ADMA and oxidative stress are responsible for endothelial dysfunction in hyperhomocyst(e)inemia: effects of L-arginine and B vitamins

OBJECTIVES: Hyperhomocyst(e)inemia is a risk factor for atherosclerotic vascular disease, and it is associated with endothelial dysfunction. Mechanisms responsible for endothelial dysfunction in hyperhomocyst(e)inemia may involve impaired bioavailability of NO, possibly secondary to accumulation of the endogenous NO synthase inhibitor asymmetric dimethylarginine (ADMA) and increased oxidative stress. We investigated whether oral treatment with B vitamins or L-arginine normalizes endothelium-dependent, flow-dependent vasodilation (FDD) in patients with peripheral arterial occlusive disease (PAOD) and hyperhomocyst(e)inemia. METHODS: 27 patients with PAOD and hyperhomocyst(e)inemia were assigned to oral treatment with combined B vitamins (folate, 10 mg; vitamin B-12, 200 microg; vitamin B-6, 20 mg/day), L-arginine (24 g/day) or placebo, for 8 weeks in a double-blind fashion. FDD was determined by high-resolution ultrasound in the radial artery. RESULTS: Vitamin B supplementation significantly lowered plasma homocyst(e)ine concentration from 15.8+/-1.8 to 8.7+/-1.1 micromol/l (P<0.01). However, B vitamins had no significant effect on FDD (baseline, 7.8+/-0.7%, B vitamins, 8.3+/-0.9%, placebo 8.9+/-0.7%; P=n.s.). In contrast, L-arginine treatment did not affect homocyst(e)ine levels, but significantly improved FDD (10.2+/-0.2%), probably by antagonizing the impact of elevated ADMA concentration (3.8+/-0.3 micromol/l) and reducing the oxidative stress by lowering urinary 8-iso-prostaglandin F(2alpha) (baseline, 76.3+/-7.1 vs. 62.7+/-8.3 pmol/mmol creatinine after 8 weeks). CONCLUSIONS: Oral supplementation with combined B vitamins during 8 weeks does not improve endothelium-dependent vasodilation in PAOD patients with hyperhomocyst(e)inemia, whereas L-arginine significantly improved endothelial function in these patients. Thus, accumulation of ADMA and increased oxidative stress may underlie endothelial dysfunction under hyperhomocyst(e)inemic conditions. These findings may have importance for evaluation of homocyst(e)ine-lowering therapy.     

Effects of oral L-arginine on endothelium-dependent vasodilation and markers of inflammation in healthy postmenopausal women

OBJECTIVES: We examined whether oral administration of L-arginine, the substrate for nitric oxide (NO) synthesis, increases NO bioactivity in healthy postmenopausal women. BACKGROUND: Nitric oxide may protect arteries against atherosclerosis, as suggested by experimental studies in animals. Estrogen therapy, which has been shown to increase NO bioactivity in the vasculature of healthy postmenopausal women, is not acceptable for long-term use by many women. METHODS: In a randomized, double-blind, crossover study, 10 postmenopausal women without additional risk factors for atherosclerosis received L-arginine 9 g or placebo daily for one month, with treatment periods separated by one month. Nitric oxide levels in serum (as an index of endothelial NO release), brachial artery endothelium-dependent dilator responses to hyperemia by ultrasonography (as an index of vascular NO bioactivity) and markers of inflammation in blood that are inhibited by NO in cell culture experiments were measured at the end of each treatment period. RESULTS: L-arginine levels in plasma were increased in all women during L-arginine treatment compared with placebo (136.8 +/- 63.1 vs. 75.2 +/- 16.2 micromol/liter, p = 0.009). However, there was no change in serum nitrogen oxide levels (42.1 +/- 24.5 vs. 39.1 +/- 16.6 micromol/liter, p = 0.61), nor was there an effect of L-arginine on flow-mediated dilation during hyperemia (3.8 +/- 3.0% vs. 4.9 +/- 4.8%, p = 0.53) compared with placebo. Our study had sufficient power (beta = 0.80) to detect a true absolute treatment difference in flow-mediated brachial artery dilation of 1.7% or larger as statistically significant at alpha = 0.05. There was no effect of L-arginine on serum levels of soluble cell adhesion molecules compared with placebo: E-selectin (50.6 +/- 14.8 vs. 52.1 +/- 17.0 ng/ml, p = 0.45), intercellular adhesion molecule-1 (230 +/- 51 vs. 230 +/- 52 ng/ml, p = 0.97) and vascular cell adhesion molecule-1 (456 +/- 62 vs. 469 +/- 91 ng/ml, p = 0.53). CONCLUSIONS: Oral administration of L-arginine may not augment endothelial NO synthesis and release in postmenopausal women and is thus unlikely to be of general benefit to healthy postmenopausal women in protection from the development of atherosclerosis.     

