In this study we confirmed that NO may induce HO-1 expression and provided a novel mechanism by which NO modulates NADPH oxidase-dependent superoxide production. This mechanism of NADPH oxidase suppression is supported by the following findings: (1) an NO donor consistently induced HO-1 expression; (2) the NO donor suppressed superoxide-generating capacity of NADPH oxidase, an effect distinct from a direct neutralizing action of NO; (3) the inhibitory effect of NO was abolished by cotreatment with HO-1 inhibitors; and (4) bilirubin, the metabolic product of HO-1, mimicked the effect of NO. The current results do not exclude the possibility that NO acts via other mechanisms to suppress NADPH oxidase activation, which can be achieved with the NO donor at concentrations ,30 μmol/L, as reported previously in neutrophils24 and more recently by our group in HMECs.9 In the present study, however, a direct inhibition of NADPH oxidase function is unlikely to be the predominant mechanism, because the ...

Phagocyte superoxide production by a multicomponent NADPH oxidase is important in host defense against microbial invasion. However inappropriate NADPH oxidase activation causes inflammation. Endothelial cells express NADPH oxidase and endothelial oxidative stress due to prolonged NADPH oxidase activation predisposes many diseases. Discovering the mechanism of NADPH oxidase activation is essential for developing novel treatment of these diseases. The p47(phox) is a key regulatory subunit of NADPH oxidase; however, due to the lack of full protein structural information, the mechanistic insight of p47(phox) phosphorylation in NADPH oxidase activation remains incomplete. Based on crystal structures of three functional domains, we generated a computational structural model of the full p47(phox) protein. Using a combination of in silico phosphorylation, molecular dynamics simulation and protein/protein docking, we discovered that the C-terminal tail of p47(phox) is critical for stabilizing its ...

Chronic Granulomatous Disease (CGD) is an inherited immunodeficiency linked with mutations in the multi-component leukocyte NADPH oxidase. Phagocytic cells deficient in NADPH oxidase fail to produce sufficient levels of the reactive oxygen species (ROS) necessary to clear engulfed pathogens. Though the hyperactive immune response associated with CGD is widely described in phagocytes, the effects of oxidase-deficiency on B cell functions has not been well studied. In B cells, recognition of microbial single-stranded RNA or unmethylated DNA by intracellular toll-like receptors (TLR) 7 and 9, respectively leads to NADPH oxidase derived ROS production. Here, studies demonstrate hyperactive responses to endosomal TLR stimulation in B cell lines derived from patients with mutations in either the p40phox or p47phox subunits of the NADPH oxidase which compromise its function. NADPH oxidase-deficient B cell lines expressed enhanced levels of TLR7 and TLR9 mRNA compared to cells reconstituted to restore ...

Oxidative stress plays a critical role in the vascular complications of type 2 diabetes. We examined the effect of type 2 diabetes on NADPH oxidase in human vessels and explored the mechanisms of this interaction. Segments of internal mammary arteries (IMAs) with their perivascular adipose tissue (PVAT) and thoracic adipose tissue were obtained from 386 patients undergoing coronary bypass surgery (127 with type 2 diabetes). Type 2 diabetes was strongly correlated with hypoadiponectinemia and increased vascular NADPH oxidase-derived superoxide anions (O2˙(-)). The genetic variability of the ADIPOQ gene and circulating adiponectin (but not interleukin-6) were independent predictors of NADPH oxidase-derived O2˙(-). However, adiponectin expression in PVAT was positively correlated with vascular NADPH oxidase-derived O2˙(-). Recombinant adiponectin directly inhibited NADPH oxidase in human arteries ex vivo by preventing the activation/membrane translocation of Rac1 and downregulating p22(phox) through a

NADPH oxidase-derived reactive oxygen species (ROS) contribute to the pathobiology of vascular disease. However, studies investigating NADPH oxidases in atherosclerosis have been limited in their ability to distinguish between the role of vascular cell and inflammatory cell -derived ROS. In this study, we examined the contribution of Nox1-derived ROS in a mouse model of atherosclerosis. Nox1 is a primary catalytic subunit of vascular cell, but not inflammatory cell, NADPH oxidase. At weaning, male apolipoprotein E deficient mice (AS, n=12) and male mice deficient in both apolipoprotein E and Nox1 (AS/Nox, n=16) received an atherogenic diet for 18 weeks. Mean blood pressures (116±3 vs. 110 ±3 mmHg; AS vs. AS/Nox, n=6), weights, and serum cholesterol levels (1578±150 vs. 1631±80 mg/dl; AS vs. AS/Nox) were similar between the AS and AS/Nox mice. As measured by lucigenin-enhanced chemiluminescence, superoxide levels were increased in segments of thoracic aorta from AS mice as compared to aorta ...

Neutrophil cytosol factor 2 is a protein that in humans is encoded by the NCF2 gene. This gene encodes neutrophil cytosolic factor 2, the 67-kilodalton cytosolic subunit of the multi-protein complex known as NADPH oxidase found in neutrophils. This oxidase produces a burst of superoxide which is delivered to the lumen of the neutrophil phagosome. Mutations in this gene, as well as in other NADPH oxidase subunits, can result in chronic granulomatous disease. GRCh38: Ensembl release 89: ENSG00000116701 - Ensembl, May 2017 GRCm38: Ensembl release 89: ENSMUSG00000026480 - Ensembl, May 2017 Human PubMed Reference:. Mouse PubMed Reference:. Entrez Gene: NCF2 neutrophil cytosolic factor 2 (65kDa, chronic granulomatous disease, autosomal 2). Wientjes FB, Segal AW (1996). NADPH oxidase and the respiratory burst. Semin. Cell Biol. 6 (6): 357-65. doi:10.1016/S1043-4682(05)80006-6. PMID 8748143. DeLeo FR, Quinn MT (1997). Assembly of the phagocyte NADPH oxidase: molecular interaction of oxidase ...

In the present study, we investigated the influence of hypoxia on vascular repair and EPC mobilization. We found that both processes depend on the NADPH oxidase Nox2. EPO alone was sufficient to mimic the effects of hypoxia on EPC mobilization and vascular repair. The signal transduction of EPO critically required Nox2, potentially as Nox2-derived ROS transiently inhibited the phosphatase SHP-2.. Injury-induced mobilization of EPCs from the bone marrow involves the local production of hormones such as VEGF and SDF-1α, and the subsequently formed gradient between the site of injury and the bone marrow is one of the factors required for EPC release. Interestingly, a role for Nox2 in the signaling of VEGF and SDF-1α has been observed: the production of VEGF23 as well as its signaling require Nox2 and explains why angiogenesis in the femoral artery ligation model of NADPH oxidase-deficient mice is attenuated.24 In addition the release of SDF-1α from the injured hindlimb is reduced in ...

TY - JOUR. T1 - Inactivation of neutrophil NADPH oxidase upon dilution and its prevention by cross-link and fusion of phox proteins. AU - Miyano, Kei. AU - Kitahara, Hiroki. AU - Ohmi, Shinobu. AU - Kakinuma, Katsuko. AU - Tamura, Minoru. PY - 2004/11/1. Y1 - 2004/11/1. N2 - Activation of the phagocyte NADPH oxidase involves assembly of p47 phox, p67 phox, Rac, and flavocytochrome b 558, and the activation can be triggered in a cell-free system with an anionic amphiphile. We find that the activated oxidase in a pure cell-free system was rapidly inactivated upon dilution. When the activated oxidase was treated with a chemical cross-linker, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, the half-life of the oxidase in dilution was extended from 1 min to 4 h at 25°C. The cross-linked oxidase was resistant to inhibition by inactive flavin analogs, indicating that cross-linking prevents flavin exchange. When a fusion protein p67N-p47N plus RacQ61L was added, flavocytochrome b 558 became ...

INTRODUCTION: Oxidative stress that is mediated through NADPH oxidase activity plays a role in the pathology of hypertension, and aerobic exercise training reduces NADPH oxidase activity. The involvement of genetic variation in the p22phox (CYBA) subunit genes in individual oxidative stress responses to aerobic exercise training has yet to be examined in Pre and Stage 1 hypertensives. METHODS: Ninety-four sedentary Pre and Stage 1 hypertensive adults underwent 6 months of aerobic exercise training at a level of 70% VO2max to determine whether the CYBA polymorphisms, C242T and A640G, were associated with changes in urinary 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha), urinary nitric oxide metabolites (NOx), and plasma total antioxidant capacity (TAC). RESULTS: Demographic and subject characteristics were similar among genotype groups for both polymorphisms. At baseline, a significant (P = 0.03) difference among the C2424T genotype groups in 8-iso-PGF2alpha levels was detected, with the TT ...

