The effects of endothelial nitric oxide synthase tagSNPs on nitrite levels and risk of hypertension and obesity in children and adolescents.

Obesity and the nitric oxide synthase 3 (NOS3) gene polymorphisms are associated with nitrite levels and hypertension. However, no study has tested the hypothesis that NOS3 tagSNPs rs3918226, rs3918188, rs743506 and rs7830 affect nitrite levels and are associated with hypertension in childhood obesity. We investigated the association of these NOS3 tagSNPs and the haplotypes formed by them with hypertension and with nitrite levels in children and adolescents with obesity and with obesity plus hypertension. We studied 355 subjects: 174 healthy (controls), 109 normotensive obese, and 72 obese children and adolescents with obesity plus hypertension. Genotypes were determined by Taqman allele discrimination assay and real-time PCR. We compared the distribution of NOS3 tagSNP genotypes, alleles and haplotypes in the three groups of subjects. Nitrite levels were determined by ozone-based chemiluminescence. Nitrite levels were affected by the rs3918226 polymorphism (P<0.05) but not by NOS3 haplotypes. There was no association between the tagSNPs studied and hypertension in children and adolescents. Our findings show that the NOS3 tagSNP rs3918226 is associated with NO production in children and adolescents, and suggest that this polymorphism may have an impact on cardiovascular health. Further studies are needed to better clarify the effects of this polymorphism on cardiovascular risk.

Simvastatin treatment increases nitrite levels in obese women: modulation by T(-786)C polymorphism of eNOS.

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.

Adiponectin associates positively with nitrite levels in children and adolescents.

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.

Haptoglobin polymorphism affects nitric oxide bioavailability in preeclampsia.

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.

PP070. Maternal flow-mediated dilation and nitrite concentration during third trimester of pregnancy and postpartum period.

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.

PP071. Evaluation of maternal-fetal doppler parameters and of whole blood nitrite throughout gestation.

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.

Aminoguanidine produces beneficial haemodynamic effects in a canine model of acute pulmonary thromboembolism.

AIM: Activating the nitric oxide (NO)-cyclic guanosine 3',5'-monophosphate (cGMP) pathway improves haemodynamics following acute pulmonary thromboembolism (APT). However, the role of NO synthase (NOS) isoforms in the responses to APT has not been determined. We examined the effects of selective and non-selective inducible NOS (iNOS) inhibition. METHODS: Haemodynamic evaluations were performed in non-embolized dogs treated with saline (control group; n = 4), L-NAME (NAME group; n = 3), or aminoguanidine (AG group; n = 3), and in dogs that received the same drugs and were embolized with 5 mL kg(-1) of clots made with autologous blood (Emb group, n = 9; NAME + Emb group, n = 4 and AG + Emb group, n = 7). The lung concentrations of nitrite/nitrate (NOx) and cGMP were determined by chemiluminescence and ELISA respectively. RESULTS: Acute pulmonary thromboembolism increased mean pulmonary arterial pressure (MPAP) and pulmonary vascular resistance index (PVRI) by 21.4 +/- 1.7 mmHg and by 843 +/- 34 dyn s cm(-5) m(-2), respectively, in Emb group. MPAP and PVRI increased to higher levels in the NAME + Emb group 15 min after APT and all dogs in this group died 15-30 min after APT. Conversely, lower MPAP and PVRI levels were found in the AG + Emb group 2 h after APT compared with the Emb group (both P < 0.05). Higher NOx concentrations were found in the Emb group compared with the other groups (all P < 0.05). Higher cGMP concentrations were found in the Emb and AG + Emb groups compared with the other groups (all P < 0.05). CONCLUSIONS: These results indicate that endogenous NO protects against APT-induced cardiovascular responses. Moreover, iNOS-derived NO possibly produces unfavourable effects, which are counteracted by aminoguanidine. However, non-NO-related mechanisms may also be involved.

Relationship between systemic nitric oxide metabolites and cyclic GMP in healthy male volunteers.

AIM: Nitric oxide (NO) is an endogenous mediator of many physiological processes, many of which are mediated by cyclic guanosine 3',5'-monophosphate (cGMP). Much effort has been made to validate clinical markers of NO production or bioavailability. While the measurement of plasma nitrate, nitrite, and cGMP concentrations have been suggested to reflect endogenous production of NO, there is no study showing whether there is correlation between these three markers. In the present study, we investigate whether there is correlation between the plasma concentrations of nitrate, nitrite, and cGMP in a relatively homogeneous group of 141 healthy subjects. METHODS: Venous blood samples were collected from healthy male subjects and plasma aliquots were then immediately removed and stored at -70 degrees C until analysed in duplicate for their nitrite and nitrate content using ozone-based chemiluminescence assays. Plasma cGMP levels were determined by using a commercial enzyme immunoassay. RESULTS: While we found no significant correlation between plasma nitrite and nitrate concentrations (P = 0.747), or between plasma nitrate and cGMP concentrations (P = 0.221), a significant positive correlation was found between plasma cGMP and nitrite concentrations (P = 0.017, r(s) = 0.270). CONCLUSIONS: The significant correlation we found between plasma nitrite and cGMP concentrations is consistent with the notion that nitrite or cGMP concentrations in plasma may be useful clinical markers of NO formation in healthy subjects.