡在25-羟基胆固醇和环己酰亚胺存在下孵育时，孕酮积累受到显著但不完全的抑制。蛋白质合成抑制剂产生的这种部分阻断表明，从外源性25-羟基胆固醇积累孕酮需要一些基础蛋白质合成。我们得出结论，卵泡壁中的细胞内胆固醇储存被用于介导两栖类卵泡中FPH诱导的孕酮积累和卵母细胞成熟。
