Alpha-Tocopherol and endothelial nitric oxide synthesis.
Nitric oxide (NO), a central regulator of vascular tone and homeostasis, is generated upon activation of endothelial NO synthase (eNOS), which is mediated by an increase of intracellular calcium and/or by eNOS phosphorylation. A reduction of NO bioavailability leads to endothelial dysfunction that has been shown to be improved by alpha-tocopherol in certain conditions. The underlying mechanisms, however, are not completely clarified. The present study was performed to investigate whether alpha-tocopherol is able to affect endothelial NO synthesis. The formation of NO was measured in human umbilical vein endothelial cells using citrulline (coproduct) and cGMP (product of the NO-activated soluble guanylate cyclase) as indicator molecules. alpha-Tocopherol (10-200 microM, 24 hr) increased ionomycin-induced citrulline and cGMP formation in intact cells in a concentration-dependent manner. In parallel, ionomycin-stimulated phosphorylation of eNOS at serine 1177, known to support enzyme activation, was increased by alpha-tocopherol, suggesting that this was the mechanism responsible for enhanced NO formation. The effect of alpha-tocopherol was dependent on its hydrophobic structure because it was mimicked by gamma-tocopherol but not by trolox, a hydrophilic derivative of alpha-tocopherol. Coincubation with ascorbic acid (100 microM, 24 hr) amplified the effects of alpha-tocopherol on eNOS phosphorylation and NO formation, which is possibly related to the regeneration of oxidized alpha-tocopherol by ascorbate. Our data suggest that vasoprotective effects of alpha-tocopherol in vivo may be related to an increase of NO formation. The effect of alpha-tocopherol seems to be dependent on tissue saturation with ascorbic acid, and both vitamins may act synergistically to provide optimal conditions for endothelial NO formation.
Heller
Regine
R
Institute of Molecular Cell Biology, Friedrich-Schiller-University of Jena, Nordhäuser Str. 78, 99089 Erfurt, Germany. heller@zmkh.ef.uni-jena.de
Werner-Felmayer
Gabriele
G
Werner
Ernst R
ER
D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism; D002478:Cells, Cultured; D002956:Citrulline / Q000096:biosynthesis; D006152:Cyclic GMP / Q000096:biosynthesis; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism; D014471:Umbilical Veins; D024502:alpha-Tocopherol / Q000494:pharmacology*
D001708:Biopterins; D002956:Citrulline; D009569:Nitric Oxide; D012694:Serine; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C003402:sapropterin; D006152:Cyclic GMP; D024502:alpha-Tocopherol