NADPH oxidase signaling and cardiac myocyte function

The NADPH oxidase family of enzymes has emerged as a major source of reactive oxygen species (ROS) that is important in diverse cellular functions including anti-microbial defence, inflammation and redox signaling. Of the five known NADPH oxidase isoforms, several are expressed in cardiovascular cells where they are involved in physiological and pathological processes such as the regulation of vascular tone, cell growth, migration, proliferation, hypertrophy, apoptosis and matrix deposition. This article reviews current knowledge regarding the role of NADPH oxidases in cardiomyocyte function in health and disease. © 2009 Elsevier Inc. All rights reserved.

The effect of growth conditions on the synthesis of Pseudomonas aeruginosa terminal oxidase.

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The upstream Variable Number Tandem Repeat polymorphism of the monoamine oxidase type A gene influences trigeminal pain-related evoked responses

Monoamines have an important role in neural plasticity, a key factor in cortical pain processing that promotes changes in neuronal network connectivity. Monoamine oxidase type A (MAOA) is an enzyme that, due to its modulating role in monoaminergic activity, could play a role in cortical pain processing. The X-linked MAOA gene is characterized by an allelic variant of length, the MAOA upstream Variable Number Tandem Repeat (MAOA-uVNTR) region polymorphism. Two allelic variants of this gene are known, the high-activity MAOA (HAM) and low-activity MAOA (LAM). We investigated the role of MAOA-uVNTR in cortical pain processing in a group of healthy individuals measured by the trigeminal electric pain-related evoked potential (tPREP) elicited by repeated painful stimulation. A group of healthy volunteers was genotyped to detect MAOA-uVNTR polymorphism. Electrical tPREPs were recorded by stimulating the right supraorbital nerve with a concentric electrode. The N2 and P2 component amplitude and latency as well as the N2-P2 inter-peak amplitude were measured. The recording was divided into three blocks, each containing 10 consecutive stimuli and the N2-P2 amplitude was compared between blocks. Of the 67 volunteers, 37 were HAM and 30 were LAM. HAM subjects differed from LAM subjects in terms of amplitude of the grand-averaged and first-block N2-P2 responses (HAM>LAM). The N2-P2 amplitude decreased between the first and third block in HAM subjects but not LAM subjects. The MAOA-uVNTR polymorphism seemed to influence the brain response in a repeated tPREP paradigm and suggested a role of the MAOA as a modulator of neural plasticity related to cortical pain processing. Monoamines have an important role in neural plasticity, a key factor in cortical pain processing that promotes changes in neuronal network connectivity. Monoamine oxidase type A (MAOA) is an enzyme that, due to its modulating role in monoaminergic activity, could play a role in cortical pain processing. © 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Endothelial Nox4 NADPH oxidase enhances vasodilatation and reduces blood pressure in vivo

Objective- Increased reactive oxygen species (ROS) production is involved in the pathophysiology of endothelial dysfunction. NADPH oxidase-4 (Nox4) is a ROS-generating enzyme expressed in the endothelium, levels of which increase in pathological settings. Recent studies indicate that it generates predominantly hydrogen peroxide (H O ), but its role in vivo remains unclear. Methods and Results- We generated transgenic mice with endothelium-targeted Nox4 overexpression (Tg) to study the in vivo role of Nox4. Tg demonstrated significantly greater acetylcholine- or histamine-induced vasodilatation than wild-type littermates. This resulted from increased H O production and H O -induced hyperpolarization but not altered nitric oxide bioactivity. Tg had lower systemic blood pressure than wild-type littermates, which was normalized by antioxidants. Conclusion- Endothelial Nox4 exerts potentially beneficial effects on vasodilator function and blood pressure that are attributable to H O production. These effects contrast markedly with those reported for Nox1 and Nox2, which involve superoxide-mediated inactivation of nitric oxide. Our results suggest that therapeutic strategies to modulate ROS production in vascular disease may need to separately target individual Nox isoforms. © 2011 American Heart Association, Inc.

NADPH oxidase and heart failure

Reactive oxygen species play important roles in the pathophysiology of chronic heart failure secondary to chronic left ventricular hypertrophy or myocardial infarction. Reactive oxygen species influence several components of the phenotype of the failing heart, including contractile function, interstitial fibrosis, endothelial dysfunction and myocyte hypertrophy. Recent studies implicate the production of reactive oxygen species by a family of NADPH oxidases in these effects. NADPH oxidases are activated in an isoform-specific manner by many pathophysiological stimuli and exert distinct downstream effects. Understanding NADPH oxidase activation and regulation, and their downstream effectors, could help to develop novel therapeutic targets.

NADPH oxidase-dependent redox signalling in cardiac hypertrophy, remodelling and failure

Markers of increased oxidative stress are known to be elevated following acute myocardial infarction and in the context of chronic left ventricular hypertrophy or heart failure, and their levels may correlate with the degree of contractile dysfunction or cardiac deficit. An obvious pathological mechanism that may account for this correlation is the potential deleterious effects of increased oxidative stress through the induction of cellular dysfunction, energetic deficit or cell death. However, reactive oxygen species have several much more subtle effects in the remodelling or failing heart that involve specific redox-regulated modulation of signalling pathways and gene expression. Such redox-sensitive regulation appears to play important roles in the development of several components of the phenotype of the failing heart, for example cardiomyocyte hypertrophy, interstitial fibrosis and chamber remodelling. In this article, we review the evidence supporting the involvement of reactive oxygen species and redox signalling pathways in the development of cardiac hypertrophy and heart failure, with a particular focus on the NADPH oxidase family of superoxide-generating enzymes which appear to be especially important in redox signalling.

Endothelial NADPH oxidase-2 promotes interstitial cardiac fibrosis and diastolic dysfunction through proinflammatory effects and endothelial-mesenchymal transition

OBJECTIVES: This study sought to investigate the effect of endothelial dysfunction on the development of cardiac hypertrophy and fibrosis. BACKGROUND: Endothelial dysfunction accompanies cardiac hypertrophy and fibrosis, but its contribution to these conditions is unclear. Increased nicotinamide adenine dinucleotide phosphate oxidase-2 (NOX2) activation causes endothelial dysfunction. METHODS: Transgenic mice with endothelial-specific NOX2 overexpression (TG mice) and wild-type littermates received long-term angiotensin II (AngII) infusion (1.1 mg/kg/day, 2 weeks) to induce hypertrophy and fibrosis. RESULTS: TG mice had systolic hypertension and hypertrophy similar to those seen in wild-type mice but developed greater cardiac fibrosis and evidence of isolated left ventricular diastolic dysfunction (p < 0.05). TG myocardium had more inflammatory cells and VCAM-1-positive vessels than did wild-type myocardium after AngII treatment (both p < 0.05). TG microvascular endothelial cells (ECs) treated with AngII recruited 2-fold more leukocytes than did wild-type ECs in an in vitro adhesion assay (p < 0.05). However, inflammatory cell NOX2 per se was not essential for the profibrotic effects of AngII. TG showed a higher level of endothelial-mesenchymal transition (EMT) than did wild-type mice after AngII infusion. In cultured ECs treated with AngII, NOX2 enhanced EMT as assessed by the relative expression of fibroblast versus endothelial-specific markers. CONCLUSIONS: AngII-induced endothelial NOX2 activation has profound profibrotic effects in the heart in vivo that lead to a diastolic dysfunction phenotype. Endothelial NOX2 enhances EMT and has proinflammatory effects. This may be an important mechanism underlying cardiac fibrosis and diastolic dysfunction during increased renin-angiotensin activation.

Evolution of protein bound Maillard reaction end-products and free Amadori compounds in low lactose milk in presence of fructosamine oxidase I

Thermal treatments and storage influence milk quality, particularly in low lactose milk as the higher concentration of reducing sugars can lead to the increased formation of the Maillard reaction products (MRPs). The control of the Amadori products (APs) formation is the key step to mitigate the Maillard reaction (MR) in milk. The use of fructosamine oxidases, (Faox) provided promising results. In this paper, the effects of Faox I were evaluated by monitoring the concentration of free and bound MRPs in low lactose milk during shelf life. Results showed that the enzyme reduced the formation of protein-bound MRPs down to 79% after six days at 37 °C. Faox I lowered the glycation of almost all the free amino acids resulting effective on basic and polar amino acids. Data here reported corroborate previous findings on the potentiality of Faox enzymes in controlling the early stage of the MR in foods.

Vitamins Reverse Endothelial Dysfunction Through Regulation of eNOS and NAD(P)H Oxidase Activities

Antioxidant vitamins C and E have protective properties in genetic hypertension associated with enhanced oxidative stress. This study investigated whether vitamins C and/or E modulate vascular function by regulating enzymatic activities of endothelial nitric oxide synthase (eNOS) and NAD(P)H oxidase using thoracic aortas of 20- to 22-week-old male spontaneously hypertensive rats (SHR) and their matched normotensive counterparts, Wistar-Kyoto rats (WKY). SHR aortas had impaired relaxant responses to acetylcholine but not to sodium nitroprusside, despite an 2-fold increase in eNOS activity and NO release. The levels of superoxide anion (O2 ), a potent NO scavenger, and NAD(P)H oxidase activity were also 2-fold higher in SHR aortas. Mechanical but not pharmacological inactivation of endothelium (by rubbing and 100 mol/L L-NAME, respectively) significantly abrogated O2 in both strains. Treatments of SHR aortas with NAD(P)H oxidase inhibitors, namely diphenyleneiodinium and apocynin, significantly diminished O2 production. The incubation of SHR aortas with different concentrations of vitamin C (10 to 100 mol/L) and specifically with high concentrations of vitamin E (100 mol/L) improved endothelial function, reduced superoxide production as well as NAD(P)H oxidase activity, and increased eNOS activity and NO generation in SHR aortas to the levels observed in vitamin C- and E-treated WKY aortas. Our results reveal endothelial NAD(P)H oxidase as the major source of vascular O2 in SHR and also show that vitamins C and E are critical in normalizing genetic endothelial dysfunction through regulation of eNOS and NAD(P)H oxidase activities.

