Low Nitric Oxide Research Articles

Rapid adiposity growth increases risks of new-onset asthma and airway inflammation in children.

We aim to (1) examine the influence of long-term adiposity status/short-term adiposity changes on asthma with high or low fractional exhaled nitric oxide (FeNO), and (2) to determine the differences in long-term adiposity status/short-term adiposity changes on atopy, airway inflammation and pulmonary function. We recruited 2450 fourth- to sixth-grade children from the nationwide Taiwan Children Health Study. Data regarding various adiposity indicators, atopic status, pulmonary function tests and asthma outcomes were collected annually. New-onset asthma was stratified by airway inflammation status using FeNO. The generalized estimating equation was used for analyzing longitudinal relationships between long-term adiposity status/short-term adiposity changes and new-onset asthma. Individual adiposity growth slopes were obtained using a hierarchical linear model to establish the relationships between short-term adiposity changes and asthma among children with high airway inflammation. We found long-term adiposity status predicted childhood asthma with low FeNO, whereas short-term adiposity changes may increase risks of childhood asthma with high FeNO. Long-term adiposity status reduced pulmonary function, whereas short-term adiposity increase were associated with atopic diseases and airway inflammation. Obesity-induced asthma could be mediated by high or low airway inflammation, depending on the velocity of increase in adiposity. Rapid adiposity growth may increase risks of childhood asthma and airway inflammation.

Further studies of the effects of aging on arginine metabolites in the rat vestibular nucleus and cerebellum

Some studies have demonstrated that aging is associated with impaired vestibular reflexes, especially otolithic reflexes, resulting in postural instability. However, the neurochemical basis of these age-related changes is still poorly understood. The l-arginine metabolic system has been implicated in changes in the brain associated with aging. In the current study, we examined the levels of l-arginine and its metabolizing enzymes and downstream metabolites in the vestibular nucleus complex (VNC) and cerebellum (CE) of rats with and without behavioral testing which were young (4months old), middle-aged (12months old) or aged (24months old). We found that aging was associated with lower nitric oxide synthase activity in the CE of animals with testing and increased arginase in the VNC and CE of animals with testing. l-citrulline and l-ornithine were lower in the VNC of aged animals irrespective of testing, while l-arginine and l-citrulline were lower in the CE with and without testing, respectively. In the VNC and CE, aging was associated with lower levels of glutamate in the VNC, irrespective of testing. In the VNC it was associated with higher levels of agmatine and putrescine, irrespective of testing. In the CE, aging was associated with higher levels of putrescine in animals without testing and with higher levels of spermine in animals with testing, and spermidine, irrespective of testing. Multivariate analyses indicated significant predictive relationships between the different variables, and there were correlations between some of the neurochemical variables and behavioral measurements. Cluster analyses revealed that aging altered the relationships between l-arginine and its metabolites. The results of this study demonstrate that there are major changes occurring in l-arginine metabolism in the VNC and CE as a result of age, as well as behavioral activity.

Sickle cell anemia - Nitric oxide related genetic modifiers of hematological and biochemical parameters.

Sickle cell anemia (SCA) is an inherited blood disorder. SCA patients present clinical and hematologic variability that cannot be only explained by the single mutation in the beta-globin gene. Others genetic modifiers and environmental effects are important for the clinical phenotype. SCA patients present arginine deficiency that contributes to a lower nitric oxide (NO) bioactivity. The aim of this work is to determine the association between hematological and biochemical parameters and genetic variants from eNOS gene, in pediatric SCA patients. 26 pediatric SCA patients were genotyped using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) techniques in three important eNOS gene polymorphisms - rs2070744, rs1799983 and intron 4 VNTR. Results from this study show a significant statistical association between some parameters and genetic variants: an increased reticulocyte count and high serum lactate dehydrogenase levels were associated with both the rs2070744_TT and the rs1799983_GG genotypes at eNOS gene and high levels of neutrophils were associated with the eNOS4a allele at intron 4 VNTR. Our results reinforce the importance of NO bioactivity in SCA. We presume that NO, and its precursors might be used as therapy to improve the quality of life of SCA patients.

Concurrent Beet Juice and Carbohydrate Ingestion: Influence on Glucose Tolerance in Obese and Nonobese Adults.

