Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.6.3.1 (NADPH oxidase)
11,281 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Inflammatory cytokines, interleukin 1beta and tumor necrosis factor-alpha, potently stimulate rat mesangial cells to express and secrete group IIA phospholipase A(2) (PLA(2)). Cytokine-induced up-regulation of PLA(2) has been blocked by inhibitors (antioxidants) of the transcription factor, nuclear factor-kappaB (NF-kappaB), suggesting a role for NF-kappaB in the regulation of group IIA PLA(2) expression. Reactive oxygen species such as H(2)O(2), which are elevated in mesangial cells after cytokine activation, can mimic cytokine-induced NF-kappaB activation. However, the source of reactive oxygen species generation in mesangial cells, produced by cytokine stimulation, has yet to be clarified. Recently, tumor necrosis factor-alpha has been demonstrated to increase superoxide radical generation in mesangial cells. Therefore, we hypothesized that a selective NADPH oxidase inhibitor, diphenyleneiodium chloride (DPI), could block cytokine-induced group IIA PLA(2) up-regulation by attenuating NF-kappaB binding. To test this hypothesis, we isolated rat mesangial cells and characterized them by ultrastructural and immunochemical methods. This homogeneous mesangial cell population was responsive to cytokine as evidenced by an increase in steady-state levels of group IIA PLA(2) mRNA and extracellular enzymatic activity over time. DPI (0.02-20 microM), added 90 min before cytokine activation, inhibited both group IIA PLA(2) mRNA and enzymatic activity in a concentration-dependent manner. By electrophoretic mobility shift analysis, cytokine activation also increased specific NF-kappaB binding to one of two NF-kappaB consensus elements in the rat group IIA PLA(2) promoter and also was suppressed by DPI pretreatment. Antibodies to NF-kappaB p65 (Rel A) and p50 (but not normal rabbit IgG) supershifted this retardation signal and verified the type of NF-kappaB species as the classical p50/p65 heterodimer.
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PMID:Diphenyleneiodium chloride blocks inflammatory cytokine-induced up-regulation of group IIA phospholipase A(2) in rat mesangial cells. 1060 58

Direct in vivo evidence for the susceptibility of human neuronal cells to dengue virus has not been reported. In this study, we demonstrated that type 2 dengue (DEN-2) virus infection induced extensive apoptosis in the human neuroblastoma cell line SK-N-SH. Phospholipase A(2) (PLA(2)) was activated by DEN-2 infection, which led to the generation of arachidonic acid (AA). Inhibition of PLA(2) activity by the PLA(2) inhibitors, AACOCF(3) and ONO-RS-082, diminished DEN-2 virus-induced apoptosis. In contrast, the cyclooxygenase inhibitors aspirin and indomethacin, thought to increase AA accumulation by blocking AA catabolism, enhanced apoptosis. Exogenous AA induced apoptosis in a dose-dependent manner. Superoxide anion, which is thought to be generated through the AA-activated NADPH oxidase, was increased after infection. Pretreatment with superoxide dismutase (SOD) protected cells against DEN-2 virus-induced apoptosis. Furthermore, generation of superoxide anion was blocked by AACOCF(3). In addition, the transcription factors, NF-kappaB and c-Jun, were found to be activated after DEN-2 virus infection. However, pretreatment of cells with oligodeoxynucleotides containing NF-kappaB, but not c-Jun, binding sites (transcription factor decoy) strongly prevented dengue virus-induced apoptosis. The finding that AACOCF(3) and SOD significantly block activation of NF-kappaB suggests that this activation is derived from the AA-superoxide anion pathway. Our results indicate that DEN-2 virus infection of human neuroblastoma cells triggers an apoptotic pathway through PLA(2) activation to superoxide anion generation and subsequently to NF-kappaB activation. This apoptotic effect can be either directly derived from the action of AA and superoxide anion on mitochondria or indirectly derived from the products of apoptosis-related genes activated by NF-kappaB.
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PMID:Potential dengue virus-triggered apoptotic pathway in human neuroblastoma cells: arachidonic acid, superoxide anion, and NF-kappaB are sequentially involved. 1095 69

