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目的探讨邻苯二甲酸二(2-乙基己基)酯(di-2-ethylhexyl phthalate,DEHP)染毒对GC-2 spd细胞半胱氨酰天冬氨酸酶(caspase)蛋白及m RNA表达的影响。方法将体外培养的处于对数生长期的GC-2 spd细胞暴露于含终浓度为0(溶剂对照)、50、100、200μmol/L DEHP的培养基中培养24 h。采用Real-time PCR法检测细胞caspase-3、caspase-8、caspase-9 m RNA的表达,采用Western blotting法检测细胞caspase-3蛋白酶原(procaspase-3)、procaspase-8、procaspase-9的表达。结果与溶剂对照组比较,100μmol/L DEHP染毒组GC-2 spd细胞caspase-3 m RNA的表达水平和200μmol/L DEHP染毒组GC-2 spd细胞caspase-8 m RNA的表达水平及50、200μmol/L DEHP染毒组GC-2 spd细胞caspase-9m RNA的表达水平均较高,差异均有统计学意义(P<0.05)。与溶剂对照组比较,100、200μmol/L DEHP染毒组GC-2 spd细胞procaspase-3、procaspase-8、procaspase-9的表达水平均较低,差异均有统计学意义(P<0.05);且随着DEHP染毒浓度的升高,GC-2 spd细胞procaspase-3、procaspase-8、procaspase-9的表达水平均呈下降趋势。结论 DEHP体外染毒可能通过启动caspase-8和caspase-9进而激活下游caspase-3级联反应,最终诱导生精细胞凋亡。
Objective To investigate the expression of caspase and m RNA in GC-2 spd cells treated with di-2-ethylhexyl phthalate (DEHP) Impact. Methods GC-2 spd cells cultured in vitro at logarithmic growth phase were exposed to medium containing final concentrations of 0 (solvent control), 50, 100 and 200 μmol / L DEHP for 24 h. The expression of caspase-3, caspase-8 and caspase-9 mRNA were detected by Real-time PCR. The expressions of caspase-3, procaspase-8 and procaspase-9 were detected by Western blotting . Results Compared with the solvent control group, the expression level of caspase-3 mRNA in GC-2 spd cells exposed to 100 μmol / L DEHP and the expression of caspase-8 mRNA in GC-2 spd cells treated with 200 μmol / L DEHP and 50 The expression of caspase-9mRNA in GC-2 spd cells exposed to 200μmol / L DEHP was higher than that of control (P <0.05). Compared with the solvent control group, the expression levels of procaspase-3, procaspase-8 and procaspase-9 in GC-2 spd cells treated with 100,200 μmol / L DEHP were lower than those in the control group (all P <0.05) The expression levels of procaspase-3, procaspase-8 and procaspase-9 in GC-2 spd cells showed a decreasing trend with the increase of DEHP concentration. Conclusion DEHP can induce the apoptosis of spermatogenic cells by activating caspase-3 cascade and activating caspase-8 and caspase-9 in vitro.