Robust protein nitration contributes to acetaminophen-induced mitochondrial dysfunction and acute liver injury
3-Nitrotyrosine; Acetaminophen; Acute liver injury; Biochemistry & Molecular Biology; covalent binding; CYP2E1; Endocrinology & Metabolism; Free radicals; immunohistochemical localization; induced hepatotoxicity; knockout mice; Mitochondrial dysfunction; mouse-liver; n-acetylcysteine; n-acetylcysteine; nitration; nitric-oxide synthase; oxidative stress; protein; superoxide-dismutase; terminal kinase
Acetaminophen (APAP), a widely used analgesic/antipyretic agent, can cause liver injury through increased nitrative stress, leading to protein nitration. However, the identities of nitrated proteins and their roles in hepatotoxicity are poorly understood. Thus, we aimed at studying the mechanism of APAP-induced hepatotoxicity by systematic identification and characterization of nitrated proteins in the absence or presence of an antioxidant, N-acetylcysteine (NAC). The levels of nitrated proteins markedly increased at 2 h in mice exposed to a single APAP dose (350 mg/kg ip), which caused severe liver necrosis at 24 h. Protein nitration and liver necrosis were minimal in mice exposed to nontoxic 3-hydroxyacetanilide or animals co-treated with APAP and NAC. Mass-spectral analysis of the affinity-purified nitrated proteins identified numerous mitochondrial and cytosolic proteins, including mitochondrial aldehyde dehydrogenase, Mn-superoxide dismutase, glutathione peroxidase, ATP synthase, and 3-ketoacyl-CoA thiolase, involved in antioxidant defense, energy supply, or fatty acid metabolism. Immunoprecipitation followed by immunoblot with anti-3-nitrotyrosine antibody confirmed that the aforementioned proteins were nitrated in APAP-exposed mice but not in NAC-cotreated mice. Consistently, NAC cotreatment significantly restored the suppressed activity of these enzymes. Thus, we demonstrate a new mechanism by which many nitrated proteins with concomitantly suppressed activity promotes APAP-induced mitochondrial dysfunction and hepatotoxicity. Published by Elsevier Inc.
Abdelmegeed M A; Jang S; Banerjee A; Hardwick J P; Song B J
Free Radical Biology and Medicine
2013
2013-07
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1016/j.freeradbiomed.2013.02.018" target="_blank" rel="noreferrer noopener">10.1016/j.freeradbiomed.2013.02.018</a>
Role of peroxisome proliferator-activated receptor-alpha in fasting-mediated oxidative stress
Aldehyde dehydrogenase; Biochemistry & Molecular Biology; differential expression; dismutase; Endocrinology & Metabolism; Fasting; fatty-acid oxidation; glutathione-s-transferase; hepatic steatosis; Lipid peroxidation; Lipid peroxidation; liver; manganese-superoxide-dismutase; mitochondrial aldehyde dehydrogenase; nitric-oxide; Null mice; oxidative stress; PPAR-alpha; PPAR-alpha; Protein nitration; Protein oxidation; rat-liver; Steatosis; Superoxide
The peroxisome proliferator-activated receptor-alpha (PPAR alpha) regulates lipid homeostasis, particularly in the liver. This study was aimed at elucidating the relationship between hepatosteatosis and oxidative stress during fasting. Fasted Ppara-null mice exhibited marked hepatosteatosis, which was associated with elevated levels of lipid peroxidation, nitric oxide synthase activity, and hydrogen peroxide accumulation. Total glutathione (GSH), mitochondrial GSH, and the activities of major antioxidant enzymes were also lower in the fasted Ppara-null mice. Consequently, the number and extent of nitrated proteins were markedly increased in the fasted Ppara-null mice, although high levels of protein nitration were still detected in the fed Ppara-null mice while many oxidatively modified proteins were only found in the fasted Ppara-null mice. However, the role of inflammation in increased oxidative stress in the fasted Ppara-null mice was minimal based on the similar levels of tumor necrosis factor-alpha change in all groups. These results with increased oxidative stress observed in the fasted Ppara-null mice compared with other groups demonstrate a role for PPAR alpha in fasting-mediated oxidative stress and that inhibition of PPAR alpha functions may increase the susceptibility to oxidative damage in the presence of another toxic agent. Published by Elsevier Inc.