Relationship between S-adenosylmethionine, S-adenosylhomocysteine, asymmetric dimethylarginine, and endothelial function in healthy human subjects during experimental hyper- and hypohomocysteinemia

Experimental hyperhomocysteinemia after an oral methionine or homocysteine load is associated with impaired nitric oxide-dependent vasodilatation in healthy human beings. However, it remains unproven that this effect is mediated by elevations in plasma homocysteine. There is evidence that an increase in plasma homocysteine may increase the formation of asymmetric dimethylarginine (ADMA), an inhibitor of nitric oxide synthase. The methyl groups within ADMA are derived from the conversion of S -adenosylmethionine to S -adenosylhomocysteine intermediates in the methionine/homocysteine pathway. No previous study has assessed the role of methylation status, its impact on ADMA formation, and their association with endothelial function in healthy human beings. In a randomized, placebo-controlled, crossover study, 10 healthy subjects (mean age, 29.1 +/- 3.9 years) were administered an oral dose of methionine (0.1 g/kg), l -homocysteine (0.01 g/kg), N-acetylcysteine (NAC) (0.1 g/kg), or placebo. Endothelial function as assessed by flow-mediated dilatation (FMD) of the brachial artery was impaired after both the methionine and homocysteine load compared with placebo at 4 hours (36 +/- 15, 67 +/- 23 vs 219 +/- 26 microm, respectively, P < .001). N-Acetylcysteine had no effect on flow-mediated dilatation. Plasma total homocysteine was significantly elevated at 4 hours after methionine (23.1 +/- 6.2) and homocysteine (41.5 +/- 8.9) loading, but significantly reduced after NAC 2.4 +/- 0.6 vs 7.1 +/- 2.1 micromol/L in the placebo (P < .001). Plasma S-adenosylmethionine/S-adenosylhomocysteine ratio was significantly (P < .001) increased at 4 hours after methionine (10.9 +/- 0.7) compared with homocysteine (5.4 +/- 0.4), NAC (5.0 +/- 0.3), and placebo (6.0 +/- 0.5). Plasma ADMA concentrations were not altered by any intervention. Our results suggest that endothelial dysfunction due to methionine or homocysteine loading is not associated with an increase in plasma ADMA or a disruption in methylation status.     

Endothelium-dependent vasodilation is independent of the plasma L-arginine/ADMA ratio in men with stable angina: lack of effect of oral L-arginine on endothelial function, oxidative stress and exercise performance

OBJECTIVES: This study was designed to determine the effect of two weeks' treatment with L-arginine on the ratio of plasma L-arginine to asymmetric dimethylarginine (ADMA), oxidative stress, endothelium-dependent vasodilatation to acetylcholine, exercise performance and heart rate variability in men with stable angina. BACKGROUND: The ratio of plasma L-arginine:ADMA has been proposed as a determinant of endothelium-dependent dilation; dietary supplementation with L-arginine has been shown to improve endothelium-dependent vasodilation and symptoms in some conditions. METHODS: Men (n = 40) with stable angina, at least one epicardial coronary artery with a stenosis >50% and a positive exercise test were randomized to receive L-arginine (15 g daily) or placebo for two weeks according to a double-blind parallel-group design. Plasma L-arginine, ADMA, 8-epi-prostaglandin F2alpha (a marker of oxidative stress) and forearm vasodilator responses to brachial artery infusion of nitroprusside and acetylcholine (+/-L-arginine) were measured. A standard Bruce protocol exercise test was performed before and at the end of the treatment period. RESULTS: Plasma L-arginine increased after oral L-arginine, whereas ADMA remained unchanged, leading to an increase in the L-arginine/ADMA ratio of 62 +/- 11% (mean +/- SE, p < 0.01). Despite a significant enhancement in acetylcholine response by intra-arterial L-arginine at baseline, this response remained unchanged after oral L-arginine. Measures of oxidative stress and exercise performance after L-arginine/placebo were similar in placebo and active groups. CONCLUSIONS: In men with stable angina, an increase in plasma L-arginine/ADMA ratio after two weeks' oral supplementation with L-arginine is not associated with an improvement in endothelium-dependent vasodilatation, oxidative stress or exercise performance.     

Asymmetric dimethylarginine determines the improvement of endothelium-dependent vasodilation by simvastatin: Effect of combination with oral L-arginine.

