Exogenous nitric oxide regulates activity and synthesis of vascular endothelial nitric oxide synthase.
BACKGROUND
Nitric oxide (NO) - a major signalling molecule of the vascular system - is constitutively produced in endothelial cells (EC) by the endothelial NO synthase (eNOS). Since a reduced NO synthesis is an early sign of endothelial dysfunction and NO delivering drugs are used to substitute the impaired endothelial NO production, we addressed the effect of exogenous NO on eNOS in human umbilical venous endothelial cell cultures.


METHODS
The synthetic NO donor DETA/NO (trade name, but in the following we refer to detNO), that releases NO in a strictly first order reaction with a half life of 20 h, was used in our experiments.


RESULTS
Short-term (20-30 min) detNO treatment of EC increases the Ser(1177) phosphorylation of the constitutively expressed endothelial NOS and the production of endogenous NO generated by eNOS from [(3)H]arginine. The phosphorylation of eNOS is Akt-dependent and completely reverted by the phosphatidylinositol-3 kinase (PI-3K) inhibitor LY294002. A prolonged continuous exposure of EC to detNO 150 micromol L(-1) over a period of 24-48 h causes a reversible cell cycle arrest at G(1)-phase associated with a larger cell volume and increased cell protein content (hypertrophic phenotype of EC). The eNOS protein and mRNA of the hypertrophic cells and the generation of endogenous NO are reduced but eNOS phosphorylation could still be elevated by stimulation with vascular endothelial growth factor.


CONCLUSIONS
Our data explain clinical studies describing a short-term but not a long-term benefit of NO treatment for patients with cardiovascular risk factors. The results could be a rational approach to develop a generation of NO donors accomplishing a retarded release from NO donors that mimic the low continuous pulsatile stress-induced release of endogenous NO.
Schmidt
A
A
Leibniz-Institute of Arteriosclerosis Research at University of Muenster, Germany. annschm@uni-muenster.de
Bilgasem
S
S
Lorkowski
S
S
Vischer
P
P
Völker
W
W
Breithardt
G
G
Siegel
G
G
Buddecke
E
E
D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects; D006801:Humans; D008954:Models, Biological; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D052250:Nitric Oxide Synthase Type III / Q000096:biosynthesis*; D015398:Signal Transduction / Q000187:drug effects; D013223:Statistics as Topic; D014226:Triazenes / Q000494:pharmacology
C105351:1-hydroxy-2-oxo-3,3-bis(2-aminoethyl)-1-triazene; D020030:Nitric Oxide Donors; D014226:Triazenes; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III