Biological Activity Of Nitric Oxide Research Articles

De-nitrosylation Coordinates Appressorium Function for Infection of the Rice Blast Fungus.

As a signaling molecule, nitric oxide (NO) regulates the development and stress response in different organisms. The major biological activity of NO is protein S-nitrosylation, whose function in fungi remains largely unclear. Here, it is found in the rice blast fungus Magnaporthe oryzae, de-nitrosylation process is essential for functional appressorium formation during infection. Nitrosative stress caused by excessive accumulation of NO is harmful for fungal infection. While the S-nitrosoglutathione reductase GSNOR-mediated de-nitrosylation removes excess NO toxicity during appressorium formation to promote infection. Through an indoTMT switch labeling proteomics technique, 741 S-nitrosylation sites in 483 proteins are identified. Key appressorial proteins, such as Mgb1, MagB, Sps1, Cdc42, and septins, are activated by GSNOR through de-nitrosylation. Removing S-nitrosylation sites of above proteins is essential for proper protein structure and appressorial function. Therefore, GSNOR-mediated de-nitrosylation is an essential regulator for appressorium formation. It is also shown that breaking NO homeostasis by NO donors, NO scavengers, as well as chemical inhibitor of GSNOR, shall be effective methods for fungal disease control.

PFKFB3 Mediated Changes in ROS/NO Balance Contribute to Endothelial Dysfunction in Obesity

Obesity is a strong risk factor for cardiovascular disease, in large part due to an altered metabolic state. One of the earliest consequences of cardiovascular dysfunction in obesity is the loss of endothelial function and impaired nitric oxide (NO) signaling. In blood vessels, NO is synthesized by endothelial nitric oxide synthase (eNOS). The biological actions of NO can be compromised by rapid inactivation by reactive oxygen species (ROS) such as superoxide that is mainly generated by NADPH oxidases (NOXs). However, the signals coordinating the increased superoxide production leading to loss of NO with altered metabolism remain unknown.We have found that the expression of 6‐phosphofructo‐2‐kinase/fructose‐2, 6‐biphosphatase 3 (PFKFB3), which is a key glycolytic regulatory enzyme, is significantly increased in endothelial cells of obese animals. Overexpression of PFKFB3 within the endothelium of blood vessels is accompanied by the loss of vascular relaxation and in decreased NO production as a result of increased eNOS phosphorylation on the inhibitory T495 residue. PFKFB3 overexpression also blunted Akt‐S473 phosphorylation, reducing stimulus‐dependent phosphorylation and activation of eNOS at S1177. While PFKFB3 exerted negative effects on NO signaling, it increased the activity and mRNA levels of NOX1, a major contributor to endothelial dysfunction. Further, we have also found that pharmacological inhibition of NOX1 attenuates the decreased vasodilator responses in PFKFB3 overexpressing aortic rings.These results demonstrate a novel functional relationship between endothelial metabolism, ROS and NO production that may contribute to endothelial dysfunction in obesity.

Electrochemical evidence of nitrate release from the nitrooxy compound 4-((nitrooxy) methyl)-3-nitrobenzoic acid and its antinociceptive and anti-inflammatory activities in mice

Considering the many biological activities of nitric oxide (NO), some lines of research focused on the modulation of these activities through the provision of this mediator by designing and synthesizing compounds coupled with an NO donor group. Thus, the objectives of the present study were to carry out an electrochemical investigation of the nitrooxy compound 4-((nitrooxy) methyl)-3-nitrobenzoic acid (1) and evaluate its activities and putative mechanisms in experimental models of pain and inflammation. Voltammetric studies performed in aprotic medium (mimetic of membranes) showed important electrochemical reduction mechanisms: nitroaromatic reduction, self-protonation, and finally reductive elimination, which leads to nitrate release. Systemic administration of the nitrooxy compound (1) inhibited the nociceptive response induced by heat and the tactile hypersensitivity and paw edema induced by carrageenan in mice. The activities in the models of inflammatory pain and edema were associated with reduced neutrophil recruitment and production of inflammatory cytokines, such as interleukin (IL)-1β, IL-6, tumor necrosis factor-α and CXCL-1, and increased production of IL-10. Concluding, electrochemical analysis revealed unequivocally that electron transfer at the nitro group of the nitrooxy compound (1) results in the cleavage of the organic nitrate, potentially resulting in the generation of NO. This electrochemical mechanism may be compared to a biochemical electron-transfer mediated nitrate release that, by appropriate in vivo bioreduction (enzymatic or not) would lead to NO production. Compound (1) exhibits activities in models of inflammatory pain and edema that may be due to reduced recruitment of neutrophils and production of inflammatory cytokines and increased production of IL-10. These results reinforce the interest in the investigation of NO donor compounds as candidates for analgesic and anti-inflammatory drugs.

