Urinary Nitric Oxide Metabolites Research Articles

Clinical and Prognostic Value of Calcium and Phosphorus Levels as Possible Markers of Endothelial Dysfunction in Preterm Infants

The system maintaining a sufficient level of calcium and phosphorus is formed only by the end of the first month of life, and in pretermure infants even later. These elements play a crucial role in many physiological processes. They also affect vascular function, endothelium, and endothelial nitric oxide (NO) synthase. Therefore, the aim of our study was to analyze the relationship of serum phosphorus and calcium levels with markers of endothelial dysfunction and features of the early neonatal period. The study included 37 preterm infants that received standard clinical, laboratory, and instrumental examination. As markers of endothelial dysfunction, the 4a/4b polymorphism of the eNOS gene was investigated and quantitative measurement of NO metabolites in urine was performed. Phosphorus and calcium levels in 37 preterm infants on the first day were 4.06 ± 1.88 mg/dL and 1.89 ± 2.08 mg/dL, and on the six day 4.89 ± 2.02 mg/dL and 1.86 ± 1.80 mg/dL, respectively. Combined hypocalcemia and hypophosphatemia were found in 27 (73.0%) of 37 examined infants. The presence of severe condition in early neonatal period was positively correlated with calcium levels on the first and sixth days after birth. There were determined positive correlation between calcium and phosphorus levels on the first day after birth and severity of clinical symptoms in neonatal period and birth gestational age. Serum calcium level in preterm birth infants was positively correlated with respiratory failure on the sixth day. Correlation between calcium and phosphorus serum levels was observed for severe and stable infants on the first day, but on the six day only in preterm birth infants with stable condition. We have not found an association between phosphorus and calcium levels and markers of endothelial dysfunction in preterm birth infants. Serum phosphorus and calcium levels and studied markers of endothelial dysfunction have been found to be independent markers of the risk of developing a complicated early neonatal period in preterm infants.

Urine Nitric Oxide Is Lower in Parents of Autistic Children.

Parents raising children with autism spectrum disorder (ASD) usually carry on their daily life under tremendous stress, but limited empirical research has been devoted to this population. It is known that parents' health status directly impacts therapeutic outcome of ASD children. As an important regulator in cardiovascular, nervous and immune systems, nitric oxide (NO) levels haven't been reported in parents of ASD children yet. In this study, we measured urine nitrite and nitrate from 43 ASD parents (ASD-P), and 43 healthy adults in the same range of age (Control) who didn't have any ASD descendants. Comparison between the ASD-P and Control groups showed that , , and / were all significantly lower in the ASD-P group. Analysis on the interaction effect of sex and group indicated that urine of mothers in ASD-P was lower than that in females of the Control group, but no significant difference was observed between males in both groups. It is for the first time that urine nitric oxide metabolites (nitrite, nitrate) levels were precisely reported to differentiate parents of autistic children from other adults without ASD descendants. This phenomenon suggests that parents (especially mothers) of autistic children might have experienced more mental and physical stressors, which led to decreased NO levels during metabolism. Further investigations are necessary to uncover the etiology of low urine NO among parents of autistic children.

Failure to vasodilate in response to salt loading blunts renal blood flow and causes salt-sensitive hypertension.

Salt-sensitive (SS) hypertension is accompanied by impaired vasodilation in the systemic and renal circulation. However, the causal relationship between vascular dysfunction and salt-induced hypertension remains controversial. We sought to determine whether primary vascular dysfunction, characterized by a failure to vasodilate during salt loading, plays a causal role in the pathogenesis of SS hypertension. Mice selectively expressing a peroxisome proliferator-activated receptor γ dominant-negative mutation in vascular smooth muscle (S-P467L) exhibited progressive SS hypertension during a 4week high salt diet (HSD). This was associated with severely impaired vasodilation in systemic and renal vessels. Salt-induced impairment of vasodilation occurred as early as 3 days after HSD, which preceded the onset of SS hypertension. Notably, the overt salt-induced hypertension in S-P467L mice was not driven by higher cardiac output, implying elevations in peripheral vascular resistance. In keeping with this, HSD-fed S-P467L mice exhibited decreased smooth muscle responsiveness to nitric oxide (NO) in systemic vessels. HSD-fed S-P467L mice also exhibited elevated albuminuria and a blunted increase in urinary NO metabolites which was associated with blunted renal blood flow and increased sodium retention mediated by a lack of HSD-induced suppression of NKCC2. Blocking NKCC2 function prevented the salt-induced increase in blood pressure in S-P467L mice. We conclude that failure to vasodilate in response to salt loading causes SS hypertension by restricting renal perfusion and reducing renal NO through a mechanism involving NKCC2 in a mouse model of vascular peroxisome proliferator-activated receptor γ impairment.

