靶向HBX基因小干扰RNA抑制乙型肝炎病毒复制的实验研究

Study of the inhibition of HBV replication of the siRNA-targeted HBX gene

目的探讨HBX特异性RNA小干扰抑制乙型肝炎病毒复制的作用。方法用脂质体转染法将干扰质粒pSilencer3.1-shHBX和通用阴性对照质粒分别转染HepG2.2.15细胞,并设立未转染的空白对照组。分别于转染后24、487、2 h收集细胞培养上清液和细胞总蛋白,Western blot检测HBx蛋白表达情况,ELISA检测3组细胞上清中HBsAg和HBeAg表达情况。结果 Western blot检测显示,干扰质粒转染组HBx蛋白表达量逐渐降低,各时间点的表达量与空白对照组和通用阴性对照组比较差异均有统计学意义(P<0.01);ELISA检测显示,各时间点干扰质粒转染组细胞培养液上清中HBsAg和HBeAg的表达均下调,表达量与空白对照组和通用阴性对照组比较差异均具有统计学意义(P<0.01)。结论 HBX特异性小干扰RNA可抑制HBV复制,为siRNA作为抗乙肝病毒感染制剂提供了实验依据。

Objective To investigate the in vitro inhibition of HBV replication of the siRNA-targeted HBX gene.Methods A pSilencer3.1-shHBX plasmid and typical negative contrast plasmids were respectively transfected into HepG2.2.15 cells with lipofectamine and untransfected cells to serve as a blank control.Cell culture supernatant and total protein were collected after 24 h,48 h,and 72 h.The expression of protein HBx was detected by Western blot in HepG2.2.15 cells.The levels of HBsAg and HBeAg in the media of 3 groups of cells were detected by ELISA.ResultsWestern blot revealed that HBx proteins in the group transfected with the interference plasmid decreased gradually;compared to the blank control group and typical negative control group,is the difference in HBx proteins was statistically significant.ELISA showed that at each time point HBsAg and HBeAg in the supernatant of the group transfected with the interference plasmid decreased gradually;compared to the blank control group and typical negative control group,the difference in the levels of HBsAg and HBeAg was statistically significant.Conclusion A specific siRNA-targeted HBX gene can inhibit HBV replication and provide evidence for use of siRNA in antiviral agents.