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Introduction: The mechanical obstruction and pulmonary vasoconstriction are major determinants of the sudden right ventricular (RV) afterload increases observed during acute pulmonary thromboembolism (APT). Vasodilators and antioxidants agents have been shown to mitigate pulmonary hypertension. We examined whether sodium nitrite and the antioxidant tempol combination could be advantageous in an APT sheep model., Methods: APT was induced in anesthetized sheep by autologous blood clots (250 mg/kg) into the right atrium. Thirty minutes after APT induction, the animals received a continuous infusion of tempol (1.0 mg/kg/min), increasing sodium nitrite infusion (5, 15, and 50 μmol/kg), or a simultaneous combination of both drugs. Saline was used as a control treatment. Hemodynamic measurements were carried out every 15 min. Also, whole blood nitrite and serum 8-isoprostanes levels were measured., Results: APT induced sustained pulmonary hypertension, increased dp/dt max , and rate pressure product (RPP). Nitrite or tempol treatments attenuated these increases (P < 0.05). When both drugs were combined, we found a robust reduction in the RV RPP compared with the treatments alone (P < 0.05). The sole nitrite infusion increased blood nitrite concentrations by 35 ± 6 μM (P < 0.05), whereas the nitrite and tempol combination produced higher blood nitrite concentrations by approximately 54 ± 7 μM. Tempol or nitrite infusions, both alone or combined, blunted the increases in 8-isoprostane concentrations observed after APT., Conclusions: Nitrite and tempol combination protects against APT-induced RV wall stress. The association of both drugs may offer an advantage to treat RV failure during severe APT., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The purpose of this study was to evaluate the relationship between 3 eNOS gene polymorphisms and training status (TS) in affecting plasma nitrite concentration (NO2) in normotensive adults over 50 years old. Resting blood pressure (BP) was measured in all participants (n = 101). Plasma was taken to analyze: lipid profile, nitrite concentration (NO2) and lipid peroxide levels (T-BARS). Also, genomic DNA was extracted from plasma for genotyping NOS3 polymorphisms (-786T>C; 894G>T; and VNTR in intron 4). TS was determined by one-mile walk test and Functional Fitness Test Battery from AAHPERD (TS1-regular TS; TS2-good TS; and TS3-very good TS). BP was not influenced by TS, but NO2 was 15% higher in TS3 (123 ± 27 nM) compared to TS-2 (106 ± 22 nM). No differences were found in plasma NO2 in the haplotype analyses. However, the presence of the C allele (T-786C) and ASP allele (Glu298Asp) was found to enhance the correlation between TS and NO2 levels (r = 0.492 in C/4b/ASP haplotype and r = 0.855 in C/4a/ASP haplotype). This study thus identifies NOS3 polymorphism-dependent sensitivity to the effects of physical training on plasma NO2. Maintenance of good levels of training status, in carriers of C allele for T-786C polymorphism, combined with ASP allele for Glu298Asp polymorphism, may result in an increase in the NO2 plasma concentrations, which may reflect improved NO bioavailability in older adult normotensive individuals.

Higher homocysteine (Hcy) levels are associated with cardiovascular risk. The aim of the present study was to evaluate the effect of simvastatin treatment on circulating Hcy levels in obese women without hypertension, diabetes or dyslipidaemia; and to determine whether the 677C>T polymorphism located in methylenetetrahydrofolate reductase (NAD(P)H) (MTHFR) gene modulates the effects of this treatment on Hcy and nitrite (as a biomarker of nitric oxide (NO) bioavailability). Twenty-five obese women (body mass index ≥ 30 kg/m(2) ) who had received 20 mg/day simvastatin for 6 weeks were enrolled in the study. Venous blood samples were collected to measure plasma biomarkers and gene polymorphisms. Simvastatin treatment significantly reduced total cholesterol, low-density lipoprotein-cholesterol, thiobarbituric acid-reactive substances, high-sensitivity C-reactive protein and Hcy, whereas nitrite levels were increased. The reduction in Hcy levels in carriers of the T allele was -20.3% compared with -9.4% in patients with the CC genotype. Importantly, before treatment, nitrite levels were significantly higher in patients with the CC genotype compared with T allele carriers, whereas after treatment these levels were similar between groups. Our findings demonstrate that obese women without comorbidities and carrying the T variant of the 677C>T polymorphism of MTHFR exhibit benefits with simvastatin treatment, mainly in terms of increased NO levels., (© 2014 Wiley Publishing Asia Pty Ltd.)