Morinda citrifolia (Noni) fruit juice promotes vascular endothelium function in hypertension via glucagon-like peptide-1 receptor-CaMKKβ-AMPK-eNOS pathway.

Morinda citrifolia (Noni) is extensively used in herbal remedies to prevent and treat various diseases, including hypertension. The purpose of this study was to investigate the vascular effects of noni fruit juice and characterize the upstream signaling pathways. We measured the systolic blood pressure, diastolic blood pressure, 24-hr urinary nitric oxide (NO) metabolite excretion, bodyweight (BW), and urine examination in SHR.Cg-Leprcp/NDmcr (SHR/cp) rats after 6 weeks noni juice (15 ml/kg) treatment. Noni juice significantly decreased blood pressure and 24-hr urinary NO metabolite without change of BW or urine volume. Furthermore, the noni juice extract (NJE) promoted endothelial vasorelaxation in rat aorta rings and NO product through an increase in phosphorylation of endothelial nitric oxide synthase (eNOS) in human umbilical vein endothelial cells (HUVECs). NJE might act on a glucagon like peptide-1 receptor (GLP-1R) via Ca2+ /calmodulin-dependent protein kinase kinase β (CaMKKβ)-AMPK signaling with pretreatment of their inhibitors or antagonist in HUVECs. Deacetylasperulosidic acid (DAA) was an active compound in noni juice to improve NO release through same pathway in HUVECs. These results suggested that noni is a novel dietary plant that probably regulates GLP-1R-induced CaMKKβ-AMPK-eNOS pathway to improve endothelium-dependent relaxation, thus reduce the blood pressure probably via one of its responsible ingredient DAA.

Tadalafil improves bladder dysfunction and object recognition in rats with pelvic venous congestion.

To examine the effects of tadalafil on bladder function and object recognition ability in rats with alterations in urinary frequency and locomotor activity as a result of pelvic venous congestion. A total of 48 female rats were divided into three groups (sham, pelvic venous congestion and pelvic venous congestion/tadalafil groups). In the pelvic venous congestion and pelvic venous congestion/tadalafil groups, the bilateral common iliac veins and uterine veins were ligated under anesthesia. Rats in the pelvic venous congestion/tadalafil group received a diet containing tadalafil, and the other rats were fed a normal diet. After 4weeks, rats underwent analysis of voiding behavior, locomotor activity, a novel object recognition test, continuous cystometry, measurement of plasma monoamines, and measurement of plasma and urinary nitric oxide metabolites. Expression of nitric oxide synthase messenger ribonucleic acid in the bladder wall was also assessed, along with histological examination of the bladder. Rats with pelvic venous congestion showed a higher urinary frequency, lower locomotor activity, and lower plasma and urinary nitric oxide levels than sham rats. The bladder wall endothelial nitric oxide synthase messenger ribonucleic acid level was low and object recognition was impaired. Pelvic venous congestion/tadalafil rats showed improvement in locomotor activity, bladder function and object recognition compared with pelvic venous congestion rats, as well as elevation of plasma and urinary nitric oxide, plasma monoamines, and bladder neuronal nitric oxide synthase messenger ribonucleic acid expression. Bladder wall vascularity was greater in pelvic venous congestion/tadalafil rats compared with sham rats. In rats with pelvic venous congestion, tadalafil might improve bladder function and the general condition by increasing blood flow to the bladder and brain, and by increasing dopamine levels.

