Mechanism of nitric oxide production induced by H2O2 in cultured endothelial cells.
This study examined the mechanism of stimulation of nitric oxide (NO) synthesis induced by hydrogen peroxide (H2O2). To determine the role of Ca2+ on H2O2-induced NO synthesis in bovine aortic endothelial cells, the increases in intracellular Ca2+ and NO production induced by H2O2 were compared with the effects of the Ca2+ ionophore ionomycin in the same batches of endothelial cells. NO production was assessed by formation of [3H]citrulline from [3H]arginine, and changes in intracellular Ca2+ were measured using fluorescent indicator fluo-3 with a confocal laser scanning system. Both H2O2 (1-10 mM) and ionomycin (10(-7)-10(-5) M) increased intracellular Ca2+ and stimulated the synthesis of L-citrulline from L-arginine. Although H2O2 induced only a small increase in intracellular Ca2+, it markedly increased L-citrulline formation compared with ionomycin. Thus, stimulation of NO synthesis induced by H2O2 may involve the mechanisms other than the increases in intracellular Ca2+ in endothelial cells. In the particulate fraction from cultured endothelial cells, addition of exogenous H2O2 (1 mM) or catalase (100 U/ml) did not affect L-citrulline formation. However, co-administration of H2O2 and catalase stimulated L-citrulline formation. These findings suggested that not only the increases in intracellular Ca2+ but also the products by the reaction with H2O2 and catalase are likely to be involved in the stimulation of NO synthesis induced by H2O2.
Shimizu
S
S
Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
Ishii
M
M
Yamamoto
T
T
Momose
K
K
D000814:Aniline Compounds / Q000737:chemistry; D000818:Animals; D001013:Aorta, Thoracic; D001120:Arginine / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D002417:Cattle; D002458:Cell Fractionation; D002478:Cells, Cultured; D002956:Citrulline / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D005456:Fluorescent Dyes / Q000737:chemistry; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007553:Isotope Labeling; D018613:Microscopy, Confocal; D009569:Nitric Oxide / Q000096:biosynthesis*; D016877:Oxidants / Q000494:pharmacology*; D014316:Tritium; D014966:Xanthenes / Q000737:chemistry
D000814:Aniline Compounds; D005456:Fluorescent Dyes; D007476:Ionophores; D016877:Oxidants; D014966:Xanthenes; D014316:Tritium; C059715:Fluo-3; D002956:Citrulline; D009569:Nitric Oxide; D015759:Ionomycin; D001120:Arginine; D006861:Hydrogen Peroxide; D002374:Catalase; D002118:Calcium