Rate Of Nitric Oxide Production Research Articles

Evaluation of Anti-Inflammatory Activity in Methanolic Seed Extracts of International Sorghum bicolor L. Resources

Sorghum is an important cereal with high value as a health food ingredient because it contains various phenolic compounds. Anti-inflammatory activity was assessed using 12 sorghum resources collected from various countries to explore their potential as medicinal resources. The findings revealed that, at extract concentrations of 25 µg/mL and 50 µg/mL, cell survival rates were observed to be between 70 and 80% for most varieties, with the exception of K159081. In the analysis of anti-inflammatory activity, measured by the rate of nitric oxide (NO) production, sorghum varieties K159041 and K159081 exhibited NO production rates of 0.46 ± 0.38% and 2.58 ± 0.20%, respectively, indicating significant anti-inflammatory properties. The investigation into anti-inflammatory effects also included examining the expression of the inducible nitric oxide synthase (iNOS) gene, which is related to the inflammatory response triggered by LPS in macrophages. Varieties K159041, K159048, K159077, K159078, K159081, K159089, and K159096 were analyzed for this purpose. Further, an expression test of the cyclooxygenase 2 (COX-2) gene revealed values less than 0.4 in K159077, K159081, and K159089, suggesting these sorghum lines possess higher anti-inflammatory activity compared to others. Additionally, the expression analysis of tumor necrosis factor alpha (TNF-α), a gene identified as an inflammatory cytokine, showed that the mRNA levels in the lines K159048, K159077, K159078, K159088, K159089, K159093, and K159096 were expressed at lower levels relative to other sorghum resources, categorizing them as having high anti-inflammatory activity. Notably, the K159081 line exhibited the lowest expression level of all genes associated with inflammation, marking it as a valuable medicinal resource with potential development as an anti-inflammatory agent.

Differential expression patterns of purinergic ectoenzymes and the antioxidative role of IL-6 in hospitalized COVID-19 patient recovery.

We have acquired significant knowledge regarding the pathogenesis of severe acute respiratory syndrome caused by coronavirus 2 (SARS-CoV-2). However, the underlying mechanisms responsible for disease recovery still need to be fully understood. To gain insights into critical immune markers involved in COVID-19 etiopathogenesis, we studied the evolution of the immune profile of peripheral blood samples from patients who had recovered from COVID-19 and compared them to subjects with severe acute respiratory illness but negative for SARS-CoV-2 detection (controls). In addition, linear and clustered correlations between different parameters were determined. The data obtained revealed a significant reduction in the frequency of inflammatory monocytes (CD14+CD16+) at hospital discharge vs. admission. Remarkably, nitric oxide (NO) production by the monocyte compartment was significantly reduced at discharge. Furthermore, interleukin (IL)-6 plasma levels were negatively correlated with the frequency of NO+CD14+CD16+ monocytes at hospital admission. However, at the time of hospital release, circulating IL-6 directly correlated with the NO production rate by monocytes. In line with these observations, we found that concomitant with NO diminution, the level of nitrotyrosine (NT) on CD8 T-cells significantly diminished at the time of hospital release. Considering that purinergic signaling constitutes another regulatory system, we analyzed the kinetics of CD39 and CD73 ectoenzyme expression in CD8 T-cells. We found that the frequency of CD39+CD8+ T-cells significantly diminished while the percentage of CD73+ cells increased at hospital discharge. In vitro, IL-6 stimulation of PBMCs from COVID-19 patients diminished the NT levels on CD8 T-cells. A clear differential expression pattern of CD39 and CD73 was observed in the NT+ vs. NT-CD8+ T-cell populations. The results suggest that early after infection, IL-6 controls the production of NO, which regulates the levels of NT on CD8 T-cells modifying their effector functions. Intriguingly, in this cytotoxic cell population, the expression of purinergic ectoenzymes is tightly associated with the presence of nitrated surface molecules. Overall, the data obtained contribute to a better understanding of pathogenic mechanisms associated with COVID-19 outcomes.