Influence of dietary nitrate food forms on nitrate metabolism and blood pressure in healthy normotensive adults

Inorganic nitrate (NO3−) supplementation has been shown to improve cardiovascular health indices in healthy adults. The purpose of this study was to investigate how the vehicle of NO3− administration can influence NO3− metabolism and the subsequent blood pressure response. Ten healthy males consumed an acute equimolar dose of NO3− (∼5.76 mmol) in the form of a concentrated beetroot juice drink (BR; 55 mL), a non-concentrated beetroot juice drink (BL; 456 mL) and a solid beetroot flapjack (BF; 60 g). A drink containing soluble beetroot crystals (BC; ∼1.40 mmol NO3−) and a control drink (CON; 70 mL deionised water) were also ingested. BP and plasma, salivary and urinary [NO3−] and [NO2−] were determined before and up to 24 h after ingestion. All NO3−-rich vehicles elevated plasma, salivary and urinary nitric oxide metabolites compared with baseline and CON (P<0.05). The peak increases in plasma [NO2−] were greater in BF (371 ± 136 nM) and BR (369 ± 167 nM) compared to BL (283 ± 93 nM; all P<0.05) and BC (232 ± 51 nM). BR, but not BF, BL and BC, reduced systolic (∼5 mmHg) and mean arterial pressure (∼3–4 mmHg; P<0.05), whereas BF reduced diastolic BP (∼4 mmHg; P < 0.05). Although plasma [NO2−] was elevated in all conditions, the consumption of a small, concentrated NO3−-rich fluid (BR) was the most effective means of reducing BP. These findings have implications for the use of dietary NO3−supplements when the main objective is to maintain or improve indices of cardiovascular health.

Smooth Muscle PPARγ Mutation Causes Salt‐sensitive Hypertension

Systemic loss of peroxisome proliferator‐activated receptor‐γ (PPARγ) function causes hypertension, whereas its activation lowers blood pressure largely via PPARγ activity in the vasculature. We have recently shown that loss of PPARγ in smooth muscle cells increases mesenteric myogenic tone, promotes vascular remodeling, and predisposes to deoxycorticosterone acetate (DOCA)‐salt‐induced hypertension in mice. However, it is unclear whether these effects were attributable to altered salt sensitivity. In this study, we examined the role of smooth muscle PPARγ in regulating renal function and salt‐sensitivity. Transgenic mice expressing dominant negative PPARγ in smooth muscle cells (S‐P467L) and non‐transgenic littermates (NT) were fed normal chow diet (containing 0.3% salt) or a 4% salt diet for a total of 8 weeks and metabolic cage studies were performed at baseline, 3 weeks, 6 weeks, and at the end of the protocol. S‐P467L and NT mice had similar food intake, feces weight, and weight gain on high salt diet throughout the study. While 24‐hour water intake tripled in both strains (6.3 ± 0.21 mL S‐P467L vs. 5.9 ± 0.40 mL NT, n=7) when they were switched to high salt diet from normal chow, S‐P467L mice excreted 32% less urine in the third week (24‐hour urine 1.96 ± 0.13 mL S‐ P467L vs. 2.90 ± 0.17 mL NT, two‐way ANOVA p&lt;0.01). In order to assess renal function, mice were subjected to a bolus i.p. injection of normal saline equal to 10% of their body weight and the urine volume excreted in the subsequent 4 hours were plotted as a percentage of saline injected. We observed a marked decline in their capacity to excrete this volume challenge in S‐ P467L mice (28.2 ± 2.1% S‐P467L vs. 39.9 ± 5.1% NT, p&lt;0.05) in the third week of high salt diet. Because nitric oxide (NO) is known to regulate sodium and water balance, we determined daily renal NO production as indicated by 24‐hour urinary nitrate/nitrite. Three‐week dietary salt induced a 4‐fold increase in urinary NO metabolites in NT mice, but this was blunted in S‐P467L mice (2.82 ± 0.19 μmoL/24 hours S‐P467L vs. 4.42 ± 0.27 μmoL/24 hours NT, p&lt;0.01). In parallel with these renal phenotypes, S‐P467L mice developed marked elevation of blood pressure that peaked in the third week of high salt feeding (telemetry systolic pressure 136.4 ± 3.4 mmHg S‐P467L vs. 124.2 ± 2.4 mmHg NT, p&lt;0.01) and remained elevated throughout the study. At the end of 8 weeks, vascular function was assessed in freshly isolated carotid and basilar arteries, and flow cytometry analysis was performed in single cell suspensions of aorta and kidney tissues. S‐P467L mice, but not NT littermates, exhibited severe impairment of acetylcholine‐ and sodium nitroprusside‐induced vascular relaxation in the carotid and basilar arteries in response to high salt. CD45+ total leukocytes, CD3+ T cells, CD4+/CD8+ T cell subsets, and F4/80+ monocytes/macrophages were not altered in the aortas but were paradoxically reduced in the kidneys of S‐P467L mice in response to high salt. These data indicate that loss of PPARγ in smooth muscle cells predispose to impaired vascular relaxation, renal dysfunction, and salt‐sensitive hypertension. This study highlights the significance of vascular PPARγ in regulating systemic physiological responses.Support or Funding InformationNIH and AHA Grants

