miR-34c逆转乳腺癌耐药细胞株MCF-7/DOX耐药性的研究

The Reversing and Molecular Mechanisms of mi R-34c on the MCF-7/DOX Cells with Doxorubicin-Resistance

miR-34在肿瘤发生发展中起着至关重要的作用,然而,mi R-34在肿瘤耐药中的作用研究不多。该研究将合成的mi R-34c成熟序列转染乳腺癌阿霉素(doxorubicin,DOX)耐药细胞MCF-7/DOX,探讨mi R-34c体外逆转MCF-7/DOX细胞耐药性作用及其可能的机制。采用Real-time RT-PCR检测mi R-34c在乳腺癌耐药细胞株MCF-7/DOX中的表达,MTS法检测miR-34c对MCF-7/DOX细胞阿霉素耐药性的影响,流式细胞术检测miR-34c对MCF-7/DOX细胞周期和凋亡的影响,Real-time RT-PCR和Western blot法检测多药耐药相关蛋白MDR、MRP以及细胞周期与凋亡相关蛋白Bcl-2、E2F3的表达。结果显示,mi R-34c在乳腺癌MCF-7/DOX耐药细胞中低表达,转染mi R-34c可明显增加耐药细胞对阿霉素的敏感性;流式分析发现,miR-34c可以促进耐药细胞G2期细胞周期阻滞和凋亡;与对照组相比较,miR-34c转染组细胞MDR、MRP蛋白表达无明显变化,而Bcl-2、E2F3 mRNA和蛋白表达均明显下调。研究表明,miR-34c直接靶向抑制Bcl-2和E2F3的表达,诱导细胞周期G2期阻滞和凋亡,进而增强MCF-7/DOX耐药细胞对阿霉素的敏感性。

mi R-34 played an important role in tumor development and progress, but there were few studies on the function of mi R-34 in drug resistance. To elucidate the function and mechanism of exogenous mi R-34 c involved in drug resistance in the breast cancer cells, mi R-34 c mimics were used to transfect into MCF-7/DOX cells with doxorubicin-resistance. The expression of mi R-34 c was analyzed by Real-time RT-PCR. Cell viability was analyzed by MTS assay. Cell cycle distribution and apoptosis were detected by PI and Annexin V/PI staining, respectively. The expressions of the drug-resistant related protein MDR and MRP, and cell proliferation and apoptosis related protein Bcl-2 and E2F3 were analyzed by Real-time RT-PCR and Western blot. Our results showed that mi R-34 c was significantly down-regulated in MCF-7/DOX cells, and overexpression of mi R-34 c increased sensitivity to doxorubicin in MCF-7/DOX cells. Flow cytometry analysis showed that mi R-34 c could induce G2 cell cycle arrest and cell apoptosis. Furthermore, mi R-34 c did not affect the expression of MDR and MRP, but sig-nificantly inhibited the m RNA and protein expression of Bcl-2 and E2F3. Our results suggested that mi R-34 c could directly target and inhibit Bcl-2 and E2F3 expression, and induced G2 cell cycle arrest and apoptosis, which in turn increased sensitivity to doxorubicin in MCF-7/DOX cells.