Bifunctional polymer brush-grafted coronary stent for anticoagulation and endothelialization

At present, cardiovascular stent intervention faces clinical complications such as delayed endothelialization, late thrombosis and restenosis after implantation. In this work, a kind of bifunctional polymer brush-grafted coronary stent with anticoagulant and endothelial functions was developed. First, a block copolymer brush with zwitterionic structure consisting of sulfoethyl methacrylate (SBMA) and glycidyl methacrylate (GMA) was surface-induced grafted onto the surface of bare metal coronary stent by atom transfer radical polymerization. The diethylenetriamine NONOate (DETA NONOate), acted as nitric oxide (NO) donor to promote endothelialization, was then attached to polyglycidyl methacrylate (PGMA) brush by a reactive epoxy group to produce NO. The process of chemical modification and the release behavior of NO were characterized in detail. Moreover, the results of anticoagulant test, cytotoxicity test, endothelial cells (ECs) proliferation test and animal experiment of this bifunctional polymer brush-grafted coronary stent we proposed indicate that the zwitterion modified and NO supplied bifunctional coatings has good anticoagulant property, no cytotoxicity and significant endothelialization effect. This work opens the door to combine biological activity of NO and anticoagulant effect of zwitterions, which has great potential to address post-operative side effects associated with restenosis and late stent thrombosis.

Dipyridamole Potentiates the Endothelium-Dependent and -Independent Vasomotion in Isolated Human Small Arteries.

BACKGROUND: To investigate the effects of dipyridamole, a drug with phosphodiesterase-, adenosine reuptake-inhibiting, and prostacyclin-stimulating activity on the biological actions of nitric oxide, 30 norepinephrine-precontracted subcutaneous arterioles were prepared from specimens removed during surgery. METHODS AND RESULTS: Specimens were mounted on a myograph and relaxes through either acetylcholine, a muscarinic agonist that stimulates endothelial nitric oxide production, or sodium nitroprusside, an endothelium-independent vasodilator. Studies were performed under control conditions and after dipyridamole which potentiated in a concentration-dependent manner the vasorelaxation induced both by acetylcholine and sodium nitroprusside, indicating an endothelium-independent mechanism of action. The contribution of nitric oxide to the relaxation produced by acetylcholine was confirmed by N-monomethyl-L-arginine, a nitric oxide synthase inhibitor. In contrast, indomethacin, a cyclo-oxygenase inhibitor, was ineffective, indicating that prostacyclin stimulation could not explain the effect of dipyridamole. CGS 21680 C, an A(2)-selective adenosine receptor agonist insensitive to tissue deaminase, did not influence the relaxations induced by acetylcholine, suggesting that interference with adenosine metabolism was not implicated in the potentiating action of dipyridamole. CONCLUSIONS: Dipyridamole potentiated the vasorelaxing effect of acetylcholine and sodium nitroprusside in human subcutaneous arterioles; neither prostacyclin stimulation nor A(2) adenosine receptor stimulation could explain this effect. The data are consistent with an increase in intracellular cyclic 3' 5'-guanosine monophosphate levels secondary to the phosphodiesterase-inhibiting properties of the drug.

