论文部分内容阅读
目的:建立小鼠急性乙型肝炎病毒感染动物模型,在此基础上观察慢病毒载体携带小干扰RNA(small interfering RNAs,siRNAs)对乙型肝炎病毒(HBV)复制和表达的影响。方法:36只Balb/c小鼠随机等分为3组(n=12):①空白对照组(NC组):通过尾静脉注射pTHBV2(包含HBV全基因组真核表达质粒),建立小鼠急性HBV感染模型;②干扰组(E组):将pTHBV2与pWPT-HBV-siRNA1(携带靶向HBVS区的siRNA1的慢病毒)通过尾静脉共注射小鼠;③无关干扰组(I组):pTHBV2和pWPT-HBV-siRNA2(携带非靶向HBV的无关siRNA2的慢病毒)通过尾静脉共注射小鼠。于注射后第4天和第7天取小鼠血清和肝脏组织。采用ELISA法检测血清中HBsAg(HBV表面抗原);选择逆转录聚合酶链反应方法(RT-PCR)检测肝组织HBVS-mRNA水平;用免疫组织化学法检测肝组织HBcAg(HBV核心抗原)和HBsAg。结果:与NC组比较,E组小鼠血清中HBsAg水平在第4和第7天分别下降89.76%和94.81%(P<0.01);RT-PCR结果提示E组肝组织HBVS-mRNA水平明显降低(P<0.05);免疫组织化学法检测结果显示E组肝组织HBcAg﹑HBsAg阳性细胞数明显减少(P<0.05)。I组与NC组比较,上述检测指标无统计学差异(P>0.05)。结论:慢病毒载体携带siRNAs能够显著抑制小鼠体内HBV的复制和表达,并且抑制作用在第7天较第4天更强,没有减弱的趋势。
OBJECTIVE: To establish an animal model of acute hepatitis B virus infection in mice and observe the effect of lentiviral vector-carrying small interfering RNAs (siRNAs) on the replication and expression of hepatitis B virus (HBV). Methods: Thirty-six Balb / c mice were randomly divided into three groups (n = 12): ① blank control group (NC group): mice were injected with pTHBV2 (including HBV genome eukaryotic expression plasmid) HBV infection model; ②interference group (group E): Co-injection of pTHBV2 and pWPT-HBV-siRNA1 (lentivirus carrying siRNA1 targeting HBVS region) through the tail vein; ③Instantial interference group (group I): pTHBV2 And pWPT-HBV-siRNA2 (lentivirus carrying non-targeted siRNA2 unrelated siRNA2) were co-injected through tail vein. Mouse serum and liver tissue were taken on the 4th and 7th day after injection. The serum HBsAg (HBV surface antigen) was detected by ELISA, the level of HBV S-mRNA was detected by reverse transcription-polymerase chain reaction (RT-PCR), the expression of HBcAg (HBcAg) and HBsAg . Results: Compared with NC group, the level of HBsAg in serum of E group decreased by 89.76% and 94.81% on the 4th and 7th day, respectively (P <0.01). The result of RT-PCR indicated that the level of HBV-mRNA in the E group decreased significantly (P <0.05). The results of immunohistochemistry showed that the number of HBcAg and HBsAg positive cells in group E decreased significantly (P <0.05). There was no significant difference between the I group and the NC group (P> 0.05). CONCLUSION: Lentivirus vectors carrying siRNAs can significantly inhibit the replication and expression of HBV in mice, and the inhibitory effect is stronger on the 7th day than on the 4th day with no decreasing trend.