Construction And Preliminary Application Of HBV-Tolerant Mice Model

Heptitis B virus(HBV)is the leading cause of acute/chronic hepatitis,cirrhosis and hepatic carcinoma(HCC).Under the influence of vaccination for prophylaxis,there were still approximately 350 million person in worldwide who were infected by HBV and at risk of chronicity and developing end-stage liver disease and HCC.Until now there isn’t effective medicine for therapeutic.Because lack of suitable animal model,the mechanisms of cirrhosis and HCC related with HBV infection are still unclear.As previous reports,the murine model which was administrated with hydrodynamic injection(HDI),was supporting HBV persistent expression.While it can highly mimic the state of HBV infection in human,and it will be used for HBV related research.So it’s necessary to construct an efficient HBV-tolerant murine model with HDI.To construct HBV-tolerant murine model,we firstly examined and compared the expression of HBV DNA in different mouse strains.According to the analysis of background of mouse strains,BALB/c,C57BL/6 and C3,B6-Tg(AAVS1)AlXob/J(AAVS1)were chose to assess in current study.All the 4 mouse strains were bred and reproduced in the lab.And lOμg of pAAV-1.2HBV DNA was injected into tail vein with HDI.The results showed that concentration of HBsAg in AAVS1 serum was the highest up to 103-4 IU/ml during the 4 mouse strains.AAVS1 mice could be divided into two groups by sequencing,AAVS1 TG(human AAVS1 locus positive)and AAVS1-/-(human AAVS1 locus negative).Further studies showed there is not significantly difference in HBsAg in both AAVS1 TG and AAVS1-/-serum.It’s suggested that AAVS1 locus has no effect on the expression of HBV.In further studies,we try to test if the cloning vector related with persistent expression of HBV in mice.The HBV DNA sequences in pAAV-1.2HBVwere cloned into pcDNA3.1.Monitoring the concentration of HBsAg in mice serum,they were cleared up 1 month later in the mice which were HDI with pcDNA3.1-1.2HBV.The results showed that the host genetic background is not the only factor affecting HBV persistent expression in hepatocytes,but instead the AAV vector also helps to maintain HBV persistence in the mouse liver.Then to test if the HBcAg could regulate the expression of HBV DNA in mice,PAAV-1.2HBV(HBcAg+)or PAAV-1.2HBV-core-null(HBcAg-)were injected with HDI in tail vein in the mice.As result shown,compared to HBcAg+ group,the concentration of HBsAg is 100-folder higher in HBcAg-group.It suggested that HBcAg could negatively regulate the expression of HBV DNA in mice.HBcAg may play the key role in breaking the persistence of HBV.Even more,we assess if the gender of the mice affect the persistent expression of HBV DNA in HBV-resistant murine model.The results demonstrated that the HBsAg in male mice is higher than that in female mice.Finally,HBV targeted SiRNA antiviral therapeutic was evaluated with HBV-tolerant murine model.It was proved that this kind of HBV-tolerant mice model have great value and it can be used to imitate the HBV chronic infection state in human.It will help understand the mechanism of HBV chronic infection and also can evaluate and screen the new antiviral drugs.