Decoupling the simultaneous effects of NO2−, pH and free nitrous acid on N2O and NO production from enriched nitrifying activated sludge

In the pursuit of energy and carbon neutrality, nitrogen removal technologies have been developed featuring nitrite (NO2−) accumulation. However, high NO2− accumulations are often associated with stimulated greenhouse gas (i.e., nitrous oxide, N2O) emissions. Furthermore, the coexistence of free nitrous acid (FNA) formed by NO2− and proton (pH) makes the consequence of NO2− accumulation on N2O emissions complicated. The concurrent three factors, NO2−, pH and FNA may play different roles on N2O and nitric oxide (NO) emissions simultaneously, which has not been systematically studied. This study aims to decouple the effects of NO2− (0-200 mg N/L), pH (6.5-8) and FNA (0-0.15 mg N/L) on the N2O and NO production rates and the production pathways by ammonia oxidizing bacteria (AOB), with the use of a series of precisely executed batch tests and isotope site-preference analysis. Results suggested the dominant factors affecting the N2O production rate were NO2− and FNA concentrations, while pH alone played a relatively insignificant role. The most influential factor shifted from NO2− to FNA as FNA concentrations increased from 0 to 0.15 mg N/L. At concentrations below 0.0045 mg HNO2-N/L, nitrite rather than FNA played a significant role stimulating N2O production at elevated nitrite concentrations. The inhibition effect of FNA emerged with further increase of FNA between 0.0045-0.015 mg HNO2-N/L, weakening the promoting effect of increased nitrite. While at concentrations above 0.015 mg HNO2-N/L, FNA inhibited N2O production especially from nitrifier denitrification pathway with the level of inhibition linearly correlated with the FNA concentration. pH and the nitrite concentration regulated the production pathways, with elevated pH promoting the nitrifier nitrification pathway, while elevated NO2− concentrations promoting the nitrifier denitrification pathway. In contrast to N2O, NO emission was less susceptible to FNA at concentrations up to 0.015 mg N/L but was stimulated by increasing NO2− concentrations. This study, for the first time, distinguished the effects of pH, NO2− and FNA on N2O and NO production, thereby providing support to the design and operation of novel nitrogen removal systems with NO2− accumulation.

Antileishmanial effects, cellular mechanisms, and cytotoxicity of Elettaria cardamomum essential oil against Leishmania major infection.

Leishmaniasis is an infectious disease with various clinical manifestations. We studied the therapeutic effects of Elettaria cardamomum essential oil (ECEO) against Leishmania major infection. In vitro effects of ECEO against L. major were examined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and macrophage assays. Nitric oxide (NO) production, infection inhibition in macrophages, and the apoptotic activity of ECEO in treated parasites were also measured. By calculating the 50% cytotoxic concentrations (CC50), we studied the cytotoxicity effects of ECEO on human macrophage cells (THP-1). The efficacy of ECEO for improving cutaneous leishmaniasis (CL) lesions in mice (BALB/c) was determined by evaluating the size of lesions and the number of amastigotes before and after four weeks of treatment. The effects of ECEO on liver and kidney function in the tested mice were also evaluated. ECEO dose-dependently (p<0.001) inhibited the viability and the mean number of promastigotes and amastigote forms of L. tropica. Four weeks of treatment with ECEO at the doses of 2.5 and 5 mg/kg/ day significantly (p<0.001) improved the CL lesions and reduced the number of parasites in the infected mice. ECEO significantly increased NO production, apoptosis induction, and infection rate in parasites. The CC50 value for ECEO and MA was 303.4 µg/mL and 835.2 µg/mL, respectively. In the mice receiving ECEO at the doses of 2.5 and 5 mg/kg/day for 28 days, no significant change was reported between the serum level of liver enzymes and kidney factors when compared with the control group. ECEO displayed promising efficacy in parasite reduction in vitro and in the animal model. ECEO can thus be used as an alternative medicine to treat CL.

Abnormal flow pattern of low wall shear stress and high oscillatory shear index in spontaneous vertebral artery dissection with vertebral artery hypoplasia.

