RNA干扰技术沉默MDR1基因逆转大肠癌耐药细胞株LoVo/5-Fu的耐药性

The reversal effect on MDR1 gene-mediated multidrug resistance in human colon carcinoma LoVo/5-Fu cells by RNA interference

目的探讨多药耐药(MDR1)基因沉默对大肠癌耐药细胞系LoVo/5-Fu耐药性的逆转作用。方法构建靶向MDR1的短发夹RNA(EASY-shRNA-MDR1)干扰质粒转染LoVo/5-Fu,实时荧光定量PCR(Realtime-PCR)在mRNA水平检测抑制效果,噻唑蓝(MTT)法检测细胞对5-Fu的敏感性,流式细胞仪检测细胞凋亡情况。结果与未转染组相比,EASY-shRNA-MDR1组细胞MDR1 mRNA表达明显下调(P<0.05);细胞对5-Fu的IC50及RI显著降低(P<0.05),敏感性的相对逆转率为74.7%;凋亡率明显升高(P<0.05)。结论 EASY-shRNA-MDR1可有效抑制大肠癌耐药细胞中MDR1的表达,进而降低了大肠癌耐药株对5-Fu的耐药性。

Objective To investigate the reversal effect on MDR1 gene-mediated multidrug resistance in human colon carcinoma LoVo/5-Fu cells by RNA interference. Methods A eukaryotic expression plasmid of shRNA targeting MDR1 was transiently transfected into human colon carcinoma LoVo/5-Fu cells. Expression of MDR1 mRNA was detected by real-time quantitative PCR. Drug sensitivity was measured by MTT. The apoptosis of cells was determined by flow cytometry assay. Results After transfection with EASY-shRNA-MDR1,compared with the un-transfected group,expression of MDR1 mRNA in LoVo/5-Fu cells was obviously reduced （P0.05）; IC50 of 5-Fu and the resistance indexes were obviously decreased（P0.05）; the relative reverse rate of sensitivity of LoVo/5-Fu cells to 5-Fu was 74.7%; and the apoptosis rate significantly increased（P0.05）. Conclusion MDR1 shRNA effectively inhibit expression of MDR1,thus reverse MDR1 gene-mediated multidrug resistance in human colon carcinoma LoVo/5-Fu cells.