Arachidonic acid (AA) generated by phospholipase A(2) (PLA(2)) is thought to be an essential cofactor for phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity. Both enzymes are simultaneously primed by cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor-alpha (TNF-alpha). The possibility that either unprimed or cytokine-primed responses of PLA(2) or NADPH oxidase to the chemotactic agents formyl-methionyl-leucyl-phenylalanine (FMLP) and complement factor 5a (C5a) could be differentially inhibited by inhibitors of the mitogen-activated protein (MAP) kinase family members p42(ERK2) (PD98059) and p38(SAPK) (SB203580) was investigated. PD98059 inhibited the activation of p42(ERK2) by GM-CSF, TNF-alpha, and FMLP, but it did not inhibit FMLP-stimulated superoxide production in either unprimed or primed neutrophils. There was no significant arachidonate release from unprimed neutrophils stimulated by FMLP, and arachidonate release stimulated by calcium ionophore A23187 was not inhibited by PD98059. In contrast, PD98059 inhibited both TNF-alpha- and GM-CSF-primed PLA(2) responses stimulated by FMLP. On the other hand, SB203580 inhibited FMLP-superoxide responses in unprimed as well as TNF-alpha- and GM-CSF-primed neutrophils, but failed to inhibit TNF-alpha- and GM-CSF-primed PLA(2) responses stimulated by FMLP, and additionally enhanced A23187-stimulated arachidonate release, showing that priming and activation of PLA(2) and NADPH oxidase are differentially dependent on both the p38(SAPK) and p42(ERK2) pathways. Studies using C5a as an agonist gave similar results and confirmed the findings with FMLP. In addition, methyl arachidonyl fluorophosphonate (MAFP), the dual inhibitor of c and iPLA(2) enzymes, failed to inhibit superoxide production in primed cells at concentrations that inhibited arachidonate release. These data demonstrate that NADPH oxidase activity can be dissociated from AA generation and indicate a more complex role for arachidonate in neutrophil superoxide production.
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PMID:Activation and priming of neutrophil nicotinamide adenine dinucleotide phosphate oxidase and phospholipase A(2) are dissociated by inhibitors of the kinases p42(ERK2) and p38(SAPK) and by methyl arachidonyl fluorophosphonate, the dual inhibitor of cytosolic and calcium-independent phospholipase A(2). 1129 Jun 12

Production of toxic oxygen metabolites provides a mechanism for microbicidal activity of the neutrophil. The NADPH oxidase enzyme system initiates the production of oxygen metabolites by reducing oxygen to form superoxide anion (O(2)()). With stimulation of the respiratory burst, cytosolic oxidase components, p47(phox), p67(phox), and Rac, translocate to the phagolysomal and plasma membranes where they form a complex with cytochrome b(558) and express enzyme activity. A 29-kDa neutrophil protein (p29) was identified by co-immunoprecipitation with p67(phox). N-terminal sequence analysis of p29 revealed homology to an open reading frame gene described in a myeloid leukemia cell line. A cDNA for p29 identical to the open reading frame protein was amplified from RNA of neutrophils. Significant interaction between p29 and p67(phox) was demonstrated using a yeast two-hybrid system. A recombinant (rh) p29 was expressed in Sf9 cells resulting in a protein with an apparent molecular weight of 34,000. The rh-p29 showed immunoreactivity with the original rabbit antiserum that detected p47(phox) and p67(phox). In addition, rh-p29 exhibited PLA(2) activity, which was Ca(2+) independent, optimal at low pH, and preferential for phosphatidylcholine substrates. The recombinant protein protected glutathione synthetase and directly inactivated H(2)O(2). By activity and sequence homology, rh-p29 can be classified as a peroxiredoxin. Finally, O(2)() production by plasma membrane and recombinant cytosolic oxidase components in the SDS-activated, cell-free NADPH oxidase system were enhanced by rh-p29. This effect was not inhibited by PLA(2) inhibitors. Thus, p29 is a novel protein that associates with p67 and has peroxiredoxin activity. This protein has a potential role in protecting the NADPH oxidase by inactivating H(2)O(2) or altering signaling pathways affected by H(2)O(2).
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PMID:A 29-kDa protein associated with p67phox expresses both peroxiredoxin and phospholipase A2 activity and enhances superoxide anion production by a cell-free system of NADPH oxidase activity. 1212 78

Experimental studies have suggested that TNF alpha, a pro-inflammatory cytokine, may contribute to the deterioration of cardiovascular function through various mechanisms, including the generation of reactive oxygen species. It has not yet been demonstrated whether TNF alpha has prooxidant activity in patients with heart failure, and what the mechanism eventually resulting in this effect are. We analyzed 42 patients (38 men and 4 women, aged 26 to 74 years) with heart failure, secondary to idiopathic dilated cardiomyopathy (n=21), coronary artery disease (n=15), and valve disease (n=6), and 20 controls (18 men and 2 women, aged 49 to 67 years). Ten patients were in class I, 9 in class II, 15 in class III and 8 in class IV according to NYHA Classification. Blood samples were obtained from each patient to evaluate basal and collagen-induced platelet O(2)(-) production, and plasma TNF alpha. In vivo results showed increased platelet O(2)(-) production and plasma TNF alpha levels in NYHA class III-IV compared with that in controls or in NYHA I-II (p<0,001); platelet O(2)(-) production correlated significantly (R=0,6; p<0,01) with TNF alpha plasma levels. In vitro studies showed TNF alpha dose-dependently (5-40 pg/ml) induced platelet O(2)(-) production, and that this effect was significantly inhibited by its specific inhibitor, WP9QY (1 microM); aspirin (100 microM), AACOCF(3), a specific PLA(2) inhibitor (14 microM), and DPI, an inhibitor of NADPH oxidase, significantly inhibited TNF alpha-mediated platelet O(2)(-) production. This study suggests that in patients with heart failure, enhanced platelet O(2)(-) production is mediated by TNF alpha via activation of arachidonic acid and NADPH oxidase pathways.
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PMID:Enhanced TNF alpha and oxidative stress in patients with heart failure: effect of TNF alpha on platelet O2- production. 1288 80

ANG II activation of phospholipase D (PLD) is required for ERK and NAD(P)H oxidase activation, both of which are involved in hypertension. Previous findings demonstrate that ANG II stimulates PLD activity through AT(1) receptors in a RhoA-dependent mechanism. Additionally, endogenous AT(2) receptors in preglomerular smooth muscle cells attenuate ANG II-mediated PLD activity. In the present study, we examined the signal transduction mechanisms used by endogenous AT(2) receptors to modulate ANG II-induced PLD activity through either PLA(2) generation of lysophosphatidylethanolamine or Galpha(i)-mediated generation of nitric oxide (NO) and interaction with RhoA. Blockade of AT(2) receptors, Galpha(i) and NO synthase, but not PLA(2), enhanced ANG II-mediated PLD activity in cells rich in, but not poor in, AT(2) receptors. Moreover, NO donors, a direct activator of guanylyl cyclase and a cGMP analog, but not lysophosphatidylethanolamine, inhibited ANG II-mediated PLD activity, whereas an inhibitor of guanylyl cyclase augmented ANG II-induced PLD activity. AT(2) receptor- and NO-mediated attenuation of ANG II-induced PLD activity was completely lost in cells transfected with S188A RhoA, which cannot be phosphorylated on serine 188. Therefore, our data indicate that AT(2) receptors activate Galpha(i), subsequently stimulating NO synthase and leading to increased soluble guanylyl cyclase activity, generation of cGMP, and activation of a protein kinase, resulting in phosphorylation of RhoA on serine 188. Furthermore, because AT(2) receptors inhibit AT(1) receptor signaling to PLD via modulating RhoA activity, AT(2) receptor signaling can potentially regulate multiple vasoconstrictive signaling systems through inactivating RhoA.
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PMID:AT2 receptors cross talk with AT1 receptors through a nitric oxide- and RhoA-dependent mechanism resulting in decreased phospholipase D activity. 1557 19

