Hylka V W, Kaki M K, diZerega G S
Livingston Reproductive Biology Laboratory, University of Southern California School of Medicine, Los Angeles 90033.
Endocrinology. 1989 Mar;124(3):1204-9. doi: 10.1210/endo-124-3-1204.
We studied the effects of porcine FSH, forskolin, and (Bu)2cAMP [agents that stimulate steroidogenesis via the adenylate cyclase-cAMP pathway (cAMP system)] either alone or with concomitant addition of phorbol 12-myristate 13-acetate (TPA; a phorbol ester that activates protein kinase-C) on steroidogenesis in porcine granulosa cells cultured from small (less than 3 mm) and medium-sized (3-6 mm) ovarian follicles. We attempted to determine if granulosa cells from different maturational states had different responses to these agonists and antagonists. Cells were cultured in serum-free medium 199 supplemented with insulin (10 micrograms/ml), transferrin ( 5 micrograms/ml), and androstenedione (2.5 X 10(-7) M) for 48 h. Levels of progesterone (P) and estradiol (E2) were determined in spent medium by RIA. We found that FSH, forskolin, and cAMP all stimulated secretion of E2 and P in a dose-dependent manner in both developmental groups. When TPA was added alone to cultures, P levels were stimulated at low doses of TPA but inhibited at higher doses in granulosa from both sized follicles, whereas cells from both small- and medium-sized follicles demonstrated reductions in E2. TPA was also found to inhibit FSH-, forskolin-, and cAMP-induced steroidogenesis in a dose-dependent manner in cells from the two groups of follicles. The stimulatory effects of any of the secretagogues on E2 secretion were inhibited by TPA to a significantly greater extent in granulosa cells from small follicles. Although inhibition of FSH- and forskolin-induced P secretion by TPA was also greater in granulosa cells from small follicles, cAMP-treated cells did not show this differential inhibition. Thus, it appears that modulators of the protein kinase-C system regulate steroidogenesis differently in granulosa cells from small and medium follicles. These differences may involve alterations in the interplay between the protein kinase-C and cAMP pathways.
我们研究了猪促卵泡素（FSH）、福斯高林和二丁酰环磷腺苷（Bu）2cAMP[通过腺苷酸环化酶 - cAMP途径（cAMP系统）刺激类固醇生成的试剂]单独使用或同时添加佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯（TPA；一种激活蛋白激酶C的佛波酯）对从小（小于3毫米）和中等大小（3 - 6毫米）卵巢卵泡培养的猪颗粒细胞类固醇生成的影响。我们试图确定来自不同成熟状态的颗粒细胞对这些激动剂和拮抗剂是否有不同反应。细胞在补充有胰岛素（10微克/毫升）、转铁蛋白（5微克/毫升）和雄烯二酮（2.5×10⁻⁷M）的无血清培养基199中培养48小时。通过放射免疫分析法测定培养液中孕酮（P）和雌二醇（E2）的水平。我们发现，FSH、福斯高林和cAMP在两个发育组中均以剂量依赖性方式刺激E2和P的分泌。当单独向培养物中添加TPA时，在两个大小卵泡的颗粒细胞中，低剂量的TPA刺激P水平，但高剂量时抑制P水平，而来自小卵泡和中等大小卵泡的细胞E2均减少。还发现TPA以剂量依赖性方式抑制两组卵泡细胞中FSH、福斯高林和cAMP诱导的类固醇生成。TPA对小卵泡颗粒细胞中任何一种促分泌剂对E2分泌的刺激作用的抑制程度明显更大。尽管TPA对小卵泡颗粒细胞中FSH和福斯高林诱导的P分泌的抑制作用也更大，但cAMP处理的细胞未表现出这种差异抑制。因此，蛋白激酶C系统的调节剂似乎对小卵泡和中等卵泡颗粒细胞中的类固醇生成调节方式不同。这些差异可能涉及蛋白激酶C和cAMP途径之间相互作用的改变。