Novel roles of PFKFB3 in mediating endothelial dysfunction in obesity

Obesity is a major and growing health problem in the US with over 93 million afflicted. Although, obesity itself is not considered a disease, it is a strong risk factor for cardiovascular disease, in large part due to an altered metabolic state. The relationship between metabolic dysfunction and impaired cardiovascular health remain incompletely understood.One of the earliest consequences of cardiovascular dysfunction in obesity is the loss of endothelial function and impaired nitric oxide (NO) signaling. NO is best known as the mediator of endothelium‐dependent relaxation, but it is also vital for maintaining blood vessel homeostasis through inhibition of leukocyte adhesion and smooth muscle cell proliferation. In blood vessels, NO is synthesized by endothelial nitric oxide synthase (eNOS) and dysregulation of eNOS has been shown to occur through changes in its expression and post‐translational phosphorylation. The biological actions of NO can be further compromised by rapid inactivation by reactive oxygen species (ROS) such as superoxide that are generated by NADPH oxidases (NOX) as well as numerous other enzyme systems.Our laboratory has previously shown that an imbalance between NO and superoxide‐generating pathways is responsible for the loss of endothelial function in obesity. However, the signals coordinating the loss of NO and increased superoxide with altered metabolism remain poorly defined.In our studies we have found that expression of the glycolytic enzyme 6‐phosphofructo‐2‐kinase/fructose‐2,6‐biphosphatase 3 (PFKFB3) is significantly increased in endothelial cells from obese animals. PFKFB3 is responsible for fructose‐2,6‐bisphosphate generation in vascular cells which is a potent allosteric activator of phosphofructokinase‐1 (PFK‐1), one of the rate‐limiting enzymes of glycolysis. We have also found that adenovirus‐mediated expression of PFKFB3 within the endothelium of mouse blood vessels is accompanied by the loss of endothelium‐dependent relaxation.In addition to its effects on blood vessels, overexpression of PFKFB3 in cultured cells resulted in decreased NO production as a result of increased eNOS phosphorylation on the well‐known inhibitory PKC phosphorylation site, threonine 495 (T495). Furthermore, PFKFB3 overexpression blunted Akt‐S473 phosphorylation, reducing stimulus‐dependent phosphorylation of eNOS at S1177. In addition to its negative effect on NO signaling overexpression of PFKFB3 increased the activity and mRNA level of NOX1, a major contributor to endothelial dysfunction in obesity.These results demonstrate a novel functional relationship between endothelial cell metabolism, nitric oxide and ROS production that may contribute to endothelial dysfunction in obesity.Support or Funding Information1R01HL147159‐01

In Cellullo and ex Vivo Availability of a Yellowish-Green-Light-Controllable NO Releaser.

Spatiotemporally controllable nitric oxide (NO) releasers are very attractive chemical tools for investigating the biological activities of NO, which is involved in the regulation of vasodilation, neurotransmission, and immune responses. We previously developed an easily synthesized, yellowish-green-light-controllable NO releaser, NO-Rosa5, and characterized its photoredox reaction mechanism. Here, we aimed to establish the biological applicability of NO-Rosa5 for in cellullo and ex vivo experiments. We successfully demonstrated yellowish-green-light-controlled NO release in HEK293T cells in vitro, as well as photomanipulation of the rat aorta response to NO in an ex vivo system. Furthermore, NO-Rosa5 showed lower toxicity than NOBL-1, a previously reported blue-light-controllable NO releaser, as determined by tetrazolium salt cell viability assay. Overall, our results indicate that NO-Rosa5 is a biocompatible, photocontrollable NO releaser with low toxicity and potentially broad applicability.

Synthesis, characterization and cytotoxicity of S-nitroso-mercaptosuccinic acid-containing alginate/chitosan nanoparticles

Nitric oxide (NO) is an endogenous free radical, which plays key roles in several biological processes including vasodilation, neurotransmission, inhibition of platelet adhesion, cytotoxicity against pathogens, wound healing, and defense against cancer. Due to the relative instability of NO in vivo (half-life of ca. 0.5 seconds), there is an increasing interest in the development of low molecular weight NO donors, such as S-nitrosothiols (RSNOs), which are able to prolong and preserve the biological activities of NO in vivo. In order to enhance the sustained NO release in several biomedical applications, RSNOs have been successfully allied to nanomaterials. In this context, this work describes the synthesis and characterization of the NO donor S-nitroso-mercaptosuccinic acid (S-nitroso-MSA), which belongs to the class of RSNOs, and its incorporation in polymeric biodegradable nanoparticles composed by alginate/chitosan. First, chitosan nanoparticles were obtained by gelation process with sodium tripolyphosphate (TPP), followed by the addition of the alginate layer, to enhance the nanoparticle protection. The obtained nanoparticles presented a hydrodynamic diameter of 343 ± 38 nm, polydispersity index (PDI) of 0.36 ± 0.1, and zeta potential of − 30.3 ± 0.4 mV, indicating their thermal stability in aqueous suspension. The negative zeta potential value was assigned to the presence of alginate chains on the surface of chitosan/TPP nanoparticles. The encapsulation efficiency of the NO donor into the polymeric nanoparticles was found to be 98 ± 0.2%. The high encapsulation efficiency value was attributed to the positive interactions between the NO donor and the polymeric content of the nanoparticles. Kinetics of NO release from the nanoparticles revealed a spontaneous and sustained release of therapeutic amounts of NO, for several hours under physiological temperature. The incubation of NO-releasing alginate/chitosan nanoparticles with human hepatocellular carcinoma (HepG2) cell line revealed a concentration-dependent toxicity. These results point to the promising uses of NO-releasing alginate/chitosan nanoparticles for anti-cancer chemotherapy.

