title: Insights into the arginine paradox: evidence against the importance of subcellular location of arginase and eNOS.
abstract: Reduced production of nitric oxide (NO) is one of the first indications of endothelial dysfunction and precedes overt cardiovascular disease. Increased expression of Arginase has been proposed as a mechanism to account for diminished NO production. Arginases consume l-arginine, the substrate for endothelial nitric oxide synthase (eNOS), and l-arginine depletion is thought to competitively reduce eNOS-derived NO. However, this simple relationship is complicated by the paradox that l-arginine concentrations in endothelial cells remain sufficiently high to support NO synthesis. One mechanism proposed to explain this is compartmentalization of intracellular l-arginine into distinct, poorly interchangeable pools. In the current study, we investigated this concept by targeting eNOS and Arginase to different intracellular locations within COS-7 cells and also BAEC. We found that supplemental l-arginine and l-citrulline dose-dependently increased NO production in a manner independent of the intracellular location of eNOS. Cytosolic arginase I and mitochondrial arginase II reduced eNOS activity equally regardless of where in the cell eNOS was expressed. Similarly, targeting arginase I to disparate regions of the cell did not differentially modify eNOS activity. Arginase-dependent suppression of eNOS activity was reversed by pharmacological inhibitors and absent in a catalytically inactive mutant. Arginase did not directly interact with eNOS, and the metabolic products of arginase or downstream enzymes did not contribute to eNOS inhibition. Cells expressing arginase had significantly lower levels of intracellular l-arginine and higher levels of ornithine. These results suggest that arginases inhibit eNOS activity by depletion of substrate and that the compartmentalization of l-arginine does not play a major role.
authors.affiliation: Department of Forensic Medicine, Nanjing Medical University, Nanjing, Jiangsu, China;
authors.forename: Shawn
authors.initials: S
authors.lastname: Elms
authors.forename: Feng
authors.initials: F
authors.lastname: Chen
authors.forename: Yusi
authors.initials: Y
authors.lastname: Wang
authors.forename: Jin
authors.initials: J
authors.lastname: Qian
authors.forename: Bardia
authors.initials: B
authors.lastname: Askari
authors.forename: Yanfang
authors.initials: Y
authors.lastname: Yu
authors.forename: Deepesh
authors.initials: D
authors.lastname: Pandey
authors.forename: Jennifer
authors.initials: J
authors.lastname: Iddings
authors.forename: Ruth B
authors.initials: RB
authors.lastname: Caldwell
authors.forename: David J R
authors.initials: DJ
authors.lastname: Fulton
mesh_terms: D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism; D001119:Arginase / Q000378:metabolism*; D001120:Arginine / Q000378:metabolism* / Q000494:pharmacology; D019556:COS Cells / Q000166:cytology / Q000378:metabolism*; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D002956:Citrulline / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D023421:Models, Animal; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D009952:Ornithine / Q000378:metabolism
chemical_list: D002956:Citrulline; D009569:Nitric Oxide; D001120:Arginine; D009952:Ornithine; D052250:Nitric Oxide Synthase Type III; D001119:Arginase
keywords: arginase; endothelial nitric oxide synthase; l-arginine; nitric oxide; urea