Deciphering the Mechanisms by which Long Chain Fatty Acid Oxidation Influences Spatial Microvesicular Steatosis

PROJECT SUMMARY Nonalcoholic fatty liver disease (NAFLD) is associated with a 4-year decrease in lifespan and is expected to become the leading cause of liver related morbidity and mortality within the next 15 years. A hallmark of NAFLD is the accumulation of both small (microvesicular steatosis) and large (macrovesicular steatosis) lipid droplets in the liver. Interestingly, however, mechanisms driving the size and zonal deposition (e.g. accumulation in periportal versus pericentral hepatocytes) of lipid droplets remains poorly understood. The carnitine palmitoyltransferase 1a (Cpt1a) enzyme facilitates long chain fatty acid oxidation, but impairments in this pathway have been associated with microvesicular steatosis and reductions in circulating polyunsaturated fatty acids in humans. Therefore, this proposal aims to identify the mechanisms explaining the associations. Using a mouse model of impaired long chain fatty acid oxidation, preliminary data shows a sexually dimorphic response to liver-specific deletion of Cpt1a characterized by panlobular microvesicular steatosis in female mice while only slight periportal steatosis in male mice. In addition, female knockout mice develop exacerbated inflammation which coincides with a reduction in ω3-polyunsaturated fatty acids in the liver. Therefore, Aim 1 utilizes a gain- and loss-of-function approach to modulate Cpt1a expression specifically in periportal and pericentral hepatocytes across male and female mice. To assess the impact of zonal-specific Cpt1a deletion on the progression from NAFLD to the more severe nonalcoholic steatohepatitis (NASH), livers from the mice will be used for dual spatial transcriptomics and lipidomics, and for flow cytometry to assess the immune cell profiles in the liver. Aim 2 determines how ablation of Cpt1a in the liver impacts polyunsaturated fatty acid metabolism in the livers of male and female mice. The purpose of this aim is to uncover a previously unrecognized role of Cpt1a in regulating either the biosynthesis or degradation of these lipids. Completion of these aims will yield mechanistic insight linking fatty acid metabolism to spatial lipid deposition in the liver. The proposed research reveals several innovative mechanisms that have yet to be explored, including how Cpt1a alters lipid droplet formation across the periportal-pericentral axis in the liver. In addition, the proposed study uses innovative approaches including the use of zonal-specific gain- and loss-of-function mouse models in conjunction with next generation spatial lipidomic and transcriptomic technology to address the overall hypothesis. Strong collaborations among the University of Kentucky Arts and Sciences Imaging Center, Oncogenomics Shared Resource Facility, COCVD Analytical research Core, Biostatistics and Bioinformatics Shared Resource Facility, and the Center for Advanced Biomolecule Research at the University of Florida ensure successful completion of the proposed research by the candidate. This R01 will support the applicant in reaching his ultimate goal of becoming an independent yet collaborative, versatile PI leveraging his unique skill set in both basic and clinical research arenas to identify novel therapeutics to treat cardiometabolic disease.