Elucidating Molecular Mechanisms Linking Fructose to Cholesterol Metabolism

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PROJECT SUMMARY Fructose consumption is not only a major risk factor for development of non-alcoholic fatty liver disease (NAFLD), but also promotes hypercholesterolemia and atherosclerosis in humans and rodents. Identification of the mechanisms linking NAFLD to cardiovascular disease (CVD) remains poorly understand. This proposal focuses on identifying the influence dietary fructose has on synthesis and metabolism of cholesterol. Using a mouse model of sugar-sweetened beverage consumption, the candidate shows that fructose metabolism increases citrate, acetyl-CoA, and hepatic cholesterol levels. In addition, the candidate demonstrates fructose decreases the protein expression of carnitine palmitoyltransferase 1a (Cpt1a), a mitochondrial fatty acid transport protein. Moreover, conditional CPT1a knockout mice exhibit similar lipid perturbations as mice fed fructose. Therefore, aim 1 utilizes dual stable isotope techniques coupled with NMR and mass spectrometry to quantify cholesterol synthesis and fructose-derived carbon enrichment into the cholesterol biosynthetic pathway in male and female mice. Livers from the mice will be used for acetyl-proteomics to delineate potential mechanisms linking fructose to cholesterol biosynthesis. Aim 2 determines how transcriptional regulation of Cpt1a alters fructose-induced suppression of fatty acid oxidation and enhanced cholesterol synthesis using both in-vitro and in-vivo approaches. The purpose of this aim is to uncover a previous unrecognized role of Cpt1a in coordinating the regulation of both lipid-signaling pathways (fatty acid oxidation and cholesterol synthesis) in response to fructose. Completion of these aims will yield mechanistic insight linking dietary sugar metabolism to hypercholesterolemia. The novelty of the proposed research is the comprehensive dual-stable isotope approach in conjunction with analytical techniques to measure cholesterol synthesis and fructose-derived carbon enrichment into the sterol synthesis pathway in the same cohort of animals. In addition, the proposed research reveals several innovative mechanisms that have yet to be explored, including acetylation of cholesterol synthesis enzymes and regulation of Cpt1a through transcriptional mechanisms. Strong collaborations among the Metabolomics Core at the University of Kentucky, Mass Spectrometry Core at the Buck Institute for Research on Aging, and scientific advisory committee members ensure successful completion of the proposed research by the candidate. This research is complimented by a career development plan in which the candidate will learn new experimental methodology in stable isotope metabolomics, broaden his scientific network through attending workshops and conferences, and develop his communication skills so that he is poised to become an independent investigator. This K01 award will allow him to reach his long-term goals of establishing a well-funded laboratory studying dietary mechanisms in cardiometabolic disease.