Reduced Nitric Oxide Synthesis Research Articles

Modulatory effect of olanzapine on neuronal nitric oxide synthase (nNOS) expression in the rat striatum.

Nitric oxide (NO) has been thought to be a novel factor involved in the mechanisms of mental disorders pathogenesis for quite some time. However, little is known about potential crosstalk between neuronal NO signaling and neuroleptics action. The present work was, therefore, focused on gene expression of neuronal NO synthase (nNOS) in the brains of rats chronically treated with olanzapine, an atypical antipsychotic drug. Studies were carried out on adult, male Sprague-Dawley rats that were divided into 2 groups: control and experimental animals treated with olanzapine (28-day-long intraperitoneal injection, at dose 5mg/kg daily). All individuals were killed under anesthesia and the whole brains excised. Immunohistochemical procedure was used for histological assessment of the whole brain, and for both descriptive and quantitative analysis of nNOS protein distribution in selected brain structures. Long-term treatment with olanzapine is reflected in different changes in the number of enzyme-expressing cells in the rat brain. Olanzapine decreased the number of nNOS-expressing cells and possibly reduced NO synthesis in the rat striatum. Olanzapine can be taken into account as a potential inhibitor of NO synthesis in the rat striatum.

Tetramerization of STAT5 regulates monocyte differentiation and the dextran sulfate sodium-induced colitis in mice.

In response to external stimuli during immune responses, monocytes can have multifaceted roles such as pathogen clearance and tissue repair. However, aberrant control of monocyte activation can result in chronic inflammation and subsequent tissue damage. Granulocyte-macrophage colony-stimulating factor (GM-CSF) induces monocyte differentiation into a heterogenous population of monocyte-derived dendritic cells (moDCs) and macrophages. However, the downstream molecular signals that dictate the differentiation of monocytes under pathological conditions is incompletely understood. We report here that the GM-CSF-induced STAT5 tetramerization is a critical determinate of monocyte fate and function. Monocytes require STAT5 tetramers to differentiate into moDCs. Conversely, the absence of STAT5 tetramers results in a switch to a functionally distinct monocyte-derived macrophage population. In the dextran sulfate sodium (DSS) model of colitis, STAT5 tetramer-deficient monocytes exacerbate disease severity. Mechanistically, GM-CSF signaling in STAT5 tetramer-deficient monocytes results in the overexpression of arginase I and a reduction in nitric oxide synthesis following stimulation with lipopolysaccharide. Correspondingly, the inhibition of arginase I activity and sustained supplementation of nitric oxide ameliorates the worsened colitis in STAT5 tetramer-deficient mice. This study suggests that STAT5 tetramers protect against severe intestinal inflammation through the regulation of arginine metabolism.

ENOS 894T allele can Contribute to Endothelial Dysfunction but not QT Interval Prolongation in Dialytic Patients

Background: Cardiovascular complications are common in chronic kidney disease (CKD) patients. Endothelial nitric oxide synthase (eNOS) is very important for the homeostasis of the cardiovascular system, and its gene polymorphism at position 894 (G>T) has not been investigated with QTc interval in patients on dialysis. Objective: This study evaluated the association of the 894G>T polymorphism with QTc prolongation and endothelial dysfunction risk in dialysis patients. Methods: Predialysis blood samples were collected for eNOS gene polymorphism, nitric oxide (NO), thiobarbituric acid reactive substances (TBARS), total antioxidant, asymmetric dimethylarginine (ADMA) and L-arginine, and 12-lead electrocardiograms were analyzed in these patients. Statistics were based on continuous and categorical variables using Fisher’s exact or Chi-square or one-way ANOVA or KruskalWallis tests. The results were considered significant when P < 0.05. Results: The study showed that the GG genotype was prevalent, with 54% of patients, followed by 41% GT and 6% TT, and the genotypic distribution was not associated with QTc prolongation. Furthermore, patients with the T allele showed increased ADMA, L-arginine and peroxidation lipid levels with reduced NO synthesis. Conclusion: Our study showed a lack of association between QTc interval and eNOS polymorphism; however, it was found that patients with the T allele had a greater risk of developing endothelial dysfunction by ADMA, which could contribute to future cardiovascular complications and worsening of CKD.

The Role of Reduced Polyamine Synthesis in Ischemic Stroke

Our aim was to determine the levels of arginine decarboxylase (ADC), ornithine decarboxylase (ODC) and agmatine, ornithine, nitric oxide (NO) and arginase activity in the acute and subacute phases of acute ischemic stroke (AIS), by revealing the possible role of these levels in neuroinflammation and neurogenesis that are postulated to begin after AIS. 35 patients diagnosed with AIS and 35 controls were included in the study. ADC, ODC, agmatinase and NO levels were determined by ELISA kits. Arginase activity and ornithine levels were measured spectrophotometrically. There was no statistically significant difference between the levels of agmatinase, ornithine and arginase activity on days 1 and 7 in the AIS group and the levels measured in the control group (p > 0.05). High ADC and low ODC and NO values were found in the patients compared to the controls in both acute and subacute periods (p < 0.05). It was shown that polyamine synthesis decreased in both acute and subacute periods after AIS; and this was due to the decrease in ODC levels, the rate-regulating step of the major pathway. This reduction may be associated with both the inadequate stimulation of neurogenesis and the immune system with reduced NO synthesis. polyamine synthesis enzymes, ornithine, ornitine decarboxylase, nitric oxide, arginine decarboxylase, agmatine, agmatinase, arginase activity, neuroinflammation, neurogenesis, immune system, acute ischemic stroke.

Abstract P127: Adverse Endothelial Cell Protein Expression is Associated With Type 1 Diabetes

Cardiovascular risk is increased in adults with type 1 diabetes (T1D), although the mechanism is poorly understood. Endothelial dysfunction may contribute to increased coronary artery calcium (CAC) in T1D, and CAC severity may be associated with reduced nitric oxide (NO) synthesis and/or increased inflammation. We tested the hypotheses that high CAC and T1D are associated with expression of endothelial NO synthase (eNOS, enzyme that produces NO), phosphorylated eNOS (p-eNOS), and NFκB (pro-inflammatory transcription factor). Participants were categorized as having high (&gt;80 Agatston units [AU]) or low CAC (&lt;15 AU), measured by CT scan. Protein expression was measured using quantitative immunofluorescence in endothelial cells harvested from 14 women with T1D (mean age 52±8 years; 7 high CAC); 13 men with T1D (mean age 58±7 years; 5 high CAC); 10 women without diabetes (mean age 61±4 years; 3 high CAC); and 10 men without diabetes (mean age 59±6 years; 4 high CAC). All analyses used ratios of endothelial cell protein expression to control human umbilical vein endothelial cell protein expression. Independent t-tests compared protein expression by sex and T1D. Linear regression tested the associations of high vs. low CAC and T1D with the 3 proteins of interest after adjustment for age and sex. In men, eNOS expression differed by T1D: men with T1D had reduced cytoplasmic (0.34 vs.0.52, p=0.03) and nuclear (0.44 vs. 0.60, p=0.02) eNOS expression compared with men without T1D, suggesting reduced NO synthesis. There were no differences in p-eNOS expression, indicating that eNOS levels were likely not due to differential activation. In women, NFκB expression similarly differed by T1D: women with T1D had higher cytoplasmic (0.81 vs. 0.63, p=0.01) and nuclear (0.99 vs. 0.75, p=0.01) NFκB expression compared with women without T1D, suggesting greater inflammation. After adjustment for age and sex, there was no significant association between high vs. low CAC in endothelial expression of any proteins. The data trend suggested that high vs. low CAC is associated with reduced cytoplasmic (0.36 vs. 0.47, p=0.06) and nuclear (0.48 vs. 0.55, p=0.12) eNOS expression. T1D trended in the same direction for both cytoplasmic (0.36 vs. 0.47, p=0.06) and nuclear (0.47 vs. 0.55, p=0.13) eNOS expression. High vs. low CAC also trended towards an association with greater cytoplasmic (0.72 vs. 0.66, p=0.21) and nuclear (0.85 vs. 0.80, p=0.40) NFκB expression, while T1D was significantly associated with greater cytoplasmic (0.76 vs. 0.63, p=0.006) and nuclear (0.92 vs. 0.73, p=0.003) NFκB expression. Neither CAC nor T1D was associated with p-eNOS expression in linear regression. In conclusion, eNOS and NFκB expression are potential mechanisms for increased CAC given their roles in reduced NO synthesis and increased inflammation, and may differ by sex in people with T1D. Regardless of high CAC, inflammation in T1D is increased.

Impaired l-arginine-nitric oxide pathway contributes to the pathogenesis of resistant hypertension.

The precise mechanisms underlying resistant hypertension remain elusive. Reduced nitric oxide (NO) bioavailability is frequently documented in chronic kidney disease, obesity, diabetes and advanced age, all of which are risk factors for resistant hypertension. Sympathetic overactivity and chronic activation of the renin-angiotensin system are salient features of resistant hypertension. Interestingly, recent data indicate that renal sympathetic overactivity can reduce the expression of neuronal nitric oxide synthase in the paraventricular nucleus. Reduced NO levels in the paraventricular nucleus can increase sympathetic outflow and this can create a vicious cycle contributing to resistant hypertension. Angiotensin II can reduce l-arginine transport and hence NO production. Reduced NO levels may reduce the formation of angiotensin 1-7 dampening the cardio-protective effects of the renin-angiotensin system contributing to resistant hypertension. In addition, interleukin-6 (IL-6) is demonstrated to be independently associated with resistant hypertension, and IL-6 can reduce NO synthesis. Despite this, NO levels have not been quantified in resistant hypertension. Findings from a small proof of concept study indicate that NO donors can reduce blood pressure in patients with resistant hypertension but more studies are required to validate these preliminary findings. In the present paper, we put forward the hypothesis that reduced NO bioavailability contributes substantially to the development of resistant hypertension.

Plasma from Patients with Rheumatoid Arthritis Reduces Nitric Oxide Synthesis and Induces Reactive Oxygen Species in A Cell-Based Biosensor.

Rheumatoid arthritis (RA) has been associated with a higher risk of developing cardiovascular (CV) diseases. It has been proposed that systemic inflammation plays a key role in premature atherosclerosis development, and is therefore crucial to determine whether systemic components from RA patients promotes endothelial cell-oxidative stress by affecting reactive oxygen species (ROS) and nitric-oxide (NO) production. The aim of this study was to evaluate whether plasma from RA patients impair NO synthesis and ROS production by using the cell-line ECV-304 as a biosensor. NO synthesis and ROS production were measured in cells incubated with plasma from 73 RA patients and 52 healthy volunteers by fluorimetry. In addition, traditional CV risk factors, inflammatory molecules and disease activity parameters were measured. Cells incubated with plasma from RA patients exhibited reduced NO synthesis and increased ROS production compared to healthy volunteers. Furthermore, the imbalance between NO synthesis and ROS generation in RA patients was not associated with traditional CV risk factors. Our data suggest that ECV-304 cells can be used as a biosensor of systemic inflammation-induced endothelial cell-oxidative stress. We propose that both NO and ROS production are potential biomarkers aimed at improving the current assessment of CV risk in RA.

NO-ergic Control of Blood Circulation in the Medulla Oblongata of Rats with Experimental Hemiparkinsonism under Prolonged Exposure to Continuous Light

The study was conducted on rats with unilateral damage to dopaminergic (DA) neurons in substantia nigra of the midbrain (experimental hemiparkinsonism). Degeneration of dopaminergic (DA) neurons was accompanied by hyperactivity of those neurons that remained intact and responded to apomorphine (Apo) test by rotational movements. Depending on the number of rotations, three groups of animals were defined. In the medulla oblongata of rats with unilateral damage to dopaminergic (DA) neurons, a significant increase in the activity of inducible NO-synthase (iNOS) was observed, while the activity of constitutive NO-synthase (cNOS) tended to decrease compared with that in control rats. An activation of neuronal NOsynthase (nNOS) in those rats by injections of L-arginine in the medullary nuclei was accompanied by weakening the hemodynamic effects compared to those in control rats. An exposure of animals to all-night light for three weeks was accompanied by increasing the number of damaged DA-ergic neurons in substantia nigra. At that, a significant decrease in cNOS activity in the medulla oblongata was observed, leading to the inhibition of de novo synthesis of nitric oxide (NO). The reduction of NO synthesis in the medulla oblongata neurons of rats with experimental hemiparkinsonism following their exposure to continuous light was also evidenced by the reduction of the amount of nitrite (NO2-) anion.