Spontaneous vertebral artery dissection (sVAD) might tend to develop in vertebral artery hypoplasia (VAH) with hemodynamic dysfunction and it is crucial to assess hemodynamics in sVAD with VAH to investigate this hypothesis. This retrospective study aimed to quantify hemodynamic parameters in patients with sVAD with VAH. Patients who had suffered ischemic stroke due to an sVAD of VAH were enrolled in this retrospective study. The geometries of 14 patients (28 vessels) were reconstructed using Mimics and Geomagic Studio software from CT angiography (CTA). ANSYS ICEM and ANSYS FLUENT were utilized for mesh generation, set boundary conditions, solve governing equations, and perform numerical simulations. Slices were obtained at the upstream area, dissection or midstream area and downstream area of each VA. The blood flow patterns were visualized through instantaneous streamline and pressure at peak systole and late diastole. The hemodynamic parameters included pressure, velocity, time-averaged blood flow, time-averaged wall shear stress (TAWSS), oscillatory shear index (OSI), endothelial cell action potential (ECAP), relative residence time (RRT) and time-averaged nitric oxide production rate (TARNO). Significant focal increased velocity was present in the dissection area of steno-occlusive sVAD with VAH compared to other nondissected areas (0.910 m/s vs. 0.449 vs. 0.566, p < 0.001), while focal slow flow velocity was observed in the dissection area of aneurysmal dilatative sVAD with VAH according to velocity streamlines. Steno-occlusive sVAD with VAH arteries had a lower time-averaged blood flow (0.499 cm3/s vs. 2.268, p < 0.001), lower TAWSS (1.115 Pa vs. 2.437, p = 0.001), higher OSI (0.248 vs. 0.173, p = 0.006), higher ECAP (0.328 Pa-1 vs. 0.094, p = 0.002), higher RRT (3.519 Pa-1 vs. 1.044, p = 0.001) and deceased TARNO (104.014 nM/s vs. 158.195, p < 0.001) than the contralateral VAs. Steno-occlusive sVAD with VAH patients had abnormal blood flow patterns of focal increased velocity, low time-averaged blood flow, low TAWSS, high OSI, high ECAP, high RRT and decreased TARNO. These results provide a good basis for further investigation of sVAD hemodynamics and support the applicability of the CFD method in testing the hemodynamic hypothesis of sVAD. More detailed hemodynamic conditions with different stages of sVAD are warranted in the future.

Transport of nitrite from large arteries modulates regional blood flow during stress and exercise.

Acute cardiovascular stress increases systemic wall shear stress (WSS)-a frictional force exerted by the flow of blood on vessel walls-which raises plasma nitrite concentration due to enhanced endothelial nitric oxide synthase (eNOS) activity. Upstream eNOS inhibition modulates distal perfusion, and autonomic stress increases both the consumption and vasodilatory effects of endogenous nitrite. Plasma nitrite maintains vascular homeostasis during exercise and disruption of nitrite bioavailability can lead to intermittent claudication. During acute cardiovascular stress or strenuous exercise, we hypothesize enhanced production of nitric oxide (NO) by vascular endothelial cells raises nitrite concentrations in near-wall layers of flowing blood, resulting in cumulative NO concentrations in downstream arterioles sufficient for vasodilation. Utilizing a multiscale model of nitrite transport in bifurcating arteries, we tested the hypothesis for femoral artery flow under resting and exercised states of cardiovascular stress. Results indicate intravascular transport of nitrite from upstream endothelium could result in vasodilator-active levels of nitrite in downstream resistance vessels. The hypothesis could be confirmed utilizing artery-on-a-chip technology to measure NO production rates directly and help validate numerical model predictions. Further characterization of this mechanism may improve our understanding of symptomatic peripheral artery occlusive disease and exercise physiology.

DRC1 deficiency caused primary ciliary dyskinesia and MMAF in a Chinese patient.

