靶向Bel-7402/ADM细胞MDR1基因高效siRNA的构建与鉴定

目的 构建并鉴定靶向人肝癌多药耐药细胞亚系Bel-7402/ADM细胞MDR1基因的高效siRNA,为逆转肝癌多药耐药提供新的分子靶点.方法 设计并化学合成4条靶向MDR1的siRNAs(mdrlsi-1、mdrlsi-2、mdrlsi-3和mdrlsi-4),通过LipofectamineTM2000介导转染Bel-7402/ADM细胞株,用RT-PCR检测Bel-7402/ADM细胞MDRlmRNA表达、western blot检测P-gp的表达,综合评价这4条siRNAs的沉默效果.结果 经siRNA干扰后,4条siRNAs均能不同程度逆转Bel-7402/ADM细胞MDR1介导的多药耐药,mdrlsi-1组mRNA和蛋白表达与其他组比较有显著性差异(P＜0.05).结论 相比较其他3条siRNAs,mdrl si-1对人肝癌Bel-7402/ADM细胞MDR1介导的耐药逆转效果最好,其靶序列有望成为逆转肝癌多药耐药新的靶点.

Construction and Identification of SiRNA Targeting to MDRl of Bel-7402/ADM

Objective To explore new molecular targets for the reversal of hepatocellular carcinoma multidrug resistance(MDR),efficient siRNA targeting MDRl gene of Bel-7402/ADM was constructed and identified.Methods Small interfering RNAs(siRNAs) of four(mdrlsi-1,mdrlsi-2,mdrlsi-3 and mdrlsi-4) targeting MDR1 were firstly designed and processed with chemical synthesis.Mediated by LipofectamineTM 2000,the siRNAs was transfected into Bel-7402/ADM cell strains.The mRNA expression of MDR-1 gene was evaluated by RT-PCR.And the product of P-glycoprotein(P-gp) was examined by Western blot.Based on these assays,the gene silencing effect of these four siRNAs was comprehensively evaluated.Results With the siRNA interference,four siRNAs reversed MDR in hepatocellular carcinoma mediated by MDR1 to varying degrees.Compared with other groups,mdr1 mRNA and P-gp expression in the mdrlsi-1 group presented significant difference(P<0.05).Conclusion: mdr1si-1 is a promising candidater for the reversal of MDR of hepatocellular carcinoma.