Plasma Nitric Oxide Metabolites Research Articles

Antioxidant and Vascular Protective Activities of Cratoxylum formosum, Syzygium gratum and Limnophila aromatica

Phytochemicals contained in dietary plants provide a variety of health benefits and may reduce the risk of cardiovascular diseases. The aqueous extracts from three popular Thai dietary and herbal plants, Cratoxylum formosum, Syzygium gratum, and Limnophila aromatica, were investigated for the antioxidant and vascular protective activities in the in vitro and in vivo models. The free radical scavenging and antioxidant activities of plant extracts were evaluated in vitro by the 1,1-diphenyl-2-picrylhydrazyl assay, the ferric reducing antioxidant power assay, the intracellular antioxidant activity in rat peritoneal macrophages by dihydrofluorescein assay, and the inhibition of nitric oxide (NO) production in RAW 264.7 macrophages. In an animal model of oxidative stress and vascular dysfunction, male Sprague-Dawley rats were orally administered with aqueous plant extracts (1 g/kg/d) or N-acetylcysteine (NAC; 300 mg/kg/d) as a control for 6 d. On day four, all animals except the normal control group, were administered with phenylhydrazine (PHZ) intraperitoneally. It was demonstrated that the plant extracts possessed high free radical scavenging and antioxidant activities. PHZ induced severe hemolysis and hemodynamic disturbances and treatment with the extracts and NAC significantly improved the hemodynamic status. Vascular responsiveness to bradykinin, acetylcholine, and phenylephrine in PHZ-control rats was markedly impaired, and the plant extracts or NAC largely restored the vascular responses. Moreover, the plant extracts prevented loss of blood reduced glutathione and suppressed formation of plasma malondialdehyde, plasma NO metabolites and blood superoxide anion. It was concluded that the plant extracts possess antioxidants and have potential roles in protection of vascular dysfunction.

Modification of CYP2E1 and CYP3A4 activities in haemoglobin E-beta thalassemia patients

Thalassemia disease is a genetic haemoglobinopathy usually associated with an iron overload and some degree of organ impairment. The impact of the disease on the drug metabolising enzyme cytochrome P450 (CYP) is not known. CYP2E1 and CYP3A4 are responsible for the metabolism of a large number of drugs and changes in their activities may have clinical consequences. Haemoglobin E-beta thalassemia paediatric, blood transfusion-dependent patients apparently without complications (n = 35) and healthy controls (n = 42) were recruited in this study. The ratios of plasma 6-hydroxychlorzoxazone to chlorzoxazone, and urinary 6-beta-hydroxycortisol (6beta-OHF) to cortisol were used as indices for CYP2E1 and CYP3A4 activities, respectively. Blood and plasma samples were assayed for parameters of clinical biochemistry, oxidants and antioxidants. There were significant increases in serum iron, protein carbonyl and lipid peroxidation in thalassemia patients, whereas there was a decrease in blood glutathione, but unchanged plasma nitric oxide metabolites. CYP2E1 activity in the patients was unchanged; however, when the patients were stratified by splenectomy status, CYP2E1 activity was increased in non-splenectomised patients in comparison with the controls and splenectomised subjects. On the other hand, 6beta-OHF/cortisol ratios increased markedly in patients associated with depressed growth hormone levels. There were no correlations between CYP2E1 activity and oxidant stress or antioxidant parameters. This report is the first demonstration that thalassemia major is associated with an alteration of CYP2E1 and CYP3A4 activities; this could modify the sensitivity of thalassemia patients to the toxic or therapeutic effects of drugs.