Primary ciliary dyskinesia (PCD) is a heterogeneous disease characterized by the failure of mucociliary clearance. Dynein regulatory complex subunit 1 (DRC1) variants can cause PCD by disrupting the nexin link connecting the outer doublets. In this study, we aimed to investigate the clinical and functional impacts of DRC1 variants on respiratory cilia and sperm. We identified and validated the DRC1 variant by using whole-exome and Sanger sequencing. High-speed video microscopy analysis (HSVA) was used to measure the nasal ciliary beating frequency and pattern in a patient and a healthy control. Hematoxylin-eosin (HE) staining and transmission electron microscopy (TEM) were applied to analyze the morphological and ultrastructural sperm defects resulting from the DRC1 variant. NM_145038.5:c.1296 G>A, p.(Trp432*), a novel homozygous DRC1 nonsense variant, was identified in a patient from a consanguineous Chinese family. The patient exhibited bronchiectasis, chronic sinusitis, situs solitus, and male infertility. The markedly reduced nasal nitric oxide production rate (3.0 nL/min) was consistent with PCD diagnosis. HSVA showed reduced bending capacity and higher beating frequency of nasal cilia in the patient compared with those in healthy control. The diagnosis of multiple morphological abnormalities of the sperm flagella (MMAF) was confirmed through sperm HE staining and TEM analysis. Following the intracytoplasmic sperm injection treatment, the patient fathered a healthy daughter. This report is the first to describe a novel DRC1 variant in a patient with PCD and MMAF, and in vitro fertilization was effective for treating infertility in this male patient. Our findings expand the genetic spectrum of PCD and MMAF, and provide a detailed clinical summary and functional analysis of patients with DRC1 variants.

ROLE OF ΚB AND AP-1 TRANSCRIPTION FACTORS IN CHANGES IN NITRIC OXIDE PRODUCTION AND UTILIZATION IN THE HEART OF RATS UNDER CHRONIC FLUORIDE INTOXICATION

Fluorides, being hazardous contaminants of soil and drinking water, can get in excessive amounts into human and animal bodies. This is especially true for regions where the fluoride content in soils is very high, for example, Poltava, Dnipropetrovsk, and Kirovohrad regions in Ukraine. Excessive fluoride intake can change the rate of nitric oxide production. The impact of fluorides on changes in nitric oxide production and metabolism in the heart and the role of redox-sensitive transcription factors in these changes are poorly understood. The aim of this study was to determine the effect of activation of κB transcription factors and activator protein 1 on the activity of inducible NO-synthase, constitutive isoforms of NO-synthase, nitrite and nitrate reductase, arginase, concentration of nitrites, peroxynitrite and nitrosothiols in the heart of rats during chronic fluoride intoxication. Materials and methods. The study was performed on 24 adult male Wistar rats weighing 220-260 grams. Animals were randomly divided into 4 groups of 6 animals in each (control, chronic fluoride intoxication group, κB blockade group and activator protein 1 blockade group). The experiment lasted 30 days. We determined the activity of inducible NO-synthase, constitutive isoforms of NO-synthase, the concentration of peroxynitrite alkali and alkaline earth metals, the concentration of nitrites and nitrosothiols, the activity of nitrite reductase, nitrate reductase and arginase. Results. Chronic fluoride intoxication increases the activity of inducible NO-synthase by 1.74 times, does not affect the activity of constitutive isoforms and reduces the activity of arginase by 35.68% compared with the control group of animals. The concentration of nitrites in the heart of rats increases 1.73 times, peroxynitrite 1.43 times, and the concentration of nitrosothiols doubled. The use of κB transcription factor blockers and activator protein 1 reduces nitric oxide production from NO synthases and reduces the concentrations of all nitric oxide metabolites in the heart of rats under conditions of chronic fluoride intoxication. Conclusions. Activation of κB transcription factors and activator protein 1 during chronic excessive intake of fluoride leads to hyperproduction of nitric oxide in the heart of rats due to increased activity of inducible NO-synthase and nitrite reductases. Excess production of nitric oxide under chronic fluoride intoxication leads to the accumulation of nitrites, peroxynitrite and nitrosothiols in the heart of rats.

Glucose availability limits microglial nitric oxide production.

