邻苯二甲酸二乙基己酯在体外对大鼠卵巢颗粒细胞凋亡Caspase通路的影响

Effects of Di-2-ethylhexyl Phthalate on the Apoptosis Caspase Pathway in Rat Ovarian Granulosa Cells in Vitro

目的通过对原代培养的大鼠卵巢颗粒细胞进行邻苯二甲酸二乙基己酯(di-2-ethylhexylphthalate,DEHP)染毒,探讨DEHP在体外对大鼠卵巢颗粒细胞凋亡通路Caspase-3和Caspase-9基因表达以及细胞凋亡的影响。方法分离和培养原代未成熟大鼠的卵巢颗粒细胞,用不同浓度的DEHP(0、10、50、100 nmol/L)染毒24 h。荧光定量PCR法检测颗粒细胞的Caspase-3和Caspase-9基因mRNA表达水平,Weastern blot检测颗粒细胞Caspase-3和Caspase-9蛋白表达水平,流式细胞术检测颗粒细胞凋亡率。结果与对照组相比,各DEHP处理组卵巢颗粒细胞Caspase-3基因mRNA和蛋白表达水平下降(P<0.05),Caspase-9基因mRNA和蛋白表达水平也降低(P<0.05),同时细胞凋亡率增加(P<0.05),呈浓度依赖关系。结论 DEHP可通过激活Caspase通路而诱导大鼠卵巢颗粒细胞凋亡,进而影响卵巢功能。

Objectives To investigate the effects of di-2-ethylhexyl phthalate （DEHP） on the expression of Caspase-3 and Caspase-9 genes, and on the apoptosis of rat ovarian granulosa cells in vitro. Methods The ovarian granulosa ceils of rats were separated ; the primary cultured granulosa ceils were treated with different concentrations of DEHP （0, 10, 50 and 100 nmol/L）. The real time PCR was used to detect the mRNA of Caspase-3 and Caspase-9 genes, and the Western blot was used to detect the protein of Caspase-3 and Caspase-9 genes. Cell ap- optosis rate was estimated by flow cytometry. Results Compared with the control group, the expressions of Caspase-3 and Caspas-9 gene mRNAs and proteins in different DEHP-treated groups increased（P 〈 0.05） in a con- centration dependent manner. The cell apoptosis rate in different DEHP-treated groups increased （ P 〈 0.05 ） in a concentration dependent manner. Conclusion DEHP could induce the apoptosis of ovarian granulosa cells through increasing the activity of caspase pathway, which results in ovarian dysfunction.