Our results show an important role of monocyte/macrophage NADPH oxidase in mediating antifungal host defense and in restraining acute neutrophilic inflammation. In the context of global NADPH oxidase deficiency, transgenic monocyte/macrophage-targeted NADPH oxidase was protective against pulmonary aspergillosis. The lethal inoculum in Ncf1*/*MN+ mice was ,100-fold greater compared with globally NADPH oxidase-deficient Ncf1*/*MN− mice. Transgenic mice developed no signs of histological fungal disease following a fungal inoculum that led to lethal fungal pneumonia in Ncf1*/*MN− mice. Consistent with these in vivo results, NADPH oxidase in isolated alveolar macrophages limited conidial growth. Additionally, monocyte/macrophage-targeted NADPH oxidase attenuated zymosan-induced neutrophilic lung inflammation. Taken together, these results support a model in which NADPH oxidase in the monocyte/macrophage lineage inhibits fungal growth and restrains inflammation induced by proinflammatory fungal ...

Introduction: NADPH oxidases are a major cardiac source of reactive oxygen species (ROS). Global deletion of NADPH oxidase type 2 (Nox2) protects against cardiac remodeling after myocardial infarction (MI) but the relevant cellular source for this effect is unknown. We investigated the relative importance of Nox2 in cardiomyocytes and endothelial cells in this process.. Methods: Mice with either endothelial-targeted or cardiomyocyte-targeted overexpression of Nox2, and their respective wild-type (WT) littermates, were studied. Permanent ligation of the left coronary artery (or sham operation) with subsequent recovery and four week follow-up was performed.. Results: Mortality post-MI was similar in all groups. Infarct size was similar in endothelial-Nox2 transgenics (Tg) and cardiomyocyte-Nox2 Tg at 2 days post-operatively (41.3 +/− 1.9 % vs. 44.8 +/− 2.6 % of LV area, by MRI late gadolinium enhancement) and at 4 weeks post-operatively (42.3 +/− 2.4 % vs. 42.1 +/− 2.5 %, by infarct length ...

TY - JOUR. T1 - The Rac effector p67phox regulates phagocyte NADPH oxidase by stimulating Vav1 guanine nucleotide exchange activity. AU - Ming, Wenyu. AU - Li, Shijun. AU - Billadeau, Daniel D.. AU - Quilliam, Lawrence A.. AU - Dinauer, Mary C.. N1 - Copyright: Copyright 2009 Elsevier B.V., All rights reserved.. PY - 2007/1/1. Y1 - 2007/1/1. N2 - The phagocyte NADPH oxidase catalyzes the reduction of molecular oxygen to superoxide and is essential for microbial defense. Electron transport through the oxidase flavocytochrome is activated by the Rac effector p67phox. Previous studies suggest that Vav1 regulates NADPH oxidase activity elicited by the chemoattractant formyl-Met-Leu-Phe (IMLP). We show that Vav1 associates with p67phox and Rac2, but not Rac1, in fMLP-stimulated human neutrophils, correlating with superoxide production. The interaction of p67 phox with Vav1 is direct and activates nucleotide exchange on Rac, which enhances the interaction between p67phox and Vav1. This provides new ...

Dysfunction of T cells and natural killer (NK) cells has been proposed to determine the course of disease in acute myeloid leukemia (AML), but only limited information is available on the mechanisms of lymphocyte inhibition. We aimed to evaluate to what extent human malignant AML cells use NADPH oxidase-derived reactive oxygen species (ROS) as an immune evasion strategy. We report that a subset of malignant myelomonocytic and monocytic AML cells (French-American-British [FAB] classes M4 and M5, respectively), recovered from blood or BM of untreated AML patients at diagnosis, expressed the NADPH oxidase component gp91(phox). Highly purified FAB M4/M5 AML cells produced large amounts of ROS on activation and triggered poly-[ADP-ribose] polymerase-1-dependent apoptosis in adjacent NK cells, CD4(+) T cells, and CD8(+) T cells. In contrast, immature (FAB class M1) and myeloblastic (FAB class M2) AML cells rarely expressed gp91(phox), did not produce ROS, and did not trigger NK or T-cell apoptosis. ...

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Essential hypertensive patients show a reduced nitric oxide availability secondary to oxidative stress generation in peripheral microcirculation. Cyclooxygenase (COX) contributes to reduce nitric oxide availability. We assessed the possible vascular sources of oxidative stress, including COX-1, COX-2, and nicotinamide adenine dinucleotide phosphate oxidase, as determinants of endothelial dysfunction in small arteries isolated from essential hypertensive patients or normotensive controls. Small arteries were dissected after subcutaneous fat biopsies and evaluated on a pressurized micromyograph. Endothelium-dependent vasodilation was assessed by acetylcholine, repeated under NG-nitro-l-arginine methyl ester, SC-560 (COX-1 inhibitor), DuP-697 (COX-2 inhibitor), ascorbic acid, or the nicotinamide adenine dinucleotide phosphate oxidase inhibitors apocynin or diphenylene iodonium. Vascular oxidative stress generation (fluorescent dihydroethidium), COX-1 and COX-2 expression (Western blot), and ...

TY - JOUR. T1 - Association of polymorphism of the p22PHOX component of NADPH oxidase in gastroduodenal diseases in Japan. AU - Tahara, Tomomitsu. AU - Shibata, Tomoyuki. AU - Wang, Fangyu. AU - Nakamura, Masakatsu. AU - Okubo, Masaaki. AU - Yoshioka, Daisuke. AU - Sakata, Mikijyu. AU - Nakano, Hiroshi. AU - Hirata, Ichiro. AU - Arisawa, Tomiyasu. PY - 2009/3. Y1 - 2009/3. N2 - Objective. Superoxide has been implicated in the pathogenesis of Helicobacter pylori-related diseases through inflammation. NADPH oxidase, a major source of superoxide generation, plays a critical role in H. pylori-related gastric inflammation. The aim of this study was to clarify the effect of the p22PHOX C242T polymorphism, an essential component of NADPH oxidase in the risk of gastroduodenal diseases, on the severity of H. pylori-induced gastritis in a Japanese population. Material and methods. The study comprised 436 patients attending the Endoscopy Center of Fujita Health University Hospital. The p22PHOX C242T ...

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Reactive oxygen species (ROS) produced by respiratory burst oxidase homologs (RBOHs) regulate numerous plant cell processes, including the symbiosis between legumes and nitrogen-fixing bacteria. Rapid and transient ROS production was reported after Phaseolus vulgaris root hairs were treated with Nod factors, indicating the presence of a ROS-associated molecular signature in the symbiosis signaling pathway. Rboh is a multigene family containing nine members (RbohA−I) in P. vulgaris. RNA interference of RbohB suppresses ROS production and attenuates rhizobial infection thread (IT) progression in P. vulgaris root hairs. However, the roles of other Rboh members in symbiotic interactions are largely unknown. In this study, we characterized the role of the NADPH oxidase-encoding gene RbohA (Phvulv091020621) in the P. vulgaris-Rhizobium tropici symbiosis. The spatiotemporal activity of the RbohA promoter colocalized with growing ITs and was associated with vascular bundles in developing nodules. Subcellular

The plasma membrane-bound peroxidases (PRX) zmprx01, zmprx66 and zmprx70 and the respiratory burst oxidase homologs (RBOH) rbohA, rbohB, rbohC and rbohD were analysed in this study. The distribution of the genes inside the roots was investigated by real-time qPCR. Therefor four different segments (root tip, elongation zone, differentiation zone and lateral roots) were in focus of the analyses. It could be observed that the genes are differently distributed in the root. The peroxidases were predominantly expressed in the elongation zone and almost not in the root tip. The rboh genes were more inhomogeneous distributed. For each RBOH a specific expression pattern could be detected. rbohA was mostly expressed in the differentiation zone. rbohB was more even expressed in the root. rbohC was even distributed as well but predominantly in the elongation zone. rbohD was mostly expressed in the differentiation zone. For a further investigation of the peroxidases plants were exposed to cadmium (short term and

TY - JOUR. T1 - NADPH oxidase inhibitors. T2 - A decade of discovery from Nox2ds to HTS. AU - Cifuentes-Pagano, Eugenia. AU - Csanyi, Gabor. AU - Pagano, Patrick J.. PY - 2012/7/1. Y1 - 2012/7/1. N2 - NADPH oxidases (Nox) are established as major sources of reactive oxygen species (ROS). Over the past two decades, Nox-derived ROS have emerged as pivotal in the development of myriad diseases involving oxidative stress. In contrast, Nox are also involved in signaling mechanisms necessary for normal cell function. The study of these enzymes in physiological and pathophysiological conditions is made considerably more complex by the discovery of 7 isoforms: Nox1 through 5 as well as Duox1 and 2, each with its own specific cytosolic components, regulatory control mechanisms, subcellular localization and/or tissue distribution. A clear understanding of the role individual isoforms play in a given system is hindered by the lack of isoform-specific inhibitors. In animal models, knockdown or knockout ...

