BRCA2基因沉默介导ATM信号通路对前列腺癌细胞增殖和凋亡的调控机制

Regulation Mechanism of BRCA2 Gene Silencing Mediating ATM Signaling Pathway on Proliferation and Apoptosis of Prostate Cancer Cells

目的探究BRCA2基因沉默介导ATM信号通路对前列腺癌细胞增殖和凋亡的影响。方法收集75例前列腺癌患者的癌组织和癌旁组织,采用免疫组化法分析不同组织中BRCA2蛋白表达的阳性率。细胞分组处理后,分别采用qRT-PCR和Western blotting法检测不同组织和细胞中BRCA2、ATM信号通路相关因子Chk2、p38MAPK及凋亡相关因子Bax、Bcl-2和caspase-3的mRNA和蛋白表达水平,CCK-8法和流式细胞仪检测各组细胞的增殖和凋亡情况。结果前列腺癌组织中的BRCA2表达水平显著高于癌旁组织(P<0.05)。与Blank组及NC组相比,BRCA2-siRNA组BRCA2、ATM、Chk2、p38MAPK、Bcl-2的mRNA和蛋白表达水平及p-ATM、p-p38MAPK、p-Chk2蛋白表达水平均显著下调,Bax和caspase-3的mRNA表达水平显著上调,Bax、cleaved-caspase-3和cleaved-caspase-9蛋白表达水平显著上调,且细胞增殖能力显著下降,凋亡率显著增加(P<0.05);ETP46464组BRCA2基因表达水平无明显变化(P>0.05),ATM、Chk2、p38MAPK、Bcl-2的mRNA和蛋白表达水平及p-ATM、p-p38MAPK、p-Chk2蛋白表达水平均显著下调,Bax和caspase-3的mRNA表达水平均显著上调,Bax、cleaved-caspase-3、cleaved-caspase-9蛋白表达水平显著上调,且细胞增殖能力显著下降,凋亡率显著增加(P<0.05);BRCA2-siRNA+ETP46464组BRCA2、ATM、Chk2、p38MAPK、Bcl-2的mRNA和蛋白表达水平及p-ATM、p-p38MAPK、p-Chk2蛋白表达水平均显著下调,Bax和caspase-3的mRNA表达水平均显著上调,Bax、cleaved-caspase-3、cleaved-caspase-9蛋白表达水平显著上调,且细胞增殖能力显著下降,凋亡率显著增加(P<0.05)。结论BRCA2在前列腺癌组织中呈高表达,BRCA2基因沉默可阻断ATM信号通路激活,从而抑制前列腺癌细胞增殖并促进其凋亡。

Objective To investigate the effect of BRCA2 gene silencing on the proliferation and apoptosis of prostate cancer cells by mediating ATM signaling pathway.Methods Cancer tissues and adjacent tissues of 75 patients with prostate cancer were collected.The positive rate of BRCA2 protein expression was analyzed by immunohistochemistry.After cell grouping,expression of BRCA2,ATM signaling pathway-related factors Chk2 and p38MAPK,and apoptosis-related factors Bax,Bcl-2 and caspase-3 in tissues and cells were analyzed by qRT-PCR and Western blotting.CCK-8 and flow cytometry were used to detect cell proliferation and apoptosis.Results BRCA2 expression in prostate cancer tissues was higher than that in adjacent tissues(P<0.05).Compared with Blank group and NC group,the mRNA and protein expression levels of BRCA2,ATM,Chk2,p38MAPK,and Bcl-2 in BRCA2-siRNA group were decreased significantly,so were the protein expression levels of p-ATM,p-p38MAPK and p-Chk2 in BRCA2-siRNA group.However,the mRNA expression levels of Bax and caspase-3 were increased,and protein expression levels of Bax,cleaved-caspase-3 and cleaved-caspase-9 were also increased markedly,accompanied by decreased cell proliferation ability and increased apoptosis(P<0.05).In ETP46464 group,there was no significant change in BRCA2 gene expression(P>0.05).But,the mRNA and protein expression levels of ATM,Chk2,p38MAPK,Bcl-2,and protein expression levels of p-ATM,p-p38MAPK and p-Chk2 were all decreased,while the mRNA levels of Bax and caspase-3 increased,and the protein levels of Bax,cleaved-caspase-3 and cleaved-caspase-9 increased.The cell proliferation ability was weakened and apoptotic rate was increased in ETP46464 group(All P<0.05).Furthermore,compared to the above two groups,the mRNA and protein expression levels of BRCA2,ATM,Chk2,p38MAPK and Bcl-2 were decreased significantly in BRCA2-siRNA+ETP46464 group.The protein expression levels of p-ATM,p-p38MAPK and p-Chk2 were also decreased in BRCA2-siRNA+ETP46464 group.However,the mRNA expression levels of Bax and caspase-3,and the protein expression levels of Bax,cleaved-caspase-3 and cleaved-caspase-9 were all increased,with remarkably decreased cell proliferation ability and increased apoptosis(All P<0.05).Conclusion BRCA2 is highly expressed in prostate cancer tissues.Silencing of BRCA2 could block the activation of ATM signaling pathway,so that could inhibit the proliferation and promote apoptosis of prostate cancer cells.