Sirotkin A V, Grossmann R
Research Institute of Animal Production, Hlohovská 2, 949 92 Nitra, Slovakia.
Anim Reprod Sci. 2006 Mar;92(1-2):169-81. doi: 10.1016/j.anireprosci.2005.05.018. Epub 2005 Jul 18.
The aim of these experiments was to study the role of protein kinase A (PKA), cyclin-dependent kinase 2 (CDC2) and insulin-like growth factor II (IGF-II) in the control of ovarian function in domestic fowl, as well as the role of PKA and CDC2 in mediating the effects of IGF-II on the ovary. For this purpose, we studied the influence of an inhibitor of PKA (KT5720; 50 ng/ml), a CDC2 blocker (olomoucine; 1 microg/ml), IGF-II (0, 1, 10 or 100 ng/ml) and their combinations on cultured fragments of chicken ovarian follicular wall. Accumulation of PKA and CDC2 and secretion of progesterone (P4), testosterone (T), estradiol (E2) and arginine-vasotocin (AVT) were evaluated by using SDS-PAGE-Western blotting and RIA/EIA. IGF-II addition to culture medium stimulated T, E2 and AVT secretion and inhibited P4 secretion. These changes were associated with an increase in PKA and a decrease in CDC2 accumulation. The PKA blocker KT5720, when given alone, increased accumulation of PKA and secretion of T and E2, but not AVT and inhibited P4 secretion. The PKA blocker also prevented and even reversed the effects of IGF-II on PKA and steroid hormones secretion, but enhanced the action of IGF-II on AVT. The inhibitor of CDC2, olomoucine, when given alone, suppressed the expression of CDC2 and the secretion of P4 and AVT (but not T and E2). When given together with IGF-II, it augmented IGF-II-induced suppression of CDC2 and reversed the effects of IGF-II on P4 (but not on T, E2 or AVT). These observations demonstrate the involvement of PKA, CDC2 and IGF-II in regulating the secretory activity of avian ovarian cells. Our data also suggest the involvement of PKA in the mediation of IGF-II effects on P4, T, E2 and AVT secretion. CDC2 can mediate the effects of IGF-II on ovarian P4 secretion but not on other hormones.
这些实验的目的是研究蛋白激酶A（PKA）、细胞周期蛋白依赖性激酶2（CDC2）和胰岛素样生长因子II（IGF-II）在调控家禽卵巢功能中的作用，以及PKA和CDC2在介导IGF-II对卵巢作用中的作用。为此，我们研究了PKA抑制剂（KT5720；50 ng/ml）、CDC2阻滞剂（olomoucine；1 μg/ml）、IGF-II（0、1、10或100 ng/ml）及其组合对鸡卵巢卵泡壁培养片段的影响。通过SDS-PAGE-蛋白质印迹法和放射免疫分析/酶免疫分析评估PKA和CDC2的积累以及孕酮（P4）、睾酮（T）、雌二醇（E2）和精氨酸加压催产素（AVT）的分泌。向培养基中添加IGF-II可刺激T、E2和AVT的分泌，并抑制P4的分泌。这些变化与PKA的增加和CDC2积累的减少有关。单独给予PKA阻滞剂KT5720时，可增加PKA的积累以及T和E2的分泌，但不影响AVT，并抑制P4的分泌。PKA阻滞剂还可预防甚至逆转IGF-II对PKA和类固醇激素分泌的影响，但增强了IGF-II对AVT的作用。CDC2抑制剂olomoucine单独使用时，可抑制CDC2的表达以及P4和AVT的分泌（但不影响T和E2）。与IGF-II一起使用时，它可增强IGF-II诱导的对CDC2的抑制作用，并逆转IGF-II对P4的影响（但不影响T、E2或AVT）。这些观察结果表明PKA、CDC2和IGF-II参与调节禽类卵巢细胞的分泌活性。我们的数据还表明PKA参与介导IGF-II对P4、T、E2和AVT分泌的影响。CDC2可介导IGF-II对卵巢P4分泌的影响，但不影响其他激素。