Increasing experimental evidence suggests that non-phagocytic cells express a potent superoxide (O2-•)-producing NADH oxidase that might be related to the phagocytic NADPH oxidase. Here we show that the cytokine tumour necrosis factor α (TNF-α) activates, in a time- and dose-dependent manner, a O2-•-producing NADH oxidase in cultured rat aortic smooth-muscle cells. Dose-response experiments for NADH showed an upward shift of the curve for TNF-α-treated cells, suggesting that TNF-α increased the amount of available enzyme. Using the anti-sense transfection technique, we further demonstrate that the molecular identity of this oxidase includes p22phox (the α subunit of cytochrome b558 and part of the electron transfer component of the phagocytic NADPH oxidase), which we recently cloned from a rat vascular smooth-muscle cell cDNA library. In addition, prolonged treatment with TNF-α increased p22phox mRNA expression without affecting p22phox mRNA stability, and only when transcriptional ...

Both cis and trans unsaturated fatty acids and sodium dodecyl sulfate activated NADPH oxidase in plasma membranes of human neutrophils in the presence of neutrophil cytosol. In contrast, 5,8,11,14-icosatetraynoic acid, saturated fatty acids, esters, peroxides and 4 beta-phorbol 12-myristate 13-acetate, a potent activator of protein kinase C, were inactive. 5,8,11,14-icosatetraynoic acid inhibited superoxide formation elicited by fatty acids. Guanosine 5[gamma-thio]triphosphate (GTP[gamma S]), a potent activator of guanine-nucleotide-binding proteins (N-proteins) enhanced superoxide formation elicited by fatty acids up to fourfold, supporting our previous suggestion that NADPH oxidase is regulated by an N-protein [Seifert, R. et al. (1986) FEBS Lett. 205, 161-165]. Cytosols from various tissues, soybean lipoxygenase and protein kinase C, purified from chicken stomach, did not substitute neutrophil cytosol. The activity of neutrophil cytosol was destroyed by heating at 95 degrees C. Superoxide ...

|p|Neutrophil cytosol factor 2 is the 67-kilodalton cytosolic subunit of the multi-protein complex known as NADPH oxidase found in neutrophils. This oxidase produces a burst of superoxide which is delivered to the lumen of the neutrophil phagosome. Mutations in this gene, as well as in other NADPH oxidase subunits, can result in chronic granulomatous disease.|/p||p|Using neutrophils from a patient with p67-deficient CGD, Okamura et al. (1990) confirmed that the p67 protein functions in the respiratory burst mediated by NADPH oxidase and that p67 may be complexed to p47-phox (NCF1) while it participates in oxidase activation.|/p|

The role of reactive oxygen species (ROS) in the regulation of signal transduction processes has been well established in many cell types and recently the fine tuning of redox signalling in neurons received increasing attention. With regard to this, the involvement of NADPH oxidase (NOX) in neuronal pathophysiology has been proposed but deserves more investigation. In the present study, we used SH-SY5Y neuroblastoma cells to analyse the role of NADPH oxidase in retinoic acid (RA)-induced differentiation, pointing out the involvement of protein kinase C (PKC) delta in the activation of NOX. Retinoic acid induces neuronal differentiation as revealed by the increased expression of MAP2, the decreased cell doubling rate, and the gain in neuronal morphological features and these events are accompanied by the increased expression level of PKC delta and p67(phox), one of the components of NADPH oxidase. Using DPI to inhibit NOX activity we show that retinoic acid acts through this enzyme to induce ...

The neutrophil cytosolic factor 1 (NCF1) gene encodes the 47 kDa cytosolic subunit of neutrophil NADPH oxidase, which produces superoxide anion. The NCF1 gene is located in close proximity to two highly similar, multi-exon pseudogenes at chromosome 7q11.23, corresponding to this gene record and GeneID:654816. The two pseudogenes contain a dinucleotide deletion (delta-GT) in exon 2 that results in a frameshift and truncation of the open reading frame, and neither pseudogene is likely to express a protein. Recombination events between the pseudogenes and the functional NCF1 gene can inactivate the NCF1 gene and result in chronic granulomatous disease. [provided by RefSeq, Nov 2009]

Neutrophil cytosol factor 4 is a protein that in humans is encoded by the NCF4 gene. The protein encoded by this gene is a cytosolic regulatory component of the superoxide-producing phagocyte NADPH-oxidase, a multicomponent enzyme system important for host defense. This protein is preferentially expressed in cells of myeloid lineage. It interacts primarily with neutrophil cytosolic factor 2 (NCF2/p67-phox) to form a complex with neutrophil cytosolic factor 1 (NCF1/p47-phox), which further interacts with the small G protein RAC1 and translocates to the membrane upon cell stimulation. This complex then activates flavocytochrome b, the membrane-integrated catalytic core of the enzyme system. The PX domain of this protein can bind phospholipid products of the PI(3) kinase, which suggests its role in PI(3) kinase-mediated signaling events. The phosphorylation of this protein was found to negatively regulate the enzyme activity. Alternatively spliced transcript variants encoding distinct isoforms have ...

PKC regulation of neutrophil NADPH oxidase was suggested 2 decades ago when it was shown to be stimulated by phorbol ester.30 It is possible that neutrophils or monocytes contribute to vascular oxidative stress. Thus, increased NADPH oxidase subunit expression in monocytes was demonstrated in diabetic patients and was partly normalized after lowering of plasma glucose.31 Vascular homologues of neutrophil NADPH oxidase are sources of a major part of ROS production in VSMC and endothelial cells during physiological and pathological conditions.29 However, this section focuses on regulation of vascular NAD(P)H oxidase.. In VSMC, angiotensin II stimulates vascular NAD(P)H oxidase.32 Angiotensin II is involved in the development of cardiovascular disease in people with and without diabetes, perhaps through mechanisms independent of hypertension.33 A very rapid angiotensin II-stimulated activation of NAD(P)H oxidase involving PKC may cause redox activation of c-Src, leading to a more sustained ...

The outdated idea that reactive oxygen species (ROS) are only dangerous products of cellular metabolism, causing toxic and mutagenic effects on cellular components, is being replaced by the view that ROS have several important functions in cell signaling. In aerobic organisms, ROS can be generated from different sources, including the mitochondrial electron transport chain, xanthine oxidase, myeloperoxidase, and lipoxygenase, but the only enzyme family that produces ROS as its main product is the NADPH oxidase family (NOX enzymes). These transfer electrons from NADPH (converting it to NADP−) to oxygen to make O2•−. Due to their stability, the products of NADPH oxidase, hydrogen peroxide, and superoxide are considered the most favorable ROS to act as signaling molecules. Transcription factors that regulate gene expression involved in carcinogenesis are modulated by NADPH oxidase, and it has emerged as a promising target for cancer therapies. The present review discusses the mechanisms by which

Over the last decade, the NADPH oxidases have been recognized as an important source of reactive oxygen species (ROS) involved in both normal physiological redox-dependent processes, and in the generation of oxidative stress, with a role in the development of cardiovascular disease. As a multisubunit enzyme complex, the NADPH oxidases have the unusual composition of multiple interchangeable homologs of the catalytic, or NADPH oxidase (Nox), subunit that directly interacts with membrane-bound p22phox. It is at the Nox subunit that NADPH binding leads to electron transfer via heme groups to oxygen resulting in the generation of superoxide anion. Recent investigations have identified 7 distinct Nox subunits, some possessing multiple splice variants, but the real challenge has been to characterize the specific cellular functions and the regulation of the various Nox enzymes. The difficulty in doing so is partly attributable to the presence of multiple Nox isoforms within cells, the low and transient ...

The present study demonstrated that the immunoreactivity of CRP was observed in DCA specimens, especially in atheromatous and thrombotic tissue, suggesting that the histological characteristics had profound influences on the immunoreactivity of CRP. In addition, DCA specimens obtained from patients with higher serum levels of CRP or fibrinogen had more intensive immunoreactivity of CRP, suggesting that clinical markers of cardiac risk are correlated with deposition of CRP in the vascular bed. Taken together, histological as well as clinical features of instability of coronary plaque are implicated in enhanced immunoreactivity of CRP in DCA specimen. Furthermore, the localization of CRP was closely associated with NADH/NADPH oxidase p22phox, and CRP directly enhanced the expression of NADH/NADPH oxidase p22phox protein as well as the generation of ROS in cultured human CASMCs. These results suggest that CRP accumulating in coronary beds might enhance vascular oxidative stress via p22phox-based ...

