Peroxynitrite is a potent inhibitor of NF-{kappa}B activation triggered by inflammatory stimuli in cardiac and endothelial cell lines.
Peroxynitrite is a potent oxidant and nitrating species proposed as a direct effector of myocardial damage in numerous cardiac pathologies. Whether peroxynitrite also acts indirectly, by modulating cell signal transduction in the myocardium, has not been investigated. Therefore, we examined a possible role for peroxynitrite on the activation of NF-kappaB, a crucial pro-inflammatory transcription factor, in cultured H9C2 cardiomyocytes. H9C2 cells were stimulated with tumor necrosis factor-alpha or lipopolysaccharide following a brief (20-min) exposure to peroxynitrite. NF-kappaB activation (phosphorylation and degradation of its inhibitor IkappaBalpha, nuclear translocation of NF-kappaB p65, and NF-kappaB DNA binding) triggered by lipopolysaccharide or tumor necrosis factor-alpha was abrogated by peroxynitrite. Peroxynitrite also inhibited NF-kappaB in two human endothelial cell lines activated with tumor necrosis factor-alpha or interleukin-1beta. These effects were related to oxidative but not nitrative chemistry and were still being observed while nitration was suppressed by epicatechin. The mechanism of NF-kappaB inhibition by peroxynitrite was a complete blockade of phosphorylation and activation of the upstream kinase IkappaB kinase (IKK) beta, required for canonical, pro-inflammatory NF-kappaB activation. At the same time, peroxynitrite activated phosphorylation of NF-kappaB-inducing kinase and IKKalpha, considered as part of an alternative, noncanonical NF-kappaB activation pathway. Suppression of IKKbeta-dependent NF-kappaB activation translated into a marked inhibition of the transcription of NF-kappaB-dependent genes by peroxynitrite. Thus, peroxynitrite has a dual effect on NF-kappaB, inhibiting canonical IKKbeta-dependent NF-kappaB activation while activating NF-kappaB-inducing kinase and IKKalpha phosphorylation, which suggests its involvement in an alternative pathway of NF-kappaB activation. These findings offer new perspectives for the understanding of the relationships between redox stress and inflammation.
Levrand
Sandra
S
Division of Critical Care, Division of Clinical Pathophysiology, Department of Internal Medicine, University Hospital, 1011 Lausanne, Switzerland.
Pesse
Benoît
B
Feihl
François
F
Waeber
Bernard
B
Pacher
Pal
P
Rolli
Joëlle
J
Schaller
Marie-Denise
MD
Liaudet
Lucas
L
D021581:Active Transport, Cell Nucleus; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival; D003593:Cytoplasm / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D042783:Endothelial Cells / Q000166:cytology*; D017930:Genes, Reporter; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D007249:Inflammation; D007375:Interleukin-1 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000737:chemistry / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008954:Models, Biological; D009206:Myocardium / Q000166:cytology*; D016328:NF-kappa B / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D030421:Peroxynitrous Acid / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D010766:Phosphorylation; D051381:Rats; D015398:Signal Transduction; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D007375:Interleukin-1; D008070:Lipopolysaccharides; D016328:NF-kappa B; D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha; D030421:Peroxynitrous Acid; D008156:Luciferases; C022616:thiazolyl blue; D010100:Oxygen