ERK1/2通路在VEGF促进绵羊卵泡颗粒细胞增殖中的作用

ERK1/2 pathway plays a important role in the proliferationon sheep's granulosa cells

选取10只1.5岁成年小尾寒羊母羊,分别取卵泡颗粒细胞并进行培养。设置5个试验组和1个空白对照组,试验组采用不同质量浓度血管内皮生长因子(VEGF)刺激,空白组不添加VEGF。并用MTT检测细胞增殖情况,Western-blotting检测细胞中P-ERK1/2表达水平。MTT比色结果表明,当VEGF质量浓度为5μg/L时对细胞增殖的促进作用最大,且试验组均高于对照组;Western-blotting结果显示,P-ERK1/2表达水平随VEGF的添加而显著增加,并在5μg/L时达到最大值(P<0.05)。但在10、15、20μg/L质量浓度下其表达量差异不显著(P>0.05)。这说明不同质量浓度VEGF对颗粒细胞增殖的促进作用各不相同,其中以5μg/L的VEGF效果最好。当颗粒细胞培养至48h时,P-ERK1/2表达水平较24h时略有提高,并在5μg/L VEGF刺激时达到最大值。VEGF可能是通过促进P-ERK1/2的表达进而促进颗粒细胞的增殖。

In this article,the role of ERK1/2 mitogen-activated protein kinases signal transduction pathway in the proliferation of sheep＇s granulosa cell enhanced by VEGF was studied.The granulosa cell from 10 adult female small fat-tail sheep was cultured and stimulated with different concentrations of VEGF.The cellular proliferation was detected by MTT and the expression of P-ERK1/2 was detected by western-blotting.The results of MTT showed a significant affection of granulosa cell proliferation by different concentration of VEGF and the 5 μg/L was the best.Western-blotting showed the expression of P-ERK1/2 increased significantly with the increasing of VEGF concentrations,and the optimized was 5 μg/L（P0.05）,the continued increasing of VEGF was insignificant effect on the expression of P-ERK1/2,such as the 10,15 and 20 μg/L.However,the inhibition of VEGF with the concentration of 25 μg/L weakened with the continued culture.The results shown that different levels of VEGF displayed different proliferation in granulosa cell.The expressions of P-ERK1/2 at 48 h is a little higher than that of 24 h.We can conclude that VEGF can activate ERK1/2 pathway and then increase the proliferation on granulosa cells of sheep.