Interactions of peroxynitrite with uric acid in the presence of ascorbate and thiols: implications for uncoupling endothelial nitric oxide synthase.
It has been suggested that uric acid acts as a peroxynitrite scavenger although it may also stimulate lipid peroxidation. To gain insight into how uric acid may act as an antioxidant, we used electron spin resonance to study the reaction of uric acid and plasma antioxidants with ONOO-. Peroxynitrite reacted with typical plasma concentrations of urate 16-fold faster than with ascorbate and 3-fold faster than cysteine. Xanthine but not other purine-analogs also reacted with peroxynitrite. The reaction between ONOO- and urate produced a carbon-centered free radical, which was inhibited by either ascorbate or cysteine. Moreover, scavenging of ONOO- by urate was significantly increased in the presence of ascorbate and cysteine. An important effect of ONOO- is oxidation of tetrahydrobiopterin, leading to uncoupling of nitric oxide synthase. The protection of eNOS function by urate, ascorbate and thiols in ONOO(-)-treated bovine aortic endothelial cells (BAECs) was, therefore, investigated by measuring superoxide and NO using the spin probe 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethyl-pyrrolidine (CMH) and the NO-spin trap Fe[DETC]2. Peroxynitrite increased superoxide and decreased NO production by eNOS indicating eNOS uncoupling. Urate partially prevented this effect of ONOO- while treatment of BAECs with the combination of either urate with ascorbate or urate with cysteine completely prevented eNOS uncoupling caused by ONOO-. We conclude that the reducing and acidic properties of urate are important in effective scavenging of peroxynitrite and that cysteine and ascorbate markedly augment urate's antioxidant effect by reducing urate-derived radicals.
Kuzkaya
Nermin
N
Department of Internal Medicine, Justus-Liebig University School of Medicine, Giessen, Germany.
Weissmann
Norbert
N
Harrison
David G
DG
Dikalov
Sergey
S
D000818:Animals; D001205:Ascorbic Acid / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004347:Drug Interactions; D004730:Endothelium, Vascular / Q000201:enzymology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D030421:Peroxynitrous Acid / Q000737:chemistry / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism*; D014475:Uncoupling Agents / Q000737:chemistry / Q000378:metabolism*; D014527:Uric Acid / Q000737:chemistry / Q000378:metabolism*
D013438:Sulfhydryl Compounds; D014475:Uncoupling Agents; D030421:Peroxynitrous Acid; D014527:Uric Acid; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; D001205:Ascorbic Acid