The neuronal nitric oxide synthase inhibitor NANT blocks acetaminophen toxicity and protein nitration in freshly isolated hepatocytes
| Autor: | Nukhet Aykin-Burns, Lynda Letzig, Stepan Melnyk, Jack A. Hinson, Kimberly J. Krager, Sudip Banerjee, Laura P. James |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
Male
Mitochondria Liver Nitric Oxide Synthase Type I Mitochondrion Guanidines Biochemistry Article Nitric oxide S-Nitrosoglutathione Mice chemistry.chemical_compound Oxygen Consumption Physiology (medical) medicine Animals Enzyme Inhibitors Chromatography High Pressure Liquid Reactive nitrogen species Acetaminophen Membrane Potential Mitochondrial digestive oral and skin physiology Glutathione Metabolism Analgesics Non-Narcotic NAD Reactive Nitrogen Species Molecular biology Oxidative Stress chemistry Toxicity Hepatocytes Chemical and Drug Induced Liver Injury Reactive Oxygen Species medicine.drug |
| Zdroj: | Free Radical Biology and Medicine. 89:750-757 |
| ISSN: | 0891-5849 |
| DOI: | 10.1016/j.freeradbiomed.2015.09.022 |
| Popis: | 3-Nitrotyrosine (3NT) in liver proteins of mice treated with hepatotoxic doses of acetaminophen (APAP) has been postulated to be causative in toxicity. Nitration is by a reactive nitrogen species formed from nitric oxide (NO). The source of the NO is unclear. iNOS knockout mice were previously found to be equally susceptible to APAP toxicity as wildtype mice and iNOS inhibitors did not decrease toxicity in mice or in hepatocytes. In this work we examined the potential role of nNOS in APAP toxicity in hepatocytes using the specific nNOS inhibitor NANT (10 µM)(N-[(4S)-4-amino-5-[(2-aminoethyl)amino]pentyl]-N'-nitroguanidinetris (trifluoroacetate)). Primary hepatocytes (1 million/ml) from male B6C3F1 mice were incubated with APAP (1mM). Cells were removed and assayed spectrofluorometrically for reactive nitrogen and oxygen species using diaminofluorescein (DAF) and Mitosox red, respectively. Cytotoxicity was determined by LDH release into media. Glutathione (GSH, GSSG), 3NT, GSNO, acetaminophen-cysteine adducts, NAD, and NADH were measured by HPLC. APAP significantly increased cytotoxicity at 1.5-3.0 h. The increase was blocked by NANT. NANT did not alter APAP mediated GSH depletion or acetaminophen-cysteine adducts in proteins which indicated that NANT did not inhibit metabolism. APAP significantly increased spectroflurometric evidence of reactive nitrogen and oxygen formation at 0.5 and 1.0 h, respectively, and increased 3NT and GSNO at 1.5-3.0 h. These increases were blocked by NANT. APAP dramatically increased NADH from 0.5-3.0 h and this increase was blocked by NANT. Also, APAP decreased the Oxygen Consumption Rate (OCR), decreased ATP production, and caused a loss of mitochondrial membrane potential, which were all blocked by NANT. |
| Databáze: | OpenAIRE |
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