Nitrated proteins are critical for acetaminophen-mediated mitochondrial dysfunction and acute liver injury

Thursday, October 11, 2012 — Poster Session IV
2:00 p.m. – 4:00 p.m.	Natcher Conference Center, Building 45	NIAAA	OXIDSTRESS-1

Authors

• MA Abdelmegeed
• S Jang
• A Banerjee
• MD Akbar
• BJ Song

Abstract

Acetaminophen (APAP) causes acute liver toxicity through increased oxidative/nitrosative stress and mitochondrial dysfunction. Herein, we aimed to identify nitrated proteins and study their roles in APAP-induced hepatotoxicity. Male mice were exposed to APAP (350/mg/kg i.p.) for 2 and 24 h. Immunoblot analysis showed markedly increased 3-NT proteins in APAP-exposed mice, with severe centrilobular liver necrosis observed at 24 h. However, the number and intensity of nitrated proteins and hepatotoxicity were markedly decreased in APAP-exposed mice pretreated with a peroxynitrite scavenger N-acetylcysteine (NAC). These data suggest important roles of nitrated proteins in APAP-mediated acute hepatotoxicity. Thus, we purified the nitrated cytosolic and mitochondrial proteins by antibody-affinity chromatography. Mass spectral data revealed that many cytosolic (>35) and mitochondrial proteins (>70 proteins) were nitrated. The nitrated proteins include mitochondrial superoxide dismutase, aldehyde dehydrogenase, ATP synthase, and 3-ketoacylCoA thiolase, involved in anti-oxidant defense, energy supply, and fat oxidation pathway, respectively. Immunoprecipitation followed by immunoblot analysis with anti-3NT antibody confirmed that these proteins were nitrated in APAP-exposed mice. In addition, the nitration and suppressed activities of these enzymes were significantly reduced and restored, respectively, by NAC pretreatment group, suggesting the important role of nitration of many cellular proteins in APAP-induced mitochondrial dysfunction and liver toxicity.

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