靶向SATB1基因的短发夹RNA诱导肺癌A549细胞凋亡

Short hairpin RNA-targeting special AT-rich sequence-binding protein-1 induces the apoptosis of human lung cancer cell line A549

目的:探讨靶向SATB1(special AT-rich sequence-binding protein-1)基因的短发夹RNA(short hairpin RNA,shRNA)对肺癌A549细胞凋亡的影响及其可能的机制。方法:构建靶向SATB1基因的shRNA重组质粒SATB1-shRNA,采用脂质体法将其转染至A549细胞;分别采用RT-PCR和蛋白质印迹法检测A549细胞中SATB1、Bcl-2、BaxmRNA和SATB1、Bcl-2、Bax、caspase3蛋白的表达水平;FCM检测A549细胞的凋亡率。结果:成功构建了SATB1-shRNA重组质粒;SATB1-shRNA转染A549细胞后,SATB1、Bcl-2 mRNA及其蛋白表达下调,Bax mRNA和Bax、caspase3蛋白表达上调(P<0.05)。SATB1-shRNA转染组细胞凋亡率[(14.18±1.59)%]较对照组[(1.84±0.57)%]明显增加(P<0.01)。结论:SATB1-shRNA可显著下调肺癌A549细胞中SATB1基因的表达水平并诱导细胞凋亡,其机制可能与下调Bcl-2基因表达所引起的级联效应有关。

Objective： To investigate the effect of short hairpin RNA （shRNA）-targeting special AT- rich sequence-binding protein-1 （SATB1） on the apoptosis of human lung cancer cell line A549, and to explore the possible mechanism. Methods： The recombined plasmid SATBI-shRNA of shRNA-targeting SATB1 was constructed and transfected into A549 cells by LipofectAMINE 2000. The expression levels of SATB1, Bcl-2 and Bax mRNAs were examined by RT-PCR, and the expression levels of SATB1, Bcl-2, Bax, and caspase 3 proteins were examined by Western-blotting. The apoptosis rate of A549 cells was detected by flow cytometry （FCM）. Results： The recombined plasmid SATBI-shRNA was successfully constructed and transfected into the A549 cells. After transfection with SATBI-shRNA, the expression levels of SATB1 and Bcl-2 mRNAs and proteins were decreased （P〈0.05）, whereas the expression levels of Bax mRNA and protein and the caspase 3 protein were increased （P〈O.05）. The apoptosis rate of A549 cells after transfection with SATBI-shRNA was higher than that of the A549 cells without transfection with SATBI-shRNA [（14.18±1.59） % vs （1.84±0.57） %, P〈0.01]. Conclusion： SATBI-shRNA can significantly down-regulate the expression level of SATB1 in human lung cancer cell line A549 and induce the apoptosis. The mechanism may be related to the cascade effect induced by the down-regulation of Bcl-2 expression.