The opposing roles of NO and oxidative stress in cardiovascular disease

Nitric oxide (NO) plays a pivotal role in the maintenance of cardiovascular homeostasis. A reduction in the bioavailability of endogenous NO, manifest as a decrease in the production and/or impaired signaling, is associated with many cardiovascular diseases including hypertension, atherosclerosis, stroke and heart failure. There is substantial evidence that reactive oxygen species (ROS), generated predominantly from NADPH oxidases (Nox), are responsible for the reduced NO bioavailability in vascular and cardiac pathologies. ROS can compromise NO function via a direct inactivation of NO, together with a reduction in NO synthesis and oxidation of its receptor, soluble guanylyl cyclase. Whilst nitrovasodilators are administered to compensate for the ROS-mediated loss in NO bioactivity, their clinical utility is limited due to the development of tolerance and resistance and systemic hypotension. Moreover, efforts to directly scavenge ROS with antioxidants has had limited clinical efficacy. This review outlines the therapeutic utility of NO-based therapeutics in cardiovascular diseases and describes the source and impact of ROS in these pathologies, with particular focus on the interaction with NO. Future therapeutic approaches in the treatment of cardiovascular diseases are highlighted with a focus on nitroxyl (HNO) donors as an alternative to traditional NO donors and the development of novel Nox inhibitors.

NO-ERGIC CONTROL OF BLOOD CIRCULATION IN THE MEDULLA OBLONGATA OF RATS WITH EXPERIMENTAL HEMIPARKINSONIZM UNDER EXPOSURE TO CONTINUOUS LIGHT.

The study was conducted on rats with unilateral damage to dopaminergic (DA) neurons in substantia nigra of the midbrain (experimental hemiparkinsonism). Degeneration of dopaminergic (DA) neurons was accompanied by hyperactivity of those neurons that remained intact and responded to apomorphine (Apo) test by rotational movements. Depending on the number of rotations, three groups of animals were defined. In the medulla oblongata of rats with unilateral damage to dopaminergic (DA) neurons, a significant increase in the activity of inducible NO-synthase (iNOS) was observed, while the activity of constitutive NO-synthase (cNOS) tended to decrease compared with that in control rats. An activation of neuronal NO-synthase (nNOS) in those rats by injections of L-arginine in the medullary nuclei was accompanied by weakening of the hemodynamic effects compared to those in control rats. An exposure of animals to continuous light for three weeks was accompanied by increasing the number of damaged DA-ergic neurons in substantia nigra. At that, a significant decrease in cNOS activity in the medulla oblongata was observed, leading to the inhibition of de novo synthesis of nitric oxide (NO). The reduction of NO synthesis in the medulla oblongata neurons of rats with experimental hemiparkinsonism following their exposure to continuous light was also evidenced by the reduction.

Asymmetric Dimethylarginine correlates significantly with tumour necrosis alfa but not with brachial ankle pulse wave velocity in the T2DM-METS