Purpose: Failure to control blood pressure (BP) despite the use of three or more drugs characterizes resistant hypertension (RHTN). Impaired endothelial function is associated with this condition and phosphodiesterase-5 inhibitors (PDE5i)-inhibiting cGMP breakdown-reduce BP in RHTN patients. We hypothesized that acute administration of PDE5i could ameliorate hemodynamic, endothelial parameters and left ventricular diastolic function (LVDF) in RHTN patients. Also, an exploratory analysis was performed to assess the influence of the T-786C endothelial NO synthase polymorphism on those responses., Methods: Subjects (n = 26) underwent a 6-month clinical screening for RHTN diagnosis. Increasing doses of oral sildenafil were given at 30 min intervals (37.5, 50 and 100 mg) while continuous non-invasive hemodynamic measures were assessed. LVDF, flow mediated dilation (FMD), nitrite and cGMP levels were also determined., Results: Mean arterial pressure and total peripheral resistance decreased in all patients (84.17 ± 21.04 to 75 ± 17.21 mmHg; 1149 ± 459.7 to 1037 ± 340 dyn.s/cm(-5), respectively). Likewise, sildenafil improved diastolic dysfunction parameters (Left atrial volume: 25 ± 5.8 to 20 ± 4.4; IVRT: 104 ± 19.33 to 88 ± 15.22; E/e' septal: 9.7 ± 3.8 to 7.9 ± 2.9; E/e' lateral: 7.7 ± 3.4 to 6.4 ± 3.2). No statistical changes were found in FMD, nitrite and cGMP with PDE5i., Conclusion: Our data suggest PDE5i acutely improves diastolic function and hemodynamic profile in RHTN subjects, despite unchanging endothelial dysfunction.

Introduction: Acute pulmonary thromboembolism (APT) is a critical condition associated with acute pulmonary hypertension. Recent studies suggest that oxidative stress and hemolysis contribute to APT-induced pulmonary hypertension, possibly as a result of increased nitric oxide (NO) consumption. We hypothesized that the antioxidant tempol could attenuate APT-induced hemolysis, and therefore attenuate APT-induced increases in plasma NO consumption., Materials and Methods: APT was induced in anesthetized sheep with autologous blood clots. The hemodynamic effects of tempol infused at 1.0mg/kg/min 30 min after APT were determined. Hemodynamic measurements were carried out every 15 min. To assess oxidative stress, serum 8-isoprostanes levels were measured by ELISA. Plasma cell-free hemoglobin concentrations and NO consumption by plasma samples were determined. An in vitro oxidative AAPH-induced hemolysis assay was used to further validate the in vivo effects of tempol., Results: APT caused pulmonary hypertension, and increased pulmonary vascular resistance in proportion with the increases in 8-isoprostanes, plasma cell-free hemoglobin concentrations, and NO consumption by plasma (all P<0.05). Tempol attenuated the hemodynamic alterations by approximately 15-20% and blunted APT-induced increases in 8-isoprostanes, in cell-free hemoglobin concentrations, and the increases in NO consumption by plasma (P<0.05). Tempol dose-dependently attenuated AAPH-induced in vitro hemolysis (P<0.05)., Conclusions: Our findings are consistent with the idea that antioxidant properties of tempol decrease APT-induced hemolysis and nitric oxide consumption, thus attenuating APT-induced pulmonary hypertension., (© 2013.)

Upregulation of inducible nitric oxide synthase (iNOS) has been reported in both experimental and clinical hypertension. However, although pro-inflammatory cytokines that up-regulate iNOS contribute to pre-eclampsia, no previous study has tested the hypothesis that a selective iNOS inhibitor (1400 W) could exert antihypertensive effects associated with decreased iNOS expression and nitrosative stress in pre-eclampsia. This study examined the effects of 1400 W in the reduced uteroplacental perfusion pressure (RUPP) placental ischaemia animal model and in normal pregnant rats. Sham-operated and RUPP rats were treated with daily vehicle or 1 mg/kg/day N-[3-(Aminomethyl) benzyl] acetamidine (1400 W) subcutaneously for 5 days. Plasma 8-isoprostane levels, aortic reactive oxygen species (ROS) levels and nicotinamide adenine dinucleotide phosphate (NADPH)-dependent ROS production were evaluated by ELISA, dihydroethidium fluorescence microscopy and lucigenin chemiluminescence respectively. Inducible nitric oxide synthase expression was assessed by western blotting analysis and aortic nitrotyrosine was evaluated by immunohistochemistry. Mean arterial blood pressure increased by ~30 mmHg in RUPP rats, and 1400 W attenuated this increase by ~50% (P < 0.05). While RUPP increased plasma 8-isoprostane levels, aortic ROS levels, and NADPH-dependent ROS production (P < 0.05), treatment with 1400 W blunted these alterations (P < 0.05). Moreover, while RUPP increased iNOS expression and aortic nitrotyrosine levels (P < 0.05), treatment with 1400 W blunted these alterations (P < 0.05). These results clearly implicate iNOS in the hypertension associated with RUPP. Our findings may suggest that iNOS inhibitors could be clinically useful in the therapy of pre-eclampsia, especially in particular groups of patients genetically more prone to express higher levels of iNOS. This issue deserves further confirmation., (© 2013 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Objective and Subjects: Evidence indicates an impairment of nitric oxide (NO) in obesity. Statins present pleiotropic effects independently of cholesterol-lowering, including increasing of eNOS expression and antioxidant effects. We evaluated the effects of simvastatin treatment at 45 days on circulating nitrite (NO marker) and TBARS-MDA levels in obese women without comorbidities (hypertension, diabetes and dyslipidemia). Moreover, we verified whether obese women carrying the C variant of T(-786)C polymorphism located in eNOS may have increased levels of nitrite after treatment compared to TT genotype., Results: After simvastatin treatment, while the plasma nitrite levels increased 42% (P=0.0008), the TBARS-MDA levels reduced 58% (P=0.0069). We observed increased levels of nitrite in both groups of genotypes (TT vs. TC+CC); however, rise in C-allele carriers was 60% comparing with 44% in TT., Conclusion: Our results demonstrated a restoration of nitrite levels in obese women treated with simvastatin, which is modulated by T(-786)C polymorphism., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Erectile dysfunction (ED) is a multifactorial disease associated with vascular dysfunction, low nitric oxide (NO) bioavailability, and oxidative stress. However, it is not known whether low NO bioavailability and oxidative stress affect the responsiveness of ED patients to sildenafil. We tested this hypothesis by studying 28 healthy subjects (control group), 26 patients with ED without comorbidities (ED group), and 18 patients with ED and diabetes mellitus (ED/DM group). The International Index for Erectile Function (IIEF) questionnaire was used to assess the erectile function of all participants, and their responsiveness to sildenafil was assessed as the percentage of change in the five-item version of IIEF score before and after sildenafil treatment. Levels of whole blood nitrite, antioxidants markers (ferric reducing ability of plasma (FRAP) and reduced glutathione), and oxidative stress markers (thiobarbituric acid reactive substance and protein carbonyl) were determined. We found a negative correlation between whole blood nitrite levels and the responses to sildenafil in both ED groups (P<0.05). FRAP correlated negatively with the responses to sildenafil in the ED/DM group (P<0.05). No other significant associations were found. Our findings show evidence that low NO bioavailability is associated with better responses to sildenafil in patients with ED (with or without DM).

Objectives: We examined whether experimental lung embolization with autologous blood clots or with the infusion of microspheres increase cell-free hemoglobin levels and nitric oxide consumption by plasma samples from anesthetized lambs. These parameters were also measured in patients with acute pulmonary thromboembolism at baseline conditions and after thrombolysis, and in healthy controls., Design: Controlled animal and clinical studies., Setting: University research laboratory and university hospital., Subjects: Sheep and humans., Interventions: Anesthetized lambs were embolized with intravenous injections of autologous blood clots or repeated injections of 300 μm microspheres. Control animals received saline. Blood samples were drawn from patients with acute pulmonary thromboembolism at baseline conditions and after thrombolytic therapy with streptokinase or alteplase., Measurements and Main Results: Hemodynamic measurements were performed and plasma cell-free hemoglobin concentrations were measured. A nitric oxide consumption assay was used to measure nitric oxide consumption by plasma samples. Embolization with blood clots or microspheres increased mean pulmonary artery pressure from ~15 to ~40 mm Hg in lambs. Both plasma hemoglobin concentrations and nitric oxide consumption increased in proportion to the hemodynamic alterations and correlated significantly. Patients with acute pulmonary thromboembolism had higher plasma hemoglobin concentrations and nitric oxide consumption than healthy controls. Thrombolysis with streptokinase or alteplase further increased both parameters, which peaked 1-3 days after thrombolysis., Conclusions: Our results show consistent evidence indicating a new mechanism involving increased hemoglobin decompartmentalization and augmented nitric oxide consumption, possibly contributing to the hemodynamic derangement of acute pulmonary thromboembolism.

Studies showed elevated cell-free hemoglobin (Hb) in preeclampsia (PE), and Hb reacts with nitric oxide (NO), decreasing its bioavailability. Haptoglobin (Hp) is a polymorphic protein (Hp1-1, Hp2-1 and Hp2-2) that binds Hb to form a complex that is removed from circulation, thus preventing Hb-driven oxidative stress and NO scavenging. Hp protein products differ in biochemical and biophysical properties, which reflects on the Hb-Hp complex clearance rate. We hypothesized that Hp phenotypes modulate NO bioavailability by influencing NO consumption in PE. We studied 92 PE subjects and 105 normal pregnant women (NP). Hp genotypes were determined using real-time PCR. To assess NO bioavailability, we measured plasma nitrite using an ozone-based chemiluminescence assay. Plasma Hb and Hp were assessed with commercial immunoassays. A NO consumption assay was used to measure NO consumption. We found no differences in Hp genotype frequencies between PE and NP groups. Hp genotypes had no effects on plasma heme levels, NO consumption and plasma nitrite in NP. However, in PE, Hp2-1 and Hp2-2 were associated with higher plasma heme levels (48 and 55% higher, respectively; P<0.05), increased NO consumption (42 and 44% more, respectively; P<0.05) and lower plasma nitrite (39% less for Hp2-2; P<0.05) compared with Hp1-1. These findings indicate that although Hp genotype does not affect the risk of PE, Hp1-1 genotype may exert a protective role in PE by reducing NO scavenging, whereas Hp2-1 and Hp2-2 further may aggravate PE by reducing NO bioavailability.

Objective: To compare the circulating levels of adiponectin and nitric oxide (NO) bioavailability in eutrophic, eutrophic hypertensive, obese, and obese hypertensive children and adolescents, and to assess whether adiponectin is associated with increased NO bioavailability in these children and adolescents., Methods: We studied 129 eutrophic, 8 eutrophic hypertensive, 91 obese, and 44 obese hypertensive children and adolescents in this cross-sectional study. Adiponectin concentrations were measured in plasma samples by enzyme-linked immunosorbent assay. To assess NO bioavailability, nitrite concentrations were measured in whole-blood samples by chemiluminescence. Multiple linear regression analysis was carried out to assess the effects of adiponectin on NO bioavailability., Results: We found no significant differences in nitrite levels among groups (P>0.05). The obese hypertensive group had the lowest adiponectin levels among groups (P<0.05). Additionally, obese subjects had lower adiponectin levels than eutrophic individuals (P<0.05). A multiple linear regression analysis showed that NO bioavailability was positively associated with adiponectin concentrations (P<0.05)., Conclusions: Our findings suggest that adiponectin increases NO bioavailability in children and adolescents. Further studies are needed to assess the cardiovascular protective role for this adipokine in childhood obesity.

Introduction: The Doppler method is extensively applied today for the evaluation of pregnancies with involvement of the uteroplacental blood flow. Although increased nitric oxide (NO) formation plays an important role in regulation of systemic vascular resistance during pregnancy, growing evidence indicates that reduced NO formation is associated with hypertensive disorders of pregnancy, especially preeclampsia., Objectives: The studies were to assess the maternal and fetal Doppler parameters and to determine the whole blood nitrite levels during pregnancy., Methods: Thirty-three healthy pregnant women were evaluated during the first (11-14 weeks), second (20-24 weeks) and third trimesters (34-36 weeks) of pregnancy. The maternal (uterine arteries) and fetal (cerebral and umbilical arteries) vessels were evaluated by Doppler velocimetry. venous blood was collected(15mL) for the determination of plasma nitrite by chemiluminescence., Results: Regarding the Doppler parameters of the uterine arteries the mean pulsatility index was 1.73, 1.06 and 0.73 in the first, second and third trimesters of pregnancy, respectively. Fetal Doppler showed a mean resistance index of 0.82 and 0.81 for the middle cerebral artery, 0.73 and 0.60 for the umbilical artery in the second and third trimesters, respectively. The mean plasma nitrite concentration was 189.10, 178.28 and 199.57 nmol/ml in the first, second and third trimesters of pregnancy, respectively., Conclusion: The study demonstrated that a fall in flow resistance occurs in the uteroplacental vessels without changes in plasma nitrite concentrations during pregnancy., (Copyright © 2012. Published by Elsevier B.V.)

Introduction: The vascular endothelium is thought to be responsible for cardiovascular adaptations in gestation, such as the decrease in peripheral vascular resistance and the decrease in arterial pressure. There is an increase of nitric oxide (NO) serum levels in normal gestation due to an increment in the activity of the enzyme endothelial nitric oxide synthase (eNOS)., Objectives: To compare maternal flow-mediated dilation of the brachial artery (FMD) and the nitrite concentration between the third trimester of pregnancy and postpartum period. Additionally we want to evaluate whether FMD correlates with nitrite concentration., Methods: Eligibility criteria were healthy pregnant women with single fetus, gestational age greater than 28 weeks, nonsmokers, and without personal or family history of vascular disease. Each pregnant woman was examined in the third trimester of pregnancy (3(rd)T) and between 8 and 12 weeks postpartum (PP) to evaluate FMD and nitrite concentrations in the whole blood. We excluded women who were not examined in both periods. We compared the values between the two periods using paired t tests. The correlation between FMD and nitrite concentration was examined by Pearson correlation coefficient. Significance level was set at p<0.05., Results: 42 pregnant women were invited for the study. 35 healthy women were elected and 7 of them were excluded for not attending the postpartum evaluation. We found a trend of decreased FMD in the PP period (10.39±5.57 % vs. 8.42±4.21 %, p=0.11; 3(rd)T vs. PP respectively). No significant change was observed in the nitrite concentration (257.41±122.95nmol/L vs.237.16±90.01nmol/L, p=0.28). We did not observe significant correlation between FMD and nitrite during 3(rd)T (r=-0.13, p=0.50) or PP (r=0.14, p=0.48)., Conclusion: Although our sample size did not permit sufficient precision, FMD seems to decrease between the third trimester and postpartum period. Nitrite concentration did not change between the third trimester of pregnancy and the postpartum period, and it was not correlated to FMD. Studies evaluating larger samples are necessary to confirm these findings., (Copyright © 2012. Published by Elsevier B.V.)