Effects of silodosin on bladder activity in rats with frequent urination induced by pelvic venous congestion.

To examine the effects of silodosin on bladder activity using female rats with frequent urination induced by pelvic venous congestion. A total of 24 female rats were divided into three groups: sham, pelvic venous congestion and pelvic venous congestion/silodosin group. Rats in the pelvic venous congestion and pelvic venous congestion/silodosin groups were anesthetized with isoflurane, after which the bilateral common iliac veins and uterine veins were ligated. In the pelvic venous congestion/silodosin group, silodosin (0.3 mg/kg/day) was given using an osmotic pump implanted into the subcutaneous space of the back. After 5-6 weeks, analysis of voiding behavior, measurements of urinary 8-hydroxydeoxyguanosine and nitric oxide metabolites, continuous cystometry under urethane anesthesia, and Evans blue dye extravasation test of the bladder were carried out. In comparison with sham rats, pelvic venous congestion rats showed an increase in urination frequency with a concomitant increase in urine volume, a shorter interval between bladder contractions on continuous cystometry, an increase in urinary 8-hydroxydeoxyguanosine, a decrease in urinary nitric oxide metabolites and an increase in vesical vascular permeability. In comparison with pelvic venous congestion rats, pelvic venous congestion/silodosin rats showed a decrease in urination frequency with a concomitant decrease in urine volume, a lower maximum bladder contraction pressure, a longer interval between bladder contractions, an increase in urinary nitric oxide metabolites and a decrease in vascular permeability. Silodosin might improve both bladder dysfunction caused by pelvic venous congestion, and the pelvic venous congestion itself.

Naftopidil improves locomotor activity and urinary frequency in rats with pelvic venous congestion.

The α1D/A receptor antagonist, naftopidil, inhibits micturition reflex by acting on various different sites. We examined the effects of naftopidil on bladder activity and changes in the induced urinary frequency using female rats with pelvic venous congestion (PC). Twenty-four female rats were divided into sham, PC, and PC/naftopidil groups. After anesthetizing rats in the PC and PC/naftopidil groups, the bilateral common iliac veins and uterine veins were ligated. Rats in the sham and PC groups were fed a standard diet, while rats in the PC/naftopidil group were fed diets containing 0.04% naftopidil. After 4 weeks of treatment, locomotor activity, urinary nitric oxide metabolites (NOx), continuous cystometry, and plasma monoamine measurements were performed. PC rats exhibited a decrease of locomotor activity, a shorter interval between bladder contractions on continuous cystometry, and decreased urinary NOx and plasma serotonin levels than the sham rats. The PC/naftopidil rats exhibited an increase of locomotor activity, a longer interval between bladder contractions, and increased urinary NOx and plasma serotonin levels. Therefore, naftopidil might improve bladder dysfunction induced by pelvic venous congestion due to several actions in the central nervous system and bladder tissue, as well as acting as an α1 blocker to cause pelvic venous dilation.

Beetroot juice increase nitric oxide metabolites in both men and women regardless of body mass

The nitrate (NO3−) present in beetroot juice (BJ) has been studied for its effect on the cardiovascular system by converting to nitric oxide (NO). In the present study, we evaluated the effect of BJ on the excretion of NO metabolites and its relationship with body mass in both men and women. NO metabolites – urinary NO3−, nitrite (NO2−) and NOx were analyzed by using a high-performance liquid chromatography system. There were significant increases in urinary NO3−, NO2− and NOx in BJ as compared to PLA (BJ without NO3−). No significant difference between men and women was observed in NO metabolites after BJ at any time point. There were no significant relationships between urinary NO3−, NO2− and NOx and body mass in BJ intervention for both men and women. In conclusion, urinary NO metabolites after BJ consumption increases in similar manner between sexes regardless of body mass.

PS-205 Inhaled Nitric Oxide Increases Urinary Nitric Oxide Metabolites And Cgmp In Premature Infants: Relationship To Pulmonary Outcome

Background/aims Inhaled NO (iNO) has been tested for prevention of bronchopulmonary dysplasia in premature infants, however the role of cGMP is not known. We hypothesised that levels of NO metabolites (NOx) and cGMP in urine, as a non-invasive source for biospecimen collection, would reflect the dose of iNO and relate to pulmonary outcome. Methods Studies were performed on 125 infants Results In NO-treated infants there was a significant dose-dependent increase of both urinary NOx and cGMP per creatinine (maximal 3.1- and 2-fold, respectively, at 10–20 ppm iNO) compared to off iNO. The ratio of cGMP to NOx, which may reflect efficiency of NO signalling via guanylate cyclase, was lower (mean 38%) at all doses of iNO compared to off NO. NOx and cGMP concentrations at both 2 ppm and off iNO were inversely related to severity of lung disease (Respiratory Severity Score) during the first month, and the NOx levels were lower in infants who died or developed BPD at term. NOx was higher in Caucasian compared to other infants at all iNO doses. Conclusions Urinary NOx and cGMP are biomarkers of endogenous NO production and lung uptake of iNO, and levels at some doses reflect the severity of lung disease in infants. These results support a role of the NO-cGMP pathway in lung development and repair from injury.

Urinary nitric oxide metabolites in children with celiac disease after long-term consumption of oats-containing gluten-free diet

Objective. Oats are accepted in the gluten-free diet (GFD) for children with celiac disease (CD). Some reports have indicated, however, that not all celiac patients tolerate oats. We have previously shown that some children still have high levels of urinary nitric oxide (NO) metabolites as markers of intestinal inflammation after 1 year on GFD with oats. In this study, we measured urinary NO metabolites in CD children who had been consuming oats-containing GFD for an extended, 2–6-year period, also taking into consideration ordinary consumption of nitrite/nitrate-rich foods close to the urine sampling. Materials and Methods. Morning urinary nitrite/nitrate concentrations were measured in 188 pediatric CD patients. A questionnaire was used to elucidate factors possibly affecting the urinary levels, for example, dietary factors, asthma, or urinary tract infection. Results. Oats were consumed by 89.4% of the patients for a median time of 3 years. The median nitrite/nitrate level was 980 μM. The majority (70.2%) who consumed oats had low levels of urinary nitrite/nitrate, that is, <1400 μM, while 29.8% demonstrated high levels, that is, >1400 μM. Nitrite/nitrate-rich foods did not significantly influence the urinary concentrations. Conclusion. The urinary levels of NO metabolites revealed two subpopulations, one with high and one with low levels. The high levels could be possibly due to poor adherence to the GFD, sensitivity to oats, or some unknown factor(s). Nitrate-rich foods, asthma, or urinary tract infection did not affect the result. The elevated levels of NO metabolites could indicate mucosal inflammation and pinpoint the need of careful follow-up of children on oats-containing GFD.

Inhaled nitric oxide increases urinary nitric oxide metabolites and cyclic guanosine monophosphate in premature infants: relationship to pulmonary outcome.

Inhaled nitric oxide (iNO) has been tested to prevent bronchopulmonary dysplasia (BPD) in premature infants, however, the role of cyclic guanosine monophosphate (cGMP) is not known. We hypothesized that levels of NO metabolites (NOx) and cGMP in urine, as a noninvasive source for biospecimen collection, would reflect the dose of iNO and relate to pulmonary outcome. Studies were performed on 125 infants who required mechanical ventilation at 7 to 14 days and received 24 days of iNO at 20-2 ppm. A control group of 19 infants did not receive iNO. In NO-treated infants there was a dose-dependent increase of both NOx and cGMP per creatinine (maximal 3.1- and 2-fold, respectively, at 10-20 ppm iNO) compared with off iNO. NOx and cGMP concentrations at both 2 ppm and off iNO were inversely related to severity of lung disease during the 1st month, and the NOx levels were lower in infants who died or developed BPD at term. NOx was higher in Caucasian compared with other infants at all iNO doses. Urinary NOx and cGMP are biomarkers of endogenous NO production and lung uptake of iNO, and some levels reflect the severity of lung disease. These results support a role of the NO-cGMP pathway in lung development.

