双酚A对体外培养小鼠睾丸睾酮合成的影响及机制

Effects of bisphenol A on testis testosterone synthesis in mouse cultured in vitro

目的探讨睾丸体外培养模型下双酚A(BPA)暴露对小鼠睾丸睾酮合成及基因表达的影响。方法采用睾丸器官体外培养方法,将睾丸随机分为DMSO溶剂对照组和4个BPA剂量染毒组(终浓度为10-7、10-6、10-5和10-4mol/L)。分别培养24、48和72h,每24h通气一次。HE染色光镜下观察睾丸组织形态学变化,放射免疫法检测培养液睾酮浓度,免疫组化法检测睾丸组织相关蛋白表达。实时荧光定量PCR检测睾丸组织相关基因表达水平。结果 10-4mol/L剂量组支持细胞数目减少,部分间质细胞核固缩或呈絮状改变;随着时间的延长睾酮分泌逐渐减少,与DMSO对照组相比,10-5mol/L剂量组在48h时睾酮合成增加(P<0.05),其余各剂量组睾酮分泌降低,以10-4mol/L剂量组较明显(P<0.01);睾酮合成相关酶3β-羟基类固醇脱氢酶(3βHSD)、胆固醇侧链裂解酶(P450scc)、细胞色素P45017a羟化酶(P450c17)及波形蛋白(Vimentin)的基因及蛋白表达水平下降。结论 BPA能通过抑制3βHSD、P450scc、P450c17及Vimentin等相关酶的表达,干扰间质细胞睾酮的合成。

Objective To investigate the effects of BPA on testosterone synthesis and the related genes expression in the fetal testis of male mouse by organ culture in vitro.Methods The testis tissues cultured in vitro were randomly divided into five groups,DMSO control group and 4 different concentration of BPA groups（10-7,10-6,10-5 and 10-4mol/L respectively）.The culture medium and gas were changed every 24 h.The histomorphological examination of the testis was conducted under optical microscope,the testosterone levels in the cultured medium were measured by radioimmunoassay,the expressions of related proteins were measured by immunohistochemistry,and the expression of 3βHSD,P450scc,P450c17 and Vimentin were determined by real-time PCR.Results Compared to the control group,the number of sertoli cells were decreased and flocculent changes appear in some Leydig ceils in 10-4mol/L dose group.Testosterone secretion was gradually reduced as time goes on.The testosterone synthesis of 10-5mol/L dose group was increased in 48h（P〈0.05）,and the rest groups were decreased,especially the 10-4mol/L dose group（P〈0.01）.The expression of genes coding for proteins involved in testosterone biosynthesis,3βHSD,P450scc,P450c17 and Vimentin were reduced.Conclusion Bisphenol A can affect testosterone synthesis of mouse cultured in vitro through suppression of 3βHSD,P450scc,P450c17 and Vimentin.