Cardiovascular diseases are associated with the induction of endoplasmic reticulum (ER) stress. The induction of ER stress in C57BL/6J and p47phox-/- mice, by the injection of tunicamycin, greatly impaired vascular endothelium-dependent relaxation in C57BL/6J than in p47phox-/- mice. To determine the mechanism by which ER stress impairs endothelium function, we incubated mice primary endothelial cells from coronary arteries (ECs) with tunicamycin (1 μg/mL) for 6 h in the presence or absence of two ER stress inhibitors (Tudca, 500 μg/mL and PBA, 10 mM). Tunicamycin increased the phosphorylation of PERK and eIF2alpha, the expression of CHOP, ATF6 and Bip, Nox2/Nox4 mRNA levels, NADPH oxidase activity and superoxide anion levels. In addition, phosphorylated eNOS and nitrites levels were reduced with tunicamycin. All these events were prevented, at least partially, with the inhibition of ER stress. Tunicamycin treatment or the transfection of ECs with plasmids that express ATF6 or CHOP reduced the ...

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in Talanta (2015). Excessive neutrophil stimulation and reactive oxygen species (ROS) production are involved in numerous human or horse pathologies. The modulation of the neutrophil NADPH oxidase (NOX) has a great ... [more ▼]. Excessive neutrophil stimulation and reactive oxygen species (ROS) production are involved in numerous human or horse pathologies. The modulation of the neutrophil NADPH oxidase (NOX) has a great therapeutic potential since this enzyme produces superoxide anion whose most of the other ROS derive. The measurement of NOX activity by cell-free systems is often used to test potential inhibitors of the enzyme. A major drawback of this technique is the possible interferences between inhibitors and the probe, ferricytochrome c, used to measure the activity. We designed the EquiNox2, a new pharmacological tool, to determine the direct interaction of potential inhibitors with equine phagocytic NOX and their effect on the enzyme activity or assembly. This method consists in ...

Tumor necrosis factor-alpha (TNF-alpha) triggers activation of cytosolic phospholipase A2 (cPLA2) and then enhancing the synthesis of prostaglandin (PG) in inflammatory diseases. However, the detailed mechanisms of TNF-alpha induced cPLA2 expression were not fully defined in human pulmonary alveolar epithelial cells (HPAEpiCs). We found that TNF-alpha-stimulated increases in cPLA2 mRNA (5.2 folds) and protein (3.9 folds) expression, promoter activity (4.3 folds), and PGE2 secretion (4.7 folds) in HPAEpiCs, determined by Western blot, real-time PCR, promoter activity assay and PGE2 ELISA kit. These TNF-alpha-mediated responses were abrogated by the inhibitors of NADPH oxidase [apocynin (APO) and diphenyleneiodonium chloride (DPI)], ROS [N-acetyl cysteine, (NAC)], NF-kappaB (Bay11-7082) and transfection with siRNA of ASK1, p47phox, TRAF2, NIK, IKKalpha, IKKbeta, or p65. TNF-alpha markedly stimulated NADPH oxidase activation and ROS including superoxide and hydrogen peroxide production which were inhibited

Chemicals. Ang II, N-acetylcysteine (NAC), DPI, and PD123319 were purchased from Sigma-Aldrich (St. Louis, Missouri, USA). Losartan was kindly provided by Merck & Co. (Rahway, New Jersey, USA). 2′,7′-Dichlorofluorescein diacetate (DCFDA) was obtained from Molecular Probes Inc. (Eugene, Oregon, USA). PD98059, SB203580, and LY294002 were purchased from Calbiochem-Novabiochem Corp. (La Jolla, California, USA). SP600125 was obtained from Celgene Corp. (San Diego, California, USA).. Animals and treatments. NADPH oxidase-deficient (p47phox-/-) C57BL/6 mice, which lack a critical cytosolic component required for assembly of an active NADPH oxidase complex, were expanded in parallel with parental inbred WT mice (30, 33). Two-month-old female mice were used for HSC isolations and for experimental liver fibrosis. Liver fibrosis was induced by bile duct ligation. Briefly, mice were anesthetized with sodium pentobarbital. After midline laparotomy, the common bile duct was doubly ligated with 4-0 silk ...

ENCODES a protein that exhibits superoxide-generating NADPH oxidase activator activity (inferred); INVOLVED IN phagocytosis (inferred); positive regulation of catalytic activity (inferred); respiratory burst (inferred); PARTICIPATES IN Leishmaniasis pathway; ASSOCIATED WITH 1q24 Deletion Syndrome (ortholog); Acute Otitis Media (ortholog); Alzheimers disease (ortholog); FOUND IN NADPH oxidase complex (inferred)

The phagocyte NADPH oxidase catalyzes the reduction of O-2 to reactive oxygen species with microbicidal activity. It is composed of two membrane-spanning subunits, gp91-phox and p22-phox (encoded by CYBB and CYBA, respectively), and three cytoplasmic subunits, p40-phox, p47-phox, and p67-phox (encoded by NCF4, NCF1, and NCF2, respectively). Mutations in any of these genes can result in chronic granulomatous disease, a primary immunodeficiency characterized by recurrent infections. Using evolutionary mapping, we determined that episodes of adaptive natural selection have shaped the extracellular portion of gp91-phox during the evolution of mammals, which suggests that this region may have a function in host-pathogen interactions. On the basis of a resequencing analysis of approximately 35 kb of CYBB, CYBA, NCF2, and NCF4 in 102 ethnically diverse individuals (24 of African ancestry, 31 of European ancestry, 24 of Asian/Oceanians, and 23 US Hispanics), we show that the pattern of CYBA diversity is ...

NADPH oxidases are major sources of reactive oxygen species (ROS) involved in the pathophysiology of many cardiovascular diseases, acting through the modulation of redox-sensitive signalling pathways as well as the inactivation of nitric oxide and generation of endothelial dysfunction. There are five oxidase isoforms (Nox1 - Nox5), which have different tissue distributions, are subject to stimulus-specific activation and may exert distinct downstream effects. Endothelial cells co-express two of these isoforms, the classical Nox isoform, Nox2, and a novel isoform, Nox4. Recent studies indicate that Nox4 generates predominantly hydrogen peroxide (H2O2) but its role in vivo remains unclear. To address this question, transgenic mice with endothelial-specific overexpression of Nox4 were studied as an approach to investigate the in vivo functional significance of Nox4 in the endothelium. Unexpectedly, Nox4 overexpression was found to significantly enhance acetylcholine-induced vasodilatation compared ...

NADPH oxidase 4 gene is mapped on chromosome 11q14.2-q21 upregulated in lungs to non-infectious injury and idiopathic pulmonary fibrosis. It functions as an oxygen sensor regulating the KCNK3 potassium channel and HIF1A activity and regulatesinsulin signaling cascade as well as role in apoptosis,bone resorption and lipopolysaccharide-mediated activation of NFKB. NADPH oxidase 4 is highly expressed at the distal tubular cells in kidney cortex with a lower amount inheart, hepatoma, adipocytes,endothelial cells, skeletal muscle, brain, brain tumor cell lines and airway epithelial cells that appear to localized at central adhesion within the nucleus or in the endoplasmic reticulum thus constitutively active to functions in resting cellular homeostasis. It produces superoxide in the nucleus and play in regulating gene expression upon cell stimulation. NADPH oxidase 4 has been implicated in the differentiation of cardiac fibroblasts to myofibroblasts and of the most induced genes in fetal lung ...

In the current studies, we used animals deficient in NADPH oxidase (28), the major pathway by which phagocytes produce H2O2 (19), to show that oxidants derived from NADPH oxidase contribute to CML formation in vivo. First, we showed that neutrophils isolated from wild-type mice, but not those from mice deficient in NADPH oxidase (CGD mice), converted l-serine into glycolaldehyde. The peroxide scavenger catalase inhibited glycolaldehyde production by wild-type neutrophils. These results indicate that phagocytes generate glycolaldehyde by a reaction dependent on H2O2 produced by the phagocyte NADPH oxidase. Second, CML was produced on RNase A when the protein was exposed to activated wild-type neutrophils in a physiological salt solution, which contained the seven most common plasma amino acids (including l-serine). In contrast, only low levels of CML were formed when we exposed RNase A to neutrophils isolated from the CGD mice. This indicates that oxidants generated by NADPH oxidase are necessary ...

