Modulation of Host Cellular Pathways by HBV-Induced ROS in Modulating Autophagy and Inflammation in Hepatic Cells

Abstract

Background and Aim: Hepatitis B virus, a ds-DNA Virus, is one of the major risk for the development of chronic liver disease. HBV modulates several cellular pathways for its own survival and replication in the host cells. Reactive oxygen species (ROS) that are produced in the host cells during HBV infection, was considered to inhibit the cellular antioxidants and SIRT-1 expression. However, the role for HBV-induced ROS in modulating autophagy, proliferation and viral replication is not known. Hence, we hypothesized that HBV might modulate host factors through ROS in hepatic cells. Methods: HepG2.2.15 cells (Hep G2 cells expressing HBV genome stably) were treated with the N-acetyl cysteine (NAC, 20 μm) and Sirt-1 inhibitor Butyrate (5 mM) overnight. Western blotting was performed for the proteins SIRT-1, HBx, p-akt, akt, p-mTOR, mTOR, Beclin-1, LC-3, ATG5, COX-2, iNOS and β-actin using specific primary and secondary antibodies. H2-DCFDA was employed to analyse the intracellular release of ROS. Autophagy assay was done using the Cyto- ID autophagy detection kit. ELISA was used to determine HBsAg and real-time RT PCR was done to quantitate HBV-DNA. Results: HBV infection increased the ROS, COX-2, iNOS expression and formation of autophagic bodies in Hep G2.2.1.5 cells. Addition of NAC inhabited the inflammatory markers, autophagy and ROS production. Further, Western blot results showed that SIRT-1 beclin-1, LC3-II and ATG-5 expression were induced by ROS, which was inhibited by incubating cells with NAC, butyrate or SIRT-1 siRNA. Inhibition of the SIRT-1 expression by butyrate or SIRT-1 si-RNA inhibited the viral replication and HBV-induced autophagy. Conclusion: In this study, the data showed that HBV infection induces autophagy and inflammatory proteins via ROS production, which can be inhibited by butyrate or NAC treatment. It may be possible that butyrate might be considered for possible therapeutic applications. The authors have none to declare.