Insulin resistance and obesity are characterized by low nitric oxide (NO) bioavailability. Insulin sensitivity is improved with stimulation of NO generating pathways. Consumption of dietary nitrate (NO3−) increases NO formation, via NO3− reduction to nitrite (NO2−) by oral bacteria. We hypothesized that acute dietary nitrate (beet juice) ingestion improves insulin sensitivity in obese but not in nonobese adults. 12 nonobese (body mass index: 26.3 ± 0.8 kg/m2 (mean ± SE)) and 10 obese adults (34.0 ± 0.8 kg/m2) ingested beet juice, supplemented with 25 g of glucose (carbohydrate load: 75 g), with and without prior use of antibacterial mouthwash to inhibit NO3− reduction to NO2−. Blood glucose concentrations after beet juice and glucose ingestion were greater in obese compared with nonobese adults at 60 and 90 minutes (P = 0.004). Insulin sensitivity, as represented by the Matsuda Index (where higher values reflect greater insulin sensitivity), was lower in obese compared with nonobese adults (P = 0.009). Antibacterial mouthwash rinsing decreased insulin sensitivity in obese (5.7 ± 0.7 versus 4.9 ± 0.6) but not in nonobese (8.1 ± 1.0 versus 8.9 ± 0.9) adults (P = 0.048). In conclusion, insulin sensitivity was improved in obese but not in nonobese adults following coingestion of beet juice and glucose when oral bacteria nitrate reduction was not inhibited. Obese adults may benefit from ingestion of healthy nitrate-rich foods during meals.

Commentary: On the Pathophysiology of Coronary Heart Disease

This article highlights some important aspects of the pathophysiology of coronary artery disease, namely: • The distribution of lesions within the arterial tree at sites of low wall shear stress. • The potential role of low flow-mediated arterial dilatation at sites of low wall stress. • Low flow-mediated dilatation leads to low nitric oxide production by the arterial endothelium and consequent reduced protection against lesion formation. • Flow-mediated dilatation is reduced by high lumenal glucose concentration. • The role of the glycocalyx dysfunction in mediating flow-mediated dilatation and consequent reduced NO production by the arterial endothelium and cell adhesion. • Stenoses cause convective acceleration of blood velocity and a consequent increase in platelet shear stress. • Increased platelet shear stress activates platelets with release of serotonin. • Serotonin activates more platelet activation via the 5HT2A platelet receptor causing a positive feedback and thrombus growth. • Arterial thrombus growth is abolished by 5HT2A receptor antagonists, the key to improved treatment of the disease. • One 5HT2A receptor antagonist has been shown in humans to be safe and to cause no excess bleeding from wounds.

Modulation of low shear stress‑induced eNOS multi‑site phosphorylation and nitric oxide production via protein kinase and ERK1/2 signaling.

Physiological shear stress has been demonstrated to serve an atheroprotective function by stimulating endothelial nitric oxide synthase (eNOS) multi‑site phosphorylation. Low shear stress (LSS) serves an atheroprone role by increasing endothelial cell apoptosis and inflammation. The present study assessed whether LSS inhibited nitric oxide (NO) production in human umbilical vein endothelial cells by modulating eNOS phosphorylation and potential signaling pathways. A parallel flow chamber imposed with 2dyn/cm2 shear stress on endothelial cells was used. Western blotting and 4,5‑diaminofluorescein diacetate were used to analyze the protein expression levels and NO production. LSS activated eNOS‑Ser1177 and eNOS‑Thr495, but inhibited eNOS‑Ser633. NO production was decreased after a transient increase at 5min. LSS‑stimulated phosphorylation of eNOS‑Ser1177 and ‑Thr495 were suppressed by the Akt inhibitor, perifosine, and extracellular signal regulated kinases1/2 (ERK1/2) inhibitor, PD98059, respectively. Additionally, the phosphorylation of eNOS‑Ser633 inhibited by LSS was restored by the protein kinase A activator, 8‑Bromo‑cAMP. PD98059 completely inhibited the LSS‑induced downregulation of NO production. NO downregulation in response to LSS was intensified by perifosine and was partly inhibited by 8‑Bromo‑cAMP. These results indicated that LSS‑induced activation of ERK1/2/eNOS‑Thr495 serves a major role in inhibiting endothelial NO synthase, which may explain the proinflammatory and proatherosclerotic properties of LSS.