Molecular Characterization and Localization of the NAD(P)H Oxidase Components gp91-phox and p22-phox in Endothelial Cells

The production of reactive oxygen species (ROS) within endothelial cells may have several effects, including alterations in the activity of paracrine factors, gene expression, apoptosis, and cellular injury. Recent studies indicate that a phagocyte-type NAD(P)H oxidase is a major source of endothelial ROS. In contrast to the high-output phagocytic oxidase, the endothelial enzyme has much lower biochemical activity and a different substrate specificity (NADH.NADPH). In the present study, we (1) cloned and characterized the cDNA and predicted amino acid structures of the 2 major subunits of rat coronary microvascular endothelial cell NAD(P)H oxidase, gp91-phox and p22-phox; (2) undertook a detailed comparison with phagocytic NADPH oxidase sequences; and (3) studied the subcellular location of these subunits in endothelial cells. Although these studies revealed an overall high degree of homology (.90%) between the endothelial and phagocytic oxidase subunits, the endothelial gp91-phox sequence has potentially important differences in a putative NADPH-binding domain and in putative glycosylation sites. In addition, the subcellular location of the endothelial gp91-phox and p22-phox subunits is significantly different from that reported for the neutrophil oxidase, in that they are predominantly intracellular and collocated in the vicinity of the endoplasmic reticulum. This first detailed characterization of gp91-phox and p22-phox structure and location in endothelial cells provides new data that may account, in part, for the differences in function between the phagocytic and endothelial NAD(P)H oxidases.

Nitric oxide synthase and NAD(P)H oxidase modulate coronary endothelial cell growth

Reactive oxygen species (ROS) including nitric oxide (NO) and superoxide anion (O2-) are associated with cell migration, proliferation and many growth-related diseases. The objective of this study was to determine whether there was a reciprocal relationship between rat coronary microvascular endothelial cell (CMEC) growth and activity/expressions (mRNA and protein) of endothelial NO synthase (eNOS) and NAD(P)H oxidase enzymes. Proliferating namely, 50% confluent CMEC possessed approximately three-fold increased activity and expression of both enzymes compared to 100% confluent cells. Treatment of CMEC with an inhibitor of eNOS (L-NAME, 100M) increased cell proliferation as assessed via three independent methods i.e. cell counting, determination of total cellular protein levels and [3H]thymidine incorporation. Similarly, treatment of CMEC with pyrogallol (0.3-3 mM), a superoxide anion (O2-)- generator, also increased CMEC growth while spermine NONOate (SpNO), a NO donor, significantly reduced cell growth. Co-incubation of CMEC with a cell permeable superoxide dismutase mimetic (Mn-III-tetrakis-4-benzoic acid-porphyrin; MnTBAP) plus either pyrogallol or NO did not alter cell number and DNA synthesis thereby dismissing the involvement of peroxynitrite (OONO-) in CMEC proliferation. Specific inhibitors of NAD(P)H oxidase but not other ROS-generating enzymes including cyclooxygenase and xanthine oxidase, attenuated cell growth. Transfection of CMEC with antisense p22-phox cDNA, a membrane-bound component of NAD(P)H oxidase, resulted in substantial reduction in [3H]thymidine incorporation, total cellular protein levels and expression of p22-phox protein. These data demonstrate a cross-talk between CMEC growth and eNOS and NAD(P)H oxidase enzyme activity and expression, thus suggesting that the regulation of these enzymes may be critical in preventing the initiation and/or progression of coronary atherosclerosis.

Fetal-maternal interface impedance parallels local NADPH oxidase related superoxide production

Blood flow assessment employing Doppler techniques is a useful procedure in pregnancy evaluation, as it may predict pregnancy disorders coursing with increased uterine vascular impedance, as pre-eclampsia. While the local causes are unknown, emphasis has been put on reactive oxygen species (ROS) excessive production. As NADPH oxidase (NOX) is a ROS generator, it is hypothesized that combining Doppler assessment with NOX activity might provide useful knowledge on placental bed disorders underlying mechanisms. A prospective longitudinal study was performed in 19 normal course, singleton pregnancies. Fetal aortic isthmus (AoI) and maternal uterine arteries (UtA) pulsatility index (PI) were recorded at two time points: 20-22 and 40-41 weeks, just before elective Cesarean section. In addition, placenta and placental bed biopsies were performed immediately after fetal extraction. NOX activity was evaluated using a dihydroethidium-based fluorescence method and associations to PI values were studied with Spearman correlations. A clustering of pregnancies coursing with higher and lower PI values was shown, which correlated strongly with placental bed NOX activity, but less consistently with placental tissue. The study provides evidence favoring that placental bed NOX activity parallels UtA PI enhancement and suggests that an excess in oxidation underlies the development of pregnancy disorders coursing with enhanced UtA impedance.

PPARα regulates endothelial progenitor cell maturation and myeloid lineage differentiation through a NADPH oxidase-dependent mechanism in mice

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