Background: Endothelial dysfunction is an important phenomenon in the pathogenesis of atherosclerosis and is related to the derangements of nitric oxide (NO) synthase in the vessel wall. Asymmetric Dimethylarginine (ADMA) is an endogenous, competitive inhibitor of nitric oxide synthase and is induced by inflammatory cytokines of tumour necrosis factor (TNF)-a in vitro. Increased ADMA levels are associated with reduced NO synthesis as assessed by impaired endothelium-dependent vasodilatation. There is cause and effect relationship between endothelial dysfunction and vascular stiffening. Measurement of brachial ankle pulse wave velocity (baPWV) is simple and applicable for cardiovascular risk screening and as a marker for the severity of atherosclerotic vascular damage. Aim: To investigate the correlation between ADMA level with TNF-a and baPWV in the type-2 diabetes mellitus (T2DM)-Metabolic Syndrome (Mets) patients. Method: This is a cross sectional study with T2DM-Mets patients who came to the outpatient clinic of Soetomo Hospital Surabaya during January 2010 to December 2012. Subjects met the inclusion and exclusion criteria were measured their ADMA, TNF-a, and blood glucose levels in plasma. Brachial ankle pulse wave velocity (ba-PWV) was determined by using the V Serra-1000. The study was approved by the local Research Ethics Committee and subjects gave written informed consent. Results: Thirty-seven T2DM-Mets patients consisted of 15 (40.5%) males and 22 (59.5%) females who met inclusion and exclusion criteria were enrolled in this study. Their mean of age was 51 ± 5.2 years old, duration of illness was 16.49 ± 23.4 months, A1C level was 8.5 ± 0.9%, BMI was 26.7 ± 4.5 kg/m2, ADMA level was 0.572 ± 0.2 μmol/L, TNF-a level was 10.0 ± 16.5 pg/mL, and ba-PWV was 1,624.5 ± 295.5 cm/s. Spearman’s correlation analysis showed that ADMA level was significantly correlated with TNF-a level (p = 0.026; r = 0.366). However, no significant correlation found with ba-PWV (p = 0.134; r = 0.251). Conclusion: ADMA level is correlated with TNF-a level in this T2DM-MetS population.

Profiling nitric oxide metabolites in patients with idiopathic pulmonary arterial hypertension.

Intact nitric oxide (NO) signalling is critical to maintaining appropriate pulmonary vascular tone. NO bioavailability is reduced in patients with pulmonary arterial hypertension. This study aimed to examine the impact of NO plasma metabolites (NOx) relative to haemodynamic dysfunction and mortality in patients with idiopathic pulmonary arterial hypertension (IPAH).A total of 104 consecutive adult IPAH patients who had undergone genetic counselling when first diagnosed were enrolled in this prospective study.The median concentration of NOx (μmol·L-1) was significantly lower in IPAH patients compared with healthy subjects, and was decreased further in 19 carriers of the bone morphogenetic protein-receptor type-2 (BMPR2) mutation compared to non-carriers. Reduced concentrations of NOx were correlated with mean pulmonary arterial pressure (mPAP), pulmonary vascular resistance (PVR) and cardiac output. Compared with higher baseline NOx concentrations, patients with a NOx concentration of ≤10 μmol·L-1 had a markedly worse survival. After adjustment for clinical features, a BMPR2 mutation and haemodynamics, a lower NOx level remained an increased risk of mortality.Patients with IPAH had lower levels of plasma NOx, which correlated inversely with mPAP, PVR and survival. Plasma NOx may be an important biomarker and prognostic indicator, suggesting that reduced NO synthesis contributes to the pathogenesis of IPAH.

Increased asymmetric dimethylarginine levels in severe transient tachypnea of the newborn.

Nitric oxide (NO) is synthesized by NO synthase (NOS), and asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of NOS. We aimed to investigate l-arginine and ADMA levels in transient tachypnea of the newborn (TTN) and their relationship with systolic pulmonary artery pressure (PAP) and disease severity. Infants born at ⩾35 weeks gestational age with clinical signs and chest X-ray findings consistent with TTN were enrolled; controls were recruited at the same time. l-arginine and ADMA levels were measured at 12 to 24 h (first samples) and at 48 to 72 h (second samples). Systolic PAP was evaluated on the second day. Patients were divided according to the duration of tachypnea and designated as group A (duration ⩽72 h) and group B (duration >72 h). In the first samples, the ADMA levels were significantly higher in patients with TTN compared with controls (P<0.001). In the second samples, the ADMA levels were significantly higher in group B compared with that in group A (P=0.019). In group A patients, the second ADMA levels were significantly lower compared with that in the first samples (P<0.001), whereas the second ADMA levels remained unchanged compared with the first samples in group B. Systolic PAP values were significantly higher in group B compared with that in group A patients (P=0.033). Increased ADMA concentration may reduce NO synthesis, leading to increased PAP and thus longer duration of tachypnea.

