Facilitative Interactions between Estradiol and Luteinizing Hormone in the Regulation of Progesterone Production by Cultured Swine Granulosa Cells: Relation to Cellular Cholesterol Metabolism*

Abstract

Well differentiated swine granulosa cells in monolayer culture were employed to investigate the mechanisms by which estradiol amplifies the stimulatory actions of LH in the later stages of follicular maturation. The facilitative interaction between estradiol and LH could not be attributed to altered rates of catabolism of progesterone to 20α-hydroxypregn- 4-en-3-one. Moreover, estradiol, LH, and estradiol combined with LH clearly stimulated pregnenolone production, measured in the presence of trilostane, to inhibit 3β-hydroxysteroid dehydrogenase- Δ^5-4isomerase activity. Thus, at least one component of the synergism between estradiol and LH must reside at or proximal to the cholesterol side-chain cleavage system. In the absence of lipoproteins, the magnitude of the synergism between estradiol and LH was significantly reduced. However, the facilitative interaction between estradiol and LH could still be observed in lipoprotein-deficient and serum-free medium and after the administration of ML-236B to suppress the de novo biosynthesis of cholesterol. In contrast, estradiol alone and LH alone significantly augmented progesterone production in the presence of the oxygenated sterol 5-cholesten-3β,25-diol, which can serve as an effective substrate for cholesterol side-chain cleavage. In addition, in the presence of 5-cholesten-3β,25-diol, the magnitude of the synergism between estradiol and LH was increased markedly. Thus, the present studies demonstrate that estradiol and LH significantly stimulate progesterone production, at least in part, by augmenting pregnenolone biosynthesis, but not by inhibiting progesterone catabolism. Studies with 5-cholesten-3β- 25-diol further suggest that estradiol and LH increase cholesterol side-chain cleavage activity. These observations delineate important mechanisms by which estradiol and LH prepare well differentiated granulosa cells for the high rates of steroidogenesis that are ultimately required in the corpus luteum.