Adiponectin and Membrane Fluidity of Erythrocytes in Normotensive and Hypertensive Men

Abnormalities in physicochemical properties of the cell membranes may underlie the defects that are strongly linked to hypertension. Recent evidence indicates that adiponectin may have protective effects against cardiovascular diseases. The purpose of the present study was to assess the possible link between plasma adiponectin and membrane fluidity in normotensive (NT) and hypertensive (HT) men. We measured the membrane fluidity (a reciprocal value of membrane microviscosity) of erythrocytes in NT and HT men by using an electron paramagnetic resonance and spin-labeling method. The order parameter (S) for the spin label agent (5-nitroxide stearate) and the peak height ratio (h0/h(-1)) for 16-nitroxide stearate in the electron paramagnetic resonance spectra of erythrocytes were significantly higher in HT men than in NT men, indicating that membrane fluidity of erythrocytes was decreased in HT men compared with NT men. Both of plasma adiponectin and nitric oxide (NO) metabolite levels were significantly lower in HT men than in NT men. The plasma adiponectin levels were correlated with plasma NO metabolites. The S and the h0/h(-1) of erythrocytes were inversely correlated with the plasma adiponectin and NO metabolite levels, indicating that the decreased membrane fluidity of erythrocytes was associated with hypoadiponectinemia and reduced plasma NO metabolites. The results of the present study demonstrated that plasma adiponectin levels were lower in HT men than in NT men and that hypoadiponectinemia was associated with decreased membrane fluidity of erythrocytes. The finding suggests that adiponectin may be linked to the rheologic behavior of the erythrocytes and the microcirculation in men, at least in part, by the NO-dependent mechanism.

Increased Plasma Nitric Oxide Metabolites in Suicide Attempters

Objective: To evaluate any correlation between plasma levels of nitric oxide metabolites (NO_x) and suicide attempt. Method: Plasma NO_x levels were measured in 53 patients who had recently attempted suicide, 58 nonsuicidal psychiatric patients, and 75 normal controls. The severity of suicidal behaviors was evaluated using Weisman and Worden’s Risk-Rescue Rating Scale. Results: Plasma NO_x levels were significantly higher in suicidal patients than nonsuicidal psychiatric patients or normal control subjects (F = 11.029, d.f. = 2, 183, p < 0.001). Among the patients with a diagnosis of major depression, suicidal depressive patients had significantly higher plasma NO_x levels than nonsuicidal depressive patients (t = –3.090, d.f. = 84, p = 0.003). Conclusion: Our study suggests that increased NO production in plasma is associated with suicide attempt, especially in depressive patients.

Plasma Leptin and its Relationship with Lipid Peroxidation and Nitric Oxide in Obese Female Patients with or without Hypertension

Recent evidence suggested that leptin-induced oxidative stress in human endothelial cells in vivo and increased oxidative stress in human essential hypertension may further contribute to both the development of atherosclerosis and other cardiovascular diseases. We investigated the association of plasma leptin levels with plasma lipid peroxidation and nitric oxide metabolites (NOx) in obese hypertensive atherosclerosis model. Plasma leptin, lipid peroxidation and NOx levels were determined in age-matched non-obese normotensive female subjects (n = 30), obese normotensive female subjects (n = 45), and obese hypertensive female subjects (n = 50). Plasma leptin levels were determined by immunoradiometric method. Lipid peroxidation was determined as thiobarbituric acid reactive substances (TBARS) using spectrophotometric method. NOx levels were determined using enzymatic method. We found that plasma leptin and TBARS levels were increased in obesity, and obese hypertensives have significantly higher plasma leptin and TBARS levels than obese normotensives (p <0.001 and p <0.001). Obese hypertensives have significantly lower plasma NOx levels than obese normotensives (p <0.001). In univariate and multivariate regression analysis, plasma leptin levels were significantly correlated with TBARS (p <0.01 and p <0.01) in obese subjects. Plasma TBARS were also inversely correlated with NOx in hypertensive obese subjects (r = -0.412, p <0.01). Our results have shown that elevated leptin levels may be associated with increased oxidative stress and free-radical-induced decreased NOx levels. Therefore, hyperleptinemia may be an important contributor to the generation of hypertension in obesity.