The effects of sildenafil citrate on some biochemical and hematological parameters in alloxan-induced diabetes (Type I) in male albino rats

Abstract Background: Sildenafil citrate is a phosphodiesterase type 5 (PDE5) inhibitor, which increases cyclic guanosine monophosphate (cGMP). It is well established that the pathogenesis of diabetic mellitus is associated with abnormalities of nitric oxide (NO) generation. Many of the biological actions of NO are mediated by cGMP, which is rapidly degraded by phosphodiesterases. Aim: The Aim of this study is to evaluate the effect of sildenafil citrate on some biochemical and hematological parameters in alloxan-induced diabetes in male albino rat. Methods: Diabetes was induced after an intraperitoneal (i.p.) injection of a single dose of alloxan (150 mg/kg body weight). Sildenafil was administered daily at a dose of (10 mg/kg body weigh) via oral gavages during a period of 14 days to normal healthy rats as well as diabetic rats. Blood samples were collected after 7 and 14 days of treatment in all experimental groups. Results: The present investigation showed that alloxan injection induced significant elevations in serum glucose level, the percent of glycohemoglobin (HbA1c) and serum malondialdehyed (MDA) level. Moreover, alloxan administration resulted in prolongation of prothrombin time (PT), activated partial thromboplastin time (APTT), as well as increase in plasma fibrinogen level. Meanwhile, protein C and protein S concentrations were significantly decreased due to alloxan treatment. Oral administration of sildenafil to diabetic rats improved most diabetic complications including hyperglycemia, oxidative stress, hemoglycation and hyperfibrinogenemia induced by alloxan. The modulatory effects of sildenafil obtained herein were partial, but significant and more pronounced when administered for14 days. In addition, sildenafil administration to normal healthy rats was found to have no effect on the studied parameters. Conclusion: The present study showed that sildenafil may have beneficial effects against some diabetic complications. The mechanism by which sildenafil citrate exerts its effect on the studied parameters might be attributed to increased cGMP which is brought out by its phosphodiesterase inhibitory action and this is discussed in details.

Conventional and Unconventional Mechanisms for Soluble Guanylyl Cyclase Signaling.

Soluble guanylyl cyclase (sGC) is the principal enzyme in mediating the biological actions of nitric oxide. On activation, sGC converts guanosine triphosphate to guanosine 3',5'-cyclic monophosphate (cGMP), which mediates diverse physiological processes including vasodilation, platelet aggregation, and myocardial functions predominantly by acting on cGMP-dependent protein kinases. Cyclic GMP has long been considered as the sole second messenger for sGC action. However, emerging evidence suggests that, in addition to cGMP, other nucleoside 3',5'-cyclic monophosphates (cNMPs) are synthesized by sGC in response to nitric oxide stimulation, and some of these nucleoside 3',5'-cyclic monophosphates are involved in various physiological activities. For example, inosine 3',5'-cyclic monophosphate synthesized by sGC may play a critical role in hypoxic augmentation of vasoconstriction. The involvement of cytidine 3',5'-cyclic monophosphate and uridine 3',5'-cyclic monophosphate in certain cardiovascular activities is also implicated.

Aliskiren improves endothelium-dependent relaxation of thoracic aorta by activating PI3K/Akt/eNOS signal pathway in SHR.

Aliskiren, a direct renin blocker, has been approved for the treatment of hypertension. However, the potential role of aliskiren on vascular endothelial function in spontaneously hypertensive rats (SHR) remains unclear. In the present study, male SHRs at 12weeks of age were orally administrated 30mg/kg per day or 60mg/kg per day aliskiren. After a 4-week treatment, aliskiren showed a significant effect on the reduction of blood pressure at a dosage of 60mg/kg per day, but not of 30mg/kg per day. Moreover, both dosages of aliskiren improved endothelium-dependent relaxation, reduced dihydroethidium fluorescence intensity, decreased level of malondialdehyde but heightened total antioxidant capacity and superoxide dismutase activity in thoracic aorta in SHR. Aliskiren also markedly increased expression of p85α, an important subunit of phosphatidylinositol 3 kinase (PI3K), enhancing phosphorylation of protein kinase B (Akt) at Ser473 and endothelial nitric oxide synthase (eNOS) at Ser1177, as well as cyclic guanosine-3'5'-monophosphate (cGMP, a sensitive index of biological activity of nitric oxide) concentration. Furthermore, both anti-oxidative and endothelium protective effects of aliskiren were diminished when PI3K was inhibited invivo. The data presented here indicates that, aliskiren improves endothelium-dependent relaxation of thoracic aorta in SHR, predominantly through attenuating oxidative stress and activation of the PI3K/Akt/eNOS pathway. These data might propose novel strategies to prevent and improve vascular endothelial dysfunction.

The effect of ascorbic acid on the efficiency of cytotoxic therapy with cytostatic drugs in combination with NaNO3 and hydroxamic acids

The present investigation is based on the paradoxical dichotomy of the biological action of nitric oxide (NO). Depending on its concentration in the organism, it can have therapeutic or pathogenic effect. It was shown that during the treatment with cisplatin (cPt) or cyclophospamide (CP), adjunct therapy with ascorbic acid (AA) known to increase the NO content in the organism extended the average lifespan (ALS) by 20—30% as compared to cytostatic monotherapy. Combination of cytostatics with sodium nitrate extended ALS by 20—38% depending on the concentration of the nitrate. The efficiency of cytotoxic therapy in combination with two chemo sensitizers (NaNO3 and hydroxamic acid) also depends on the concentration of injected NaNO3. Addition of AA to these combinations extended ALS by 16%, or inhibited it by 40%. Variation of the dose of injected NaNO3 that is a NO donor in combination with endogenous NO stimulated with AA controls the action of cytostatics and combinations thereof.

Transcriptional and Posttranslational Regulation of eNOS in the Endothelium.

Nitric oxide (NO) is a highly reactive free radical gas and these unique properties have been adapted for a surprising number of biological roles. In neurons, NO functions as a neurotransmitter; in immune cells, NO contributes to host defense; and in endothelial cells, NO is a major regulator of blood vessel homeostasis. In the vasculature, NO is synthesized on demand by a specific enzyme, endothelial nitric oxide synthase (eNOS) that is uniquely expressed in the endothelial cells that form the interface between the circulating blood and the various tissues of the body. NO regulates endothelial and blood vessel function via two distinct pathways, the activation of soluble guanylate cyclase and cGMP-dependent signaling and the S-nitrosylation of proteins with reactive thiols (S-nitrosylation). The chemical properties of NO also serve to reduce oxidation and regulate mitochondrial function. Reduced synthesis and/or compromised biological activity of NO precede the development of cardiovascular disease and this has generated a high level of interest in the mechanisms controlling the synthesis and fate of NO in the endothelium. The amount of NO produced results from the expression level of eNOS, which is regulated at the transcriptional and posttranscriptional levels as well as the acute posttranslational regulation of eNOS. The goal of this chapter is to highlight and integrate past and current knowledge of the mechanisms regulating eNOS expression in the endothelium and the posttranslational mechanisms regulating eNOS activity in both health and disease.

Why is nitric oxide important for our brain?

The freely diffusible gaseous compound nitric oxide (NO) has been shown to be an important messenger in many organ systems throughout the body, and particularly in the central nervous system (CNS). The importance of NO as an intermediary in cell communication in the brain is highlighted by the fact that the excitatory amino acid glutamate, the most abundant CNS neurotransmitter, is an initiator of the reaction that forms NO. Because of its numerous physiological and pathophysiological roles, the impact of NO on clinical medicine is developing. NO can act as a "double-edged sword" and it has been demonstrated that clarification of the dual effect of NO might have implications for clinical medicine, and could lead to the emergence of therapeutic opportunities. Accordingly, NO was proclaimed "Mole cule of the Year" in 1992 by the journal Science, while discovery of the pathways and roles of NO was acknowledged with the Nobel Prize in 1998. Additionally, the ubiquity of NO in the CNS implies that drugs designed to modify the biological activity of NO may have distinct effects. Thus, further clinical applications of NO, of its analogs or of newly developed NOS inhibitors are forthcoming. The therapeutic challenge would be to succeed in manipulating the NO pathways selectively.

