Institution: | 1. Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA;2. Proteomics and Metabolomics core, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA;3. Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA
Departments of Pulmonary Medicine, Cleveland Clinic, Cleveland, Ohio, USA;4. Cardiovascular and Metabolic Sciences, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA;5. Department of Pharmacology, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA;6. Department of Inflammation and Immunity, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA
Critical Care Medicine, Respiratory Institute, Cleveland Clinic, Cleveland, Ohio, USA |
Abstract: | Perturbed metabolism of ammonia, an endogenous cytotoxin, causes mitochondrial dysfunction, reduced NAD+/NADH (redox) ratio, and postmitotic senescence. Sirtuins are NAD+-dependent deacetylases that delay senescence. In multiomics analyses, NAD metabolism and sirtuin pathways are enriched during hyperammonemia. Consistently, NAD+-dependent Sirtuin3 (Sirt3) expression and deacetylase activity were decreased, and protein acetylation was increased in human and murine skeletal muscle/myotubes. Global acetylomics and subcellular fractions from myotubes showed hyperammonemia-induced hyperacetylation of cellular signaling and mitochondrial proteins. We dissected the mechanisms and consequences of hyperammonemia-induced NAD metabolism by complementary genetic and chemical approaches. Hyperammonemia inhibited electron transport chain components, specifically complex I that oxidizes NADH to NAD+, that resulted in lower redox ratio. Ammonia also caused mitochondrial oxidative dysfunction, lower mitochondrial NAD+-sensor Sirt3, protein hyperacetylation, and postmitotic senescence. Mitochondrial-targeted Lactobacillus brevis NADH oxidase (MitoLbNOX), but not NAD+ precursor nicotinamide riboside, reversed ammonia-induced oxidative dysfunction, electron transport chain supercomplex disassembly, lower ATP and NAD+ content, protein hyperacetylation, Sirt3 dysfunction and postmitotic senescence in myotubes. Even though Sirt3 overexpression reversed ammonia-induced hyperacetylation, lower redox status or mitochondrial oxidative dysfunction were not reversed. These data show that acetylation is a consequence of, but is not the mechanism of, lower redox status or oxidative dysfunction during hyperammonemia. Targeting NADH oxidation is a potential approach to reverse and potentially prevent ammonia-induced postmitotic senescence in skeletal muscle. Since dysregulated ammonia metabolism occurs with aging, and NAD+ biosynthesis is reduced in sarcopenia, our studies provide a biochemical basis for cellular senescence and have relevance in multiple tissues. |