Ui M, Shimonaka M, Shimasaki S, Ling N
Department of Molecular Endocrinology, Whittier Institute, La Jolla, California 92037.
Endocrinology. 1989 Aug;125(2):912-6. doi: 10.1210/endo-125-2-912.
An inhibitor of FSH action on granulosa cells has been purified from porcine follicular fluid using a combination of ammonium sulfate precipitation, dialysis in 30% (vol/vol) acetic acid, gel filtration chromatography under acidic conditions, and several steps of reverse phase HPLC. Activity was monitored by using an in vitro granulosa cell bioassay, measuring the effect of the inhibitor on FSH-stimulated estradiol production. The purified polypeptide dose-dependently inhibited production of both estradiol (EC50 = 0.7 nM) and progesterone (EC50 = 1.3 nM) by rat granulosa cells cultured in the presence of 20 ng/ml FSH. N-Terminal sequence analysis revealed a high degree of homology with the 53,000 mol wt human GH-dependent insulin-like growth factor-binding protein (IGF-BP). Coincubation of stoichiometric amounts of IGF-I or -II and the inhibitor (IGF-BP) resulted in complete neutralization of the inhibitory effect. Since both IGFs are produced locally in the ovary and exert stimulatory effects on granulosa cells, local production of IGF-BP may provide an important means of regulating ovarian follicle growth.
已通过硫酸铵沉淀、在30%（体积/体积）乙酸中透析、酸性条件下的凝胶过滤色谱以及几步反相高效液相色谱法相结合的方法，从猪卵泡液中纯化出一种抑制卵泡刺激素对颗粒细胞作用的物质。通过体外颗粒细胞生物测定法监测活性，测量该抑制剂对卵泡刺激素刺激的雌二醇产生的影响。纯化的多肽以剂量依赖方式抑制在20 ng/ml卵泡刺激素存在下培养的大鼠颗粒细胞产生雌二醇（半数有效浓度=0.7 nM）和孕酮（半数有效浓度=1.3 nM）。N端序列分析显示与53,000分子量的人生长激素依赖性胰岛素样生长因子结合蛋白（IGF-BP）高度同源。化学计量的胰岛素样生长因子-I或-II与抑制剂（IGF-BP）共同孵育导致抑制作用完全中和。由于两种胰岛素样生长因子均在卵巢局部产生并对颗粒细胞发挥刺激作用，因此IGF-BP的局部产生可能是调节卵巢卵泡生长的重要手段。
