Agostino Virdis

Introduction Vascular endothelial production of nitric oxide (NO) plays an important role in the modulation of vessel tone and structure, protecting the vascular wall from atherosclerosis. In pathological conditions, however, the endothelium also produces pro-atherogenic substances (mainly reactive oxygen species), which inactivate NO. The Endothelial dysfunction, induced by reduced NO availability, is known to contribute to the development and progression of vascular damage. For this reason, endothelial function has been a major focus of cardiovascular research in the last few decades. Because NO has a very short half-life and its in vivo measurement is difficult, many researchers prefer to measure its biological activity, particularly the NO-dependent vasodilation, at the level of the coronary and peripheral circulation by endothelial stimuli. The most widely used technique involves measurement of brachial artery flow-mediated dilation. This test allows non-invasive evaluation of endothelium-dependent vasodilation in the peripheral macrocirculation induced by a mechanical stimulus (increase in shear stress caused by 5 minutes of forearm ischemia). The vasodilatatory response is reduced in the presence of major cardiovascular risk factors, particularly essential hypertension. Conclusions Studies conducted mainly in high-risk patients have demonstrated that endothelial dysfunction within the coronary or peripheral circulation is predictive of cardiovascular events (independently of classical risk factors). Drug therapy can improve endothelial function by increasing the availability of NO (a possible adjunctive benefit in terms of preventing vascular damage and improving the prognosis). Future studies will establish whether the evaluation of endothelial function by non-invasive, standardized, reproducible, low-cost techniques is an important test for cardiovascular risk stratification in clinical practice.

The endothelium plays a crucial role in modulating vascular function and structure. In healthy conditions, nitric oxide produced by endothelial cells exerts not only vasodilating properties, but also several other protective actions toward the vessel wall against the development of atherosclerosis and thrombosis. Traditional cardiovascular risk factors are characterized by endothelial dysfunction caused by an enhanced production of oxidative stress leading to destroy nitric oxide, thus reducing its availability. Abdominal obesity is associated with endothelial dysfunction, through direct mechanisms, such as insulin resistance and the association with risk factors (including diabetes mellitus, hypertension and dyslipidemia), and direct, by the production of adipokines and pro-inflammatory cytokines, which in turn induce oxidative stress leading to a reduced nitric oxide availability. A reduced endothelium-dependent relaxation is a predictor of cardiovascular events in high-risk patients and represents a putative clinical parameter to stratify the cardiovascular risk and a useful marker for therapy efficacy. Weight loss and a modification of lifestyle ameliorate endothelial function in obese patients, an effect due not only to a better glycemic profile, but also secondary to reduced plasma levels of inflammatory markers and adipokines. At present, whether an improvement of endothelial dysfunction secondary to weight loss is significantly associated with a better cardiovascular prognosis is still unknown. (G Ital Cardiol 2006; 7 (11): 715-723)

We investigated whether cyclooxygenase (COX) isoforms (COX-1 and COX-2) and decreased NO availability contribute to endothelial dysfunction in endotoxemic rats. The involvement of reactive oxygen species (ROS) was also evaluated. Rats were injected with Salmonella-derived lipopolysaccharide or saline. After 6 h, endothelial function of mesenteric resistance arteries was evaluated. In controls, acetylcholine (ACh)-induced relaxation was inhibited by the nitric-oxide synthase inhibitor N^G-monomethyl-l-arginine (l-NMMA) and unaffected by 5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)-phenyl-2(5H)-furanone (DFU) (COX-2 inhibitor). In lipopolysaccharide (LPS)-treated rats, the response to ACh was blunted compared with controls, less sensitive to l-NMMA, and enhanced by DFU. COX-2 blockade also improved the inhibitory effect of l-NMMA on cholinergic relaxation. SC-560 [5-(4-clorophenyl)-1-(4-metoxyphenyl)-3-trifluoromethylpirazole] (COX-1 inhibitor) did not modify the response to ACh in both groups. LPS-induced endothelial dysfunction was unaffected by the thromboxane A₂ (TxA₂) receptor antagonist SQ-29548 (7-[3-[[2-[(phenylamino)carbonyl]hydrazino]methyl]-7-oxabicyclo[2.2.1] hept-2-yl]-[1S(1alpha,2alpha(Z),3alpha,4alpha)]-5-heptenoic acid). In vivo inducible nitric-oxide synthase (iNOS) inhibition by S-methylisothiourea partly attenuated LPS-induced endothelial dysfunction. The antioxidants ascorbic acid and superoxide dismutase normalized endothelium-dependent relaxation and restored the inhibitory action of l-NMMA on ACh. Responses to sodium nitroprusside were similar in both groups. In LPS-treated rats, reverse transcription-polymerase chain reaction showed a marked increase in mesenteric iNOS and COX-2 expressions, whereas endothelial nitric-oxide synthase and COX-1 were unchanged. LPS-induced COX-2 overexpression was reduced but not abrogated by S-methylisothiourea. LPS-induced COX-2 up-regulation was also documented by immunohistochemistry. In conclusion, mesenteric resistance vessels from endotoxemic rats show impaired endothelial function due to reduced NO availability, a condition that is partly ascribable to an iNOS-dependent enhanced COX-2 expression, whereas TxA₂ does not seem to be involved. Oxidative stress is the main mechanism responsible for reduced NO availability, and COX-2 might act as a source of ROS.