Metabolic intermediates influence inflammation not only through signaling effects, but also by fueling the production of pro-inflammatory molecules. Microglial production of nitric oxide (NO) requires the consumption of NADPH. NADPH consumed in this process is regenerated from NADP+ primarily through the hexose monophosphate shunt, which can utilize only glucose as a substrate. These factors predict that glucose availability can be rate-limiting for glial NO production. To test this prediction, cultured astrocytes and microglia were incubated with lipopolysaccharide and interferon-γ to promote expression of inducible nitric oxide synthase, and the rate of NO production was assessed at defined glucose concentrations. Increased NO production was detected only in cultures containing microglia. The NO production was markedly slowed at glucose concentrations below 0.5mM, and comparably reduced by inhibition of the hexose monophosphate shunt with 6-aminonicotinamide. Reduced NO production caused by glucose deprivation was partly reversed by malate, which fuels NADPH production by malate dehydrogenase, and by NADPH itself. These findings highlight the role of the hexose monophosphate shunt in fueling NO synthesis and suggest that microglial NO production in the brain may be limited at sites of low glucose availability, such as abscesses or other compartmentalized infections.

Acute Hemodynamic Improvement by Thermal Vasodilation inside the Abdominal and Iliac Arterial Segments of Young Sedentary Individuals

Objective: To study the hemodynamic response to lower leg heating intervention (LLHI) inside the abdominal and iliac arterial segments (AIAS) of young sedentary individuals. Methods: A Doppler measurement of blood flow was conducted for 5 young sedentary adults with LLHI. Heating durations of 0, 20, and 40 min were considered. A lumped parameter model (LPM) was used to ascertain the hemodynamic mechanism. The hemodynamics were determined via numerical approaches. Results: Ultrasonography revealed that the blood flow waveform shifted upwards under LLHI; in particular, the mean flow increased significantly (p < 0.05) with increasing heating duration. The LPM showed that its mechanism depends on the reduction in afterload resistance, not on the inertia of blood flow and arterial compliance. The time-averaged wall shear stress, time-averaged production rate of nitric oxide, and helicity in the external iliac arteries increased more significantly than in other segments as the heating duration increased, while the oscillation shear index (OSI) and relative residence time (RRT) in the AIAS declined with increasing heating duration. There was a more obvious helicity response in the bilateral external iliac arteries than the OSI and RRT responses. Conclusion: LLHI can effectively induce a positive hemodynamic environment in the AIAS of young sedentary individuals.

Beet root (Beta vulgaris) protects lipopolysaccharide and alcohol-induced liver damage in rat.

The purpose of this study was to investigate the protective effects of beet root (Beta vulgaris var. rubra) in lipopolysaccharide (LPS) and alcohol induced liver damage. Beta vulgaris ethanol extract (BVEE) showed good antioxidant activity in the contents of polyphenol and flavonoid compounds, and the electron-donating ability and ABTS+ radical scavenging activity. As for anti-inflammatory effect in RAW 264.7 cells, inhibition rate of nitric oxide production was increased in dose dependent manner. In hepatotoxicity model induced by LPS and alcohol in rat, BVEE significantly decreased serum AST, ALT and γ-GTP concentrations in a dose-dependent manner. The histopathological changes after H&E staining showed that fat accumulation and inflammatory cell infiltration were decreased by BVEE. The collagen fibers around the central lobule observed by Masson's trichrome staining were also decreased by BVEE. In addition, as for the immunohistochemical staining and Transmission electron microscopy, BVEE improved morphological characteristics of damaged liver lesion. The increased mRNA expressions of NF-κB, MAPK1, MAPK3, CYP2E1, and α-SMA were significantly decreased in BVEE treated group. These results indicated that BVEE would have protective effects in hepatotoxicity by altering various indicators related to the liver damage induced by LPS or alcohol.

