双酚A对颗粒细胞激素生成及相关甾体生成酶的影响

Effect of bisphenol A on progesterone,estradiol and correlative steroidogensis enzyme mRNA expression in granulosa cells

目的探讨双酚A(BPA)对大鼠颗粒细胞雌、孕激素生成及相关甾体生成酶的影响。方法采用大鼠卵巢颗粒细胞进行体外培养,72h后给予不同浓度的BPA(0、10-7、10-6、10-5、10-4M)处理,继续培养48h后,收集细胞培养液,用超敏感的固相放射免疫方法测定孕酮和雌二醇(E2)的浓度;用荧光实时定量PCR法测定颗粒细胞内芳香化酶(P450arom)、胆固醇侧链裂解酶(P450Scc)和类固醇合成急性调节蛋白(StAR)的mRNA表达水平。结果在BPA作用下,颗粒细胞孕酮分泌量在10-7M到10-5M组逐渐升高(P<0.05),但在10-4M剂量组出现意外陡降(P<0.01);而E2的分泌量则随BPA浓度的增加而下降(P<0.01)。P450arom mRNA表达显著降低;P450Scc表达量亦在10-7M到10-5M组逐渐升高(P<0.05),而10-4M组出现显著降低(P<0.01)。StARmRNA表达在10-4M组增高(P<0.05)。结论BPA能直接作用于卵巢颗粒细胞,通过作用于P450arom,P450Scc等相关酶的表达以及类固醇合成的限速因子-StAR的表达,而影响颗粒细胞雌孕激素的分泌。

Objective To investigate the effect of BPA on progesterone, estradiol production and expression of P450arom, P450scc and steroid acute regulator （STAR） genes in rat granulosa cells. Methods The granulosa cells were isolated from rat ovarian and cultured with MCcoy＇ 5A medium. After cultured for 72 hours, the cells were treated with various concentrations of BPA （0,10^-7 , 10^-6 , 10^-5 ,10^-4 M）. The culture media at 48 h after treatment were collected for the measurements of progesterone,estradiol level with high sensitive RIA. Gene expression of two key enzymes related with progesterone, estradiol production, P450arom and P450Scc, as well as STAR, in the cultured granulose cells was detected by RT-PCR. Results Progesterone levels in medium rised gradully as BPA concentration increased （P〈0. 05 ）, but dropped unexpectedly in 10^-4 M group （P〈0. 01 ）. However,estradiol level was decreased at the concentrations 10^-7 M to 10^-4 M（P〈0.01）. The expression of P450Scc mRNA was increased as BPA increased from 10^-7 M to 10^-5 M（P〈0.05）, but signficantly decreased at BPA 10^-4 M（P（0.01）. So did for progesterone. The expression of StAR mRNA in granulosa cells was increased markedly at BPA 10^-4 M. Conclusion The results indicate that BPA may interrupt ovarian steroidogenesis though acting on steroidogenesis enzymes and STAR.