HBV induces liver fibrosis through the generation of reactive oxygen species in a pyruvate-dependent manner

Background and Aims: Chronic hepatitis B (CHB) is a major etiology of liver cirrhosis. We previously found that pyruvate, a key intermediate in many metabolic pathways, increases HBV replication. However, the mechanism by which pyruvate mediates HBV-induced liver fibrosis is not well characterized. We hypothesize that HBV induces liver fibrosis through a pyruvate-peroxisome proliferator-activated receptor α (PPARα) - reactive oxygen species (ROS) pathway. Approach and Results: We evaluated HBV-induced fibrogenesis in HepAD38, HBV-infected NTCP-HepG2, primary human hepatocytes (PHHs), and hepatic stellate cell lines (HSC, LX2) in mono- and co-culture models and in patient sera. We also evaluated the effects of PPARα agonist/antagonist and ROS inhibition on HBV-induced liver fibrosis in cell culture, HBV carrier mouse, HBV-Transgenic mouse (HBV-Tg), humanized liver mouse, and human precision-cut liver slice (PCLS) models. We found that HBV increased pyruvate levels and fibrosis-related gene expression in both HBV-infected hepatocytes and patient sera. Supernatants from HBV-infected cells (HBVsup) and pyruvate supplementation independently and additively increased expression of profibrotic genes (TGF-β1, TIMP-1, COL1A1, and α-SMA), activated ROS production, and inhibited PPARα expression. Notably, PPARα inhibition and siRNA knockdown increased ROS production and profibrogenic gene expression, while activation of PPARα blocked HBVsup- and pyruvate-induced ROS generation and fibrogenesis. ROS was confirmed to be downstream of PPARα-related fibrogenesis, as ROS inhibition abrogated HBVsup-, pyruvate supplementation-, or PPARα inhibition-induced liver fibrosis. Conclusions: HBV infection induces pyruvate production and decreases PPARα expression, leading to increased ROS generation and liver fibrosis. Pyruvate and PPARα represent novel targets for antifibrotic therapeutic development in CHB.