title: Laminar flow stimulates ATP- and shear stress-dependent nitric oxide production in cultured bovine endothelial cells.
abstract: Based on the fact that nitric oxide (NO) production is associated with changes in intracellular cGMP levels and is selectively inhibited by N omega-methyl L-arginine (L-NME), we investigated the shear stress dependency of NO production in endothelial cells (ECs) from its cGMP responses to various shear stress loads. Cultured fetal bovine aortic ECs treated with a phosphodiesterase inhibitor, isobutylmethylxanthine (IBMX; 1 mM), were exposed to a laminar flow of Krebs buffer solution for 5 minutes in a parallel-plate flow chamber and examined for changes in intracellular cGMP levels by radioimmunoassay using an [125I] cGMP kit. Application of flow increased the cGMP levels. The increase was significant in the presence of extracellular ATP (1 microM)(control, 286.1 +/- 43.6; flow, 506.5 +/- 44.9 fmol/10(7) cells; p < 0.001), but not in its absence (control, 256.6 +/- 60.6; flow, 301.5 +/- 91.4 fmol/10(7) cells; N.S.). The cGMP levels increased significantly as the magnitude of shear stress applied increased. Treatment of ECs with a specific inhibitor of NO production, L-NMA (200 microM), completely inhibited the flow-induced increase in cGMP, and L-arginine reversed the L-NMA-induced inhibition, indicating that the increase in cGMP was due to NO produced by the flow. The flow-induced increase in NO production was markedly suppressed when extracellular Ca++ was chelated by adding EGTA to the perfusate. These findings suggest that flow stimulates NO production to increase cGMP levels shear stress-dependently in ECs and that extracellular Ca++ and ATP modulate the effects of flow.
authors.affiliation: Dept. of Cardiovascular Biomechanics, Faculty of Medicine, University of Tokyo, Japan.
authors.forename: R
authors.initials: R
authors.lastname: Korenaga
authors.forename: J
authors.initials: J
authors.lastname: Ando
authors.forename: H
authors.initials: H
authors.lastname: Tsuboi
authors.forename: W
authors.initials: W
authors.lastname: Yang
authors.forename: I
authors.initials: I
authors.lastname: Sakuma
authors.forename: T
authors.initials: T
authors.lastname: Toyo-oka
authors.forename: A
authors.initials: A
authors.lastname: Kamiya
mesh_terms: D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D001013:Aorta, Thoracic; D001120:Arginine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D005333:Fetus; D007700:Kinetics; D009569:Nitric Oxide / Q000096:biosynthesis*; D013314:Stress, Mechanical; D019323:omega-N-Methylarginine
chemical_list: D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D004533:Egtazic Acid; D000255:Adenosine Triphosphate; D001120:Arginine; D006152:Cyclic GMP; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine