Changes In Nitric Oxide Metabolism Research Articles

Systemic Hypoxia and Interventions on Nitric Oxide Metabolism Affect Retinal Vasomotion in Healthy Subjects.

Previous in vitro studies on porcine retinal arterioles have shown that the frequency and amplitude of retinal vasomotion can be affected by hypoxia and nitric oxide (NO). However, it is unknown whether these effects can be reproduced in humans in vivo. Video recordings of retinal arterioles from 40 healthy subjects were studied before and during breathing of a hypoxic gas mixture consisting of 12.5% oxygen and 87.5% nitrogen. The experiments were repeated before and after the addition of either an NO donor or continuous infusion of an NO synthesis inhibitor. The harmonic content of spontaneous diameter changes in retinal arterioles was extracted by Fourier analysis, and the effects of the interventions were studied on the amplitudes of diameter oscillations in the frequency bands 1: 0.5/min-3.5/min, 2: 4/min-7.5/min, 3: 8/min-17.5/min, and 4: 18/min-30/min. Hypoxia significantly increased the amplitude of spontaneous diameter oscillations within the low (1) and high (3-4) frequency bands (P < 0.006 for all comparisons), and the effect in frequency band 1 was eliminated by the NO donor. In frequency band 2, hypoxia had no significant effect on the amplitude of diameter oscillations, but the amplitudes were significantly reduced by the NO donor (P < 0.01) and significantly increased by the NO synthesis inhibitor (P = 0.03). Hypoxia and interventions on NO metabolism can affect spontaneous diameter oscillations in retinal arterioles. Disturbances in vasomotion may play a role in hemodynamic changes in retinal diseases in which hypoxia and changes in NO metabolism are involved in the disease pathogenesis.

Endothelin-1 May Be a Potential Biomarker for Monitoring Changes in Nitric Oxide Metabolism in Pulmonary Hypertension

Pulmonary hypertension (PH) is characterized by endothelial cell dysfunction and dysregulation of endogenous nitric oxide (NO) synthesis. NO bioavailability is further decreased due to dysregulation of the oral microbiome and reduction of dietary nitrate (NO3-) to nitrite (NO2-). PH biomarkers such as NT-proBNP have been associated with disease severity, prognosis and response to therapy. Emerging biomarkers like Endothelin-1 (ET-1) reflect NO signaling and cellular dysregulation. We evaluated the effect of a single dose of nitrate on NT-proBNP, ET-1, NO3- and NO2- levels in PH patients versus controls. Patients with prior right heart catheterization were administered 1000mg of sodium nitrate orally. Plasma NT-proBNP, ET-1, NO3- and NO2- were measured at baseline, 2, 6, and 24 hours following nitrate drug administration. Response of patients with PH (mPAP ≥ 25mmHg, TPG ≥ 12mmHg) was compared to controls (mean PAP Nine patients were evaluated (control: n=4, age 53 ± 14 years, 100% female, EF 55 ± 0%, mPAP 19 ± 4 mmHg, PCWP 11 ± 3 mmHg, TPG 9 ± 4 mmHg, PVR 1.5 ± 0.6 WU; PH: n=5, age of 59 ± 9 years, 80% female, EF 58 ± 8%, mPAP 41 ± 8 mmHg, PCWP 26 ± 7 mmHg, TPG 14 ± 3 mmHg, PVR 2.9 ± 0.7 WU). Following oral nitrate dose, mean plasma NO3- and NO2- levels increased in both groups. NO3- levels were significantly greater in the control vs PH group at 2 hrs (803 ± 404 µm vs 571 ± 202 µm) and 6 hrs (634 ± 254 µm vs 482 ± 183 µm) (overall P

Inter- and intra-subject variability of nitric oxide levels in leukocyte subpopulations

Assessment of nitric oxide (NO) dynamics in immune cells, commonly measured using NO surrogates such as inducible nitric oxide synthase (iNOS) rather than NO itself, has been effective in understanding pathophysiology across a wide range of diseases. Although the intracellular measurement of NO is now feasible, many technical issues remain unresolved. The principle aim of our study was to determine the effect of storage time of whole blood on nitric oxide (NO) level expression in leukocytes. This is important because immune cells remain chemically dynamic even after they are removed from the circulation, and the impact of storage time must be known to optimally quantify the effect of a disease or condition on NO dynamics in circulating leukocytes. We measured NO levels using the fluorescent probe, diaminofluorescein (DAF-2DA), and flow cytometry in monocytes, neutrophils, and natural killer cells from healthy subjects immediately after blood draw (Time 0) and 30, 60, and 120 min following the blood draw. There was no significant difference among the 4 study time points in NO (DAF-2) levels, though there was wide intra-subject variability at all time points. Using LPS stimulation, we compared iNOS (the more traditional surrogate marker of NO dynamics) with NO (by DAF-2) in natural killer cells and monocytes and, we found no difference in the response patterns. In summary, we did find that within a 2-hour interval from blood draw to sample processing, there was a remarkably wide intra-subject variability in expression of intracellular NO (DAF-2) in leukocytes of healthy individuals at baseline and over time. The mechanism(s) for these differences are not known but could clearly confound efforts to detect changes in NO metabolism in white blood cells. We speculate that rapid pulsatility of NO could explain the wide variability seen.

Early stage of obesity potentiates nitric oxide reduction during the development of renal failure

Obesity is a serious health problem associated with the pathogenesis of various metabolic diseases. Nitric Oxide (NO) plays an important role in kidney function and altered NO levels have been associated with the pathogenesis of obesity. Therefore, we aimed to study whether an early stage of obesity contributes with progression of renal failure through further NO impairment. Male C57BL/6 mice were fed with a high-fat diet (HFD) or a normal diet (ND) during 2 weeks. All mice underwent either sham surgery (sham) or 5/6 nephrectomy (Np). One group of HFD Np mice was treated with antioxidants plus L-arginine. Kidney damage parameters were assessed and eNOS metabolism was evaluated. Mice on a HFD increased body weight, eNOS protein and mRNA expression, and radical oxygen species (ROS). Urine nitrites excretion, urine volume, and plasma BH4 were decreased. In HFD mice, 5/6 Np further increased BH2 and urine protein concentration, ROS levels, and eNOS mRNA expression. The decrease in BH4 plasma levels and urine nitrites excretion was accentuated. NO synthesis stimulation with the antioxidants + L-arginine treatment prevented all these changes. The early changes in NO metabolism are associated with an early stage of obesity. This effect on NO potentiates kidney damage development.

Clopidogrel results in favourable changes in nitric oxide metabolism in patients undergoing percutaneous coronary intervention

Clopidogrel results in favourable changes in nitric oxide metabolism in patients undergoing percutaneous coronary intervention -

Effects of angiotensin receptor blockers on endothelial nitric oxide release: the role of eNOS variants

• Angiotensin II receptor blockers improve endothelial cell-dependent vasodilation in patients with hypertension through suppression of angiotensin II type 1 receptors but may have additional and differential effects on endothelial nitric oxide synthase (eNOS) function. • The key finding from this study is that angiotensin II receptor blockers (ARBs) differentially enhanced nitric oxide (NO) release in a manner influenced by certain genetic variants of eNOS. This finding provides new insights into the effects of ARBs on endothelial cell-dependent vasodilation and eNOS function that are of high importance in vascular medicine and clinical pharmacology. AIM Angiotensin II receptor blockers (ARBs) improve endothelial cell (EC)-dependent vasodilation in patients with hypertension through suppression of angiotensin II type 1 receptors but may have additional and differential effects on endothelial nitric oxide (NO) synthase (eNOS) function. To investigate this question, we tested the effects of various ARBs on NO release in ECs from multiple donors, including those with eNOS genetic variants linked to higher cardiovascular risk. The effects of ARBs (losartan, olmesartan, telmisartan, valsartan), at 1 µm, on NO release were measured with nanosensors in human umbilical vein ECs obtained from 18 donors. NO release was stimulated with calcium ionophore (1 µm) and its maximal concentration was correlated with eNOS variants. The eNOS variants were determined by a single nucleotide polymorphism in the promoter region (T-786C) and in the exon 7 (G894T), linked to changes in NO metabolism. RESULTS All of the ARBs caused an increase in NO release as compared with untreated samples (P < 0.01, n= 4-5 in all eNOS variants). However, maximal NO production was differentially influenced by eNOS genotype. Olmesartan increased maximal NO release by 30%, which was significantly greater (P < 0.01, n= 4-5 in all eNOS variants) than increases observed with other ARBs. The ARBs differentially enhanced NO release in ECs in a manner influenced by eNOS single nucleotide polymorphisms. These findings provide new insights into the effects of ARBs on EC-dependent vasodilation and eNOS function.