Renin-Angiotensin System Inhibitors Reduce Serum Asymmetric Dimethylarginine Levels and Oxidative Stress in Normotensive Patients with Chronic Kidney Disease

Background/Aims: The purpose of our study was to elucidate the relationship between asymmetric dimethylarginine (ADMA) and intrarenal lesions and to determine the effect of renin-angiotensin system inhibitors (RAS-Is) on serum ADMA levels, nitric oxide (NO) synthesis and oxidative stress in normotensive patients with chronic kidney disease (CKD). Methods: This study included 23 normotensive patients with chronic glomerulonephritis and normal or mildly impaired renal function who underwent renal biopsy. We evaluated the relationship between serum ADMA levels and intrarenal lesions, and examined renal function, urinary protein excretion, ADMA levels, NO synthesis, oxidative stress and blood pressure (BP) before and 3 months after starting the treatment with RAS-Is. Results: Serum ADMA levels were correlated only with arterial intimal fibroplastic thickness. Despite comparable renal function and BP, serum ADMA levels and excretion of urinary protein excretion significantly decreased, and urinary NO metabolite excretion significantly increased after starting the treatment with RAS-Is. Oxidative stress markers also tended to be reduced by the treatment. Conclusion: These findings suggest that RAS-Is improve the NO system and decrease oxidative stress in normotensive patients with CKD. In addition, ADMA may be associated with intrarenal lesions and can be a useful marker for the effects of treatment in the early stages of CKD.

Crocetin, a Carotenoid from Gardenia jasminoides Ellis, Protects against Hypertension and Cerebral Thrombogenesis in Stroke‐prone Spontaneously Hypertensive Rats

Crocetin is a natural carotenoid dicarboxylic acid that is found in the fruit of Gardenia jasminoides Ellis (Cape Jasmine) and in the stamen and pistil of Crocus sativus L. (saffron). It is used worldwide as an important spice, food colorant, and herbal medicine. In the current investigation, we have examined the cardiovascular effects of crocetin using stroke-prone spontaneously hypertensive rats (SHRSPs). Male SHRSPs (6 weeks old) were classified into three groups: a control group and two crocetin groups (25 and 50 mg/kg/day). The animals were given crocetin for 3 weeks. Body weights in each group were not significantly different during the treatment period, but the increase in systolic blood pressures observed with age was significantly moderated by crocetin. Thrombogenesis, assessed using a He-Ne laser technique in pial vessels, was significantly decreased. Antioxidant activity, assessed by measuring urinary 8-hydroxy-2'-deoxyguanosine levels, together with urinary nitric oxide (NO) metabolite levels, was increased significantly after treatment. Acetylcholine-induced vasodilation was measured using the aorta and indicated that endothelial function was significantly improved by crocetin. These results strongly suggest that the antihypertensive and antithrombotic effects of crocetin were related to an increase in bioavailable NO, possibly mediated by decreased inactivation of NO by reactive oxygen species.

Abstract 273: Increased Blood Pressure and Proteinuria and Absence of Increased Nitric Oxide (NO) Production During Pregnancy in the Dahl Salt Sensitive (S) Rat

In normal pregnancy, systemic vasodilation due to increased NO production allows a drop in blood pressure (BP) despite increased volume retention. Little is known about the pathogenesis of preeclampsia, defined by increased BP and proteinuria, due to a lack of animal models that spontaneously develop the disease. Here we tested the hypothesis that the Dahl S rat, a genetic model of hypertension and kidney disease, is also a spontaneous model of preeclampsia. Female Dahl S rats were implanted with a telemetry unit, and baseline BP was recorded. Rats were placed in metabolic cages for 24 hr urine collection while on a low nitrate diet, and urinary protein and NO metabolite concentrations were measured via Bradford and Greiss assays, respectively. There were no differences in baseline BP (152±1 vs 151±4 mmHg) or proteinuria (61±10 vs 60±17 mg/d) in the rats selected for mating vs virgin rats (n=5-7). Pregnancy was confirmed by presence of sperm (day 1). Measurements were made during mid and late pregnancy (days 10-11, 17-18), and terminal measurements were taken on day 19. Pregnant rats exhibited an increase in BP and proteinuria with no change in urinary NOx excretion (Table), while no changes were observed in age-matched virgin rats. Kidney cortex abundance of neither NOS1 nor NOS3 was increased at late pregnancy; however, plasma concentration of the endogenous NOS inhibitor ADMA was increased in late pregnant compared to virgin rats (0.82±0.06 vs. 0.62±0.06 μM, p&lt;0.05). These data suggest that the Dahl S rat cannot upregulate NO production during pregnancy; therefore, this relative NO deficiency may contribute to worsening hypertension and proteinuria during pregnancy in this strain.