Endothelium-derived nitric oxide is not only a potent vasodilator but also inhibits platelet aggregation, vascular smooth muscle cell migration and proliferation, monocyte adhesion and adhesion molecule expression, thus protecting the vessel wall from the development of atherosclerosis and thrombosis. Major cardiovascular risk factors are associated with endothelial dysfunction, which involves enhanced production of oxygen free radicals, that can destroy nitric oxide and reduce its availability, and release of endothelium-derived contracting factors including prostanoids and endothelin-1. Endothelial dysfunction is a promoter of atherosclerotic and thrombotic damage and in prospective studies on patients with high cardiovascular risk impaired endothelium-dependent vasodilation is associated with an increased incidence of cardiovascular events. However, endothelial function cannot yet be included among the surrogate endpoints which need to be measured for cardiovascular risk stratification. This limitation springs from the fact that available tests to assess endothelium-dependent vasodilation are invasive or, if noninvasive, they have no sufficient sensitivity and specificity to be proposed for clinical practice. Moreover, no study is available demonstrating that reversal of endothelial dysfunction, which can be obtained by appropriate treatment, is independently associated with a better clinical outcome. However it is conceivable that in the future, by the utilization of a non-invasive method such as the determination of brachial artery flow-mediated dilation, large-scale multicenter trials might provide a definitive answer to the real prognostic value of endothelial dysfunction, in terms of cardiovascular risk and therapeutic approach.

Essential hypertension is associated with impaired endothelium-dependent vasodilation caused by oxidative stress-induced nitric oxide (NO) breakdown and compensatory production of a hyperpolarizing factor. To test whether calcium antagonist treatment can restore NO availability and prevent hyperpolarization through antioxidant properties, in 15 healthy subjects and 15 patients with essential hypertension, we studied forearm blood flow (strain-gauge plethysmography) modifications induced by intrabrachial bradykinin (5, 15, 50 ng/100 mL per minute), an endothelium-dependent vasodilator, in basal conditions, during infusion of N ^G -monomethyl- l -arginine (L-NMMA, 100 μg/100 mL per minute), an NO-synthase inhibitor, and ouabain (0.72 μg/100 mL per minute), an Na ⁺ -K ⁺ ATPase inhibitor to prevent hyperpolarization. These infusions were repeated in the presence of the antioxidant vitamin C (8 mg/100 mL/min). The response to sodium nitroprusside was also evaluated. In controls, vasodilation to bradykinin was inhibited by L-NMMA and remained unchanged by ouabain or vitamin C. In hypertensive patients, vasodilation to bradykinin was blunted and resistant to L-NMMA but sensitive to ouabain. Vitamin C increased the response to bradykinin and restored the inhibiting effect of L-NMMA while preventing the effect of ouabain. In hypertensive patients, infusions were repeated after 3-month treatment with lercanidipine (10 to 20 mg daily). Lercanidipine decreased plasma lipoperoxides, isoprostanes, and malondialdehyde and increased plasma antioxidant capacity. Moreover, lercanidipine increased the vasodilation to bradykinin and restored the inhibiting effect of L-NMMA on bradykinin-induced vasodilation while preventing the effect of ouabain. Finally, vitamin C no longer exerted its facilitating activity. These results indicate that in essential hypertension, lercanidipine increases endothelium-dependent vasodilation by restoring NO availability and preventing hyperpolarization, an effect probably determined by antioxidant activity.