利用siRNA对β细胞上胰岛素受体的抑制作用

Inhibition of insulin receptor expression in β cells by siRNA

目的:利用siRNA抑制βTC-3细胞中胰岛素受体基因的表达.方法:化学合成胰岛素受体(IR)基因的4对特异siRNAs(实验组:siRNA-1组,siRNA-2组,siRNA-3组,siR-NA-4组)和1对非特异siRNA(NC组).通过阳离子脂质体将siRNAs分别转染βTC-3细胞,24 h后荧光显微镜下观察不同浓度siRNA转染细胞的效率;应用逆转录PCR检测IRmRNA表达.结果:①50,100,150,200 nmol/L siRNA转染βTC-3细胞24 h后,转染效率分别为(73.1±4.1)%,(92.9±3.4)%,(90.8±4.0)%,(93.9±2.8)%,选择100 nmol/L浓度作为siRNA转染浓度.②转染100 nmol/L siRNAs 24 h后,与对照组相比siRNA-1,2,3,4组βTC-3细胞IR mRNA表达都有不同程度下调,其中以siRNA-2组抑制效果最为明显,表达减少了70.5%.结论:靶向IR mRNA的siRNA能在体外特异性的抑制IR的表达,为进一步研究β细胞上IR的功能提供了实验基础.

AIM： To inhibit the expression of insulin receptor （IR） gene in mouse insulinoma β TC-3 cells by siRNA. METHODS： Four pairs of siRNA targeting IR gene, named siRNA-1, siRNA-2, siRNA-3, siRNA-4, and one pair non-specific siRNA （control group） were chemically synthesized and then transfected into 13 TC-3 cells using cation liposome transfection reagent. The efficacies of transfection were observed with fluorescence microscope, and the expression levels of IR mRNA were detected by reverse transcriptase-polymerase chain reaction after transfection for 24 h. RESULTS： ①The transfection efficacies with 50, 100, 150, 200 nmol/L siRNA into β TC-β cells were （73.1 ±4.1）%, （92.9±3.4）%, （90.8 ±4.0）%, （93.9± 2.8）% , respectively. Finally 100 nmol/L was chosen. ② After transfection with 100 nmol/L siRNAs for 24 h, the expression of IR mRNA in 13 TC-3 cells was decreased in all 4 groups compared with control group; among them, the effect was the most obvious in siRNA-2 group, in which the expression was decreased by 70.5%. CONCLUSION： The siRNAs targeting IR mRNA can effectively inhibit IR expression in vitro, which offers an experimental basis for further researches on IR in β cells.