Carboxy-PTIO increases the tetrahydrobiopterin level in mouse brain microvascular endothelial cells.
The aim of the present study was to characterize the increase in tetrahydrobiopterin (BH4), which is a cofactor for nitric oxide synthase (NOS), by carboxy-PTIO, a scavenger of nitric oxide (NO), in vascular endothelial cells. BH4 level was determined by oxidation under acidic conditions as biopterin. Addition of lipopolysaccharide (LPS) to endothelial cells increased mRNA levels of inducible NOS (iNOS) and GTP-cyclohydrolase I (GTPCH), which is a rate-limiting enzyme for BH4 synthesis, and the biopterin level. NOS inhibitors, NO-donors and L-arginine, a substrate of NOS, did not affect the increase in the biopterin level induced by LPS, suggesting that BH4 synthesis is unlikely to be modulated by NO produced by iNOS during LPS treatment. However, carboxy-PTIO increased the biopterin level in the absence and the presence of LPS. Carboxy-PTIO did not affect the expression of GTPCH mRNA level. Moreover, 2,4-diamino-6-hydroxypyrimidine, an inhibitor of GTPCH, inhibited only about 30% of the carboxy-PTIO-induced increase in the biopterin level. Whereas, N-acetylserotonin, an inhibitor of sepiapterin reductase, strongly inhibited the increase in biopterin level. Carboxy-PTIO inhibited the accumulation of pterin, a decomposition product of BH4 in endothelial cells. These findings suggest that carboxy-PTIO accumulates BH4 under basal and LPS-treated conditions in vascular endothelial cells due to both inhibition of the decomposition of BH4 to pterin and activation of the salvage pathway of BH4 synthesis via sepiapterin reductase.
Shimizu
S
S
Department of Pathophysiology, School of Pharmaceutical Sciences, Showa University, Tokyo, Japan. shun@pharm.showa-u.ac.jp
Ishii
M
M
Iwasaki
M
M
Shiota
K
K
Yamamoto
T
T
Kiuchi
Y
Y
D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D001565:Benzoates / Q000494:pharmacology*; D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism*; D001921:Brain / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D006136:GTP Cyclohydrolase / Q000037:antagonists & inhibitors / Q000378:metabolism; D007093:Imidazoles / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D009569:Nitric Oxide / Q000096:biosynthesis; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011622:Pterins / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism
D001565:Benzoates; D007093:Imidazoles; D008070:Lipopolysaccharides; D020030:Nitric Oxide Donors; D011622:Pterins; D012333:RNA, Messenger; C079393:1,3-dihydroxy-4,4,5,5-tetramethyl-2-(4-carboxyphenyl)tetrahydroimidazole; D001708:Biopterins; D009569:Nitric Oxide; D001120:Arginine; D000429:Alcohol Oxidoreductases; C022630:sepiapterin reductase; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D006136:GTP Cyclohydrolase; C003402:sapropterin