蛋白酶体抑制剂MG132诱导卵巢癌SKOV3细胞凋亡和自噬的作用机制

The Mechanism of Proteasome Inhibitor MG132 Induces Ovarian Carcinoma Cells Autophagic and Apoptotic

目的:观察MG132对卵巢癌SKOV3细胞株生长的影响,以及自噬、凋亡相关因子的表达,初步探讨MG132抑制卵巢癌细胞生长的作用机制。方法:以0.5,1.5,2.5,3.5μg/mL浓度MG132作用于SKOV3细胞,采用噻唑蓝(MTT)比色法检测细胞生长情况;流式细胞术检测细胞凋亡率;免疫组织化学(IHC)、蛋白质印迹(Western blotting)、逆转录聚合酶链反应(RT-PCR)检测自噬Beclin1因子、凋亡Caspase-3因子的表达。结果:MG132作用SKOV3细胞后细胞生长受到抑制;MG132的作用随细胞浓度和时间逐渐增加,细胞凋亡率逐渐增加;IHC检测发现Beclin1、Caspase-3阳性表达;Western blotting检测发现Beclin1、Caspase-3、Bim、Bax蛋白表达增加,与MG132浓度的增加呈正相关,Bcl-2蛋白表达减少,与MG132浓度的增加呈负相关;MG132组Caspase-3和Beclin1的mRNA表达量均高于对照组(P<0.05)。结论:蛋白酶体抑制剂MG132对卵巢癌SKOV3细胞生长有抑制作用,呈浓度和时间依赖性,其抑制作用与凋亡和自噬有关。

Objective： To observe the growth of ovarian cancer SKOV3 cells after treated with proteasome inhibitor MG132, and the expression of autophagic and apoptotic factors, for further investigating the inhibited mechanism of MG132 on ovarian cancer cells. Methods： SKOV3 cells were treated with MG132 at the concentrations of 0.5 μg/m L, 1.5 μg/m L, 2.5 μg/m L and 3.5 μg/m L, then the growth of cells were detected by MTT assay; Apoptotic rates of cells were detected by flow cytometry;The expression of autophagic and apoptotic factors（Beclin1 and Caspase-3） in cells were detected by IHC, Western blotting and RT-PCR. Results： MTT assay demonstrated that the growth of SKOV3 cells were inhibited by MG132 and dependented on concentrations and time; FCM demonstrated that the apoptotic rates were gradually increased with the increased MG132 concentrations and time. Expression of Beclin1 and Caspase-3 were positive by IHC; Western blotting detected Caspase-3,Bim, Bax and Beclin1 were highly expressed while the expression of Bcl-2 was decreased in SKOV3 cells after treated with MG132. The expression of Beclin1, Caspase-3, Bim and Bax protein were positively correlated with MG132 concentrations, the expression of Bcl-2 was negatively correlated with concentrations; RT-PCR detected the m RNA relative quantity of Beclin1 and Caspase-3 in MG132 groups were both higher than the control group（P〈0.05）. Conclusions： The growth of ovarian carcinoma SKOV3 cells can be inhibited by proteasome inhibitor MG132 with concentration and time dependent. The inhibition effect is related with apoptosis and autophagy.