The Soluble Guanylate Cyclase Genetic Variant rs13139571 Is Associated with an “Unfavorable” Metabolic Phenotype in an US General Community

Metabolic syndrome and dyslipidemia are important risk factors for heart failure. Nitric oxide (NO) induces vasodilation and also exerts a metabolic action by stimulating lipogenesis. In rodents, lipid accumulation and lipogenic enzymes are induced by NO and insulin-stimulated glucose uptake is dependent on intact NO synthesis in white adipose tissue. Mice that are knock-out for the inducible NO synthase gene are also protected from high-fat diet induced insulin resistance and NO inhibition in rodents on a high fat diet reduces hepatic lipid accumulation. These biological actions of NO are mediated by soluble guanylate cyclase (sGC). Importantly human adipocytes express the genes GUCY1A2, GUCY1A3, GUCY1B3 encoding for the alpha and beta subunits of sGC. The single nucleotide polymorphism rs13139571 is located within an intron of GUCY1A3 and its minor allele is associated with lower blood pressure and odds of hypertension in a recent genome wide association study. To date, no studies have investigated the metabolic phenotype associated with rs13139571. In accordance with lipogenic properties of NO, the genetic variant rs13139571 will be associated with a metabolic penotype characterized by increased lipid plasma levels and visceral adiposity. We investigated the metabolic phenotype associated with rs13139571 in a random sample (n= 1854) of the general population from Olmsted County, MN. Frequencies of rs13139571 genotypes were CC: 59%, AC: 36%, AA: 5%. In a multivariate analysis adjusted for age, sex, BMI and antilipemic treatment, carriers of the A minor allele had significantly larger waist circumference (CC: 91±14, AC: 92±15, AA: 94±14 cm, p value= 0.01), higher plasma values of total cholesterol (CC: 202 ± 36, AC: 204±36, AA: 208 ±35 mg/dl, p value= 0.02) and triglycerides (CC: 146±87, AC: 151±86, AA: 173±104 mg/dl, p value= 0.02). Prevalence of metabolic syndrome was significantly higher among the carriers of the A minor allele (CC: 21%, AC: 24%, AA: 29%, p value= 0.04). Genotypes did not differ in terms of systolic blood pressure (CC: 132 ± 22, AC: 134 ± 21, AA: 132 ± 21 mmHg, p value= 0. 5), diastolic blood pressure (CC: 73 ± 10, AC: 73 ± 10, AA: 73 ± 12 mmHg, p value= 0.71), BMI (CC: 28 ± 5, AC: 29 ± 5, AA: 28 ± 5 Kg/m2, p value= 0.43) and percentage of subjects on antilipemic therapy (CC: 17%, AC: 19%, AA: 18%, p value= 0.56). The A minor allele of rs13139571 is associated with higher values of waist circumference, plasma levels of total cholesterol and tryglicerides. Metabolic syndrome is more prevalent among carriers of the A allele. The genetic variant rs13139571 may be associated with increased NO activity and influence lipid metabolism by favoring lipogenesis. Additional studies are warranted to confirm our findings and further investigate the metabolic phenotype associated with rs13139571.

A genetic variant of soluble guanylate cyclase is associated with higher plasma cholesterol and triglycerides in the general population

Nitric oxide (NO) is a vasodilator that also possesses lipogenic properties. In rats, lipid accumulation and lipogenic enzymes are induced by NO and insulin-stimulated glucose uptake is dependent on intact NO synthesis in white adipose tissue. Mice in which the gene encoding inducible NO synthase was disrupted are protected from high-fat induced insulin resistance and NO inhibition in rodents on a high fat diet reduces hepatic lipid accumulation. These biological actions of NO are mediated by soluble guanylate cyclase (sGC) and importantly human adipocytes express the genes GUCY1A2, GUCY1A3, GUCY1B3 encoding for the alpha and beta subunits of sGC. The single nucleotide polymorphism rs13139571 is located within an intron of GUCY1A3 and its minor allele is associated with higher blood pressure and odds of hypertension in a recent genome wide association study. For the first time, we investigated the metabolic phenotype associated with rs13139571 in a random sample (n1⁄4 2007) of the general population from Olmsted County, MN. Frequencies of rs13139571 genotypes were CC: 59%, AC: 36%, AA: 5%. In age and gender adjusted analysis, carriers of the A minor allele had significantly higher plasma values of total cholesterol (CC: 202 36, AC: 204 36, AA: 208 34 mg/dl, p value1⁄4 0.04) and triglycerides (CC: 145 85, AC: 150 86, CC: 169 101 mg/dl, p value1⁄4 0.01). Metabolic syndrome trended toward increased prevalence in carriers of the minor allele (CC: 21%, AC: 23%, AA: 28%, p value1⁄4 0.06). Genotypes did not differ in terms of systolic blood pressure (CC: 132 22, AC: 134 21, CC: 132 21 mmHg, p value1⁄4 0.45), diastolic blood pressure (CC: 74 10, AC: 74 10, CC: 73 12 mmHg, p value1⁄4 0.77), BMI (CC: 28 5, 29 5, 28 5 Kg/m, p value1⁄4 0.43) and percentage of subjects on antilipemic therapy (CC: 17%, AC: 19%, AA: 18%, p value1⁄4 0.56). These studies suggest that the minor allele of rs13139571 may be associated with increased NO activity and influence lipid metabolism by favoring lipid accumulation and insulin resistance. Alternatively, rs13139571 may be in linkage disequilibrium with another genetic locus that affects cardiometabolic phenotype. Additional studies are warranted to confirm our findings and further investigate the metabolic phenotype associated with this genetic variant.

Assessing the incidence of probable ischemic colitis in treated adult hypertensive patients

Nitric oxide (NO) is a vasodilator that also possesses lipogenic properties. In rats, lipid accumulation and lipogenic enzymes are induced by NO and insulin-stimulated glucose uptake is dependent on intact NO synthesis in white adipose tissue. Mice in which the gene encoding inducible NO synthase was disrupted are protected from high-fat induced insulin resistance and NO inhibition in rodents on a high fat diet reduces hepatic lipid accumulation. These biological actions of NO are mediated by soluble guanylate cyclase (sGC) and importantly human adipocytes express the genes GUCY1A2, GUCY1A3, GUCY1B3 encoding for the alpha and beta subunits of sGC. The single nucleotide polymorphism rs13139571 is located within an intron of GUCY1A3 and its minor allele is associated with higher blood pressure and odds of hypertension in a recent genome wide association study. For the first time, we investigated the metabolic phenotype associated with rs13139571 in a random sample (n1⁄4 2007) of the general population from Olmsted County, MN. Frequencies of rs13139571 genotypes were CC: 59%, AC: 36%, AA: 5%. In age and gender adjusted analysis, carriers of the A minor allele had significantly higher plasma values of total cholesterol (CC: 202 36, AC: 204 36, AA: 208 34 mg/dl, p value1⁄4 0.04) and triglycerides (CC: 145 85, AC: 150 86, CC: 169 101 mg/dl, p value1⁄4 0.01). Metabolic syndrome trended toward increased prevalence in carriers of the minor allele (CC: 21%, AC: 23%, AA: 28%, p value1⁄4 0.06). Genotypes did not differ in terms of systolic blood pressure (CC: 132 22, AC: 134 21, CC: 132 21 mmHg, p value1⁄4 0.45), diastolic blood pressure (CC: 74 10, AC: 74 10, CC: 73 12 mmHg, p value1⁄4 0.77), BMI (CC: 28 5, 29 5, 28 5 Kg/m, p value1⁄4 0.43) and percentage of subjects on antilipemic therapy (CC: 17%, AC: 19%, AA: 18%, p value1⁄4 0.56). These studies suggest that the minor allele of rs13139571 may be associated with increased NO activity and influence lipid metabolism by favoring lipid accumulation and insulin resistance. Alternatively, rs13139571 may be in linkage disequilibrium with another genetic locus that affects cardiometabolic phenotype. Additional studies are warranted to confirm our findings and further investigate the metabolic phenotype associated with this genetic variant.