In a cat model of acute experimental esophagitis, resting in vivo lower esophageal sphincter (LES) pressure and in vitro tone are lower than in normal LES, and the LES circular smooth muscle layer contains elevated levels of IL-1beta that decrease the LES tone of normal cats. We now examined the mechanisms of IL-1beta-induced reduction in LES tone. IL-1beta significantly reduced acetylcholine-induced Ca(2+) release in Ca(2+)-free medium, and this effect was partially reversed by catalase, demonstrating a role of H(2)O(2) in these changes. IL-1beta significantly increased the production of H(2)O(2), and the increase was blocked by the p38 MAPK inhibitor SB-203580, by the cytosolic phospholipase A(2) (cPLA(2)) inhibitor AACOCF3, and by the NADPH oxidase inhibitor apocynin, but not by the MEK1 inhibitor PD-98059. IL-1beta significantly increased the phosphorylation of p38 MAPK and cPLA(2). IL-1beta-induced cPLA(2) phosphorylation was blocked by SB-203580 but not by AACOCF3, suggesting sequential activation of p38 MAPK-phosphorylating cPLA(2). The IL-1beta-induced reduction in LES tone was partially reversed by AACOCF3 and by the Ca(2+)-insensitive PLA(2) inhibitor bromoenol lactone (BEL). IL-1beta significantly increased cyclooxygenase (COX)-2 and PGE(2) levels. The increase in PGE(2) was blocked by SB-203580, AACOCF3, BEL, and the COX-2 inhibitor NS-398 but not by PD-98059 or the COX-1 inhibitor valeryl salicylate. The data suggested that IL-1beta reduces LES tone by producing H(2)O(2), which may affect Ca(2+)-release mechanisms and increase the synthesis of COX-2 and PGE(2). Both H(2)O(2) and PGE(2) production depend on sequential activation of p38 MAPK and cPLA(2). cPLA(2) activates NADPH oxidases, producing H(2)O(2), and may produce arachidonic acid, converted to PGE(2) via COX-2.
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PMID:IL-1beta signaling in cat lower esophageal sphincter circular muscle. 1664 61

Electrophysiological events are of central importance during the phagocyte respiratory burst, because NADPH oxidase is electrogenic and voltage sensitive. We investigated the recent suggestion that large-conductance, calcium-activated K(+) (BK) channels, rather than proton channels, play an essential role in innate immunity (Ahluwalia, J., A. Tinker, L.H. Clapp, M.R. Duchen, A.Y. Abramov, S. Page, M. Nobles, and A.W. Segal. 2004. Nature. 427:853-858). In PMA-stimulated human neutrophils or eosinophils, we did not detect BK currents, and neither of the BK channel inhibitors iberiotoxin or paxilline nor DPI inhibited any component of outward current. BK inhibitors did not inhibit the killing of bacteria, nor did they affect NADPH oxidase-dependent degradation of bacterial phospholipids by extracellular gIIA-PLA(2) or the production of superoxide anion (O(2*)(-)). Moreover, an antibody against the BK channel did not detect immunoreactive protein in human neutrophils. A required role for voltage-gated proton channels is demonstrated by Zn(2+) inhibition of NADPH oxidase activity assessed by H(2)O(2) production, thus validating previous studies showing that Zn(2+) inhibited O(2*)(-) production when assessed by cytochrome c reduction. In conclusion, BK channels were not detected in human neutrophils or eosinophils, and BK inhibitors did not impair antimicrobial activity. In contrast, we present additional evidence that voltage-gated proton channels serve the essential role of charge compensation during the respiratory burst.
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PMID:The antibacterial activity of human neutrophils and eosinophils requires proton channels but not BK channels. 1670 53