OBJECTIVES: We hypothesized that the level of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of endothelial nitric oxide (NO) synthase (eNOS), might determine the endothelial effects of statins. BACKGROUND: Endothelial NO synthase is up-regulated by statins. However, statins failed to improve endothelial function in some studies. Asymmetric dimethylarginine inhibits eNOS by a mechanism that is reversible by L-arginine. METHODS: Ninety-eight clinically asymptomatic elderly subjects had their plasma ADMA levels screened. Those in the highest (high ADMA, n = 15) and lowest quartiles of the ADMA distribution (low ADMA, n = 13) were eligible to receive, in a randomized order, simvastatin (40 mg/day), L-arginine (3 g/day), or a combination of both, each for 3 weeks. Endothelium-dependent vasodilation (EDD) was assessed by brachial artery ultrasound. RESULTS: Simvastatin had no effect on EDD in subjects with high ADMA (6.2 +/- 1.2% vs. 6.1 +/- 0.9%), whereas simvastatin plus L-arginine significantly improved EDD (9.8 +/- 1.5% vs. 5.3 +/- 0.8%; p < 0.01). In subjects with low ADMA, simvastatin improved endothelial function when given alone (9.5 +/- 3.2% vs. 6.1 +/- 3.8%; p < 0.001) or in combination with L-arginine (9.0 +/- 3.1% vs. 6.3 +/- 3.3%; p = 0.001). L-arginine alone improved endothelial function in both groups. Endothelium-independent vasodilation was not affected. CONCLUSIONS: Simvastatin does not enhance endothelial function in subjects with elevated ADMA, whereas it does so in patients with low ADMA. Combination of simvastatin with oral L-arginine improves endothelial function in subjects with high ADMA, but has no additional effect in subjects with low ADMA. As NO-mediated effects may play a major role in the therapeutic effects of statins, ADMA concentration is an important factor that influences the "pleiotropic" effects of simvastatin.     

Endothelial dysfunction in patients with peripheral arterial disease and chronic hyperhomocysteinemia: potential role of ADMA

Hyperhomocysteinemia is associated with an enhanced risk for cardiovascular disease. Patients with peripheral arterial disease (PAD) show an increased prevalence of hyperhomocysteinemia. A decreased biological activity of nitric oxide (NO) may contribute to homocysteine-associated endothelial dysfunction. This study was designed to investigate whether elevated levels of the endogenous NO synthase inhibitor asymmetric dimethylarginine (ADMA) are involved in endothelial dysfunction in patients with chronic hyperhomocysteinemia and PAD. A total of 76 patients (58 males and 18 females; mean age 65.2 +/- 2.0 years) with PAD were included in the analysis and characterized according to demographic variables and cardiovascular risk factors. Flow-dependent vasodilation (FDD) was determined by high-resolution ultrasound in the radial artery. Total plasma homocysteine (plasma tHcy) and ADMA levels were measured by HPLC. Urinary nitrate was quantified using gas chromatography-mass spectrometry. Patients with plasma tHcy in the highest tertile (n = 27; i.e. > 10.6 micromol/l) had a mean plasma level of 14.4 +/- 1.21 mol/l compared with 9.9 +/- 0.1 micromol/l in those patients in the middle tertile (n = 22; p < 0.05) and 9.4 +/- 0.1 micromol/l in those in the lowest tertile (n = 27; i.e. <9.6 micromol/l; p < 0.05). The hyperhomocysteinemic individuals (highest tertile) had a significantly decreased FDD compared with healthy age-matched controls (n = 15) (7.6 +/- 1.0 vs 13.0 +/- 0.4%; p < 0.05), higher plasma ADMA concentrations (4.0 +/- 0.3 vs 2.6 +/- 0.3 micromol/l; p < 0.05), and a lower urinary nitrate excretion rate (89.5 +/- 13.4 vs 131.3 +/- 17.9 micromol/mmol creatinine; p < 0.05) compared with patients with plasma tHcy in the lowest tertile. Multivariate regression analysis including plasma tHcy, ADMA, total cholesterol, diabetes mellitus, smoking, and systolic blood pressure revealed ADMA as the only significant factor determining FDD (p < 0.05). In conclusion, we demonstrated a stronger relationship between impaired endothelial function and elevated ADMA levels in comparison with plasma tHcy concentrations in patients with PAD and chronic hyperhomocysteinemia. This may raise the question of whether different therapeutical options that interact indirectly with plasma tHcy, i.e. treatment with ACE inhibitors and AT1-receptor blockers to reduce ADMA plasma concentrations or L-arginine, could be a beneficial tool for treating patients with hyperhomocysteinemia.     

Salt loading on plasma asymmetrical dimethylarginine and the protective role of potassium supplement in normotensive salt-sensitive asians

Asymmetrical dimethylarginine (ADMA) is an endogenous inhibitor of NO synthase. Because endothelial NO pathway is compromised in patients with salt-sensitive hypertension, we investigated whether the plasma ADMA can be modulated by chronic salt loading in normotensive salt-sensitive persons and its relationship with NO, and we further determined whether or not dietary potassium supplementation can reverse them. Sixty normotensive subjects (aged 20 to 60 years) were selected from a rural community of Northern China. All of the people were sequentially maintained on a low-salt diet for 7 days (3 g/day, NaCl), then a high-salt diet for 7 days (18 g/day), and high-salt diet with potassium supplementation for another 7 days (4.5 g/day, KCl). After salt loading, the plasma ADMA concentrations increased significantly in salt-sensitive subjects (0.89+/-0.02 micromol/L versus 0.51+/-0.02 micromol/L; P<0.05), whereas the plasma NOx levels reduced considerably (41.8+/-2.1 micromol/L versus 63.5+/-2.1 micromol/L; P<0.01). All of the abnormalities normalized when dietary potassium were supplemented (0.52+/-0.03 micromol/L versus 0.89+/-0.02 micromol/L for ADMA and 58.1+/-0.9 micromol/L versus 41.8+/-2.1 micromol/L for NOx). Statistically significant correlations were found among plasma ADMA level, the mean blood pressure, and the level of NO after salt loading in normotensive salt sensitive individuals. Our study indicates that high dietary potassium intake reduces blood pressure and ADMA levels while increasing NO bioactivity in normotensive salt-sensitive but not salt-resistant Asian subjects after salt loading.     

