Shinohara O, Knecht M, Feng P, Catt K J
J Steroid Biochem. 1986 Jan;24(1):161-8. doi: 10.1016/0022-4731(86)90046-4.
Gonadotropin-stimulated steroidogenesis in the differentiating ovarian granulosa cell is mediated through the activation of cAMP-dependent protein kinase, and is also modulated by calcium-dependent mechanisms. Granulosa cells contain calcium-activated, phospholipid-dependent protein kinase (C kinase), and show an increase in phosphatidylinositol turnover in response to GnRH agonist analogs. To evaluate the role of C kinase in ovarian steroidogenesis, the potent phorbol ester, TPA, and the permeant diacylglycerol, OAG, were used to activate C kinase in granulosa cells from PMSG-treated immature rats. Both TPA and OAG caused dose-dependent stimulation of progesterone production without affecting intra- or extracellular cAMP levels. However, the maximum steroid responses to these compounds were less than those stimulated by cAMP. The ED50 for TPA-stimulated progesterone production was 3 nM, which is close to the known Km for activation of C kinase. Stimulation of steroidogenesis was only observed with biologically-active phorbol esters and permeant diacylglycerols such as OAG and DOG. Exposure of granulosa cells to phospholipase C also increased progesterone production in a dose-dependent manner without changing the cAMP content. Although TPA and OAG did not increase basal cAMP production, both agents enhanced the cAMP responses stimulated by hCG and forskolin; likewise, phospholipase C alone did not change cAMP production but caused a dose-dependent increase in the cAMP responses to hCG and forskolin. These results demonstrate that activation of C kinase promotes steroidogenesis in ovarian granulosa cells, and potentiates the activation of adenylate cyclase by hCG and forskolin. Such findings support the possibility that the calcium, phospholipid-dependent enzyme could be involved in the regulation of progesterone production by hormonal ligands such as gonadotropins and GnRH.
促性腺激素刺激分化中的卵巢颗粒细胞进行类固醇生成是通过环磷酸腺苷（cAMP）依赖性蛋白激酶的激活介导的，并且也受到钙依赖性机制的调节。颗粒细胞含有钙激活的、磷脂依赖性蛋白激酶（C激酶），并且对促性腺激素释放激素（GnRH）激动剂类似物有反应时，磷脂酰肌醇周转率增加。为了评估C激酶在卵巢类固醇生成中的作用，使用强效佛波酯（TPA）和可渗透的二酰基甘油（OAG）来激活来自孕马血清促性腺激素（PMSG）处理的未成熟大鼠的颗粒细胞中的C激酶。TPA和OAG均引起孕酮产生的剂量依赖性刺激，而不影响细胞内或细胞外cAMP水平。然而，这些化合物对类固醇的最大反应小于cAMP刺激的反应。TPA刺激孕酮产生的半数有效剂量（ED50）为3 nM，这接近已知的C激酶激活的米氏常数（Km）。仅在具有生物活性的佛波酯和可渗透的二酰基甘油（如OAG和DOG）存在时才观察到类固醇生成的刺激。颗粒细胞暴露于磷脂酶C也以剂量依赖性方式增加孕酮产生，而不改变cAMP含量。虽然TPA和OAG没有增加基础cAMP产生，但两种试剂均增强了人绒毛膜促性腺激素（hCG）和福斯高林刺激的cAMP反应；同样，单独的磷脂酶C没有改变cAMP产生，但导致对hCG和福斯高林的cAMP反应呈剂量依赖性增加。这些结果表明，C激酶的激活促进卵巢颗粒细胞中的类固醇生成，并增强hCG和福斯高林对腺苷酸环化酶的激活。这些发现支持了钙、磷脂依赖性酶可能参与诸如促性腺激素和GnRH等激素配体对孕酮产生的调节的可能性。