S-Nitrosylation of Ascorbate Peroxidase Is Part of Programmed Cell Death Signaling in Tobacco Bright Yellow-2 Cells

Nitric oxide (NO) is a small redox molecule that acts as a signal in different physiological and stress-related processes in plants. Recent evidence suggests that the biological activity of NO is also mediated by S-nitrosylation, a well-known redox-based posttranslational protein modification. Here, we show that during programmed cell death (PCD), induced by both heat shock (HS) or hydrogen peroxide (H2O2) in tobacco (Nicotiana tabacum) Bright Yellow-2 cells, an increase in S-nitrosylating agents occurred. NO increased in both experimentally induced PCDs, although with different intensities. In H2O2-treated cells, the increase in NO was lower than in cells exposed to HS. However, a simultaneous increase in S-nitrosoglutathione (GSNO), another NO source for S-nitrosylation, occurred in H2O2-treated cells, while a decrease in this metabolite was evident after HS. Consistently, different levels of activity and expression of GSNO reductase, the enzyme responsible for GSNO removal, were found in cells subjected to the two different PCD-inducing stimuli: low in H2O2-treated cells and high in the heat-shocked ones. Irrespective of the type of S-nitrosylating agent, S-nitrosylated proteins formed upon exposure to both of the PCD-inducing stimuli. Interestingly, cytosolic ascorbate peroxidase (cAPX), a key enzyme controlling H2O2 levels in plants, was found to be S-nitrosylated at the onset of both PCDs. In vivo and in vitro experiments showed that S-nitrosylation of cAPX was responsible for the rapid decrease in its activity. The possibility that S-nitrosylation induces cAPX ubiquitination and degradation and acts as part of the signaling pathway leading to PCD is discussed.

Inaccuracies of nitric oxide measurement methods in biological media.

Despite growing reports on the biological action of nitric oxide (NO) as a function of NO payload, the validity of such work is often questionable due to the manner in which NO is measured and/or the solution composition in which NO is quantified. To highlight the importance of measurement technique for a given sample type, NO produced from a small-molecule NO donor (N-diazeniumdiolated l-proline, PROLI/NO) and a NO-releasing xerogel film were quantified in a number of physiological buffers and fluids, cell culture media, and bacterial broth by the Griess assay, a chemiluminescence analyzer, and an amperometric NO sensor. Despite widespread use, the Griess assay proved to be inaccurate for measuring NO in many of the media tested. In contrast, the chemiluminescence analyzer provided superb kinetic information in most buffers but was impractical for NO analysis in proteinaceous media. The electrochemical NO sensor enabled greater flexibility across the various media with potential for spatial resolution, albeit at lower than expected NO totals versus either the Griess assay or chemiluminescence. The results of this study highlight the importance of measurement strategy for accurate NO analysis and reporting NO-based biological activity.

High serum nitric oxide metabolites and incident metabolic syndrome

Background. Endothelial dysfunction, synonymous with reduced biological activity of nitric oxide, is related to all cardiovascular risk factors. Association between metabolic syndrome and nitric oxide metabolites (nitrite + nitrate = NOx) has been previously shown in cross-sectional studies. The aim of this study was to determine the ability of serum NOx levels in predicting the incidence of metabolic syndrome in a population-based study. Methods. Serum NOx levels measured in 2098 adult subjects, participants of Tehran Lipid and Glucose Study, without metabolic syndrome at baseline. After 3.3 years follow-up, logistic regression analysis was used to calculate the odds ratio (OR) and 95% confidence interval for developing metabolic syndrome, with serum NOx quartiles as independent variables. Results. Incident metabolic syndrome was diagnosed in 23.0% of men and 16.3% of women after 3.3 years. Age-adjusted risk for developing metabolic syndrome in women who had higher NOx values (above 75th percentile vs. lower 75th percentile) at baseline, was significantly higher [OR: 1.59 (1.11–2.27), p = 0.011]; OR remained significant after multivariable-adjustment including adjustment for components of metabolic syndrome [OR: 1.75 (1.19–2.59), p = 0.005]. Serum NOx could not predict the incidence of metabolic syndrome in men. Conclusions. High serum NOx level is an independent predictor of incident metabolic syndrome in women, but not in men, a finding which presents serum NOx level as a potential biomarker for assessing cardiometabolic disturbances.