Effect of folic acid and antioxidant vitamins on endothelial dysfunction in patients with coronary artery disease

OBJECTIVES: The purpose of this study was to determine whether lowering homocysteine levels with folic acid, with or without antioxidants, will improve endothelial dysfunction in patients with coronary artery disease (CAD). BACKGROUND: Elevated plasma homocysteine levels are a risk factor for atherosclerosis. Homocysteine may promote atherogenesis through endothelial dysfunction and oxidative stress. METHODS: In a double-blind, placebo-controlled, randomized trial, we used vascular ultrasound to assess the effect of folic acid alone or with antioxidants on brachial artery endothelium-dependent flow-mediated dilation (FMD). Seventy-five patients with CAD (screening homocysteine level > or =9 micromol/liter) were randomized equally to one of three groups: placebo, folic acid alone or folic acid plus antioxidant vitamins C and E. Patients were treated for four months. Plasma folate, homocysteine, FMD and nitroglycerin-mediated dilation were measured before and after four months of treatment. RESULTS: Plasma folate, homocysteine and FMD were unchanged in the placebo group. Compared with placebo, folic acid alone increased plasma folate by 475% (p < 0.001), reduced plasma homocysteine by 11% (p = 0.23) and significantly improved FMD from 3.2 +/- 3.6% to 5.2 +/- 3.9% (p = 0.04). The improvement in FMD correlated with the reduction in homocysteine (r = 0.5, p = 0.01). Folic acid plus antioxidants increased plasma folate by 438% (p < 0.001), reduced plasma homocysteine by 9% (p = 0.56) and insignificantly improved FMD from 2.6 +/- 2.4% to 4.0 +/- 3.7% (p = 0.45), as compared with placebo. Nitroglycerin-mediated dilation did not change significantly in any group. CONCLUSIONS: Folic acid supplementation significantly improved endothelial dysfunction in patients with coronary atherosclerosis. Further clinical trials are required to determine whether folic acid supplementation may reduce cardiovascular events.     

Folic acid improves endothelial dysfunction in type 2 diabetes--an effect independent of homocysteine-lowering

Diabetes is associated with endothelial dysfunction, which in part may be related to uncoupling of the endothelial nitric oxide (NO) synthase enzyme, thus reducing the availability of NO. As folates may potentially reverse the uncoupling of NO synthase, we wanted to determine whether folic acid supplementation could modulate endothelial function and markers of inflammation in patients with type 2 diabetes without vascular disease. Nineteen patients with type 2 diabetes were treated with folic acid (10mg/day for 2 weeks) versus placebo in a randomized, placebo-controlled, cross-over study with an 8-week washout period between treatments. Fasting endothelium-dependent flow-mediated dilatation (FMD) of the brachial artery, endothelium-independent nitroglycerin-mediated dilatation (NMD), plasma homocysteine, serum lipids, folate, and inflammatory markers (high-sensitivity C-reactive protein, soluble intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, interleukin-18, tumor necrosis factor-alpha) were assessed after each 2-week treatment period. Folic acid supplementation significantly increased folate levels and lowered plasma homocysteine levels. Folic acid significantly improved FMD compared to placebo (5.8 +/- 4.8% vs 3.2 +/- 2.7%, p = 0.02). There were no significant effects of folic acid supplementation on lipids, NMD, or the inflammatory markers. There was no relationship between the change in homocysteine and the improvement in FMD. Thus, 2 weeks of folic acid supplementation can improve endothelial dysfunction in type 2 diabetics independent of homocysteine-lowering, but does not modulate markers of inflammation.     

