小干扰RNA有效抑制乙型肝炎病毒的动物实验

Inhibition of Hepatitis B virus replication by small interfering RNA in vivo

目的以乙型肝炎病毒(HBV)S区为靶位,观察小干扰RNA(siRNA)在动物体内抗HBV的效果。方法以流体动力学法建立HBV感染的动物模型,将pcDNA3．1-HBV和细胞体外实验证明有效的siRNA尾静脉共注射Balb/c小鼠,用时间分辨免疫荧光分析法(IFMA)检测小鼠血清中HBsAg,用定量聚合酶链反应法(FQ-PCR)检测血清HBV DNA,用逆转录聚合酶链反应(RT- PCR)法检测HBV S-mRNA,用免疫组织化学法检测肝组织HBsAg和HBcAg。结果在小鼠体内, siRNA能有效抑制HBsAg的分泌,降低HBV DNA的滴度,免疫组织化学结果也证实HBsAg、HBcAg阳性细胞数明显减少,干扰效果至少持续3d,而无关siRNA则无抑制作用。结论在动物体内靶向HBV S区的siRNA能有效特异抑制HBV。

Objective To evaluate the inhibitory effect of the small interfering RNA （siRNA） on hepatitis B virus （HBV） in vivo which targets HBV S gene region. Methods An animal model of HBV infection was developed hydrodynamically by injecting peDNA3. 1-HBV together with siRNA through the tail vein of Balb/e. HBsAg was analyzed by time-resolved immunofluorometrie assay. HBV DNA was analyzed by fluorogenie quantitative PCR （FQ-PCR）, HBV SmRNA was detected by senti-quantitative RT-PCR, and viral specific proteins （ HBsAg and HBcAg） in the liver were assayed by immunohistoehemieal staining. Results In the mice, the siRNA could effectively inhibit the seeretion of HBsAg, reduce the titers of HBV DNA, and immunohistoehemieal results also indicated that the number of HBsAg and HBeAg positive cells was reduced. The inhibitory effect of siRNA on HBV lasted 3 days at least. Conclusion These results demonstrate that the siRNA targeting HBV S gene region can substantially and specifically inhibit HBV replication and expression in vivo.