Modulating Oxidative Stress and Inflammation in Elders: The MOXIE Study

ABSTRACTCardiovascular disease (CVD) is the leading cause of death among women in the United States. Endothelial dysfunction and arterial stiffness increase with advancing age and are early predictors of future CVD outcomes. We designed the Modulating Oxidative Stress and Inflammation in Elders (MOXIE) study to examine the effects of 100% watermelon juice as a “food-first” intervention to reduce CVD risk among African American (AA) and European American (EA) women aged 55–69 years. Vascular dysfunction is more pronounced in AA compared to EA women due in part to lower nitric oxide bioavailability caused by higher oxidative stress. However, bioactive compounds in watermelon may improve vascular function by increasing nitric oxide bioavailability and antioxidant capacity. This trial will use a randomized, placebo-controlled, crossover design to investigate the potential of 100% watermelon juice to positively impact various robust measures of vascular function as well as serum biomarkers of oxidative stress and antioxidant capacity. This nutrition intervention and its unique methodology to examine both clinical and mechanistic outcomes are described in this article.

Culture supernatants of cervical cancer cells induce an M2 phenotypic profile in THP-1 macrophages

Patients with cervical cancer (CxCa) typically present an infiltrate of tumor-associated macrophages, which is associated with a poor prognosis. We found that CxCa cell lines (HeLa, SiHa, and C-33A) secreted factors involved in regulating tumor growth including IL-6, IL-4, PDGFAA, HGF, VEGF, ANG-2, and TGF-β3. We assessed the effects of culture supernatants from these cell lines on macrophages derived from the THP-1 cell line. Macrophages treated with culture supernatants from CxCa cells developed an M2-like phenotype with expression of CD163, low nitric oxide release, and high secretion of IL-6, PDGFAA, HGF, ANG-2, and VEGF. The macrophages continued to produce PDGFAA, PDGFBB, and VEGF 48h after the CxCa cell culture supernatants were removed. The induction of M2 macrophages in vivo favors tumor growth, angiogenesis, tissue remodeling, and metastasis. These results demonstrated that factors secreted by CxCa cells induced a stable M2 phenotype in THP-1 macrophages.

Phenotype-Specific Treatment of Heart Failure With Preserved Ejection Fraction: A Multiorgan Roadmap.

Heart failure (HF) with preserved ejection fraction (EF; HFpEF) accounts for 50% of HF cases, and its prevalence relative to HF with reduced EF continues to rise. In contrast to HF with reduced EF, large trials testing neurohumoral inhibition in HFpEF failed to reach a positive outcome. This failure was recently attributed to distinct systemic and myocardial signaling in HFpEF and to diversity of HFpEF phenotypes. In this review, an HFpEF treatment strategy is proposed that addresses HFpEF-specific signaling and phenotypic diversity. In HFpEF, extracardiac comorbidities such as metabolic risk, arterial hypertension, and renal insufficiency drive left ventricular remodeling and dysfunction through systemic inflammation and coronary microvascular endothelial dysfunction. The latter affects left ventricular diastolic dysfunction through macrophage infiltration, resulting in interstitial fibrosis, and through altered paracrine signaling to cardiomyocytes, which become hypertrophied and stiff because of low nitric oxide and cyclic guanosine monophosphate. Systemic inflammation also affects other organs such as lungs, skeletal muscle, and kidneys, leading, respectively, to pulmonary hypertension, muscle weakness, and sodium retention. Individual steps of these signaling cascades can be targeted by specific interventions: metabolic risk by caloric restriction, systemic inflammation by statins, pulmonary hypertension by phosphodiesterase 5 inhibitors, muscle weakness by exercise training, sodium retention by diuretics and monitoring devices, myocardial nitric oxide bioavailability by inorganic nitrate-nitrite, myocardial cyclic guanosine monophosphate content by neprilysin or phosphodiesterase 9 inhibition, and myocardial fibrosis by spironolactone. Because of phenotypic diversity in HFpEF, personalized therapeutic strategies are proposed, which are configured in a matrix with HFpEF presentations in the abscissa and HFpEF predispositions in the ordinate.

Effects of shear stress on endothelial cells: go with the flow.