Oral L-arginine and vitamins E and C improve endothelial function in women with type 2 diabetes

Endothelial dilator function is impaired in people with type 2 diabetes mellitus (T2DM). Prior research indicates that this can be improved with intravenous administration of ascorbate or L-arginine, but whether these agents have this effect when administered by the clinically practical oral route is unknown. To investigate this question, 10 premenopausal women with T2DM and 10 healthy, premenopausal, non-diabetic women received, in random sequence, a 1-week administration of oral L-arginine (9 g daily) or vitamins E (1800 mg) and C (1000 mg) with an intervening 1-week washout period. Flow-mediated brachial artery dilation (FMD) was measured by ultrasonography and forearm blood flow was measured by plethysmography before and following blood pressure cuff-induced forearm ischemia before and after each week of treatment. At baseline, the women with T2DM had lesser FMD responses (0.028 +/- 0.006 cm vs 0.056 +/- 0.008 cm, p < 0.05). Post-ischemic forearm hyperemia was reduced at baseline in T2DM compared with controls (16.4 +/- 1.8 vs 26.0 +/- 1.4 ml 100 ml(-1) min(-1), p < 0.05). Administration of L-arginine caused a 50 +/- 12% increase in FMD in T2DM (p < 0.05) and raised post-ischemic forearm blood flow by 29 +/- 8% (p < 0.05). No significant changes were seen in controls. Administration of vitamins E and C in women with T2DM produced an increase in the brachial artery diameter response of 79 +/- 15% (p < 0.05), but did not significantly increase the hyperemic blood flow response (p = NS). No significant changes in the responses of controls from pre to post vitamin administration were observed. We concluded that administration of two types of oral agents improved measures of endothelial function in people with T2DM.     

Oral snuff impairs endothelial function in healthy snuff users

AIMS: Oral snuff is a less dangerous drug and therefore a good substitute for cigarette smoking. The aim of this study was to determine whether oral moist snuff induces acute endothelial dysfunction. Previous studies have shown that endothelial dysfunction predicts cardiovascular morbidity. METHODS AND RESULTS: Twenty healthy middle-aged snuff users underwent ultrasound assessment of endothelial-dependent flow-mediated dilatation (FMD) of the brachial artery. FMD measurements were performed in duplicate at baseline and then 20 and 35 min after the administration of 1 g portion-bag-packed moist snuff or placebo. Ten of the subjects were examined twice according to a randomized cross-over procedure, once with snuff and once with placebo. All images were arbitrarily analysed off-line by a single blinded observer. FMD values declined significantly from 3.4 +/- 2.0% to 2.3 +/- 1.3% (P < 0.05) 35 min after the administration of 1 g oral moist snuff. Heart rate, systolic and diastolic blood pressure increased significantly (P < 0.05) after snuff administration. All parameters remained unchanged after placebo. CONCLUSIONS: Oral moist snuff significantly impaired FMD of the brachial artery. As endothelial dysfunction predicts cardiovascular morbidity, use of oral snuff should be discouraged.     

Effects of vitamin E on chronic and acute endothelial dysfunction in smokers

OBJECTIVES: The aims of this study were to determine whether chronic or acute impairment of flow mediated vasodilation (FMD) in the brachial artery of smokers can be restored or preserved by the antioxidant vitamin E. BACKGROUND: Transient impairment of endothelial function after heavy cigarette smoking and chronic endothelial dysfunction in smokers result at least in part from increased oxidative stress. METHODS: We studied 22 healthy male smokers (mean +/- SD, 23 +/- 9 cigarettes per day) randomly assigned to receive either 600 IU vitamin E per day (n = 11, age 28 +/- 6 years) or placebo (n = 11, age 27 +/- 6 years) for four weeks and 11 age-matched healthy male nonsmokers. Flow mediated vasodilation and endothelium-independent, nitroglycerin-induced dilation were assessed in the brachial artery using high resolution ultrasound (7.5 MHz) at baseline and after therapy. Subjects stopped smoking 2 h before the ultrasound examinations. At the end of the treatment period, a third scan was obtained 20 min after smoking a cigarette (0.6 mg nicotine, 7 mg tar) to estimate transient impairment of FMD. RESULTS: Flow mediated vasodilation at baseline was abnormal in the vitamin E (5.3 +/- 3.8, p < 0.01) and in the placebo group (6.4 +/- 3.5, p < 0.05) compared with nonsmoking controls (11.6 +/- 4.7). Using a two-way repeated measures analysis of variance (ANOVA) to examine the effects of vitamin E on FMD, we found no effect for the grouping factor (p = 0.5834) in the ANOVA over time but a highly significant difference with respect to time (p = 0.0065). The interaction of the time factor and the grouping factor also proved to be significant (p = 0.0318). Flow mediated vasodilation values remained similar after treatment for four weeks in both groups but declined faster after smoking a cigarette in subjects taking placebo compared with those receiving vitamin E (p values from successive differences for the time/group factor: 0.0001/0.0017). The transient attenuation of FMD (calculated as the percent change in FMD) was related to the improvement of the antioxidant status, estimated as percent changes in thiobarbituric acid-reactive substances (r = -0.67, p = 0.0024). Nitroglycerin-induced dilation did not differ between study groups at baseline or after therapy. CONCLUSIONS: These results demonstrate that oral supplementation of vitamin E can attenuate transient impairment of endothelial function after heavy smoking due to an improvement of the oxidative status but cannot restore chronic endothelial dysfunction within four weeks in healthy male smokers.     

