解放军医学杂志2017，Vol.42Issue(11)：979-984,6.DOI:10.11855/j.issn.0577-7402.2017.11.08
微环DNA载体介导的shRNA在HBV转基因小鼠中的抗病毒作用
Antiviral activity of minicircle DNA-mediated shRNA in HBV transgenic mice
摘要
Abstract
Objective To investigate whether minicircle DNA-mediated shRNA can inhibit the replication and expression of HBV in HBV transgenic mice.Methods First,a universal plasmid shRNA,pU6-shRNA HBV and a minicircle shRNA vector pMC-U6-shRNA HBV targeting HBV gene was prepared by molecular cloning technique.Then,the HBV transgenic mice were divided into three groups and the pMC-U6-shRNA HBV,pU6-shRNA HBV and control vector pU6-control were transfected into them respectively by hydrodynamic injection (HDI).Sera were collected at different time points after injection.The changes of HBV DNA and HBsAg in serum of transgenic mice were detected by real-time PCR and chemiluminescence microparticle immunoassay (CMIA).Immunohistochemistry was used to analyze the expression of HBcAg in the liver of transgenic mice.Results Compared with pU6-control group,HBsAg and HBV DNA in serum of transgenic mice were significantly inhibited by pU6-shRNA HBV and pMC-U6-shRNA from day 7 to day 21 after HDI (P＜0.05).After that,the serum HBsAg and HBV DNA recovered and returned to the control level on the 35th day in pU6-shRNA HBV group.However,pMC-U6-shRNA HBV still maintained a strong inhibitory effect in vivo until 35 days post-injection compared with the other two groups (P＜0.05).Immunohistochemical results also suggest that pMC-U6-shRNA HBV could more significantly inhibit HBcAg expression than pU6-shRNA HBV in mouse liver tissue.Conclusion The minicircle DNA-based shRNA vector pMC-U6-shRNA HBV is of longer inhibitory effect on HBV replication and gene expression in HBV transgenic mice than pU6-shRNA HBV.Minicircle DNA is superior to pRNAT-U6.1/Neo for shRNA delivery in vivo.关键词
微环载体/shRNA/转基因小鼠/乙型肝炎Key words
minicircle vector/shRNA/transgenic mice/hepatitis B分类
医药卫生引用本文复制引用
李秀梅,刘光泽,谢勇,孔祥平..微环DNA载体介导的shRNA在HBV转基因小鼠中的抗病毒作用[J].解放军医学杂志,2017,42(11):979-984,6.基金项目
广东省科技计划项目(2014A020212639) （2014A020212639）
广州市科技计划项目(201510010004,201604046020)This work was supported by the Science and Technology Planning Project of Guangdong Province (2014A020212639) and the Science and Technology Program of Guangzhou (201510010004,201604046020) （201510010004,201604046020）
