Research Summary/Abstract Non-alcoholic fatty liver disease (NAFLD) is one of the leading causes of chronic liver disease that can progress into a more advanced form- non-alcoholic steatohepatitis (NASH) featuring inflammation and fibrosis. There is no specific medicine for advanced NASH patients, making it critical to find supplements to mitigate liver damage. Non- psychoactive cannabinoids cannabidiol (CBD) and cannabigerol (CBG) has shown anti- inflammatory effect in other diseases, but their therapeutic effect in NAFLD/NASH is unknown. The goal of this project is to evaluate the efficacy of CBG with or without CBD intervention to reduce NAFLD/NASH. We have found that administration of CBG in diet-induced NAFLD/NASH leads to decreases in hepatic steatosis, fibrosis, immune cell infiltration, and cytokine secretion in the liver. Based on these data, we hypothesize that Cannabinoids attenuate immune cell recruitment, suppresses inflammatory mediator release, and thus mitigate liver damage caused by diet in mice. We propose three specific aims to address the hypothesis: (i) Evaluate the administration of CBG with or without CBD in alleviating the symptoms of NAFLD and NASH, specifically liver steatosis, inflammation, fibrosis, and oxidative stress, in two mouse models. (ii) Assess the expansion of T cells and mast cells across immune compartments. (iii) Evaluate crosstalk between PPARg activation and the TGF-β1 pathway in different cell populations in the liver. C57BL/6 mice will receive special diets to develop NAFLD/NASH symptoms and then be administered with CBG with or without the combination of CBD. The liver, peripheral blood, and spleen will be examined for cellular and secreted indicators of inflammation. Cytokines, including TGF-β1 and its pathway, will be evaluated in isolated hepatocytes, cholangiocytes and immune cell subsets via total RNA sequencing, multiplex cytometry bead assay, and qRT-PCR. Further, cannabinoid receptors and their downstream pathways will be evaluated with or without cannabinoid treatment in vivo and in vitro. We predict CBG alone or with CBD will protect the liver from NAFLD/NASH symptoms and inhibit immune cell infiltration into the liver via TGF-b1 involved pathway and PPARg (CBG receptor) activation. This research will provide therapeutic efficacy data about CBG and CBD for NAFLD/NASH for the first time in mice, which will benefit future cannabinoid-related animal and human studies. Further, we will provide in-depth information about molecular mechanisms in which CBG alone or with CBD diminishes NAFLD/NASH progression. Through these aims, we will build a rigorous body of evidence about the effect of cannabinoids during the pathogenesis of NAFLD/NASH.

Project narrative Obesity-induced non-alcoholic fatty liver disease (NAFLD) and NAFLD-induced liver cancer are increasingly prevalent in the United States and worldwide with no FDA-approved medicine developed. This project aims to investigate the efficacy of Cannabigerol (CBG, a non- psychoactive component of cannabis) with or without combination of cannabidiol (CBD) in treatment of NAFLD/NASH, considering promising preliminary results. Successful completion of the studies can provide basis for a prospective treatment for chronic liver diseases and expand our understanding on how these diseases progress.