Effect of chronic oral supplementation with vitamins on the endothelial function in chronic smokers

Cigarette smoking has been associated with endothelial dysfunction including impaired endothelium-dependent flow-mediated dilation (FMD). In cigarette smokers, increased oxygen-derived free radicals have been suspected of being one of the major causes of endothelial dysfunction, owing possibly to the inactivation of nitric oxide by free radicals. Vitamins C and E are widely used antioxidant vitamins, which have also been reported to effectively improve the endothelial function in several conditions. To test the effect of moderate-term oral antioxidant vitamin supplementation on the endothelial function in smokers, the authors evaluated the combined effect of vitamins C and E, administered in normal dosages, on FMD in young male smokers. A prospective interventional study was performed. In 15 healthy male subjects (mean age, 24.4 +/-2.5 years old). They studied FMD in the brachial artery by using high-resolution ultrasound. The vascular effects of moderate-term oral supplementation with vitamin C (1.0 g/day) and vitamin E (500 mg/day) were determined during reactive hyperemia, which causes endothelium-dependent FMD. They performed a vascular function study 3 times including before vitamin supplement, after 25 days of vitamin supplement, and 4 weeks after the cessation of the vitamin supplement. The flow-mediated dilator response measurements were repeated twice a day before vitamin supplements, and the repeatability obtained from these measurements was found acceptable (variability of FMD <2%). The oral antioxidant vitamin supplement significantly restored FMD (3.8 +/-2.2% vs 5.9 +/-2.5%; p<0.05), however, this effect disappeared 4 weeks after the vitamin supplementations ended. The combined usual dosage of vitamins C and E supplements was found to improve the endothelial function in chronic smokers.     

Endothelial function in patients with obstructive sleep apnea syndrome but without hypertension

BACKGROUND: Obstructive sleep apnea syndrome (OSAS) influences endothelial function and causes hypertension. OBJECTIVES: Our aim was to evaluate the role of endothelial dysfunction in the pathogenesis of hypertension in OSAS. METHODS: Twenty-three patients with OSAS but without hypertension and 15 healthy normotensive subjects were investigated. The presence or absence of OSAS was evaluated with a sleep study. Endothelial function was investigated with brachial artery ultrasound examination. RESULTS: Baseline characteristics were equivalent between the two groups. Minimal oxygen saturation and apnea-hypopnea indexes in the OSAS and control groups were 62.9 +/- 16.5 versus 94.9 +/- 1.1% (p < 0.0001) and 53.1 +/- 20.3 versus 3.8 +/- 0.9 (p < 0.0001), respectively. There was not statistically significant difference between basal brachial artery diameters measured in the morning and in the evening in all groups. Flow-mediated dilation (FMD) values measured in the morning were lower than those measured in the evening in both OSAS patients and the control group: FMD of OSAS patients was 6.04 +/- 3.18% in the morning and 10.38 +/- 4.23% in the evening hours (p = 0.001), and FMD of control subjects was 10.9 +/- 2.6% in the morning and 13.9 +/- 2.32 in the evening hours (p = 0.002). Differences in FMD values measured both in the morning and evening hours in OSAS patients were lower compared with those in control subjects (p < 0.0001 in the morning hours and p = 0.003 in the evening hours). CONCLUSIONS: We detected a prominent diurnal deterioration in endothelial function in normotensive OSAS patients compared with healthy subjects. This deterioration may occur due to ongoing hypoxemia during the night and it may be a possible cause of hypertension and atherosclerotic cardiovascular diseases in patients with OSAS.     

Plasma nitric oxide level and its role in slow coronary flow phenomenon

Previous studies have suggested that microvascular abnormalities and endothelial dysfunction cause slow coronary flow (SCF). The objective of this study was to assess the plasma nitric oxide (NO) level and determine its role in the pathogenesis of SCF phenomenon. Thirty-six patients with SCF (group 1) and otherwise patent coronary arteries and 34 subjects with normal coronary flow (group 2) were included in the study. Coronary flow was quantified according to the TIMI Frame Count (TFC) method. Brachial artery endothelium-dependent flow-mediated dilatation (FMD) and nitroglycerin (NTG)-induced endothelium-independent dilatation were studied in both groups. In addition, plasma NO levels were measured and their contribution to FMD was determined. The sex, age, body mass index, arterial blood pressure, and heart rate distributions were similar in both groups. TFC was significantly higher in group 1 compared to group 2 for each artery. The plasma NO level was lower in patients with SCF than in control subjects (18.4 +/- 4.4 versus 25.2 +/- 6.3 micromol/L P = 0.001). FMD was significantly smaller in group 1 than in group 2 (4.0 +/- 3.2% versus 10.6 +/- 5.8%, P = 0.0001). The percent NTG-induced dilatation was similar in the two groups (16.8 +/- 1.1% versus 17.1 +/- 1.1%, P = 0.42). In group 1, the plasma NO level was correlated with percent of FMD. Also, the plasma NO level was inversely correlated with TFC for each artery. Reduced NO bioactivity as well as impaired FMD support the presence of endothelial damage in the pathogenesis of SCF phenomenon.     