H2S-induced NO/SNO positively promotes betulin production in Betula platyphylla

S-nitrosoglutathione reductase (GSNOR) is the pivotal enzyme that mediates cellular levels of nitric oxide (NO) and S-nitrosylation (SNO) in plants. We have previously reported that GSNOR deficiency enhanced betulin production. However, the physiological function of NO/SNO in plants remains unclear, especially for SNO. This study aimed to reveal the role of hydrogen sulfide (H2S)-induced NO/SNO on betulin production in birch (Betula platyphylla). Results showed that the H2S donor sodium hydrosulfide(NaHS)enhanced the production rates of NO, SNO and betulin in birch suspension cells to maximum levels, which was 92.59%, 50.09% and 122.58% higher than that in the control, at 24 h after 1 mmol·L−1 NaHS treatment. NaHS also promoted NO burst, SNO production and the gene expression of farnesyl pyrophosphate synthase, lupeol synthase and squalene synthetase related to betulin synthesis in birch GSNOR transgenic plants by RNA interference silencing. Similarly, pretreatment of the birch suspension cells with the GSNOR inhibitor 3-(5-(4-(1H-imidazol-1-yl) phenyl)-1-(4-carbamoyl-2-methylphenyl)- 1H-pyrrol-2-yl) propionic acid (N6022) promoted NaHS-triggered NO/SNO and betulin production, and the betulin content under NaHS and N6022 treatment was significantly higher than that under NaHS treatment or NaHS and NO specific scavenger 2-(4-carboxyphenyl)-4,4,5,5 -tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). However, pretreatment of the birch suspension cells with cPTIO or NO synthase inhibitor (sodium azide or NG-nitro-L-Arg methyl ester) significantly decreased NaHS-triggered NO/SNO and betulin production. Overall, the above results first verified that endogenous SNO mediated hydrogen sulfide-induced betulin production at the pharmacological and genetic levels, and the homeostasis of NO/SNO mediated hydrogen sulfide-induced betulin production. This study provides new insights into the homeostasis of NO/SNO in plant secondary metabolite production.

Acute Soy Supplementation Improves 20-km Time Trial Performance, Power, and Speed.

Isoflavones, a chemical class of phytoestrogens found in soybeans and soy products, may have biological functions similar to estradiol. After binding with ERβ or perhaps independently of estrogen receptors, isoflavones may augment vascular endothelial relaxation, contributing to improved limb blood flow. To determine if acute fermented soy extract supplementation influences 20-km time trial cycling performance and cardiac hemodynamics compared with a placebo. Subjects included 25 cyclists and triathletes (31 ± 8 yr, V˙O2peak: 55.1 ± 8.4 mL·kg·min). Each subject completed a V˙O2peak assessment, familiarization, and two 20-km time trials in randomized order after ingestion of a fermented soy extract supplement or placebo. The fermented soy extract consisted of 30 g powdered supplement in 16 fl. ounces of water. The placebo contained the same quantities of organic cocoa powder and water. Each trial consisted of 60 min of rest, 30 min at 55% Wpeak, and a self-paced 20-km time trial. Soy supplementation elicited a faster time to 20-km completion (-0.22 ± 0.51 min; -13 s), lower average HR (-5 ± 7 bpm), and significantly greater power (7 ± 3 W) and speed (0.42 ± 0.16 km·h) during the last 5 km of the time trial compared with placebo. Analysis of the results by relative fitness level (<57 vs ≥ 57 mL⋅kg⋅min) indicated that those with a higher level of fitness reaped the largest performance improvement alongside a reduced HR (-5 ± 7 bpm). Ingestion of a fermented soy extract supplement improved sprint-distance performance through improvements in both power and speed. For those with great aerobic fitness, soy supplementation may help to decrease cardiac demand alongside performance improvement.

Hemodynamic Performance of Multilayer Stents in the Treatment of Aneurysms with a Branch Attached

Although multilayer stents (MSs) can be used to treat aneurysm effectively, for some aneurysms with branches attached, the hemodynamic mechanisms are still unclear. In this work, we modeled five cases that involve 1–4-layer stents implanted in aneurysms with side branches, and the numerical approach was used. Case 1 corresponds to an aneurysm without a stent, and cases 2–5 represent 1–4-layer stents being employed within aneurysms, respectively. The results showed that the velocity within the sac declined dramatically and the eddies’ intensity weakened with increased number of stent layers, time-averaged wall shear stress (TAWSS), and nitric oxide production rate (TARNO) dropped linearly with increase in stent porosity, and oscillatory shear index (OSI) and relative residence time (RRT) increased evidently with MS intervention. Moreover, the MSs had a slight effect on the patency of the side branch; its flow rate was still above the normal case than without aneurysm. It can be concluded that MSs are helpful in promoting the growth of thrombus within the aneurysm through an isolated hemodynamic environment and keeping the branch unobstructed, but more clinical evidences are required.