Increased plasma nitric oxide level associated with suicide attempt in depressive patients

Background Nitric oxide (NO) is known to influence cerebral monoaminergic activity, including the activity of serotonin. We evaluated plasma NO metabolite (NO x ) levels in depressive patients with and without a recent history of suicide attempt. Method Plasma NO x levels were measured in 39 depressive patients who had recently attempted suicide, 44 non-suicidal depressed patients, and 70 normal controls. The severity of depression was measured with the Hamilton's Depression Rating Scale. The lethality of the suicide attempt was scored using Weisman and Worden's risk-rescue rating scale and Lethality Suicide Attempt Rating Scale. Results Plasma NO x levels were significantly higher in suicidal depressive patients than non-suicidal depressive patients or normal control subjects ( Z = − 2.472, p = 0.013). However, higher plasma NO x levels in suicidal depressive patients were significantly related to a lower lethality of suicide attempts and lower severity of depression. Conclusions Our study suggests that increased plasma NO x level is associated with suicide attempts in depressive patients. Moreover, higher plasma NO x level is related with suicide attempts in mild depressed patients. However, further studies are required to understand the role of NO system in depression and suicidal behavior.

Estrogen therapy replenishes vascular tetrahydrobiopterin and reduces oxidative stress in ovariectomized rats

We investigated whether the effect of estrogen therapy on vascular endothelial function is mediated through increasing the bioavailability of tetrahydrobiopterin (BH4) and associated antioxidant capacity in ovariectomized (Ovx) rats. Aortas of sham-operated, Ovx, and Ovx plus estrogen therapy (Ovx + ET) female Sprague-Dawley rats were used to measure vascular reactivity. Plasma levels of nitric oxide (NO) metabolites, total antioxidant capacity, aortic superoxide anion (O2.), and BH4 contents were determined. Vascular reactivity, assessed on isolated aortic segments, indicated that phenylephrine-induced contraction in the Ovx group was significantly greater than that in the sham and Ovx + ET groups. The vasodilator responses to acetylcholine (10 to 10 M) and L-arginine (L-Arg; 10 M) in the sham and Ovx + ET groups were significantly greater than those in the Ovx group. Pretreatment with BH4 (10 M) enhanced the vasodilator responses to L-Arg in the Ovx group compared with the untreated Ovx group. An inhibitor of BH4 synthesis, 2,4-diamino-6-hydroxypyrimidine (2 mM), significantly attenuated the vasodilator response to L-Arg in the sham and Ovx + ET groups. In addition, Ovx significantly increased O2. production in aortic tissues and decreased plasma NO metabolites levels, whereas ET significantly prevented these effects. Pretreatment with BH4 also significantly decreased aortic O2. production in the Ovx group; both plasma total antioxidant capacity and aortic BH4 contents in the Ovx group decreased significantly compared with those in the sham group, which were also improved by ET. There were no significant differences in the protein expression of endothelial NO synthase in aortas in these groups. ET increases the availability of vascular BH4 to attenuate O2. production and restores total antioxidant capacity, leading to improved NO-mediated vasodilation in Ovx rats.

Role of nitric oxide and endothelium-derived hyperpolarizing factor (EDHF) in the regulation of blood pressure by leptin in lean and obese rats

We investigated the role of nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF) in hemodynamic action of leptin. The effect of leptin (1 mg/kg i.p.) on systolic blood pressure (SBP) was examined in lean rats and in rats made obese by feeding highly palatable diet for either 1 or 3 months. Separate groups received NO synthase inhibitor, l-NAME, or EDHF inhibitors, the mixture of apamin + charybdotoxin or sulfaphenazole, before leptin administration. Leptin increased NO production, as evidenced by increase in plasma and urinary NO metabolites and cyclic GMP. This effect was impaired in both obese groups. In lean rats either leptin or EDHF inhibitors had no effect on blood pressure. l-NAME increased blood pressure in lean animals and this effect was prevented by leptin. However, when leptin was administered to animals pretreated with both l-NAME and EDHF inhibitors, blood pressure increased even more than after l-NAME alone. In the 1-month obese group leptin had no effect on SBP, however, pressor effect of leptin was observed in animals pretreated with EDHF inhibitors. In the 3-month obese group leptin alone increased SBP, and EDHF inhibitors did not augment its pressor effect. The results suggest that leptin may stimulate EDHF when NO becomes deficient, e.g. after NOS blockade or in short-term obesity. Although the effect of leptin on NO production is impaired in the 1-month obese group, BP does not increase, probably because EDHF compensates for NO deficiency. In contrast, leptin increases BP in 3-month obesity because its effect on EDHF is also attenuated.