The relationship of nitric oxide synthesis capacity, oxidative stress, and albumin-to-creatinine ratio in black and white men: the SABPA study.

Inadequate substrate availability and increased nitric oxide synthase inhibitor levels attenuate nitric oxide (NO) synthesis, whereas increased vascular oxidative stress may lead to inactivation of NO. We compared markers of NO synthesis capacity and oxidative stress in a bi-ethnic male population. Inter-relationships of ambulatory blood pressure and urinary albumin-to-creatinine ratio with NO synthesis capacity and oxidative stress markers were investigated. NO synthesis capacity markers (L-arginine, asymmetric dimethylarginine (ADMA), and symmetric dimethylarginine (SDMA)) and oxidative stress markers (serum peroxides, total glutathione, glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD), and catalase) were measured. Black men displayed higher blood pressure and albumin-to-creatinine ratio (all p<0.001), while NO synthesis capacity was more favorable (higher L-arginine and lower ADMA (p≤0.003)). Antioxidant enzyme activities were similar except for the redox status markers (GR activity and GR/GPx ratio), which were upregulated in black men (p<0.001). In black men, ADMA was inversely related to GPx activity (R (2)=0.15; β=-0.20; p=0.050) and GPx/SOD ratio (R (2)=0.24; β=-0.37; p<0.001), but none of these markers related to blood pressure or albumin-to-creatinine ratio. In white men, albumin-to-creatinine ratio was positively associated with ADMA (R (2)=0.18; β=0.39; p<0.001) while ADMA was inversely related to GR activity (R (2)=0.26; β=-0.29; p=0.002) and GR/GPx ratio (R (2)=0.25; β=-0.28; p=0.003). Black men with elevated blood pressure and albumin-to-creatinine ratio displayed a favorable NO synthesis capacity. This may be counteracted by increased inactivation of NO, although it was not linked to vascular or renal phenotypes. In white men, reduced NO synthesis capacity may lower NO bio-availability, thereby influencing the albumin-to-creatinine ratio.

Dimethylarginine dimethylaminohydrolase 1 is involved in spinal nociceptive plasticity

Activation of neuronal nitric oxide synthase, and consequent production of nitric oxide (NO), contributes to spinal hyperexcitability and enhanced pain sensation. All NOS isoforms are inhibited endogenously by asymmetric dimethylarginine, which itself is metabolised by dimethylarginine dimethylaminohydrolase (DDAH). Inhibition of DDAH can indirectly attenuate NO production by elevating asymmetric dimethylarginine concentrations. Here, we show that the DDAH-1 isoform is constitutively active in the nervous system, specifically in the spinal dorsal horn. DDAH-1 was found to be expressed in sensory neurons within both the dorsal root ganglia and spinal dorsal horn; L-291 (NG-[2-Methoxyethyl]-L-arginine methyl ester), a DDAH-1 inhibitor, reduced NO synthesis in cultured dorsal root ganglia neurons. Spinal application of L-291 decreased N-methyl-D-aspartate-dependent postdischarge and windup of dorsal horn sensory neurons--2 measures of spinal hyperexcitability. Finally, spinal application of L-291 reduced both neuronal and behavioral measures of formalin-induced central sensitization. Thus, DDAH-1 may be a potential therapeutic target in neuronal disorders, such as chronic pain, where elevated NO is a contributing factor.

Substrate-dependent nitric oxide synthesis by secreted endoplasmic reticulum aminopeptidase 1 in macrophages.