Picolinic acid (PA) potentiates macrophage (MPhi) antimicrobial activity against intracellular Mycobacterium avium complex (MAC). Here, we studied the mechanisms of this phenomenon using human THP-1 MPhis. First, when PA-treated MAC-infected MPhis were cultured in the presence or absence of reactive oxygen intermediate (ROI) scavengers, nitric oxide synthase (NOS) inhibitors or phospholipase A(2) (PLA(2)) inhibitors, none of these agents blocked the activity of PA in potentiating MPhi anti-MAC activity. Second, when PA was added to the in vitro anti-MAC bactericidal system consisting of either ROIs, reactive nitrogen intermediates (RNIs) or free fatty acid (FFA) molecules, which are the major MPhi antimicrobial effectors, PA inhibited the activity of ROIs and conversely potentiated the activity of RNIs; PA did not affect the activity of FFAs. Third, PA reduced mRNA expression of NADPH oxidase and beta-defensin-1 by MAC-infected MPhis, whilst neither cytosolic PLA(2) nor CAP37 mRNA expression was affected. Notably, inducible NOS and secretory PLA(2) mRNA expression was not detected for MAC-infected MPhis even when given PA treatment. These findings suggest that ROIs, RNIs, FFAs and beta-defensin-1 do not play important roles in the PA-induced potentiation of MPhi anti-MAC activity.
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PMID:Effects of picolinic acid on the antimicrobial functions of host macrophages against Mycobacterium avium complex. 1729 87

Hypotonic exposure provokes the mobilization of arachidonic acid, production of ROS, and a transient increase in taurine release in Ehrlich Lettre cells. The taurine release is potentiated by H(2)O(2) and the tyrosine phosphatase inhibitor vanadate and reduced by the phospholipase A(2) (PLA(2)) inhibitors bromoenol lactone (BEL) and manoalide, the 5-lipoxygenase (5-LO) inhibitor ETH-615139, the NADPH oxidase inhibitor diphenyl iodonium (DPI), and antioxidants. Thus, swelling-induced taurine efflux in Ehrlich Lettre cells involves Ca(2+)-independent (iPLA(2))/secretory PLA(2) (sPLA(2)) plus 5-LO activity and modulation by ROS. Vanadate and H(2)O(2) stimulate arachidonic acid mobilization and vanadate potentiates ROS production in Ehrlich Lettre cells and NIH3T3 fibroblasts under hypotonic conditions. However, vanadate-induced potentiation of the volume-sensitive taurine efflux is, in both cell types, impaired in the presence of BEL and DPI and following restoration of the cell volume. Thus, potentiation of the volume-sensitive taurine efflux pathway following inhibition of tyrosine phosphatase activity reflects increased arachidonic acid mobilization and ROS production for downstream signaling. Vanadate delays the inactivation of volume-sensitive taurine efflux in NIH3T3 cells, and this delay is impaired in the presence of DPI. Vanadate has no effect on the inactivation of swelling-induced taurine efflux in Ehrlich Lettre cells. It is suggested that increased tyrosine phosphorylation of regulatory components of NADPH oxidase leads to increased ROS production and a subsequent delay in inactivation of the volume-sensitive taurine efflux pathway and that NADPH oxidase or antioxidative capacity differ between NIH3T3 and Ehrlich Lettre cells.
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PMID:Activation and inactivation of the volume-sensitive taurine leak pathway in NIH3T3 fibroblasts and Ehrlich Lettre ascites cells. 1753 4


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