Computational analysis of nitric oxide biotransport in a microvessel influenced by red blood cells

It is pivotal that endothelium-dependent Nitric Oxide (NO) consumed by hemoglobin (Hb) inside red blood cells (RBCs) membrane, regulates the vascular tone. The whole processes of NO transport in vessel containing flowing RBCs is still not clear, such as NO production in endothelium, diffusion in plasma and consumption inside RBCs. In this work, the motion of RBCs in a microvessel is investigated by using immersed boundary lattice Boltzmann method (IB-LBM) first and the deformability of RBCs is expressed by using spring network model which is based on the minimum energy principle. Furthermore, the interaction between RBCs is considered. Based on the wall shear stress (WSS), NO production rate originated from endothelium was obtained by using a hyperbolic model. NO distribution inside the microvessel with multiple RBCs was computed by using immersed boundary finite difference method (IB-FDM). The result shows that a large (small) WSS exists at locations with a relatively wide(narrow) gap between the wall and cell. In terms of mass transfer, an increase of RBC membrane permeability leads to a decrease of NO concentration in the vessel and the surrounding endothelium significantly. In addition, with the increasing of hematocrit (Hct) value, NO concentration distribution in the whole vessel decreases both in the lumen and vascular wall. Finally, the thickness of RBCs-depleted layer gradually decreases with the weakened deformability of RBCs membrane, and the change degree of cell free layer (CFL) thickness decreases as the bending stiffness is relatively higher. Thus, when bending stiffness is higher, the NO concentration in vascular wall is reduced resulting from the thinner CFL.

42 - How does ascorbate improve endothelial dysfunction?

Low levels of ascorbate (ASC, reduced form of vitamin C) are associated with several diseases linked to increased oxidative stress, such as cancer, diabetes, HIV, sepsis and other cardiovascular diseases (CVD’s). In these diseases, either the systemic or localized cellular ASC deficiency is reported as a cause for endothelial dysfunction (ED). It is well established that ED is primarily caused due to reduction in nitric oxide (NO) and increase in oxidative stress. The synthesis and bioavailability of NO are sensitive to ASC, yet the mechanism by which ASC improves ED is unclear. In the present study, we examined putative mechanisms that might account for the ability of ASC to increase NO bioavailability and thus improve ED. Since the biochemical interactions among the reactive species are complex and rapid, we used the computational modeling approaches to understand these mechanistic interactions. In the present study, a single cell computational model was developed that involves eNOS biochemical pathway, downstream reactions involving NO and various reactive oxygen and nitrogen species, cellular oxidative stress and ASC. Using this model, we analyzed the role of ASC in ED by determining, i) the NO bioavailability; ii) optimal levels of intracellular ASC required to improve ED and iii) analyzing interactions of ASC with BH4 and other reactive species including O2•– and ONOO–. The model results show, (i) oxidative stress decreased the NO production and biopterin ratio and increased O2•– production; (ii) ASC supplementation in the range of 10 – 100 µM improved NO production rate, BH4 bioavailability and biopterin ratio under varying oxidative stress conditions; (iii) scavenging of O2•– and ONOO– by ASC was negligible. This study suggests that ASC improves ED by enhancing BH4 bioavailability and stabilizing eNOS under oxidative stress conditions.

Managing the power grid: how myoglobin can regulate PO2 and energy distribution in skeletal muscle.

Managing the power grid: how myoglobin can regulate PO2 and energy distribution in skeletal muscle.

Experimental Evidence for in Situ Nitric Oxide Production in Anaerobic Ammonia-Oxidizing Bacterial Granules.

Although nitric oxide (NO) emissions from anaerobic ammonium oxidation (anammox)-based processes were reported previously, the NO production pathways are poorly understood. Here, we investigated the NO production pathways in anammox granules in detail by combining 15N-stable isotope tracer experiments with various inhibitors, microsensor measurements, and transcriptome analysis for key genes of NO2- reduction. NO was emitted from the anammox granules, which account for 0.07% of the N2 emission. 15N-stable isotope-tracer experiments indicated that most of the N2 was produced by anammox bacteria, whereas NO was produced from NO2- reduction by anammox and denitrifying bacteria. The NO emission rate was highest at pH 8.0 and accelerated by increasing NH4+ and NO2- concentrations in the culture media. The microsensor analyses showed the in situ NO production rate was highest in the outer layer of the anammox granule where anammox activity was also highest. The detected in situ NO concentrations of up to 2.7 μM were significantly above physiological thresholds known to affect a wide range of microorganisms present in wastewater. Hence, NO likely plays pivotal roles in the microbial interactions in anammox granules, which needs to be further investigated.