Haemodynamic forces influence the functional properties of vascular endothelium. Endothelial cells (ECs) have a variety of receptors, which sense flow and transmit mechanical signals through mechanosensitive signalling pathways to recipient molecules that lead to phenotypic and functional changes. Arterial architecture varies greatly exhibiting bifurcations, branch points and curved regions, which are exposed to various flow patterns. Clinical studies showed that atherosclerotic plaques develop preferentially at arterial branches and curvatures, that is in the regions exposed to disturbed flow and shear stress. In the atheroprone regions, the endothelium has a proinflammatory phenotype associated with low nitric oxide production, reduced barrier function and increased proadhesive, procoagulant and proproliferative properties. Atheroresistant regions are exposed to laminar flow and high shear stress that induce prosurvival antioxidant signals and maintain the quiescent phenotype in ECs. Indeed, various flow patterns contribute to phenotypic and functional heterogeneity of arterial endothelium whose response to proatherogenic stimuli is differentiated. This may explain the preferential development of endothelial dysfunction in arterial sites with disturbed flow.

Influence of multiple fuel injection strategies on performance and combustion characteristics of a diesel fuelled HCCI engine – An experimental investigation

Homogeneous charge compression ignition (HCCI) engines have the potential to operate with low nitric oxide (NOx) and soot emissions. Diesel HCCI engines pose challenges related to too early combustion, high combustion rates and wall wetting of the fuel. Injecting diesel in pulses has been shown to improve combustion in these engines. In this work, diesel injected in five timed pulses has been investigated in a compression ignition (CI) engine operating in HCCI mode. The influence of varying the first, middle and last injection pulse durations alone, in sequence, while maintaining all the other injection pulse durations equal has been studied. In addition, the influence of the injection timing of the last injection pulse was also studied. A comparison of the results has been made with HCCI operation using diesel injection in a single pulse at an injection timing of 100°CA before top dead centre (TDC) and also with the conventional compression ignition mode of operation under similar outputs. The results show that, multiple pulse (MP) injection is better than injecting the fuel in a single pulse (SP) as it leads to lower emissions and higher thermal efficiency. This is because of better combustion phasing and higher heat release rates.

TCF7L2 polymorphism is associated with low nitric oxide release, endothelial dysfunction and enhanced inflammatory response after myocardial infarction

BackgoundThe favorable effects of insulin during myocardial infarction (MI) remain unclear due to the divergence between mechanistic studies and clinical trials of exogenous insulin administration. The rs7903146 polymorphism of the transcription factor 7-like 2 (TCF7L2) gene is associated with attenuated insulin secretion. MethodsIn non-diabetic patients with ST-elevation MI (STEMI), using such a model of genetically determined down-regulation of endogenous insulin secretion we investigated the change in plasma insulin, C-peptide, interleukin-2 (IL-2), C-reactive protein (CRP), and nitric oxide (NOx) levels between admission (D1) and the fifth day after MI (D5). Coronary angiography and flow-mediated dilation (FMD) were performed at admission and 30days after MI, respectively. Homeostasis Model Assessment estimated insulin secretion (HOMA2%β) and insulin sensitivity (HOMA2%S). ResultsAlthough glycemia did not differ between genotypes, carriers of the T-allele had lower HOMA2%β and higher HOMA2%S at both D1 and D5. As compared with non-carriers, T-allele carriers had higher plasma IL-2 and CRP at D5, higher intracoronary thrombus grade, lower FMD and NOx change between D1 and D5 and higher 30-day mortality. ConclusionIn non-diabetic STEMI patients, the rs7903146 TCF7L2 gene polymorphism is associated with lower insulin secretion, worse endothelial function, higher coronary thrombotic burden, and higher short-term mortality. General significanceDuring the acute phase of MI, a lower capacity of insulin secretion may influence clinical outcome.

Neonatal malnutrition programs the oxidant function of macrophages in response to Candida albicans

Experimental maternal nutrition restriction models are used to investigate short or long-term consequences of nutritional deficiency on puppies' growth. By assuming that the immune function is directly related to host's nutritional status, the current study aims to investigate the effects of neonatal malnutrition on oxidative stress and on the cell death of the alveolar macrophage after in vitro infection by Candida albicans. Wistar rats were suckled by mothers fed on diets containing 17% protein (Nourished group) or 8% protein (Malnourished group) in the current assay. Both groups received the standard diet used in the vivarium until adulthood, after weaning. The results showed that the offspring from mothers fed on low-protein diet presented lower body weight from 5 days of life on. Their low weight remained until adulthood when it was compared to that of rats in the nourished group. Superoxide and nitric oxide production was lower in malnourished animals and it was accompanied by low inducible nitric oxide synthase gene expression levels in systems in which the alveolar macrophages were challenged by immunogenic stimulus. No significant differences were observed in comparisons performed between the nourished and malnourished groups in any of the analyzed cell viability (apoptosis/necrosis) parameters. The fungal inoculum-stimulated system induced higher oxidative stress and cell death by necrosis. The current study demonstrated that dietary restriction during lactation alters the oxidant function of alveolar macrophages in puppies; It happens from the gene transcription step to the release of mediators, thus compromising the host's defenses against Candida albicans. It raises the possibility that Candida albicans may cease to be a commensal fungus to become a pathogen in offspring that have suffered nutritional deficiency during critical developmental periods, due to impaired immune responses.

Noninvasive Analysis of the Sputum Transcriptome Discriminates Clinical Phenotypes of Asthma.

It is increasingly recognized that asthma is a heterogeneous disease. Therefore, it is possible that analysis of gene expression in the airway will reveal clinically meaningful transcriptional endotypes of asthma (TEA clusters). We measured whole transcriptome gene expression profiles in the sputum and whole blood of 100 individuals with asthma and 12 control subjects using the Affymetrix HuGene ST 1.0 gene arrays. Unsupervised clustering was conducted using pathways from Kyoto Encyclopedia of Genes and Genomes (KEGG). This identified three TEA clusters that were correlated with clinical, physiologic, and inflammatory characteristics of the disease. TEA cluster 1 is a cluster of patients with asthma with a significantly higher rate of intubation (P = 0.05), a lower prebronchodilator FEV1 (P = 0.006), a higher bronchodilator response (P = 0.03), and higher exhaled nitric oxide levels (P = 0.04) than the other two TEA clusters. TEA cluster 2 has a higher rate of hospitalization for asthma (P = 0.04) and is heterogeneous. TEA cluster 3 is the largest cluster and has normal lung function, low exhaled nitric oxide levels, and lower inhaled steroid requirements. TEA cluster 1 had the highest sputum Th2 gene signature (IL-4, -5, and -13) compared with the other clusters. A classifier was developed that predicts TEA cluster assignment using 53 predictive genes in the circulation. The classifier was applied to gene expression data of children from the Asthma Biorepository for Integrative Genomic Exploration (Asthma BRIDGE) consortium cohort and confirmed that TEA clusters 1 and 2 are associated with history of intubation (P = 5.58 × 10(-06)) and hospitalization (P = 0.01), respectively. Analysis of the sputum transcriptome reveals three TEA clusters with different clinical and physiologic characteristics of disease in children and adults with asthma. This suggests that there are common transcriptomic signatures in the blood in children and adults with asthma that are associated with features of severe asthma.

Indications and contraindications for infusing specific amino acids (leucine, glutamine, arginine, citrulline, and taurine) in critical illness.

The review assesses the utility of supplementing parenteral or enteral nutrition of ICU patients with each of five specific amino acids that display pharmacological properties. Specifying indications implies also stating contraindications.Combined supplementation of amino acids with ω3-fatty acids and/or trace elements (immune-enhancing diets) will not be considered in this review because these mixtures do not allow the role of amino acids in the effect (positive or negative) of the mixture to be isolated, and so cannot show whether or not supplementation of a given amino acid is indicated. After decades of unbridled use of glutamine (GLN) supplementation in critically ill patients, recent large trials have brought a note of caution, indicating for example that GLN should not be used in patients with multiple organ failure. Yet these large trials do not change the conclusions of recent meta-analyses. Arginine (ARG), as a single dietary supplement, is probably not harmful in critical illness, in particular in a situation of ARG deficiency syndrome with low nitric oxide production. Citrulline supplementation strongly improves microcirculation in animal models with gut injury, but clinical studies are lacking. Taurine has a potent protective effect against ischemic reperfusion injury. Amino acid-based pharmaconutrition has displayed familiar 'big project' stages: enthusiasm (citrulline and taurine), doubt (GLN), hunt for the guilty (ARG), and backpedalling (leucine). Progress in this field is very slow, and sometimes gives way to retreat, as demonstrated by recent large trials on GLN supplementation.

