Peroxynitrite formation from macrophage-derived nitric oxide.
Peroxynitrite formation by rat alveolar macrophages activated with phorbol 12-myristate 13-acetate was assayed by the Cu,Zn superoxide dismutase-catalyzed nitration of 4-hydroxyphenylacetate. The inhibitor of nitric oxide synthesis N-methyl-L-arginine prevented the Cu,Zn superoxide dismutase-catalyzed nitration of 4-hydroxyphenylacetate by stimulated macrophages, while Cu-depleted Zn superoxide dismutase did not catalyze the formation of 3-nitro-4-hydroxyphenylacetate either in vitro or in the presence of activated macrophages. The rate of phenolic nitration by activated macrophages was 9 +/- 2 pmol x 10(6) cells-1 x min-1 (mean +/- STD). Only 8% of synthetic peroxynitrite was trapped by superoxide dismutase, which suggested that the rate of peroxynitrite formation may have been as high as 0.11 nmol x 10(6) cells-1 x min-1. This upper estimate was consistent with N-methyl-L-arginine increasing the amount of superoxide detected with cytochrome c by 0.12 nmol x 10(6) cells-1 x min-1. The rate of nitrite and nitrate accumulation was 0.10 +/- 0.001 nmol x 10(6) cells-1 x min-1, suggesting that the majority of nitric oxide produced by activated macrophages may have been converted to peroxynitrite. The formation of a relatively long lived, strong oxidant from the reaction of nitric oxide and superoxide in activated macrophages may contribute to inflammatory cell-mediated tissue injury.
Ischiropoulos
H
H
Department of Anesthesiology, University of Alabama, Birmingham 35294.
Zhu
L
L
Beckman
J S
JS
D000818:Animals; D002417:Cattle; D004912:Erythrocytes / Q000201:enzymology; D007700:Kinetics; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000378:metabolism*; D009566:Nitrates / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D010101:Oxygen Consumption; D051381:Rats; D017207:Rats, Sprague-Dawley; D013482:Superoxide Dismutase / Q000097:blood / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D009566:Nitrates; D013481:Superoxides; C039642:peroxynitric acid; D009569:Nitric Oxide; D013482:Superoxide Dismutase; D013755:Tetradecanoylphorbol Acetate