Homocysteine and nitric oxide in patients undergoing diagnostic coronary angiography.

We evaluated the plasma homocysteine (tHcy) and nitric oxide metabolites (nitrite plus nitrate; NOx) data of consecutive patients undergoing diagnostic coronary angiography (n=79) with respect to the presence and severity of coronary artery disease (CAD), the presence of acute coronary syndromes (ACS), and the risk status of patients. Hyperhomocysteinemia (>15 micromol/L) was detected in 11% of the controls (n=19) and 37% of CAD patients (n=60) (p=0.03). Plasma tHcy in CAD patients was not significantly different from controls, but those with 3-vessel disease had a significantly higher tHcy concentrations than did controls (p=0.049). The patients with 3-vessel disease and ACS had the highest concentrations of tHcy (16.9 +- 4.4 micromol/L), and the difference from the ACS patients with 1- and 2-vessel involvement was significant (p=0.03). In patients with 1-vessel involvement, tHcy was correlated with NOx (r=0.62, p=0.005); in patients with 2- and 3-vessel disease this correlation could not be observed. The high-risk patients (n=51) had a higher mean number of vessel involvement and tHcy (p<0.001, p<0.05, respectively) but lower NOx (p<0.05) when compared to the low-risk patients (n=28). It appears that in the early stages of atherosclerosis hyperhomocysteinemia causes an increase in NOx production, but with progression of the disease this compensatory increase disappears.

P.3.a.012 Low concentration of plasma nitric oxide metabolites in schizophrenic patients

Composition of Indian Russell's viper (Daboia russelii russelii) venom, a medically important snake and member of “Big Four” snakes of India was done by gel filtration chromatography followed by tandem mass spectrometry. The MS/MS analyses of tryptic digested gel filtration peaks divulged the presence of 63 different proteins belonging to 12 families. Phospholipase A2 (PLA2), serine proteases, metalloproteases, cysteine-rich secretory proteins, l-amino acid oxidase, C-type lectin-like proteins, kunitz-type serine protease inhibitor, disintegrin, nucleotidase, phosphodiesterase, vascular endothelial growth factor and vascular nerve growth factor families were identified. PLA2 enzymes with isoforms of N-, S- and H-type based on their first N-terminal amino acid residue were observed. The venom is also found to be rich in RVV-X, RVV-V and thrombin-like enzymes. Homologues of disintegrins with RGD and RTS motifs were also observed. The high percentage of PLA2 and proteases in the venom proteome could be responsible for the observed coagulopathy, haemorrhage and edema which can be correlated with the clinical manifestations of Russell's viper envenomation. This is the first proteomic analysis of Indian D. russelii venom which might assist in understanding the pathophysiological effects of viper envenomation. Such study will also be important for developing more effective antivenom for viper bite management.