Objectives: We compared nitrite, B-type natriuretic peptide (BNP), and cGMP levels in preeclamptic with those found in healthy pregnant., Methods: We studied 21 healthy pregnant and 27 preeclamptic. Plasma cGMP and BNP levels were determined by ELISA. Nitrite levels were determined by chemiluminescence., Results: Higher cGMP and BNP, and lower nitrite levels were found in preeclamptic versus healthy pregnant., Conclusions: Altered cGMP levels reflect increased BNP levels and not impaired nitric oxide activity in preeclampsia., (Copyright © 2011 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

Nitric oxide (NO) is a major regulator of the cardiovascular system. However, the effects of endothelial nitric oxide synthase (eNOS) gene polymorphisms or haplotypes on the circulating concentrations of nitrite (a sensitive marker of NO formation) and cGMP are unknown. Here we examined the effects of eNOS polymorphisms in the promoter region (T-786C), in exon 7 (Glu298Asp), and in intron 4 (4b/4a) and eNOS haplotypes on the plasma levels of nitrite and cGMP. We hypothesized that eNOS haplotypes could have a major impact on NO formation. We genotyped 142 healthy subjects by PCR-RFLP. To assess NO formation, the plasma concentrations of nitrite and cGMP were determined using an ozone-based chemiluminescence assay and an enzyme immunoassay. Haplotypes were inferred using the PHASE 2.1 program. No significant differences were found in age, body mass index, systolic and diastolic arterial blood pressure, heart rate, total cholesterol, triglycerides, cGMP, or nitrite among the genotype groups for the three polymorphisms studied here (all p>0.05). Interestingly, the C-4b-Glu haplotype was associated with lower plasma nitrite concentrations than those found in the other haplotype groups (p<0.05), but not with different cGMP levels (p>0.05). These findings suggest that eNOS gene variants combined within a specific haplotype modulate NO formation, although individual eNOS polymorphisms probably do not have major effects.

Statins inhibit cholesterol synthesis and produce pleiotropic, cholesterol-independent effects including endothelial NO synthase (eNOS) stimulation and increased expression. However, a functional polymorphism in the promoter of the eNOS gene (T-786C) reduces its activity and could modulate the response to statins. Here, we examined whether this polymorphism modulates the effects of atorvastatin on the plasma levels of markers of NO formation and oxidative stress. We genotyped 200 healthy subjects for this polymorphism, and 15 subjects with the TT genotype and 15 with the CC genotype were selected to receive placebo or atorvastatin 10 mg/day po for 14 days. To assess NO bioavailability, the plasma concentrations of nitrate, nitrite, and cGMP and the whole blood nitrite concentrations were determined after placebo or atorvastatin using an ozone-based chemiluminescence assay and an enzyme immunoassay. Thiobarbituric acid-reactive species (TBA-RS) were measured in the plasma to assess oxidative stress. Atorvastatin decreased cholesterol concentrations independent of genotype. Whereas atorvastatin produced no significant changes in plasma nitrite, nitrate, or cGMP concentrations in both genotype groups, atorvastatin increased whole blood nitrite concentrations and decreased plasma TBA-RS concentrations in the CC (but not in the TT) genotype group. These findings suggest that the T-786C polymorphism modulates the effects of atorvastatin on NO bioavailability and oxidative stress.