IFN-γ Prolongs The Persistence Of HBV And Its Mechanism

ObjectiveIt has been shown that IFN-y could prolong the persistence of HBV in hydrodynamic injection mouse model, however, the mechanism underling this phenomenon remains unclear. Therefore, in this study, we aimed to explore the virological and immunological mechanism mediating the persistence of HBV after IFN-y treatment.Methods1. pAAV/HBV1.2combined with pCMV2/IFN-null or pAAV/HBV1.2together with pCMV2/IFN-y were hydrodynamically injected (HI) into the tail veins of BALB/c. The serum and liver and spleen samples were collected at different time points. The HBV antigens and antibodies in sera were detected by ELISA, the expression of HBcAg in the liver was identified by immunohistochemistry, the replication of HBV and mRNA levels of the immune molecules were detected by Real-Time PCR, the HBV special cellular immune response was measured by ELISPOT, the expression of PD-1and Foxp3in the lymphocyte were analysed by FACS, the infiltration of inflammatory injury in the liver were obsever by H&E stain, the level of serum ALT was detected by IFCC. we aimed to investigate the effect of IFN-y on the persistence of HBV from special immune response and negative regulation of immunity.2. BALB/c mice were immunized with plasmid pcDNA3.1(+)/HBs using elecroporation method, two weeks after immunization, pAAV/HBV1.2and pCMV2/IFN-y were HI into BALB/c mice. Circulating HBV antigens and antibodies were detected by ELISA.3. To investigate the effect of IFN-y on the persistence of HBsAg, the BALB/c mice were injected with HBeAg/core-null pAAV/HBV1.2and pcDNA3.1(+)/HBs respectively. The serum samples were collected at different time points. The HBV antigens and antibodies were detected by ELISA.Results1. We found that the average duration of HBV was28,22.8and44.6days in pAAV/HBV1.2, pCMV2/EFN-null and pCMV2/IFN-y group, respectively. This result demonstrated that IFN-y could significantly prolong the persistent of HBV in BALB/c model.2. IFN-y had an anti-HBV effect. IFN-y inhibited the replication and expression of HBV at early stage, but prolonged the persistence of HBV at late stage.3. IFN-y inhibited the humoral immune response to HBV and the production of HBV antibody. IFN-y also inhibited the HBcAg-specific cellular immune responses at10day post injection (dpi), and there was no significant difference among those three gourps at20dpi, however, IFN-y promoted the HBcAg-specific cellular immune responses at30dpi. The HBsAg-specific cellular immune responses were not detected at10,20,30dpi.4. IFN-y did not increase the expression of PD-1and FoxP3in the lymphocyte of both liver and spleen, but, IFN-y increased the expression of PD-L1, NOS2, IDO and SOCS in the liver.5. IFN-y prolonged the persistence of HBV in both lower dose (2.5p.g) and higher dose (50μg) pAAV/HBV1.2HI mouse model.6. After immunized with pcDNA3.1(+)/HBs sucessfully, The mice were HI with pAAV/HBV1.2and pCMV2/IFN-y, IFN-y promoted the production of HBsAb and contributed to the clearance of HBV. 7. In the HBeAg/core-null pAAV/HBV1.2HI mouse model, IFN-y promoted the clearance of HBsAg (P=0.036). However, in the pcDNA3.1(+)/HBs HI mouse model, IFN-y had no effect on the clearance of HBsAg.Conclusion1. In the pAAV/HBV1.2injected BALB/c mouse model, IFN-y did not only inhibite the replication and expression of HBV at early stage, but also inhibite the humoral immune response and prolonged the persistence of HBV.2. In this study, there was no direct relationship between the HBcAg-specific immune responses and the clearance of HBV. In addition, IFN-y did not induce the negative immune regulation.3. The persistence of HBV was prolonged by IFN-y due to the inhibition of HBcAg immune responses and the production of HBsAb.