邻苯二甲酸二丁酯对成年大鼠睾丸Leydig细胞中Cx43蛋白表达及睾酮分泌的影响

目的:观察邻苯二甲酸二丁酯(DBP)对成年大鼠睾丸Leydig细胞中连接蛋白43(Cx43)的表达及睾酮分泌的影响,探讨DBP对睾酮的调节是否与Cx43有关联。方法:将SD大鼠无菌脱臼处死,原代培养纯化Leydig细胞,将细胞分为溶剂对照组(0.1%DMSO)、DBP组(50 mg·L^-1 DBP)、Cx43增强剂组(50 mg·L^-1 DBP+10 μmol·L^-1 PGE2)和特异睾酮阻断剂组(40 μmol·L^-1氟他胺),分别处理细胞24 h;放射免疫法测定Leydig细胞睾酮水平;细胞免疫荧光和Western blotting法检测细胞中Cx43蛋白表达。结果:细胞染毒24 h后,与DMSO组比较,DBP组Leydig细胞中Cx43蛋白表达水平和睾酮水平均降低(P<0.05);与DBP组比较,DBP+PGE2组Leydig细胞中Cx43表达水平升高(P<0.05),但睾酮水平变化不明显(P>0.05);与DMSO组比较,氟他胺组Leydig细胞中睾酮水平和Cx43蛋白表达水平均明显降低(P<0.05)。结论:DBP对睾酮分泌及Cx43表达均有影响,Cx43对睾酮合成无抑制作用,而睾酮可反向调节Cx43表达;DBP对Leydig细胞睾酮合成抑制作用并不一定以Cx43为靶点。

Effects of dibutyl phthalate on expression of Cx43 protein and testosterone excretion in Leydig cells of testicle in adult rats

Objective To explore the effect of dibutyl phthalate(DBP) on the expression of Cx43 protein and testosterone excretion in the Leydig cells of testicle in the adult rats,and to disscuss whether there is relationship between the regulation of testosterone by DBP and Cx43.Methods After the adult SD rats were killed, the cultured primary pure Leydig cells were divided into solvent control group(0.1%DMSO),DBP group (50 mg·L^-1 DBP),Cx43 enhancer group(50 mg·L^-1 DBP+10 μmol·L^-1 PGE2),and specific testosterone blocker group (40 mg·L^-1 flutamid).After cultured for 24 h,radioimmunoassay was used to measure the levels of testosterone in the Leyding cells; cell immunofluorescence and Western blotting method were performed to detect the expressions of Cx43 protein in the Leydig cells.Results Compared with DMSO group,the level of testosterone and the expression level of Cx43 protein in the Leydig cells in DBP group were decreased after 24 h exposure(P<0.05).Compared with DBP group,the testosterone level in DBP+PGE2 group had no obvious change(P>0.05),but the expression level of Cx43 protein was increased(P<0.05).Compared with DMSO group,the level of testosterone and the expression level of Cx43 protein in flutamide group were decreased significantly(P<0.05).Conclusion DBP has the effects on the Cx43 expression and testosterone excreation,and Cx43 has no inhibitory effect on the synthesis of testosterone.However,testosterone can regulate the expression of Cx43 reversely.Therefore,it's not necessary to take Cx43 as the target of DBP in adjusting the synthesis of testosterone in Leydig cells.