ROLE OF INTERLEUKIN 18 IN NITRIC OXIDE PRODUCTION AND PANCREATIC DAMAGE DURING ACUTE PANCREATITIS

The release of the immunomodulator, interleukin 18 (IL-18) into sera early in acute pancreatitis (AP) corresponds to disease severity. IL-18 induces nitric oxide (NO), which is involved in the pathophysiology of pancreatitis. The objective of this study was to clarify the role of IL-18 in pathogenesis and NO production during early AP using recombinant mouse (rm) IL-18 protein and IL-18 gene knockout (KO) mice. After pretreatment with phosphate-buffered saline or rmIL-18, wild-type (WT) or KO mice were injected intraperitoneally with phosphate-buffered saline (sham) or cerulein (AP) hourly for 3 h. Blood, pancreas, spleen, and liver were collected until 24 h after the first dose. Main outcome measures were serum IL-18, amylase and lipase levels, histological evaluation of the pancreas with parenchyma vacuolization of acinar cells, mRNA expression of inducible NO synthase (iNOS) in the pancreas, and spleen, liver, and plasma NO metabolite level. Serum IL-18 was significantly increased immediately after induction of AP in WT mice. Serum amylase, lipase, and the numbers of acinar cells with parenchyma vacuolization were significantly higher in the group AP/KO than in the group AP/WT, but these parameters were improved by dose-dependent pretreatment with rmIL-18 administration in both groups. Pancreatic iNOS gene expression and plasma NO metabolites were significantly increased by 6 h after the initiation of AP, but were significantly lower in the group AP/KO than in the AP/WT mice. Pretreatment with rmIL-18 also significantly increased these levels in both groups. Splenic and hepatic iNOS expression was not changed after the initiation of AP in WT mice, whereas pretreatment with rmIL-18 also increased these levels. Administration of aminoguanidine, a selective iNOS inhibitor, before AP induction abolished the protective effect of pretreatment with rmIL-18 on pancreatic injury. IL-18 appears to protect the pancreas during early induced-induced AP in mice, probably through induction of NO release from an iNOS source. IL-18 may be a target for new AP therapeutics.

Alteration of Decreased Plasma NO Metabolites and Platelet NO Synthase Activity by Paroxetine in Depressed Patients

Although major depression (MD) and cardiovascular disease (CVD) have been conclusively linked in the literature, the mechanism associating MD and CVD is yet undetermined. The purpose of this paper is to further investigate a potential mechanism involving nitric oxide (NO) and to examine the effect of the selective serotonin reuptake inhibitor paroxetine on NO production by both platelets and the endothelium. In total, 17 subjects with MD and 12 healthy controls (HCs) with no known history of cardiovascular illness completed the study. Paroxetine was administered to both the MD patients and HCs over an 8-week period, and then medication was discontinued. Blood samples were taken at various times throughout paroxetine treatment and after discontinuation. Plasma NO metabolite (NOx) levels were measured by a chemiluminescence method. Platelet endothelial NO synthase (eNOS) activity was examined through the conversion of L-[14C]arginine to L-[(14)C]citrulline. Data were analyzed using t-tests and a linear mixed effects model. Baseline levels of both plasma NOx and platelet NOS activity were significantly lower in subjects with MD compared to HCs. Throughout paroxetine treatment, plasma NOx levels increased in both HCs and MD patients. However, platelet eNOS activity decreased in HCs, while no statistically significant change was evidenced in MD patients. These data suggest that, in MD patients, decreased peripheral production of NO, a potential contributor to increased cardiovascular risk, is modified by administration of the antidepressant paroxetine.

Modulation of erythrocyte arginase activity in sickle cell disease patients during hydroxyurea therapy

An elevated erythrocyte arginase activity with a corresponding decrease in nitric oxide (NO) level has been implicated in the pathophysiology of sickle cell disease (SCD). Recent studies have shown that hydroxyurea (HU) increases the production of NO, which increases the soluble guanylate cyclase activity and fetal haemoglobin (HbF) synthesis. To study the effects of HU on the arginase and nitric oxide synthase (NOS) activities in SCD patients, we compared levels of arginase activity and NO metabolites in red blood cells and plasma, respectively, from 23 patients with SCD (HbSS) receiving HU therapy, with those of 12 SCD patients not receiving HU treatment. Patients on HU therapy showed significantly lower arginase activity than that of HbSS patients not on HU therapy (1.36+/-0.2 U/10(8) cells vs. 3.31+/-0.29 U/10(8) cells). NOS activity was higher in patients on HU therapy than in untreated patients (0.72+/-0.4 nmol/ml/min vs. 0.35+/-0.15 nmol/ml/min, P<0.05). Among the HU-treated patients, the decreased level of arginase activity correlated (r=0.71) with HbF level as well as the mean corpuscular haemoglobin content. These data suggest that one of the beneficial effects of HU in vivo may involve the regulation of arginase activity and a concomitant induction of NOS activity, which may lead to an increased production of NO. The outcome of this study may lead to the development of improved NO-based treatments for SCD.