Nitric oxide generating copper–chitosan particles for wound healing applications

AbstractBACKGROUNDNitric oxide (NO) is a signaling molecule that plays many roles during infection, inflammation, and wound healing processes. Due to the role of NO in wound repair, a novel NO generation system was developed based on copper–chitosan complexes that can be used for the topical generation of NO. Chitosan, a biocompatible polymer, chelates copper ions. Copper in the +1 state can reduce nitrite (NO2–) and convert it into NO. With glucose, a reducing sugar, present in the system, Cu+2 can be returned to Cu+1 to complete the catalytic cycle.RESULTSCopper–chitosan milli‐ and micro‐sized particles were produced using microfluidic techniques. Copper–chitosan milli‐particles (Cu‐chito) did produce nitric oxide (NO). The maximum rates of NO production were ∼ 1.40 nmol min‐1 g‐1 (Cu‐chito) and 1.08 nmol min‐1 g‐1 (Cu‐chito + glucose). The milli‐particles were tested with ARPE‐19 cell lines in cell proliferation assays. Cu‐chito particle treatments with nitrite showed 130% more growth in comparison with chitosan milli‐particles not containing copper. Furthermore, Cu‐chito treatments of nitrite + glucose showed 152% more growth in comparison with control groups, and 118% in comparison with Cu‐chito with nitrite alone. The activity of intracellular NO target, matrix metalloproteinases (MMP‐2 and ‐9), were shown to increase by 60% after 48 h of Cu‐chito ± glucose treatments.CONCLUSIONNO‐releasing copper–chitosan derivatives were produced, with proof of concept for nitric oxide release and positive effects on a cell culture model of wound healing. © 2018 Society of Chemical Industry

13 - A computational analysis of interactions of oxidative stress and antioxidant system in endothelial dysfunction

Introduction Oxidative stress and endothelial dysfunction have been associated in the development and progression of the cardiovascular diseases (CVDs). Oxidative stress is characterized by significant increase in the reactive oxygen species (predominantly superoxide (O2•‒ )) and the decrease in the nitric oxide (NO) bioavailability and antioxidant system of the cell. The major antioxidant system affected in endothelial dysfunction are enzymatic system such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and non-enzymatic systems such as glutathione (GSH) and ascorbate ((ASC). Studies on the therapeutic potential of increasing the antioxidants capacity to improve endothelial dysfunction have reported mixed results. This is because we have very little quantitative information on the complex interplay between the reactive species and antioxidant system involved in the disease progression. The role of antioxidants in oxidative stress is a complicated problem and can be addressed by computational modeling approaches. Material and Methods In the present study, we developed a computational model that involves eNOS biochemical pathway, downstream reactions involving NO and various reactive oxygen and nitrogen species, cellular oxidative stress and antioxidant systems. The role of GSH, GPX, ASC, SOD and CAT in the NO and O2•‒ co-generated system was analyzed to investigate the changes in NO and O2•- production rates, biopterin ratio with respect to changes in cellular oxidative stress. Results and Discussions The model results show that oxidative stress decreased the NO production and biopterin ratio and increased O2•‒ production. The upregulation of ASC, GSH, SOD improved the NO production rates and biopterin ratio but the decrease in O2•‒ production was insignificant when higher cellular oxidative stress was present. ASC supplementation improved endothelial dysfunction by improving biopterin ratio and making tetrahydrobiopterin (BH4) available. The extent of improvement in endothelial dysfunction by antioxidants was dependent on extent of cellular oxidative stress. Conclusion The present study provides quantitative insights on how the up- or down- regulation of key antioxidant enzymes affects the cellular ROS levels and endothelial function. The thorough mechanistic analysis of each antioxidant performed under endothelial dysfunction reveals the important antioxidant systems relevant for oxidative stress.