RNA干扰survivin基因对宫颈癌细胞中survivin蛋白表达及caspase-3酶活性的影响

The effects of RNA interference survivin gene on the expression of survivin protein and caspase-3 enzyme activity in cervical cancer cells

目的观察RNA干扰survivin基因对宫颈癌细胞中survivin蛋白表达及caspase-3酶活性的影响。方法针对survivin mRNA序列设计合成3对编码小干扰RNA(siRNA)的DNA模板,构建shRNA重组质粒,转染Hela细胞,筛选获得转染效率最高的阳性克隆,阴性对照为阳性克隆随机打乱序列。Western blot方法检测survivin蛋白表达,分光光度法检测caspase-3酶活性。结果干扰48 h时,siRNA-168载体干预组干预效果最好,siRNA阳性质粒干扰组(实验组)survivin蛋白表达量为阴性对照组中survivin蛋白表达量的13%,是空白组survivin蛋白表达量的16%。干扰后caspase-3活性表达升高,实验组与阴性对照组、实验组与空白组相比差异均有统计学意义(P<0.05)。结论siRNA可有效抑制Hela细胞survivin蛋白表达,升高caspase-3酶活性。

Objective To investigate the effect of RNA survivin interference gene on the expression of survivin protein and the activity of caspase-3 enzyme in cervical cancer cells. Methods According to the survivin mRNA sequence, 3 pairs of DNA template for small interference RNA （siRNA） was designed and synthetized, shRNA recombination plasmid was built, Hela cells was transfected. The positive clones with the highest transfection efficiency was isolated, and negative control was the randomly upseted sequence of positive clones. Western blot method was used to detect the expression of survivin protein and spectrophotometry was used to detect the activity of caspase-3 enzyme. Results At 48 h of interference, the siRNA-168 vectors intervention group had the best effect. The survivin protein expression of the siRNA positive plasmid interfern group （experimental group） was 13% of the negative control group, which was 16% of the blank control group. After interference, the activity of caspase-3 rised and there was significant difference not only between experimental group and negative control group, but also between experimental group and blank control control （P 〈 0.05）. Conclusion siRNA can inhibit the expression of survivin protein in Hela cells effectively and increase caspase-3 activity.