Circulating and excretory nitrite and nitrate as indicators of nitric oxide synthesis in humans: methods of analysis

Nitric oxide (NO) is produced in various cells from l-arginine by the catalytical action of NO synthases (NOS). The main metabolic fate of NO includes oxidation to nitrate by oxyhemoglobin in red blood cells and autoxidation to nitrite. Nitrate and nitrite circulate in blood and are excreted in urine. The concentration of these NO metabolites in plasma, serum, and urine can be used to assess NO synthesis in humans. Circulating nitrite reflects constitutive endothelial NOS activity. Excretory nitrate indicates systemic NO production. Nitrite and nitrate can be measured in plasma, serum, and urine of humans by various analytical methods which are based on different analytical principles. These methods include colorimetry, spectrophotometry, fluorescence, chemiluminescence, gas and liquid chromatography, electrophoresis, and mass spectrometry. The present article gives an overview of the most significant currently used quantitative methods of analysis of nitrite and nitrate in human plasma, serum, and urine in the framework of clinical studies and discusses their significance.

Exposure to a moderate intensity static magnetic field enhances the hypotensive effect of a calcium channel blocker in spontaneously hypertensive rats

We investigated the combined effects of a moderate intensity static magnetic field (SMF) and an L-type voltage-gated Ca(2+) channel blocker, nicardipine in stroke-resistant spontaneously hypertensive rats during the development of hypertension. Five-week-old male rats were exposed to SMF intensity up to 180 mT (B(max)) with a peak spatial gradient of 133 mT/mm for 14 weeks. Four experimental groups of 14 animals each were examined: (1) sham exposure with intraperitoneal (ip) saline injection (control); (2) SMF exposure with ip saline injection (SMF); (3) sham exposure with ip nicardipine injection (NIC); (4) SMF exposure with ip nicardipine injection (SMF + NIC). A disc-shaped permanent magnet or a dummy magnet was implanted in the vicinity adjacent to the left carotid sinus baroreceptor region in the neck of each rat. Nicardipine (2 mg/kg ip) was administered three times a week for 14 weeks, and then 15 min after each injection, arterial blood pressure (BP), heart rate (HR), baroreflex sensitivity (BRS), skin blood flow (SBF), skin blood velocity (SBV), plasma nitric oxide (NO) metabolites (NO(x) = NO(2) (-) + NO(3) (-)), plasma catecholamine levels and behavioral parameters of a functional observational battery were monitored. The action of nicardipine significantly decreased BP, and increased HR, SBF, SBV, plasma epinephrine and norepinephrine in the NIC group compared with the control respective age-matched group without changing plasma NO(x) levels. Neck exposure to SMF alone for 5-8 weeks significantly suppressed or retarded the development of hypertension together with increased BRS in SMF group. Furthermore, the exposure to SMF for 1-8 weeks significantly promoted the nicardipine-induced BP decrease in the SMF + NIC group compared with the respective NIC group. Moreover, the SMF induced a significant increase in plasma NO(x) in the nicardipine-induced hypotension. There were no significant differences in any of the physiological or behavioral parameters measured between the SMF + NIC and the NIC groups, nor between the SMF and the control groups. These results suggest that the SMF may enhance nicardipine-induced hypotension by more effectively antagonizing the Ca(2+) influx through the Ca(2+) channels compared with the NIC treatment alone. Furthermore, the enhanced antihypertensive effects of the SMF on the nicardipine-treated group appear to be partially related to the increased NO(x). Theoretical considerations suggest that the applied SMF (B(max) 40 mT, 0 Hz) can be converted into a changing magnetic field (B(max) 30-40 mT, 5.7-6.5 Hz or 7.5-8.3 Hz) in the baroreceptor region by means of the carotid artery pulsation. Therefore, we propose that the moderate intensity changing magnetic field, i.e., the magnetic field modulated by the pulse rate, may influence the activity of baroreceptor and baroreflex function.