Effect of soy isoflavone supplementation on nitric oxide metabolism and blood pressure in menopausal women

Isoflavones, having chemical structures similar to estrogens, are believed to stimulate nitric oxide production and thus lower blood pressure. The efficacy of soy isoflavone supplementation to stimulate nitric oxide production and lower blood pressure in menopausal women with high normal blood pressure remains unknown. The objective was to test the effect of soy isoflavone supplementation on nitric oxide production and blood pressure in menopausal women with high normal blood pressure. A randomized, double-blind, parallel, placebo-controlled 6-wk trial was conducted to assess the effects of daily supplementation with 80 mg soy hypocotyl isoflavones (in aglycone units) on nitric oxide metabolism and blood pressure in 24 menopausal women with 12 women per group. Changes in nitric oxide metabolism were assessed via a primed, constant-infusion protocol with [15N]arginine and [13C]- and [2H]citrulline. Changes in blood pressure and associated vascular hemodynamics were assessed via office and 24-h ambulatory blood pressure monitoring, forearm blood flow, and indexes of arterial compliance. When compared with placebo and after control for pretreatment values, soy isoflavone supplementation had no effect on arginine flux, citrulline flux, nitric oxide synthesis, blood pressure, forearm blood flow, or estimates of arterial stiffness. Daily supplementation with 80 mg soy hypocotyl isoflavones over a 6-wk period had no effect on nitric oxide metabolism or blood pressure and associated vascular hemodynamics in menopausal women with high normal blood pressure.

Changes in nitric oxide metabolism in co-morbidity of chronic obstructive pulmonary disease and chronic cerebral ischemia

Summary. The study was aimed at investigation of nitric oxide (NO) metabolism including total and separate concentrations of stable NO metabolites (nitrate anions and nitrite anions) and 3-nitrotirosine concentration in stable chronic obstructive pulmonary disease (COPD) and in COPD with concomitant chronic cerebral ischemia (CCI). The study involved 55 patients aged 51 to 67 years divided into 2 groups: 28 patients with COPD II–III stage (the 1st group) and 27 patients with COPD II–III stage and concomitant CCI II–III stage (the 2nd group). The control group included 25 healthy non-smoking volunteers. A significant increase in nitrate anion concentration and in the total nitrite and nitrate concentration was found in the blood of patients of both groups compared to the controls. A statistically significant inverse relationship was found between the blood nitrite anion concentration and FEV1 (r = –0.77; p &lt; 0.05) in COPD patients. These results demonstrate that changes in NO metabolism related to nitrous stress could emerge even in stable COPD. In co-existing COPD and CCI, changes in these biomarkers could be also related to functional particularities of nitrite reductase apart from the nitrous stress.

Relationship Between Endothelial Dependent Vasodilation and Size of Abdominal Aortic Aneurysms

Flow-mediated dilation of the brachial artery (FMDB) indirectly reflects the action of nitric oxide liberated by the endothelium. In patients with abdominal aortic aneurysms (AAA), the changes in nitric oxide metabolism in close association with inflammation, appear to play a leading role in the aetiopathology of this disease, although it is still not clear. The objective was to study the correlation and behavior of FMDB relative to the aneurysm diameter (AD). To evaluate the relationship between C-reactive protein (CRP) and the FMDB in these patients. To study the correlation and behavior of FMDB relative to the aneurysm diameter (AD). To evaluate the relationship between C-reactive protein (CRP) and the FMDB in these patients. The FMDB value and the CRP were determined in a total of 30 patients with an AAA > or =30 mm, confirmed by computed tomography. The cardiovascular and treatment history was recorded, together with the lipid and renal profile and leucocyte count. The median AD in the sample was 43 mm (25 percentile: 37 mm; 75 percentile: 60 mm). The primary variables of the study, FMDB and CRP, were the only ones that differed statistically when we stratified the sample according to AD quartiles (p < 0.001). There was a negative correlation between the FMDB and the AD (R = -0.78 [p < 0.001]) and a positive one for CRP (0.74 [p < 0.001]). The CRP/FMDB gave an R value of -0.56 (p = 0.001). Endothelium dependent vasodilation has a linear and negative correlation with the AD. The positive correlation between the FMDB and CRP supports the hypothesis that inflammation and endothelial dysfunction are processes associated with the physiopathology of AAA and vary with their growth.

A 12-Month Evaluation of Nitrite Oxide Metabolism Around Immediate and Conventionally Loaded Dental Implants

During bone remodeling, alignment of bone is affected by loading direction. Considerable amount of data also suggest nitric oxide (NO) to be involved in bone metabolism. This study was conducted to evaluate the potential changes in NO metabolism in relation to the loading style of dental implants by analyzing the nitrite content of peri-implant sulcus fluid. Dental implants were placed in 12 patients who had first molar loss bilaterally in the mandibular area. One site of the patient determined as immediately loaded and the other side were conventionally loaded. Clinical parameters were recorded and peri-implant sulcus fluid samples were obtained. Peri-implant sulcus fluid nitrite levels were spectrophoto-metrically determined. Clinical measurements and nitrite analysis were repeated at 1, 3, 6, 9, and 12 months. In both groups, there was a reduction between baseline and 12 months in all clinical parameters, except probing depth. Although, steady decrease was noticed in total nitrite levels during the whole experimental period in immediately loaded implants, more fluctuations were observed in conventionally loaded ones. The findings of this 12-month follow-up study support the association of NO in the bone metabolism around dental implants and further suggest the impact of different loading regimens on NO metabolism.

