Sun Q Y, Wang W H, Hosoe M, Taniguchi T, Chen D Y, Shioya Y
Department of Animal Reproduction, National Institute of Animal Industry, Ibaraki, Japan.
Dev Growth Differ. 1997 Aug;39(4):523-9. doi: 10.1046/j.1440-169x.1997.t01-2-00014.x.
The effects of protein kinase C (PKC) activation on meiotic resumption and cortical granule (CG) exocytosis as well as its dependence on Ca2+ in porcine eggs matured in vitro were studied. Cortical granule release was judged by both confocal laser microscopy after the eggs were labeled with fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and electron microscopy. Meiotic resumption and pronuclear formation were observed after eggs were stained with acetic orcein. When eggs were treated with PKC activators, 1-oleyl-2-acetyl-glycerol (OAG) or phorbol 12-myristate 13-acetate (PMA), the pronuclear formation percentage was significantly lower than that of Ca2+ ionophore A23187-treated group, but not statistically different from that in negative control group (P > 0.05), and most of the eggs were still arrested at metaphase II stage, suggesting that PKC activation does not induce the resumption of meiosis and pronuclear formation. In contrast, PKC activation induced 89.1% to 100% of the eggs completely or partially released their CG in different groups, not statistically different from A23187-treated group, and this effect could be overcome by PKC inhibition. When the intracellular free Ca2+ was chelated with acetoxymethal ester form of 1,2-bis(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM), and then treated with PMA or OAG in Ca(2+)-free medium, the proportions of eggs with CG release were 90.9% and 78.1%, respectively, not statistically different from the above-treated groups, suggesting that CG exocytosis induced by PKC activation is independent of Ca2+ rise. The results indicate that different events of porcine egg activation may be uncoupled from one another.
研究了蛋白激酶C（PKC）激活对体外成熟猪卵母细胞减数分裂恢复、皮质颗粒（CG）胞吐作用的影响及其对Ca2+的依赖性。在用异硫氰酸荧光素-花生凝集素（FITC-PNA）标记卵母细胞后，通过共聚焦激光显微镜和电子显微镜判断皮质颗粒释放情况。在用醋酸洋红染色后观察减数分裂恢复和原核形成情况。当用PKC激活剂1-油酰-2-乙酰甘油（OAG）或佛波酯12-肉豆蔻酸酯13-乙酸酯（PMA）处理卵母细胞时，原核形成百分比显著低于钙离子载体A23187处理组，但与阴性对照组无统计学差异（P>0.05），且大多数卵母细胞仍停滞在中期II期，表明PKC激活不会诱导减数分裂恢复和原核形成。相反，PKC激活在不同组中诱导89.1%至100%的卵母细胞完全或部分释放其CG，与A23187处理组无统计学差异，且这种作用可被PKC抑制所克服。当用1,2-双（O-氨基苯氧基）乙烷-N,N,N',N'-四乙酸乙酰甲酯（BAPTA-AM）螯合细胞内游离Ca2+，然后在无Ca(2+)培养基中用PMA或OAG处理时，CG释放的卵母细胞比例分别为90.9%和78.1%，与上述处理组无统计学差异，表明PKC激活诱导的CG胞吐作用与Ca2+升高无关。结果表明，猪卵母细胞激活的不同事件可能相互解偶联。