A Model for Predicting the Lift-Off Height of Premixed Jets in Vitiated Cross Flow

Lean premixed single-stage combustion is state of the art for low pollution combustion in heavy-duty gas turbines with gaseous fuels. The application of premixed jets in multistage combustion to lower nitric oxide emissions and enhance turn-down ratio is a novel promising approach. At the Lehrstuhl für Thermodynamik, Technische Universität München, a large-scale atmospheric combustion test rig has been set up for studying staged combustion. The understanding of lift-off (LO) behavior is crucial for determining the amount of mixing before ignition and for avoiding flames anchoring at the combustor walls. This experiment studies jet LO depending on jet equivalence ratio (0.58–0.82), jet preheat temperature (288–673 K), cross flow temperature (1634–1821 K), and jet momentum ratio (6–210). The differences to existing LO studies are the high cross flow temperature and applying a premixed jet. The LO height of the jet flame is determined by OH* chemiluminescence images, and subsequently, the data is used to analyze the influence of each parameter and to develop a model that predicts the LO height for similar staged combustion systems. A main outcome of this work is that the LO height in a high temperature cross flow cannot be described by one dimensionless number like Damköhler- or Karlovitz-number. Furthermore, the ignition delay time scale τign also misses part of the LO height mechanism. The presented model uses turbulent time scales, the ignition delay, and a chemical time scale based on the laminar flame speed. An analysis of the model reveals flame stabilization mechanisms and explains the importance of different time scale.

Computational and Experimental Study of Neuroglobin and Carbon Monoxide

Neuroglobin is a hexacoordinated protein normally found in the brain and nervous system that is closely related to myoglobin and hemoglobin. We have previously shown that neuroglobin (Ngb) reduces nitrite to nitric oxide 50x faster than myoglobin, 500x faster than hemoglobin, and tightly binds to carbon monoxide (CO) with an association rate that is 500x faster than hemoglobin. In the beginning phase of my research, I will be computationally analyzing structures of Ngb and mutants which improve these qualities. The computational simulation will be used to help predict the following questions:1) Why are these Ngb mutants so much better at reducing nitrite than myoglobin and hemoglobin?2) Why do they have a higher binding affinity to CO than hemoglobin?3) What are the optimal mutants one can make to maximize nitrite reduction and CO binding?To analyze the structure of neuroglobin and the characteristics causing these phenomena, focusing on CO binding in this work, I am running 3 sets of molecular dynamic simulations of wild-type human Ngb (4mpm), reduced human Ngb (10j6), 4mpm mutants (C46A, C55A, H64L, H64Q), and 1OJ6 mutants (G46A, S55A, H64L, H64Q). I will also confirm my predictions with time-resolved absorption spectroscopy. These studies will help identify treatments for diseases involving low nitric oxide and carbon monoxide poisoning.

Tetrahydrobiopterin Role in human umbilical vein endothelial dysfunction in maternal supraphysiological hypercholesterolemia

Maternal physiological hypercholesterolemia (MPH) allows a proper foetal development; however, maternal supraphysiological hypercholesterolemia (MSPH) associates with foetal endothelial dysfunction and early development of atherosclerosis. MSPH courses with reduced endothelium-dependent dilation of the human umbilical vein due to reduced endothelial nitric oxide synthase activity compared with MPH. Whether MSPH modifies the availability of the nitric oxide synthase cofactor tetrahydrobiopterin is unknown. We investigated whether MSPH-associated lower umbilical vein vascular reactivity results from reduced bioavailability of tetrahydrobiopterin. Total cholesterol <7.2mmol/L was considered as maternal physiological hypercholesterolemia (n=72 women) and ≥7.2mmol/L as MSPH (n=35 women). Umbilical veins rings were used for vascular reactivity assays (wire myography), and primary cultures of human umbilical vein endothelial cells (HUVECs) to measure nitric oxide synthase, GTP cyclohydrolase 1, and dihydrofolate reductase expression and activity, as well as tetrahydrobiopterin content. MSPH reduced the umbilical vein rings relaxation caused by calcitonine gene-related peptide, a phenomenon partially improved by incubation with sepiapterin. HUVECs from MSPH showed lower nitric oxide synthase activity (l-citrulline synthesis from l-arginine) without changes in its protein abundance, as well as reduced tetrahydrobiopterin level compared with MPH, a phenomenon reversed by incubation with sepiapterin. Expression and activity of GTP cyclohydrolase 1 was lower in MSPH, without changes in dihydrofolate reductase expression. MSPH is a pathophysiological condition reducing human umbilical vein reactivity due to lower bioavailability of tetrahydrobiopterin leading to lower NOS activity in the human umbilical vein endothelium.