Role of asymmetrical dimethylarginine in inflammation-induced endothelial dysfunction in human atherosclerosis

We explored the role of asymmetrical dimethylarginine (ADMA) as a cause of endothelial dysfunction induced by systemic inflammation. In vitro data suggest that ADMA bioavailability is regulated by proinflammatory stimuli, but it is unclear whether ADMA is a link between inflammation and endothelial dysfunction in humans. In study 1 we recruited 351 patients with coronary artery disease (CAD) and 87 healthy controls. In study 2 we recruited 69 CAD, 69 healthy, and 10 patients with rheumatoid arthritis, whereas in study 3, 22 healthy and 70 CAD subjects were randomly assigned to Salmonella typhii vaccination (n=11 healthy and n=60 CAD) or placebo (n=11 healthy and n=10 CAD). Circulating interleukin 6/ADMA and flow-mediated dilation (FMD) were measured at 0 and 8 hours. In study 1, ADMA was inversely correlated with FMD in healthy individuals and CAD patients (P<0.0001 for both). However, interleukin 6 was inversely correlated with FMD (P<0.0001) in healthy subjects but not in CAD patients. The positive correlation between ADMA and interleukin 6 was stronger in healthy (r=0.515; P<0.0001) compared with CAD (r=0.289; P=0.0001) subjects. In study 2, both patients with rheumatoid arthritis and CAD had higher interleukin 6 (P<0.0001) and ADMA (P=0.004) but lower FMD (P=0.001) versus healthy subjects. In study 3, vaccination increased interleukin 6 in healthy (P<0.001) and CAD (P<0.001) subjects. FMD was reduced in healthy subjects (P<0.05), but its reduction in CAD was borderline. Vaccination increased ADMA only in healthy subjects (P<0.001). Systemic, low-grade inflammation leads to increased ADMA that may induce endothelial dysfunction. This study demonstrated that ADMA may be a link between inflammation and endothelial dysfunction in humans.     

Oral vitamin C and endothelial function in smokers: short-term improvement, but no sustained beneficial effect

OBJECTIVES: To test the hypothesis that antioxidant therapy would improve endothelial function in smokers. BACKGROUND: Several studies have documented a beneficial effect of short-term oral or parenteral vitamin C on endothelial physiology in subjects with early arterial dysfunction. Possible long-term effects of vitamin C on endothelial function, however, are not known. METHODS: We studied the effects of short- and long-term oral vitamin C therapy on endothelial function in 20 healthy young adult smokers (age 36 +/- 6 years, 8 male subjects, 21 +/- 10 pack-years). Each subject was studied at baseline, 2 h after a single dose of 2 g vitamin C and 8 weeks after taking 1 g vitamin C daily, and after placebo, in a randomized double-blind crossover study. Blood samples were analyzed for plasma ascorbate levels and endothelial function was measured as flow-mediated dilation of the brachial artery, using high resolution ultrasound. Nitroglycerin-mediated dilation (endothelium-independent) was also measured at each visit. RESULTS: At baseline, plasma ascorbate level was low in the smokers (42 +/- 21 micromol/liter; normal range, 50 to 150 micromol/liter), increased with vitamin C therapy after 2 h to 120 +/- 54 micromol/liter (p < 0.001) and remained elevated after eight weeks of supplementation at 92 +/- 32 micromol/liter (p < 0.001, compared with placebo). Flow-mediated dilation, however, increased at 2 h (from 2.8 +/- 2.0% to 6.3 +/- 2.8%, p < 0.001), but there was no sustained beneficial effect after eight weeks (3.9 +/- 3.2%, p = 0.26). Nitroglycerin-mediated dilation was unchanged throughout. CONCLUSION: Oral vitamin C therapy improves endothelial dysfunction in the short term in healthy young smokers, but it has no beneficial long-term effect, despite sustained elevation of plasma ascorbate levels.     