NADPH oxidases are a family of enzymes that generate reactive oxygen species (ROS). The NOX1 (NADPH oxidase 1) and NOX2 oxidases are the major sources of ROS in the artery wall in conditions such as hypertension, hypercholesterolaemia, diabetes and ageing, and so they are important contributors to the oxidative stress, endothelial dysfunction and vascular inflammation that underlies arterial remodelling and atherogenesis. In this Review, we advance the concept that compared to the use of conventional antioxidants, inhibiting NOX1 and NOX2 oxidases is a superior approach for combating oxidative stress. We briefly describe some common and emerging putative NADPH oxidase inhibitors. In addition, we highlight the crucial role of the NADPH oxidase regulatory subunit, p47phox, in the activity of vascular NOX1 and NOX2 oxidases, and suggest how a better understanding of its specific molecular interactions may enable the development of novel isoform-selective drugs to prevent or treat cardiovascular ...

NADPH oxidases contribute to brain injury, yet they may also have a role in brain repair, particularly in vascular signaling and angiogenesis. This study determined the temporal and spatial profile of NADPH oxidase subunit expression/activity concurrently with angiogenesis in the brain following transient ischemic stroke induced by prolonged constriction of the middle cerebral artery by perivascular injection of endothelin-1 in conscious Hooded Wistar rats (n = 47). VEGF mRNA expression was increased in the ipsilateral cortex and striatum between 6 h and 28 days post-stroke concurrently with a marked increase in Nox2 mRNA expression up to 7 days, and increased Nox4 mRNA expression detected between 7 and 28 days. Point counting of blood vessels using Metamorph imaging software showed increased vascular sprouting between 3 and 7 days after stroke with new vascular networks detected in the core infarct region by 14 days. Angiogenic blood vessels 3 and 7 days post-stroke were observed to co-localise with

Production of reactive oxygen species, often by NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidases, plays a role in the signaling responses of cells to many receptor stimuli. Here, we describe the function of the calcium-dependent, nonphagocytic NADPH oxidase Duox1 in primary human CD4+ T cells and cultured T cell lines. Duox1 bound to inositol 1,4,5-trisphosphate receptor 1 and was required for early T cell receptor (TCR)-stimulated production of hydrogen peroxide (H2O2) through a pathway that was dependent on TCR-proximal kinases. Transient or stable knockdown of Duox1 inhibited TCR signaling, especially phosphorylation of tyrosine-319 of ζ chain-associated protein kinase of 70 kilodaltons (ZAP-70), store-operated entry of calcium ions (Ca2+), and activation of extracellular signal-regulated kinase. The production of cytokines was also inhibited by knockdown of Duox1. Duox1-mediated inactivation of Src homology 2 domain-containing protein tyrosine phosphatase 2 ...

TY - CONF. T1 - HYPERGLYCAEMIA ALTERS NOX4 NADPH OXIDASE-MEDIATED ENDOTHELIAL CELL REGULATION IN VITRO. AU - Gill, Eleanor. AU - Edgar, Kevin. AU - Wilson, Adam J. AU - Patterson, Ellen. AU - Grieve, David. PY - 2018/6/6. Y1 - 2018/6/6. M3 - Abstract. T2 - British Cardiovascular Society Annual Conference. Y2 - 4 June 2018 through 5 June 2018. ER - ...

8-Hydroxy-2-deoxyguanosine (8-OHdG), a marker of oxidative stress, has been recently rediscovered to inhibit Rac1 in neutrophils and macrophages, thereby inhibiting Rac1-linked functions of these cells, including reactive oxygen species production through NADPH oxidase activation, phagocytosis, chemotaxis, and cytokine release. In vascular smooth muscle cells (VSMCs), reactive oxygen species also induce abnormal proliferation and migration leading to progression of atherosclerosis. Based upon the involvement of reactive oxygen species in phagocytic cells and VSMCs during the atherosclerotic process, we hypothesized that 8-OHdG could have antiatherosclerotic action and tested this hypothesis in an experimentally induced atherosclerosis in mice. Partially ligated ApoE knockout mice, a more physiologically relevant model of low and oscillatory flow, developed an advanced lesion in 2 weeks, and orally administered 8-OHdG significantly reduced plaque formation along with reduced superoxide formation, ...

Key Points. Sickle RBC ROS production is mediated in part by NADPH oxidase activity.Sickle RBC ROS production can be induced by plasma signaling molecules.

TY - JOUR. T1 - Apocynin prevents cyclooxygenase 2 expression in human monocytes through NADPH oxidase and glutathione redox-dependent mechanisms. AU - Barbieri, Silvia S.. AU - Cavalca, Viviana. AU - Eligini, Sonia. AU - Brambilla, Marta. AU - Caiani, Alessia. AU - Tremoli, Elena. AU - Colli, Susanna. PY - 2004/7/15. Y1 - 2004/7/15. N2 - In the present study we report the preventive effect of apocynin, an active constituent of the Himalayan herb Picrorhiza kurrooa, on cyclooxygenase-2 (Cox-2) synthesis and activity in human adherent monocytes exposed to serum treated zymosan (STZ) and phorbol myristate acetate (PMA). Apocynin markedly decreases the intracellular reduced/oxidized glutathione ratio (GSH/GSSG) and prevents nuclear factor-κB (NF-κB) activation in stimulated monocytes. Moreover, it reduces intracellular reactive oxygen species (ROS) generation, NADPH oxidase activity in monocyte homogenates and translocation of p47phox subunit in monocyte membranes. p47phox levels are also reduced ...

During pneumococcal pneumonia, NADPH oxidase-derived reactive oxygen species are redundant for host defense but limit neutrophil recruitment and survival. Decreased NADPH oxidase-dependent reactive oxygen species production is well tolerated and improves disease outcome during pneumococcal pneumonia by removing neutrophils from the tight constraints of reactive oxygen species-mediated regulation.. ...

TY - JOUR. T1 - Lipoteichoic acid induces HO-1 expression via the TLR2/MyD88/c-Src/NADPH oxidase pathway and Nrf2 in human tracheal smooth muscle cells. AU - Lee, I-Ta. AU - Wang, Shyi Wu. AU - Lee, Chiang Wen. AU - Chang, Chia Chi. AU - Lin, Chih Chung. AU - Luo, Shue Fen. AU - Yang, Chuen Mao. PY - 2008/10/1. Y1 - 2008/10/1. N2 - Heme oxygenase (HO)-1 is a stress-inducible rate-limiting enzyme in heme degradation that confers cytoprotection against oxidative injury and provides a vital function in maintaining tissue homeostasis. Increasing reports have indicated that lipoteichoic acid (LTA) exerts as LPS as an immune system-stimulating agent and plays a role in the pathogenesis of severe inflammatory responses induced by Gram-positive bacterial infection. We report that LTA is an inducer of HO-1 expression mediated through the signaling pathways in human tracheal smooth muscle cells (HTSMCs). LTA-induced HO-1 protein levels, mRNA expression, and promoter activity were attenuated by ...

Diabetic cardiovascular disease is associated with decreased adiponectin and increased oxidative stress. This study investigated the effect of telmisartan on the expression of adiponectin receptor 2 (adipoR2) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits in the heart and the expression of adiponectin receptor 1 (adipoR1) in aorta in type 2 diabetic rats. Type 2 diabetes was induced by high-fat and high-sugar diet and intraperitoneal injection of a low dose of streptozotocin (STZ). Heart function, adipoR2, p22phox, NOX4, glucose transporter 4(GLUT4), monocyte chemoattractant protein-1(MCP-1) and connective tissue growth factor (CTGF)in the heart, and adipoR1, MCP-1 and nuclear factor kappa B (NF-κB) in aorta were analyzed in controls and diabetic rats treated with or without telmisartan (5mg/kg/d) by gavage for 12 weeks. Heart function, plasma and myocardial adiponectin levels, the expression of myocardial adipoR2 and GLUT4 were significantly decreased in diabetic rats (P |0

Previous studies from this laboratory described the kinetic characteristics of the inhibition by tosylphenylalanine chloromethane (TosPheCH2Cl) on superoxide anion production by human neutrophils (PMN) stimulated with a phorbol ester (PMA). In this study we present further evidence concerning the potential role of the chloromethane target in the normal cellular activation of NADPH oxidase. When PMN are treated with TosPheCH2Cl and subsequently PMA, or with the two reagents in the reverse order, the inhibition of superoxide production by the intact cells is still present in a particulate NADPH oxidase fraction prepared from these cells. Nevertheless, when cells incubated only with the chloromethane and not with PMA are disrupted, both their cytosolic and membrane fractions are fully competent in the cell-free activation assay. Thus, the chloromethane target has a role in NADPH oxidase activation exclusively at the cellular level. This observation constitutes additional evidence in favour of the ...

Neutrophil extracellular traps (NETs) play an important role in innate immunity to microbial infections. NETs have been described in several species, but the molecular details of NET formation and their role in infection has not been addressed, partly because we lack optimal experimental models. Here we describe tools to investigate NET formation in neutrophils isolated from mice. Upon in vitro stimulation of wild-type mouse neutrophils with PMA, we analyzed 3 important steps in the process of NET formation: reactive oxygen species (ROS) production, NET cell death and NET release. As expected, neutrophils from NADPH oxidase-deficient mice failed to produce ROS and did not die nor release NETs upon stimulation. We found that neutrophils from several mouse strains produced NETs with different efficiency and that NET formation correlated with the amount of ROS produced. Activation with Candida albicans also resulted in ROS production and NET cell death. The hyphal form of this fungus induced NETs ...

TY - JOUR. T1 - NADPH oxidases in traumatic brain injury - Promising therapeutic targets?. AU - Ma, Merry W.. AU - Wang, Jing. AU - Dhandapani, Krishnan Michael. AU - Wang, Ruimin. AU - Brann, Darrell W. PY - 2018/6/1. Y1 - 2018/6/1. N2 - Traumatic brain injury (TBI) is a major cause of death and disability worldwide. Despite intense investigation, no neuroprotective agents for TBI have yet translated to the clinic. Recent efforts have focused on identifying potential therapeutic targets that underlie the secondary TBI pathology that evolves minutes to years following the initial injury. Oxidative stress is a key player in this complex cascade of secondary injury mechanisms and prominently contributes to neurodegeneration and neuroinflammation. NADPH oxidase (NOX) is a family of enzymes whose unique function is to produce reactive oxygen species (ROS). Human post-mortem and animal studies have identified elevated NOX2 and NOX4 levels in the injured brain, suggesting that these two NOXs are ...

Chronic granulomatous disease What every physician needs to know: Background Chronic granulomatous disease (CGD) is a deficiency of microbicidal oxidant production affecting neutrophils, eosinophils, monocytes, and some tissue macrophages. There are five genetic forms of CGD affecting different subunits of the phagocyte NADPH (nicotinamide adenine dinucleotide phosphate) oxidase (gp91phox, p47phox, p22phox, p67phox, p40phox). X-linked CGD…. ...

Background: The X-linked form of chronic granulomatous disease (CGD) is a primary immunodeficiency that affects phagocytes of the innate immune system and is characterized by an increased susceptibility to severe bacterial and fungal infections. It is caused by mutations in the CYBB gene, which encodes the 91-kD subunit of phagocyte NADPH oxidase. Aim: To identify the mutation in the CYBB gene in two unrelated patients from Chile with, the diagnosis of X-linked CGD and their families. Patients and methods: The molecular genetic defects of two unrelated patients from Chile with X-linked CGD caused by defects in the CYBB gene were investigated. The underlying mutation was investigated by single strand conformation polymorphism (SSCP) analysis of PCR-amplified genomic DNA and by sequencing of the affected gene region. Results: We found an insertion c.1267_1268insA in exon 10 leading to a frameshift mutation. This mutation is a novel report. We also identified a splice site mutation in the other ...

Our β-Nicotinamide-Adenine Dinucleotide Phosphate, Monopotassium Salt (β-NADP-K) is applicable for making Creatine Kinase N-acetylcysteine (CK-NAC), Creatine Kinase Myoglobin (CK-MB), and Glucose reagents ...

Hypoxia-inducible factor 1 (HIF-1) mediates many of the systemic and cellular responses to intermittent hypoxia (IH), which is an experimental model that simulates O2 saturation profiles occurring with recurrent apnea. IH-evoked HIF-1α synthesis and stability are due to increased reactive oxygen species (ROS) generated by NADPH oxidases, especially Nox2. However, the mechanisms by which IH activates Nox2 are not known. We recently reported that IH activates xanthine oxidase (XO) and the resulting increase in ROS elevates intracellular calcium levels. Since Nox2 activation requires increased intracellular calcium levels, we hypothesized XO-mediated calcium signaling contributes to Nox activation by IH. We tested this possibility in rat pheochromocytoma PC12 cells subjected to IH consisting alternating cycles of hypoxia (1.5% O2 for 30 sec) and normoxia (21% O2 for 5 min). Kinetic analysis revealed that IH-induced XO preceded Nox activation. Inhibition of XO activity either by allopurinol or by siRNA

NADPH-oxidases (Nox) and the related Dual oxidases (Duox) play varied biological and pathological roles via regulated generation of reactive oxygen species (ROS). Members of the Nox/Duox family have been identified in a wide variety of organisms, including mammals, nematodes, fruit fly, green plants, fungi, and slime molds; however, little is known about the molecular evolutionary history of these enzymes. We assembled and analyzed the deduced amino acid sequences of 101 Nox/Duox orthologs from 25 species, including vertebrates, urochordates, echinoderms, insects, nematodes, fungi, slime mold amoeba, alga and plants. In contrast to ROS defense enzymes, such as superoxide dismutase and catalase that are present in prokaryotes, ROS-generating Nox/Duox orthologs only appeared later in evolution. Molecular taxonomy revealed seven distinct subfamilies of Noxes and Duoxes. The calcium-regulated orthologs representing 4 subfamilies diverged early and are the most widely distributed in biology. Subunit

Serum amyloid A (SAA) amounts are elevated highly in acute phase response and elevated slightly and persistently in chronic diseases such as rheumatoid arthritis and diabetes. findings support the notion that, in chronic inflammatory sites, SAA activated fibroblast proliferation and ROS production. 0001 for comparison between control and SAA treatment. ## 001 for comparison between SAA and SAA plus DPI treatments. There was a positive correlation between SAA concentration and O2- production (Fig. 1b), and Pearsons correlation found = 099 and = 0001 (Fig. 1a). ROS production seems to occur, at least in part, due to activation of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enzymes, as diphenyleneiodonium (DPI), a specific inhibitor of NADPH oxidase, reduced O2- production induced by SAA by 50% (Fig. 1c). Also, the expression of p47-was up-regulated by SAA, as evaluated by qPCR (Table 1). The mechanism of O2- production by a membrane-bound NADPH oxidase entails translocation of ...

The major findings in the present study include the following: (1) vascular superoxide is responsible for the selective impairment of endothelium-dependent dilation after H/R in isolated cerebral arteries; (2) apocynin-sensitive NADPH oxidase in the cerebral vascular wall is a potential source of superoxide production; and (3) our data are consistent with activation of NF-κB playing a role in H/R-induced cerebral endothelial dysfunction. To our best knowledge, this is the first study to document that NF-κB activation in response to H/R may contribute to functional regulation of resistance arteries from cerebral circulation.. It is likely that intracellular O2− is the primary mediator of H/R-induced cerebral endothelial dysfunction. Endothelial NADPH oxidase increases superoxide intracellularly.34,35 Furthermore, superoxide is charged and may not easily diffuse across cell membranes. Exogenous SOD enzyme activity is primarily extracellular, whereas the cell-permeable SOD mimetic Tempo is ...

We aimed to investigate specific roles of mitogen-activated protein kinases (MAPK) in the deterioration of endothelial function during the progression of diabetes and the potential therapeutic effects of MAPK inhibitors and agonists in the amelioration of endothelial function. Protein expression and phosphorylation of p38, c-Jun NH2-terminal kinase (JNK), and extracellular signal-regulated kinase (Erk) were assessed in mesenteric arteries of 3- (3M) and 9-month-old (9M) male diabetic and control mice. The expression of p38, JNK, and Erk was comparable in all groups of mice, but the phosphorylation of p38 and JNK was increased in 3M and further increased in 9M diabetic mice, whereas the phosphorylation of Erk was substantially reduced in 9M diabetic mice. NADPH oxidase-dependent superoxide production was significantly increased in vessels of two ages of diabetic mice. Inhibition of either p38 with SB203580 or JNK with SP600125 reduced superoxide production and improved shear stress-induced ...