Bioavailability of epicatechin and effects on nitric oxide metabolites of an apple flavanol‐rich extract supplemented beverage compared to a whole apple puree: a randomized, placebo‐controlled, crossover trial

Flavanol-rich foods are known to exert beneficial effects on cardiovascular health. The biological effects depend on bioavailability of flavanols which may be influenced by food matrix and dose ingested. We compared the bioavailability and dose-response of epicatechin from whole apple and an epicatechin-rich extract, and the effects on plasma and urinary nitric oxide (NO) metabolites. In a randomized, placebo-controlled, crossover trial, subjects consumed drinks containing 70 and 140 mg epicatechin from an apple extract and an apple puree containing 70 mg epicatechin. Blood and urine samples were collected for 24 h post ingestion. Maximum plasma concentration, AUC(0-24 h) , absorption and urinary excretion were all significantly higher after ingestion of both epicatechin drinks compared with apple puree (p < 0.05). Time to maximum plasma concentration was significantly later for the puree compared with the drinks (p < 0.01). Epicatechin bioavailability was >2-fold higher after ingestion of the 140 mg epicatechin drink compared to the 70 mg epicatechin drink (p < 0.05). Excretion of NO metabolites was higher for all test products compared with placebo, which was significant for the high dose drink (p = 0.016). Oral bioavailability of apple epicatechin increases at higher doses, is reduced by whole apple matrix and has the potential to increase NO bioavailability.

Abstract 93: Hypertension and Renal Injury Induced by Nitric Oxide Depletion are Ameliorated by Concomitant Depletion of Hydrogen Sulfide

Chronic nitric oxide (NO) depletion induces hypertension and renal damage. Chronic kidney disease is associated with decreased NO availability and less renal H 2 S production. We hypothesized that combined depletion of NO and H 2 S aggravates hypertension and renal injury. Male 8-wk old Sprague Dawley rats were treated with vehicle, NO synthase inhibitor L-NG-nitroarginine (LNNA; 125 mg/L in drinking water), cystathionine-γ-lyase (CSE) inhibitor propargylglycine (PAG; 37.5 mg/kg BW ip daily) or LNNA + PAG for 1 and 4 weeks (6 rats/group). LNNA after 4w increased systolic blood pressure (SBP; 223±10 vs . 137±3 mmHg in controls; P&lt;0.01), proteinuria (144±35 vs. 17±2 mg/d; P&lt;0.01), uremia (16.6±4.2 vs . 7.0±0.4 mmol/L; P&lt;0.05) and tubulo-interstitial injury (P&lt;0.01). LNNA reduced urinary NO metabolite (NOx) excretion by ∼85% after 1w and 4w. PAG alone had no effect on SBP, renal function or injury, but did reduce urinary NOx excretion. Co-treatment with PAG ameliorated LNNA-induced hypertension (182±10 mmHg; P&lt;0.01) and prevented proteinuria (27±3 mg/d), uremia (8.3±0.4 mmol/L) and tubulo-interstitial injury, but did not further reduce urinary NOx excretion. Renal H 2 S production was almost absent in all PAG groups after 1w and 4w (P&lt;0.01) and was reduced in LNNA-treated rats after 4w (4.6±1.4 vs . 9.2±0.5 μmol/hr/mg; P&lt;0.01). Renal HO-1 gene expression was strongly induced in all PAG-treated groups after 1w and 4w (4 to 19-fold; P&lt;0.01) whereas LNNA only increased HO-1 gene expression at 4w (P&lt;0.01). Immunohistochemistry showed that renal HO-1 protein was primarily interstitial in all PAG-treated groups at 1w and 4w. In contrast, LNNA only showed HO-1 in tubular epithelium in conjunction with protein casts. Depleting NO caused hypertension and renal damage followed by reduced renal H 2 S production and increased renal HO-1 expression. Surprisingly, concomitant inhibition of CSE ameliorated hypertension and prevented renal injury. PAG almost completely blocked renal H 2 S production and caused strong induction of renal HO-1, independently of injury, suggesting that H 2 S suppresses renal HO-1 expression. In conclusion, concomitant upregulation of HO-1 expression by inhibition of H 2 S production, prevents LNNA-induced hypertension and renal injury.