Asymmetric dimethylarginine and coronary collateral vessel development

INTRODUCTION: Nitric oxide (NO) plays a major role in collateral vessel development. Asymmetric dimethylarginine (ADMA) that is an endogenous inhibitor of NO synthesis may impair the effective coronary collateral vessel development. The aim of this study was to evaluate the relationship between plasma ADMA level and coronary collateral vessel development. METHODS: The patients with a greater than or equal to 95% obstruction in at least one epicardial coronary artery were included in the study. Degree of coronary collateral development was determined according to Rentrop method. Patients with grade 2-3 collateral development were regarded as good collateral group and formed group I. The patients with grade 0-1 collateral development were regarded as poor collateral group and were included in group II. Group III that had been formed as a control group included the patients with a normal coronary angiogram. We compared the plasma ADMA, symmetric dimethylarginine, L-arginine/ADMA ratio among three groups. RESULTS: Seventy-four patients have been included in the study. Patients with good collateral development had lower plasma ADMA level in comparison with patients with poor collateral development (0.41+/-0.25 micromol/l vs. 0.70+/-0.23 micromol/l, P=0.001) and had similar plasma ADMA levels with the patients who have normal coronary arteries. When we compared L-arginine/ADMA ratio between good and poor collateral groups, we found that the patients with higher L-arginine/ADMA ratio have significantly better collateral development (270.8+/-168.0 vs. 120.9+/-92.1, P<0.001). In the analyses comparing Rentrop score with ADMA level and L-arginine/ADMA ratio, there were significant correlations (r=-0.444, P=0.008 and r=0.553, P=0.001, respectively). In multivariate analysis, ADMA level (odds ratio, 0.009; 95% confidence interval, 0.000-0.466, P=0.020) and L-arginine/ADMA ratio (odds ratio, 1.010; 95% confidence interval, 1.001-1.020, P=0.032) were independent predictors of collateral development. CONCLUSION: Increased plasma ADMA levels are related with poor coronary collateral development. ADMA may be responsible for the difference in coronary collateral vessel development among different patients with coronary artery disease. NO inhibitors that have a determinative relation with endothelial cell functions may be integral prerequisite in all steps of collateral development.     

Improved endothelium dependent vasodilation in endurance athletes and its relation with ACE I/D polymorphism.

BACKGROUND: Aerobic exercise enhances endothelium-dependent vasodilation in healthy individuals. It is thought that exercise increases nitric oxide (NO) production and decreases NO inactivation, leading to an increase in NO bioavailability. Angiotensin II and NO have important roles in maintaining vascular tone. There are polymorphisms of the angiotensin converting enzyme (ACE) gene and the presence of the deletion (D) allele has been associated with higher concentrations of circulating and tissue ACE. In this study, the relationship between endothelial function and ACE gene polymorphisms was investigated in athletes and sedentary subjects. METHODS AND RESULTS: The study group comprised 56 endurance athletes and 46 sedentary subjects who underwent brachial artery ultrasonographic examination. ACE insertion (I) and D allele frequencies were analyzed in all patients. Baseline brachial artery diameter and resting blood flow were similar in athletes and controls (p > 0.05). The flow-mediated dilation (FMD) was 8.48+/-3.65% in athletes and 5.16+/-2.5% in controls (p = 0.0001). FMD was significantly different between ACE genotypes in the athletes (p < 0.0001): it was higher in ACE II (10.5+/-1.6%) subjects than in the DI (8.4+/-2.3%) or DD (7+/-1.2%) subgroups. CONCLUSION: Regular isotonic exercise can improve endothelium-dependent vasodilation especially in those with the ACE II genotype.     

Impaired progenitor cell activity in age-related endothelial dysfunction

OBJECTIVES: We investigated whether human age-related endothelial dysfunction is accompanied by quantitative and qualitative alterations of the endothelial progenitor cell (EPC) pool. BACKGROUND: Circulating progenitor cells with an endothelial phenotype contribute to the regeneration and repair of the vessel wall. An association between the loss of endothelial integrity and EPC modification may provide a background to study the mechanistic nature of such age-related vascular changes. METHODS: In 20 old and young healthy individuals (61 +/- 2 years and 25 +/- 1 year, respectively) without major cardiovascular risk factors, endothelial function, defined by flow-mediated dilation of the brachial artery via ultrasound, as well as the number and function of EPCs isolated from peripheral blood, were determined. RESULTS: Older subjects had significantly impaired endothelium-dependent dilation of brachial artery (flow-mediated dilation [FMD] 5.2 +/- 0.5% vs. 7.1 +/- 0.6%; p < 0.05). Endothelium-independent dilation after glycerol trinitrate (GTN) was not different, but the FMD/GTN ratio was significantly lower in old subjects (49 +/- 4% vs. 37 +/- 3%; p < 0.05), suggesting endothelial dysfunction. There were no differences in the numbers of circulating EPCs, defined as CD34/KDR or CD133/KDR double-positive cells in peripheral blood. In contrast, lower survival (39 +/- 6 cells/mm(2) vs. 65 +/- 11 cells/mm(2); p < 0.05), migration (80 +/- 12 vs. 157 +/- 16 cells/mm(2); p < 0.01), and proliferation (0.20 +/- 0.04 cpm vs. 0.44 +/- 0.07 cpm; p < 0.05) implicate functional impairment of EPCs from old subjects. The FMD correlated univariately with EPC migration (r = 0.52, p < 0.05) and EPC proliferation (r = 0.49, p < 0.05). Multivariate analysis showed that both functional features represent independent predictors of endothelial function. CONCLUSIONS: Maintenance of vascular homeostasis by EPCs may be attenuated with age based on functional deficits rather than depletion of CD34/KDR or CD133/KDR cells.