Background: CGD is a disorder of NADPH oxidase (NADPHO) characterized by severe bacterial and fungal infections. We used p47phox-/- (CGD) mice (M) to evaluate NADPHO-independent host defense against Aspergillus fumigatus (Af). Methods: Survival, histology, fungal burden, and cytokine responses were compared in these M: CGD alone, CGD + cyclophosphamide (Cy), CGD + anti-Gr1 neutrophil depleting antibody, wildtype (WT) + Cy, and WT + anti-Gr1. Results: Susceptibility to intratracheal (IT) Af (1.25 x 104 CFU) based on mortality was: CGD + Cy , CGD alone , WT + Cy. In separate studies, M were administered IT Af (1.25 x 104 CFU) and sacrificed on day 5. CGD alone M developed focal neutrophilic infiltrates surrounding hyphae, and no vascular invasion. CGD + Cy M developed acellular coagulative necrosis and hyphal angioinvasion. CGD + anti-Gr1 M, developed histiocytic infiltrates with few neutrophils and hyphal angioinvasion. WT + Cy and WT + anti-Gr1 mice had no invasive disease. Serum galactomannan ...

The NADPH oxidase generates microbicidal superoxide in phagocytes, and when defective it leads to chronic granulomatous disease (CGD). Oxidase specific proteins in the cytosol, p47phox and p67phox, as well as the small GTP binding protein p21rac are important for activation of superoxide production. Because the activity of this oxidase is normally tightly restricted to the phagocytic vacuole, and its temporal and spatial organisation might be regulated by cytoskeletal proteins, we examined the cytosolic phox proteins for interactions with cytoskeletal elements. p67phox copurified with a 57 kDa protein, identified as coronin, an actin binding protein that is important for movement and phagocytosis in Dictyostelium. Binding studies revealed that coronin attaches to the C-terminal half of p40phox, a binding partner of p67phox. The phox proteins and coronin had a similar distribution in the cell, and both accumulated around the phagocytic vacuole. PMA activation of adherent neutrophils resulted in a ...

The NADPH oxidases are important transmembrane proteins producing reactive oxygen species (ROS). Within the Nox family, different modes of activation can be discriminated. Nox1-3 are dependent on different cytosolic subunits, Nox4 seems to be constitutively active and Nox5 is directly activated by calcium. With the exception of Nox5, all Nox family members are thought to depend on the small transmembrane protein p22phox. With the discovery of the CRISPR/Cas9-system, a tool to alter genomic DNA sequences has become available. So far, this method has not been widely used in the redox community. On such basis, we decided to study the requirement of p22phox in the Nox complex using CRISPR/Cas9-mediated knockout. Knockout of the gene of p22phox, CYBA, led to an ablation of activity of Nox4 and Nox1 but not of Nox5. Production of hydrogen peroxide or superoxide after knockout could be rescued with either human or rat p22phox, but not with the DUOX-maturation factors DUOXA1/A2. Furthermore, different ...

TY - JOUR. T1 - Endothelial Nox4-based NADPH oxidase regulates atherosclerosis via soluble epoxide hydrolase. AU - Hu, Pingping. AU - Wu, Xiaojuan. AU - Khandelwal, Alok R.. AU - Yu, Weimin. AU - Xu, Zaicheng. AU - Chen, Lili. AU - Yang, Jian. AU - Weisbrod, Robert M.. AU - Lee, Kin Sing Stephen. AU - Seta, Francesca. AU - Hammock, Bruce D.. AU - Cohen, Richard A.. AU - Zeng, Chunyu. AU - Tong, Xiaoyong. PY - 2017/6/1. Y1 - 2017/6/1. N2 - Nox4-based NADPH oxidase is a major reactive oxygen species-generating enzyme in the vasculature, but its role in atherosclerosis remains controversial. Objective Our goal was to investigate the mechanisms of endothelial Nox4 in regulating atherosclerosis. Approach and results Atherosclerosis-prone conditions (disturbed blood flow, type I diabetes, and Western diet) downregulated endothelial Nox4 mRNA in arteries. To address whether the downregulated endothelial Nox4 was directly involved in the development of atherosclerosis, we generated mice carrying a human ...

Poster (2013, September 11). Neutrophils (PMNs) produce reactive oxygen species (ROS) to kill pathogenic agents. After appropriate stimulation, leading to the activation of protein kinase C (PKC), the cytosolic subunits of the NADPH ... [more ▼]. Neutrophils (PMNs) produce reactive oxygen species (ROS) to kill pathogenic agents. After appropriate stimulation, leading to the activation of protein kinase C (PKC), the cytosolic subunits of the NADPH oxidase (Nox2) are phosphorylated and translocated to the membrane flavocytochrome b558, forming the active enzyme which produces superoxide anion (O2●-). From O2●- derives H2O2 used by the PMNs myeloperoxidase (MPO) to form strong oxidant species. Many human and animal pathologies with fatal issue are associated with uncontrolled activation of PMNs. The modulation of enzymes implied in ROS production is thus a primary target to manage excessive inflammatory events. For this purpose, we evaluated the effects of NDS27, a water-soluble salt of ...

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Objective: Reactive oxygen species (ROS) have been proposed as signaling molecules mediating exercise training adaptation, but the ROS source has remained unclear. This study aimed to investigate if increased NADPH oxidase (NOX)2-dependent activity during exercise is required for long-term high-intensity interval training (HIIT) in skeletal muscle using a mouse model lacking functional NOX2 complex due to absent p47phox (Ncf1) subunit expression (ncf1* mutation).. Methods: HIIT was investigated after an acute bout of exercise and after a chronic intervention (3x/week for 6 weeks) in wild-type (WT) vs. NOX2 activity-deficient (ncf1*) mice. NOX2 activation during HIIT was measured using an electroporated genetically-encoded biosensor. Immunoblotting and single-fiber microscopy was performed to measure classical exercise-training responsive endpoints in skeletal muscle.. Results: A single bout of HIIT increased NOX2 activity measured as p47-roGFP oxidation immediately after exercise but not 1 h or ...

Enzyme can be run in node using jsdom to simulate a browser. Why? 1. org Simulated Biological Fluids with Possible Application in Dissolution Testing Margareth R. It also has cool features to check the props and state of a component or call methods of class directly. bio-rad. A common pattern when testing React component methods using the AirBnB enzyme library, is to figure out what event triggers the method through normal usage of the component and simulate that event to indirectly trigger it. NADPH oxidase (nicotinamide adenine dinucleotide phosphate oxidase) is a membrane-bound enzyme complex that faces the extracellular space. simulate (click); The difference here between shallow() is you dont need to provide event object by yourself and the event can propagate. Only . A) Standard M&M Enzyme (Straight forward) E + S ES E + P Assume an enzyme (E) binds a substrate (S) and isomerizes it into a product (P) according to the equation above. They added a large molecular weight substance to ...

Here, we demonstrate that endothelial dysfunction has a profound ROS-dependent impact on the development of cardiac fibrosis, independent of cardiomyocyte hypertrophy. One mechanism underlying these effects may be the proinflammatory properties of activated endothelium. The healthy endothelium is anti-inflammatory and antithrombotic, but in pathological settings such as renin-angiotensin system activation, it becomes activated and promotes increased interaction with circulating inflammatory cells (17). Previous studies suggested that NOX2 is involved in cytokine- and AngII-induced EC activation and the expression of adhesion molecules such as VCAM-1 (18,19), as well as in enhanced monocyte binding to ECs under oscillatory shear stress (20). Here, we found that NOX2-overexpressing CMECs had a larger increase in VCAM-1 expression after AngII stimulation than did wild-type CMECs, which were associated with greater endothelial-leukocyte adhesion in an in vitro flow assay. Consistent with a similar ...

Flavocytochrome b558 of the NADPH oxidase which generates superoxide in phagocytic cells, is a α1β1 heterodimer of gp91phox and p22phox, which together form a membrane-spanning electron-transport chain that transfers electrons from NADPH in the cytosol to oxygen. The C-terminal portion of gp91phox is a member of the ferredoxin-NADP+ reductase family of reductases. Little is known of the organization of the N-terminal section of this molecule, which is associated with the two haem structures. It is N-glycosylated, and site-directed mutagenesis has been used to eliminate the five potential N-linked glycosylation consensus sites. Mutated cDNAs were expressed in vitro. This approach provided evidence for glycosylation of residues Asn131, Asn148 and Asn239, but not of Asn96 and Asn429.. ...

Rho GTPases are frequent targets of virulence factors as they are keystone signaling molecules. Herein, we demonstrate that AMPylation of Rho GTPases by VopS is a multifaceted virulence mechanism that counters several host immunity strategies. Activation of NFκB, Erk, and JNK kinase signaling pathways were inhibited in a VopS-dependent manner during infection with Vibrio parahaemolyticus. Phosphorylation and degradation of IKBα were inhibited in the presence of VopS as was nuclear translocation of the NFκB subunit p65. AMPylation also prevented the generation of superoxide by the phagocytic NADPH oxidase complex, potentially by inhibiting the interaction of Rac and p67. Furthermore, the interaction of GTPases with the E3 ubiquitin ligases cIAP1 and XIAP was hindered, leading to decreased degradation of Rac and RhoA during infection. Finally, we screened for novel Rac1 interactions using a nucleic acid programmable protein array and discovered that Rac1 binds to the protein C1QA, a protein ...