High-fat meal impairs vascular compliance in a subgroup of young healthy subjects

Postprandial hypertriglyceridemia impairs endothelial function and may possibly worsen vascular compliance by increasing oxidative stress. Large (C 1) and small (C 2) artery compliance, glucose, insulin, and triglycerides (TGs) were measured hourly for 6 hours in 18 young healthy volunteers after a low-fat meal and a high-fat meal, with and without antioxidant vitamins. C 1 and C 2 declined significantly for 6 hours after fat ingestion in 8 subjects (“fat reactors”) and increased in 10 (“nonreactors”). Fat reactors had higher fasting and peak serum TGs after fat loading and increased baseline glucose and insulin levels and homeostasis model assessment of insulin resistance (HOMA IR). Fasting insulin correlated with C 1 and C 2 only in fat reactors. After fat intake, plasma nitric oxide metabolites decreased more in fat reactors than in nonreactors (17.0% ± 5.1% vs 4.8% ± 2.1%; P < .05). In fat reactors, pretreatment with antioxidant vitamins before the high-fat meal blunted the fall in C 1 but not in C 2. Compliance was unchanged after the low-fat meal. Normal weight young subjects with an insulin resistance phenotype show significantly decreased vascular compliance, increased postprandial TG peaks, and markedly reduced plasma nitric oxide metabolites after a high-fat meal.

An Association Between Plasma Asymmetric Dimethylarginine and Membrane Fluidity of Erythrocytes in Hypertensive and Normotensive Men: An Electron Paramagnetic Resonance Investigation

Evidence indicates that increased levels of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide (NO) synthase, are associated with increased risk of vascular dysfunction. The present study was performed to assess the possible relationship between plasma ADMA levels and membrane fluidity in hypertensive and normotensive men. Membrane fluidity (a reciprocal value of membrane microviscosity) of erythrocytes in hypertensive (n = 38) and normotensive (n = 35) men were measured using an electron paramagnetic resonance (EPR) and spin-labeling method. The order parameter (S) for the spin-label agent (5-nitroxide stearate) in the EPR spectra of erythrocyte membranes was significantly higher in hypertensive men than in normotensive men (mean +/- SEM: hypertensive subjects, 0.725 +/- 0.001,n = 38; normotensive subjects, 0.713 +/- 0.001, n = 35, P < .001), indicating that membrane fluidity of erythrocytes was decreased in hypertensive men compared with normotensive men. Plasma NO metabolite (nitrite and nitrate) concentration was lower in hypertensive than in normotensive men. In contrast, plasma ADMA levels were higher in hypertensive than in normotensive men. In the overall analysis of hypertensive and normotensive men, the order parameter (S) of erythrocytes was inversely correlated with plasma NO metabolites (r = -0.36, n = 73, P < .01) and positively correlated with plasma ADMA levels (r = 0.44, n = 73, P < .001), indicating that the reduced membrane fluidity of erythrocytes was associated with decreased NO and increased ADMA levels in plasma. When correlation analysis was performed separately for hypertensive and normotensive men, there was a positive correlation between plasma ADMA levels and the order parameter (S) of erythrocytes in both groups. The results of the present study showed that the plasma ADMA levels were elevated in hypertensive men and associated with lower membrane fluidity of erythrocytes, suggesting that ADMA might have a close correlation with the rheologic behavior of erythrocytes and the microcirculation in hypertension.