Plasma levels of nitric oxide and stroke outcome

Production of reactive oxygen species after cerebral blood flow disruption may enhance tissue damage through multiple molecular pathways. Changes in nitric oxide (NO) metabolism and oxidative stress status were investigated in 47 patients with ischemic stroke by measuring plasma nitric oxide (NO) and peroxynitrite (ONOO(-)) levels.A correlation was sought between these two parameters and i) baseline stroke severity based on the National Institute of Health stroke scale (NIHSS) and ii) neurological outcome in terms of NIHSS changes from entry (T(0)) to 30 days after symptom onset (T(1)). The control group consisted of 30 age- and sex-matched healthy subjects. Mean plasma levels of ONOO(-) (arbitrary fluorescence number +/- SD) were significantly higher in patients (7.70 +/- 1.71 vs 5.35 +/- 0.69, p < 0.001), whereas mean NO levels (nmol/mg protein) were significantly higher in controls (115.40 +/- 12.40 vs. 51.10 +/- 12.50, p < 0.001). Plasma ONOO(-) was significantly higher among patients with non-lacunar stroke (8.48 +/- 1.50 vs. 6.95 +/- 1.58 in those with lacunar stroke; p = 0.001), whereas NO levels were significantly higher among lacunar stroke patients (60.00 +/- 7.86, vs. 41.77 +/- 9.29 in patients with nonlacunar stroke; p < 0.001). Nitric oxide plasma levels were also associated with an unfavorable evolution in non-lacunar stroke, since a 10 unit increase in NO predicted a 1 point reduction in the NIHSS score at T1. Findings show that changes in NO metabolism may be considered as markers of brain injury in patients with ischemic stroke. Further work is needed to establish whether the amount of biochemical changes related to oxidative stress may influence outcome in these patients.

Azithromycin reduces Chlamydia pneumoniae‐induced attenuation of eNOS and cGMP production by endothelial cells

Intracellular infections with cytomegalovirus (CMV) or Chlamydia pneumoniae (Cp) may play a role in the aetiology of atherosclerosis. Nitric oxide (NO) is a key regulator of endothelial function. Under pathological conditions uncoupling of endothelial nitric oxide synthase (eNOS) leads to vessel damage as a result of production of oxygen radicals instead of NO. We hypothesized that infection-induced atherosclerosis is initiated by changes in NO metabolism and may be reversed by azithromycin treatment. Confluent human umbilical vein endothelial cells (HUVECs) were infected with Cp or CMV. After 48 h of infection, production of eNOS, cyclic guanosine monophosphate (cGMP) and reactive oxygen species (ROS) was measured. Detection of cGMP was used as a reporter assay for the bioavailability of NO. Subsequently, Cp- and CMV-infected HUVECs were coincubated with 0.016 mg L(-1) and 1 mg L(-1) azithromycin. Infection with Cp (MOI 1 and MOI 0.1) and CMV (MOI 1) caused a dose- and time-dependent reduction of eNOS production in the HUVECs: Cp MOI 1: 1141 +/- 74 pg mL(-1) (P < 0.01); Cp MOI 0.1: 3189 +/- 30 pg mL(-1) (P < 0.01); CMV: 3213 +/- 11 pg mL(-1) (P < 0.01) vs. 3868 +/- 83 pg mL(-1) for uninfected HUVECs. Chlamydia pneumoniae- but not CMV-infection also reduced cGMP-production (Cp: 0.195 +/- 0.030 pmol mL(-1) (P < 0.01); CMV: 0.371 +/- 27 pmol mL(-1) (P > 0.05) vs. 0.378 +/- 0.019 pmol mL(-1) for uninfected HUVECs). CMV-infection did not affect ROS production either, but Cp-infection reduced ROS-production by 21% (P > 0.05; Cp MOI 0.1) to 68% (P < 0.01; Cp MOI 1). Azithromycin treatment restored Cp-induced eNOS, cGMP and ROS production in a dose-dependent manner. Infection with Cp in endothelial cells in vitro attenuates eNOS, cGMP and ROS production in HUVECs and azithromycin reverses Cp-induced effects on eNOS, cGMP and ROS-production. The results from our in vitro research support the role of antibiotic therapy for infection-induced atherosclerosis by indicating that azithromycin does actually improve endothelial function.