Abdelmegeed M A; Moon K H; Hardwick J P; Gonzalez F J; Song B J
Free Radical Biology and Medicine
2009
2009-09
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1016/j.freeradbiomed.2009.06.017" target="_blank" rel="noreferrer noopener">10.1016/j.freeradbiomed.2009.06.017</a>
Increased Nitroxidative Stress Promotes Mitochondrial Dysfunction in Alcoholic and Nonalcoholic Fatty Liver Disease
aldehyde-dehydrogenase; ascorbate-dependent artifact; biotin-switch assay; Cell Biology; cytochrome-c-oxidase; hepatoma-cells; lipid-peroxidation; mediated oxidative stress; nitric-oxide; rat-liver; s-nitrosylation
Increased nitroxidative stress causes mitochondrial dysfunctions through oxidative modifications of mitochondrial DNA, lipids, and proteins. Persistent mitochondrial dysfunction sensitizes the target cells/organs to other pathological risk factors and thus ultimately contributes to the development of more severe disease states in alcoholic and nonalcoholic fatty liver disease. The incidences of nonalcoholic fatty liver disease continuously increase due to high prevalence of metabolic syndrome including hyperlipidemia, hypercholesterolemia, obesity, insulin resistance, and diabetes. Many mitochondrial proteins including the enzymes involved in fat oxidation and energy supply could be oxidatively modified (including S-nitrosylation/ nitration) under increased nitroxidative stress and thus inactivated, leading to increased fat accumulation and ATP depletion. To demonstrate the underlying mechanism(s) of mitochondrial dysfunction, we employed a redox proteomics approach using biotin-N-maleimide (biotin-NM) as a sensitive biotin-switch probe to identify oxidized Cys residues of mitochondrial proteins in the experimental models of alcoholic and acute liver disease. The aims of this paper are to briefly describe the mechanisms, functional consequences, and detection methods of mitochondrial dysfunction. We also describe advantages and limitations of the Cys-targeted redox proteomics method with alternative approaches. Finally, we discuss various applications of this method in studying oxidatively modified mitochondrial proteins in extrahepatic tissues or different subcellular organelles and translational research.
Song B J; Abdelmegeed M A; Henderson L E; Yoo S H; Wan J; Purohit V; Hardwick J P; Moon K H
Oxidative Medicine and Cellular Longevity
2013
2013
Journal Article
<a href="http://doi.org/10.1155/2013/781050" target="_blank" rel="noreferrer noopener">10.1155/2013/781050</a>
Mitochondrial dysfunction and tissue injury by alcohol, high fat, nonalcoholic substances and pathological conditions through post-translational protein modifications
Biochemistry & Molecular Biology; cell death; cytochrome-c-oxidase; hepatic ischemia-reperfusion; induced liver-injury; Mitochondrial dysfunction; Mitochondrial proteins; nadp(+)-dependent isocitrate dehydrogenase; nitric-oxide; Nitroxidative stress; oxidative stress; Post-translational modifications; rat-liver; Redox; synthase; targeted antioxidant mitoq; terminal kinase; Tissue injury
Mitochondria are critically important in providing cellular energy ATP as well as their involvement in anfi-oxiclant defense, fat oxidation, intermediary metabolism and cell death processes lt is well-established that mitochondrial functions are suppressed when living cells or organisms are exposed to potentially toxic agents including alcohol, high fat diets, smoking and certain drugs or in many pathophysiological states through increased levels of oxidative/nitrative stress. Under elevated nitroxidative stress, cellular macromolecules proteins, DNA, and lipids can undergo different oxidative modifications, leading to disruption of their normal, sometimes critical, physiological functions. Recent reports also indicated that many mitochondrial proteins are modified via various post-translation modifications (PTMs) and primarily inactivated. Because of the recently-emerging information, in this review, we specifically focus on the mechanisms and roles of five major PTMs (namely oxidation, nitration, phosphorylation, acetylation, and adduct formation with lipid-peroxides, reactive metabolites, or advanced glycation end products) in experimental models of alcoholic and nonalcoholic fatty liver disease as well as acute hepatic injury caused by toxic compounds. We also highlight the role of the ethanol-inducible cytochrome P450-2E1 (CYP2E1) in some of these PTM changes. Finally, we discuss translational research opportunities with natural and/or synthetic anti-oxidants, which can prevent or delay the onset of mitochondial dysfunction, fat accumulation and tissue injury. Published by Elsevier B.V.
Song B J; Akbar M; Abdelmegeed M A; Byun K; Lee B; Yoon S K; Hardwick J P
Redox Biology
2014
2014
Journal Article
<a href="http://doi.org/10.1016/j.redox.2014.10.004" target="_blank" rel="noreferrer noopener">10.1016/j.redox.2014.10.004</a>