In 1980, Furchgott and Zawadzki demonstrated that the relaxation of vascular smooth muscle cells in response to acetylcholine is dependent on the anatomical integrity of the endothelium. Endothelium-derived relaxing factor was identified 7 years later as the free radical gas nitric oxide (NO). In endothelium, the amino acid L-arginine is converted to L-citrulline and NO by one of the three NO synthases, the endothelial isoform (eNOS). Shear stress and cell proliferation appear to be, quantitatively, the two major regulatory factors of eNOS gene expression. However, eNOS seems to be mainly regulated by modulation of its activity. Stimulation of specific receptors to various agonists (e.g., bradykinin, serotonin, adenosine, ADP/ATP, histamine, thrombin) increases eNOS enzymatic activity at least in part through an increase in intracellular free Ca2+. However, the mechanical stimulus shear stress appears again to be the major stimulus of eNOS activity, although the precise mechanisms activating the enzyme remain to be elucidated. Phosphorylation and subcellular translocation (from plasmalemmal caveolae to the cytoskeleton or cytosol) are probably involved in these regulations. Although eNOS plays a major vasodilatory role in the control of vasomotion, it has not so far been demonstrated that a defect in endothelial NO production could be responsible for high blood pressure in humans. In contrast, a defect in endothelium-dependent vasodilation is known to be promoted by several risk factors (e.g., smoking, diabetes, hypercholesterolemia) and is also the consequence of atheroma (fatty streak infiltration of the neointima). Several mechanisms probably contribute to this decrease in NO bioavailability. Finally, a defect in NO generation contributes to the pathophysiology of pulmonary hypertension. Elucidation of the mechanisms of eNOS enzyme activity and NO bioavailability will contribute to our understanding the physiology of vasomotion and the pathophysiology of endothelial dysfunction, and could provide insights for new therapies, particularly in hypertension and atherosclerosis.

Angiotensin II (ANG II) produces vasoconstriction by a direct action on smooth muscle cells via AT1 receptors. These receptors are also present in the endothelium, but their function is poorly understood. This study was therefore undertaken to determine whether ANG II elicits the release of nitric oxide (NO) from cultured rat aortic endothelial cells. NO production, measured by the accumulation of nitrite and nitrate, was enhanced by 10−7 M ANG II. The biological activity of the NO released by ANG II action was evaluated by measuring its guanylate cyclase-stimulating activity in smooth muscle cells. The guanosine 3′,5′-cyclic monophosphate (cGMP) content of smooth muscle cells was significantly increased by exposure of supernatant from ANG II-stimulated endothelial cells. These effects resulted from the activation of NO synthase, as they were inhibited by the l-arginine analogs. These ANG II actions were mediated by the AT1 receptor, as shown by their inhibition by the AT1 antagonist losartan. The cGMP production by reporter cells was inhibited by the calmodulin antagonist W-7, suggesting that ANG II activates endothelial calmodulin-dependent NO synthase. This hypothesis is also supported by the increase of intracellular free calcium induced by ANG II in endothelial cells. ANG II also stimulated luminol-enhanced chemiluminescence in endothelial cells. This effect was inhibited by N ω-monomethyl-l-arginine and superoxide dismutase, suggesting that this luminol-enhanced chemiluminescence reflected an increase in peroxynitrite production. Thus ANG II stimulates NO release from macrovascular endothelium, which may modulate the direct vasoconstrictor effect of ANG II on smooth muscle cells. However, this beneficial effect may be counteracted by the simultaneous production of peroxynitrite, which could contribute to several pathological processes in the vascular wall.

Nasal NO concentration is increased in patients with allergic rhinitis. Interestingly, patients without symptoms on the day of the test also showed a clear-cut increase in nasal NO production, which could reflect a permanent inflammation of the sinus mucosa.

Abstract-Although estradiol (E 2 ) has been recognized to exert several vasculoprotective effects in several species, its effects in mouse vasomotion are unknown, and consequently, so is the estrogen receptor subtype mediating these effects. We investigated the effect of E 2 (80 g/kg/day for 15 days) on NO production in the thoracic aorta of ovariectomized C57Bl/6 mice compared with those given placebo. E 2 increased basal NO production. In contrast, the relaxation in response to ATP, to the calcium ionophore A23187, and to sodium nitroprusside was unaltered by E 2 , whereas acetylcholine-elicited relaxation was decreased. The abundance of NO synthase I, II, and III immunoreactive proteins (using Western blot) in thoracic aorta homogenates was unchanged by E 2 . To determine the estrogen receptor (ER) subtype involved in these effects, transgenic mice in which either the ER␣ or ER␤ has been disrupted were ovariectomized and treated, or not, with E 2 . Basal NO production was increased and the sensitivity to acetylcholine decreased in ER␤ knockout mice in response to E 2 , whereas this effect was abolished in ER␣ knockout mice. Finally, these effects of E 2 on vasomotion required long-term and/or in vivo exposure, as short-term incubation of aortic rings with 10 nmol/L E 2 in the isolated organ chamber did not elicit any vasoactive effects. In conclusion, this study demonstrates that ER␣, but not ER␤, mediates the beneficial effect of E 2 on basal NO production. Key Words: nitric oxide synthase Ⅲ endothelial cell Ⅲ estrogens Ⅲ estrogen receptor T he incidence of cardiovascular disease is higher in men than in premenopausal women but increases in postmenopausal women. Until recently, an abundance of epidemiological data suggested a role for estrogens in this atheroprotective effect. 1,2 However, the protective effects of estrogen on cardiovascular diseases has become controversial these last years. The Heart and Estrogen/progestin Replacement Study (HERS) recently demonstrated the failure of hormonal replacement therapy (HRT) to increase survival in secondary prevention. 3 Because long-term trials evaluating the effect of HRT in primary prevention will conclude in several years, it is important to get insight into the vascular effects and mechanisms in the meantime.The mechanism whereby this protection is attributable to favorable changes in blood lipids and lipoproteins accounts for approximatively one third of its effect, 1 but a number of studies in humans 4 as well as animals 5-7 strongly suggest a direct action on the arterial wall.Endothelium is now recognized to play a crucial role in the physiology of circulation. In particular, endothelium generates nitric oxide (NO). This free radical messenger inhibits platelet aggregation, interferes with mononuclear cell adhesion, and relaxes the underlying smooth muscle cells. 8 -10 NO is generated by a family of 3 isoenzymes: neuronal NOsynthase (type I), inducible NO-synthase (type II), and endothelial NO-synthase (type III). NO synthase (NOS) III activity can be ...