Vasoactive enzymes and blood flow responses to passive and active exercise in peripheral arterial disease

BackgroundPeripheral arterial disease (PAD) is characterised by impaired leg blood flow, which contributes to claudication and reduced exercise capacity. This study investigated to what extent vasoactive enzymes might contribute to altered blood flow in PAD (Fontaine stage II). MethodsWe compared femoral artery blood flow during reactive hyperaemia, leg-extension exercise and passive leg movement, and determined the level of vasoactive enzymes in skeletal muscle samples from the vastus lateralis in PAD (n = 10, 68.5 ± 6.5 years) and healthy controls (CON, n = 9, 62.1 ± 12.3 years). Leg blood flow was measured with Doppler ultrasound and muscle protein levels of phosphorylated endothelial nitric oxide synthase, NADPH oxidase, cyclooxygenase 1 and 2, thromboxane synthase, and prostacyclin synthase were determined. ResultsLeg blood flow during the initial 90 s of passive leg movement (242 ± 33 vs 441 ± 75 ml min−1, P = 0.03) and during reactive hyperaemia (423 ± 100 vs 1255 ± 175 ml min−1, P = 0.002) was lower in PAD than CON, whereas no significant difference was observed for leg blood flow during exercise (1490 ± 250 vs 1887 ± 349 ml min−1, P = 0.37). PAD had higher NADPH oxidase than CON (1.04 ± 0.19 vs 0.50 ± 0.06 AU, P = 0.02), with no differences for other enzymes. Leg blood flow during exercise was correlated with prostacyclin synthase (P = 0.001). ConclusionElevated NADPH oxidase indicates that oxidative stress may be a primary cause of low nitric oxide availability and impaired blood flow in PAD.

Increased RhoA/Rho-Kinase Activity and Markers of Endothelial Dysfunction in Young Adult Subjects with Metabolic Syndrome.

Metabolic syndrome, a chronic condition associated with higher risk of cardiovascular diseases, is increasingly prevalent in young adults. Dyslipidemia, proinflammatory cytokines, endothelial dysfunction signs, and RhoA/Rho-kinase (ROCK) activation are considered risk factors of cardiovascular diseases. The occurrence of these factors in young patients with metabolic syndrome but without type 2 diabetes or hypertension has not been fully studied. The objective of this study was to evaluate young subjects with enlarged waist circumference and dyslipidemia but without type 2 diabetes or hypertension,for markers associated with a higher risk of cardiovascular diseases. Thirty-two male patients aged 31 ± 1.3 years diagnosed with metabolic syndrome according to the National Cholesterol Education Program Adult Treatment Panel III guide for enlarged waist circumference, elevated triglycerides, and low HDL levels, but with blood pressure and fasting glucose within normal ranges, were evaluated for RhoA/ROCK activity in leukocytes, serum fatty acid methyl esters profile, proinflammatory cytokines, and oxidative stress markers in addition to thrombin generation and biochemical analysis. Age- and gender-matched healthy subjects were equivalently evaluated. Patients showed higher RhoA/ROCK activity, elevated levels of interleukin-6, soluble CD40L, monocyte chemoattractant protein, and high-sensitivity C-reactive protein (P < 0.001) as well as parameters of endogenous thrombin generation potential (P < 0.05) compared with healthy subjects. Increased thiobarbituric acid reactive substances, advanced oxidation protein product, and insulin levels and low nitric oxide biodisponibility (P < 0.001) were also found in patients as compared with controls. Palmitic acid was one of the saturated fatty acids found to be significantly elevated in patients compared with control subjects (P = 0.0087). Increased markers of cardiovascular risk are already present in young adults with metabolic syndrome but without type 2 diabetes or hypertension.