In this study, we examined the role of aminopeptidases with reference to endoplasmic reticulum aminopeptidase 1 (ERAP1) in nitric oxide (NO) synthesis employing murine macrophage cell line RAW264.7 cells activated by lipopolysaccharide (LPS) and interferon (IFN)-γ and LPS-activated peritoneal macrophages derived from ERAP1 knockout mouse. When NO synthesis was measured in the presence of peptides having N-terminal Arg, comparative NO synthesis was seen with that measured in the presence of Arg. In the presence of an aminopeptidase inhibitor amastatin, NO synthesis in activated RAW264.7 cells was significantly decreased. These results suggest that aminopeptidases are involved in the NO synthesis in activated RAW264.7 cells. Subsequently, significant reduction of NO synthesis was observed in ERAP1 knockdown cells compared with wild-type cells. This reduction was rescued by exogenously added ERAP1. Furthermore, when peritoneal macrophages prepared from ERAP1 knockout mouse were employed, reduction of NO synthesis in knockout mouse macrophages was also attributable to ERAP1. In the presence of amastatin, further reduction was observed in knockout mouse-derived macrophages. Taken together, these results suggest that several aminopeptidases play important roles in the maximum synthesis of NO in activated macrophages in a substrate peptide-dependent manner and ERAP1 is one of the aminopeptidases involved in the NO synthesis.

Liver plays a central role in asymmetric dimethylarginine-mediated organ injury

Asymmetric-dimethylarginine (ADMA) competes with L-arginine for each of the three isoforms of nitric oxide synthase: endothelial; neuronal; inducible. ADMA is synthesized by protein methyltransferases followed by proteolytic degradation. ADMA is metabolized to citrulline and dimethylamine, by dimethylarginine dimethylaminohydrolase (DDAH) and enters cells through cationic amino-acid transporters extensively expressed in the liver. The liver plays a crucial role in ADMA metabolism by DDAH-1 and, as has been recently demonstrated, it is also responsible for ADMA biliary excretion. A correlation has been demonstrated between plasma ADMA levels and the degree of hepatic dysfunction in patients suffering from liver diseases with varying aetiologies: plasma ADMA levels are increased in patients with liver cirrhosis, alcoholic hepatitis and acute liver failure. The mechanism by which liver dysfunction results in raised ADMA concentrations is probably due to impaired activity of DDAH due to severe inflammation, oxidative stress, and direct damage to DDAH. High plasma ADMA levels are also relevant as they are associated with the onset of multi-organ failure (MOF). Increased plasma concentration of ADMA was identified as an independent risk factor for MOF in critically-ill patients causing enhanced Intensive Care Unit mortality: a significant reduction in nitric oxide synthesis, leading to malperfusion in various organs, eventually culminating in multi organs dysfunction.

Endothelial dysfunction: the link between homocysteine and hydrogen sulfide.

High level of homocysteine (hyperhomocysteinemia, HHcy) is associated with increased risk for vascular disease. Evidence for this emerges from epidemiological studies which show that HHcy is associated with premature peripheral, coronary artery and cerebrovascular disease independent of other risk factors. Possible mechanisms by which homocysteine causes vascular injury include endothelial injury, DNA dysfunction, proliferation of smooth muscle cells, increased oxidative stress, reduced activity of glutathione peroxidase and promoting inflammation. HHcy has been shown to cause direct damage to endothelial cells both in vitro and in vivo. Clinically, this manifests as impaired flow-mediated vasodilation and is mainly due to a reduction in nitric oxide synthesis and bioavailability. The effect of impaired nitric oxide release can in turn trigger and potentiate atherothrombogenesis and oxidative stress. Endothelial damage is a crucial aspect of atherosclerosis and precedes overt manifestation of disease. In addition, endothelial dysfunction is also associated with hypertension, diabetes, ischemia reperfusion injury and neurodegenerative diseases. Homocysteine is a precursor of hydrogen sulfide (H2S) which is formed by transulfuration process catalyzed by the enzymes, cystathionine β-synthase and cystathionine γ-lyase. H2S is a gasotransmitter that has emerged recently as a novel mediator in cardiovascular homeostasis. As a potent vasodilator, it plays several roles which include regulation of vessel diameter, protection of endothelium from redox stress, ischemia reperfusion injury and chronic inflammation. However, the precise mechanism by which it mediates these beneficial effects is complex and still remains unclear. Current evidence indicates H2S modulates cellular functions by a variety of intracellular signaling processes. In this review, we summarize the mechanisms of HHcy-induced endothelial dysfunction and the metabolism and physiological functions of H2S as a protective agent.

