siRNA靶向抑制RP L6基因表达对人宫颈癌HeLa细胞增殖及凋亡的影响  

Effect of siRNA targeting RPL6 gene on the proliferation and apoptosis of human cervical cancer He La cells

作　　者：闫峰[1] 席艳妮[1] 

机构地区：[1]陕西榆林榆林市第二医院妇产科,719000

出　　处：《临床肿瘤学杂志》2015年第8期679-684,共6页Chinese Clinical Oncology

摘　　要：目的探讨小干扰RNA(siRNA)靶向抑制核糖体蛋白L6(RPL6)基因表达对人宫颈癌He La细胞增殖及凋亡的影响。方法优化siRNA转染条件,将2条靶向抑制RPL6基因的siRNA载体片段(siRPL6-1、siRPL6-2)分别高效转染人宫颈癌He La细胞(抑制组),同时设转染无义序列(si Control)的空转染组及不进行任何处理的对照组。分别于转染24、48、72、96 h后采用实时定量PCR(qRT-PCR)检测RPL6表达水平以筛选抑制率高的siRPL6载体用于后续实验。噻唑蓝比色(MTT)法检测各组转染24、48、72、96 h后的增殖抑制率,流式细胞术检测各组转染48、96 h后的细胞凋亡和细胞周期情况,Western blotting检测各组转染96 h后的cyclin A、CDK2和p21CIP1表达情况。结果抑制组的RPL6 mRNA水平均低于空转染组和对照组(P<0.05),siRPL6-2片段的干扰效率高于siRPL6-1,故后续实验选择siRPL6-2片段;与其余两组相比,抑制组转染后的增殖抑制率、凋亡率及caspase-3活化率均升高,且G0/G1期细胞比例升高,但S期和G2/M期细胞比例均降低,以上差异均有统计学意义(P<0.05);抑制组转染后的p21CIP1水平升高,cyclin A和CDK2水平降低,与空转染组和对照组的差异有统计学意义(P<0.05)。空转染组和对照组以上指标的差异均无统计学意义(P>0.05)。结论通过siRNA抑制RPL6基因表达可抑制增殖并诱导凋亡和细胞周期阻滞,对宫颈癌防治有一定价值。Objective To explore the effect of siRNA targeting RPL6 gene on the proliferation and apoptosis of human cervi-cal cancer HeLa cells. Methods After the optimization of the transfection conditions, 2 siRNA vector fragments （ siRPL6-1, siRPL6-2） targeting RPL6 gene were effectively transfected into HeLa cells of human cervical cancer（inhibition group）, respectively. Mean-while, there were also empty transfection group whose cells were transfected with the antisense sequence （ siControl） and control group without any treatment. The RPL6 expression levels were detected by real-time quantitative PCR （ qRT-PCR） at 24, 48, 72 and 96 h after transfection, and the siRPL6 vector with the higher inhibition rate was used for the following experiments. Thiazolyl blue （ MTT） was used to detect the proliferation inhibition rate at 24, 48, 72, 96 h after transfection. The apoptosis and cell cycle of the cells were detected by flow cytometry after 48 and 96 h. The expressions of CDK2, cyclin A and p21CIP1 after 96 h were detected by Western blot-ting. Results The level of mRNA RPL6 in the inhibition group was lower than that in the empty transfection group and the control group （ P〈0.05） . Since the interference efficiency of the siRPL6-2 fragment was higher than that of the siRPL6-1, so the subsequent trials chose the siRPL6-2 fragment. Compared with other two groups, the proliferation inhibitory rate, apoptosis rate and caspase-3 acti-vation rate together with the proportion of cells in G0/G1 phase increased, but the proportion of cells in S phase and G2/M phase de-creased in inhibition group with statistically significant difference （ P〈0.05）;After transfection, the levels of p21CIP1 increased but the levels of CDK2 and cyclin A were decreased in inhibition group versus other groups （ P〈0.05） . Conclusion The inhibition of RPL6 gene expression by siRNA can inhibit the proliferation and induce apoptosis and cell cycle arrest, and it can be valuable for the preven-tion and treatment of cervi

关 键 词：小干扰RNA 核糖体蛋白L6 宫颈癌 HELA细胞 增殖 凋亡 

分 类 号：R737.33[医药卫生—肿瘤]