Although safe at therapeutic doses, overdoses of acetaminophen produce a centrilobular hepatic necrosis that can be fatal. Each year, more than 9,000 individuals in the US sustain liver damage due to acetaminophen, with 53 deaths reported in 1996. The rationale for this proposal is based on key findings from our preliminary work that challenge the currently accepted paradigm of toxicity. It is generally accepted that acetaminophen is metabolized by CYP-450 to the reactive metabolite, N-acetyl-p-benzoquinone imine (NAPQI), which reacts with GSH, leading to its depletion (and thus decreased peroxide detoxification), and subsequently forming acetaminophen-protein adducts. Our findings indicate that metabolism of acetaminophen to NAPQI may not be the sole determinant of cell lysis and death. We find that the metabolic stress activates resident cells, leading to increase synthesis of nitric oxide (NO) and superoxide, which combine to form peroxynitrite. This entity reacts to form nitrotyrosine-protein adducts and has hydroxyl radical like activity. We detect nitrotyrosine-protein adducts in the hepatic centrilobular cells of acetaminophen-treated mice, the site of the toxicity. Thus, based on this and other preliminary data, we propose a paradigm of acetaminophen hepatotoxicity whereby peroxynitrite generation, coupled with acetaminophen-protein adduct formation, act synergistically to cause cell lysis and death. We hypothesize that peroxynitrite generated during or as a result of acetaminophen metabolic activation is a major determinant of acetaminophen hepatotoxicity. To test this hypothesis, we plan to SA1) Determine the time and dose relationships between acetaminophen metabolism, NO formation, and development of toxicity; SA2) Investigate the roles of NO, superoxide, and peroxynitrite generation in acetaminophen hepatotoxicity by inhibiting NO formation and by using NO, superoxide, and peroxynitrite scavengers; and SA3) Identify the liver cells responsible for NO and superoxide generation during acetaminophen hepatotoxicity. By understanding the role of peroxynitrite in acetaminophen hepatotoxicity, new treatment paradigms may be developed for hepatotoxicity, and this mechanism may be important with other toxins.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM058884-01
Application #
2744671
Study Section
Special Emphasis Panel (ZRG4-ALTX-1 (01))
Program Officer
Serrano, Jose
Project Start
1998-12-01
Project End
2002-11-30
Budget Start
1998-12-01
Budget End
1999-11-30
Support Year
1
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Arkansas for Medical Sciences
Department
Pharmacology
Type
Schools of Medicine
DUNS #
City
Little Rock
State
AR
Country
United States
Zip Code
72205
James, Laura P; Donahower, Brian; Burke, Angela S et al. (2006) Induction of the nuclear factor HIF-1alpha in acetaminophen toxicity: evidence for oxidative stress. Biochem Biophys Res Commun 343:171-6
James, Laura P; Kurten, Richard C; Lamps, Laura W et al. (2005) Tumour necrosis factor receptor 1 and hepatocyte regeneration in acetaminophen toxicity: a kinetic study of proliferating cell nuclear antigen and cytokine expression. Basic Clin Pharmacol Toxicol 97:8-14
James, Laura P; Simpson, Pippa M; Farrar, Henry C et al. (2005) Cytokines and toxicity in acetaminophen overdose. J Clin Pharmacol 45:1165-71
Gu, Jun; Cui, Huadong; Behr, Melissa et al. (2005) In vivo mechanisms of tissue-selective drug toxicity: effects of liver-specific knockout of the NADPH-cytochrome P450 reductase gene on acetaminophen toxicity in kidney, lung, and nasal mucosa. Mol Pharmacol 67:623-30
Gujral, Jaspreet S; Hinson, Jack A; Farhood, Anwar et al. (2004) NADPH oxidase-derived oxidant stress is critical for neutrophil cytotoxicity during endotoxemia. Am J Physiol Gastrointest Liver Physiol 287:G243-52
Gujral, Jaspreet S; Liu, Jie; Farhood, Anwar et al. (2004) Functional importance of ICAM-1 in the mechanism of neutrophil-induced liver injury in bile duct-ligated mice. Am J Physiol Gastrointest Liver Physiol 286:G499-507
James, Laura P; McCullough, Sandra S; Knight, Tamara R et al. (2003) Acetaminophen toxicity in mice lacking NADPH oxidase activity: role of peroxynitrite formation and mitochondrial oxidant stress. Free Radic Res 37:1289-97
James, Laura P; Lamps, Laura W; McCullough, Sandra et al. (2003) Interleukin 6 and hepatocyte regeneration in acetaminophen toxicity in the mouse. Biochem Biophys Res Commun 309:857-63
James, Laura P; Mayeux, Philip R; Hinson, Jack A (2003) Acetaminophen-induced hepatotoxicity. Drug Metab Dispos 31:1499-506
James, Laura P; McCullough, Sandra S; Lamps, Laura W et al. (2003) Effect of N-acetylcysteine on acetaminophen toxicity in mice: relationship to reactive nitrogen and cytokine formation. Toxicol Sci 75:458-67

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