title: Continuous exposure to L-arginine induces oxidative stress and physiological tolerance in cultured human endothelial cells.
abstract: The therapeutic benefits of L-arginine (ARG) supplementation in humans, often clearly observed in short-term studies, are not evident after long-term use. The mechanisms for the development of ARG tolerance are not known and cannot be readily examined in humans. We have developed a sensitive in vitro model using a low glucose/low arginine culture medium to study the mechanisms of ARG action and tolerance using two different human endothelial cells, i.e., Ea.hy926 and human umbilical venous endothelial cells. Cultured cells were incubated with different concentrations of ARG and other agents to monitor their effects on endothelial nitric oxide synthase (eNOS) expression and function, as well as glucose and superoxide (O2(·-) ) accumulation. Short-term (2 h) exposure to at least 50 μM ARG moderately increased eNOS activity and intracellular glucose (p < 0.05), with no change in eNOS mRNA or protein expression. In contrast, 7-day continuous ARG exposure suppressed eNOS expression and activity. This was accompanied by increase in glucose and O2(·-) accumulation. Co-incubation with 100 μM ascorbic acid, 300 U/ml polyethylene glycol-superoxide dismutase (PEG-SOD), 100 μM L-lysine or 30 μM 5-chloro-2-(N-2,5-dichlorobenenesulfonamido)-benzoxazole (a fructose-1,6-bisphosphatase inhibitor) prevented the occurrence of cellular ARG tolerance. Short-term co-incubation of ARG with PEG-SOD improved cellular nitrite accumulation without altering cellular ARG uptake. These studies suggest that ARG-induced oxidative stress may be a primary causative factor for the development of cellular ARG tolerance.
authors: Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, State University of New York, Buffalo, NY 14260, USA.
authors: Srinidi
authors: S
authors: Mohan
authors: Chia-Ching
authors: CC
authors: Wu
authors: Soyoung
authors: S
authors: Shin
authors: Ho-Leung
authors: HL
authors: Fung
mesh_terms: D001119:Arginase / Q000378:metabolism; D001120:Arginine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000494:pharmacology; D002478:Cells, Cultured; D002956:Citrulline / Q000378:metabolism; D003470:Culture Media; D015536:Down-Regulation / Q000187:drug effects; D004361:Drug Tolerance; D016166:Free Radical Scavengers / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D005947:Glucose / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D009566:Nitrates / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D011092:Polyethylene Glycols / Q000494:pharmacology; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism
chemical_list: D003470:Culture Media; D016166:Free Radical Scavengers; D009566:Nitrates; D009573:Nitrites; D013481:Superoxides; D002956:Citrulline; D011092:Polyethylene Glycols; C018524:N,N-dimethylarginine; D001120:Arginine; D052250:Nitric Oxide Synthase Type III; D013482:Superoxide Dismutase; C063471:polyethylene glycol-superoxide dismutase; D001119:Arginase; D005947:Glucose; D001205:Ascorbic Acid