In this study, we demonstrate that resveratrol produced significant neuroprotection on DA neurons against LPS-induced neurotoxicity. Using multiple primary cell cultures, we have elucidated novel mechanisms and advanced our understanding of the relationship between the anti-inflammatory and the neuroprotective effects of resveratrol. First, our study showed that resveratrol protected DA neurons against LPS-induced neurotoxicity in the midbrain neuron-glia cultures through inhibiting the activation of microglia and the production of proinflammatory factors. Second, our study is the first report indicating that microglial NADPH oxidase is a major target mediating resveratrol-elicited anti-inflammation and neuroprotection. Further mechanistic study showed that resveratrol reduced the ROS production through suppressing the translocation of cytosolic subunit p47PHOX to the cell membrane. Third, our results also suggest that inhibition of NADPH oxidase by resveratrol may link to the inhibition of ...

Learn about the structure and functions of NADPH (Nicotinamide Adenine Dinucleotide Phosphate) in human body as well is process of its formation.

Reactive oxygen species (ROS) can have divergent effects in cerebral and peripheral circulations. We found that Ca(2+)-permeable transient receptor potential ankyrin 1 (TRPA1) channels were present and colocalized with NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidase 2 (NOX2), a major source of ROS, in the endothelium of cerebral arteries but not in other vascular beds. We recorded and characterized ROS-triggered Ca(2+) signals representing Ca(2+) influx through single TRPA1 channels, which we called TRPA1 sparklets. TRPA1 sparklet activity was low under basal conditions but was stimulated by NOX-generated ROS. Ca(2+) entry during a single TRPA1 sparklet was twice that of a TRPV4 sparklet and ~200 times that of an L-type Ca(2+) channel sparklet. TRPA1 sparklets representing the simultaneous opening of two TRPA1 channels were more common in endothelial cells than in human embryonic kidney (HEK) 293 cells expressing TRPA1. The NOX-induced TRPA1 sparklets activated ...

A decrease in reactive oxygen species (ROS) production has been associated with extended life span in animal models of longevity. Mice deficient in the p66Shc gene are long-lived, and their cells are both resistant to oxidative stress and produce less ROS. Our microarray analysis of p66Shc(−/−) mouse tissues showed alterations in transcripts involved in heme and superoxide production and insulin signaling. Thus, we carried out analysis of ROS production by NADPH oxidase (PHOX) in macrophages of control and p66Shc knock-out mice. p66Shc(−/−) mice had a 40% reduction in PHOX-dependent superoxide production. To confirm whether the defect in superoxide production was a direct consequence of p66Shc deficiency, p66Shc was knocked down with siRNA in the macrophage cell line RAW264, and a 30% defect in superoxide generation was observed. The pathway of PHOX-dependent superoxide generation was investigated. PHOX protein levels were not decreased in mutant macrophages; however, the rate and extent ...

OBJECTIVE: Oxidative stress is implicated in hypertension and the NADPH oxidase systems constitute the main source of superoxide in vascular wall. We searched for new polymorphisms within the CYBA promoter, the human gene that encodes the p22phox protein, and studied their potential association with essential hypertension. DESIGN: A case-control study in a random sample of the general population. METHODS: CYBA polymorphisms were determined by restriction fragment length polymorphism and allelic discrimination. NADPH oxidase activity was quantified in phagocytic cells by chemiluminescence. RESULTS: We identified three novel polymorphisms, at positions -852, -675 and -536 from the ATG codon. Only the -675(A/T) polymorphism associated with essential hypertension. The prevalence of the TT genotype and the T allele frequency were significantly higher (P < 0.05) in hypertensives than in normotensives. Furthermore, TT hypertensives exhibited higher (P < 0.05) systolic blood pressure values than TA/AA ...

In addition to granule proteins, the production of superoxide free radicals by the NADPH oxidase is an important contributor to the antimicrobial action of neutrophils. Gp91phox (CYBB), a membrane-bound component of the NADPH oxidase known to partially localize to secondary granules (19), and the other membrane-bound subunit p22phox (CYBA) were not significantly downregulated in Nbeal2-/- neutrophils (Supplemental Table 1). Interestingly, the cytosolic components of the NADPH oxidase (NCF1, NCF2, NCF4, and RAC2) were increased in Nbeal2-/- neutrophils (Figure 1D, Supplemental Table 1, and Supplemental Figure 5C).. Nbeal2-deficient neutrophils generated increased ROS in response to soluble ROS-inducing agonists, such as fMLP and PMA, or opsonized particulate zymosan, whether detected by chemiluminescent luminol (Figure 2, B-D) or lucigenin (Supplemental Figure 5D). Several factors may contribute to this enhanced phagocyte respiratory burst. First, generation of superoxide is enhanced in the ...

ALPHA CHEMIKA is a manufacturer, supplier and exporter of Nicotinamide adenine dinucleotide phosphate disodium salt based in Mumbai, Maharashtra, India.

MalaCards based summary : Chronic Granulomatous Disease, Autosomal, Due to Deficiency of Cyba, is also known as granulomatous disease, chronic, autosomal recessive, cytochrome b-negative, and has symptoms including hepatomegaly, splenomegaly and hepatic abscesses due to immunodeficiency. An important gene associated with Chronic Granulomatous Disease, Autosomal, Due to Deficiency of Cyba is CYBA (Cytochrome B-245 Alpha Chain). Affiliated tissues include neutrophil, testes and skin ...

Reactive oxygen species (ROS) produced by the NADPH oxidase RESPIRATORY BURST OXIDASE HOMOLOG (RBOH) trigger signal transduction in diverse biological processes in plants. However, the functions of RBOH homologues in rice (Oryza sativa) and other gramineous plants are poorly understood. Ethylene induces the formation of lysigenous aerenchyma, which consists of internal gas spaces created by programmed cell death of cortical cells, in roots of gramineous plants under oxygen-deficient conditions. Here, we report that, in rice, one RBOH isoform (RBOHH) has a role in ethylene-induced aerenchyma formation in roots. Induction of RBOHH expression under oxygen-deficient conditions was greater in cortical cells than in cells of other root tissues. In addition, genes encoding group I calcium-dependent protein kinases (CDPK5 and CDPK13) were strongly expressed in root cortical cells. Co-expression of RBOHH with CDPK5 or CDPK13 induced ROS production in Nicotiana benthamiana leaves. Inhibitors of RBOH ...

Semantic Scholar extracted view of Heat Stress and Ischemia/reperfusion Cause Oxidative Stress via Nadph Oxidase in Hypothalamic Neurons by Colin Rogers et al.

Aim: To examine the effect of NADPH oxidase Nox4 deletion in diabetic nephropathy (DN) using a global Nox4 deficient mice as well as podocyte specific Nox4 deficient mice. Background: Chronic kidney failure is a major complication of diabetes. However, the underlying causes remain unclear. NADPH oxidase, particularly Nox4 derived reactive oxygen species (ROS) in the kidney play a crucial role in the development and progression of diabetic nephropathy. Albuminuria is a key feature of diabetic nephropathy and podocyte injury leads to the development of albuminuria in diabetes. Methods: Nox4-/-ApoE-/- and Nox4+/+ApoE-/- mice as well as podNox4KO and floxedNox4 mice were rendered diabetic via streptozotocin injection. At week 20 urine samples were collected for the measurement of albuminuria. Animals were culled after 20 weeks and kidneys were removed for assessment of structural damage, oxidative stress markers, as well as measurement of the protein expression of extracellular matrix (ECM), ...

Pharmacological approaches also suggest that different parts of the overall ROS production in response to infection appear to be mediated by different mechanisms. Though the involvement of an NADPH oxidase has been predominant in most cases (Bolwell et al., 1998; Grant et al., 2000b; Torres and Dangl, 2005), both NADPH oxidases and cell wall peroxidases might mediate ROS production in response to the same pathogen (Grant et al., 2000a). A more detailed temporal resolution of the activity of each system may reveal that the pools of ROS produced by each mechanism do not functionally overlap. For example, differential effects of DPI on ROS accumulation during the HR- and MAMP-mediated basal defense responses were reported, with the latter being considerably less attenuated by DPI (Soylu et al., 2005). These results suggest that alternative mechanisms might be activated to produce ROS during some basal defense responses, while NADPH oxidases might have later effects following R-mediated pathogen ...