title: Aurintricarboxylic acid protects against cell death caused by lipopolysaccharide in macrophages by decreasing inducible nitric-oxide synthase induction via IkappaB kinase, extracellular signal-regulated kinase, and p38 mitogen-activated protein kinase inhibition.
abstract: To elucidate the mechanisms involved in cell protection by aurintricarboxylic acid (ATA), an endonuclease inhibitor, high nitric oxide (NO)-induced macrophage apoptosis was studied. In RAW 264.7 macrophages, a high level of NO production accompanied by cell apoptosis was apparent with lipopolysaccharide (LPS) treatment. Direct NO donor sodium nitroprusside (SNP) also dramatically induced cell death, with an EC(50) of 1 mM. Coincubation of ATA (1-500 microM) in LPS-stimulated RAW 264.7 cells resulted in a striking reduction of NO production and cell apoptosis, whereas only a partial cell protection was achieved in response to SNP. This suggests that abrogation of inducible nitric-oxide synthase (iNOS)-dependent NO production might contribute to ATA protection of LPS-treated cells. Immunoblotting and reverse transcription-polymerase chain reaction analysis revealed that ATA down-regulated iNOS protein through transcriptional inhibition of iNOS gene expression but was unrelated to iNOS protein stability. ATA not only inhibited nuclear factor-kappaB (NF-kappaB) activation through impairment of the targeting and degradation of IkappaBs but also reduced LPS-induced activator protein-1 (AP-1) activation. These actions of ATA were not caused by the influence on LPS binding to macrophage membrane. Kinase assays indicated that ATA inhibited IkappaB kinase (IKK), extracellular signal-regulated kinase (ERK), and p38 mitogen-activated protein kinase (MAPK) activity both in vivo and in vitro, suggesting a direct interaction between ATA and these signaling molecules. Taken together, these results provide novel action targets of ATA and indicate that ATA protection of macrophages from LPS-mediated cell death is primarily the result of its inhibition of NO production, which closely relates to the inactivation of NF-kappaB and AP-1 and inhibition of IKK, ERK and p38 MAPK.
authors.affiliation: Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan.
authors.forename: Chin-Ju
authors.initials: CJ
authors.lastname: Tsi
authors.forename: Yee
authors.initials: Y
authors.lastname: Chao
authors.forename: Ching-Wen
authors.initials: CW
authors.lastname: Chen
authors.forename: Wan Wan
authors.initials: WW
authors.lastname: Lin
mesh_terms: D000818:Animals; D017209:Apoptosis; D001312:Aurintricarboxylic Acid / Q000494:pharmacology*; D001665:Binding Sites; D002478:Cells, Cultured; D004347:Drug Interactions; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D009599:Nitroprusside / Q000494:pharmacology; D020011:Protective Agents / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D051381:Rats; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D048051:p38 Mitogen-Activated Protein Kinases
chemical_list: D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D020011:Protective Agents; D018808:Transcription Factor AP-1; D009599:Nitroprusside; D009569:Nitric Oxide; D001312:Aurintricarboxylic Acid; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D017346:Protein Serine-Threonine Kinases; D051550:I-kappa B Kinase; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D016650:Fluorescein-5-isothiocyanate