Hyperfiltration and effect of nitric oxide inhibition on renal and endothelial function in humans with uncomplicated type 1 diabetes mellitus

Studies of experimental diabetes mellitus (DM) suggest that increased nitric oxide (NO) bioactivity contributes to renal hyperfiltration. However, the role of NO in mediating hyperfiltration has not been fully elucidated in humans. Our aim was to examine the effect of NO synthase inhibition on renal and peripheral vascular function in normotensive subjects with uncomplicated type 1 DM. Renal function and brachial artery flow-mediated vasodilatation (FMD) were measured before and after an intravenous infusion of the NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NMMA) in 21 healthy control and 37 type 1 DM patients. Measurements in DM participants were made under clamped euglycemic conditions. The effect of l-NMMA on circulating and urinary NO metabolites (NO(x)) and cGMP and on urinary prostanoids was also determined. Baseline characteristics were similar in the two groups. For analysis, the DM patients were divided into those with hyperfiltration (DM-H, n = 18) and normal glomerular filtration rate (GFR) levels (DM-N, n = 19). Baseline urine NO(x) and cGMP were highest in DM-H. l-NMMA led to a decline in GFR in DM-H (152 ± 16 to 140 ± 11 ml·min(-1)·1.73 m(-2)) but not DM-N or healthy control participants. The decline in effective renal plasma flow in response to l-NMMA (806 ± 112 to 539 ± 80 ml·min(-1)·1.73 m(-2)) in DM-H was also exaggerated compared with the other groups (repeated measures ANOVA, P < 0.05), along with declines in urinary NO(x) metabolites and cGMP. Baseline FMD was lowest in DM-H compared with the other groups and did not change in response to l-NMMA. l-NMMA reduced FMD and plasma markers of NO bioactivity in the healthy control and DM-N groups. In patients with uncomplicated type 1 DM, renal hyperfiltration is associated with increased NO bioactivity in the kidney and reduced NO bioactivity in the systemic circulation, suggesting a paradoxical state of high renal and low systemic vascular NO bioactivity.

Vitamin D and Parathyroid Hormone Relationships with Urinary Nitric Oxide Metabolites and Plasma Isoprostanes in African-Americans

Background: Vitamin D deficiency and secondary rises in parathyroid hormone (PTH) are highly prevalent in obese African-Americans. Endothelial dysfunction related to oxidative stress is more common in African-Americans compared to whites. Currently, the association of vitamin D (25-hydroxyvitamin D, 25-OH D) and PTH to nitric oxide metabolites (NOx) – nitrate and nitrite – and oxidative stress in African-Americans is unknown. Objective: A cross-sectional design was utilized to determine the association of 25-OH D and PTH with urinary NOx (UNOx) (n = 101) and plasma isoprostanes (n = 125), an oxidative stress marker, in overweight (body mass index of 25–39.9), normotensive African-Americans aged ≥35 years. Measurements: Multivariable linear regression analysis adjusted for age, sex, body mass index, and season was used to determine the relationship of 25-OH D and PTH to UNOx and isoprostanes. General linear models, adjusted for the same covariates, contrasted UNOx across three mutually exclusive vitamin D/PTH groups: (1) normal 25-OH D (51–249 nmol/l) and normal PTH (≤65 pg/ml); (2) low 25-OH D and normal PTH, and (3) low 25-OH D and high PTH. Results: 25-OH D was directly associated with UNOx before (p = 0.02) and after (p = 0.03) adjustment for PTH levels. A borderline significant association was observed between PTH and isoprostanes (p = 0.08). UNOx was 424, 290, and 270 µmol/8 h, respectively, across vitamin D/PTH groups 1–3 (p = 0.08). Conclusion: 25-OH D was directly associated with NO availability and PTH was positively, though borderline, associated with isoprostanes in overweight, normotensive adult African-Americans.