shRNA SIRT3基因对乳腺癌细胞MCF-7侵袭能力的影响及机制研究

Effect of shRNA SIRT3 gene on invasive ability of breast cancer cell MCF-7 and its mechanism

目的:探讨shRNA SIRT3基因对乳腺癌细胞MCF-7侵袭能力的影响及机制。方法:以人乳腺癌细胞系MCF-7细胞为实验对象。按照处理方法将乳腺癌MCF-7细胞分成3组,即空白对照组(不转染慢病毒的乳腺癌MCF-7细胞组)、阴性对照组(乳腺癌MCF-7细胞转染无序序列慢病毒载体)和shRNA干扰组(乳腺癌MCF-7细胞转染shRNA SIRT3序列慢病毒载体)。采用实时荧光定量PCR和Western blotting等方法检测SIRT3和凋亡相关因子caspase-3、caspase-9的表达水平;采用Transwell试剂盒检测细胞的侵袭能力。结果:shRNA干扰组SIRT3 mRNA相对表达量均明显低于阴性对照组和空白对照组(P<0.01),caspase-3 mRNA、caspase-9 mRNA相对表达量均明显高于阴性对照组和空白对照组(P<0.01)。Western-blotting检测结果显示,shRNA干扰组SIRT3蛋白相对表达量均明显低于阴性对照组和空白对照组,caspase-3、caspase-9蛋白相对表达量均明显高于阴性对照组和空白对照组(P<0.01)。Transwell实验结果显示,shRNA干扰组迁移细胞数均明显少于阴性对照组和空白对照组(P<0.01)。结论:shRNA-SIRT3的慢病毒干扰载体可促进人乳腺癌细胞的凋亡,抑制人乳腺癌细胞的侵袭。

Objective:To explore the effect of shRNA SIRT3 gene on invasive ability of breast cancer cell MCF-7 and its mechanism.Methods:Human breast cancer cell line MCF-7 cells were used as experimental subjects.According to the treatment method,breast cancer MCF-7 cells were divided into three groups:blank control group(breast cancer MCF-7 cells without lentivirus transfection),negative control group(breast cancer MCF-7 cells transfected with disordered sequence lentivirus vector)and shRNA interference group(breast cancer MCF-7 cells transfected with shRNA SIRT3 sequence lentivirus vector).The expression levels of SIRT3 and apoptosis-related factors caspase-3 and caspase-9 were detected by RT-PCR and Western blotting,and the invasive ability of cells was detected by Transwell kit.Results:The relative expression of SIRT3 mRNA in the shRNA interference group was lower than that in the negative control group and the blank control group,and the relative expression of caspase-3 mRNA and caspase-9 mRNA was higher than those in the negative control group and the blank control group(P<0.01).The protein expression levels of SIRT3,caspase-3 and caspase-9 detected by Western blotting showed the same trend.The results of the Transwell experiment showed that the number of migrated cells in the shRNA interference group was less than that in the negative control group and the blank control group(P<0.01).Conclusions:The lentivirus interference vector shRNA-SIRT3 can promote the apoptosis of human breast cancer cells and inhibit the invasion of human breast cancer cells.