Mechanisms of HIF1 alpha mediated dysregulated skeletal muscle proteostasis in alcoholic liver disease

The prevalence of alcohol use disorders and consequent tissue injury, primarily alcoholic liver disease (ALD) continue to increase. Skeletal muscle loss or sarcopenia is a consistent abnormality in patients with ALD and is associated with adverse clinical outcomes that include increased mortality, other complications of liver disease and poor post-liver transplant outcomes. We have recently reported more severe muscle loss and a greater rate of muscle loss in patients with alcoholic cirrhosis compared with those in other causes of cirrhosis. Despite the high clinical significance of sarcopenia in ALD there are no effective therapeutic options because the underlying mechanisms are not well understood. We also reported that ethanol, directly and indirectly via impaired hepatic ammonia disposal and consequent hyperammonemia, results in a sarcopenic phenotype with dysregulated protein homeostasis (proteostasis). In preliminary studies, we have shown mitochondrial dysfunction in response to ethanol and hyperammonemia. We also noted that ethanol results in cataplerosis or loss of tricarboxylic acid (TCA) cycle intermediates, specifically α-ketoglutarate (αKG) an inhibitor of HIF1α. Consistently, unbiased approaches (assay for transposase accessible chromatin sequencing), and targeted experiments showed oxygen independent stabilization of muscle hypoxia inducible factor-1α (HIF1α) with ammonia. In pilot studies, we observed an increased expression of REDD1, a transcriptional target of HIF1α and a negative regulator of the mammalian target of rapamycin complex 1 (mTORC1) that maintains skeletal muscle proteostasis with functional responses. We also noted relative preservation of muscle mass during hyperammonemia in muscle specific deletion of HIF1α mice. These observations formed the basis for our hypothesis that ethanol induced hyperammonemia causes cataplerosis of αKG with oxygen independent stabilization and impaired proteostasis and sarcopenia. We will test this hypothesis by testing if ethanol stabilizes HIF1α in skeletal muscle, and determine the mechanisms of stabilization of muscle HIF1α. Ethanol treatment in vitro in myotubes and in vivo in mice with loss and gain of function of HIF1α and its regulatory molecules will be used for these studies We will also test how metabolic perturbations regulate HIF1α stabilization and consequent molecular and functional responses in our preclinical models. Validation of key observations will be done in human muscle tissue from our biorepository. The proposed studies will enhance our understanding of the mechanisms of sarcopenia in ALD and lay the foundation for targeted therapeutics. This award will provide the support and time for the applicant for a supervised research career development in translational research. The applicant works with NIAAA funded independent investigators in the Northern Ohio Alcohol Center and her mentor developed the field of sarcopenia in liver disease. The institutional environment is highly supportive of her career path towards becoming an independent physician scientist focusing on mechanistic approaches to address unmet clinical needs in patients with alcohol use disorders.