摘要
目的:利用流体动力学法构建小鼠HBV感染模型,观察pSiHBV/P在体内对HBV的抑制作用.方法:以流体动力学法构建小鼠HBV感染模型,将pSiHBV/P与pHBV1.3尾静脉共注射Balb/c小鼠.转染后第6天,用酶联免疫分析法检测小鼠血清乙肝表面抗原(HBsAg),用荧光定量PCR检测血清HBVDNA的水平,用免疫组织化学方法检测肝内HBsAg、乙肝病毒核心抗原(HBcAg)的表达,RT-PCR法半定量检测肝内HBV3.5kb mRNA、S-mRNA及0.7kb mRNA的水平.结果:感染组和无关干扰组血清HBsAg表达量平均值分别为9.11±3.34和8.19±5.41,pSiHBV/P干扰组平均值1.94±1.78;pSiHBV/P干扰组血清中HBV DNA的含量较感染组明显下降(2.91±0.55vs4.32±0.57,P<0.05),抑制率为33%;干扰组肝细胞中HBsAg和HBcAg阳性细胞数较对照组明显减少;干扰组3.5kb mRNA、S-mRNA的水平较感染组和无关干扰组均有明显下降(0.48±0.19vs1.06±0.40,0.88±0.54;0.46±0.18vs1.05±0.38,0.90±0.51,P<0.05).结论:RNAi技术在体内能高效、特异地抑制小鼠体内HBV.
AIM： To develop a mouse model of hepatitis B virus （HBV） infection by hydrodynamic method and to observe the inhibition and replication of HBV induced by pSiHBV/P in the model. METHODS： Hydrodynamic method was used to establish the model of HBV infection in Balb/c mice. pHBV1.3 was injected into the mice via tail veins together with pSiHBV/P, which could specifically target the P gene sequence of HBV by synthesizing small interfering RNA （siRNA） in vivo. On the 6th day of transfection, blood samples liver specimens were collected. Serum HBsAg were analyzed by enzyme-linked immunosorbent assay （ELISA）, and the contents of HBsAg and HBcAg in liver specimens were measured by immunohistochemical staining. HBV DNA and mRNA levels were detected by fluorescence quantitative polymerase chain reaction （FQ-PCR） and semi-quantitative RT-PCR. RESULTS： The mean levels of serum HBsAg expression in the infected group, the irrelative interference group and the pSiHBV/P interfering group were 9.11 ± 3.34, 8.19 ± 5.41 and 1.94 ± 1.78, respectively. The HBV DNA loads were reduced by 33% in the pSiHBV/P interfering group, sig- nificantly lower than that in the infected group （2.91 ± 0.55 vs 4.32 ± 0.57, P 〈 0.05）. The count of HbsAg- or HbcAg-positive cells in the pSiHBV/ P interfering group was lower than that in the control groups. The levels of 3.5 kb mRNA and S-mRNA in the pSiHBV/P interfering group were reduced remarkably in comparison with those in the infected group and the irrelative interference group （0.48 ± 0.19 vs 1.06 ± 0.40, 0.88 ± 0.54; 0.46 ± 0.18 vs 1.05 ± 0.38, 0.90 ± 0.51; all P 〈 0.05）. CONCLUSION： RNAi can inhibit HBV expression and replication effectively and specifically in vivo.
出处
《世界华人消化杂志》
CAS
北大核心
2008年第22期2448-2452,共5页
World Chinese Journal of Digestology
基金
四川省科技厅资助项目
No.05JY029-145~~
关键词
肝炎病毒
模型
动物
基因治疗
RNA干扰
Hepatitis B virus
Models
Animal
Gene therapy
RNA interference