Ocular blood flow changes after dynamic exercise in humans

To investigate control mechanisms for ocular blood flow changes after dynamic exercise using two different methods. Changes over time in the tissue blood flow in the retina and choroid-retina of healthy volunteers were determined after dynamic exercise (Master's double two-step test), using scanning laser Doppler flowmetry (SLDF) and laser speckle flowgraphy (LSFG). Changes in intraocular pressure (IOP), blood pressure, plasma CO(2) gas concentration (pCO(2)), and levels of nitric oxide (NO) metabolites were examined. Retinal blood flow measured by SLDF increased significantly only at 15 min after exercise. In contrast, normalized blur (NB) values in the choroid-retina, obtained by LSFG, increased significantly up to 60 min after exercise. Ocular perfusion pressure (OPP), calculated from IOP and blood pressure, increased significantly immediately and 15 min after exercise. The plasma NO metabolite levels increased significantly, although pCO(2) levels were unchanged. Dynamic exercise changes OPP and produces increased tissue blood flow in the retina in the immediate postexercise period, while blood flow increases more persistently in the choroid-retina. Difference in control of blood flow in these two regions may be related to stronger autoregulatory mechanism of blood flow in the retina. Nitric oxide may play a role in the regulation of blood flow.

Age‐dependent effect of plasma nitric oxide on parasite density in Ghanaian children with severe malaria

Nitric oxide (NO) has toxic properties against Plasmodium falciparum. While high blood levels have been associated with protection against severe malarial disease, they may also contribute to the pathophysiology of cerebral malaria and severe anaemia. Promoter variants in the inducible nitric oxide synthase (iNOS) gene have been shown to influence NO concentrations and disease manifestation. However, findings are conflicting. We examined associations of plasma NO metabolites (NOx) with symptoms of severe malaria, particularly malarial anaemia and cerebral malaria, and with iNOS promoter variants. In 210 Ghanaian children with severe malaria, we measured plasma nitrite, nitrate, and S-nitrosothiol, and genotyped the iNOS promoter variants -954G-->C, -1173C-->T, and the -2.5 kb (CCTTT)(n) microsatellite. NOx levels decreased with age. In young children (<24 months), high NOx was associated with reduced parasite density. This was not seen in patients of 24-48 months of age and reversed in older children. Subgroup analysis revealed that in children with severe anaemia but without cerebral involvement (prostration, impaired consciousness, convulsions), high NOx levels correlated with low parasitaemia (P = 0.02). In these children, elevated NOx levels were also associated with the iNOS-954C-->T/(CCTTT)(8) haplotype (P = 0.03). No association between NOx or iNOS genotypes and cerebral malaria was observed. Our findings suggest that in young children with severe malaria NOx reduces parasitaemia. This effect wanes at higher ages and may reflect a predominance of unspecific immune responses to infection in early childhood. This finding may have importance for the understanding of associations between iNOS variants and severe malaria in regions of differing disease manifestation.

Association of p22 phox polymorphism with cholesterol levels in African American hypertensives

The p22 phox subunit of NAD(P)H oxidase is a major source of superoxide anions, and contributes to lower biological levels of nitric oxide in the vasculature. Gene polymorphisms of p22 phox and a susceptibility to cardiovascular risk factors, such as increased cholesterol and blood pressure levels remain unclear. The association between C242T polymorphism of p22 phox and plasma nitric oxide metabolites (NOx), cholesterol and blood pressure was determined in 105 normotensive (44±10 yr) and 129 hypertensive (49±10 yr) African Americans excluded for diabetes, prevalent cardiovascular disease, drug abuse and prescribed lipid-lowering medication. The C242T polymorphism was determined by polymerase chain reaction and restriction enzyme digestion assays. The prevalence of the T allele in this population was comparable to that described for Caucasians. Fasting cholesterol levels were higher in hypertensive carriers of the TT genotype (205±25, 199±47, 188±35 mg/dl for TT, CT, CC genotypes respectively, P=0.03), but not among normotensives (190±32, 204±41, 196±32, P=ns). Using a multivariate linear regression model, cholesterol levels were independently associated with the 242T allele (β coefficient = 9.2, P=0.04), after adjusting for age, body mass index and systolic blood pressure. The genotype distribution of the 242T allele did not differ by NOx levels. Findings from this study suggest a relationship between the p22 phox polymorphism and cholesterol levels, manifested when blood pressure levels are elevated.