Viral and Lipid Regulation of Hepatocyte Signaling Pathways
Authors:   , Guarnieri Joseph W. , Bouchard Michael , Noguchi Eishi 
Published: July 2021

Primary liver cancer is the sixth most common cancer and the third highest cause of cancer-related deaths, globally. Hepatocellular carcinoma (HCC) is the predominate form of primary liver cancer, comprising 75-85% of the cases. Approximately 60% of HCCs are associated with a chronic hepatitis B virus (HBV) infection. Additionally, the increasing number of obese individuals in the United States has been linked to an increase in obesity-associated liver diseases such as non-alcoholic fatty liver disease (NAFLD). In Western countries, the incidence of NAFLD associated HCC is also on the rise and has been linked to 4-22% of HCC. In studies described here, we examined whether localization of the Hepatitis B Virus (HBV) HBx protein with mitochondria affects HBV replication and separately analyzed the interplay of changes in the gut microbiome and the liver transcriptome in the context of NAFLD development. HBx is required for efficient HBV replication and is thought to contribute to the development of HBV-associated HCC. Several studies have shown that a fraction of cytosolic HBx localizes to mitochondria in established cell lines and primary hepatocytes. Numerous studies have shown that HBx localizes to mitochondria and affects mitochondrial physiology. However, prior to our study, it was not known whether HBx affects on mitochondria or on HBV replication requires HBx localization to mitochondria. We now report that HBx localization to mitochondria is associated with efficient HBV replication in HepG2 cells and cultured primary rat hepatocytes. We also show that HBx localization to mitochondria is associated with activating signaling pathways directly or indirectly linked with HBx-induced elevation of cytosolic calcium levels, an essential function of HBx that is also required for HBV replication. Cumulatively, our research suggests that HBx association with mitochondria contributes to efficient HBV replication and is responsible for activating signaling pathways associated with HBx-induced elevation of calcium. NAFLD arises from the accumulation of lipids in hepatocytes, the major cell type of the liver, and the resulting liver inflammation as immune cells enter the liver in response to lipid accumulation. Recently, studies in mice and humans have shown that there are alterations in the composition of the gut microbiome in individuals with NAFLD as compared to healthy individuals. However, the mechanism by which the gut microbiome contributes to NAFLD is unclear. While previous studies characterized microbiome changes during NAFLD development, no previous studies have correlated changes in the gut microbiome with changes in liver gene expression. Feeding mice a high-fat diet (HFD) is a model system for studying obesity and associated metabolic disorders such as NAFLD. We characterized changes in gut bacteria and liver gene expression associated with the development of NAFLD in both male and female mice fed a HFD. We also analyzed the effect of non-fermentable fiber on the development of NAFLD. We analyzed the effect of supplementing the HFD and a standard diet (SD) with non-fermentable fiber on liver lipid content. Our research demonstrated that only SD supplemented with non-fermentable fiber resulted in decreased liver lipid content as compared to HFD, with or without non-fermentable fiber supplementation, and SD without non-fermentable fiber supplementation. While numerous studies have shown positive effects of supplementing diets with fiber, before our study, it was unknown whether supplementing diets with non-fermentable fiber could affect the accumulation of lipids in the liver and, consequently, the development of NAFLD. Our results suggest that fiber content, specifically non-fermentable fiber, in the diet is important in preventing the accumulation of lipids in hepatocytes and the development of NAFLD.