Citrulline ingestion did not improve the age‐associated reduction in nitric oxide synthesis (698.6)

A reduction in nitric oxide (NO) bioavailability that impairs peripheral blood flow regulation is a natural consequence of aging. Therefore, we investigated the effect of ingestion of citrulline, the precursor of arginine, on de novo arginine and NO synthesis at baseline and during citrulline ingestion. A total of 3g of citrulline was ingested every 15 min for 3 h. A 6‐h stable isotope infusion protocol was performed in young (n=8, 27 ± 1.1 y) and elderly subjects (n=8, 76.6 ± 3.1 y) using primed constant infusions of ring‐ L‐[guanidine‐15N2]arginine, L‐[ureido‐13C,5,5‐2H2]citrulline,and ring‐[2H5]phenylalanine. Basal NO synthesis [0.32 ± 0.03 vs. 0.47 ± 0.05 μmol/kg ffm/h; p &lt; 0.02], but not de novo arginine [12.5 ± 1.60 vs. 16.4 ± 1.47 μmol/kg ffm/h], was significantly lower in the elderly vs. the young. Citrulline ingestion increased de novo arginine synthesis in both the elderly and the young, but to a greater extent in the young [55.4 ± 9.64 vs. 72.3 ± 5.96 μmol/kg ffm/h; p &lt; 0.01]. Citrulline ingestion increased NO synthesis in the young [3.47 ± 0.65 μmol/kg ffm/h; p &lt; 0.01], but not in the elderly [1.26 ± 0.42 μmol/kg ffm/h; p = 0.08]. We conclude that aging reduces NO synthesis. This decrease in NO synthesis cannot be attributed to limited availability of citrulline but to the inability to synthesize NO via NOS.Grant Funding Source: NIH/NIA P30 AG028718

Effects of fructose-1, 6-diphosphate pretreatment on lung injury induced by hepatic cold ischemia-reperfusion in rats

Objective To evaluate the effects of fructose-1 ,6-diphosphate （FDP） pretreatment on lung injury induced by hepatic cold liver ischemia-reperfusion in rats .Methods Eighteen healthy male Sprague-Dawley rats were randomly divided into 3 groups （n= 6 each） using a random number table ：sham operation group （S group） ,hepatic cold liver ischemia-reperfusion model group （M group） ,and FDP pretreatment group （FP group） . The animals were anesthetized with intraperitoneal chloral hydrate and kept spontaneous breathing .Laparotomy was performed ,and the related blood vessels were only isolated in group S .Hepatic cold ischemia-reperfusion was induced in M and FP groups .In FP group ,FDP 250 mg/kg was injected via the caudal vein at 15 min before skin incision .At 6 h of reperfusion ,the bronchoalveolar lavage fluid （BALF） was collected to detect the levels of tumor necrosis factor-α（TNF-α） ,interleukin-10 （IL-10） and nitric oxide （NO） by ELISA .Lungs were removed for microscopic examination of the pathological changes by light microscopy .Real-time PCR was used to detect the expression of iNOS mRNA .Results Compared with S group , the levels of TNF-α and NO in BALF were significantly increased , the expression of iNOS mRNA was up-regulated , and the level of IL-10 in BALF was decreased in M and FP groups （ P〈0.05 ） .Compared with M group ,the levels of TNF-αand NO in BALF were significantly decreased ,the expression of iNOS mRNA was down-regulated ,and the level of IL-10 in BALF was＆nbsp;increased in FP group （ P〈0.05 ） .The pathological changes of lungs were significantly attenuated in FP group as compared with M group .Conclusion FDP pretreatment can obviously attenuate lung injury induced by hepatic cold ischemia-reperfusion in rats ,and inhibition of iNOS expression ,reduction of NO synthesis ,and decrease in inflammatory responses are involved in the mechanism . Key words: Fructose; Liver; Lung; Reperfusion injury